An Autonomous Chemically-Fuelled

Rotary Motor

A thesis submitted to The University of Manchester for the

degree of

Doctor of Philosophy

in the Faculty of Engineering and Physical Sciences

2015

Miriam Ruth Wilson

School of Chemistry

2

Contents

Abstract ......................................................................................................................................... 4

Declaration and Copyright Statement .......................................................................................... 5

Acknowledgements ....................................................................................................................... 6

Abbreviations ................................................................................................................................ 7

General Experimental ................................................................................................................. 11

Chapter One ................................................................................................................................ 13

1.1. Synopsis........................................................................................................................... 14

1.2. Introduction .................................................................................................................... 15

1.3. Unidirectional Rotation Around C-C Single Bonds .......................................................... 15

1.4. Unidirectional Rotation Around C=C Double Bonds ....................................................... 19

1.4.1. Applications ................................................................................................................. 27

1.4.1.1. From relative to absolute rotation .......................................................................... 27

1.4.1.2. Switching chirality ................................................................................................... 29

1.4.1.3. Cumulative work ..................................................................................................... 32

1.5. Unidirectional Rotation Around C=N Imine Bonds ......................................................... 34

1.6. Unidirectional Rotation in Interlocked Architectures ..................................................... 37

1.7. Conclusion ....................................................................................................................... 42

1.8. References ...................................................................................................................... 43

Chapter Two ................................................................................................................................ 48

2.1. Synopsis........................................................................................................................... 49

2.2. Introduction .................................................................................................................... 50

2.3. Design .............................................................................................................................. 56

2.4. Retrosynthesis ................................................................................................................. 63

2.5. Synthesis ......................................................................................................................... 64

2.6. Operation and Analysis ................................................................................................... 68

2.6.1. Directionality ............................................................................................................... 68

2.6.2. Autonomy ................................................................................................................... 73

2.7. Conclusion ....................................................................................................................... 79

2.8. Experimental Section ...................................................................................................... 80

2.9. References .................................................................................................................... 100

Chapter Three ........................................................................................................................... 101

3.1. Synopsis......................................................................................................................... 102

3.2. Retrosynthesis ............................................................................................................... 103

3

3.3. Synthesis ....................................................................................................................... 104

3.4. Operation and Analysis ................................................................................................. 109

3.4.1. Directionality Studies ................................................................................................ 109

3.4.2. Autonomy ................................................................................................................. 116

3.5. Conclusion and Outlook ................................................................................................ 128

3.6. Experimental Section .................................................................................................... 130

3.7. References .................................................................................................................... 178

FINAL WORD COUNT

33461

4

Abstract

An Autonomous Chemically-Fuelled Rotary Motor

Miriam Ruth Wilson

A thesis submitted to The University of Manchester for the degree of Doctor of Philosophy in the Faculty of Engineering and Physical Sciences

2015

Biological processes commonly use molecular motors to drive chemical systems away from equilibrium thus enabling work to be done. This has inspired efforts to create synthetic rotary motors which mimic the key properties of their biological counterparts, namely autonomy and directionality with the use of a chemical fuel. Thus far, attempts to combine all three properties in a synthetic rotary motor have proven unsuccessful. This thesis describes the design, synthesis and operation of an autonomous, chemically-fuelled, directional rotary motor. In this two-compartment [2]catenane an information ratchet mechanism operates. Directional transport of the small macrocycle around the larger one is promoted by an acylation reaction using a sterically demanding pyridine-based catalyst. To achieve autonomy, conditions for a one-pot, directional, de-acylation/re-acylation reaction were developed. Under autonomous operation conditions the macrocycle displacement was followed by 1H NMR.

Figure 1: Catenane motor and schematic operation cycle. The acylation step creates a directional bias resulting in

net clockwise motion (as drawn) of the small macrocycle around the larger one.

Chapter One describes the previous strategies that have been employed to realise unidirectional rotary motion in synthetic systems and aims to give the reader an overview of the relevant literature in the field of synthetic rotary motors.

Chapter Two describes the concept and design of the project. Previous work which formed the basis for this research is also discussed. The synthesis and successful operation of a molecular information ratchet fuelled by chemical energy is reported.

Chapter Three describes the first autonomous, synthetic rotary motor fuelled by chemical energy. The autonomous nature of the operation is determined by 1H NMR, mass spectrometry and HPLC.

5

Declaration and Copyright Statement

Unless otherwise stated at the beginning of each chapter, the work referred to in this thesis has not been submitted in support of an application for another degree or qualification of this or any other university or other institute of learning.

i. The author of this thesis (including any appendices and/or schedules to this thesis) owns certain copyright or related rights in it (the “Copyright”) and s/he has given The University of Manchester certain rights to use such Copyright, including for administrative purposes.

ii. Copies of this thesis, either in full or in extracts and whether in hard or electronic copy, may be made only in accordance with the Copyright, Designs and Patents Act 1988 (as amended) and regulations issued under it or, where appropriate, in accordance with licensing agreements which the University has from time to time. This page must form part of any such copies made.

iii. The ownership of certain Copyright, patents, designs, trade marks and other intellectual property (the “Intellectual Property”) and any reproductions of copyright works in the thesis, for example graphs and tables (“Reproductions”), which may be described in this thesis, may not be owned by the author and may be owned by third parties. Such Intellectual Property and Reproductions cannot and must not be made available for use without the prior written permission of the owner(s) of the relevant Intellectual Property and/or Reproductions.

iv. Further information on the conditions under which disclosure, publication and commercialisation of this thesis, the Copyright and any Intellectual Property and/or Reproductions described in it may take place is available in the University IP Policy (see http://documents.manchester.ac.uk/DocuInfo.aspx?DocID=487), in any relevant Thesis restriction declarations deposited in the University Library, The University Library’s regulations (see http://www.manchester.ac.uk/library/aboutus/regulations) and in The University’s policy on Presentation of Theses

Miriam Ruth Wilson September 2015

6

Acknowledgements

I would firstly like to thank my supervisor Prof. David Leigh for allowing me to work on such a

fascinating project in such a wonderful laboratory environment.

When I first joined the group I had the pleasure of working alongside some amazing chemists:

Armando, Adam and Jordi, thank you for teaching me all I know. I would also like to thank

Nathalie and Steve for their contribution to the project before my arrival in the group.

Thanks to Louise, Sau Yin, Stuart and Valérie for keeping our lab running in good order. Thanks

to all the instrument monkeys: Bartek, Sonja, Steffen, Salma and Simone for the NMR; Dave H,

Gus, Malcolm, Jon D, Charlie and Ulvi for MS; and Vanesa and Javier for HPLC. I would also like

to thank Javier and Guillaume for help with Spartan and image production.

I would like to thank everybody else I had the pleasure of working with during my time in the

Leigh group: Alan, Alina, Alex, Angel, Anneke, Antonio, Ara, Barney, Bartek, Bea, Chris, Chris

Martin, Colm, Craig, Dan, Daniela, Dave H, Diederik, Gen, Guillaume, Guzman, Jack, Jason, Jeff

A, Jeff L, Jhenyi, John, Jon B, Kathleen, Leo, Louise, Marcus, Maria, Matt, Max, Mike, Mustafa,

Patrick, Paul, Philipp, Ramon, Shoufeng, Steve, Stewart, Sundus, Sunny, Taisuke, Tom, Tuba,

Tugrul, Ula, Valerie, Victor and Yusuf.

I would like to thank all the instrument wizards who have helped me at some point along my

journey: Anne, Carlo, Carole, Gareth, Ilya, Juraj, Lorna and Mohammed – your help has been

invaluable.

And finally, I thank my family for all of their support and encouragement over the years. This is

for you!

7

Abbreviations

(Note: conventional abbreviations for units and physical quantities not included here)

APCI Atomospheric pressure chemical ionisation

Aq. Aqueous

ATP Adenosine Triphosphate

BOC tert-butylcarbonyl

Boc2O Di-tert-butyl dicarbonate

Bz Benzoyl

Bz2O Benzoic anhydride

CBS Corey-Bakshi-Shibata

COSY Correlation spectroscopy

δ Chemical shift

DBU 1,8-Diazabicyclo[5.4.0]undec-7-ene

DCM Dichloromethane

DIPEA N,N-Diisopropylethylamine

DMAP 4-(Dimethylamino)pyridine

DMF Dimethylformamide

DMSO Dimethylsulfoxide

DNA deoxyribonucleic acid

8

Eq. Equivalents

ESI Electrospray ionisation

Et2O Diethyl ether

EtOH Ethanol

EtOAc Ethyl acetate

Fmoc Fluorenylmethyloxycarbonyl

Fmoc-Cl Fluorenylmethyloxycarbonyl Chloride

FRET Fluorescence resonance energy transfer

fum Fumaramide

h Hour(s)

HATU 1-[Bis(dimethylamino)methylene]-1H-2,3-triazolo[4,5-b]pyridinium 3-oxid

hexafluorophosphate

HMBC Heteronuclear multiple bond coherence

HOBt Hydroxybenzotriazole

HPLC High performance liquid chromatograhy

HRMS High resolution mass spectrometry

HSQC Heteronuclear single quantum coherence

HZ, MHz Hertz, Megahertz

IPA 2-propanol (isopropyl alcohol)

LC liquid crystal

9

LRMS Low resolution mass spectrometry

mal maleamide

MeOH Methanol

min Minute(s)

MP Melting point

m/z mass-to-charge ratio

NEt3 Triethylamine

NMR Nuclear magnetic resonance

NOESY Nuclear Overhauser effect spectroscopy

NSI Nanospray ionisation

Nu Nucleophile

Pet Petroleum Ether (40–60 °C)

PEG Polyethylene glycol

Ph Phenyl

PMB para-methyoxybenzyl ether

ppm part per million

quant. quantitative yield

Py pyridine

rt room temperature

succ succinamide

10

satd. Saturated

STM Scanning tunnel microscope

TBAF Tetra-n-butylammonium fluoride

TBDMS tert-butyldimethylsilyl

TBDPS tert-butyldiphenylsilyl

TBDPSCl tert-butyldiphenylsilyl chloride

TBAT Tetrabutylammonium difluorotriphenylsilicate

TBTU O-(Benzotriazol-1-yl)-N,N,N’,N’-tetramethyluronium tetrafluoroborate

TFA Trifluoroacetic acid

THF Tetrahydrofuran

TLC thin layer chromatography

TsCl 4-toluenesulfonyl chloride

UV Ultraviolet

wt. weight

11

General Experimental

Unless stated otherwise, reagents were obtained from commercial sources and used without

purification. Anhydrous THF (HPLC grade, Fischer scientific), CHCl3 (AR grade, Fisher scientific,

stabilised with amylene) and CH2Cl2 (HPLC grade, Fischer scientific) were obtained by passing

the solvent through an activated alumina column on a Phoenix SDS (solvent drying system; JC

Meyer Solvent Systems, CA, USA) or through an activated alumina column on a PureSolv™

solvent purification system (InnovativeTechnologies, Inc., MA, USA). DMF (Peptide synthesis

grade, Merck) was used throughout. 1H NMR spectra were recorded on Bruker AV 400, Bruker

AV 500 (equipped with a cryoprobe) and Bruker Avance III with an Oxford AS600 magnet

equipped with a cryoprobe [5mm CPDCH 13C-1 H/D] (600 MHz) instruments. Chemical shifts

are reported in parts per million (ppm) from high to low frequency using the residual solvent

peak as the internal reference (CDCl3 = 7.26 ppm, CD2Cl2 = 5.32 ppm and CD3OD = 3.31 ppm).

All 1H resonances are reported to the nearest 0.01 ppm. The multiplicity of 1H signals are

indicated as: s = singlet; d = doublet; t = triplet; quint = quintet; m = multiplet; br = broad; or

combinations of thereof. Coupling constants (J) are quoted in Hz and reported to the nearest

0.1 Hz. 13C NMR spectra were recorded on the same spectrometer with the central resonance

of the solvent peak as the internal reference (CDCl3 = 77.16 ppm, CD2Cl2 = 54.00 ppm, and

CD3OD = 49.00 ppm). All 13C resonances are reported to the nearest 0.01 ppm to aid in the

differentiation of closely resolved signals. DEPT, COSY, HSQC, HMBC and NOE experiments

were used to aid structural determination and spectral assignment. Fully characterized

compounds were chromatographically homogeneous. Flash column chromatography was

carried out using Silica 60 Å (particle size 40-63 μm, Sigma Aldrich, UK) as the stationary phase.

Automated purification was performed using prepacked silica columns (Reveleris® Flash

Cartridges) on the Reveleris® automated Flash Chromatography System. Preparative TLC was

12

performed using either PLC 20×20 cm, 60 F254 Prep plates (Merck) or Silica Gel GF 20×20 cm,

U254 Prep plates (Analtech) of various thicknesses. TLC was performed on precoated silica gel

plates (0.25 mm thick, 60 F254, Merck, Germany) and visualized using both short and long

waved ultraviolet light in combination with standard laboratory stains (acidic potassium

permanganate, acidic ammonium molybdate and ninhydrin). Low resolution ESI mass

spectrometry was performed with a Thermo Scientific LCQ Fleet Ion Trap Mass Spectrometer

or an Agilent Technologies 1200 LC system with 6130 single quadrupole MS detector. High-

resolution mass spectrometry was carried out by the EPSRC National Mass Spectrometry

Service Centre (Swansea, UK). Melting points (MP) were determined using a Büchi M-565

apparatus and are corrected. Optical rotations were measured using a Rudolph Research

Analytical Autopol I polarimeter with both AP Accuracy (±0.004°) and resolution upgrades with

a built in thermoprobe for temperature measurement/control. Measurements were

conducted using a sodium lamp ( 589 nm, D-line); [α]25𝐷

values were reported in 10 deg cm2

g-1, concentration (c) in g per 100 ml. Analytical HPLC was performed on an Agilent

Technologies (1200 LC system with photodiode array detector) instrument. A normal-phase

column (SUPELCO Ascentis-Si, analytical: 250 × 4.6 mm, 5 µm) was used with combined

isocratic and gradient elution (1.0 mL/min, CH2Cl2/ iPrOH, 5% → 5% → 40% → 40% → 5%

iPrOH; UV detection @ 254 nm). DFT calculations have been performed using Spartan ‘14. The

ground state geometries have been optimised using the B3LYP exchange-correlation functional

with the 6-31G(d) basis set.

Chapter One

Synthetic Rotary Motors

Acknowledgements

Dr Thomas A. Singleton and Dr John W. Ward are gratefully acknowledged for examining and

proofreading this chapter.

14

1.1. Synopsis

In this chapter the current strategies for producing synthetic motors capable of unidirectional

rotary motion, and their applications are reviewed. Although these motors are primitive by

Nature’s standards the basic principles of rotary motion have been demonstrated and the first

examples which exert control over microscopic and macroscopic properties are emerging. The

design and efficient operation of molecular motors holds promise and possible future

developments are briefly discussed.

15

1.2. Introduction

The design and synthesis of molecular machines and motors is among the most challenging

fields in modern science.1 Nature’s molecular machines rank among the most complex and

fascinating structures known and lie at the heart of virtually all important biological

processes.2–7 In the last two decades a number of synthetic molecular machines have been

developed which enable controlled movement, including Brownian ratchets,8 molecular

synthesisers9,10 and walking molecules.11 Inspired by Nature’s powerhouse ATP synthase2 and

the bacteria flagellar motor,3 several synthetic rotary motors have also been developed. Here

synthetic molecular motors capable of unidirectional rotation around single, double or

mechanical bonds are reviewed.

1.3. Unidirectional Rotation Around C-C Single Bonds

As a first attempt at achieving directional rotary motion, Kelly and co-workers prepared

molecular “cog-wheel” 1 (Figure 1a)12,13. In a molecular realisation of Feynman’s adiabatic

ratchet and pawl their “cog-wheel” consists of a triptycene rotor bound to a helicene stator

whose inherent chirality was intended to direct the rotation of the triptycene blade.

Computational studies revealed an asymmetric potential energy surface for rotation, with

clockwise rotation experiencing a steady increase in resistance, while anticlockwise motion

resulted in a much steeper barrier (Figure 1b). To determine the directional bias of 1, Kelly et

al. employed the elegant NMR technique of spin polarisation transfer. The rotation of the

triptycene around the helicene is slow on the NMR timescale, leading to three distinct signals

being observed. By selectively polarising of one of these signals and then observing the

spectrum after an appropriate time delay, it was possible to follow the rotation of the

16

polarised triptycene blade. Directional rotation should result in a disproportionate amount of

polarisation being transferred to one of the two other resonances. Rotation with no directional

bias would result in polarisation transfer at equal rates to both other resonances.

Disappointingly, but not surprisingly, the triptycene blade rotates in both directions with equal

ease, consistent with Feynman’s thought experiment.14

Figure 1: Kelly’s first generation molecular “cog-wheel”.12,13

In an effort to find a solution and create a true directional motor, a modified version of 1 was

synthesised in which a chemical reaction would provide the required energy input.15,16 The

new design (2, Scheme 1) incorporates an amine on the triptycene blade, and an alcohol tether

on the helicene stator. If the amino group is ignored, the three different rotamers of 2

represent three equal energy minima. Reaction of 2 with phosgene gives isocyanate 3, which

as a result of random rotation, will encounter the alcohol tether and react to form urethane 4.

This irreversible step “ratchets” the motion part way up the rotational energy barrier.

Subsequent thermal relaxation continues rotation in the same direction over the energy

barrier and yields 4’. Cleavage of the urethane linker gives the original molecule rotated by

120°. Although this molecular machine only performs one third of a turn it incorporates the

features necessary for cumulative work and was a major development in the field of molecular

machines. Unfortunately, despite extensive work 360° rotation has not yet been achieved with

this system.17

17

Scheme 1: 120° rotation about a C-C bond powered by chemical energy.15

An alternative approach to a chemically driven rotary motor was reported by Dahl and

Branchaud.18 Their system relied on the stereoselective ring-opening and ring-closing of biaryl

lactones; however, a full rotary cycle remained a challenge (Scheme 2). Racemic biaryl lactone

(R)-5/(S)-5 was subjected to lactone cleavage in the presence of cumylamine, affording ring-

opened amide (M-S)-6/(P-R)-6 with very high diastereoselectivity. Through chemoselective

carboxylic acid activation, relactonization proceeded to give (S)-7, representing a 180° rotation

about the aryl-aryl bond relative to (S)-1. Chemoselective hydrolysis of the cumylamide bond

in the presence of the lactone would have regenerated 5 and allowed repetitive rotation;

however, the authors were unable to develop such experimental conditions.

Scheme 2: Unidirectional rotation around a C-C bond based on ring-opening/ring-closing of a chiral lactone.18

18

In an independent yet almost simultaneous effort, Feringa and co-workers successfully

extended this strategy to yield the first example of a synthetic molecular motor which was

capable of full and repetitive 360° rotation fuelled by chemical energy.19 Through the choice of

chemical reagents and order of reactions the directionality of the motor could be controlled.

Scheme 3: 360° unidirectional rotation around a C-C bond fuelled by chemical energy. Step 1) Stereoselective

reduction with (S)-CBS then allyl protection. Step 2) Chemoselective cleavage of PMB group. Spontaneous

lactonisation follows. Step 3) Stereoselective reduction with (S)-CBS then PMB protection. Step 4) Chemoselective

cleavage of the allyl group. Spontaneous lactonisation follows. Step 5) Oxidation to carboxylic acid.19

Reduction of the lactone (Scheme 3, step 1 and 3) was achieved using BH3·THF and the chiral

reagent (S)-CBS in excellent enantioselectivities. The resulting phenol and alcohol moieties are

protected and oxidised to the acids, respectively. The ortho substitution of species 9 and 11

blocks free rotation around the central C-C bond for steric reasons. After selective

deprotection, the liberated phenol group spontaneously undergoes lactonisation to give 8 or

10. The orthogonal nature of the protecting group strategy employed allows for selective

deprotection-ring closure of the lactone ensuring complete unidirectionality is achieved.

19

Although 8 demonstrates the principle of chemically-fuelled 360° unidirectional rotation, the

number of chemical reactions and purifications required to complete a sequence significantly

reduces the practicality.

1.4. Unidirectional Rotation Around C=C Double Bonds

The first synthetic rotary motor capable of autonomous (i.e. the components move

directionally as long as fuel is present) repetitive unidirectional rotation was developed by

Feringa and co-workers.20 Their motor is based on an overcrowded alkene, and uses

photochemical energy to fuel rotation around the central C=C bond. Upon irradiation with

light, overcrowded alkene (3R-3′R)-(P-P)-trans-12 can undergo a full 360° rotation of the top

half of the molecule (rotor) relative to the bottom half (stator). These so-called first generation

overcrowded alkene molecular motors have several important features: (i) the inherent

helicity of the sterically crowded alkene; (ii) the central carbon-carbon double bond which

connects the two identical halves and acts as the axis of rotation; and (iii) two methyl-

substituted stereogenic centres which control the direction of rotation. In the stable isomers of

the motor these methyl substituents adopt a low energy pseudo-axial conformation in order to

minimise steric repulsion with the other half of the molecule (Scheme 4).

20

Scheme 4: The first light-powered unidirectional molecular motor.20 Step 1) and Step 3) Photochemical trans → cis

isomerisation. Step 2) and Step 4) Thermally-induced helical inversion.

Irradiation of (3R-3′R)-(P-P)-trans-12 with UV light (λ > 280 nm) causes photochemical trans →

cis isomerisation of the central carbon-carbon double bond, which induces helical inversion

(P,P → M,M) of the molecule. In this isomer (3R-3′R)-(M-M)-cis-12, the methyl substituents are

forced into a pseudo-equatorial conformation where they experience significant steric

crowding with the naphthalene rings. Thermally-activated helix inversion allows the molecule

to spontaneously relax to the stable isomer (3R-3′R)-(P-P)-cis-12 where the methyl

substituents are in the more favoured axial conformation. The large difference in free energy

between these two isomers renders this step irreversible ensuring the unidirectionality of the

process. Further irradiation (λ > 280 nm) again initiates a photochemical trans → cis

isomerisation and generates unstable isomer (3R-3′R)-(M-M)-trans-12 where the methyl

substituents are again in an unfavoured pseudo-equatorial conformation. Gentle heating (60

°C) facilitates the last helix inversion, thus restoring the molecule to its original stable

conformation (3R-3′R)-(P-P)-trans-12. Remarkably, heating combined with sustained

irradiation results in continual repetitive unidirectional rotation of one half of the molecule

relative to the other; it operates autonomously. A number of modifications to the basic

molecular design of the first generation motor were made in an attempt to tune the rate of

the thermal helical inversion and ultimately optimise the rotation process. The most dramatic

21

increase in rotation was observed upon contraction of the rings fused to the central olefin (t1/2

from 439 h to 74 min).21

Concurrently with the refinements in the design of 12, a new “second generation” of

overcrowded alkene molecular motors was explored which consist of distinct upper and lower

halves.22 The upper half was similar to that used in the first generation motors: i.e., a

naphthalene unit fused to a six-membered ring with a methyl substituent at the stereogenic

centre, responsible for controlling the direction of rotation. The lower half however is now

derived from a symmetric tricyclic molecule – an alteration which offered practical advantages

including milder synthetic conditions and more selective functionalization. The second

generation motors operate in the same fashion as the first generation: (i) irradiation of (M)-

trans-13 initiates photoisomerisation of the central carbon-carbon bond from trans → cis to

form unstable (P)-cis-13 in which the substituent at the stereogenic centre is trapped in an

unfavourable conformation; (ii) thermal helix inversion releases the strain and forms (M)-cis-

13; (iii) in analogy to step (i) photoisomerisation generates the strained, unstable isomer (P)-

trans-13; (iv) unidirectional 360° rotation is complete upon thermal helix inversion

regenerating (M)-trans-13.

Scheme 5: Rotary cycle of a second generation molecular motor.22

22

As with the first generation overcrowded alkene molecular motors, many structural

modifications of the second generation design were made to explore the limits of the system.

Moving the position of the stereogenic centre away from the fjord region to the 3′ position of

the upper ring was anticipated to reduce the steric crowding and increase the rate of thermal

helix inversion. However, it was found that this simple modification reduced the energy

difference between the stable and unstable isomers to the extent that the unidirectionality of

the system was significantly compromised.23 In the series of molecular motors 14—19, simple

variations in bridging atoms X and Y were found to have a dramatic effect on the rate of

thermal helix inversion.24 For example, simply exchanging X from sulfur (14) to a methylene

(17) group resulted in a 300-fold increase in the rate of thermal helix inversion at room

temperature, from 215 hours to only 39 minutes (Table 1). Motor 20 was designed to probe

electronic control over the rate of thermal helix inversion.25 It was envisioned that by

introducing an electron-withdrawing ketone moiety in the lower half and an electron-donating

amine moiety into the upper half the push-pull effect could be exploited to elongate the

central carbon-carbon bond. These modifications had the desired effect generating the fastest

motor at the time of its publication (t1/2 = 40 s). Contraction of the ring on the upper half (21)

resulted in significant reduction in the overall rate of rotation producing a motor capable

rotation in the MHz scale.26

23

Figure 2: Structural modification of the second generation molecular motor.24–26

Table 1: Half-life measurements for thermal helix inversion step of various second generation molecular motors.

Motor X Y R t1/2 (20 °C) h

13 S S OMe 184 14 S S H 215 15 S O H 26.3 16 S C(CH3)2 H 233 17 CH2 S H 0.67 18 CH2 C(CH3)2 H 2.01 19 CH2 CH=CH H 60.1 20 NBoc C=O H 40 s 21 Deletion S H 19 ns

In continuing efforts towards designing faster molecular motors, a number of structural

changes were made and interesting structure-rate trends emerged. For example, exchanging

the six-membered ring in the upper half of the molecule for a five-membered ring dramatically

increased the rate of rotation (22 vs 23).27 Employing a p-xylyl (28)28 or benzothiophene (27)29

instead of the naphthyl group in the upper half also increased the rotation (Figure 3).

24

Figure 3: Various second generation molecular motors and their rotation rates expressed in terms of half-life of the

thermal helical inversion at room temperature.27–32

In studying the effects of size of the substituent at the stereogenic centres it was revealed that

increased steric bulk accelerated the rotation.30 DFT calculations were performed on the stable

and unstable isomers of these molecules and revealed that the more sterically demanding

groups have more of an effect on the energy of the unstable form than on the stable form,

thus reducing the barrier to rotation (Figure 4). By combining increased bulk at the stereogenic

centre with a p-xylyl motif on the upper half (two modifications which were both known to be

rate-accelerating, 23 vs 26 and 28 respectively) an acceleration of the rate was observed which

was greater than that obtained when making the modifications individually (29).31

25

Figure 4: DFT optimised energy profiles for the thermal isomerisation of 23 and 26. A greater destabilisation of the

unstable isomer in 26 results in a lower barrier and thus an acceleration in the rate of rotation.

More recently, metal-ligand interactions have also been demonstrated to have an effect on

the photophysical and thermodynamic properties of molecular motors.32 Upon coordination to

a Ru(II)-bipyridine complex, motor 30 was driven by visible light instead of UV, and a large

relative increase in the speed of rotation was observed. The photophysical properties of the

molecular motor were probed with 31—34 where various substituents were located at the 5′

position to be in direct conjugation with the central carbon-carbon double bond.33,34 The

nature of the substituent was shown to alter the photochemical yield of the isomerisation by a

factor of four in the order OMe < H ≈ Cl < CN. By modifying the electronic nature of the

substituent the efficiency of the motors could be improved without affecting the speed of

rotation.

Figure 5: Second generation motor designed to probe the photoisomerisation step.34

Table 2: Quantum yield for the photoisomerisation step of 31—34.34

Motor Quantum Yield

31 0.048 32 0.14 33 0.15 34 0.2

26

A key characteristic of ATP synthase and the bacteria flagellar motor is the ability to reverse

the direction of rotation. In ATP synthase, the hydrolysis of ATP drives the motor in one

direction, while ATP synthesis reverses the direction of rotation.2 In the bacteria flagellar

motor the direction of rotation is thought to be controlled by conformational changes in the

rotor-mounted protein assembly termed the “switch complex”.35,36 This ability raises the

question of whether control over the direction of rotation is also possible in artificial molecular

motors. Since the direction of rotation in the previously discussed molecular motors is strongly

governed by the stereochemistry of the methyl substituent, it was postulated that by changing

the configuration of this centre during the operation the direction of rotation could be

reversed. It was anticipated this could be achieved through base-catalysed epimerisation of a

suitable substituent at the stereogenic centre. To promote the epimerisation it was necessary

to increase the acidity of the relevant proton and so the usual methyl motif was replaced with

an electron-withdrawing amide group. To avoid an unwanted allylic proton shift the

stereogenic centre was moved from the 2′ to the 3′ position. With these design considerations

in mind, molecular motor 35 was prepared (Scheme 6).37

Scheme 6: Controlled clockwise and anti- clockwise motion of rotary motor 35.37

Clockwise motion can be initiated by photo-induced isomerisation of stable (3′S)-(M)-35 to

unstable (3′S)-(P)-35 which then undergoes thermal helix inversion to reform stable (3′S)-(M)-

27

35. Repeating these steps would result in full 360° clockwise rotation. The anti-clockwise cycle

starts with stable (3′R)-(P)-35 which can be obtained by base-catalysed epimerisation of

unstable (3′S)-(P)-35. Anti-clockwise motion now proceeds in the same fashion as clockwise

motion by cycling between photo-induced isomerisation and thermal helix inversions. The

ability to change the directionality of synthetic molecular motors is a significant development

towards their use in functional systems.

1.4.1. Applications

Overcrowded alkene motors possess distinctive features which may be exploited for a variety

of applications. They are chiral and their rotation proceeds in only one direction, switching

between four distinctive intermediate states in the process of completing one rotation.

Furthermore, under continuous irradiation and heating they are capable of continuous

rotation making them suitable for repetitive or cumulative work. To date there have been

many successful examples which rely on these molecules’ ability to act as four-state molecular

switches but only a couple which exploit their capability of cumulative work.

1.4.1.1. From relative to absolute rotation

All of the systems discussed above operate in solution and are constantly buffeted by

Brownian motion making it nearly impossible to harness any useful work. A further challenge is

to make an ensemble of molecules act in concert to transfer the effect of directed motion of a

single molecule on the nanoscale to the micro- or macroscale. Immobilisation of molecular

motors on a surface is a potential solution to convert relative rotation of the rotor to the stator

to absolute rotation relative to a surface. To this end, various molecular motors have been

attached covalently to gold and quartz surfaces through thiol (e.g. 36, Figure 6),38–40 amide,41

silane bonds,42 azide-alkyne Huisgen cycloadditions43,44 and more recently through

electrostatic interactions.45

28

Figure 6: Second generation molecular motor 36 azimuthally immobilised on a gold nanoparticle.38

Two types of attachment mode can be distinguished: azimuthal, with the rotation parallel to

the surface, or altitudinal, with the rotation perpendicular to the surface. The latter was

exploited to reversibly alter the wettability of a surface by presenting or retracting a

hydrophobic perfluorobutyl moiety upon switching between the trans and cis configurations

37. (Figure 7).46,47

29

Figure 7: Top: Altitudinal immobilisation of molecular motor 37 onto a gold film in a) trans and b) cis configuration.

