THE RAPID POTENTIOMETRIC DETECTION OF CATALASE POSITIVE MICROORGANISMS

The rate of fluoride ion release from the enzymatic cleavage of fluori de ion from 4-fluorophenol by horseradish peroxidase, in the presence of hydrogen peroxide, was measured using a fluoride ion selective elec trode. Monitoring the utilisation of hydrogen peroxide by catalase (in tracellularly present in almost all aerobic microorganisms) in the pre sence of 4-fluorophenol demonstrated the inhibition of the enzyme. Hor seradish peroxidase appeared to impart a partial protective mechanism of this inhibition. The development of a sequential assay demonstrated the applicability of the proposed method in the assessment of aerobic microorganism numbers. The judicious variation of three parameters, t he length of incubation, the concentration of the primary substrate (h ydrogen peroxide) and the indicator enzyme activity (horseradish perox idase), affected both the detection limit and the sensitivity of the a ssay. Typically with a 15 minute incubation, a detection limit for cat alase activity of 1.5 x 10(-6) Uml-1 was obtained together with a sens itivity of 2.42 mumol l-1 s-1 per decade change in activity. Applicati on of the developed catalase assay to the detection of Escherichia col i achieved a detection limit of 1 x 10(2) colony forming units (cfu) m l-1 with a sensitivity of 3.26 mumol l-1 s-1 per decade change in inta ct microorganisms. By lysis of the microorganisms the detection limit was further reduced to less than 10 cfu ml-1, indicating the future po ssibilities of the assay.