Determination of proteolytic enzymes by flow-injection analysis.
Quantitation of proteolytic enzymes using N-succinyl-L-Ala-L-Ala-L-Pro-L-Phe-p-nitroanilide has been adapted to flow-injection analysis. This procedure has been developed using two different proteases: subtilisin and chymotrypsin. For both enzymes the influence of substrate concentration on spectrophotometric response has been studied. The assay is based on the merging zones technique combined with a washing step. Results are obtained in less than 15 s and samples may be run at a rate of 90/h with good reproducibility. A linear relation between peak heights and enzyme concentrations was observed for 0-0.15 Anson unit/liter of subtilisin and for 0-30 mg/liter of a commercial preparation of chymotrypsin. The method requires only small sample volumes, and the consumption of the chromogenic substrate is reduced to a minimum by using intermittent pumping.
Nicholas
P
P
Nestlé Research Centre, Nestec Ltd, Lausanne, Switzerland.
Lamy
A
A
Reymond
S
S
D000595:Amino Acid Sequence; D002138:Calibration; D002918:Chymotrypsin / Q000378:metabolism; D010450:Endopeptidases / Q000032:analysis*; D008969:Molecular Sequence Data; D015203:Reproducibility of Results; D012697:Serine Endopeptidases / Q000378:metabolism; D013379:Substrate Specificity; D013381:Subtilisins / Q000378:metabolism
D010450:Endopeptidases; D012697:Serine Endopeptidases; D013381:Subtilisins; D002918:Chymotrypsin