Bottom: Water droplet on trans-37 SAM (left) and cis-37 SAM (right). Upon irradiation with UV light the contact

angle changes from 60° to 75° and 82° to 68° for trans and cis respectively. Adapted from reference47: © American

Chemical Society 2014.

1.4.1.2. Switching chirality

A major development was the demonstration that the switchable helicity of the molecular

motors can be exploited in a number of ways. The photoswitchable helicity of molecular

motors has been shown to play a key role in the enantioselectivity of reactions.48 38 features

thiourea and DMAP motifs, which when in close proximity are known to catalyse the 1,4-

addition of thiophenols to cyclohexenones. Using UV light the catalyst can be switched

between its ON or OFF states (cis and trans respectively), and control over the thermal helix

inversion allows the chirality of product to be selected, generating the (S) enantiomer of the

product in 50% ee with the M-helix catalyst and the (R) enantiomer in 54% ee with the P-helix

catalyst (Figure 8). This catalyst marked the first example of being able to obtain either the

racemate or each individual enantiomer of a product. This system has been extended to the

Henry reaction49 and phosphine-ligand Pd catalysis.50

38

30

Figure 8: Illustration of integrated molecular motor and bifunctional catalyst 38 (top) and molecular structure

(bottom). Adapted from reference48: © American Association for the Advancement of Science 2011.

Switching the helicity of molecular motors has also been shown to be capable of creating an

effect at the supramolecular and even microscopic level. By attaching molecular motor 39 to

the terminus of a poly(isocyanate) chain, which normally exists as a racemic mixture of two

helical conformations, the polymer conformation could be switched between a racemic state,

P helicity or M helicity, as determined by low-temperature circular dichroism (CD)

spectroscopy, Figure 9.51,52

Figure 9: Polymer-modified molecular motor 39 and illustration of the reversible inversion of helicity of a polymer

by a light-powered molecular motor. Adapted from reference51: © John Wiley & Sons, Inc. 2007.

Molecular motors are also excellent dopants for a variety of liquid crystal (LC) phases,

converting the nematic phase to a cholesteric phase. The change in helicity of the molecular

motor 40 during the rotary process induces a reorganisation of the LC film which in turn causes

a reversible colour change in the material (Figure 10).53

31

Figure 10: Molecular motor 40 and the colour changes of a LC film doped with 40. Adapted from reference53: ©

John Wiley & Sons, Inc. 2006.

More remarkably, it was found that the change in helicity of molecular motors could be

amplified from the molecular level to the microscale level by the LC matrix (Figure 11). Upon

photochemical and thermal isomerisation the helicity of dopant 40 could be switched, which in

turn could switch the chirality of the cholesteric phase. As a result, irradiation of an LC film

doped with molecular motor 40 led to unidirectional rotation of a microscopic glass rod placed

on top of the film.54,55 In this way, through harnessing energy from light and by acting

synchronously, molecular motor 40 is capable of rotating an object thousands of times larger

than itself! However, it should be noted that when the photostationary state is reached, the

rotation halts. Subsequent thermal isomerisation of the motor dopant results in rotation of the

glass rod in the opposite direction. It is the switching helical chirality of the motor molecules

used as dopant which is reflected in the rotation of the glass rod, not the unidirectional

character of the rotation.

32

Figure 11: Pictures of an LC film doped with motor 40 (1 wt%) and a glass rod deposited on top of it, taken at 15 s

intervals after UV irradiation commenced. Scale bar = 50 µm. Adapted from reference54: © Nature Publishing Group

2006.

1.4.1.3. Cumulative work

Recently, the first example in which the unidirectional molecular motor is truly applied was

reported.56 Molecule 41 consists of four unidirectional motors which in response to electronic

and vibrational excitation act as paddlewheels to propel the molecule across a copper surface

(Figure 12). After sublimation onto a Cu(111) surface the molecular “nanocars” were visualised

with a scanning tunnel microscope (STM). A voltage pulse is then applied to the “nanocars”

using the STM tip, which induces conformational changes in the motor units. In the meso-(R,S-

R,S) isomer the four motor units act in unison and these conformational changes result in

conrotatory motion, propelling the molecule along the surface in a linear fashion which was

visualised by STM imaging. In contrast, the (R,R-R,R) or (S,S-S,S) isomer produces disrotatory

motion of the four paddlewheels, resulting in the molecule spinning or moving randomly.

33

Figure 12: a) Structure and illustration of the meso-(R,S-R,S) isomer. b) Chemical structure of the motor unit. c) 360°

rotation of the motor unit. d) Schematic representation of STM tip excitation of the molecule. e) Molecular model

representation of the paddlewheel-like motion. Adapted from reference56: © Nature Publishing Group 2011.

Molecule 41 synchronised the motion of molecular motors to produce directed motion at the

molecular level and although this was a truly remarkable achievement, the key challenge of

coordinating multiple molecular motors to produce an effect at the macroscale remained.

Recently, researchers led by Nicolas Guiseppone incorporated a molecular motor into a

polyethylene glycol (PEG) gel which contracts in response to UV irradiation.57 By integrating

unidirectional molecular motor 42 as reticulating points in their gel it was possible to amplify

the motion at the molecular level to the macroscopic level (Figure 13). 42 comprises a second

generation overcrowded alkene molecular motor functionalised with azide- or alkyne-

terminated PEG chains on their upper and lower part respectively. Under concentrated

conditions, copper-catalysed Huisgen[3+2] cycloaddition leads to a crosslinked gel with

molecular motors as the reticulation units. When irradiated with UV light the molecular

34

motors in the gel rotate unidirectionally and twist the entangled polymer strands resulting in

macroscopic contraction of the gel by approximately 80%.

Figure 13: a) Polymer motor conjugate 42. Bottom: b) Cartoon representation of crosslinked polymer-motor

conjugate. Upon irradiation with UV light the polymer chains coil up and reduce the entire size of the polymer

network. d) Pictures showing time-dependent contraction of gel. Adapted from reference57: © Nature Publishing

Group 2015.

1.5. Unidirectional Rotation Around C=N Imine Bonds

Moving from rotation around C=C double bonds to rotation around C=N double bonds, Lehn

and co-workers developed an imine-based, light-driven molecular motor.58 By exploiting a

combination of photo-induced E/Z isomerisation with two orthogonal thermal isomerisation

processes it was possible to achieve 360° rotation around the central C=N double bond

(Scheme 7).59 Irradiation of 43 with UV light induces out-of-plane rotation around the central

imine bond. This is followed by thermal isomerisation which occurs through ring inversion (RI)

of the cycloheptatriene stator moiety. Further photochemical then thermal isomerisation

restores the molecule to its original conformation. By decreasing the conformational flexibility

35

of the stator (44) the mechanism of thermal isomerisation now favours an in-plane nitrogen

(NI) pathway. Crucially, both RI and NI pathways are orthogonal to the photochemical out-of-

plane isomerisation. Given the ease of synthesis of these motors, and the possibility of

combining motional dynamics with constitutional dynamics, imine-based molecular motors

present a valuable addition to the toolbox of molecular motors.

Scheme 7: a) Photo and thermal isomerisation of 43, which proceeds through out of plane rotation then ring

inversion. b) Photo and thermal isomerisation of 44 which proceeds through out of plane rotation then nitrogen

inversion.

A system published by Haberhauer demonstrates directional rotation of a molecular blade

through a cycle of trans/cis isomerisations of an azobenzene motif and metal

coordination/demetallation (Figure 14).60 In the starting state trans-(P)-45 has a N-C-C-N

dihedral angle of approximately 180° and a trans/cis ratio of azobenzene isomers of 80:20.

Irradiation of this species with UV light causes the azobenzene moiety to reach a new

photostationary state of 42:58 trans/cis. Addition of Zn2+ ions causes a movement of the blade

in the form of 180° rotation around the bipyridine C-C bond. Computational analysis on a

model compound reveals that steric bulk on one side of the molecule presents an

insurmountable barrier to rotation in that direction, rendering this step essentially

unidirectional.

36

Figure 14: A molecular four-stroke motor. Chemical structures and schematic illustration of each state of the

operation. Adapted from reference60: © John Wiley and Sons, Inc. 2011.

The initial trans/cis ratio is then restored by irradiating the resulting complex cis-(M)-45-Zn2

with visible light. Removal of the Zn2+ ions by addition of a competing ligand restores the

molecule to its original state having rotated the blade through a full 360° turn. By alternating

between the switching processes in this way, the blade moves through four different positions,

returning to its starting point via a different path from which it left.

37

1.6. Unidirectional Rotation in Interlocked Architectures

Catenanes are molecules in which two or more rings are mechanically interlocked, an

architecture which restricts certain degrees of freedom but still allows the components to

move with large amplitude in a motion termed pirouetting (Figure 16). The relative positions

and orientations of the rings can be biased by use of an external stimulus – for example

solvent61, pH62, electrochemistry63–68, or metal coordination69. This control of relative

submolecular motion makes this class of compounds excellent systems with which to study

and develop molecular machines.

Figure 16: A [2]catenane exhibiting large amplitundinal motion in a pirouetting fashion as shown by the arrows.

The key to understand how a catenane can be considered as a molecular motor is not to

consider the whole structure as the molecular machine, but rather to consider one ring (the

motor) which directionally transports the other ring (the substrate) around itself. The first

example of stimuli-driven sequential rotation in a catenane was demonstrated with 46 (Figure

17).70 Three stations were incorporated into the larger macrocycle: fumaramide (A, green),

tertiary fumaramide (B, red) and succinamide (C, orange), each of which has a different

binding affinity for the smaller benzylic amide macrocycle. Isomerisation of fumaramide

stations A and B to their maleamide counterparts disrupts the hydrogen bonding and

drastically changes their binding affinities. Fumaramide station A is located next to a

benzophenone unit allowing selective photosensitized isomerisation at 350 nm before

38

photoisomerisation of tertiary fumaramide B at 254 nm. In its initial state the binding affinity

of the stations to the small macrocycle is in the order A>B>C and consequently the macrocycle

predominantly resides over station A. Photosensitized isomerisation of this station (350 nm,

green → dark green) changes the relative binding affinities to B>C>A and the macrocycle now

predominantly resides over station B. Subsequent photoisomerisation of this station (254 nm,

red → pink) results in macrocycle relocation to station C, with the station binding affinities

now in the order C>B≈A. Reisomerisation of station A and B back to their fumaramide forms

reinstates the initial station binding affinities and the macrocycle returns to its original position

over station A. Although 1H NMR revealed excellent positional integrity at each state, the

rotation of the smaller benzylic amide macrocycle around the larger one is not directional —

an equal number of macrocycles will travel from A to B and C clockwise as travel anticlockwise.

Figure 17: The small macrocycle in [2]catenane 46 can move between the three stations with high fidelity, but with

no preferred direction.70

To bias the direction in which the smaller macrocycle travels, barriers must exist to restrict

Brownian motion to a particular direction. However, to allow complete circumrotation these

barriers must be transient. Such a situation is inherently present in [3]catenane 47 which

differs from 46 only by the addition of a second benzylic amide macrocycle (dark blue, Figure

39

18).70 The amide group now acts as a fourth binding station, D. In its initial state the binding

affinity of the stations to the small macrocycles is in the order A>B>C>D and consequently the

macrocycles predominantly reside over stations A and B. Irradiation at 350 nm induces

translocation of the blue macrocycle to succinamide station C. Crucially, the presence of the

second macrocycle forces this translocation to occur anti-clockwise (as drawn in Figure 18).

Subsequent isomerisation of the tertiary fumaramide station at 254 nm causes the dark blue

macrocycle to shuttle to the amide station D. Likewise, to reach its destination, translocation

of the dark blue ring can only proceed in the anti-clockwise direction. Thermal re-isomerisation

restores the original binding order A>B>C>D and regenerates (E,E-47) but with the positions of

the macrocycles swapped. Hence, this “follow-the-leader” process in which each small ring

controls the direction of circumrotation of the other is used to directionally move the smaller

rings around the larger one. The synthetic sequence must be completed twice for full

unidirectional rotation of both small macrocycles to occur. This system is particularly elegant

since it is self-compartmentalising.

40

Figure 18: Directional circumrotation of two rings in [3]catenane 47.70

Selective rotation in either direction was achieved with [2]catenane 48 (Figure 19) which

consists of a small benzylic amide macrocycle interlocked with a large macrocycle bearing a

fumaramide residue, and a succinamide residue flanked by two different bulky substituents: a

silyl protected hydroxyl and a trityl protected hydroxyl.71 The direction the small macrocycle

travels is determined by the order in which the bulky blocking groups are removed.

Isomerisation of fum-(E)-48 to mal-(Z)-48 significantly disrupts the binding and creates a

thermodynamic driving force for the macrocycle to shuttle to the succinamide station.

Removal of one of the protecting groups (silyl for clockwise, trityl for anti-clockwise) results in

directional motion of the small macrocycle to the now preferred succinamide station.

Appropriate reprotection traps the macrocycle in the succinamide compartment. The

41

maleamide station can be re-isomerised back to fumaramide thus restoring the

thermodynamic preference for the macrocycle to shuttle back to the fumaramide station.

Removal of the appropriate hydroxyl protecting group results in full 360° rotation of the small

macrocycle around the larger one.

Figure 19: Reversible 360° rotation in [2]catenane 48.71

In 2013 a catenane motor which used single stranded DNA as its construction material was

reported (Figure 20).72 This [2]catenane acted as a rotary motor by moving between three

defined states in response to different stimuli. The formation or disassembly of an i-motif in

response to pH acted as the first stimuli, while addition of Hg2+/cysteine switched Hg2+-bridge-

assisted base pairing on and off. By choosing the order in which to apply the stimuli, the

system could be driven in a clockwise or anticlockwise fashion with high yield and selectivity as

evidenced by a series of fluorescence experiments.

42

Figure 20: Reversible, stimuli-induced motion of ring α across three stations I, II and III. Adapted from reference72:

© American Chemical Society 2013.

1.7. Conclusion

Since the discovery of the first synthetic rotary motor there have been significant

developments in the field and scientists now have a toolbox filled with a variety of genuine

motors. Although these motors are primitive by Nature’s standards the basic principles of

rotary motion have been demonstrated and the first examples exploiting their properties are

emerging. There have been several successful applications of the class of overcrowded alkene

molecular motors, mainly exploiting their ability to act as four-state switches but arguably the

most important advance of rotary motors was the development of examples that could

perform work and drive a system out of thermodynamic equilibrium to produce directed

motion across a surface or which impressively transfer a conformational change at the

molecular scale to a visible change at the macroscale. It is quite remarkable that by

coordinating motion at the nanoscale it is possible for these motors to effect the microscale

and even the macroscopic scale. An as-yet unmet challenge for rotary motors is a chemically-

driven system that can operate autonomously, that is rotate directionally so long as fuel is

present, as is the case with Nature’s motors. The field of synthetic rotary motors is still in its

infancy, and only imagination and time will reveal the true potential of these motors.

43

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72 C.-H. Lu, A. Cecconello, J. Elbaz, A. Credi, I. Willner, Nano Lett. 2013, 13, 2303–2308.

Chapter Two

A [2]rotaxane Molecular Information Ratchet

Compound 77 has been reported in the thesis of Miriam Wilson in support of an application for

the degree of Master of Chemistry with Industrial Experience awarded by The University of

Edinburgh, 2010. Its synthesis is shown here for clarity only.

Acknowledgements

Dr Armando Carlone, Dr Stephen Goldup, Dr Natalie Lebrasseur and Dr Jordi Solà are gratefully

acknowledged for their intellectual and synthetic contribution to this chapter.

Dr Guillaume De Bo, Dr Matthew O. Kitching, Dr Thomas A. Singleton, Dr Daniel Tetlow and Dr

John W. Ward are gratefully acknowledged for examining and proofreading this chapter.

49

2.1. Synopsis

This Chapter describes the synthesis and operation of a chemically-driven, autonomous,

[2]rotaxane molecular information ratchet. Steric information was revealed to be the driving

force of the information ratchet with acylation reactions taking place preferentially when the

macrocycle is far away from the reactive site. It is hoped these results will prove useful in

designing an autonomous, chemically-fuelled information ratchet which operates over greater

distances and a higher number of compartments or in a rotary fashion by incorporation into a

catenane architecture.

50

2.2. Introduction

In the field of molecular machines, it is important to distinguish between molecular switches

and molecular motors. When a switch is returned to its original state, any mechanical work

which has been performed is undone. In contrast, when a motor returns to its original state

the work performed by the motor is not undone: it can drive a system away from equilibrium.

The ability to control the motion of components within interlocked structures unlocks the

possibility for them to act as ratchets – an essential feature of any machine more complex than

a simple switch. Brownian ratchets fall into two main classes: energy ratchets and information

ratchets. A classic example of an energy ratchet is a flashing ratchet, which forms the basis of

[2]catenane rotary motor 48 (see section 1.6), and is illustrated in Figure 21. It comprises an

asymmetric potential energy surface, consisting of a periodic series of energy maxima and

minima. Sequentially raising and lowering each set of energy minima and maxima directionally

transports a Brownian particle along the surface. The particle starts in a green or yellow well

(Figure 21, a or c respectively). Raising that energy well while simultaneously lowering the

adjacent minima provides the driving force for the particle to move position by Brownian

motion (Figure 21, b to c or d to e). By repeatedly varying the relative heights of energy

barriers and minima of the energy wells in this way the particle can be directionally

transported along the potential energy surface.

51

Figure 21: A flashing ratchet. In (a) and (c) the particle starts in a green or orange well respectively. Raising this

energy minima while simultaneously lowering the adjacent energy maxima provides the driving force for the

particle to move position by Brownian motion. By repeatedly varying the energy barriers in this way the particle can

be directionally transported. Reproduced from reference1 in agreement with a Creative Commons Attribution (CC-

BY) License.

Note that the modulation of the energy surface proceeds irrespective of the particle’s position.

However, in the case of an information ratchet the potential energy surface does respond to

the position of the particle on the surface. In this case, directional motion is achieved by

selectively lowering kinetic barriers to transport in front of the particle, or by selectively raising

barriers to transport behind the particle (Figure 22).

Figure 22: An information ratchet. In (a) and (d) the dashed lines indicate transfer of information about the location

of the particle. (b) The position of the particle selectively lowers the energy maxima to the right, but not to the left.

(c) The particle moves by Brownian motion. Reproduced from reference1 in agreement with a Creative Commons

Attribution (CC-BY) License.

52

This requires information to be passed between the particle and the potential energy surface.

Such information can come from asymmetry in the distance of the particle between two

barriers, or from a well-established form of chemical information: chirality. This latter

technique was exploited in the chemically-fuelled information ratchet 49 (Figure 23).2

Figure 23. Information ratchet 49 shown in its hydroxyl deprotected form.

Rotaxane 49 consists of two energetically equivalent fumaramide stations, evenly spaced

around a prochiral hydroxyl group. Although the bare thread is achiral (discounting the

deuterium labels), presence of the macrocycle in the rotaxane form lends it chirality, and the

two positional co-conformers are in fact enantiomers. In the free hydroxyl form of the

rotaxane the macrocycle freely shuttles between the two isoenergetic fumaramide stations:

the enantiomers are in an equilibrium ratio of 50:50 FumH2:FumD2. This equilibrium makes 49

a perfect candidate for dynamic kinetic resolution: where two interconverting enantiomers

have different transition states in the presence of a chiral catalyst and the lower transition

state becomes the kinetically favoured pathway (Figure 24).

53

Figure 24. Energy diagram of an (R) and (S) isomer. In the presence of a chiral catalyst one transition state is lower

and becomes the kinetically favoured pathway.

Acylation of the hydroxyl group of 49 creates a kinetic barrier to shuttling and traps the

macrocycle in either compartment. An achiral acylation catalyst results in an equilibrium

distribution of 50:50 FumH2:FumD2. However, a chiral DMAP-based acylation catalyst was

shown to drive the distribution away from equilibrium to an unequal distribution of 33:66

FumH2:FumD2. Use of the chiral catalyst’s enantiomer gave the equal and opposite ratio of

66:33 FumH2:FumD2. This directional transport results from the ability of the chiral acylation

catalyst to use information on the position of the macrocycle to preferentially raise the kinetic

barrier to shuttling behind the macrocycle. In this case, the potential energy of the

fumaramide stations or the macrocycle remains unchanged throughout the process (Scheme

8).

54

Conditions Catalyst Product Distribution (±3%)

FumH2-52: FumD2-52

(a) DMAP 50:50 (b) (S)-51 33:67 (c) (R)-51 67:33

Scheme 8. Molecular information ratchet 49 and its distribution after acylation under different conditions: a) with

an achiral catalyst 2, b) with chiral catalyst (S)-51, c) with the catalyst’s enantiomer, (R)-51.

This concept has been further developed to incorporate multiple ratchet sites into a one-

dimensional track to allow cumulative, directional transport (Scheme 9)3.

55

Conditions Catalyst Product Distribution (±3%) left-56:centre-56:right-56

(a) (S)-54 <1:21:79 (b) (R)-54 75:25:<1 (c) DMAP 39:18:43 (d) 50:50 (S)/(R)-54 10:77:13

Scheme 9. Molecular information ratchet 53 and its distribution after acylation under different conditions: a) with a

chiral catalyst, (S)-54, b) with the catalyst’s enantiomer, (R)-54, c) with an achiral catalyst, d) with a racemic mixture

of the chiral catalyst.

Rotaxane 53 was capable of transporting the macrocycle with high efficiency, with >75%

reaching the desired terminal station.

56

2.3. Design

The aim of this project is to design, synthesise and operate a two station [2]catenane which

will operate as a unidirectional chemically-driven autonomous rotary motor. A smaller

macrocycle would circumrotate around a larger one autonomously and continuously without

outside intervention. The machine would use chemical energy to directionally transport the

macrocycle from one compartment to the next, using the position of the macrocycle to effect

useful mechanical work. One can envisage incorporation of an information ratchet core into a

catenane architecture would help achieve the goal of an autonomous, chemically-fuelled,

directional rotary motor. Thus, taking rotaxane 49 and linking the two ends together would

result in a [2]catenane capable of operating under the same chemical information ratchet

principle. This would result in the [2]catenane 57, depicted in Figure 25. At the same time the

alternative [2]catenane 58 was proposed. Both designs are based on a heterocircuit

[2]catenane, two mechanically interlocked molecular rings of different sizes. Fumaramide

residues (shown in green) on the larger ring (the ‘track’) serve as binding sites for a smaller

benzylic amide macrocycle (blue). Hydroxyls protected with bulky protecting groups (red)

block passage of the small ring and, when both are attached, trap it in one compartment of the

cyclic track or the other. In the first design, 57, opening and closing of the top gate using DKR

conditions should result in directional motion of the macrocycle. However, when operating the

bottom gate, the macrocycle will be too far away to create a well-defined chiral environment,

and the gate will close indiscriminately thus reducing the overall directional efficiency of the

motor. In comparison, the directional bias for 58 would not rely on DKR and a chiral catalyst,

but instead a bias exists between the two stations already owing to the asymmetry in distance

between the stations and a gate. The difference in the distance of the fumaramide groups to

each hydroxyl group (one very close, where the presence of the macrocycle should inhibit

57

nucleophilic attack by the OH group on an acylating agent, and one too far away for a bound

macrocycle to significantly influence rates of reaction) should result in dissimilar reaction rates

for when the macrocycle is close to (kclose) or far from (kfar) the hydroxyl group and create a

directional bias for the small macrocycle around the large one. Furthermore, 58 has an

identical station-gate arrangement at both gates therefore directional motion will be more

efficient. For this reason it was decided to pursue the design shown in Figure 25b.

Figure 25. Proposed [2]catenane designs 57 and 58. Ignoring the deuterium label and the presence of the blue

macrocycle, 57 has C2v symmetry and 58 has C2h symmetry.

[2]Catenane 58 bears similarities to the two-compartment information ratchet 49 (see Figure

23). Fumaramide residues serve as binding sites for an amide macrocycle. The pre-organised

58

hydrogen bond network offered by the fumaramide residue is also useful as a template for the

formation of the macrocycle around the thread. Development of the original chemically-driven

information ratchets into an autonomous system would require the ratchet mechanism to

operate continuously, meaning that the barriers must be repeatedly raised and lowered under

a single set of reaction conditions. For this goal to be achieved a blocking group that attaches

and detaches through dissimilar reaction mechanisms must be used, with one reaction (e.g.

attachment) proceeding at rates that vary according to the position of the small macrocycle,

the other (e.g. detachment) occurring at a rate independent of the small macrocycle position.

The first law of thermodynamics dictates that the total energy of an isolated system is

constant; energy can be transformed from one form to another, but cannot be created or

destroyed. So for work to be done (macrocycle moving into a higher energy distribution) while

the benzoyl group is deprotected and reprotected energy must be transferred from some

species to the machine. Therefore the deprotection and reprotection cannot exist in

equilibrium. A fuel must be consumed and a by-product generated. With this in mind

analogues of the benzoyl protecting group were investigated and it was found that the SiOMB

group 60 (Scheme 10) was shown by Morvan to be cleaved by a mild fluoride source to liberate

the alcohol and phthalide waste product.4 If the SiOMB were to be activated as its fluoride

ester 60 then the liberated fluoride could subsequently cleave the silyl ether 61. In this elegant

approach, a constant supply of SiOMB fluoride could be used to repeatedly protect and

deprotect the hydroxyl moiety.

59

Scheme 10. Protection of a secondary alcohol with SiOMB activated as a fluoride ester and subsequent

deprotection mediated by fluoride anion.

Prior to my arrival in the group exhaustive studies revealed that the SiOMB protecting group

and other derivatives were unsuitable so an alternative was sought. Various acyl protecting

groups were investigated before 9-Fluorenylmethoxycarbonyl chloride (Fmoc-Cl) was chosen

as the chemical fuel. The Fmoc protecting group is more traditionally used as an N-protecting

group in solid phase peptide synthesis; however it has also been employed to some extent for

the protection of alcohols in carbohydrate chemistry.5–7 Its potential mechanism of attachment

to the molecular motor is significantly different to that of cleavage of the resulting

fluorenylmethoxycarbonate group (Scheme 11). The former occurs by nucleophilic attack of a

catenane hydroxyl group directly on the C=O of the chloroformate activated with an acylation

catalyst such as DMAP (66), where the presence or absence of the bulky benzylic amide

macrocycle on the adjacent fumaramide group would be expected influence the reaction rate.

In contrast, the detachment reaction occurs by a reaction cascade (eliminating CO2 and

dibenzofulvene as waste products) initiated by base abstraction of a proton from the fluorenyl

methine group, five bonds remote from the site of attachment to the [2]catenane and so the

influence of the position of the macrocycle on the reaction rate should be minimal.

60

Scheme 11. Fmoc protection of a secondary alcohol and subsequent base-catalysed deprotection.

In the Fmoc protected form of 58 the macrocycle is unable to pass over the steric barrier of the

Fmoc group, leaving it trapped in one of two compartments: top or bottom. Double Fmoc

cleavage of 58 would lead to the diol form leaving the macrocycle free to shuttle over the full

length of the catenane. Owing to the inherent symmetry of the molecule the two positional

isomers top and bottom are chemically indistinguishable, therefore a non-chromatographic

technique had to be found which would enable one to determine the ratio of the isomers in

the mixture. As was demonstrated with rotaxane 49, fumaramide protons residing inside the

cavity of a benzylic amide macrocycle experience a significant upfield shift in the 1H NMR

spectrum. Integration of the shielded fumaramide protons, in reference to an internal

standard in the molecule, can be used to calculate the ratio of macrocycle over the two

stations. To make this analysis possible one of the stations must be deuterium-labelled. The

two stations will now be described as FumH2 and FumD2 (Figure 26).

61

Figure 26. FumH2 and FumD2 positional isomers of [2]catenane 58.

Other than the nature of the interlocked architecture employed, one key structural difference

between rotaxanes 49, 53 and the proposed catenane motor 58 is the arrangement of the two

fumaramide stations around the hydroxyl moiety. In 58, the macrocycle creates a high steric

demand around the top hydroxyl group when residing over the FumH2 station, and a low

demand in the FumD2 isomer. It was postulated that this asymmetry in the station-gate-station

arrangement would result in a directional bias governed by sterics, rather than chirality in the

case of 49 and 53. A further development is that for the system to be described as

autonomous, deprotection and reprotection of the hydroxyl must occur in the same reaction

vessel through a non-equilibrium pathway.

The proposed autonomous motor is based on a [2]catenane structure, the synthesis of which is

non-trivial and time consuming. It was anticipated that a [2]rotaxane could be used as a model

system as it would be more easily synthesised and would allow more straightforward analysis

(Figure 27).

62

Figure 27. The desired [2]catenane 58 and a [2]rotaxane model 69, with their respective cartoon representations.

We envisaged the chosen [2]rotaxane 69 would be a good model for the proposed [2]catenane

autonomous motor 58 as it has a very similar station-to-gate arrangement, conserving the

distance between the station, gate and next station.

63

2.4. Retrosynthesis

The target [2]rotaxane 69 and its retrosynthesis is shown in Scheme 12. Building block 72 is

known in the literature3, and is prepared in 9 steps from commercially available (R)-3-amino-

1,2-propanediol, mono-ethyl fumarate, and N-benzylaniline. Deuterium-labelled fumaramide

building block 75 has been reported previously,2 and is prepared from dimethyl

acetylenedicarboxylate and N-benzylaniline. Rotaxane 70 is prepared using a five-component

clipping procedure from thread 71.8,9 The target molecules would be assembled using peptide

coupling procedures. Amines would be protected as tert-butylcarbamates, and acids protected

as their methyl or ethyl esters. The free amines would be liberated by TFA-mediated

deprotection and the acids by saponification with lithium hydroxide as needed.

64

Scheme 12. Retrosynthesis of [2]rotaxane 69.

2.5. Synthesis

Amine 76 was prepared from 1-aminoundecanoic acid by thionyl chloride-assisted ester

formation. Peptide coupling of this unit with acid 75 yielded 73 in 69% yield after column

chromatography. Saponification with lithium hydroxide in a THF/water mixture proceeded

65

quantitatively to yield the right hand portion 77, which was used without further purification

(Scheme 13).10

Scheme 13. (i) SOCl2, MeOH, r.t. to reflux, 4 h, quant.; (ii) DIPEA, TBTU, HOBt, DMF, 0 °C to rt, 18 h, 69%; (iii)

LiOH·H2O, THF:H2O 3:1, r.t., 21 h, quant.

