Determination of proteolytic enzymes by flow-injection analysis

✍ Scribed by Pierre Nicolas; Annick Lamy; Sylviane Reymond

• Publisher: Elsevier Science
• Year: 1991
• Tongue: English
• Weight: 363 KB
• Volume: 192
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(91)90185-v

✦ Synopsis

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.