Introduction to Spectroscopic methods

Large group of analytical methods based on atomic and molecular spectroscopy

Spectroscopy?The science that deals with interactions of

various types of radiation with matter. (electromagnetic radiation & matter)

Electromagnetic radiation?Wave particle duality Electromagnetic spectrum

Electromagnetic SpectrumEncompasses an enormous range of

wavelength and frequency (energy)

Energy states of chemical species1. Atoms, ions & molecules can exist only in

certain discrete states, characterized by definite amounts of energy. When species change states, it absorbs/emits an amount of E exactly equal to the E difference between the states.

2. When atoms, ions & molecules absorb/emit radiation , the frequency, , or the wavelength , of the radiation is related to the energy difference between the states by:

E1 - E2 = h = hc/l

UV and Visibile Spectroscopy

How does it work?

6

How does it work?A beam of UV and VIS light source is

separated into its components by a prism/diffraction grating.

The monochromatic beam is then split into two beams of equal intensity by a half mirror.

One passes through cuvette containing sample solution and the other through another cuvette containing only solvent

Intensities of two beams measured and compared – difference indicates absorption and presence of functional group

7

BEER’S LAW-ABSORPTION LAW

Absorption is proportional to the number of absorbing molecules.

The amount of UV light absorbed is expressed as the samples molar absorptivity (), defined by the equation:

= e A/bc Molar absorptivity is a physical constant, characteristic of the

paritcular substance being observed and thus characteristic of the particular p electron system in the molecule.

0I Ic

b

Transmitted radiation

Incident

radiation

Pathlength concentration

BEER’S LAW-ABSORPTION LAW…continue

Note: in absorption spectrum, the absorbance varies with wavelength in direct proportion to a (b & c are held constant). The product of the absorptivity and the molecular weight of the absorbing species is called the molar absorptivity,

Thus log I0/I =

bcA

)( 11 Lmolcm

Example

Amines, RNH2 react with picric acid to form amine picrates, which absorb strongly at 359 nm (ε=1.25 x 104 ). An unknown amine (0.1155 g) is dissolved in water and dilute to 100 mL. A 1 ml aliquot of this is diluted to 250 ml for measurement. If this final solution exhibit an absorbance of 0.454 at 359 nm using a 1.00 cm cell, what is the formula weight of the amine? What is a probable formula?

Solution:

in original flask

bcA ccmLmolcm 00.11025.1454.0 114

Lmolc /1063.3 5

mL

mLLLmol

00.1

100)250.0)(/1065.3( 5

mol41008.9

14

1271008.9

1155.0

gmol

mol

g

Measurement of the UV Spectrum:

Using a dilute solution: 1 mg of compound with MW (100-400 g/mol) is dissolved in 100 mL volumetric flask. Hexane is used as a solvent. Portion of the solution is transferred to a cuvette quartz of 1 cm (Styrene, MW = 104 g/mol)

max=250 nm

How does it work?When UV and VIS light is absorbed by the

sample chromophores (system containing the electrons responsible for absorption in UV-VIS) electrons held in π bonds are readily excited to higher energy.

Energy absorbed is related to the type of functionality within compound

12

How does matter absorb radiation

Three basic processes by which a molecule can absorb radiation; all involve raising the molecule to a higher internal energy level, the increase in energy being equal to the energy of the absorbed radiation, . Three type of internal energy are quantized;

Rotational transition: the molecule rotates about various axes, the energy of rotation being a definite energy levels, so the molecule may absorb radiation and be raised to a higher rotational energy level.

Vibrational transition: the atoms or groups of atoms within a molecule vibrate relative to each other, and the energy of this vibration occurs at definite quantized levels. The molecules may then absorb a discrete amount of energy and be raised to a higher vibrational energy level.

Electron transition: the electron of a molecule may be raised to higher electron energy. UV-VIS spectroscopy principal/theory

hv

Energy level diagram illustrating energy changes associated with absorption of electromagnetic radiation

A = pure rotational changer (far IR)B = rotational + vibrational changes (near IR)C = rotational + vibrational + electronic transition (Vis + UV)Eo = electronic ground stateE1 = first electronic exited state

Electronic Spectra and Molecule Structure

The electronic transition in the UV-VIS regions are due to the absorption of radiation by specific types of: Groups, bonds and functional groups within the molecule

Kinds of transitions: Electrons in a molecule can be classified into 4 different types:

i. Closed shell electron – not involved in bondingii. Covalent single bond electron, s

iii. Paired non bonding outer – shell electron (n electrons) such as those on N, O, S, and halogens. These are less tightly held then electrons and can be excited by visible or UV radiation

iv. Electron in p orbitals, in = or ≡ bonds. These are the most readily excited and are responsible for a majority of electronic spectra in the UV-VIS regions.