Following a modified literature procedure3 72 was treated with TBAF to cleave the TBDPS

protecting group. A sodium citrate work-up removed any tetrabutylammonium salts and

column chromatography furnished 78 in 95% yield. Reprotection of the hydroxyl group with 9-

fluorenylmethyl chloroformate in the presence of DMAP and pyridine gave 71, an ideal

template for rotaxane formation using the clipping method.8 This is a five-component reaction

in which four amide bonds are formed. At high dilution, thread 71 was allowed to react with p-

xylylenediamine and isophthaloyl chloride which were added separately by syringe pumps.

High dilution and slow addition were used in order to minimise the formation of polypeptide

oligomers. The hydrogen bonding preorganisation offered by the fumaramide station

encourages the components to react together and form a macrocycle around the thread

yielding a [2]rotaxane.9 The steric bulk of the N-benzylaniline group on one side and the Fmoc

group on the other prevents the macrocycle from dethreading. Careful control of the amount

66

of triethylamine added, (which can cleave the Fmoc protecting group), allows good yields to be

achieved (Scheme 14).

Scheme 14. (i) TBAF, THF, r.t., 18 h, 95%; (ii) Fmoc-Cl, DMAP, pyridine, CH2Cl2, 18 h, 73%; (iii) isophthaloyl chloride,

p-xylylene diamine, NEt3, CHCl3, 3 h, 75%.

In the 1H NMR spectrum of 70 there is a significant upfield shift of the fumaramide protons

(H11 + H12) compared to those in non-interlocked thread 71 owing to shielding by the

macrocycle (ΔδH11 = -1.11 ppm, ΔδH12 -1.25 = ppm). The downfield shift of amide NH proton

(ΔδH14= 1.24 ppm) is a result of hydrogen bonding between the amide macrocycle and the

fumaramide carbonyls. Furthermore, the presence of the macrocycle makes the diasterotopic

nature of Fmoc protons H23, H26 and H29 more pronounced.

67

Figure 28. Partial 1H NMR spectra (600 MHz, CD2Cl2, 300 K) of (a) thread 71 and (b) rotaxane 70. Residual solvent

peaks are shown in grey. The lettering corresponds to proton labelling in Scheme 14. (c) DFT optimized geometry of

molecule 70 showing the distance between H9 and the macrocycle (3.5 Å).

A further significant difference is the large upfield shift of H9 (Δδ = -0.91 ppm). DFT calculations

revealed H9 was in close proximity to an aryl ring from the macrocycle (Figure 28 c), and likely

experiencing shielding from the π-cloud.

The final step in the synthesis was the assembly of the final rotaxane. The BOC group of

rotaxane 70 was cleaved with TFA and the resulting primary ammonium TFA salt was coupled

with acid 77 using peptide coupling conditions to give 69 in excellent yield. Owing to the

synthetic route chosen, rotaxane 69 had a starting macrocycle distribution of 100:0

FumH2:FumD2.

68

Scheme 15. (i) a) 70, TFA, CH2Cl2, r.t., 20 h; b) 77, DIPEA, TBTU, HOBt, DMF, r.t., 18 h, 87%.

2.6. Operation and Analysis

2.6.1. Directionality

To measure the efficacy of information ratchet 69, the directional bias upon acylation had to

be determined. Deprotection of 69 with TBAF, gave hydroxyl rotaxane 79; the reprotection of

which could then be studied under various acylating conditions (Scheme 18). The DMAP-type

acylation catalysts previously used by Leigh and co-workers (see Scheme 8 and 9) were found

to be unsuitable – after overnight reactions very little or no product was observed.2,3 DMAP is

known to be a very good acylation catalyst so one possible reason for it not working in this

case is that it was acting as a base and deprotecting any product which may have formed. The

pyridine analogues were found to be more suitable and can be prepared in two steps from

both enantiomers of commercially available α-α-bis[3,5-bis(trifluoromethyl)phenyl]-2-

pyrrolidinemethanol and nicotinoyl chloride hydrochloride (Scheme 16 and 17).

69

Scheme 16: (i) Nicotinoyl chloride hydrochloride, NEt3, CH2Cl2, r.t., 18 h, 71%.

Scheme 17: (i) Nicotinoyl chloride hydrochloride, NEt3, CH2Cl2, r.t., 2 h, 86%.

The typical procedure for Fmoc protection was to dissolve rotaxane 79, acylation catalyst (5

eq.) and Fmoc-Cl (5 eq.) in dry dichloromethane. Once TLC indicated the starting material had

been consumed the reaction was diluted with dichloromethane, washed with 1M HCl (aq.),

and purified by preparative TLC (CH2Cl2:EtOH 95:5) (Scheme 18). The result of acylation was a

mixture of kinetically trapped positional isomers FumH2 and FumD2 the ratio of which was

determined by 1H NMR.

70

Scheme 18. Operation of molecular information ratchet 69. Reaction conditions: (i) TBAF, THF, r.t., 18 h, 95%; (ii)

Fmoc-Cl (5 eq.), (S)-81 (5 eq.) or (R)-81 (5 eq.) or pyridine 82 (5 eq.), CH2Cl2, r.t., 20 h.

Analysis of the acylation products by 1H NMR spectroscopy revealed notable differences when

compared to 69 FumH2:FumD2 100:0 (Figure 29). Firstly, the signals corresponding to shielded

H11 and H12 did not integrate to equal 1. Signals corresponding to the macrocycle protons HD

appeared at two different chemical shifts, indicating the macrocycle was in two different

environments. After full characterisation it was possible to attribute these changes to the

FumD2 positional isomer and determine the ratio of positional isomers simply by integration of

the mixture. The regions from 5.3 to 6.6 and 8.0 to 8.6 ppm were chosen as an analysis

window.

71

Figure 29. Partial 1H NMR spectra (600 MHz, CD3OD:CDCl3 3:1, 300 K) of (a) molecular shuttle 69 and rotaxane 79

reprotected to give 69 in the presence of (b) Py, (c) (R)-81, (d) (S)-81. Residual solvent peaks are shown in grey. The

lettering corresponds to proton labelling in Scheme 14.

The results obtained for the acylation of hydroxyl rotaxane 79 are given in Table 3.

Table 3. Product distribution upon acylation of 79 using various catalysts.

Conditions Catalyst Product Distribution

FumH2:FumD2

(a) Pyridine 39:61 (b) (R)-81 17:83 (c) (S)-81 19:81

Reaction of 79 with Fmoc-Cl in the presence of pyridine resulted in the macrocycle being

kinetically locked predominantly in the FumD2 compartment (40:60). Acylation in the presence

of (R)-81 again locked the macrocycle in the FumD2 compartment, but with a higher directional

bias (17:83). Use of the antipode catalyst (S)-81 gave a similar result. Since both hands of

catalyst 81 gave similar results and the acylation complex clearly reacts preferentially when

72

the macrocycle is far away from the reactive site this reveals that directional movement is

governed by sterics rather than chirality. Evidently the catalyst, in its acylated intermediate

form, is able to discriminate between the two positional isomers which interconvert through

the macrocycle shuttling between the two fumaramide residues, and preferentially acylates

the hydroxyl group when the macrocycle is on the FumD2 group. This directional discrimination

appears to be independent of catalyst handedness and rather stems from the inherent

asymmetry of the thread.

It has been established that the position of the macrocycle can affect the rate of protection,

therefore it is reasonable to postulate that the macrocycle’s position may also influence the

rate of deprotection. If deprotection is more likely to occur on the favoured positional isomer

(in an equal and opposite amount to the bias upon protection) then net motion would not

occur. To investigate this, rotaxane 69 FumH2:FumD2 20:80 was subjected to deprotection

conditions, and the reaction stopped before reaching completion (Scheme 19).

Scheme 19. (i) NEt3 (5 eq.), CH2Cl2, r.t., 5h, 67%.

If the recovered unreacted rotaxane has the same positional ratio as the starting rotaxane

then both positional isomers must have been consumed at the same rate and it can be

concluded that the position of the macrocycle doesn’t influence the rate of deprotection.

When 69 was treated with NEt3 and the unreacted starting material recovered, 1H NMR

73

analysis revealed the ratio of FumH2:FumD2 to be the same as in the starting material rotaxane

(Figure 30).

Figure 30. Partial 1H NMR spectra (600 MHz, CD3OD:CDCl3 3:1, 300 K) of (a) FumH2-69:FumD2-69 20:80 and (b)

FumH2-69:FumD2-69 20:80 recovered from partial deprotection reaction. The integrals for shielded H11 and H12

remain unchanged over the course of the reaction indicating both positional isomers (FumH2-69 and FumD2-69) are

consumed at the same rate. Residual solvent peaks are shown in grey. The lettering corresponds to proton labelling

in Scheme 14. The section 6.6—5.4 ppm has been scaled vertically 3× compared to the section 8.6—8.0 ppm.

This leads to the conclusion that both positional isomers were consumed at the same rate and

therefore the position of macrocycle doesn’t affect the rate of deprotection.

2.6.2. Autonomy

All the experiments described so far have involved stepwise reactions, but as previously

mentioned one of the aims of this project is to operate the motor autonomously.

Development into an autonomous system would require both the protection (Scheme 18) and

deprotection (Scheme 19) steps of the ratchet mechanism to operate in one-pot continuously,

meaning that the barriers must be repeatedly raised and lowered under a single set of reaction

conditions. Or in other words, so long as the fuel source is maintained, the deprotection and

reprotection, and associated macrocycle shuttling, should occur continuously. The following is

a series of experiments designed to show that under the developed conditions the blocking

groups are continuously exchanging and the macrocycle is transported directionally.

74

It was envisaged that the information ratchet could be operated autonomously by adding the

required reagents to a solution containing the ratchet via syringe pumps ensuring a constant

supply of fuel. After screening various conditions (base, stoichiometry, rate of addition, and

addition by syringe pump or not) it was found those shown in Scheme 20 worked well.

Scheme 20. Autonomous operation of 69. Reaction conditions: (R)-81 (5 eq.), KHCO3 (20 eq.), CH2Cl2, r.t., Fmoc-Cl

(solution in CH2Cl2, added via syringe pump at 20 μL/h, 2.4 eq./h) then NEt3 (6 eq.) added after 1 h of Fmoc-Cl

addition.

The role of NEt3 is to deprotect the Fmoc group on the rotaxane and allow the macrocycle to

shuttle. The catalyst reacts with the incoming Fmoc-Cl to form an activated acylation complex

which then reprotects the alcohol as the Fmoc carbonate, preferentially when the macrocycle

is on FumD2. The supply of Fmoc-Cl is continuously replenished via syringe pump. Rather than

replenish the stocks of NEt3 via syringe pump, KHCO3 was added to regenerate the NEt3 which

otherwise may form hydrochloride salts under the reaction conditions. Typical autonomous

operation conditions were to dissolve the rotaxane, acylation catalyst (R)-81 and KHCO3 in

CH2Cl2, start the addition of Fmoc-Cl, then after 1 hour add NEt3. Subjecting rotaxane 69 with a

starting macrocycle distribution of FumH2:FumD2 100:0 to operation conditions for 24 hours

resulted in a new distribution of FumH2:FumD2 35:65. The difference in positional isomer ratio

from that obtained in Table 3 can be rationalised by not all of the starting material reacting

during the time frame of the autonomous operation. Despite that, these results do confirm

75

that under the developed conditions the Fmoc protecting group is exchanging and the

macrocycle is shuttling (Figure 31).

Figure 31. Partial 1H NMR spectra (600 MHz, CD3OD:CDCl3 3:1, 300 K) of (a) starting material, (b) after operation for

24 h. The change in integrals for shielded H11 and H12 over the 24h period of the reaction indicates a change in the

ratio of positional isomers (FumH2-69 and FumD2-69) as a result of the ratchet mechanism. Residual solvent peaks

are shown in grey. The lettering corresponds to proton labelling in Scheme 14.

To further demonstrate the continuous nature of the operation of 69, isotope labelling

experiments were performed focussing on monitoring continuous operation with in situ

exchange of the blocking Fmoc groups. Autonomous operation of rotaxane 69 with d2-Fmoc-Cl

will generate the isotopologue d2-69. Further operation in the presence of Fmoc-Cl should

regenerate the original isotopologue 69, (Scheme 21). This can be observed by mass

spectrometry, observing a change from [69+X]+ to [d2-69+X]+ and back to [69+X]+. Although this

method alone will not reveal the composition of the positional isomers in the reaction mixture,

it should allow the observation of continuous operation by switching between the

isotopologues in situ.

76

Scheme 21. Autonomous operation of 69. Reaction conditions: (R)-81 (5 eq.), KHCO3 (20 eq.), CH2Cl2, rt, Fmoc-Cl,

CH2Cl2, added via syringe pump at 20 μL/h, 2.4 eq./h then NEt3 (6 eq.) added after 1 h of Fmoc-Cl addition.

Deuterium-labelled 9-fluorenylmethyl chloroformate was synthesised in two steps from

commercially available fluorene-9-carboxylic acid. Reduction of the carboxylic ester in the

presence of sodium borodeuteride gave the corresponding alcohol with >95% deuterium

incorporation. Subsequent reaction with phosgene generated the desired chloroformate.

Scheme 22: (i) EtOH, H2SO4, reflux, 18 h, quant.; (ii) NaBD4, MeOH, 0 °C to r.t., 18 h, 72%; (iii) 20% phosgene solution

in toluene, CH2Cl2, 0 °C to r.t., 18 h, 43%.

To test the suitability of 86 to participate in autonomous operation a test reaction was

performed as in Scheme 20 but with 86 as fuel. This provided stocks of d2-69 for the following

experiment. Rotaxane d2-69 ([69+Na]+:[d2-69+Na]+ 37:63), as approximated by isotope pattern

simulation, Figure 32) was submitted to autonomous operation conditions in the presence of

Fmoc-Cl. After 18 hours an aliquot was taken and analysed by mass spectrometry and a clear

change was observed from [69+Na]+:[d2-69+Na]+ 37:63, to 90:10. The operation was continued

77

in the presence of d2-Fmoc-Cl and after 48 hours another sample was taken for mass

spectrometric analysis and revealed the ratio of isotopologues to be [69+Na]+:[d2-69+Na]+

34:66. Figure 32 shows a) starting isotope distribution b) isotope distribution after 18 hours

and c) isotope distribution after a further 48 hours. A clear shift of 2 mass units is observed

between samples, as would be expected with the loss and subsequent reincorporation of the

deuterium label. Therefore, over the period of 66 hours, the Fmoc group is being cleaved and

reprotected continuously; the machine is operating autonomously.

Figure 32. Partial MS showing [M+Na]+ of rotaxane 69 a, t = 0 h, b, after 18 h operation in the presence of Fmoc-Cl

and c, after a further 48 h operation in the presence of d2-Fmoc-Cl. Orange lines indicate simulated isotope patterns

of mixtures of [69+Na] and [d2-69+Na]. Peaks shown in grey correspond to [2M+K+NH4]2+.

Operation of equilibrium mixtures of positional isomers should show no change in the 1H NMR.

However, by taking advantage of isotopologues it is possible to demonstrate that the system is

indeed still under continuous operation. A sample of rotaxane 69 (at an equilibrium

distribution of positional isomers, FumH2:FumD2 15:85) was exposed to operation conditions

(rotaxane, acylation catalyst (R)-81 and KHCO3 in CH2Cl2, start the addition of Fmoc-Cl, then

after 1 hour add NEt3) for 22.5 hours and the 1H NMR sample before and after operation

78

(Figure 33) was compared. Indeed, the fumaramide signals which are indicative of the position

of the macrocycle do not change significantly. However, a change in the mass spectrum is

observed from [69+Na]+ to mixtures of [69+Na]+ and [d2-69+Na]+ (Figure 34) revealing that

rotaxane at equilibrium mixtures of positional isomers can still operate.

Figure 33. Partial 1H NMR spectra (600 MHz, CD3OD:CDCl3 3:1, 300 K) of (a) starting material, (b) after operation for

22.5 h. Residual solvent peaks are shown in grey. The lettering corresponds to proton labelling in Scheme 14.

Figure 34. Partial MS showing [M+Na]+ of a) rotaxane 69 at equilibrium distribution of positional isomers and b)

after operation showing incorporation of the deuterium label. Peaks shown in grey correspond to [2M+K+NH4]2+.

79

2.7. Conclusion

Rotaxane 69 has been demonstrated to be an autonomous, chemically-fuelled information

ratchet system. Steric information was revealed to be the driving force of the information

ratchet, with acylation reactions taking place preferentially when the macrocycle is far away

from the reactive site. The deacylation reaction was shown to be unbiased. Deuterium

labelling experiments were exploited as evidence for the autonomous nature of the operation.

It is hoped these results will prove useful in designing an autonomous chemically-fuelled

information ratchet which operates over greater distances and a higher number of

compartments or in a rotary fashion by incorporation into a catenane architecture.

Development of an autonomous chemically-fuelled rotary motor would represent a major

advance in the field of synthetic motors.

80

2.8. Experimental Section

Scheme 13: (i) SOCl2, MeOH, r.t. to reflux, 4 h, quant.; (ii) DIPEA, TBTU, HOBt, DMF, 0 °C to r.t., 18 h, 69%; (iii)

LiOH·H2O, THF:H2O 3:1, r.t., 21 h, quant.

81

Scheme 14: (i) TBAF, THF, r.t., 18 h, 95%; (ii) Fmoc-Cl, DMAP, Py, CH2Cl2, 18 h, 73%; (iii) isophthaloyl chloride, p-

xylylene diamine, NEt3, CHCl3, 3 h, 75%.

82

Scheme 23: (i) a) 70, TFA, CH2Cl2, r.t., 20 h; b) 77, DIPEA, TBTU, HOBt, DMF, r.t., 18 h, 87% over two steps; (ii) TBAF,

THF, r.t., 18 h, 95%.

Scheme 22: (i) EtOH, H2SO4, reflux, 18 h, quant.; (ii) NaBD4, MeOH, 0 °C to r.t., 18 h, 72%; (iii) 20% phosgene solution

in toluene, CH2Cl2, 0 °C to r.t., 18 h, 43%.

Scheme 16: (i) Nicotinoyl chloride hydrochloride, NEt3, CH2Cl2, r.t., 18 h, 71%.

83

Scheme 17: (i) Nicotinoyl chloride hydrochloride, NEt3, CH2Cl2, r.t., 2 h, 86%.

84

To a suspension of 11-aminoundecanoic acid (7.0 g, 34.8 mmol) in MeOH (70 mL), SOCl2 (1.6

eq., 4.02 mL, 56.0 mmol) was added slowly. The resulting solution was stirred under N2 at

room temperature for 2 hours and then heated at reflux for 2 hours. Upon cooling, the solvent

was removed in vacuo giving 76 (8.7 g, 34.8 mmol, quant.) as a colourless solid.

MP: 154–158 °C.

1H NMR (600 MHz, CD3OD): δ = 3.67 (s, 3H, H1), 2.93 (t, J = 7.7 Hz, 2H, H12), 2.34 (t, J = 7.4 Hz,

2H, H3), 1.70 – 1.59 (m, 4H, H4+11), 1.45 – 1.30 (m, 12H, H5+6+7+8+9+10).

13C NMR (151 MHz, CD3OD): δ = 174.58 (C2), 50.55 (C1), 39.36 (C12), 33.36 (C3), 29.01

(C5/6/7/8/9/10), 28.99 (C5/6/7/8/9/10), 28.92 (C5/6/7/8/9/10), 28.77 (C5/6/7/8/9/10), 28.74 (C5/6/7/8/9/10), 27.18

(C11), 26.02 (C5/6/7/8/9/10), 24.59 (C4).

LRMS: (ESI+) m/z 216 [M+H]+ (100%).

HRMS: (NSI+) calculated for C12H26NO2 [M+H]+: 216.1958; observed: 216.1954. Δ = -1.9 ppm.

A flask containing 75 (1 g, 3.55 mmol) and 76 (1.2 eq., 1.07 g , 4.25 mmol) was purged with N2

and anhydrous DMF (40 mL) was added. After cooling to 0 °C, DIPEA (2 eq., 1.25 mL, 7.1

mmol), TBTU (1.2 eq., 1.365 g, 4.25 mmol) and HOBt (1.2 eq., 0.575 g, 4.25 mmol) were added.

The solution was stirred under N2 and allowed to return to room temperature. After 18 hours,

1M HCl (aq.) was added and the aqueous layer was extracted with EtOAc. The combined

85

organic layers were washed with NaHCO3 (sat. aq.), brine, dried over MgSO4 and concentrated

under reduced pressure. Purification by flash column chromatography (SiO2, CH2Cl2:EtOAc 8:2)

gave 73 (1.17 g, 2.43 mmol, 69%) as a colourless solid.

MP: 99–101 °C.

1H NMR (600 MHz, CD2Cl2): δ = 7.40 – 7.33 (m, 3H, H25+26), 7.29 – 7.22 (m, 3H, H21+22), 7.21 –

7.17 (m, 2H, H20), 7.06 – 7.03 (m, 2H, H24), 5.97 (s, 1H, H13), 4.98 (s, 2H, H18), 3.62 (s, 3H, H1),

3.21 (m, 2H, H12), 2.27 (t, J = 7.6 Hz, 2H, H3), 1.61 – 1.54 (m, 2H, H4), 1.46 (m, 2H, H11), 1.31 –

1.22 (m, 12H, H5+6+7+8+9+10).

13C NMR (151 MHz, CD2Cl2): δ = 174.58 (C2), 164.95 (C17), 164.12 (C14), 141.87 (C23), 137.75

(C19), 130.13 (C25/26), 128.92 (C20/21/22), 128.88 (C20/21/22), 128.66 (C24/25/26), 128.63 (C24/25/26),

127.91 (C21/22), 53.77 (C18), 51.77 (C1), 40.19 (C12), 34.51 (C3), 29.95 (2×CH2), 29.87 (CH2), 29.74

(2×CH2), 29.62 (CH2), 27.38 (CH2), 25.47 (C4). NOTE: C15 and C16 not observed.

LRMS: (ES-) m/z 515 [M+Cl]- (100%).

HRMS: (NSI+) calculated for C29H37D2N2O4 [M+H]+: 481.3030; observed: 481.3022. Δ = -1.7 ppm.

73 (1.17 g, 2.43 mmol) was dissolved in THF:H2O (18:6 mL) and LiOH·H2O (3 eq., 0.31 g, 7.29

mmol) was added. The resulting solution was stirred at room temperature. After 21 hours 1M

HCl (aq.) was added and the aqueous phase was extracted with EtOAc. The combined organic

layers were washed with brine, dried over MgSO4 and concentrated under reduced pressure to

give 77 (1.12 g, 2.40 mmol, 95%) as a colourless solid.

86

MP: 120–124 °C.

1H NMR (600 MHz, CD3OD): δ = 7.44 – 7.36 (m, 3H, H25+26), 7.31 – 7.24 (m, 3H, H21+22), 7.21 (d, J

= 6.5 Hz, 2H, H20), 7.10 (d, J = 6.6 Hz, 2H, H24), 5.04 (s, 2H, H18), 3.21 (t, J = 7.1 Hz, 2H, H12), 2.29

(t, J = 7.4 Hz, 2H, H3), 1.61 (m, 2H, H4), 1.51 (m, 2H, H11), 1.37 – 1.29 (m, 12H, H5+6+7+8+9+10).

13C NMR (151 MHz, CD3OD): δ = 177.70 (C2), 166.40 (C17), 166.22 (C14), 142.23 (C23), 138.13

(C19), 130.83 (C25/26), 129.69 (C20), 129.57 (C21/22/25/26), 129.52 (C21/22/25/26), 129.42 (C24), 128.66

(C21/22), 54.28 (C18), 40.65 (C12), 34.95 (C3), 30.55 (CH2), 30.50 (CH2), 30.38 (CH2), 30.31 (CH2),

30.22 (CH2), 30.15 (C11), 27.96 (CH2), 26.09 (C4). NOTE: C15 and C16 not observed.

LRMS: (ES+) m/z 465 [M-H]- (100%).

HRMS: (NSI+) calculated for C28H35D2N2O4 [M+H]+: 467.2873; observed: 467.2866. Δ = -1.5 ppm.

72 (650 mg 0.94 mmol) was dissolved in TBAF (1.0 M in THF, 2.1 eq., 2 mL, 2.0 mmol). After 18

hours the reaction mixture was diluted with CH2Cl2. The organic layer was washed with NH4Cl

(sat. aq.), citrate solution (sat. aq.), brine, dried over Na2SO4 and concentrated under reduced

pressure. Purification by flash column chromatography (SiO2, CH2Cl2:EtOH 95:5 to 90:10) gave

78 (415 mg, 0.91 mmol, 95%) as a colourless solid.

MP: 66–72 °C.

1H NMR (600 MHz, CD2Cl2): δ = 7.39 – 7.33 (m, 3H, H8+9), 7.29 – 7.22 (m, 3H, H1+2), 7.21 – 7.19

(m, 2H, H3), 7.07 – 7.03 (m, 2H, H7), 6.98 – 6.93 (m, 1H, H11/12), 6.75 – 6.68 (m, 2H, H11/12+14),

5.09 (s, 1H, H18), 4.98 (s, 2H, H5), 3.69 (p, J = 5.1 Hz, 1H, H16), 3.52 – 3.45 (m, 1H, H22), 3.42 –

87

3.35 (m, 1H, H15), 3.26 (dt, J = 14.2, 5.8 Hz, 1H, H15), 3.19 – 3.12 (m, 1H, H17), 3.08 (dt, J = 14.6,

5.9 Hz, 1H, H17), 1.41 (s, 9H, H21).

13C NMR (151 MHz, CD2Cl2): δ = 165.86 (C13), 164.74 (C10), 157.79 (C19), 141.81 (C6), 137.70 (C4),

134.21 (C11/12), 131.75 (C11/12), 130.16 (C8/9), 128.94 (2×C1/2/3), 128.72 (C7/8/9), 128.66 (C7/8/9),

127.93 (C1/2), 80.22 (C20), 71.31 (C16), 53.26 (C5),* 44.03 (C17), 43.36 (C15), 28.57 (C21).

*Determined by HSQC.

LRMS: (ES+) m/z 354 [M-C5H9O2]+ (100%); 454 [M+H]+ (65%); 476 [M+Na]+ (25%); 492 [M+K]+

(51%); 907 [2M+H]+ (44%); 929 [2M+Na]+ (39%); 945 [2M+K]+ (35%).

HRMS: (NSI+) calculated for C25H32N3O5 [M+H]+: 454.2336; observed: 454.2324. Δ = -2.6 ppm.

[α] 𝟐𝟓𝑫

+4.70 (c 0.36, CH2Cl2).

To a solution of 78 (280 mg, 0.62 mmol) in CH2Cl2 (2 mL) was added 9-

fluorenylmethoxycarbonyl chloride (2.5 eq., 398 mg, 1.55 mmol), DMAP (0.1 eq, 7.5 mg, 0.062

mmol) and pyridine (5 eq., 0.27 mL, 3.10 mmol). After 18 hours the reaction mixture was

diluted with CH2Cl2 and 1M HCl (aq.). The aqueous layer was extracted with CH2Cl2 and the

combined organic layers were washed with brine, dried over Na2SO4 and concentrated under

reduced pressure. Purification by flash column chromatography (SiO2, CH2Cl2:EtOH 100:0 to

98:2) gave 71 (301 mg, 0.45 mmol, 73%) as a colourless solid.

MP: 73–91 °C.

88

1H NMR (600 MHz, CD2Cl2): δ = 7.80 (d, J = 7.6 Hz, 2H, H29), 7.60 (d, J = 7.5 Hz, 2H, H26), 7.42 (t, J

= 7.5 Hz, 2H, H28), 7.38 – 7.31 (m, 5H, H8+9+27), 7.29 – 7.22 (m, 3H, H1+2), 7.21 – 7.18 (m, 2H, H3),

7.05 – 7.00 (m, 2H, H7), 6.91 (d, J = 14.9 Hz, 1H, H11), 6.72 (d, J = 14.8 Hz, 1H, H12), 6.64 – 6.59

(m, 1H, H14), 5.03 (t, J = 6.7 Hz, 1H, H18), 4.98 (s, 2H, H5), 4.63 (p, J = 5.7 Hz, 1H, H16), 4.55 – 4.46

(m, 2H, H23), 4.25 (t, J = 6.5 Hz, 1H, H24), 3.60 (dt, J = 13.2, 6.0 Hz, 1H, H15), 3.36 (ddd, J = 15.0,

7.5, 4.1 Hz, 1H, H17), 3.31 – 3.24 (m, 1H, H15), 3.14 (dt, J = 15.0, 5.8 Hz, 1H, H17), 1.41 (s, 9H,

H21).

13C NMR (151 MHz, CD2Cl2): δ = 164.93 (C13), 164.68 (C10), 156.97 (C19), 154.79 (C22), 143.96

(C25/30), 143.87 (C25/30), 141.86 (C6/25/30), 141.82 (C6/25/30), 137.76 (C4), 134.21 (C11) , 131.83 (C12),

130.14 (C8/9/27), 128.95 (C1/2/3), 128.92 (C1/2/3), 128.64 (C7), 128.41 (C28), 127.91 (C8/9/27), 127.72

(C8/9/27), 125.55 (C26), 125.52 (C26), 120.57 (C29), 80.29 (C20), 75.54 (C16), 70.01 (C23), 47.44 (C24),

40.47 (C17), 39.19 (C15), 28.55 (C21).

LRMS: (ES-) m/z 710 [M+Cl]- (100%).

HRMS: (NSI+) calculated for C40H42N3O7 [M+H]+: 676.3016; observed: 676.3017. Δ = 0.1 ppm.

[α]𝟐𝟓𝑫

-6.24 (c 1.41, CH2Cl2).

To a stirred solution of 71 (188 mg, 0.28 mmol) in anhydrous CHCl3 (140 mL) under N2 was

added simultaneously a solution of isophthaloyl chloride (8 eq., 454 mg 2.24 mmol) in

anhydrous CHCl3 (28 mL) and a solution of p-xylylene diamine (8 eq., 304 mg 2.24 mmol) and

NEt3 (16 eq., 0.62 mL, 4.48 mmol) over 3 hours using syringe pumps. The mixture was filtered

89

through a celite pad and washed with CH2Cl2. The combined organic phase was washed with

1M HCl (aq.), dried over Na2SO4 and concentrated under reduced pressure. Purification by

flash column chromatography (SiO2, Pet. Ether:EtOAc 50:50 to 0:100) gave 70 (253 mg, 0.21

mmol, 75%) as a colourless solid.

MP: 130–135 °C, 138–145 °C.

