9780521516358book.pdf

Sample derivatisation

Some functional groups, especially hydroxyl, present in a test analyte may comprom-

ise the quality of its behaviour in a chromatographic system. The technique of analyte

pre-orpost-column derivatisationmay facilitate better chromatographic separation

and detection by masking these functional groups. Common derivatisation reagents

are shown in Table 11.1.

Table 11.1Examples of derivatising agents

Analyte Reagent

Pre-column

Ultraviolet detection

Alcohols, amines, phenols 3,5-Dinitrobenzoyl chloride

Amino acids, peptides Phenylisothiocyanate, dansyl chloride

Carbohydrates Benzoyl chloride

Carboxylic acids 1-p-Nitrobenzyl-N,N^0 -diisopropylisourea

Fatty acids, phospholipids Phenacyl bromide, naphthacyl bromide

Electrochemical detection

Aldehydes, ketones 2,4-Dinitrophenylhydrazine

Amines, amino acids o-Phthalaldehyde, fluorodinitrobenzene

Carboxylic acids p-Aminophenol

Fluorescent detection

Amino acids, amines, peptides Dansyl chloride, dabsyl chloride, fluoroescamine,

o-phthalaldehyde

Carboxylic acids 4-Bromomethyl-7-methoxycoumarin

Carbonyl compounds Dansylhydrazine

Post-column

Ultraviolet detection

Amino acids Phenylisothiocyanate

Carbohydrates Orcinol and sulphuric acid

Penicillins Imidazole and mercuric chloride

Fluorescent detection

Amino acids o-Phthalaldehyde, fluorescamine, 6-aminoquinolyl-

N-hydroxysuccinimidyl carbamate

445 11.2 Chromatographic performance parameters