MARINE ORGANIC CHEMISTRY

SL20103

PRACTICAL TITLE : HIGH-PERFORMANCE

LIQUID CHROMATOGRAPHY AND THIN-

LAYER CHROMATOGRAPHY

NAME MATRIX NO.

ENGRID PACHECO DANING TUZAN BS14161084

SHARMINA BINTI JEBIN BS14110781

SITI SYAHIRAH BINTI MUZLI BS14110802

MULLAISUDHARR A/L MUNIANDY BS14110554

SHARIFAH AZIAH JASMINE BINTI SHARIF JAKARIA

BS14110110

1.0 INTRODUCTION

Chromatography is a pretty accurate description of what happens to in on wet paper

because it literally means “colour writing”. Chromatography is actually a way of separating

out a mixture of chemicals which are in gas or liquid form by letting them to creep slowly

past another substance which is typically a liquid or solid. The essential thing about

chromatography is that we have some mixture in one state of matter moving over the

surface of something else in another state of matter (liquid or solid) that stays where it is.

The moving substance is called the mobile phase and the substance that stays put is the

stationary phase. As the mobile phase moves, it separates out into components on the

stationary phase.

Gas chromatography is a term used to describe the group of analytical separation

techniques used to analyse volatile substances in the gas phase. In gas chromatography, the

components of a sample are dissolved in a solvent and vaporized in order to separate the

analytes by distributing the sample between two phases which are stationary phase abd a

mobile phase. The mobile phase is a chemically inert gas that serves to carry the molecules

of the analyte through the heated column. Gas chromatography is one of the sole forms of

chromatography that does not utilize the mobile phase for interacting with the analyte. The

stationary phase is either a solid adsorbant termed gas- solid chromatography (GSC) or a

liquid on an inert support termed gas- liquid chromatography (GLC).

Liquid chromatography is a technique used to separate a sample into its individual

parts. This separation occurs based on the interactions of the sample with the mobile and

stationary phases. There are many stationary and mobile phase combinations that can be

employed when separating a mixture, there are several different types of chromatography

hat are classified based on the physical states of those phases. Liquid- solid column

chromatography is the most popular chromatography technique and the one discussed here

features a liquid mobile phase which slowly filters down through the solid stationary phase

bringing the separated components with it.

OBJECTIVE

1. To acknowledge the analytical separation techniques use to separate marine organic

compounds.

2. To study the uses and functions of each chromatography machines.

MATERIAL

1) High Performance Liquid Chromatography machine

2) Mass spectrometry machine

3) Gas chromatography machine

PROCEDURE

1. All of us are instructed to assemble at Biotechnology Research Institute (BRI),

University Malaysia Sabah (UMS) at 8:00am.

2. Dr. Shafiqquzzaman Sidiqquee then gave us a short briefing about the practical and

he led us to the BRI building into the lab.

3. Proper attire should be worn in the laboratory and a notepad with pen should be

brought.

4. A PhD student named Yong then gave us a briefing on how to use the Liquid

Chromatography machine and precautions that should be taken when handling the

machine.

5. After the briefing on the LCG, we are then brought to another lab where the Gas

Chromatography is kept and a further briefing on the machine is done by Young.

6. Pictures and notes are taken during the practical as evidence and further reference.

RESULT

Figure 1 : HPLC System

Capillary pump

Degasser 2

Solvent Cabinet 1

Auto Sampler

Photodiode Detector

!

Figure 2 Mass Spectrometer

Figure 3: Gas Chromatography Machine

Glass Capillary

Double Electron System

Flight Tube

Figure 4: Computer is used as an online system to monetize the sample

Figure 5: Computer is used as an online system to monetize the sample

Figure 6: The auto sampler and the injector.

Figure 7: Mass Spectrometry

DISCUSSION

Liquid Chromatography Machine

An LC-MS is an High Performance Liquid Chromatography (HPLC) system with a mass

spec detector.The HPLC separates chemicals by conventional chromatography on a column.