1H NMR (600 MHz, CD2Cl2): δ = 8.30 (s, 2H, HD), 8.20 (d, J = 7.8, 2H, HB), 8.18 (d, J = 7.8, 2H, HB),

7.89 – 7.81 (m, 1H, H14), 7.71 (d, J = 10.2, 1H, H29), 7.70 (d, J = 10.2, 1H, H29), 7.63 (t, J = 7.7 Hz,

2H, HA), 7.51 (d, J = 7.5 Hz, 1H, H26), 7.45 – 7.28 (m, 7H, H26+28+NHmac), 7.25 (t, 7.5 Hz, 2H, H2/27),

7.21 (t, 7.4 Hz, 2H, H2/27), 7.07 – 6.99 (m, 11H, HH+1+3), 6.89 (t, J = 7.6 Hz, 2H, H8), 6.76 (d, J = 7.7

Hz, 2H, H7), 6.42 (t, J = 7.5 Hz, 1H, H9), 5.80 (d, J = 14.8 Hz, 1H, H11), 5.47 (d, J = 14.7 Hz, 1H,

H12), 4.98 (t, J = 6.8 Hz, 1H, H18), 4.72 – 4.61 (m, 3H, H5+16), 4.55 – 4.26 (m, 10H, HF+23), 4.13 (t, J

= 6.5 Hz, 1H, H24), 3.69 (dt, J = 14.8, 6.1 Hz, 1H, H15), 3.38 (m, 4.4 Hz, 2H, H15+17), 3.31 (dt, J =

14.7, 5.9 Hz, 1H, H17), 1.27 (s, 9H, H21).

13C NMR (151 MHz, CD2Cl2): δ = 166.18 (C13), 166.07 (CE), 166.01 (CE), 165.02 (C10), 157.49 (C19),

154.85 (C22), 143.81 (C25), 143.44 (C25), 141.84 (C30), 141.76 (C30), 140.41 (C6), 138.35 (CG),

138.23 (CG), 136.49 (C4), 134.48 (CC), 134.43 (CC), 132.36 (C11), 131.98 (CB), 131.90 (CB), 129.84

(C8), 129.72 (CA/H/1/3), 129.68 (CA/H/1/3), 129.59 (CA/H/1/3), 129.54 (CA/H/1/3), 129.02 (C2), 128.76

(C28/9), 128.66 (C28/9), 128.40 (C28/9), 128.39 (C28/9), 128.12 (C12), 127.86 (C7), 127.67 (C27), 127.57

(C27), 125.43 (C26), 125.30 (C26), 123.89 (CC), 120.59 (C29), 120.56 (C29), 81.14 (C20), 75.46 (C16),

70.19 (C23), 54.80 (C5), 47.29 (C24), 44.03 (CF), 43.91 (CF), 40.76 (C17), 39.51 (C15), 28.35 (C21).

LRMS: (ES-) m/z 1243 [M+Cl]- (100%).

HRMS: (NSI+) calculated for C72H69N7O11Na [M+Na]+: 1230.4947; observed: 1230.4943. Δ = -0.3

ppm.

90

[α]𝟐𝟓𝑫

-2.72 (c 0.45, CH2Cl2).

70 (193 mg, 0.16 mmol) was stirred in a solution of CH2Cl2:TFA (3:1, 2 mL). After 20 hours the

reaction mixture was concentrated under reduced pressure and azeotroped with CHCl3. The

obtained residue was dissolved in anhydrous DMF (2 mL) under N2. Acid 77 (1.5 eq., 111 mg,

0.24 mmol), DIPEA (4 eq., 0.11 mL, 0.64 mmol), TBTU (2 eq., 102 mg, 0.32 mmol) and HOBt (2

eq., 43 mg, 0.32 mmol) were added. After 18 hours, 1M HCl (aq.) was added and the aqueous

layer was extracted with EtOAc. The combined organic layers were washed with NaHCO3 (sat.

aq.), 5 % LiCl (aq.), dried over Na2SO4 then concentrated under reduced pressure. Purification

by flash column chromatography (SiO2, CH2Cl2:EtOH 97:3 to 95:5) gave 69 (213 mg, 0.14 mmol,

87%) as a colourless solid.

MP: 128–137 °C.

91

1H NMR (600 MHz, CD2Cl2): δ = 8.28 (s, 2H, HD), 8.20 (d, J = 7.8 Hz, 2H, HB), 8.18 (d, J = 7.8 Hz,

2H, HB), 7.98 – 7.92 (m, 1H, H14), 7.72 (t, J = 8.4 Hz, 2H, H51), 7.63 (t, J = 7.7 Hz, 2H, HA), 7.55 –

7.43 (m, 4H, H48+NH), 7.39 – 7.16 (m, 16H, H2+37+38+39+42+43+19+50+NH), 7.07 – 6.98 (m, 13H, H1+3+41+H),

6.89 (t, J = 7.7 Hz, 2H, H8), 6.78 (d, J = 7.6 Hz, 2H, H7), 6.54 – 6.48 (m, 1H, H30), 6.46 (t, J = 7.5

Hz, 1H, H9), 6.18 (m, 1H, H18), 5.86 (d, J = 14.7 Hz, 1H , H11/12), 5.53 (d, J = 14.8 Hz, 1H, H11/12),

4.99 – 4.93 (m, 2H, H35), 4.74 – 4.61 (m, 3H, H5+16), 4.57 – 4.18 (m, 10H, H45+F), 4.15 (t, J = 6.3

Hz, 1H, H46), 3.51 – 3.43 (m, 2H, H15+17), 3.37 (ddd, J = 14.3, 6.7, 4.8 Hz, 1H, H17), 3.28 – 3.19 (m,

3H, H15+29), 2.12 – 2.06 (m, 2H, H20), 1.45 (dp, J = 20.4, 7.3 Hz, 4H, H21+28), 1.30 – 1.12 (m, 12H,

H22+23+24+25+26+27).

13C NMR (151 MHz, CD2Cl2): δ = 175.77 (C19), 166.39 (CE), 166.35 (CE), 166.11 (C13), 165.05 (C34),

164.85 (C10), 164.39 (C31), 154.82 (C44), 143.78 (C47), 143.48 (C47), 141.86 (C52/40), 141.78 (C52/40),

140.54 (C6), 138.27 (CG), 138.10 (CG), 137.73 (C36), 136.63 (C4), 134.62 (CC), 134.61 (CC), 132.53

(C11/12), 131.96 (CB), 131.90 (CB), 130.09 (CH), 129.84 (C8), 129.73 (CA/H/1/3/41), 129.63 (CA/H/1/3/41),

129.57 (CA/H/1/3/41), 129.55 (CA/H/1/3/41), 129.01 (CH), 128.92 (CH), 128.84 (CH), 128.79 (CH),

128.62 (CH), 128.59 (CH), 128.58 (CH), 128.42 (CA/H/1/3/41), 128.28 (C11/12), 127.92 (CH), 127.91

(CH), 127.68 (C49), 127.59 (C49), 125.40 (C48), 125.29 (C48), 124.08 (CD), 120.61 (C51), 120.57

(C51), 75.13 (C16), 70.17 (C45), 54.68 (C5), 53.25 (C35),* 47.30 (C46), 44.20 (CF), 44.03 (CF), 40.09

(C29), 39.87 (C17), 39.48 (C15), 36.87 (C20), 29.67 (C28), 29.45 (CH2), 29.39 (CH2), 29.38 (CH2),

29.36 (CH2), 29.31 (CH2), 27.05 (CH2), 25.94 (C21). *Determined by HSQC. NOTE: C32 and C33 not

observed.

LRMS: (ES-) m/z 1592 [M+Cl]- (100%).

HRMS: (NSI+) calculated for C95H94D2N9O12 [M+H]+: 1556.7298; observed: 1556.7291. Δ = -0.4

ppm.

92

[α]𝟐𝟓𝑫

-1.32 (c 0.81, CH2Cl2).

To a stirred solution of 69 (1.29 g , 0.82 mmol) in anhydrous THF (10 mL) under N2 was added

TBAF (1.0 M in THF, 3 eq.,2.5 mL, 2.5 mmol). After 18 hours the reaction mixture was

quenched with sat. NH4Cl (aq.). The aqueous layer was extracted with EtOAc and the combined

organic layers were washed with citrate solution, brine, dried over Na2SO4 before being

concentrated under reduced pressure. The residue was purified by trituration from Et2O to

give 79 (1.06 g, 0.79 mmol, 95%) as a colourless solid.

MP: 124–132 °C.

1H NMR (600 MHz, CD3OD:CDCl3 1:1) δ = 8.44 (m, 2H, HD), 8.24 – 8.17 (m, 4H, HB), 7.69 (m, 2H,

HA), 7.39 – 6.75 (m, 26H, H1+2+3+7+8+9+37+38+39+41+42+43+H), 6.56 – 6.42 (m, H9FumH2), 5.96 (m,

H11FumH2), 5.47 (m, H12

FumH2), 4.88 – 4.78 (m, 4H, H5+35), 4.53 – 4.24 (m, 8H, HF), 3.75 (m, 1H, H16),

93

3.30 – 3.16 (m, 6H, H15+17+29), 2.14 (t, J = 7.6 Hz, 2H, H20), 1.53 (m, 4H, H21+28), 1.38 – 1.15 (m,

12H, H22+23+24+25+26+27).

13C NMR (151 MHz, CDCl3:CD3OD 1:1): δ = 176.26 (C19/19'), 176.18 (C19/19'), 167.3 (CE), 166.4

(C13/31), 165.9 (C13/31), 165.7 (C10/34), 165.5 (C10/34), 141.2 (C6/40), 141.1 (C6/40), 137.8 (CG), 136.94

(C4/36), 136.91 (C4/36), 134.16 (CC), 133.08 (C11), 132.21 (CB), 130.50 (CH), 130.14 (CH), 130.09

(CH), 129.98 (CA), 129.78 (CH), 129.52 (CH), 129.44 (CH), 129.32 (CH), 129.09 (CH), 129.05 (CH),

128.97 (CH), 128.51 (CH), 128.45 (CH), 128.41 (CH), 128.33 (CH), 128.08 (CH), 127.32 (C12),

124.77 (CD), 69.59 (C16), 55.08 (C5/5'/35/35'), 54.65 (C5/5'/35/35'), 54.54 (C5/5'/35/35'), 54.13 (C5/5'/35/35'),

44.17 (CF), 43.73 (C15/17), 43.56 (C15/17), 40.42 (C29), 36.73 (C20), 30.00 (CH2), 29.93 (CH2), 29.79

(CH2), 29.74 (CH2), 29.65 (CH2), 27.55 (CH2), 26.34 (C21/28). NOTE: C32 and C33 not observed.

LRMS: (NSI+) m/z 668 [M+2H]2+ (100%); 1334 [M+H]+ (30%).

HRMS: (NSI+) calculated for C80H84D2N9O10 [M+H]+: 1334.6618; observed: 1334.6608. Δ = -0.7

ppm.

[α]𝟐𝟓𝑫

+129.64 (c 1.42, CH2Cl2).

94

Fluorene-9-carboxylic acid solution (2 g, 9.5 mmol) was dissolved in EtOH (16 mL). H2SO4 (0.2

mL) was added and the solution heated at reflux for 18 hours. The reaction was cooled to r.t.

before the majority of EtOH was removed under reduced pressure. H2O was added and the

resulting aqueous phase was extracted with CH2Cl2. The combined organic layers were washed

with 1M NaOH, dried over MgSO4 and concentrated under reduced pressure to give 84 (2.14g,

95%) as a yellow oil which was used without further purification.

1H NMR (600 MHz, CDCl3): δ = 7.76 (d, J = 7.6 Hz, 2H, H2), 7.67 (d, J = 6.7 Hz, 2H, H5), 7.42 (t, J =

7.5 Hz, 2H, H3), 7.35 (td, J = 7.5, 1.2 Hz, 2H, H4), 4.86 (s, 1H, H7), 4.23 (q, J = 7.1 Hz, 2H, H9), 1.29

(t, J = 7.1 Hz, 3H, H10).

13C NMR (151 MHz, CDCl3): δ = 171.01 (C8), 141.55 (C1), 140.87 (C6), 128.21 (C3), 127.47 (C4),

125.74 (C5), 120.16 (C2), 61.54 (C9), 53.61 (C7), 14.35 (C10).

LRMS: (ES-) m/z 237 [M-H]- (100%).

HRMS: (ES+) calculated for C16H14O2Na [M+Na+]: 261.0891; observed: 261.0896. Δ = 1.9 ppm.

84 (5.6 g, 23.8 mmol) was dissolved in MeOH (20 mL) and cooled to 0 °C under N2. NaBD4 (2.2

eq., 2.2 g, 47.6 mmol) was added in batches. The reaction was allowed to warm to r.t. and

stirred for 18 hours. After addition of 1M HCl the majority of MeOH was removed under

reduced pressure and the resulting aqueous fraction extracted with CH2Cl2. The combined

95

organic layers were dried over MgSO4 and concentrated under reduced pressure.

Recrystallization (Heptane:EtOH 97:3) gave 85 as colourless needles (3.7 g, 72%).

MP: 103–105 °C.

1H NMR (600 MHz, CDCl3): δ = 7.78 (d, J = 7.5 Hz, 2H, H2), 7.62 (d, J = 7.4 Hz, 2H, H5), 7.41 (t, J =

7.4 Hz, 2H, H3), 7.33 (td, J = 7.5, 1.2 Hz, 2H, H4), 4.12 (s, 1H, H7).

13C NMR (151 MHz, CDCl3): δ = 144.43 (C1), 141.68 (C6), 127.75 (C3), 127.24 (C4), 124.83 (C5),

120.23 (C2), 64.61 (C8), 50.33 (C7).

LRMS: (ESI+) m/z 216 [M+H]+ (100%).

HRMS: (ESI+) calculated for C14H14D2NO [M+NH4] +: 216.1352; observed: 216.1349. Δ = -1.4

ppm.

To a solution of 85 (1.46 g, 7.38 mmol) in CH2Cl2 (90 mL) at 0 °C was added phosgene (20% in

toluene, 90 mL). The solution was allowed to warm to r.t.. After 16 hours the mixture was

concentrated under reduced pressure and the resulting residue triturated with heptane. The

liquor was concentrated under reduced pressure to give 86 (820.0 mg, 43%) as an off-white

solid.

MP: 54–58 °C.

1H NMR (600 MHz, CDCl3): δ = 7.79 (d, J = 7.6 Hz, 2H, H2), 7.60 (dd, J = 7.5, 0.6 Hz, 2H, H5), 7.44

(t, J = 7.5 Hz, 2H, H3), 7.35 (td, J = 7.5, 1.0 Hz, 2H, H4), 4.30 (s, 1H, H7).

96

13C NMR (151 MHz, CDCl3): δ = 150.95 (C9), 142.53 (C6), 141.46 (C1), 128.34 (C2), 127.44 (C5),

125.15 (C4), 120.47 (C3), 46.15 (C7).

HRMS: (APCI+) calculated for C15H13D2ClNO2 [M+NH4] +: 278.0911; observed: 278.0905. Δ = -2.6

ppm.

To a solution of (R)-80 (947 mg, 1.80 mmol) in CH2Cl2 (40 mL) was added Nicotinyl Chloride

Hydrochloride (1.2 eq., 385 mg, 2.16 mmol) NEt3 (4 eq., 1 mL, 7.2 mmol). After 18 hours the

reaction was diluted with CH2Cl2, washed with NaHCO3 (sat. aq.), brine, dried over Na2SO4 and

concentrated under reduced pressure. Flash column chromatography (SiO2, Pet. Ether:EtOAc

1:1) gave (R)-81 as a white solid (805.2 mg, 1.28 mmol, 71%).

MP: 174–177 °C.

1H NMR (600 MHz, CDCl3): δ = 8.68 (dd, J = 4.9, 1.7 Hz, 1H, H2), 8.54 (s, 1H, H1), 8.06 (d, J = 1.9

Hz, 2H, H14), 7.96 (s, 1H, H16), 7.87 (s, 3H, H14+16), 7.62 (d, J = 8.2 Hz, 1H, H4), 7.36 (dd, J = 7.8,

4.9 Hz, 1H, H3), 7.23 (s, 1H, OH), 5.27 (t, J = 8.5 Hz, 1H, H10), 3.50 (ddd, J = 10.6, 7.8, 2.5 Hz, 1H,

H7), 3.20 (td, J = 10.7, 6.2 Hz, 1H, H7), 2.08 (dtd, J = 13.6, 8.0, 3.3 Hz, 1H, H9), 1.88 (ddt, J = 13.6,

10.0, 7.8 Hz, 1H, H9), 1.79 – 1.62 (m, 2H, H8).

13C NMR (151 MHz, CDCl3): δ = 171.22 (C6), 151.80 (C2), 147.82(C1), 146.46 (C13), 144.88 (C13’),

134.98 (C4), 131.91 (C15), 131.41 (C5), 127.88 (C14), 127.61 (C14), 123.61 (C3), 123.22 (C17),

122.40 (C16), 80.93 (C12), 68.28 (C10), 52.49 (C7), 30.70 (C9), 24.81 (C8).

LRMS: (APCI+) m/z 631 [M+H]+ (100%).

97

HRMS: (APCI+) calculated for C27H19F12N2O2 [M+H]+: 631.1249; observed: 631.1247. Δ = -0.63

ppm.

[α]𝟐𝟓𝑫

+20.76 (c 0.73, CH2Cl2).

To a solution of (S)-80 (526 mg, 1 mmol) in CH2Cl2 (24 mL) was added Nicotinyl Chloride

Hydrochloride (1.2 eq., 213 mg, 1.2 mmol) NEt3 (4 eq., 0.56 mL, 4 mmol). After 2 hours the

reaction was diluted with CH2Cl2, washed with NaHCO3 (sat. aq.), brine, dried over Na2SO4 and

concentrated under reduced pressure. Flash column chromatography (SiO2, Pet. Ether:EtOAc

1:1) gave (S)-81 as a white solid (540.3 mg, 0.86 mmol, 86%).

MP: 176–178 °C.

1H NMR (600 MHz, CDCl3): δ = 8.69 (dd, J = 5.0, 1.7 Hz, 1H, H2), 8.49 (d, J = 2.2 Hz, 1H, H1), 8.06

(s, 2H, H14), 7.97 (s, 1H, H16), 7.88 (s, 1H, H16), 7.85 (s, 2H, H14), 7.59 (dt, J = 8.0, 2.0 Hz, 1H, H4),

7.35 (dd, J = 7.9, 4.9 Hz, 1H, H3), 5.25 (t, J = 8.5 Hz, 1H, H10), 3.51 (ddd, J = 10.7, 8.1, 2.4 Hz, 1H,

H7), 3.17 (td, J = 10.6, 6.6 Hz, 1H, H7), 2.09 (dtd, J = 16.6, 8.1, 3.0 Hz, 1H, H9), 1.88 (tt, J = 13.6,

8.0 Hz, 1H, H9), 1.79 – 1.69 (m, 1H, H8), 1.65 (ddq, J = 13.3, 7.0, 3.7, 3.2 Hz, 1H, H8).

13C NMR (151 MHz, CDCl3): δ = 171.39 (C6), 152.08 (C2), 147.95 (C1), 146.46 (C13), 144.81 (C13),

134.74 (C4), 132.06 (C15), 131.08 (C15), 127.93 (C14), 127.61 (C14), 123.53 (C3), 123.21 (C17),

122.42 (C16), 80.94 (C12), 68.44 (C10), 52.47 (C7), 30.81 (C9), 24.78 (C8).

LRMS: (NSI+) m/z 631 [M+H]+ (100%); 653 [M+Na]+ (7%).

HRMS: (NSI+) calculated for C27H19F12N2O2 [M+H]+: 631.1249; observed: 631.1245. Δ = -0.6

ppm.

98

[α]𝟐𝟓𝑫

-19.58 (c 1.17, CH2Cl2).

General method for protection of molecular information ratchet 79.

To a solution of 79 (5 mg, 3.7 µmol) in CH2Cl2 (0.5 mL) was added Fmoc-Cl (5 eq., 4.8 mg, 18.6

µmol) and acylation catalyst (pyridine (5 eq., 2.5 µL, 18.6 µmol) or (R)-81 (5 eq., 11.8 mg, 18.6

µmol) or (S)-81 (5 eq., 11.8 mg, 18.6 µmol)). After 18 hours, 1M HCl (aq.) was added and the

aqueous layer was extracted with CH2Cl2. The combined organic layers were washed with

brine, dried over Na2SO4 then concentrated under reduced pressure. Purification by

preparative TLC (SiO2, CH2Cl2:EtOH 95:5) gave 69 which was submitted for NMR analysis.

Method for the partial deprotection of molecular information ratchet 69.

To a solution of 69 (5 mg, 3.2 µmol) in CH2Cl2 (0.2 mL) was added NEt3 (5 eq., 2.2 µL, 16.1

µmol). After 2 hours, the reaction was diluted with CH2Cl2, 1M HCl (aq.) was added and the

aqueous layer was extracted with CH2Cl2. The combined organic layers were washed with

brine, dried over Na2SO4 then concentrated under reduced pressure. Purification by

preparative TLC (SiO2, CH2Cl2:EtOH 95:5) gave 69 which was submitted for NMR analysis.

General method for autonomous operation of molecular information

ratchet 69.

To a solution of 69 (6 mg, 3.9 µmol) in CH2Cl2 (0.2 mL) was added (R)-81 (5 eq., 12.1 mg, 19.2

µmol) and KHCO3 (20 eq., 7.7 mg, 78 µmol). A solution of Fmoc-Cl (58 mg, 0.22 mmol) in CH2Cl2

(0.24 mL) was added at a rate of 10 µL/h. After 45 min NEt3 (6 eq., 3.25 µL, 23.4 µmol) was

added and Fmoc-Cl addition was continued. After 24 hours, 1M HCl (aq.) was added and the

aqueous layer was extracted with CH2Cl2. The combined organic layers were washed with

99

brine, dried over Na2SO4 then concentrated under reduced pressure. Purification by

preparative TLC (SiO2, CH2Cl2:EtOH 95:5) gave 69 which was submitted for NMR analysis.

100

2.9. References

1 S. Erbas-Cakmak, D. A. Leigh, C. T. McTernan, A. L. Nussbaumer, Chem. Rev. 2015, 115,

10081–10206.

2 M. Alvarez-Pérez, S. M. Goldup, D. A. Leigh, A. M. Z. Slawin, J. Am. Chem. Soc. 2008,

130, 1836–1838.

3 A. Carlone, S. M. Goldup, N. Lebrasseur, D. A. Leigh, A. Wilson, J. Am. Chem. Soc. 2012,

134, 8321–8323.

4 T. Guerlavais-Dagland, A. Meyer, J.-L. Imbach, F. Morvan, Eur. J. Org. Chem. 2003, 2003,

2327–2335.

5 F. Roussel, L. Knerr, R. R. Schmidt, Eur. J. Org. Chem. 2001, 2001, 2067–2073.

6 F. Roussel, L. Knerr, M. Grathwohl, R. R. Schmidt, Org. Lett. 2000, 2, 3043–3046.

7 K. C. Nicolaou, N. Winssinger, J. Pastor, F. De Roose, J. Am. Chem. Soc. 1997, 119, 449–

450.

8 A. G. Johnston, D. A. Leigh, A. Murphy, J. P. Smart, M. D. Deegan, J. Am. Chem. Soc.

1996, 118, 10662–10663.

9 F. G. Gatti, D. A. Leigh, S. A. Nepogodiev, A. M. Z. Slawin, S. J. Teat, J. K. Y. Wong, J. Am.

Chem. Soc. 2001, 123, 5983–5989.

10 M. R. Wilson, MChem thesis, The University of Edinburgh (UK), 2010.

Chapter Three

A Chemically-Fuelled, Autonomous Rotary Motor

Acknowledgements

Dr Armando Carlone, Dr Stephen Goldup, Dr Natalie Lebrasseur and Dr Jordi Solà are gratefully

acknowledged for their intellectual and synthetic contribution to this chapter.

Dr Matthew O. Kitching, Dr Thomas A. Singleton and Dr John W. Ward are gratefully

acknowledged for examining and proofreading this chapter.

102

3.1. Synopsis

The successful operation of a rotaxane information ratchet was an important milestone in the

progress of this project: conditions were developed which allowed a deacylation/reacylation

reaction to operate autonomously. This Chapter describes the synthesis and operation of a

[2]catenane rotary motor. Autonomous operation conditions developed earlier in the project

were successfully employed for the operation over a cyclic arrangement of compartments in a

rotary motor.

103

3.2. Retrosynthesis

The desired [2]catenane and its retrosynthesis is shown in Scheme 24. Catenane 58 can be

synthesised in 26 steps with 20 steps in the longest linear sequence. The catenane will be

formed through the macrocyclisation of the thread of [2]rotaxane precursor 87. A five-

component clipping procedure will be employed to form the [2]rotaxane 87.1,2 Thread 88 will

be assembled using a series of peptide coupling reactions from building blocks 89, 90, 91 and

92. Building block 89 is reported in the literature,3 and is prepared in 6 steps from

commercially available (R)-3-amino-1,2-propanediol and mono-ethyl fumarate. 90 can be

synthesised in 7 steps from (R)-3-amino-1,2-propanediol and dimethyl acetylenedicarboxylate.

91 and 92 are both prepared from 11-bromoundecanoic acid. Amines would be protected as

tert-butylcarbamates, and acids protected as their methyl, ethyl or tert-butyl esters.

104

Scheme 24: Retrosynthesis of [2]catenane 58.

3.3. Synthesis

The synthesis of 97 is shown in Scheme 25. Ester 94 was prepared from 11-bromoundecanoic

acid by thionyl chloride-assisted ester formation. Palladium catalysed C-C bond formation

conditions were employed to couple bromide 94 and 4-bromobenzaldehyde to yield 95.

Reductive amination of methylamine and aldehyde 95 gave 96 in 84% yield after purification.

105

The resulting amine was protected as its Boc carbamate in 73% yield. Saponification with

lithium hydroxide in a THF/water mixture proceeded quantitatively to yield the free acid 97,

which was used without further purification.

Scheme 25: (i) SOCl2, MeOH, r.t. to reflux, 4 h, quant.; (ii) I2, Zn, DMF, 80 °C, 4 h then 4-bromobenzaldehyde,

Pd(PPh3)4, DMF, 60 °C, 17 h, 56%; (iii) Methylamine, MeOH, NaBH4, 0 °C to r.t., 2 h, 84%; (iv) Boc2O, DMAP, r.t., 22 h,

73%; (v) LiOH·H2O, MeOH:H2O 3:1, r.t., 12 h, 95%.

The synthesis of 92 is shown in scheme 26. Also starting from 11-bromoundecanoic acid, ester

99 was prepared by activating the acid as its trifluoracetic anhydride before adding t-BuOH.

The subsequent Negishi coupling yielded 100 in 48% yield after purification. Reductive

amination of methylamine and aldehyde 100 gave 92 which would be used in a future peptide

coupling reaction.

106

Scheme 26: (i) TFAA, t-BuOH, –40 °C to r.t., 16 h, 93%; (ii) I2, Zn, DMF, 80 °C, 4 h then 4-bromobenzaldehyde,

Pd(PPh3)4, DMF, 60 °C, 24 h, 48%; (iii) methylamine, MeOH, NaBH4, 0 °C to r.t., 2 h, 82%.

Peptide coupling between acid and amine fragments 1014,5 and 1026, gave the deuterium

labelled station-gate unit 90. Subsequent saponification with lithium hydroxide in a THF/water

mixture proceeded quantitatively to yield the free acid 103, which was used without further

purification (Scheme 27).

Scheme 27: (i) HOBt, TBTU, DMF, CHCl3, 0 °C then DIPEA, r.t., 18 h, 90%; (ii) LiOH·H2O, THF:H2O 1:1, r.t., 1 h, 97%.

With the main components 89, 90, 91 and 92 in hand, the assembly of the full thread could

begin (Scheme 28). 89 was prepared by following a literature procedure.3 The Boc group of 89

was cleaved with TFA and the resulting primary ammonium TFA salt was coupled with acid 97

using peptide coupling conditions to give 104. Following a modified literature procedure3 104

was treated with TBAF to cleave the silyl ether protecting group. A sodium citrate work up

removed any tetrabutylammonium salts and column chromatography furnished 105 in 78%

107

yield. TFA-mediated Boc deprotection of 105 gave primary amine as its TFA salt which was

coupled directly with acid 103 using peptide coupling conditions to give 106. Saponification

and peptide coupling with amine fragment 92 yielded the full thread with the free hydroxyl

group, 108. Addition of fresh HOBt, TBTU then DIPEA after 18 hours and stirring the reaction

for a further 24 hours improved the yield of this step from 64% to 83%. Protection of the

hydroxyl with 9-fluoroenylmethyl chloroformate gave 88, an ideal template for rotaxane

formation using the clipping method.

Scheme 28: (i) TFA, CH2Cl2, 12 h, then 97 HOBt, TBTU, DIPEA, DMF, 0 °C to r.t., 18 h, 69%; (ii) TBAF, THF, rt, 40 min,

78%; (iii) TFA, CH2Cl2, 16 h, then 103, HOBt, TBTU, DIPEA, DMF, 0 °C to r.t., 12 h, 84%; (iv) LiOH·H2O, MeOH:H2O 3:1,

108

1 h, 97%; (v) 92, HOBt, TBTU, DIPEA, DMF, 0 °C to r.t., 18 h, then HOBt, TBTU, DIPEA, 24 h, 83%; (vi) Fmoc-Cl, DMAP,

Py, r.t., 18 h, 93%;

Similarly to that described in Section 2.5, at high dilution thread 88 was allowed to react with

p-xylylenediamine and isophthaloyl chloride which were added separately by syringe pumps

(Scheme 29). High dilution and slow addition were used to minimise the formation of

polypeptide oligomers. The hydrogen bonding preorganisation offered by the fumaramide

station encourages the components to react together and form a macrocycle around the

thread yielding a [2]rotaxane. Both FumH2 and FumD2 are suitable templates for macrocycle

formation, but only clipping around FumD2 yields rotaxane since dethreading is possible from

the FumH2 portion of the molecule. The steric bulk of the TBDPS group on one side of FumD2,

and the Fmoc group on the other prevents the macrocycle from dethreading. Although this

step is low yielding, it was possible to isolate and recycle any unreacted thread. The next key

step in the synthesis was the macrocyclisation of [2]rotaxane 87 to give [2]catenane 109

(Scheme 29). Firstly, rotaxane 87 was stirred in a TFA/CH2Cl2 mixture to cleave the Boc group

and the tert-butyl ester yielding the free amine and acid respectively. After excess TFA was

removed by azeotrope with CHCl3, the residue was dissolved in DMF and added via syringe

pump to a solution containing HATU and DIPEA resulting in macrocyclisation of the thread to

give [2]catenane 109 in 50% yield over two steps.

109

Scheme 29: (i) Isophthaloyl chloride, p-xylylene diamine, NEt3, CHCl3, 3 h, 26%; (ii) TFA, CHCl3, r.t., 3 h then DMF,

HATU, DIPEA, r.t., 2 h, 50%.

