derivatisation techniques ppt
TRANSCRIPT
DERIVATIZATION IN GAS CHROMATOGRAPHY
Presented by
Sowmya k balan
TOPICS TO BE DISCUSSED
• DEFINITION • CRITERIA FOR THE COMPOUNDS • REASONS FOR DERIVATIZATION• TYPES OF DERIVATIZATION• DERIVATIZATION REACTIONS
DEFINITION
• Derivatization is the process of chemically
modifying a compound to produce a new
compound which has properties that are
suitable for analysis using gas
chromatography.
CRITERIA / PROPERTIES OF SAMPLE
• VOLATILITY
• POLARITY
• THERMOSTABILITY
The ideal derivatization procedure will:
Accomplish the desired modification.
Proceed quantitatively, or at least reproducibly.
Produce products that are readily distinguishable and separable from the starting materials.
Proceed rapidly with simple and straightforward laboratory techniques, and will be both selective and applicable to a number of similar compounds.
Involve reagents and reactions that present no unusual hazards.
Compounds containing functional groups with active hydrogen (-
COOH, -OH, -NH and -SH) are usually derivatized for analysis by
gas chromatography.
REASONS FOR DERIVATIZATION
• Improve peak shape• Suitable volatility • Better sensitivity and detectability • Increased thermostability • Relocalization of interfering peaks
Types of derivatization
• Pre column derivatization ; For improve
separation by column
• Post column derivatization ; For improve
response by detector.
Precolumn derivatisation:
• Here the compound is converted into more volatile and thermostable derivatives.
• Improve the separation and less tailing will be seen.
In the following condition, pre derivatization is done:
• The compound is less volatile• The compounds are thermolabile
• To reduce tailing• To improve separation factor
Post column derivatization:
• Improve the response shown by detector
• The compound may be converted in such a way that their ionisation or affinity towards electrons is increased.
Derivatization reactions:
• Esterification• Acylation• Perfluoroacylation• Alkylation • Silylation• Condensation• Chiral derivatization
ESTERIFICATION:
• Use to prepare derivatives of carboxylic acids.
• Eg . Analgesics , Prostaglandins , amiono acids etc. • Derivatization – Fischer Esterification Method under
acidic conditions
R-COOH + R’- OH R-COOR’
Methyl ester frequently synthesizedReagents ; diazomethane ,diazoethane
ACYLATION:Acylation Reactions
Popular reaction for the production of volatile derivatives of highly polar and involatile organic materials.
Advantages.:
• Reduces the polarity of the substance
• Improve the peak shape and reduce peak tailing.
• Improves the stability of those compounds that are thermally labile.
• Disadvantage• Acylation derivatives difficult to prepare• Reagents are moisture sensitive and hazardous• Acid byproducts need to be removed before analysis
Applications
• Render extremely polar materials such as sugars amenable to separation by GC
• Used to derivatize amines, amides, alcohols, thiols, phenols, enols, and glycols etc…….
ACYLATING REAGENTS
Fluorinated anhydrides fluoro acyl imidazolesPentafluoro propanolol
PERFLUOROACYLATION:
• Increases the molecular weight of the sample
relative to the analogous hydrocarbon.
• Increases the retention time.
eg; Direct acylation of the trifluoroacetic anhydride in
trifluoroacetic acid followed by methylation with
diazomethane in methanol.
ALKYLATION
• Used to derivatize functional groups such as alcohols,
phenols, primary and secondary amines, imides etc in which
there is labile hydrogen
• Reduces molecular polarity by replacing active hydrogen with
alkyl group
• Principal reaction ; nucleophillic displacement
.eg. Williamson ether synthesis….. Alcohol/phenol +
alkyl/benzyl halide in presence of base.
• Alkylating reagents
• Dialkyl acetals ( DMF)
• Tetrabutyl ammonium hydroxide(TBH)
• Pentafluoro benzyl bromide
• Advantage
• Wide range of reagents available
• Reactions done in aqueous solutions
• Derivatives formed are stable
• Disadvantage• Limited to amines and acidic hydroxyls• Reaction conditions are severe• Reagents ate toxic
Silylation • Most prevalent method• Produces silyl derivatives which are more volatile
and thermally stable• Occurs through nucleophillic attack• Used for the analysis of the non volatile polar
compounds.• Order of functional group by silyl acceptor ability is
alcohols > Phenols > carboxylic acids > amines > amides
• Reagents• Hexamethyl disilane (HMDS)• Tri methyl clorosilane(TMCS• Advantage• Easily prepared• Large number of reagents available• Disadvantage• Reagents are moisture sensitive• Must use aprotic organic solvents( pyridine)
• Sample or solution of sample is heated with slight
excess of silylating agent. Reaction must be carried
out in the sealed vial with inert closure. Solvent used
must be free from active hydrogen. Popular solvent
like hexane, ether, benzene will dissolve the reagent
but are not conductive to rapid reactions
Condensation
• Usually done for aldehydes or ketones present in the
sample.
• Commonly used reagent is methoxylamine and is
used to protect keto groups in steroids by forming
methoximes .
GC CHIRAL DERIVATIZATION
• They target one specific functional group and produce individual diasteromers
• Enantiopure derivatization reagents• menthyl chloroformate(MCF)
• N trifluro acetyl L propyl chloride (TPC)
REFERENCES
Practical pharmaceutical chPractical pharmaceutical chPrinciples of instrumental analysis,5th edition,Skoog,Holler.
Vogel’s textbook of chemical analysis
Practical pharmaceutical chemistry,A.H.Beckett,J.B.Stenlake