PRESENTED BY

RAVAL BHAVIN
DEPARTMENT OF PHARMACEUTICS
K.L.E. COLLEGE OF PHARMACY
BELGAUM
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Gas

CONTENTS
Chromatography

Type Of Derivatization
Method Of Derivatization
References
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GAS CHROMATOGRAPHY
Gas chromatography is an analytical technique
used for separation of thermally stable and volatile
substances.

In gas chromatography the mobile phase is gas

and the stationary phase used may be solid or
liquid.
Mobile phase used in gas chromatography are
Hydrogen, Helium, Nitrogen, Oxygen and Air.
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PRINCIPLE: Principle involved in gas chromatography is

adsorption or partition which is depend upon
the stationary phase used.
If stationary phase is liquid so it is called
Gas liquid chromatography and here
separation done by partition.
If stationary phase is solid than it is called
Gas solid chromatography and here
separation done by adsorption.

What is Derivatization?
Derivatization is the process of chemically modifying
a compound to produce a new compound which has
properties that are suitable for analysis using a GC.

Derivatization techniques are frequently employed

in chemical analysis of mixtures and in surface
analysis. e.g. in x-ray photoelectron spectroscopy.

GC samples are usually derivatized to render

highly polar materials sufficiently volatile so that
they can be eluted at reasonable temperatures
without thermal decomposition or molecular
rearrangement.

The objective of Derivatization is to increase the

volatility of the analyte.

 Improve chromatographic
detectability.

behavior

or

Derivatization is a useful tool allowing the use

of GC to be done on samples that would
otherwise not be possible in various areas of
chemistry such as medical, forensic, and
environmental.

To permit analysis of compounds not directly

able to analysis due to, for example, insufficient
volatility or stability.
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Examples of such materials that need to be

derivatized are the organic acids, amides, poly
hydroxy compounds, amino acids etc.

In order to render such materials more volatile,

they are either esterified, silanated or acetylated
using one of a number of different methods of
Derivatization .

There are two types of Derivatization-

1.
2.

Pre-column Derivatization.
Post-column Derivatization.

Pre-column Derivatization-

By this technique the components are converted

to more volatile and thermostable derivatives.
Improved separation and less tailing will be
seen after such treatment.

In the following conditions this technique is

done-

The component is less volatile.

The compounds are thermolabile.

To reduce tailing.

To improve separation factor.

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Post-column Derivatization-

It is done to improve the response shown by

detector.

The components may be converted in such way

that their ionization or affinity towards
electrons is increased.

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 Esterification :It is used to prepare the

group like prostaglandins,
has carboxyl group, which
that increases the volatility
bonding.

derivatives of carboxyl
amino acids, analgesics
is converted to ESTER
by decreasing hydrogen

For e.g. Fischer Esterification process.

R-COOH + R-OH

H+
BF3

R-COOR
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 Acylation and perfluro Acylation

When the compounds contain an amino, hydroxyl

or thio group Derivatization by acylation or
perfluro acylation is used.
The polarity will be reduced so compounds will
have greater volatility and stability than the
original compound.
This is performed by using acetic anhydride in
acylation & trifluroacetate, pentafluropropionate,
in case of perfluro acylation. It increases the
sensitivity of the detector.
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 Acylating Reagents
Pentafluoropropionic acid
Pentafluoropropionic anhydride
Pivalic anhydride

Butylboronic acid
Trifluoroacetic anhydride
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 Advantages Addition of halogenated carbons

detectability by ECD.
Derivatives are hydrolytically stable.
Increased the sensitivity.

increased

Disadvantages Acylation derivatives can be difficult to prepare.

Acylation reagents are moisture sensitive.
Reagents are hazardous and odorous.
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Alkylation

This method is used to derivatize the compounds

containing primary or secondary phenols, alcohols
or amines in which there is a liable H2.

Ether derivative can be made by Williamson

Ether synthesis in which alcohol or phenol is
treated with alkyl or benzyl halide in the presence
of base.

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Example of alkylation

R -C

DMSO

R -C

NH

N-CH3

R -C

R -C

imide

Methyl imide

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 Alkylating Reagent
PFBBr(Pentafluorobenzyl bromide)

TBH(Tetrabutylammonium hydroxide)
BF3 in methanol or butanol

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Advantages
Wide range of alkylation reagents available.

Some reactions can be done in aqueous solutions.

Alkylation derivatives are generally stable.

Disadvantages
Limited to amines and acidic hydroxyls.

Reaction conditions are frequently severe.

Reagents are often toxic.
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Silylation
Silylation produces silyl derivatives which are
more volatile, less polar, and more thermally
stable.
Replaces active hydrogens
(trimethylsilyl group).

with

TMS

Silylation occurs through nucleophilic attack

(SN2). The better the leaving group, the better the
Silylation.
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 Silylation reagents will react with water and

alcohols first. Care must be taken to ensure that
both sample and solvents are dry.

Solvents should be as pure as possible. This will

eliminate excessive peaks.
Pyridine is the most commonly used solvent.

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 In many cases, the need for a solvent

eliminated with silylating reagents. (If
sample readily dissolves in the reagent,
usually is a sign that the derivatization
complete).

is
a
it
is

Ease of reactivity of functional groups

towards Silylation follows the order:

Alcohol > Phenol > Carboxyl > Amine >

Amide hydroxyl.
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 The order of alcohols being:-

Primary > Secondary > Tertiary

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 Silylating Reagents
HMDS(Hexamethyldisilzane)
TMCS(Trimethylchlorosilane)
TMSI(Trimethylsilylimidazole)
BSA(Bistrimethylsilylacetamide)
BSTFA(Bistrimethylsilyltrifluoroacetamide)
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 Advantages and Disadvantages

Advantages
Ability to silylate a wide variety of compounds
Large number of silylating reagents available
Easily prepared
Disadvantages
Silylation Reagents are moisture sensitive
Must use aprotic (no protons available) organic
solvents
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 References
Principles of Instrumental Analysis by Skoog.D.A;
Holler.F.J.
Quantitative Analysis of drug-III edition by
P.D.Sethi.
Pharmaceutical analysis- vol II by Dr. A.V.kasture
www.google.com
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