3.4. Operation and Analysis

3.4.1. Directionality Studies

Now, all that remained in the synthesis was deprotection of the TBDPS group, and

reprotection of the resulting alcohol with Fmoc. To determine the directional bias of the motor

the reprotection step had to be studied over one gate only, therefore it was important to be

able to selectively cleave the TBDPS group in the presence of Fmoc. TBAF- and TBAT-mediated

deprotection proved unsuitable with both affording undesirable cleavage of the Fmoc group.

However, methanolic HCl (~1.25 M) was successful in selectively cleaving the TBDPS group at

room temperature within a few hours to yield 110. To establish the directionality of the motor

it was necessary to screen various conditions so a model compound was proposed where one

of the Fmoc carbonate groups is replaced with an Fmoc ester, 111 (Scheme 30). This was

prepared by reacting 110 with the appropriate Fmoc derivative 112 activated as its acyl

chloride. 112 was synthesised in 35% yield from fluorene and bromopropionic acid.

110

Scheme 30: (i) 1.25 M HCl in MeOH, CH2Cl2, 4 h, quant.; (ii) a) 112, (COCl)2, r.t., 3 h, then b) 110, Py, CH2Cl2, r.t., 92%;

(iii) n-BuLi, bromopropionic acid, –5 °C to r.t., 5 h, 35%.

The Fmoc ester in 111 was expected to provide a similar steric environment to the Fmoc

carbonate but be stable to traditional Fmoc deprotection conditions. The advantage of

screening conditions on this model rather than the real system is that the material can be

recovered and recycled until screening is complete, preserving precious material (Scheme 31).

111

Scheme 31: (i) NEt3, CH2Cl2, r.t., 18 h, 68%.; (ii) Fmoc-Cl, acylation catalyst, solvent, 20 h.

The typical procedure for operation was to dissolve catenane 114, an acylation catalyst and

Fmoc-Cl in dry solvent. After the indicated time the reaction was diluted with

dichloromethane, washed with 1 M HCl (aq.), and purified by preparative TLC (CH2Cl2:EtOH

95:5). The result of acylation was a mixture of kinetically trapped positional isomers FumH2

and FumD2 the ratio of which was determined by 1H NMR. The initial test employed pyridine as

112

the acylation catalyst in CH2Cl2 at room temperature. Unfortunately only a modest directional

bias was observed, with a FumH2:FumD2 distribution of 56:44. A screen of different solvents

(acetone, acetonitrile, chloroform, dimethylformamide, nitromethane, pyridine,

tetrachloroethane, toluene) revealed that the directionality was independent of solvent

choice, with most examples giving a FumH2:FumD2 distribution of ~60:40. No reaction was

observed in the case of DMF. Varying the temperature also proved ineffective in increasing the

directionality of the ratchet. Reactions performed with pyridine as catalyst at –78 °C (acetone,

CH2Cl2), –60 °C (CHCl3), – 40 °C (acetonitrile, DMF), –30 °C (acetone, CHCl3) and 50 °C (acetone,

CHCl3) showed no significant improvement in directional bias compared to room temperature.

No variation in directional bias was observed when reactions were stopped at different time

points (1 hr vs. 4 hr, Py, CH2Cl2, –78 °C). Next, a screen of pyridine-based nucleophiles was

performed. Use of 2,4,6-collidine, 2,6-lutidine or 3,5-lutidine did not improve the directional

bias. Concurrently with this screen of catalysts for the acylation of 114, the acylation of

rotaxane 69 was also studied (see Section 2.6.1), and 81 was revealed to be a suitable

candidate. However, before this result had been obtained, 4-pyrrolidinopyridine derivatives

51 and 54 were tested for the acylation of 114 but as was observed for rotaxane 69, they were

found to be too basic and no product was observed in the reaction.

Figure 35: Acylation catalysts screened.

The results obtained for the acylation of 114 are given in Table 4 and Figure 36.

113

Table 4. Product distribution upon acylation of 114 using various catalysts.

Conditions Catalyst Product Distribution

(a) Pyridine FumH2:FumD2 56:44 (b) (R)-51/(S)-51 No product observed (c) (R)-54/(S)-54 No product observed (d) (R)-81 FumH2:FumD2 25:75 (e) (S)-81 FumH2:FumD2 27:73

Reaction of 114 with Fmoc-Cl in the presence of (R)-81 resulted in the macrocycle being

kinetically locked predominantly in the FumD2 compartment (25:75). Use of the antipode

catalyst (S)-81 gave a similar result. Since both hands of catalyst 81 gave similar results and the

acylation complex clearly reacts preferentially when the macrocycle is far away from the

reactive site, this reveals that directional movement is governed by sterics rather than

chirality, as with the rotaxane model. With directional bias on the model compound 114

established, these optimised conditions (catenane, CH2Cl2, Fmoc-Cl (5 eq.), (R)-81 (5 eq.), r.t.,

19 hr) were then tested on 110. Pleasingly, in the presence of (R)-81 the reaction proceeded

with a directional bias of FumH2:FumD2 81:19 (Figure 36).

Scheme 32: (i) (R)-81, Fmoc-Cl, CH2Cl2, r.t., 19 h, 93%;

114

Figure 36: Partial 1H NMR spectra (500 MHz, CD2Cl2:CD3OD: 1:1, 300 K) of acylation of 114 in the presence of (a) Py,

(b) (R)-81, (c) (S)-81 and (d) acylation of 110 in the presence of (R)-81. The lettering corresponds to labelling in

Scheme 31 and 32. The section 6.30—5.75 ppm has been scaled vertically 3× compared to the section 8.80—8.35

ppm.

As with the rotaxane model it has been established that the position of the macrocycle can

affect the rate of protection, therefore it is reasonable to postulate that the position of the

macrocycle may also influence the rate of deprotection. If deprotection is more likely to occur

on the favoured positional isomer (in an equal and opposite amount to the bias upon

protection) then net motion would not occur. To investigate this, catenane 111 FumH2:FumD2

30:70 was subjected to deprotection conditions, and the reaction stopped before reaching

completion (Scheme 33).

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Scheme 33: Reaction conditions: (i) NEt3, CH2Cl2, r.t., 1h, 75%.

If the recovered unreacted catenane has the same positional ratio as the starting catenane

then both positional isomers must have been consumed at the same rate and it can be

concluded that the position of the macrocycle does not influence the rate of deprotection.

When 111 was treated with NEt3 (10 eq.) for 1 hour and the unreacted starting material

recovered, 1H NMR analysis revealed the ratio of FumH2:FumD2 to be the same as in the

starting material catenane (Figure 37).

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Figure 37. Partial 1H NMR spectra (600 MHz, CD3OD:CDCl3 3:1, 300 K) of (a) starting material and (b) recovered

starting material. Residual solvent peaks are shown in grey. The lettering corresponds to proton labelling in Scheme

31. The section 6.30—5.75 ppm has been scaled vertically 3× compared to the section 8.80—8.35 ppm.

This leads to the conclusion that both positional isomers were consumed at the same rate and

therefore the position of macrocycle does not affect the rate of deprotection. The

directionality of the [2]catenane rotary motor was found to be comparable to that in the

[2]rotaxane model. At this stage the results were promising mirroring those obtained on the

rotaxane model.

3.4.2. Autonomy

The ultimate aim of this project is to operate the motor autonomously. That is, so long as the

fuel source is maintained, the deprotection and reprotection, and associated macrocycle

shuttling, should occur continuously. With each full rotation of the small macrocycle around

the large one cumulative work is possible. However, if during operation significant amounts of

diol are formed any work performed is undone. An operation cycle is depicted in Figure 38.

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Figure 38. Schematic illustration of the operation cycle of a [2]catenane motor. The ring preferentially resides on

one or other of the two binding sites (green). The red spheres represent bulky groups which sterically block the

motion of the ring between the two binding sites and trap it in one compartment of the track or the other. Removal

of a red sphere allows the ring to shuttle between the two stations. Reattachment of the sphere under appropriate

conditions ratchets a quantity of rings into the next compartment. The blue arrow indicates the direction of

transport in the situation where kfar>kclose for protection and kfar≅kclose for deprotection.

Before discussing the operation cycle it is important to keep in mind two points: 1) protection

results in directional motion of the small macrocycle away from the reactive site (kfar>kclose) and

2) the rate of deprotection of positional isomers is equal (kfar=kclose). Starting from FumH2-58,

Fmoc deprotection will result in 110 and 110’ in equal amounts. The macrocycle is now able to

shuttle: both 110 and 110’ will exist as mixtures of their FumH2 and FumD2 positional isomers.

Since reprotection is favoured when the macrocycle is far away, reprotection of 110 will result

in mostly FumD2-58. The macrocycle has now completed a half rotation. Under autonomous

operation conditions deprotection and reprotection should happen continuously, so

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deprotection of FumD2-58 will occur. This again generates 110 and 110’ in equal amounts as

mixtures of their positional isomers. Now reprotection of 110’ will regenerate FumH2-58 and

the macrocycle completes a full rotation. Since 110 and 110’ are formed in equal amounts and

110 will react to preferentially give FumD2-58 and 110’ will react to preferentially give FumH2-

58 then over time, the distribution should tend towards a FumH2:FumD2 ratio of 1:1. The

change in distribution can be followed by 1H NMR and is an indication of autonomous

operation. When the rate of protection is faster than the rate of deprotection the mechanism

of operation should proceed as described. However, in the case where the rate of

deprotection is faster than the rate of protection we need to consider the implication of

forming diol. The problem of generating diol is two-fold: 1) it reduces the efficiency of the

system by half, and 2) the system can no longer be described as a motor since cumulative work

is not possible. Upon forming diol the macrocycle is free to shuttle along the entire larger

macrocycle, with no preference for direction. The first protection (to give 110 or 110’, Figure

38) doesn’t result in any directional motion of the smaller macrocycle. It isn’t until 110/110’ is

reprotected again does the ratchet mechanism work and result in directional motion. For

every two protections, only one results in directional motion, significantly reducing the

efficiency. Secondly, any work done by the ratchet is undone when the macrocycle is ‘released’

and allowed to shuttle over the entire large macrocycle in its diol form. The key distinction

between a switch and a motor is that a motor is able to perform cumulative work. That is,

when the machine is reset any work performed is not undone. Therefore, by operating

through the diol the system can only be described as a switch.

Catenane 58 was submitted to operation conditions and isolated at various time points to

monitor the macrocycle distribution over FumH2:FumD2. A typical procedure was to dissolve 58

(FumH2:FumD2 ~80:20 as dictated by the chosen synthetic route), (R)-81 (5 eq.) and KHCO3 (20

eq.) in CH2Cl2. Fmoc-Cl in CH2Cl2 was added via syringe pump (2.4 eq./h) for 1 hour when NEt3

(1.5 eq.) was added as a stock solution in CH2Cl2. Fmoc-Cl addition was continued until the

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given time when the mixture was diluted with CH2Cl2 and washed with 1M HCl. Purification by

preparative TLC (SiO2, CH2Cl2:EtOH 95:5) gave 58 as a mixture of kinetically trapped isomers

FumH2 and FumD2, the ratio of which was determined by 1H NMR (Figure 39). Since formation

of diol would be detrimental to the operation mechanism, these conditions differ slightly from

those used on rotaxane 69; notably less NEt3. To minimise the formation of diol during the

reaction it was decided to use a substoichiometric amount of NEt3 (1.5 eq. or 0.75 eq. per

Fmoc protected hydroxyl).

Figure 39: Partial 1H NMR spectra (500 MHz, CD2Cl2:CD3OD: 1:1, 300 K) of 58 (FumH2:FumD2 80:20) and 58 after

operation for 4 h, 24 h and 48 h. The lettering corresponds to labelling in Scheme 32. Peaks in blue correspond to

the internal standard on the macrocycle, and peaks in green correspond to fumaramide protons shielded by the

macrocycle. The section 6.3—5.7 ppm has been scaled vertically 6× compared to the section 8.8—8.3 ppm.

These results clearly show the macrocycle distribution is tending to a 1:1 mixture over a period

of 48 hours. As deprotection is unbiased, the only way the distribution can be changing is if

deprotection and reprotection is occuring. Therefore, over a period of 48 hours the reaction is

operating autonomously. This change in macrocycle distribution could also be obtained if all of

the catenane was deprotected to the diol then reprotected again. However, operation through

this mechanism would result in no net work. It was therefore necessary to use further

techniques to monitor the composition of the mixture during operation. By using an

isotopically-labelled fuel during operation, mass spectrometry should reveal insights into the

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autonomous nature of operation. As a reminder, operation of catenane 58 with d2-Fmoc-Cl 86

should generate the isotopologue d4-58. Further operation in the presence of Fmoc-Cl should

regenerate the original isotopologue 58, (Scheme 34). This can be followed by mass

spectrometry, observing a change from [58+X]+ to [d4-58+X]+ and back to [58+X]+.

Scheme 34: Conditions: (i) (R)-81 (10 eq.), NEt3 (12 eq.), KHCO3 (40 eq.), CH2Cl2, d2-Fmoc-Cl (1M in CH2Cl2, 4.8 eq/h

via syringe pump), r.t., 24 h; then (ii) Fmoc-Cl (1M in CH2Cl2, 4.8 eq/h via syringe pump), r.t., 48 h.

A typical procedure was to dissolve 58, (R)-81 (10 eq.) and KHCO3 (40 eq.) in CH2Cl2. Fmoc-Cl in

CH2Cl2 was added via syringe pump (4.8 eq./hr) for 1 hour when NEt3 (12 eq.) was added.

Fmoc-Cl addition was continued and aliquots taken for MS analysis. Pleasingly after overnight

operation a change was observed in the MS from [58+Na]+ to mixtures of [58+Na]+, [d2-

58+Na]+ and [d4-58+Na]+ (Figure 40).

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Figure 40: Partial mass spectra of operation of catenane 58 showing [58+Na]+ at (a) t = 2 h; (b) t = 19 h; (c) t = 144 h

(isolated).

Unfortunately when the d2-Fmoc-Cl was swapped for Fmoc-Cl a change back to [58+Na]+ was

not observed (Figure 40 c). After 19 h of operation at a rate of addition of 4.8 eq./h, 91.2 eq of

d2-Fmoc-Cl should have been added to the reaction mixture. When the fuel was changed to

Fmoc-Cl it would take a further 19 h before the concentration of Fmoc-Cl exceeds d2-Fmoc-Cl

and any incorporation of the new fuel to be noticeable by mass spectrometry. Further, as the

amount of (d2)-Fmoc-Cl in the reaction increases, the chance of NEt3 reacting with catenane

rather than Fmoc-Cl is significantly reduced so it is not surprising to not see further changes in

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the mass spectrum. These mass spectrometry results show that over a period of 19 h the

deprotection and reprotection of the Fmoc protecting group is happening in one pot, i.e. the

motor is operating autonomously. Although changes are not observed after 19 h this does not

necessarily mean the motor stalls at that stage, rather that further operation results in very

subtle changes in the mass spectrum.

To gain further insight into the mechanism of operation a reaction was monitored every 2

hours (58, (R)-81 (10 eq.) and KHCO3 (40 eq.) in CH2Cl2. Fmoc-Cl in CH2Cl2 added via syringe

pump (4.8 eq./hr) for 30 min when NEt3 (12 eq.) was added ). Select MS results are presented

in Figure 41.

Figure 41: Partial mass spectra of operation of catenane 58 (a) t = 0 h; (b) t = 2 h; (c) t = 4 h; (d) t = 8 h; (e) t = 22 h.

The most noticeable result is that within 8 hours all of the catenane has been deprotected with

the majority existing as the diol form. This is a disappointing result: as previously mentioned it

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renders the catenane a switch, not a motor. However, after 22 hours most of the catenane has

been reprotected by d2-Fmoc-Cl, as indicated by the appearance of the [d4-58+Cl]– peak. Peaks

highlighted in orange which correspond to [58+Cl]– before and after operation are shown in

Figure 42 with a clear shift of 4 mass units, representing incorporation of the labelled Fmoc.

Figure 42: Partial mass spectra of operation of 58 (a) before and (b) after operation.

Although these results show that almost all the material reacted with the diol as an

intermediate, it is clear that the deprotection and reprotection reactions are compatible in one

pot, supporting the argument for autonomous operation. An interesting difference can be

seen when comparing Figure 40 b) with Figure 42 b). The isotope distribution of Figure 40 b)

indicates a mixture of d2-58 and d4-58 whereas that in Figure 42 b) appears to be almost

entirely d4-58. This implies not all of the reaction in Figure 40 has operated through the diol,

meaning it is possible to reduce the formation of diol by optimising the amount and timing of

NEt3 addition (after 120 mins for Figure 40 vs 30 mins for Figure 42).

It can be expected that catenane 58, mono-OH catenane 110 and diol catenane 115 have

different ionisation efficiencies in the mass spectrometer, so to obtain quantitative results it

was decided to follow the reactions by HPLC and determine the molar ratio of catenane

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58/mono-OH 110/diol 115 in the reaction at any given time. Since each species is sequentially

losing a UV-active fluorene motif, it can be expected that each would have a different

absorbance value at the same concentration, therefore the absorbance coefficient for each

species had to be determined. Separation of the three different catenane species was achieved

using a normal-phase column: SUPELCO, Analytical, Ascentis® Si, 250 × 4.6 mm, 5 µm, 1

mL/min, with an eluent gradient of DCM:IPA 95:5 for 5 min initially increasing to 40:60 over

30 minutes, followed by 10 minutes post time to return to 95:5 (Figure 43).

Figure 43: HPLC traces showing catenane 58, mono-OH 110 and diol 115 respectively.

With separation conditions optimised a stock solution of each species was prepared. From this

stock solution of known concentration, a series of solutions of decreasing concentrations were

prepared, injected into the HPLC instrument and eluted using the conditions mentioned above

and their corresponding absorbance at 254 nm was measured. Figure 44 displays the

absorbance versus concentration plot for catenane 58, mono-OH 110 and diol 115. Their

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gradient ε (molar absorbance coefficient) and R2 value is also shown. Reassuringly, the series

follows the trend expected for relative absorbance; catenane 58>mono-OH 110>diol 115.

Figure 44: A plot of absorbance as a function of concentration for catenane 58, mono-OH 110 and diol 115.

The behaviour of the catenane motor in response to stepwise-addition of fuel is shown in

Figure 45. Through the addition of NEt3 then (d2-)Fmoc-Cl activated with (R)-81 two

deprotection-reprotection cycles were performed. High performance liquid chromatography

(HPLC) was used to monitor the composition of the mixture during the reaction and it was

found that the amount of catenane species in its double Fmoc protected form 58 ranged from

32-100 %; mono-protected 110 ranged from 0-57 % while the amount of diol was kept to a

minimum (≤11 %). Mass spectrometry analysis after each deprotection-reprotection cycle

showed incorporation then loss of the deuterium label with the ratio of [58+Na]:[d2-

58+Na]:[d4-58+Na] changing from 100:0:0 to 32:50:18 to 65:30:5 (as approximated by isotope

pattern simulation, Figure 46).

y = 8.5686x + 60.413R² = 0.9928

y = 6.0111x + 44.243R² = 0.9919

y = 4.7259x - 29.45R² = 0.9972

0

200

400

600

800

1000

1200

1400

0 50 100 150 200 250

Ab

sorb

ance

L·m

ol-1

·cm

-1

Concentration (µM)

catenane 58 mono-OH 110 diol 115

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Cycles Time (h)

Composition of the mixture Incorporation of isotope label

58 mono-OH 110 diol [58+Na]+ [d2-58+Na]+ [d4-58+Na]+

0 100 0 0 100 0 0

1 { 6 32 57 11 - - -

24 94 6 0 32 50 0

2 { 29 34 55 11 - - -

32 100 0 0 65 30 0 Figure 45: Exchange of blocking groups during stepwise operation of catenane 58. Reaction conditions: (i) NEt3 (8

eq.), (ii) d2-Fmoc-Cl (16 eq.), (R)-81 (16 eq.), (iii) NEt3 (10 eq.), (iv) Fmoc-Cl (16 eq.), (R)-81 (16 eq.).

Figure 46: Mass spectrum of (a) [58+Na]+, (b) [58+Na]+ at t=24 h and (c) [58+Na]+ at t=32 h. Orange lines indicate

simulated isotope patterns of mixtures of [58+Na], [d2-58+Na] and [d4-58+Na].

Thus, in the presence of the appropriate fuel, the catenane motor 58 operates repetitively. In

addition to operating through the desired mono-hydroxyl mechanism, the presence of diol

(albeit in small amounts, ≤11 %) represents a minor pathway in the stepwise operation cycle.

Interestingly, even the biological rotary motor F1-ATPase has been observed to rotate 120° in

the wrong direction occasionally during its ‘unidirectional’ cycle. This experiment

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demonstrates that 58 is able to undergo two cycles of deprotection-reprotection in the same

pot so long as the appropriate reagents are present. In conjunction with the directionality

studies in Section 3.4.1. and the NMR evidence showing macrocycle distribution change over

time in Section 3.4.2. it can be concluded that 58 possesses all the qualities required to be

called an autonomous, chemically-fuelled rotary molecular motor.

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3.5. Conclusion and Outlook

A [2]catenane information ratchet rotary motor has been successfully synthesised and

operated. As with rotaxane 69, the catalyst in its acylated intermediate form is able to

discriminate between the two postitional isomers of 58 which interconvert through the

macrocycle shuttling between the two fumaramide residues, and preferentially acylates the

hydroxyl group when the macrocycle is on the FumD2 group. This directional discrimination

appears to be independent of catalyst handednesss, and rather stems from the inherent

asymmetry of the thread. Temperature or solvent dependency was not observed to affect the

directional bias of the macrocycle upon acylation for 58. The directionality of the [2]catenane

rotary motor was found to be as effective as in the [2]rotaxane model. Through a series of 1H

NMR, HPLC and mass spectrometry experiments it has been demonstrated that the Fmoc

protecting groups on 58 can be cleaved and reprotected repeatedly in one-pot. This leads to

the conclusion that 58 can act as a chemically-fuelled autonomous rotary motor.

The autonomous chemically-fuelled rotary motor presented within is built around an

information ratchet core where a protecting group is utilised as a gate to selectively trap the

macrocycle when on one side. Fundamental to this project is the ability to open and close this

gate (deprotect and protect) in the same pot. For work to be possible these steps must occur

through different mechanisms and not be in equilibrium. Although all these goals have been

achieved, this rotary motor is still very primitive in comparison to Nature’s machines. Exploring

new chemistries which fit this brief could lead to more directional motors with increased fuel

efficiency.

The molecular machines capable of autonomous chemically-fuelled operation presented in this

thesis represent a significant advance in the development of molecular machines. In fact, these

machines are the first of their kind: to date no other machine reported has been able to mimic

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Nature’s use of chemical energy to autonomously drive a system out of thermodynamic

equilibrium.

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3.6. Experimental Section

Scheme 25: (i) SOCl2, MeOH, r.t. to reflux, 4 h, quant.; (ii) I2, Zn, DMF, 80 °C, 4 h then 4-bromobenzaldehyde,

Pd(PPh3)4, DMF, 60 °C, 17 h, 56%; (iii) Methylamine, MeOH, NaBH4, 0 °C to r.t., 2 h, 84%; (iv) Boc2O, DMAP, r.t., 22 h,

73%; (v) LiOH·H2O, MeOH:H2O 3:1, r.t., 12 h, 95%.

Scheme 26: (i) TFAA, t-BuOH, –40 °C to r.t., 16 h, 93%; (ii) I2, Zn, DMF, 80 °C, 4 h then 4-bromobenzaldehyde,

Pd(PPh3)4, DMF, 60 °C, 24 h, 48%; (iii) methylamine, MeOH, NaBH4, 0 °C to r.t., 2 h, 82%.

131

Scheme 27: (i) HOBt, TBTU, DMF, CHCl3, 0 °C then DIPEA, r.t., 18 h, 90%; (ii) LiOH·H2O, THF:H2O 1:1, r.t., 1 h, 97%.

132

Scheme 28: (i) TFA, CH2Cl2, 12 h, then 97 HOBt, TBTU, DIPEA, DMF, 0 °C to r.t., 18 h, 69%; (ii) TBAF, THF, rt, 40 min,

78%; (iii) TFA, CH2Cl2, 16 h, then 103, HOBt, TBTU, DIPEA, DMF, 0 °C to r.t., 12 h, 84%; (iv) LiOH·H2O, MeOH:H2O 3:1,

1 h, 97%; (v) 92, HOBt, TBTU, DIPEA, DMF, 0 °C to r.t., 18 h, then HOBt, TBTU, DIPEA, 24 h, 83%; (vi) Fmoc-Cl, DMAP,

Py, r.t., 18 h, 93%.

Scheme 35: (i) TBAF, THF, r.t., 18 h, 66%; (ii) Fmoc-Cl, Py, r.t., 18 h, 58%.

Scheme 29: (i) Isophthaloyl chloride, p-xylylene diamine, NEt3, CHCl3, 3 h, 26%; (ii) TFA, CHCl3, r.t., 3h then DMF,

HATU, DIPEA, r.t., 2 h, 50%.

133

Scheme 36: (i) 1.25M HCl in MeOH, CH2Cl2, 5 h, quant.; (ii) (R)-81, Fmoc-Cl, CH2Cl2, r.t., 19 h, 93%; (iii) a) 112,

C2O2Cl2, r.t., 3 h, then b) 110, Py, CH2Cl2, r.t., 92%;

134

Scheme 37: (i) NEt3, CH2Cl2, r.t., 18 h, 68%.

Scheme 38: (i) n-BuLi, bromopropionic acid, –5 °C to r.t., 5 h, 35%.

135

To a suspension of 11-bromoundecanoic acid (10.0 g, 38 mmol) in MeOH (75 mL), SOCl2 (1.6

eq., 4.4 mL, 61 mmol) was added slowly. The resulting solution was stirred under N2 at room

temperature for 2 hours and then heated at reflux for 2 hours. Upon cooling, the solvent was

removed in vacuo and the residue filtered through a pad of silica (EtOAc). Concentration under

reduced pressure gave 94 (10.5 g, 38 mmol, quant.) as a colourless oil.

1H NMR (400 MHz, CDCl3): δ = 3.62 (s, 3H, H1), 3.36 (t, J = 6.9 Hz, 2H, H12), 2.26 (t, J = 7.5 Hz,

2H, H3), 1.87 – 1.75 (m, 2H, H11), 1.63 – 1.52 (m, 2H, H4), 1.42 – 1.33 (m, 2H, H10), 1.25 (s, 10H,

H5+6+7+8+9).

13C NMR (101 MHz, CDCl3): δ = 173.95 (C2), 51.25 (C1), 33.90 (CH2), 33.76 (CH2), 32.71 (CH2),

29.25 (CH2), 29.21 (CH2), 29.10 (CH2), 28.99 (CH2), 28.62 (CH2), 28.04 (CH2), 24.80 (CH2).

LRMS: (APCI+) m/z 279 [M(79Br)+H]+ (100%), 281 [M(81Br)+H]+ (100%).

HRMS: (APCI+) calculated for C12H2479BrO2 [M+H]+: 279.0954; observed: 279.0953. Δ = –0.4

ppm.

A solution of I2 (0.05 eq., 38 mg, 0.15 mmol) and Zn (1.5 eq., 294 mg, 4.5 mmol) in anhydrous

DMF (3 mL) under N2 was stirred at r.t. for 3 minutes. 94 (879 mg, 3 mmol) was added and the

reaction mixture heated to 80 °C. After 4 hours the mixture was added to a solution of 4-

bromobenzaldehyde (0.6 eq., 335 mg, 1.8 mmol) and Pd(PPh3)4 (0.05 eq., 173 mg, 0.15 mmol)

in anhydrous DMF (3.6 mL) under N2 and the resulting mixture heated at 60 °C. After 17 hours

136

the mixture was cooled to r.t. and filtered through a pad of silica (EtOAc). The combined

organics were washed with water, dried over MgSO4 and concentrated under reduced

pressure. Flash chromatography (SiO2, CH2Cl2:Pet. Ether 3:1) gave 95 as a white solid (309 mg,

1.0 mmol, 56%).

MP: 74–79 °C.

1H NMR (400 MHz, CDCl3): δ = 9.96 (s, 1H, H18), 7.79 (d, J = 8.1 Hz, 2H, H15), 7.33 (d, J = 8.1 Hz,

2H, H14), 3.66 (s, 3H, H1), 2.72 – 2.63 (m, 2H, H12), 2.29 (t, J = 7.5 Hz, 2H, H3), 1.61 (dt, J = 14.9,

8.1 Hz, 4H, H4+11), 1.34 – 1.23 (m, 12H, H5+6+&+8+9+10).

13C NMR (101 MHz, CDCl3): δ = 192.22 (C17), 174.48 (C2), 150.62 (C16), 134.48 (C13), 130.03 (C15),

129.21 (C14), 51.60 (C1), 36.34 (CH2), 34.22 (CH2), 31.21 (CH2), 29.57 (CH2), 29.52 (CH2), 29.51

(CH2), 29.35 (2×CH2), 29.24 (CH2), 25.06 (CH2).

LRMS: (APCI+) m/z 305 [M+H]+ (100%); 322 [M+NH4]+ (32%).

HRMS: (APCI+) calculated for C19H29O3 [M+H]+: 305.2111; observed: 305.2105. Δ = –2.0 ppm.

To a solution of 95 (100 mg, 0.33 mmol) in MeOH (1 mL) was added methylamine (1.3 eq., 38

µL of 40% aq. solution, 0.43 mmol). After stirring at r.t. for 15 minutes the mixture was cooled

to 0 °C and NaBH4 (0.5 eq., 7 mg, 0.18 mmol) was added portionwise. The resulting solution

was stirred at r.t. for 2 hours. After addition of H2O, the MeOH was removed under reduced

pressure and the resulting aqueous fraction extracted with CH2Cl2. The combined organic

layers were dried over MgSO4 and concentrated under reduced pressure. Flash

chromatography (SiO2, CH2Cl2:MeOH 5:1) gave 96 as an oil (90 mg, 0.28 mmol, 84%).

137

1H NMR (400 MHz, CDCl3): δ = 7.21 (d, J = 7.8 Hz, 2H, H15), 7.13 (d, J = 7.8 Hz, 2H, H14), 3.71 (s,

2H, H17), 3.65 (s, 3H, H1), 2.60 – 2.54 (m, 2H, H12), 2.44 (s, 3H, H19), 2.29 (t, J = 7.5 Hz, 2H, H3),

2.00 (s, 1H, H18), 1.67 – 1.52 (m, 4H, H4+11), 1.37 – 1.17 (m, 12H, H5+6+7+8+9+10).

13C NMR (101 MHz, CDCl3): δ = 174.40 (C2), 141.79 (C13), 137.13 (C16), 128.51 (C14/15), 128.25

(C14/15), 55.83 (C17), 51.52 (C1), 35.99 (C19), 35.71 (CH2), 34.20 (CH2), 31.62 (CH2), 29.59 (CH2),

29.57 (CH2), 29.52 (CH2), 29.40 (CH2), 29.34 (CH2), 29.24 (CH2), 25.05 (CH2).

