A method for the quantitation of tryptophan in Escherichia coli fermentation broth by isocratic high-performance liquid chromatography with ultraviolet detection.

A rapid, isocratic method for the determination of tryptophan in Escherichia coli fermentation broths by reversed-phase HPLC is described. Tryptophan can be measured in fermentations containing either chemically defined media or media with hydrolyzed protein supplements. The procedure was rugged and rereproducible (RSD = 1.7%). The sample response was found to be linear up to 10 mcg of tryptophan/ml. Two different columns--Vydac C18 30 mm and "deactivated" SupelcoSil LC-18-DB--were compared and evaluated for use in the analysis. The deactivated columns had the residual silanols on the silica gel chemically inactivated to reduce the interaction with basic groups or analytes. The deactivated column was found to provide better peak shape (peak assymetry factor less than 1.1) and superior efficiency (plate count greater than 40000/m) and durability (greater than 3000 injections per column) than the non-deactivated column. The procedure described was found to be more selective than a fluorometric procedure.