DEVELOPMENT OF NEAR REAL-TIME MONITORING SYSTEMS FOR SOME SERINE-PROTEASE ENZYMES IN THE INDUSTRIAL ATMOSPHERE

Rapid, sensitive and reusable assay systems have been developed which can be used for near real-time monitoring of some protease enzymes in the workplace. The systems are based on fluorescence measurement with flow analysis using a mini-bioreactor containing a chemically immobili zed fluorophore-labelled protein substrate. Protease enzymes such as s ubtilisin digest the fluoro-substrate and release fluorescent fragment s which are detected downstream by a fluorimeter. The assay systems we re optimized and used for measurement of standard solutions of proteas e enzymes and enzyme-spiked non-biological detergent solutions. These exhibited satisfactory performance in terms of speed, sensitivity, rep roducibility and linearity. Detection is fast with response times of 4 -5 min. The relationship between the enzyme (subtilisin) concentration and the observed fluorescence signal is linear within the measured ra nge (0-20 ng for a flow injection analysis system and 0-20 ng ml(-1) f or a simulated air sampling open-loop system). For the latter system u sing a bioreactor with 0.9 ml immobilized matrix, the limit of detecti on at 95% confidence is 0.26 ng ml(-1). The signal is reproducible wit h s(r) of 0.93% (n=8) for continuous use over a 16 h period with repea ted exposure to 2 ng ml(-1) subtilisin episodes. Copyright (C) 1996 Br itish Occupational Hygiene Society.