LRMS: (ES+) m/z 320 [M+H]+ (100%).

HRMS: (ES+) calculated for C20H34NO2 [M+H]+: 320.2589; observed: 320.2605. Δ = 5.0 ppm.

To a solution of 96 (319 mg, 1 mmol) in CHCl3 (10 mL) was added Boc2O (1.2 eq, 262 mg, 1.2

mmol) and DMAP (0.05 eq., 6.1 mg, 0.05 mmol). After stirring at r.t. for 22 hours 1M HCl was

added and the aqueous layer extracted with CHCl3. The combined organic layers were washed

with NaHCO3 (sat. aq.) dried over MgSO4 and concentrated under reduced pressure. Flash

chromatography (SiO2, Pet. Ether:EtOAc 9:1) gave 91 as a colourless oil (308 mg, 0.73 mmol,

73%).

On the NMR timescale, 91 exists as a mixture of rotamers.

1H NMR (600 MHz, CDCl3): δ = 7.13 (s, 4H, H14+15), 4.38 (s, 2H, H17), 3.66 (s, 3H, H1), 2.80 (br s,

3H, H18), 2.60 – 2.55 (m, 2H, H12), 2.30 (t, J = 7.6 Hz, 2H, H3), 1.59 (m, 4H, H4+11), 1.48 (s, 9H,

H21), 1.34 – 1.23 (m, 12H, H5+6+7+8+9+10).

13C NMR (151 MHz, CDCl3): δ = 174.41 (C2), 156.23 (C19), 155.90 (C19), 141.96 (C13), 135.30 (C16),

128.59 (C14), 127.78 (C15), 127.36 (C15), 79.69 (C20), 79.59 (C20), 52.44 (C17), 51.69 (C17), 51.52

138

(C1), 35.70 (C12), 34.18 (C3), 33.87 (C18), 31.61 (C11), 29.58 (CH2), 29.56 (CH2), 29.52 (CH2), 29.40

(CH2), 29.33 (CH2), 29.22 (CH2), 28.56 (C21), 25.03 (C4).

LRMS: (NSI+) m/z 420 [M+H]+ (100%); 437 [M+NH4]+; 364 [M–C4H9]+ (22%); 320 [M-C5H9O2]+

(14%).

HRMS: (NSI+) calculated for C25H42NO4 [M+H]+: 420.3108; observed: 420.3112. Δ = 1.0 ppm.

To a solution of 91 (2.7 g, 6.4 mmol) in MeOH (22.5 mL) was added a solution of LiOH·H2O (6.3

eq., 1.7 g, 40.5 mmol) in H2O (7 mL). After 12 hours 1M HCl was added and the aqueous

fraction was extracted with CH2Cl2. The combined organic fractions were washed with brine,

dried over MgSO4 and concentrated under reduced pressure to give 97 as a colourless oil (2.47

g, 6.1 mmol, 95%).

On the NMR timescale, 97 exists as a mixture of rotamers.

1H NMR (600 MHz, CDCl3): δ = 7.13 (s, 4H, H14+15), 4.38 (s, 2H, H17), 2.80 (s, 3H, H18), 2.60 – 2.55

(m, 2H, H12), 2.34 (t, J = 7.5 Hz, 2H, H3), 1.65 – 1.56 (m, 4H, H4+11), 1.48 (s, 9H, H21), 1.33 – 1.24

(m, 12H, H5+6+7+8+9+10).

13C NMR (151 MHz, CDCl3): δ = 179.41 (C2), 156.35 (C19), 156.03 (C19), 142.02 (C13), 135.30 C16),

128.66 (C14), 127.83 (C15), 127.40 (C15), 79.86 (C20), 79.72 (C20), 52.50 (C17), 51.75 (C17), 35.72

(C12), 34.10 (C3), 33.94 (C18), 31.60 (C11), 29.85 (CH2), 29.59 (CH2), 29.55 (CH2), 29.49 (CH2),

29.35 (CH2), 29.18 (CH2), 28.61 (C21), 24.82 (C4).

LRMS: (NSI–) m/z 404 [M-H]– (100%).

HRMS: (NSI–) calculated for C24H38NO4 [M-H]–: 404.2806; observed: 404.2801. Δ = –1.2 ppm.

139

A solution of 11-bromoundecanoic acid (40 g, 0.15 mol) in dry THF (228 mL) was cooled to –40

°C and trifluoroacetic anhydride (2 eq., 42 mL, 0.3 mol) was added slowly. After 30 minutes

stirring at –40 °C tert-Butanol (13.4 eq., 192 mL, 2.0 mol) was added and the solution allowed

to warm to r.t. and stirred for 16 hours. The reaction mixture was poured slowly onto a

mixture of crushed ice and NaHCO3 (sat. aq.). The aqueous layer was extracted with EtOAc and

the combined organics were washed with brine, dried over MgSO4 and concentrated under

reduced pressure. Flash chromatography (SiO2, Hexane:CH2Cl2 92:8) gave 99 as a colourless oil

(46 g, 0.14 mol, 93%).

1H NMR (500 MHz, CDCl3): δ = 3.37 (t, J = 6.9 Hz, 2H, H13), 2.17 (t, J = 7.5 Hz, 2H, H4), 1.82 (dt, J

= 14.3, 6.9 Hz, 2H, H12), 1.59 – 1.49 (m, 2H, H5), 1.41 (s, 11H, H1+11), 1.25 (s, 10H, H6+7+8+9+10).

13C NMR (126 MHz, CDCl3): δ = 173.34 (C3), 79.93 (C2), 35.67 (CH2), 34.04 (CH2), 32.91 (CH2),

29.44 (CH2), 29.42 (CH2), 29.32 (CH2), 29.14 (CH2), 28.81 (CH2), 28.24 (CH2), 28.20 (H1), 25.17

(CH2).

LRMS: (ES+) m/z 343, [M(79Br)+Na]+ (100%); 345, [M(81Br)+Na]+ (100%); 265 [M(79Br)-C4H9]+

(87%); 265 [M(81Br)-C4H9]+ (87%).

HRMS: (ES+) calculated for C15H2979BrO2Na [M+Na]+: 343.1248; observed: 343.1255. Δ = 2.0

ppm.

140

A solution of I2 (0.05 eq., 1.27 g, 5 mmol) and Zn (1.5 eq., 9.81 g, 150 mmol) in anhydrous DMF

(100 mL) under N2 was stirred at r.t. for 3 minutes. 99 (32.1 g, 100 mmol) was added and the

reaction mixture heated to 80 °C. After 4 hours the mixture was added to a solution of 4-

bromobenzaldehyde (0.6 eq., 11.2 g, 60 mmol) and Pd(PPh3)4 (0.1 eq., 11.5 g, 10 mmol) in

anhydrous DMF (120 mL) under N2 and the resulting mixture heated at 60 °C. After 24 hours

the mixture was cooled to r.t. and filtered through a pad of silica (EtOAc). The combined

organics were washed with water, dried over MgSO4 and concentrated under reduced

pressure. Flash chromatography (SiO2, Pet. Ether:EtOAc 9:1) gave 100 as an off-white solid

(10.1 g, 29 mmol, 48%).

1H NMR (400 MHz, CDCl3): δ = 9.97 (s, 1H, H19), 7.80 (d, J = 8.1 Hz, 2H, H16), 7.33 (d, J = 8.0 Hz,

2H, H15), 2.74 – 2.63 (m, 2H, H13), 2.19 (t, J = 7.5 Hz, 2H, H4), 1.73 – 1.50 (m, 4H, H5+12), 1.44 (s,

9H, H1), 1.35 – 1.23 (m, 12H, H6+7+8+9+10+11).

13C NMR (101 MHz, CDCl3): δ = 192.09 (C18), 173.36 (C3), 150.51 (C17), 134.35 (C14), 129.90 (C16),

129.08 (C15), 79.92 (C2), 36.23 (CH2), 35.62 (CH2), 31.10 (CH2), 29.47 (CH2), 29.43 (CH2), 29.42

(CH2), 29.29 (CH2), 29.25 (CH2), 29.08 (CH2), 28.13 (C1), 25.10 (CH2).

LRMS: (ES+) m/z 385 [M+K]+ (100%); 369 [M+Na]+ (67%).

HRMS: (ES+) calculated for C22H34O3Na [M+Na]+: 369.2406; observed: 369.2419. Δ = 3.5 ppm.

To a solution of 100 (11.8 g, 34.2 mmol) in MeOH (114 mL) was added methylamine (1.3 eq.,

3.9 mL of 40% aq. solution, 45 mmol). After stirring at r.t. for 15 minutes the mixture was

cooled to 0 °C and NaBH4 (0.5 eq., 650 mg, 17.1 mmol) was added portionwise. The resulting

141

solution was stirred at r.t. for 2 hours. After addition of H2O the MeOH was removed under

reduced pressure and the resulting aqueous fraction extracted with CH2Cl2. The combined

organic layers were dried over MgSO4 and concentrated under reduced pressure to give 92 as

a white solid (10.1 g, 27.9 mmol, 82%). The product was used without further purification.

MP: 43–48 °C.

1H NMR (400 MHz, CDCl3): δ = 7.22 (d, J = 8.0 Hz, 2H, H16), 7.13 (d, J = 8.0 Hz, 2H, H15), 3.72 (s,

2H, H18), 2.60 – 2.54 (m, 2H, H13), 2.45 (s, 3H, H20), 2.19 (t, J = 7.5 Hz, 2H, H4), 1.89 (s, 1H, H19),

1.62 – 1.51 (m, 4H, H5+12), 1.44 (s, 9H, H1), 1.33 – 1.22 (m, 12H, H5+6+7+8+9+10+11).

13C NMR (101 MHz, CDCl3): δ = 173.47 (C3), 141.92 (C17), 136.76 (C14), 128.56 (C15), 128.35 (C16),

80.00 (C2), 55.65 (C18), 35.78 (C20), 35.73 (2×CH2), 31.65 (CH2), 29.63 (CH2), 29.59 (CH2), 29.58

(CH2), 29.44 (CH2), 29.41 (CH2), 29.20 (CH2), 28.23 (C1), 25.23 (CH2).

LRMS: (NSI+) m/z 362 [M+H]+ (100%).

HRMS: (NSI+) calculated for C23H40NO2 [M+H]+: 362.3054; observed: 362.3054. Δ = 0.0 ppm.

101 (206 mg, 1.56 mmol), 102 (1.2 eq., 809 mg, 1.89 mmol), HOBt (1.5 eq., 316 mg, 2.34

mmol) and TBTU (1.5 eq., 751 mg, 2.34 mmol) were dissolved in DMF (0.5 mL, peptide

synthesis grade) and CHCl3 (15 mL) under N2 and cooled to 0 °C. DIPEA (1.2 eq., 0.33 mL, 1.87

mmol) was added dropwise and the resulting solution allowed to warm to r.t. and stirred for

18 hours. 1M HCl was added and the aqueous layer extracted with CHCl3:IPA 3:1. The

142

combined organics were washed with NaHCO3 (sat. aq.), LiCl (5% w/w aq.) dried over MgSO4

and concentrated under reduced pressure. Dry column vacuum chromatography (SiO2,

heptane:EtOAc 7:3) gave 90 as a white solid (762 mg, 1.41 mmol, 90%).

MP: 140–142 °C.

1H NMR (600 MHz, CDCl3): δ = 7.66 (dd, J = 13.7, 6.4 Hz, 4H, H18), 7.45 (t, J = 6.8 Hz, 2H, H19),

7.44 – 7.37 (m, 4H, H17), 6.64 (t, J = 6.4 Hz, 1H, H6), 4.86 – 4.81 (m, 1H, H10), 3.90 (dt, J = 10.1,

5.0 Hz, 1H, H8), 3.79 (s, 3H, H1), 3.64 (ddd, J = 13.1, 7.3, 4.6 Hz, 1H, H7), 3.31 (ddd, J = 14.7, 8.3,

3.5 Hz, 1H, H9), 3.07 (dt, J = 13.2, 6.1 Hz, 1H, H7), 2.92 (dt, J = 14.7, 5.1 Hz, 1H, H9), 1.43 (s, 9H,

H13), 1.09 (s, 9H, H15).

13C NMR (151 MHz, CDCl3): δ = 166.12 (C2), 163.70 (C5), 156.93 (C11), 135.79 (C18), 133.50 (C16),

130.26 (C19), 130.21(C19), 128.09 (C17), 128.06 (C17), 79.99 (C12), 70.70 (C8), 52.27 (C1), 43.13

(C9), 41.65 (C7), 28.47 (C13), 27.12 (C15), 19.45 (C14). NOTE: C3 and C4 not observed.

LRMS: (NSI+) m/z 543 [M+H]+ (100%); 565 [M+Na]+ (25%).

HRMS: (NSI+) calculated for C29H39D2N2O6Si [M+H]+: 543.2854; observed: 543.2849. Δ = –0.9

ppm.

[α]𝟐𝟓𝑫

+20.20 (c 0.29, CH2Cl2).

To a solution of 90 (5.3 g, 9.8 mmol) in THF (30 mL) was added a solution of LiOH·H2O (3 eq.,

1.23 g, 29.4 mmol) in H2O (16 mL). After stirring at r.t. for 1 hour 2M HCl was added and the

143

aqueous fraction was extracted with EtOAc. The combined organic fractions were washed with

brine, dried over MgSO4 and concentrated under reduced pressure to give 103 as a white solid

(5.02 g, 9.5 mmol, 97%).

MP: 78–87 °C.

1H NMR (400 MHz, CDCl3): δ = 7.66 (td, J = 7.9, 1.6 Hz, 4H, H, H18), 7.48 – 7.35 (m, 6H, H17+19),

7.29 (dd, J = 7.8, 4.6 Hz, 1H, H6), 4.90 (dd, J = 8.4, 3.7 Hz, 1H, H10), 3.91 (dq, J = 7.8, 3.8 Hz, 1H,

H8), 3.76 (ddd, J = 13.6, 7.8, 3.6 Hz, 1H, H7), 3.43 (ddd, J = 14.3, 8.5, 4.0 Hz, 1H, H9), 3.10 (ddd, J

= 13.5, 8.8, 4.6 Hz, 1H, H7), 2.97 (dt, J = 14.3, 3.9 Hz, 1H, H9), 1.43 (s, 9H, H13), 1.10 (s, 9H, H15).

13C NMR (101 MHz, CDCl3): δ = 168.38 (C2), 164.11 (C5), 157.12 (C11), 135.86 (C18), 135.76 (C18),

133.53 (C16), 133.12 (C16), 130.25 (C19), 130.16 (C19), 128.04 (C17), 128.01 (C17), 80.94 (C12),

70.20 (C8), 43.13 (C9), 41.62 (C7), 28.49 (C13), 27.11 (C15), 19.46 (C14). NOTE: C3 and C4 not

observed.

LRMS: (NSI–) m/z 587 [M+C2H3O2]– (100%); 573 [M+CHO2]– (65%); 527 [M-H]– (59%).

HRMS: (NSI+) calculated for C28H35D2N2O6Si [M-H]–: 527.2552; observed: 527.2550. Δ = –0.4

ppm.

[α]𝟐𝟓𝑫

+38.42 (c 0.28, CH2Cl2).

144

To a solution of 89 (2.65 g, 4.8 mmol) in CH2Cl2 (16 mL) an open flask was added TFA (2 mL).

After 12 hours 1M NaOH was added until the solution was basic by pH paper. The aqueous

layer was extracted with EtOAc and the combined organics were dried over MgSO4 and

concentrated under reduced pressure. The resulting residue was redissolved in DMF (22 mL,

peptide synthesis grade) under N2 and cooled to 0 °C. 97 (1.3 eq., 2.5 g, 6.2 mmol), HOBt (1.5

eq., 979 mg, 7.25 mmol), TBTU (1.55 eq., 2.4 g, 7.5 mmol) and DIPEA (1.55 eq., 1.3 mL, 7.5

mmol) were added. The reaction mixture was allowed to warm to r.t. and stirred for 18 hours

when 1M HCl was added. The aqueous layer was extracted with EtOAc and the combined

organics were washed with NaHCO3 (sat. aq.), brine, dried over MgSO4 and concentrated

under reduced pressure. Flash column chromatography (SiO2, hexane:EtOAc 5:1) gave 104 as a

colourless oil (2.78 g, 3.3 mmol, 69%).

On the NMR timescale, 104 exists as a mixture of rotamers.

1H NMR (600 MHz, CDCl3): δ = 7.69 – 7.64 (m, 4H, H36), 7.49 – 7.43 (m, 2H, H37), 7.43 – 7.38 (m,

4H, H35), 7.13 (s, 4H, H24+25), 6.80 (d, J = 15.4 Hz, 1H, H4), 6.77 – 6.71 (m, 2H, H5+7), 5.84 (dd, J =

8.1, 5.0 Hz, 1H, H11), 4.41 – 4.36 (m, 2H, H27), 4.23 (q, J = 7.1 Hz, 2H, H2), 3.99 – 3.94 (m, 1H, H9),

3.71 (ddd, J = 13.2, 8.1, 4.4 Hz, 1H, H8), 3.58 (ddd, J = 14.2, 8.1, 3.4 Hz, 1H, H10), 2.92 (ddd, J =

145

13.7, 7.2, 4.9 Hz, 1H, H8), 2.86 (dt, J = 14.4, 5.3 Hz, 1H, H10), 2.84 – 2.75 (m, 3H, H28), 2.58 (t, J =

7.8 Hz, 2H, H22), 2.13 (td, J = 7.4, 2.4 Hz, 2H, H13), 1.62 – 1.53 (m, 4H, H14+21), 1.50 – 1.45 (m, 9H,

H31), 1.33 – 1.22 (m, 15H, H1+15+16+17+18+19+20), 1.09 (s, 9H, H33).

13C NMR (151 MHz, CDCl3): δ = 174.45 (C12), 165.57 (C3), 164.10 (C6), 156.30 (C29), 155.96 (C29),

142.02 (C23), 136.30 (C4), 135.78 (C36), 135.74 (C36), 135.34 (C26), 133.62 (C34), 133.39 (C34),

130.57 (C5), 130.33 (C37), 130.28 (C37), 128.64 (C24), 128.14 (C35), 128.11 (C35), 127.82 (C25),

127.40 (C25), 79.73 (C30), 79.61 (C30), 70.39 (C9), 61.26 (C2), 52.49 (C27), 51.72 (C27), 41.79 (C8/10),

41.65 (C8/10), 36.90 (C13), 35.75 (C22), 33.94 (C28), 31.67 (C21), 29.67 (CH2), 29.62 (CH2), 29.61

(CH2), 29.47 (2×CH2), 29.43 (CH2), 28.61 (C31), 27.09 (C33), 25.87 (C14), 19.47 (C32), 14.30 (C1).

LRMS: (ESI+) m/z 865 [M+Na]+ (100%); 1705 [2M+Na]+ (36%); 842 [M+H]+ (6%).

HRMS: (ESI+) calculated for C49H72N3O2Si [M+H]+: 842.5134; observed: 842.5130. Δ = –0.5 ppm.

[α]𝟐𝟓𝑫

+8.16 (c 1, CH2Cl2).

To a solution of 104 (996 mg, 1.17 mmol) in anhydrous THF (12 mL) under N2 was added TBAF

(4 eq., 4.7 mL of a 1M solution in THF, 4.7 mmol). After 40 minutes NH4Cl (sat. aq.) was added

and the aqueous phase extracted with EtOAc. The combined organics were washed with 1M

HCl, brine, dried over MgSO4 and concentrated under reduced pressure. Flash column

146

chromatography (SiO2, Pet. Ether:EtOAc 2:1 to 0:1) gave 105 as a colourless solid (550 mg, 0.91

mmol, 78%).

MP: 82–84 °C.

On the NMR timescale, 105 exists as a mixture of rotamers.

1H NMR (600 MHz, CDCl3): δ = 7.49 – 7.42 (m, 1H, H7), 7.12 (s, 4H, H24+25), 6.99 (dd, J = 15.5, 4.5

Hz, 1H, H4), 6.79 (dd, J = 15.4, 3.1 Hz, 1H, H5), 6.67 – 6.62 (m, 1H, H11), 4.41 – 4.34 (m, 2H, H27),

4.26 – 4.20 (m, 2H, H2), 3.82 (p, J = 5.2 Hz, 1H, H9), 3.46 (dt, J = 12.0, 5.7 Hz, 1H, H8), 3.36 (ddq,

J = 25.1, 13.2, 5.9 Hz, 2H, H8+10), 3.27 (dq, J = 14.1, 5.4, 4.9 Hz, 1H, H10), 2.83 – 2.73 (m, 3H, H28),

2.56 (t, J = 7.7 Hz, 2H, H22), 2.21 (t, J = 7.7 Hz, 2H, H13), 1.59 (dt, J = 17.1, 7.8 Hz, 4H, H14+21), 1.50

– 1.44 (m, 9H, H31), 1.32 – 1.22 (m, 15H, H1+15+16+17+18+19+20).

13C NMR (151 MHz, CDCl3): δ = 175.49 (C12), 165.61 (C3), 165.20 (C6), 156.34 (C29), 155.97 (C29),

141.99 (C23), 136.20 (C4), 136.13 (C4), 135.22 (C26), 130.81, (C5) 130.74 (C5), 128.64 (C24) 127.67

(C25), 127.39 (C25), 79.81 (C30), 79.69 (C30), 70.03 (C9), 69.93 (C9), 61.36 (C2), 52.48 (C27), 51.72

(C27), 42.81 (C8), 42.71 (C10), 36.72 (C13), 35.70 (C22), 33.95 (C28), 31.60 (C21), 29.56 (CH2), 29.41

(CH2), 29.38 (CH2), 28.59 (C31), 25.85 (C14), 14.26 (C1).

LRMS: (NSI+) m/z 604 [M+H]+ (100%); 626 [M+Na]+ (34%).

HRMS: (NSI+) calculated for C33H54N3O7 [M+H]+: 604.3956; observed: 604.3953. Δ = –0.5 ppm.

[α]𝟐𝟓𝑫

+5.82 (c 0.42, CH2Cl2).

147

To a solution of 105 (6.03 g, 10 mmol) in CHCl3 (60 mL) an open flask was added TFA (60 mL).

After 16 hours the reaction mixture was concentrated under reduced pressure and azeotroped

with CHCl3. The resulting residue (402 mg, 0.76 mmol) was redissolved in DMF (7.6 mL, peptide

synthesis grade) under N2 and cooled to 0 °C. Acid 103 (402 mg, 0.76 mmol), HOBt (1.2 eq.,

132 mg, 0.91 mmol), TBTU (1.2 eq., 385 mg, 0.91 mmol) and DIPEA (2 eq., 265 µL, 1.52 mmol)

were added. The reaction mixture was allowed to warm to r.t. and stirred for 12 hours when

1M HCl was added. The aqueous layer was extracted with EtOAc and the combined organics

were washed with NaHCO3 (sat. aq.), brine, dried over MgSO4 and concentrated under reduced

pressure. Flash column chromatography (SiO2, CH2Cl2:MeOH 97:3) gave 106 as a white solid

(648 mg, 0.64 mmol, 84%).

MP: 64–68 °C.

On the NMR timescale, 106 exists as a mixture of rotamers.

1H NMR (600 MHz, CDCl3): δ = 7.70 – 7.61 (m, 4H, H45), 7.47 – 7.42 (m, 2H, H46), 7.39 (tt, J = 7.3,

4.3 Hz, 4H, H44), 7.16 – 7.02 (m, 4H, H24+25), 6.98 (dd, J = 16.3, 11.0 Hz, 1H, H4), 6.87 – 6.82 (m,

1H), 6.79 (dd, J = 15.4, 3.5 Hz, 1H, H5), 6.71 (t, J = 6.2 Hz, 1H), 6.58 (t, J = 6.7 Hz, 1H), 6.47 (s,

1H), 4.93 (dt, J = 13.1, 6.9 Hz, 1H), 4.68 – 4.53 (m, 2H, H27), 4.22 (p, J = 6.8 Hz, 2H, H2), 3.90 (dt,

J = 11.0, 5.7 Hz, 1H, H35), 3.81 (q, J = 5.1 Hz, 1H, H9), 3.68 – 3.56 (m, 1H, H34), 3.49 – 3.24 (m,

148

5H, H8+10+36), 3.13 – 2.88 (m, 5H, H28+34+36), 2.57 (t, J = 7.5 Hz, 2H, H22), 2.21 (td, J = 7.4, 2.5 Hz,

2H, H13), 1.65 – 1.54 (m, 4H, H14+21), 1.41 (d, J = 7.3 Hz, 9H, H40), 1.32 – 1.20 (m, 15H,

H1+15+16+17+18+19+20), 1.08 (d, J = 6.1 Hz, 9H, H42).

13C NMR (151 MHz, CDCl3): δ = 175.58 (C12), 175.54 (C12), 165.71 (C3/6/29/32), 165.61 (C3/6/29/32),

165.56 (C3/6/29/32), 165.37 (C3/6/29/32), 165.14 (C3/6/29/32), 164.64 (C3/6/29/32), 164.55 (C3/6/29/32),

156.75 (C38), 142.76 (C26), 142.41 (C23), 136.22 (C4), 136.11 (C4), 135.80 (C45), 133.75 (C43),

133.57 (C43), 133.53 (C43), 133.22 (C43), 130.85 (C5), 130.75 (C5), 130.20 (C46), 130.16 (C46),

129.12 (C24/25/44), 128.94 (C24/25/44), 128.05 (C24/25/44), 128.02 (C24/25/44), 127.95 (C24/25/44), 126.74

(C24/25/44), 79.85 (C39), 70.76 (C9/35), 70.16 (C9/35), 61.34 (C2), 53.46 (C27), 51.28 (C27), 43.13

(C8/10/34/36), 42.79 (C8/10/34/36), 42.66 (C8/10/34/36), 41.83 (C8/10/34/36), 36.73 (C13), 35.58 (C22), 35.27

(C28), 34.14 (C28), 31.35 (C21), 31.26 (C21), 29.39 (CH2), 29.35 (CH2), 29.28 (2×CH2), 29.06 (CH2),

28.85 (CH2), 28.48 (C42), 27.13 (C40), 25.85 (C14), 19.45 (C41), 14.29 (C1). NOTE: C30 and C31 not

observed.

LRMS: (ESI+) m/z 1037 [M+Na]+ (100%); 1014 [M+H]+ (6%).

HRMS: (ESI+) calculated for C56H80D2N5O10Si [M+H]+: 1014.5951; observed: 1014.5940. Δ = –1.1

ppm.

[α] 𝟐𝟓𝑫

+17.06 (c 0.69, CH2Cl2).

149

To a solution of 106 (2.4 g, 2.37 mmol) in MeOH (36 mL) was added a solution of LiOH·H2O (3

eq., 300 mg, 7.11 mmol) in H2O (12 mL). After stirring at r.t. for 1 hour the mixture was

concentrated under reduced pressure, redissolved in EtOAc and 2M HCl was added. The

aqueous fraction was extracted with EtOA and the combined organic fractions were washed

with brine, dried over MgSO4 and concentrated under reduced pressure to give 107 as a white

solid (2.27 g, 2.30 mmol, 97%).

MP: 123–129 °C.

On the NMR timescale, 107 exists as a mixture of rotamers.

1H NMR (600 MHz, CD2Cl2:CD3OD): δ = 7.74 – 7.65 (m, 4H, H44), 7.47 – 7.36 (m, 6H, H43+45), 7.22

– 7.08 (m, 4H, H23+24), 6.99 (d, J = 15.5 Hz, 1H, H3), 6.73 (d, J = 15.5 Hz, 1H, H4), 4.67 – 4.64 (m, J

= 6.5 Hz, 2H, H26), 3.97 – 3.84 (m, 1H, H8), 3.75 (ddd, J = 25.0, 11.4, 5.6 Hz, 1H, H34), 3.36 – 3.30

(m, 4H, H7/9/33/35), 3.27 – 3.21 (m, 2H, H7/9/33/35), 3.13 (dd, J = 14.5, 4.5 Hz, 2H, H7/9/33/35), 3.03 (d,

J = 44.7 Hz, 3H, H27), 2.59 (t, J = 7.5 Hz, 2H, H21), 2.22 (t, J = 7.6 Hz, 2H, H12), 1.60 (s, 4H, H13+20),

1.40 (d, J = 8.8 Hz, 9H, H39), 1.35 – 1.26 (m, 12H, H14+15+16+17+18+19), 1.08 (d, J = 10.6 Hz, 9H, H41).

13C NMR (151 MHz, CD2Cl2:CD3OD): δ = 176.34 (C11), 168.04 (C2/5/28/31), 167.01 (C2/5/28/31), 166.59

(C2/5/28/31), 166.18 (C2/5/28/31), 157.54 (C37), 143.53 (C22/25), 143.23 (C22/25), 137.02 (C3/4), 136.54

150

(C44), 134.41 (C22/25/42), 134.25 (C22/25/42), 134.20 (C22/25/42), 134.06 (C22/25/42), 131.23 (C3/4),

130.61 (C45), 129.61 (C23/24), 129.38 (C23/24), 128.52 (C23/24/43), 128.45 (C23/24/43), 127.33 (C23/24),

80.04 (C38), 71.48 (C34), 69.67 (C8), 53.86 (C26)*, 51.72 (C27), 43.92 (C7/9/33/35), 43.72 (C7/9/33/35),

43.41 (C7/9/33/35), 43.15 (C7/9/33/35), 36.93 (C12), 36.19 (C21), 36.16 (C21), 35.61 (C27), 34.47 (C27),

32.25 (C13/20), 32.22 (C13/20), 30.21 (CH2), 30.17 (CH2), 30.16 (CH2), 30.14 (CH2), 30.02 (CH2),

29.95 (CH2), 28.56 (C39), 27.25 (C41), 26.52 (C13/20), 19.74 (C40). NOTE: C29 and C30 not observed.

*Determined by HSQC.

LRMS: (ESI+) m/z 1009 [M+Na]+ (100%); 1041 [M+Na]+ (76%); 986 [M+H]+ (9%).

HRMS: (ESI+) calculated for C54H76D2N5O10Si [M+H]+: 986.5638; observed: 986.5637. Δ = –0.1

ppm.

[α] 𝟐𝟓𝑫

+35.72 (c 0.29, CH2Cl2).

A solution of 107 (2.20 g, 2.23 mmol) and 92 (1.2 eq., 967 mg, 2.68 mmol) in DMF (25 mL,

peptide synthesis grade) under N2 was cooled to 0 °C. HOBt (1.2 eq., 361 mg, 2.68 mmol),

TBTU (1.2 eq., 860 mg, 2.68 mmol) and DIPEA (1.2 eq., 0.47 mL, 2.68 mmol) were added. The

reaction mixture was allowed to warm to r.t. and stirred for 18 hours when further HOBt (1.2

151

eq., 361 mg, 2.68 mmol), TBTU (1.2 eq., 860 mg, 2.68 mmol) and DIPEA (1.2 eq., 0.47 mL, 2.68

mmol) were added. The reaction mixture was stirred for 24 hours when 2M HCl was added.