* p and n electrons responsible for UV-VIS spectrum

Other electron remain/occupate in orbitals

A molecule also possesses unoccupied orbitals called antibonding orbitals; these correspond to excited- state energy levels (either * s or *p ). Therefore, absorption of radiation results in an electronic transition to antibonding orbitals.

The most common transition: *

*

above 200 nmnm

*occur at very short λ nm

n

n

HOMO: Highest Occupied Molecular OrbitalLUMO: Lowest Unoccupied Molecular Orbital

Example, molecule H2

Example of transition * and n * in ketone (acetone)

* *The probability of is higher than n

*Molar absorptivities,

* (1000-100,000)

n * (less than 1000)

*

Measurement of the spectrumTaken in dilute solution

Compound diluted in solvent of choiceSolvent: transparent & less interaction with

absorbing speciesMost common 95% ethanolOthers: cyclohexane, acetonitrile

Spectrum plotted as log10 (Io/I) vs λFor publication and comparison converted to e

vs λ or log e vs λ Intensity measured as area under curveemax maximum intensity adopted in everyday use

Selection rule and intensityIrradiation does not always give rise to excitation of

electrons from any filled orbital to any unfilled orbital

Due to symmetry governing transitions which are allowed.

Intensity is a function of allowedness (electronic transition of target area to capture light)

e = 0.87 x 1020 P.aP: transition probabilitya: target area of chromophore

The longer the conjugated system the more intense the absorption

ABSORPTION BY ISOLATED CHROMOPHORES

The absorbing groups (the electrons responsible for the absorption) in a molecules are called chromophores

A molecule containing a chromophore – chromogen

The present of auxochrome enhance the absorption by a chromophore or shift the λ of absorption.

Example: hydroxyl (OH), amine (-NH2), halogens (X)

posses n electrons – interact with electrons in the chromophores (n ) conjugation.

AuxochromeFunctional group that does not itself absorb

in the UV region but has the effect of shifting chromophore peaks to longer wavelengths and increasing intensities

Ex: -OH (270nm, e = 1450), -NH2 (280nm, e = 1430) on benzene(256nm, e = 200)

Spectra changes can be classed as:

Bathochromic shift (red shift) – shifted to longer wavelength. It may caused by addition of conjugation.

Hypsochromic shift (Blue shift) - shifted to shorter wavelength. It may caused by removal of conjugation.

Eg. The conjugation of the lone pair electrons on the N atom on aniline with the bond system of the benzene ring is removed on protonationNeutral aniline absorbs at = 230 nm

= 8600But in acid solution (protonated) = 203 nm

= 7500

max

max

max

max

Solvent effectπ-π*:

transition result in an excited state more polar than ground state. Dipole interaction with solvent molecule will lower energy of excited state – longer wavelength maxima

Red shift from hexane to ethanoln-π*:

Weak transition of O lone pair in ketones (n-π* transition) shows solvent effect in opposite direction

Solvent effect is lesser extent to which solvents can hydrogen bond to carbonyl group in excited state – blue shift

24

Spectra changes can be classed as: continue….

Hyperchromic effect – increased absorption intensity ( ↑)

Hypochromic effect – decreased absorption intensity ( ↓)

Note: = the wavelength of the absorption maximum

= Absorption [ ] of a 1% solution in a cell with 1 cm pathlength- used when MW of a compound is not known

max

%11cm )(log 0

10 II

Structure Determination in Conjugated Systems: UV SpectroscopyConjugated compounds can absorb light in the

ultraviolet region of the spectrumThe electrons in the highest occupied molecular

orbital (HOMO) undergo a transition to the lowest unoccupied molecular orbital (LUMO)

The region from 2 x 10-7m to 4 x 10-7m (200 to 400 nm) is most useful in organic chemistry

A plot of absorbance (log of the ratio of the intensity of light in over light transmitted) against wavelength in this region is an ultraviolet spectrum

26

Ultraviolet Spectrum of 1,3-Butadiene Example: 1,4-butadiene has four molecular

orbitals with the lowest two occupiedElectronic transition is from HOMO to LUMO

at 217 nm (peak is broad because of combination with stretching, bending)

27

Interpreting UV Spectra: The Effect of Conjugationmax: wavelength where UV absorbance for a

compound is greatestEnergy difference between HOMO and LUMO

decreases as the extent of conjugation increasesmax increases as conjugation increases (lower

energy)1,3-butadiene: 217 nm, 1,3,5-hexatriene: 258 nm

Substituents on system increase max

28

Conjugation, Color and the Chemistry of VisionVisible region is about 400 to 800 nmExtended systems of conjugation absorb in

visible region b-Carotene, 11 double bonds in conjugation, max = 455 nm

Visual pigments are responsible for absorbing light in eye and triggering nerves to send signal to brain

29