Fluorescence labelling of alcohols

1 7-Methoxycoumarin-3-carbonyl azide (3-MCCA) and 7-methoxycoumarin-4-carbonyl azide (4-MCCA) 

3-MCCA and 4-MCCA were developed for the labelling of hydroxyl functions [498]. They react with primary and secondary alcohols in dichloromethane to form car-bamic esters, which are sufficiently stable to be separated by reversed phase column chromatography. For cholesterol, a detection limit of 50 fg per 100 y sample volume was reported. 

7-(Chlorocarbonylmethoxy)-4-melhylcoumarin is a reagent related to the above mentioned carbonyl azides [499]. It is prepared from 7-hydroxy-4-methylcoumarin and bromoacelic acid. The resulting carboxylic acid is transformed into the carbonyl chloride by refluxing with thionyl chloride. Derivatives (of prostaglandins and steroids) are formed in carefully dried methylene chloride containing 4-dimethylaminopyridine as catalyst. The esters formed are sufficiently stable for chromatographic separation. 

4-Dimethylamino-l-naphthoylnitrile [500], 1- and 9-anthroylnitrile [501] and 2-methyl-l,l -binaphthalene-2 -carbonylnitrile [502] have been suggested as fluorescent labels for primary and secondary alcohols. 2-Methyl-l,l-binaphthalene-2 -carbonylnitrile is a chiral reagents, and is therefore suited for the separation of enantiomers. Quantitative reaction is achieved by heating in the presence of triethylamine, whereby the diastereo-meric esters are formed. 

Separations of the diastereomers were performed using a silica gel column (excitation at 342 nm, emission at 420nnri) [502]. 

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