The aqueous layer was extracted with EtOAc and the combined organics were washed with

NaHCO3 (sat. aq.), LiCl (5% w/w aq.), dried over MgSO4 and concentrated under reduced

pressure. Flash column chromatography (SiO2, EtOAc:MeOH 95:5) gave 108 as a white solid

(2.46 g, 1.85 mmol, 83%).

MP: 71–76 °C.

On the NMR timescale, 108 exists as a mixture of rotamers.

1H NMR (600 MHz, CD2Cl2:CD3OD 1:1) δ = 7.70 – 7.65 (m, 4H, H62), 7.56 – 7.50 (m, 1H, NH),

7.46 – 7.36 (m, 7H, H21 or H22 and H61, H63), 7.20 – 7.07 (m, 8H, H15, H16, H41, H42), 6.91 (dd, J =

15.0, 8.7 Hz, 1H, H21 or H22), 5.73 – 5.66 (m, 1H, NH), 4.66 – 4.60 (m, 4H, H18, H44), 3.90 (dq, J =

14.8, 5.5 Hz, 1H, H52), 3.75 (dt, J = 13.2, 5.7 Hz, 1H, H26), 3.35 – 3.08 (m, 8H, H25, H27, H51, H53),

3.08 – 2.96 (m, 6H, H19, H45), 2.60 – 2.55 (m, 4H, H13, H39), 2.22 – 2.16 (m, 4H, H4, H30), 1.63 –

1.52 (m, 8H, H5, H12, H31, H38), 1.42 (s, 9H, H1), 1.38 (d, 9H, H57), 1.32 – 1.25 (m, 24H, CH2), 1.06

(d, 9H, H59).

13C NMR (151 MHz, CD2Cl2:CD3OD 1:1): δ = 176.23 (C3), 174.79 (C29), 167.03 (C=O), 166.95

(C=O, 166.63 (C=O), 166.53 (C=O), 166.49 (C=O), 166.41 (C=O), 166.16 (C=O), 166.04 (C=O),

157.53, 157.48 (C55), 143.54 (Cq), 143.50 (Cq), 143.21 (Cq), 143.20 (Cq), 134.38 (Cq), 134.34 (Cq),

134.21 (Cq), 134.16 (Cq), 134.02 (Cq), 134.00 (Cq), 136.50 (C62), 135.06 (C21), 134.93 (C21), 130.77

(C22), 130.69 (C22), 130.59 (CHAr), 129.58 (CHAr), 129.35 (CHAr), 128.49 (CHAr), 128.42 (CHAr),

127.30 (CHAr), 80.93 (C2), 80.02 (C56), 71.44 (C26/52), 71.42 (C26/52), 69.71 (C26/52), 69.68 (C26/52),

53.96 (C18/44), 51.70 (C18/44), 43.97 (C25/27/51/53), 43.86 (C25/27/51/53), 43.78 (C25/27/51/53), 43.74

(C25/27/51/53), 43.38 (C25/27/51/53), 43.34 (C25/27/51/53), 43.12 (C25/27/51/53), 43.07 (C25/27/51/53), 36.92

(C4+30), 36.22 (C13/39), 36.16 (C13/39), 36.14 (C13/39), 35.61 (C19/45), 35.60 (C19/45), 34.45 (C19/45),

32.22 (C5/12/31/38), 32.19 (C5/12/31/38), 26.48 (C5/12/31/38), 25.77 (C5/12/31/38), 30.19 (CH2), 30.18 (CH2),

152

30.14 (CH2), 30.10 (CH2), 30.09 (CH2), 29.99 (CH2), 29.92 (CH2), 29.90 (CH2), 29.67 (CH2), 28.55

(C57), 28.23 (C1), 27.23 (C59), 19.70 (C58). NOTE: C47 and C48 not observed.

LRMS: (ES+) m/z 685 [M+H+K]+ (100%); 1353 [M+Na]+ (67%).

HRMS: (ESI+) calculated for C77H113D2N6O11Si [M+H]+: 1329.8513; observed: 1329.8498. Δ = –1.1

ppm.

[α] 𝟐𝟓𝑫

+16.02 (c 0.28, CH2Cl2).

To a solution of 108 (200 mg, 0.15 mmol) in CH2Cl2 (5 mL) under N2 was added DMAP (0.2 eq.,

4 mg, 0.03 mmol) and Fmoc-Cl (2.5 eq., 98 mg, 0.38 mmol). Py (5 eq., 60 µL, 0.75 mmol) was

added dropwise. The reaction was stirred at r.t. for 18 hours, diluted with CH2Cl2 and washed

with 1M HCl. The aqueous fraction was extracted with CH2Cl2 and the combined organics were

washed with brine, dried over MgSO4 and concentrated under reduced pressure. Flash column

chromatography (SiO2, CH2Cl2:MeOH 100:0 to 96:4) gave 88 as a white solid ( 215 mg, 0.14

mmol, 93%).

MP: 66–74 °C.

153

On the NMR timescale, 88 exists as a mixture of rotamers.

1H NMR (600 MHz, CD2Cl2:CD3OD 1:1): δ = 7.81 – 7.77 (m, 2H, H71), 7.72 – 7.67 (m, 4H, H62),

7.65 – 7.60 (m, 2H, H68), 7.47 – 7.37 (m, 9H, H22, H61, H63, H70), 7.34 – 7.29 (m, 2H, H69), 7.19 –

7.05 (m, 8H, H15, H16, H41, H42), 6.93 (ddd, J = 15.0, 9.5, 2.2 Hz, 1H, H21), 4.93 – 4.85 (m, 1H, H26),

4.66 – 4.58 (m, 4H, H18, H44), 4.52 – 4.40 (m, 2H, H65), 4.25 (dt, J = 14.1, 7.0 Hz, 1H, H66), 3.93

(dp, J = 16.3, 5.5 Hz, 1H, H52), 3.63 – 3.10 (m, 8H, H25, H27, H51, H53), 3.08 – 2.97 (m, 6H, H19, H45),

2.62 – 2.53 (m, 4H, H13, H39), 2.23 – 2.16 (m, 4H, H4, H30), 1.62 – 1.52 (m, 8H, H5, H12, H31, H38),

1.44 (s, 9H, H1), 1.40 (d, J = 9.9 Hz, 9H, H57), 1.35 – 1.21 (m, 24H, CH2), 1.07 (d, J = 12.1 Hz, 9H,

H59).

13C NMR (151 MHz, CD2Cl2:CD3OD 1:1): δ = 176.30 (C3), 176.29 (C3), 174.96 (C29), 167.18 (C=O),

167.17 (C=O), 166.75 (C=O), 166.71 (C=O), 166.66 (C=O), 166.60 (C=O), 166.44 (C=O), 166.34

(C=O), 157.73 (C55), 155.70 (C64), 155.67 (C64), 144.42 (Cq), 144.40 (Cq), 144.30 (Cq), 143.60 (Cq),

143.30 (Cq), 142.21 (Cq), 142.19 (Cq), 136.69 (C62), 136.67 (C62), 135.03 (C21), 134.92 (C21),

134.60 (Cq), 134.56 (Cq), 134.40 (Cq), 134.34 (Cq), 134.29 (Cq), 134.20 (Cq), 131.13 (C22), 131.10

(C22), 130.73 (CHAr), 129.74 (CHAr), 129.71 (CHAr), 129.51 (CHAr), 128.68 (CHAr), 128.64 (CHAr),

128.56 (CHAr), 127.45 (CHAr), 127.97 (C69), 125.95 (C68), 125.86 (C68), 120.77 (C71), 81.09 (C2),

80.08 (C56), 76.18 (C26), 76.16 (C26), 71.65 (C52), 71.63 (C52), 70.66 (C65), 54.13 (C18/44), 51.81

(C18/44), 47.66 (C66), 44.13 (C25/27/51/53), 43.42 (C25/27/51/53), 43.38 (C25/27/51/53), 40.94 (C25/27/51/53),

40.57 (C25/27/51/53), 40.53 (C25/27/51/53), 36.96 (C4/13/30/39), 36.32 (C4/13/30/39), 36.30 (C4/13/30/39), 36.24

(C4/13/30/39), 35.65 (C19/45), 35.63 (C19/45), 34.53 (C19/45), 34.50 (C19/45), 32.37 (C5/12/31/38), 32.33

(C5/12/31/38), 32.30 (C5/12/31/38), 26.66 (C5/12/31/38), 25.92 (C5/12/31/38), 30.32 (CH2), 30.28 (CH2), 30.24

(CH2), 30.14 (CH2), 30.03 (CH2), 30.00 (CH2), 29.80 (CH2), 28.64 (C57), 28.30 (C1), 27.34 (C59),

19.84 (C58). NOTE: C47 and C48 not observed.

LRMS: (ES+) m/z 804 [M+NH4+K]2+ (100%); 796 [M+H+K]2+ (87%); 1574 [M+Na]+ (28%).

154

HRMS: (ESI+) calculated for C92H126D2N7O13Si [M+NH4]+: 1568.9459; observed: 1568.9440. Δ = –

1.2 ppm.

[α] 𝟐𝟓𝑫

+16.02 (c 8.80, CH2Cl2).

To a solution of 108 (47 mg, 0.036 mmol) in CH2Cl2 (0.5 mL) under N2 was added TBAF (1.1 eq.,

36 µL of a 1.0 M in THF, 0.036 mmol). After 18 hours the reaction was diluted with CH2Cl2 and

washed with NH4Cl (sat. aq.). The combined organics were washed with brine, dried over

Na2SO4 and concentrated under reduced pressure. Preparative TLC (SiO2, CH2Cl2:EtOH 94:6)

gave 116 (26 mg, 0.024 mmol, 66%).

On the NMR timescale, 116 exists as a mixture of rotamers.

1H NMR (600 MHz, CD2Cl2:CD3OD 1:1): δ = 7.75 (ddd, J = 15.1, 5.5, 2.4 Hz, 1H, H21/22), 7.55 –

7.42 (m, 8H, H15+16+41+42), 7.28 (ddd, J = 15.2, 9.0, 2.4 Hz, 1H, H21/22), 5.01 – 4.93 (m, 4H, H18+44),

4.15 – 4.04 (m, 2H, H26+52), 3.74 – 3.63 (m, 4H H25/27/51/53), 3.62 – 3.56 (m, 2H, H25/27/51/53), 3.49 –

3.43 (m, 2H, H25/27/51/53), 3.42 (s, 3H, H19/45), 3.33 (s, 3H, H19/45), 2.97 – 2.89 (m, 4H, H13+39), 2.58

– 2.51 (m, 4H, H4+30), 2.03 – 1.86 (m, 8H, H5+12+31+38), 1.80 – 1.74 (m, 18H, H1+57), 1.69 – 1.59 (m,

24H, CH2).

155

13C NMR (151 MHz, CD2Cl2:CD3OD 1:1): δ = 176.31 (C3), 174.84 (C29), 167.14, 167.11, 166.75,

166.71, 166.58, 166.55, 166.49, 166.45 (C20, C23, C46, C49), 158.08 (C55), 143.56 (C14/17/40/42),

143.55 (C14/17/40/42), 143.24 (C14/17/40/42), 143.23 (C14/17/40/42), 135.16 (C2/3), 135.03 (C2/3), 134.47

(C14/17/40/42), 134.14 (C14/17/40/42), 130.83 (C2/3),, 130.75 (C2/3),, 129.65 (C15/16/41/42), 129.42

(C15/16/41/42), 128.56 (C15/16/41/42), 127.38 (C15/16/41/42), 80.99 (C2), 80.12 (C56), 70.01 (C26/52), 69.98

(C26/52), 69.77 (C26/52), 69.74 (C26/52), 54.06 (C18/44), 51.74 (C18/44), 44.41 (C25/27/51/53), 44.39

(C25/27/51/53), 43.92 (C25/27/51/53), 43.87 (C25/27/51/53), 43.78 (C25/27/51/53), 43.75 (C25/27/51/53), 43.53

(C25/27/51/53), 43.49 (C25/27/51/53), 36.95 (C4+30), 36.27 (C13+39), 36.23 (C13+39), 36.20 (C13+39), 35.64

(C19/45), 35.62 (C19/45), 34.48 (C19/45), 32.27 (C5/12/31/38), 30.24 (CH2), 30.21 (CH2), 30.19 (CH2),

30.16 (CH2), 30.06 (CH2), 29.97 (CH2), 29.74 (CH2), 28.59 (C1/57), 28.27 (C1/57), 26.57 (C5/12/31/38),

25.85 (C5/12/31/38). NOTE: C47 and C48 not observed.

LRMS: (ES)+ m/z 1113 [M+Na]+ (100%).

To a solution of 116 (23 mg, 0.021 mmol) in CH2Cl2 (2 mL) was added Fmoc-Cl (10 eq., 55 mg,

0.21 mmol) and pyridine (10 eq., 17 µL, 0.21 mmol). After 25 hours 1M HCl was added and the

aqueous phase extracted with CH2Cl2. The combined organics were washed with brine, dried

156

over Na2SO4 and concentrated under reduced pressure. Preparative TLC (SiO2, CH2Cl2:EtOH

95:5) gave 117 (18.7 mg, 0.012 mmol, 58%).

On the NMR timescale, 117 exists as a mixture of rotamers.

1H NMR (600 MHz, CD2Cl2:CD3OD 1:1): δ = 7.81 – 7.76 (m, 4H, H65+74), 7.65 – 7.60 (m, 4H,

H62+71), 7.43 – 7.36 (m, 5H, H21+64+73), 7.34 – 7.29 (m, 4H, H63+72), 7.16 – 7.11 (m, 6H, H15+16+41+42),

7.08 – 7.04 (m, 2H, H15+16+41+42), 6.97 – 6.90 (m, 1H, H22), 6.37 (s, 1H, NH), 4.92 – 4.78 (m, 2H,

H26+52), 4.64 – 4.57 (m, 4H, H18+44), 4.53 – 4.40 (m, 4H, H59+68), 4.25 (dt, J = 11.9, 6.5 Hz, 2H,

H60+69), 3.65 – 3.55 (m, 4H, H25+27+51+53), 3.54 – 3.44 (m, 2H, H25+27+51+53), 3.43 – 3.25 (m, 2H,

H25+27+51+53), 3.01 (s, 3H, H19/45), 2.98 (s, 3H, H19/45), 2.61 – 2.51 (m, 4H, H13+39), 2.23 – 2.15 (m,

4H, H4+30), 1.63 – 1.51 (m, 8H, H5+12+31+38), 1.47 – 1.41 (m, 18H, H1+57), 1.34 – 1.22 (m, 24H, CH2).

13C NMR (151 MHz, CD2Cl2:CD3OD 1:1): δ = 176.08 (C29), 174.81 (C3), 167.18 (C16), 167.15 (C16),

166.76 (C20), 166.73 (C20), 166.64 (C23/49), 166.60 (C23/49), 166.58 (C23/49), 166.52 (C23/49), 155.60

(C58+67), 144.39 (C61/70), 144.33 (C61/70), 144.31 (C61/70), 143.60 (C14/39), 143.28 (C14/39), 143.27

(C14/39), 142.20 (C66+75), 142.19 (C66+75), 134.86 (C22), 134.76 (C22), 134.58 (C17/43), 134.22 (C17/43),

131.24 (C21), 131.21 (C21), 129.64 (C15/16/41/42), 129.43 (C15/16/41/42), 128.62 (C15/16/41/42+64+73),

128.60 (C15/16/41/42+64+73), 127.91 (C63+72), 127.47 (C15/16/41/42), 125.83 (C62/71), 125.81 (C62/71),

125.77 (C62/71), 120.70 (C65/74), 120.69 (C65/74), 81.02 (C2+56), 76.55 (C26/52), 76.22 (C26/52), 70.68

(C59/68), 70.63 (C59/68), 55.11 (C18/44), 51.82 (C18/44), 47.78 (C60+69), 47.76 (C60+69), 40.91

(C25/27/51/53), 40.77 (C25/27/51/53), 40.55 (C25/27/51/53), 40.51 (C25/27/51/53), 36.97 (C4/30), 36.34 (C4/30),

36.22 (C13/39), 36.18 (C13/39), 35.58 (C19/45), 34.38 (C19/45), 32.15 (C5/12/31/38), 32.12 (C5/12/31/38),

32.09 (C5/12/31/38), 32.06 (C5/12/31/38), 30.19 (CH2), 30.17 (CH2), 30.14 (CH2), 30.13 (CH2), 30.02

(CH2), 29.95 (CH2), 29.92 (CH2), 29.75 (CH2), 28.62 (C1/57), 28.33 (C1/57), 26.52 (CH2), 25.87 (CH2).

NOTE: C47 and C48 not observed.

LRMS: (ES+) m/z 1559 [M+Na]+ (100%); 1536 [M+H]+ (9%).

157

To a solution of 88 (100 mg, 0.074 mmol) in anhydrous CHCl3 (22 mL) under N2, was added

simultaneously a solution of isophthaloyl chloride (8 eq., 120.2 mg, 0.59 mmol) in anhydrous

CHCl3 (7 mL) and a solution of p-xylylene diamine (8 eq., 80.5 mg, 0.59 mmol) in anhydrous

CHCl3 (7 mL) over 3 hours using syringe pumps. After 3 hours the mixture was filtered through

celite and the residue washed with CHCl3. The combined organic phase was washed with 1M

HCl, brine, dried over MgSO4 and concentrated under reduced pressure. Preparative TLC (SiO2,

EtOAc:MeOH 99:1) gave 87 as a colourless solid (40 mg, 0.019 mmol, 26%).

MP: 91–97 °C.

On the NMR timescale, 87 exists as a mixture of rotamers.

1H NMR (600 MHz, CD2Cl2:CD3OD 1:1): δ = 8.56 (m, 2H, HD), 8.25 – 8.09 (m, 4H, HB), 7.78 (t, J =

5.6 Hz, 2H, H71), 7.68 (m, 5H, H62+A), 7.65 – 7.59 (m, 3H, H68+A), 7.43 – 7.30 (m, 11H,

H21/22+61+63+69+70), 7.17 – 7.09 (m, 4H, H15/16/41), 7.06 (d, J = 6.3 Hz, 2H, H15/16/41), 6.99 – 6.84 (m,

158

10H, H15/16/21/22/41/H), 6.55 (s, 1H, H42), 4.91 – 4.83 (m, 1H, H26), 4.62 – 4.58 (m, 2H, H18/44), 4.55 –

4.32 (m, 8H, H65+F), 4.23 (m, 4H, H18/44+66+F), 4.11 (s, 1H, H18/44), 3.89 (s, 1H, H52), 3.63 – 3.55 (m,

1H, H25/27/51/53), 3.54 – 3.43 (m, 2H, H25/27/51/53), 3.42 – 3.26 (m, 4H, H25/27/51/53), 3.14 – 3.04 (m,

1H, H25/27/51/53), 2.97 (m, 4.5H, H19/45), 2.68 (s, 1.5H, H19/45), 2.63 – 2.47 (m, 4H, H13+39), 2.17 (m,

4H, H4+30), 1.66 – 1.47 (m, 8H, H5+12+31+38), 1.44 (s, 9H, H1), 1.35 (d, J = 15.4 Hz, 9H, H57), 1.32 –

1.11 (m, 24H, H6+7+8+9+10+11+32+33+34+35+36+37), 1.05 (d, J = 4.8 Hz, 9H, H59).

13C NMR (151 MHz, CD2Cl2:CD3OD 1:1): δ = 176.13 (C=O), 174.86 (C=O), 174.85 (C=O), 167.59

(C=O), 167.03 (C=O), 166.65 (C=O), 166.57 (C=O), 166.51 (C=O), 166.43 (C=O), 165.64 (C=O),

157.96 (C=O), 157.92 (C=O), 155.55 (C=O), 155.53 (C=O), 144.28 (Cq), 144.26 (Cq), 144.18 (Cq),

143.54 (Cq), 143.23 (Cq), 142.08 (Cq), 142.07 (Cq), 138.09 (Cq), 138.05 (Cq), 136.51 (CH), 136.50

(CH), 136.48 (CH), 136.46 (CH), 134.93 (CH), 134.81 (CH), 134.63 (Cq), 134.53 (Cq), 134.41 (Cq),

134.17 (Cq), 134.03 (Cq), 133.81 (CB), 132.29 (CH), 132.23 (CH), 132.15 (CH), 131.01 (CH),

130.97 (CH), 130.83 (CH), 130.24 (CH), 130.05 (CH), 129.61 (CH), 129.55 (CH), 129.46 (CH),

129.41 (CH), 129.30 (CH), 128.57 (CH), 128.53 (CH), 128.47 (CH), 127.86 (CH), 127.33 (CH),

126.17 (CH), 125.83 (CH), 125.74 (CH), 125.06 (CD), 125.04 (CD), 120.68 (CH), 80.99 (C2/56),

80.47 (C2/56), 76.05 (C26), 71.17 (C52), 71.07 (C52), 70.54 (C65), 55.14 (C18/44), 54.05 (C18/44), 53.00

(C18/44), 51.74 (C18/44), 47.55 (C66), 44.61 (CF/25/27/51/53), 44.43 (CF/25/27/51/53), 44.29 (CF/25/27/51/53),

43.11 (C25/27/51/53), 40.79 (C25/27/51/53), 40.42 (C25/27/51/53), 36.86 (C4/13/30/39), 36.47 (C19/45), 36.26

(C4/13/30/39), 36.21 (C4/13/30/39), 36.16 (C4/13/30/39), 36.11 (C4/13/30/39), 35.93 (C19/45), 35.61 (C19/45),

34.47 (C19/45), 32.28 (C5/12/31/38), 32.22 (C5/12/31/38), 32.08 (C5/12/31/38), 30.35 (CH2), 30.26 (CH2),

30.19 (CH2), 30.15 (CH2), 30.04 (CH2), 29.95 (CH2), 29.72 (CH2), 28.55 (C57), 28.50 (C57), 27.24

(C1), 26.47 (C59), 25.83 (C5/12/31/38), 19.74 (C58). NOTE: C47 and C48 not observed.

LRMS: (ESI+) m/z 2107 [M+Na]+ (100%).

HRMS: (ESI+) calculated for C124H154D2N11O17Si [M+NH4]+: 2101.1570; observed: 2101.1543. Δ =

–1.3 ppm.

159

[α] 𝟐𝟓𝑫

+7.02 (c 0.57, CH2Cl2).

To a solution of 87 (15 mg, 7.2 µmol) in CHCl3 (0.65 mL) in an open flask was added TFA (0.65

mL). After 3 hours the reaction mixture was concentrated under reduced pressure and the TFA

160

azeotroped with CHCl3. The residue was redissolved in anhydrous DMF (0.2 mL) and was added

via syringe pump to a solution of HATU (3 eq., 8.2 mg, 21.6 mmol) and DIPEA (6 eq., 7 µL, 43.2

µmol) in DMF (2 mL) over 2 hours. Once the addition was complete NH4Cl (sat. aq.) was

added. The aqueous layer was extracted with EtOAc and the combined organics were washed

with LiCl (5% w/w aq.), dried over MgSO4 and concentrated under reduced pressure.

Preparative TLC (SiO2, EtOAc:MeOH 98:2) gave 109 (6.8 mg, 3.6 µmol, 50%).

On the NMR timescale, 109 exists as a mixture of rotamers.

1H NMR (600 MHz, CD2Cl2:CD3OD 1:1): δ = 8.66 – 8.41 (m, 2H, HD), 8.27 – 8.10 (m, 4H, HB), 7.81

– 7.74 (m, 2H, H60), 7.72 – 7.57 (m, 8H, H57+66+A), 7.45 – 7.27 (m, 11H, H2/3+58+59+65+67), 7.20 – 7.09

(m, 4H, H2/3+22+23+48+49+H), 7.08 – 7.01 (m, 2H, H2/3+22+23+48+49+H), 6.98 – 6.82 (m, 10H,

H2/3+22+23+48+49+H), 6.53 (s, 1H, H2/3+22+23+48+49+H), 4.92 – 4.80 (m, 1H, H7), 4.61 (m, 3H, H25/51+F), 4.55

– 4.36 (m, 5H, H54+F), 4.32 – 4.18 (m, 4H, H25/51+55+F), 4.16 – 4.02 (m, 3H, H25/51+F), 3.87 (ddt, J =

17.4, 8.7, 4.2 Hz, 1H, H33), 3.58 (tt, J = 7.0, 4.5 Hz, 1H, H6/8/32/34), 3.54 – 3.44 (m, 1H, H6/8/32/34),

3.43 – 3.36 (m, 1H, H6/8/32/34), 3.36 – 3.17 (m, 5H, H6/8/32/34), 3.05 – 2.62 (m, 6H, H26+52), 2.61 –

2.42 (m, 4H, H20+46), 2.27 – 2.00 (m, 4H, H11+37), 1.64 – 1.39 (m, 8H, H12+19+38+45), 1.38 – 1.08 (m,

24H, H13+14+15+16+17+18+39+40+41+42+43+44), 1.06 (s, 9H, H63).

13C NMR (151 MHz, CD2Cl2:CD3OD 1:1): δ = 176.50 (C=O), 176.38 (C=O), 176.14 (C=O), 176.07

(C=O), 167.75 (CE), 167.69 (CE), 167.08 (C=O), 166.58 (C=O), 166.56 (C=O), 166.53 (C=O), 166.51

(C=O), 166.45 (C=O), 166.35 (C=O), 166.30 (C=O), 166.26 (C=O), 165.50 (C=O), 155.54 (C53),

155.52 (C53), 144.32 (Cq), 144.27 (Cq), 144.23 (Cq), 144.20 (Cq), 143.74 (Cq), 143.51 (Cq), 143.48

(Cq), 143.16 (Cq), 142.11 (Cq), 138.12 (Cq), 138.09 (Cq), 137.97 (Cq), 136.54 (CH), 136.52 (CH),

134.95 (CH), 134.86 (CH), 134.72 (Cq), 134.70 (Cq), 134.67 (Cq), 134.64 (Cq), 134.61 (Cq), 134.59

(Cq), 134.51 (Cq), 134.34 (Cq), 134.28 (Cq), 134.14 (Cq), 134.07 (Cq), 133.84 (Cq), 133.80 (Cq),

133.64 (Cq), 132.30 (CB), 132.23 (CB), 132.15 (CB), 131.03 (CH), 130.94 (CH), 130.20 (CH), 130.05

(CH), 129.70 (CH), 129.67 (CH), 129.61 (CH), 129.58 (CH), 129.54 (CH), 129.51 (CH), 129.49

161

(CH), 129.47 (CH), 129.41 (CH), 129.35 (CH), 128.64 (CH), 128.60 (CH), 128.56 (CH), 128.53

(CH), 127.89 (CH), 127.33 (CH), 126.20 (CH), 125.86 (CH), 125.83 (CH), 125.77 (CH), 125.21 (CD),

125.15 (CD), 120.71, 76.04 (C7), 76.02 (C7), 71.20 (C33), 71.14 (C33), 70.54 (C54), 53.03 (CH2),

52.92 (CH2), 51.76 (CH2), 47.56 (C55), 44.45 (CF), 44.38 (CF), 43.61 (C6/8/32/34), 43.51 (C6/8/32/34),

43.35 (C6/8/32/34), 43.21 (C6/8/32/34), 42.99 (C6/8/32/34), 42.95 (C6/8/32/34), 42.79 (C6/8/32/34), 40.77

(C6/8/32/34), 40.72 (C6/8/32/34), 40.31 (C6/8/32/34), 40.26 (C6/8/32/34), 40.21 (C6/8/32/34), 36.90 (C20/46),

36.86 (C20/46), 36.84 (C20/46), 36.82 (C20/46), 36.52 (C26/52), 36.37 (C26/52), 36.15 (C11/37), 36.12

(C11/37), 36.09 (C11/37), 36.01 (C11/37), 35.94 (C26/52), 35.66 (C26/52), 34.56 (C26/52), 32.21

(C12/19/38/45), 32.19 (C12/19/38/45), 32.16 (C12/19/38/45), 32.13 (C12/19/38/45), 31.95 (C12/19/38/45), 30.22

(CH2), 30.20 (CH2), 30.19 (CH2), 30.13 (CH2), 30.08 (CH2), 29.99 (CH2), 29.97 (CH2), 29.95 (CH2),

29.91 (CH2), 29.90 (CH2), 29.87 (CH2), 29.84 (CH2), 29.80 (CH2), 29.76 (CH2), 29.73 (CH2), 29.63

(CH2), 27.26 (C63), 26.55 (C12/19/38/45), 26.51 (C12/19/38/45), 26.49 (C12/19/38/45), 26.42 (C12/19/38/45),

19.74 (C62). NOTE: C28 and C29 not observed.

HRMS: (NSI+) calculated for C115H136D2N11O14Si [M+NH4]+: 1927.0314; observed: 1927.0282. Δ =

-1.7 ppm.

162

A solution of 109 (38.9 mg, 20.4 µmol) in HCl (4 mL, 1.25M in MeOH) in an open flask was

stirred at r.t.. After 4.5 hours the mixture was diluted with CH2Cl2, washed with brine, dried

over Na2SO4 and concentrated under reduced pressure. Preparative TLC (SiO2, CH2Cl2:EtOH

96:4) gave 110 (34.0 mg, 20.3 µmol, quant.).

On the NMR timescale, 110 exists as a mixture of rotamers and positional isomers.

1H NMR (600 MHz, CD2Cl2:CD3OD 1:1): δ = 8.74 – 8.54 (m, 2H, HD), 8.26 – 8.08 (m, 4H, HB), 7.76

(dd, J = 12.2, 5.4 Hz, 2H, H60), 7.67 – 7.50 (m, 4H, H57+A), 7.38 (m, 2H, H59), 7.34 – 7.20 (m, 2H,

H58), 7.17 – 6.84 (m, 16.6H, H2FumD2+3FumD2+22+23+48+49+H), 6.55 (br s, 1H, H23/49), 6.13 – 5.82 (br s,

0.4H, H2FumH2+3FumH2), 4.81 (d, J = 5.1 Hz, 1H, H7), 4.66 – 4.40 (m, 11H, HF/25/51/54), 4.40 – 4.26 (m,

3H, HF/25/51/54), 4.22 (s, 1H, H55), 3.82 – 3.71 (m, 1H, H33), 3.57 (ddd, J = 14.5, 12.6, 3.8 Hz, 1H,

H6/8/32/34), 3.49 – 3.15 (m, 7H, H6/8/32/34), 3.02 – 2.78 (m, 6H, H26+52), 2.56 (ddd, J = 19.1, 14.6, 7.1

Hz, 4H, H20+46), 2.22 – 2.00 (m, 4H, H11+37), 1.63 – 1.41 (m, 8H, H12+19+38+45), 1.35 – 1.03 (m, 24H,

H13+14+15+16+17+18+39+40+41+42+43+44).

