High-performance liquid chromatographic determination of delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine in complex media by precolumn derivatisation with dansylaziridine.
A novel method is described for the trace level quantitation of the tripeptide delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine (ACV) in complex fermentation media, using a high-performance liquid chromatographic, pre-column derivatisation technique. The procedure is based upon the reaction of the ACV monomer with 5-dimethylaminonaphthalene-1-sulphonylaziridine (dansylaziridine) and produces a highly fluorescent product. Reaction conditions between the reagent and tripeptide were investigated and optimal conditions established. Linear calibration graphs were obtained over the ranges 227-0.56 micrograms/ml and 227-5.6 ng/ml. The extracellular ACV levels produced in fermentation broths of several different fungal strains and species were determined using this technique. The method was compared using ACV standards in buffer solutions for ease of use, sensitivity and selectivity with two other pre-column derivatisation procedures, using dithionitrobenzoic acid and monobromobimane, which also exploit the reaction with the sulphydryl group of the ACV monomer.
Orford
C D
CD
School of Biotechnology, Polytechnic of Central London, U.K.
Perry
D
D
Adlard
M W
MW
D000164:Acremonium / Q000378:metabolism; D001233:Aspergillus nidulans / Q000378:metabolism; D001388:Aziridines; D002851:Chromatography, High Pressure Liquid; D003470:Culture Media; D003619:Dansyl Compounds; D005285:Fermentation; D005456:Fluorescent Dyes; D006863:Hydrogen-Ion Concentration; D009842:Oligopeptides / Q000032:analysis*; D010408:Penicillium chrysogenum / Q000378:metabolism; D013056:Spectrophotometry, Ultraviolet
D001388:Aziridines; D003470:Culture Media; D003619:Dansyl Compounds; D005456:Fluorescent Dyes; D009842:Oligopeptides; C011638:5-(2-aminoadipyl)cysteinylvaline; C017173:dansylaziridine