Rapid assay for tryptophanase using reversed-phase high-performance liquid chromatography.
A rapid and sensitive high-performance liquid-chromatographic assay for tryptophanase based upon the fluorometric measurement of the enzymatically liberated indole was developed. The total incubation time is 20 min, and the reversed-phase separation is fast (elution time of indole in 8 min) and reproducible. The sensitivity of the method is in the nanomole range. This method was tested in the assay of tryptophanase activity in E. coli, giving an average activity of 6589.6 U/g of cells. Because of its speed, high sensitivity and minimal sample preparation, this method circumvents several problems commonly encountered in standard spectrophotometric methods of analysis.
D002851:Chromatography, High Pressure Liquid; D004926:Escherichia coli / Q000201:enzymology; D007211:Indoles / Q000032:analysis; D008190:Lyases / Q000302:isolation & purification*; D013050:Spectrometry, Fluorescence; D014368:Tryptophanase / Q000302:isolation & purification*
D007211:Indoles; D008190:Lyases; D014368:Tryptophanase