163

13C NMR (151 MHz, CD2Cl2:CD3OD 1:1): δ = 176.27 (C=O), 176.22 (C=O), 176.20 (C=O), 167.57

(C=O), 167.54 (C=O), 167.52 (C=O), 167.45 (C=O), 166.80 (C=O), 166.70 (C=O), 166.61 (C=O),

166.52 (C=O), 155.41 (C53), 144.10 (Cq), 143.98 (Cq), 143.50 (Cq), 143.47 (Cq), 142.04 (Cq),

142.02 (Cq), 138.14 (Cq), 137.98 (Cq), 134.59 (Cq), 134.57 (Cq), 134.55 (Cq), 134.52 (Cq), 134.48

(Cq), 134.28 (Cq), 134.26 (Cq), 132.18 (CH), 132.14 (CH), 130.14 (CH), 130.06 (CH), 129.96 (CH),

129.67 (CH), 129.56 (CH), 129.45 (CH), 128.86 (CH), 128.55 (CH), 127.82 (CH), 127.79 (CH),

125.60 (CH), 125.00 (CH), 120.67 (CH), 75.96 (C7), 75.92 (C7), 70.45 (C54), 69.69 (C33), 52.39

(C25/51), 47.47 (C55), 44.38 (CF), 44.30 (CF), 43.77 (C6/8/32/34), 43.73 (C6/8/32/34), 43.53 (C6/8/32/34),

43.51 (C6/8/32/34), 40.70 (C6/8/32/34), 40.63 (C6/8/32/34), 40.27 (C6/8/32/34), 36.83 (C11/37), 36.77 (C11/37),

36.05 (C20/26/46/52), 36.02 (C20/26/46/52), 36.00 (C20/26/46/52), 35.12 (C26/52), 34.61 (C26/52), 34.55

(C26/52), 32.11 (C12/19/38/45), 32.10 (C12/19/38/45), 32.06 (C12/19/38/45), 31.98 (C12/19/38/45), 31.95

(C12/19/38/45), 30.33 (CH2), 30.31 (CH2), 30.09 (CH2), 30.05 (CH2), 29.99 (CH2), 29.97 (CH2), 29.92

(CH2), 29.89 (CH2), 29.88 (CH2), 29.85 (CH2), 29.83 (CH2), 29.81 (CH2), 29.77 (CH2), 29.75 (CH2),

29.73 (CH2), 29.60 (CH2), 26.43 (CH2), 26.41 (C12/19/38/45). NOTE: C28 and C29 not observed.

LRMS: (NSI+) m/z 836 [M+2H]2+ (100%); 1695 [M+Na]+ (3%).

HRMS: (NSI+) calculated for C99H116D2N10O14 [M+2H]2+: 836.4493; observed: 836.4472. Δ = –2.5

ppm.

164

To a solution of 110 (42.1 mg, 25.2 µmol) in CH2Cl2 (7.5 mL) was added (R)-81 (6 eq., 95.2 mg,

0.15 mmol) and Fmoc-Cl (5 eq., 32.5 mg, 0.13 mmol). After 19 hours 1M HCl was added and

the aqueous phase extracted with CH2Cl2. The combined organics were washed with brine,

dried over Na2SO4 and concentrated under reduced pressure. Preparative TLC (SiO2,

CH2Cl2:EtOH 94:6) gave 58 (44.3 mg, 23.4 µmol, 93%).

On the NMR timescale, 58 exists as a mixture of rotamers and positional isomers.

1H NMR (600 MHz, CD2Cl2:CD3OD 1:1): δ = 8.56 – 8.37 (m, 2H, HD), 7.98 (dt, J = 12.2, 6.4 Hz, 4H,

HB), 7.59 (ddd, J = 28.2, 14.7, 6.5 Hz, 4H, H60+69), 7.51 – 7.30 (m, 6H, H57+66+A), 7.27 – 7.17 (m,

4H, H59+68), 7.11 (ddd, J = 19.3, 15.0, 7.2 Hz, 4H, H58+67), 7.02 – 6.66 (m, 15.4H, H2+3+22+23+48+49+H),

6.39 (s, 1H, H2+3+22+23+48+49+H), 6.00 – 5.67 (m, 1.6H, H2+3+22+23+48+49+H), 4.77 – 4.62 (m, 2H, H7+33),

4.47 – 4.39 (m, 3H, H25/51/54/63/F), 4.38 – 4.13 (m, 12H, H25/51/54/63/F), 4.07 (dd, J = 19.1, 9.6 Hz, 1H,

H55/64), 3.99 (dd, J = 16.9, 9.2 Hz, 1H, H55/64), 3.94 (d, J = 13.4 Hz, 1H, H25/51), 3.46 – 3.37 (m, 1H,

165

H6/8/32/34), 3.36 – 3.27 (m, 2H, H6/8/32/34), 3.22 (ddd, J = 18.1, 10.9, 5.5 Hz, 3H, H6/8/32/34), 3.13 –

3.00 (m, 2H, H6/8/32/34), 2.83 (t, J = 5.3 Hz, 3H, H26/52), 2.76 (d, J = 15.4 Hz, 1.5H, H26/52), 2.52 (d, J

= 21.2 Hz, 1.5H, H26/52), 2.39 (dt, J = 14.6, 6.2 Hz, 2H, H20/46), 2.33 (t, J = 7.5 Hz, 2H, H20/46), 2.06 –

1.85 (m, 4H, H11+37), 1.45 – 1.27 (m, 8H, H12+19+38+45), 1.17 – 0.92 (m, 24H,

H13+14+15+16+17+18+39+40+41+42+43+44).

Figure S2. Partial 1H NMR spectra (600 MHz, CD2Cl2, 300 K) of (a) thread 117 and (b) catenane 58. The lettering

corresponds to proton labelling in Scheme 32.

In the 1H NMR spectrum of 58 there is a significant upfield shift of the fumaramide protons (H2

+ H3) compared to those in non-interlocked thread 117 owing to shielding by the macrocycle. A

further significant difference is the large upfield shift of H23, as observed with rotaxane 69.

13C NMR (151 MHz, CD2Cl2:CD3OD 1:1): δ = 176.52 (C=O), 176.50 (C=O), 176.46 (C=O), 176.41

(C=O), 176.24 (C=O), 176.17 (C=O), 167.77 (C=O), 167.75 (C=O), 167.72 (C=O), 167.61 (C=O),

167.16 (C=O), 167.12 (C=O), 167.05 (C=O), 166.98 (C=O), 166.61 (C=O), 166.59 (C=O), 166.55

(C=O), 165.79 (C=O), 165.74 (C=O), 155.64 (C=O), 155.60 (C=O), 144.39 (Cq), 144.34 (Cq), 144.30

(Cq), 144.28 (Cq), 144.26 (Cq), 144.20 (Cq), 144.04 (Cq), 143.99 (Cq), 143.80 (Cq), 143.57 (Cq),

143.51 (Cq), 143.23 (Cq), 143.21 (Cq), 142.20 (Cq), 142.18 (Cq), 142.17 (Cq), 138.41 (Cq), 138.36

(Cq), 138.10 (Cq), 134.80 (Cq), 134.72 (Cq), 134.64 (Cq), 134.61 (Cq), 134.56 (Cq), 134.43 (Cq),

134.37 (Cq), 134.20 (Cq), 133.78 (CH), 133.49 (Cq), 133.06 (Cq), 132.28 (Cq), 132.23 (Cq), 132.15

(Cq), 131.11 (Cq), 130.26 (Cq), 130.08 (Cq), 129.85 (CH), 129.77 (CH), 129.72 (CH), 129.62 (CH),

166

129.58 (CH), 129.55 (CH), 129.53 (CH), 129.45 (CH), 128.68 (CH), 128.66 (CH), 128.57 (CH),

127.95 (CH), 127.87 (CH), 127.36 (CH), 126.32 (CH), 125.93 (CH), 125.89 (CH), 125.83 (CH),

125.65 (CH), 125.60 (CH), 125.57 (CH), 125.28 (CH), 125.23 (CH), 120.81 (CH), 120.79 (CH),

120.77 (CH), 120.76 (CH), 76.28 (C7/33), 76.23 (C7/33), 76.16 (C7/33), 76.14 (C7/33), 70.61 (C54),

70.56 (C54), 70.55 (C54), 70.52 (C54), 70.51 (C54), 54.16 (C25/51), 53.14 (C25/51), 53.06 (C25/51), 51.79

(C25/51), 47.62 (C55), 47.59 (C55), 44.50 (CF), 44.34 (CF), 41.08 (C6/8/32/34), 40.97 (C6/8/32/34), 40.85

(C6/8/32/34), 40.64 (C6/8/32/34), 40.60 (C6/8/32/34), 40.57 (C6/8/32/34), 40.48 (C6/8/32/34), 40.44 (C6/8/32/34),

40.34 (C6/8/32/34), 36.88 (C11/37), 36.82 (C11/37), 36.67 (C26/52), 36.55 (C26/52), 36.19 (C20/46), 36.16

(C20/46), 36.12 (C20/46), 36.11 (C20/46), 36.08 (C20/46), 35.66 (C26/52), 34.58 (C26/52), 32.23

(C12/19/38/45), 32.20 (C12/19/38/45), 32.12 (C12/19/38/45), 32.08 (C12/19/38/45), 32.03 (C12/19/38/45), 32.02

(C12/19/38/45), 30.22 (CH2), 30.19 (CH2), 30.14 (CH2), 30.13 (CH2), 30.08 (CH2), 30.03 (CH2), 29.98

(CH2), 29.95 (CH2), 29.93 (CH2), 29.90 (CH2), 29.88 (CH2), 29.84 (CH2), 29.83 (CH2), 29.79 (CH2),

29.70 (CH2), 29.68 (CH2), 29.61 (CH2), 26.64 (C12/19/38/45), 26.60 (C12/19/38/45), 26.54 (C12/19/38/45),

26.47 (C12/19/38/45). NOTE: C28 and C29 not observed.

LRMS: (NSI+) m/z 948 [M+2H]2+ (71%); 970 [M+2Na]2+ (100%); 1917 [M+Na]+ (90%).

HRMS: (NSI+) calculated for C114H124D2N10O16Na [M+Na]+: 1915.9371; observed: 1915.9356. Δ =

–0.8 ppm.

167

[α]𝟐𝟓𝑫

–2.51 (c 0.28, CH2Cl2).

To a solution of 112 (27 mg, 0.115 mmol) in CH2Cl2 (2.5 mL) was added oxalyl chloride (3 eq.,

28 µL, 0.345 mmol). After 3 hours the mixture was concentrated under reduced pressure and

azeotroped with toluene. The residue was redissolved in CH2Cl2 (0.63 mL) and a portion (0.5

mL, 0.0913 mmol) added to a solution of 110 (30.7 mg, 18.4 µmol) and Pyridine (9 eq., 14 µL,

0.173 mmol) in CH2Cl2 (3 mL). After 18 hours 1M HCl was added and the aqueous phase

extracted with CH2Cl2. The combined organics were washed with brine, dried over Na2SO4 and

concentrated under reduced pressure. Preparative TLC (SiO2, CH2Cl2:EtOH 94:6) gave 111 (32.1

mg, 17.0 µmol, 92%).

On the NMR timescale, 111 exists as a mixture of rotamers and positional isomers.

168

1H NMR (600 MHz, CD2Cl2:CD3OD 1:1): δ = 8.68 – 8.47 (m, 2H, HD), 8.20 – 8.08 (m, 4H, HB), 7.81

– 7.70 (m, 4H, H60+70), 7.67 – 7.46 (m, 6H, H57+67+A), 7.44 – 7.21 (m, 8H, H58+59+69+69), 7.18 – 6.86

(m, 16H, H2/3/22/23/48/49/H), 6.54 (s, 1H, H2/3/22/23/48/49/H), 6.12 – 5.84 (m, 1H, H2/3/22/23/48/49/H), 4.90 –

4.74 (m, 2H, H7+33), 4.64 – 4.15 (m, 15H, H25+51+54+55+F), 4.12 – 4.02 (m, 1H, H65), 3.67 – 3.09 (m,

8H, H6+8+32+34), 3.05 – 2.97 (m, 3H, H26/52), 2.95 – 2.87 (m, 1.5H, H26/52), 2.72 – 2.62 (m, 1.5H,

H26/52), 2.61 – 2.47 (m, 4H, H20+46), 2.46 – 2.35 (m, 2H, H64), 2.23 – 2.02 (m, 4H, H11+37), 2.00 –

1.89 (m, 2H, H63), 1.63 – 1.41 (m, 8H, H12+19+38+45), 1.36 – 1.08 (m, 24H,

H13+14+15+16+17+18+39+40+41+42+43+44).

13C NMR (151 MHz, CD2Cl2:CD3OD 1:1): δ = 176.39 (C=O), 176.23 (C=O), 176.13 (C=O), 176.00

(C=O), 175.95 (C=O), 173.99 (C=O), 167.59 (C=O), 167.57 (C=O), 167.48 (C=O), 166.97 (C=O),

166.93 (C=O), 166.47 (C=O), 166.39 (C=O), 166.32 (C=O), 165.54 (C=O), 155.43 (C=O), 155.40

(C=O), 146.87 (Cq), 146.66 (Cq), 144.13 (Cq), 143.87 (Cq), 143.73 (Cq), 143.50 (Cq), 143.14 (Cq),

142.06 (Cq), 142.04 (Cq), 141.98 (Cq), 141.97 (Cq), 138.26 (Cq), 138.21 (Cq), 138.15 (Cq), 138.01

(Cq), 137.96 (Cq), 134.88 (CH), 134.78 (CH), 134.64 (CH), 134.58 (CH), 134.49 (CH), 132.94 (CH),

132.19 (CH), 132.12 (CH), 132.09 (CH), 132.04 (CH), 131.99 (CH), 130.94 (CH), 130.13 (CH),

129.95 (CH), 129.70 (CH), 129.63 (CH), 129.60 (CH), 129.57 (CH), 129.52 (CH), 129.50 (CH),

129.43 (CH), 129.41 (CH), 128.54 (CH), 128.49 (CH), 128.45 (CH), 128.42 (CH), 128.00 (CH),

127.98 (CH), 127.93 (CH), 127.83 (CH), 127.75 (CH), 127.25 (CH), 126.20 (CH), 125.77 (CH),

125.74 (CH), 125.69 (CH), 125.52 (CH), 125.46 (CH), 125.14 (CH), 125.09 (CH), 125.03 (CH),

124.93 (CH), 124.84 (CH), 124.82 (CH), 120.70 (CH), 120.68 (CH), 120.65 (CH), 120.52 (CH),

120.51 (CH), 120.48 (CH), 75.99 (C7/33), 75.94 (C7/33), 75.90 (C7/33), 71.83 (C7/33), 71.78 (C7/33),

71.75 (C7/33), 70.47 (C54), 70.44 (C54), 70.40 (C54), 52.95 (C25/51), 51.72 (C25/51), 47.51 (C55), 47.47

(C55), 46.97 (C65), 46.84 (C65), 44.38 (CF), 44.36 (CF), 44.30 (CF), 40.65 (C6/8/32/34), 40.59 (C6/8/32/34),

40.56 (C6/8/32/34), 40.50 (C6/8/32/34), 36.83 (C11+37), 36.76 (C11+37), 36.73 (C11+37), 36.50 (C26/52),

36.39 (C26/52), 36.08 (C20/46), 36.05 (C20/46) (C20/46), 36.00 (C20/46), 35.66 (C26/52), 34.52 (C26/52),

32.12 (C12/19/38/45), 32.03 (C12/19/38/45), 31.96 (C12/19/38/45), 30.13 (CH2+C63), 30.09 (CH2+C63), 30.03

169

(CH2+C63), 30.00 (CH2+C63), 29.83 (CH2+C63), 29.80 (CH2+C63), 29.78 (CH2+C63), 29.73 (CH2+C63),

29.65 (CH2+C63), 28.01 (C64), 27.78 (C64), 26.44 (C12/19/38/45), 26.39 (C12/19/38/45), 26.32 (C12/19/38/45).

NOTE: C28 and C29 not observed.

LRMS: (NSI+) m/z 947 [M+2H]2+ (100%); 969 [M+2Na]2+ (48%); 958 [M+H+Na]2+ (44%); 1915

[M+Na]+ (5%).

HRMS: (NSI+) calculated for C115H127D2N10O15 [M+H]+: 1891.9759; observed: 1891.9800. Δ = 2.2

ppm.

170

To a solution of 111 (34.0 mg 18.0 µmol) in CH2Cl2 (0.4 mL) was added NEt3 (5 eq., 13 µL 90

µmol). After 18 hours 1M HCl was added and the aqueous phase extracted with CH2Cl2. The

combined organics were washed with brine, dried over Na2SO4 and concentrated under

reduced pressure. Preparative TLC (SiO2, CH2Cl2:EtOH 95:5) gave 114 (20.4 mg, 12.2 µmol,

68%).

On the NMR timescale, 114 exists as a mixture of rotamers and positional isomers.

1H NMR (600 MHz, CD2Cl2:CD3OD 1:1): δ = 8.76 – 8.50 (m, 2H, HD), 8.25 – 8.09 (m, 4H, HB), 7.75

(s, 2H, H62), 7.71 – 7.58 (m, 2H, HA), 7.50 (br s, 2H, H59), 7.41 – 7.25 (m, 4H, H60+61), 7.20 – 6.78

(m, 16H, H22+23+48+49+H), 4.87 (dd, J = 11.1, 5.6 Hz, 1H, H33), 4.58 – 4.42 (m, 4H HF), 4.41 – 4.27

(m, 4H, HF), 4.12 – 3.98 (m, 1H, H57), 3.77 (dd, J = 10.3, 4.8 Hz, 1H, H7), 3.45 – 3.14 (m, 8H,

H6+8+32+34), 3.04 – 2.76 (m, 6H, H26+52), 2.63 – 2.47 (m, 4H, H20+46), 2.41 (br s, 2H, H56), 2.23 – 2.01

(m, 4H, H11+37), 1.95 (br s, 2H, H55), 1.65 – 1.41 (m, 8H, H12+19+38+45), 1.36 – 1.05 (m, 24H,

H13+14+15+16+17+18+39+40+41+42+43+44). NOTE: H2 + H3 not observed. Macrocycle shuttling is fast on nmr

timescale causing broadening of these signals.

171

13C NMR (151 MHz, CD2Cl2:CD3OD 1:1): δ = 176.36 (C=O), 176.28 (C=O), 176.09 (C=O), 174.08

(C=O), 174.05 (C=O), 167.64 (C=O), 167.61 (C=O), 166.58 (C=O), 146.82 (Cq), 143.51 (Cq), 141.96

(Cq), 138.16 (Cq), 138.06 (Cq), 134.62 (Cq), 134.5 (Cq), 134.35 (Cq), 132.20 (CH), 132.16 (CH),

130.17 (CH), 130.08 (CH), 130.00 (CH), 129.68 (CH), 129.60 (CH), 129.49 (CH), 128.96 (CH),

128.90 (CH), 128.00 (CH), 127.79 (CH), 125.16 (CH), 125.13 (CH), 125.01 (CH), 124.97 (CH),

120.54 (CH), 71.88 (C33), 71.84 (C33), 69.77 (C7), 52.33 (C25/51), 46.94 (C57), 44.38 (CF), 43.85

(C6/8/32/34), 43.80 (C6/8/32/34), 43.64 (C6/8/32/34), 40.84 (C6/8/32/34), 40.33 (C6/8/32/34), 36.85 (C11/37),

36.79 (C11/37), 36.09 (C20/46), 36.06 (C20/46), 35.89 (C26/52)*, 34.98 (C26/52)*, 32.15 (C12/19/38/45/55),

32.12 (C12/19/38/45/55), 32.04 (C12/19/38/45/55), 30.35 (CH2), 30.12 (CH2), 30.03 (CH2), 29.93 (CH2),

29.87 (CH2), 29.85 (CH2), 29.79 (CH2), 29.73 (CH2), 29.65 (CH2), 27.95 (C56), 26.48 (C12/19/38/45).

*Determined by HSQC. NOTE: C28 and C29 not observed.

LRMS: (NSI+) m/z 836 [M+2H]2+ (100%); 847 [M+H+Na]2+ (19%); 858 [M+2Na]2+ (18%); 1693

[M+Na]+ (5%).

HRMS: (NSI+) calculated for C100H117D2N10O13 [M+H+]: 1669.9078; observed: 1669.9066. Δ = –

0.7 ppm; calculated for C100H118D2N10O13 [M+2H]2+: 835.4575; observed: 835.4576. Δ = 0.1 ppm.

172

To a solution of 58 (2 mg 1.1 µmol) in CH2Cl2 (0.2 mL) was added NEt3 (6.55 eq., 1 µL, 7.2

µmol). After 18 hours 1M HCl was added and the aqueous phase extracted with CH2Cl2. The

combined organics were washed with brine, dried over Na2SO4 and concentrated under

reduced pressure. Preparative TLC (SiO2, CH2Cl2:EtOH 93:7) gave 115 (1.3 mg, 0.9 µmol, 82%).

On the NMR timescale, 115 exists as a mixture of rotamers and positional isomers.

1H NMR (600 MHz, CD2Cl2:CD3OD 1:1): δ = 8.74 – 8.63 (m, 2H, HD), 8.21 – 8.09 (m, 4H, HB), 7.69

– 7.58 (m, 2H, HA), 7.16 – 7.11 (m, 8H, H22+23+48+49), 7.10 – 6.98 (m, 8H, HH), 4.58 – 4.44 (m, 6H,

HF+25/51), 4.41 – 4.30 (m, 6H, HF+25/51), 3.78 (tt, J = 11.6, 5.7 Hz, 2H, H7+33), 3.35 (dd, J = 12.8, 4.3

Hz, 1H, H6/8/32/34), 3.33 – 3.16 (m, 7H, H6/8/32/34), 3.01 – 2.94 (m, 3H, H26/52), 2.89 (s, 3H, H26/52),

2.56 (dt, J = 22.1, 7.5 Hz, 4H, H20+46), 2.21 – 2.09 (m, 4H, H11+37), 1.54 (ddd, J = 20.5, 13.8, 7.1 Hz,

8H, H12+19+38+45), 1.36 – 1.11 (m, 24H, H13+14+15+16+17+18+39+40+41+42+43+44). NOTE: H2 + H3 not

observed. Macrocycle shuttling is fast on nmr timescale causing broadening of these signals.

13C NMR (151 MHz, CD2Cl2:CD3OD 1:1): δ = 176.38 (C=O), 176.32 (C=O), 176.29 (C=O), 167.69

(C=O), 167.65 (C=O), 167.58 (C=O), 166.89 (C=O), 166.72 (C=O), 166.63 (C=O), 166.28 (C=O),

143.56 (Cq), 143.51 (Cq), 143.47 (Cq), 138.15 (Cq), 138.12 (Cq), 138.08 (Cq), 134.65 (Cq), 134.62

173

(Cq), 134.60 (Cq), 134.57 (Cq), 134.55 (Cq), 134.53 (Cq), 134.41 (Cq), 134.39 (Cq), 132.23 (CB),

132.20 (CB), 132.17 (CB), 130.02 (CH), 129.82 (CH), 129.79 (CH), 129.76 (CH), 129.62 (CH),

129.51 (CH), 129.02 (CH), 128.95 (CH), 126.83 (CH), 125.13 (CD), 69.74 (C7+33), 52.43 (C25/51),

44.41 (CF), 44.38 (CF), 43.89 (C6/8/32/34), 43.84 (C6/8/32/34), 43.65 (C6/8/32/34), 43.63 (C6/8/32/34), 36.85

(C11/37), 36.24 (C26/52), 36.16 (C26/52), 36.09 (C20/46), 36.06 (C20/46), 35.18 (C26/52), 32.15

(C11/20/37/46), 32.09 (C11/20/37/46), 32.03 (C11/20/37/46), 30.12 (CH2), 30.10 (CH2), 30.06 (CH2), 30.03

(CH2), 29.99 (CH2), 29.94 (CH2), 29.90 (CH2), 29.87 (CH2), 29.85 (CH2), 29.84 (CH2), 29.78 (CH2),

29.61 (CH2), 29.60 (CH2), 26.50 (C11/20/37/46). NOTE: C28 and C29 not observed.

LRMS: (NSI+) m/z 725 [M+2H]2+ (100%); 736 [M+H+Na]2+ (19%); 747 [M+2Na]2+ (16%).

HRMS: (NSI+) calculated for C84H105D2N10O12 [M+H]+: 1449.8190; observed: 1449.8187. Δ = –0.2

ppm; calculated for C84H106D2N10O12 [M+2H]2+: 725.4131; observed: 725.4131. Δ = 0.0 ppm.

174

A solution of Fluorene (1 g, 6 mmol) in THF (7 mL) under N2 was cooled to –5 °C. n-BuLi (1 eq.,

2.4 mL, 2.5M in hexane) was added slowly while maintaining the reaction temperature at –5

°C. The solution was stirred for 10 minutes before Bromopropionic acid (0.76 eq., 703 mg, 4.6

mmol) was added. The reaction was allowed to warm to r.t. and stirring was continued for 5

hours then NH4Cl was added and the aqueous layer was extracted with EtOAc. The combined

organic layers were dried over MgSO4 and concentrated under reduced pressure. The residue

was triturated with hexane to give 112 (498.9 mg, 35%) as a yellow powder.

MP: 132–137 °C.

1H NMR (600 MHz, CDCl3): δ = 7.75 (d, J = 7.5 Hz, 2H, H2), 7.50 (d, J = 7.4 Hz, 2H, H5), 7.37 (t, J =

7.4 Hz, 2H, H3), 7.31 (t, J = 7.3 Hz, 2H, H4), 4.09 (t, J = 5.1 Hz, 1H, H7), 2.43 (dt, J = 8.1, 5.7 Hz,2H,

H8), 2.00 – 1.95 (m, 2H, H9).

13C NMR (151 MHz, CDCl3): δ = 179.18 (C10), 145.80 (C6), 141.32 (C1), 127.36 (C3), 127.12 (C4),

124.26 (C5), 120.00 (C2), 46.10 (C7), 29.10 (C9), 27.24 (C8).

LRMS: (ES–) m/z 237 [M–H]– (100%).

HRMS: (ES–) calculated for C16H13O2 [M–H]–: 237.0916; observed: 237.0914. Δ = –0.8 ppm.

General methods for protection of molecular information ratchet 114.

To a solution of 114 (5 mg, 3.0 µmol) in CH2Cl2 (0.5 mL) was added Fmoc-Cl (10 eq., 7.7 mg,

29.8 µmol) and pyridine (10 eq., 2.4 µL, 30 µmol). After 24 hours, 1M HCl (aq.) was added and

the aqueous layer was extracted with CH2Cl2. The combined organic layers were washed with

brine, dried over Na2SO4 then concentrated under reduced pressure. Purification by

preparative TLC (SiO2, CH2Cl2:EtOH 96:4) gave 111 which was submitted for NMR analysis.

175

To a solution of 114 (10 mg, 6.0 µmol) in CH2Cl2 (0.5 mL) was added Fmoc-Cl (5 eq., 7.7 mg,

29.8 µmol) and acylation catalyst (R)-81 (5 eq., 19.0 mg, 30.1 µmol) or (S)-81 (5 eq., 19.0 mg,

30.1 µmol). After 24 hours, 1M HCl (aq.) was added and the aqueous layer was extracted with

CH2Cl2. The combined organic layers were washed with brine, dried over Na2SO4 then

concentrated under reduced pressure. Purification by preparative TLC (SiO2, CH2Cl2:EtOH 96:4)

gave 111 which was submitted for NMR analysis.

Method for the partial deprotection of molecular information ratchet

111.

To a solution of 111 (4.5 mg, 2.4 µmol) in CH2Cl2 (0.3 mL) was added NEt3 (10.75 eq., 3.6 µL,

25.8 µmol). After 1 hour, the reaction was diluted with CH2Cl2, 1M HCl (aq.) was added and the

aqueous layer was extracted with CH2Cl2. The combined organic layers were washed with

brine, dried over Na2SO4 then concentrated under reduced pressure. Purification by

preparative TLC (SiO2, CH2Cl2:EtOH 96:4) gave 111 which was submitted for NMR analysis.

General methods for protection of molecular information ratchet 110.

To a solution of 110 (35 mg, 20.9 µmol) in CH2Cl2 (6.3 mL) was added Fmoc-Cl (5 eq., 27 mg,

0.10 mmol) and (R)-81 (5 eq., 65.8 mg, 0.10 mmol). After 24 hours, 1M HCl (aq.) was added

and the aqueous layer was extracted with CH2Cl2. The combined organic layers were washed

with brine, dried over Na2SO4 then concentrated under reduced pressure. Purification by

preparative TLC (SiO2, CH2Cl2:EtOH 96:4) gave 58 which was submitted for NMR analysis.

176

General method for autonomous operation of molecular information

ratchet 58.

To a solution of 58 (5 mg, 2.6 µmol) in CH2Cl2 (0.3 mL) was added (R)-81 (5 eq., 8.2 mg, 13.0

µmol) and KHCO3 (20 eq., 5.2 mg, 52 µmol). A solution of Fmoc-Cl (240 mg, 0.93 mmol) in

CH2Cl2 (1.0 mL) was added at a rate of 6.7 µL/h. After 1 hour NEt3 (1.5 eq., 0.55 µL, 3.9 µmol)

was added and Fmoc-Cl addition was continued. After the given amount of time 1M HCl (aq.)

was added and the aqueous layer was extracted with CH2Cl2. The combined organic layers

were washed with brine, dried over Na2SO4 then concentrated under reduced pressure.

Purification by preparative TLC (SiO2, CH2Cl2:EtOH 95:5) gave 58 which was submitted for NMR

analysis.

177

Figure S3: Autonomous operation cycle of catenane motor 58.

178

3.7. References

1 A. G. Johnston, D. A. Leigh, A. Murphy, J. P. Smart, M. D. Deegan, J. Am. Chem. Soc.

1996, 118, 10662–10663.

2 F. G. Gatti, D. A. Leigh, S. A. Nepogodiev, A. M. Z. Slawin, S. J. Teat, J. K. Y. Wong, J. Am.

Chem. Soc. 2001, 123, 5983–5989.

3 A. Carlone, S. M. Goldup, N. Lebrasseur, D. A. Leigh, A. Wilson, J. Am. Chem. Soc. 2012,

134, 8321–8323.

4 K. M. Lee, K. Ramalingam, J. K. Son, R. W. Woodard, J. Org. Chem. 1989, 54, 3195–3198.

5 S. Niwayama, J. Org. Chem. 2000, 65, 5834–5836.

6 M. Alvarez-Pérez, S. M. Goldup, D. A. Leigh, A. M. Z. Slawin, J. Am. Chem. Soc. 2008,

130, 1836–1838.