Components of an HPLC system are solvent cabinet, degasser, capillary pump, injector,

column, detector, fraction collector and an integrator. Usually the method will be reverse

phase chromatography, where the metabolite binds to the column by hydrophobic

interactions in the presence of a hydrophilic solvent (for instance water) and is eluted off by

a more hydrophobic solvent (methanol or acetonitrile). As the metabolites appear from the

end of the column they enter the mass detector, where the solvent is removed and the

metabolites are ionised. The metabolites must be ionised because the detector can only

work with ions, not neutral molecules. And ions only fly through a very good vacuum, so

removal of the solvent is a vital first step. The mass detector then scans the molecules it

sees by mass and produces a full high-resolution spectrum, separating all ions that have

different masses . 1

LC system is responsible for the separation. The solvent cabinet is use for filtered the

sample. HPLC uses a liquid to push the sample. This liquid is called the “Mobile Phase” or

“solvent’. Before use the solvent may be filtered through micron pore size filters or the

container can have a frit filter. Based, degasser is a component of an HLPC system. The

function of degasser is degassed a solvents to eliminate formation of bubbles. Pump is to

deliver the mobile phase through the system before dilute into the auto sampler. After that,

the sample will go to the auto sampler after its break out. In the auto sampler, there is a

sample collector chamber. Some sample such as fat or organic compound was quite

impossible to ionize. Its can be ionized when add some amphoteric acid. The sample will not

recognized if the amphoteric acid is not added.

MS system is to ionize the sample again. There is double electron spray system

where there have two needles. One of the needles is for the reference of the sample mass

and the other is for the sample. The needles is a injector which is to put the sample in the

mobile phase.

Computer is used as an online system to monetize the sample. It uses an online

system to store data, temperature and to observe the ongoing process that occurs on the

sample. A chromatogram will display on the screen of the computer and it will show the

http://www.chemir.com/liquid-chromatography-mass-spectrometry.html1

profile of the sample during the process. The chromatogram shows the good and poor

separation of the sample. Each peak shown on the chromatogram represents only one

profile of a compound. Profile of the sample depends on the volatility.

Gas Chromatography Machine

The gas chromatography machine starts with injecting the liquid sample into a small

sample bottle. It is then placed into the auto sampler. The samples are then pushed into the

injector. The injector have a temperature of about to 200°C and the sample is injected into

the gas chromatography. Inside, the samples will evaporate the liquid. The separation

process is based on the temperature which usually starts from 15°C and can be up to

300°C. The liquid in the sample should be removed before it enters the column.

Next the sample will enter the column. The column is about 30m long with a

diameter of about 0.25µm and the column has a temperature of up to 355°C. It is where

the coating will be done upon the sample. The temperature has to be adjusted according to

the boiling point of sample . If the temperature exceeds the sample then the result will be 2

loose.

Similar to the Liquid Chromatography Machine, it uses an online system to store

data, temperature and to observe the ongoing process that occurs on the sample. During

the process, a chromatogram will display on the screen of the computer and it will show the

profile of the sample. The chromatogram shows the good and poor separation of the

sample. Each peak shown on the chromatogram represents only one profile of a compound.

Profile of the sample depends on the volatility.

www.teaching.shu.ac.uk/hwb/chemistry/tutorials/chrom/gaschrm.htm2

CONCLUSION

The analytical separation techniques used to separate marine organic compounds are

acknowledged and the functions of each chromatography machines are studied.

REFERENCES

1.Chromatography. (no date) . What is Chromatography? in http://

www.explainthatstuff.com/chromatography.html

Retrieved on 3rd October 2015.

2.UCDAVIS CHEMWiki. (no date). Liquid Chromatography in http://chemwiki.ucdavis.edu/

Analytical_Chemistry/Instrumental_Analysis/Chromatography/Liquid_Chromatography

Retrieved on 3rd October 2015.

3. Dr. Shafiquzzaman Siddiquee. (2015). Lecture 2 – Separations of Marine Products.

Powerpoint slide. Malaysia, Sabah. Universiti Malaysia Sabah.