Rapid Method for the Quantitative Estimation of Microbial Lipases

Abstract

THE investigation of microbial lipases has been considerably impeded by a lack of knowledge about their natural substrates and their physiological role in the metabolism of the organisms. Any substrate chosen to detect microbial lipases must therefore be arbitrary and it is difficult to evaluate data of different workers because of the wide choice of substrates, assay conditions and methods employed. The underlying principle in the assay of microbial lipases has usually been the estimation of free fatty acids liberated from triglycerides after a suitable incubation time. We have found that direct titration methods are not suitable, because most cultures contain alkaline or acid degradation products. This can be overcome by extracting the fatty acids from the medium after incubation with the substrate¹ but the procedure is laborious and difficult to carry out quantitatively. A long incubation time is also open to the criticisms that the products of lipolysis may inhibit lipase activity^2–4 or be used in the reverse, synthetic reaction⁵. Short duration assays have been developed such as the indirect manometric method employed by Forster and co-workers⁶ or the direct method of continuous automatic titration of the acid liberated from an appropriate substrate in a pH. stat⁷, but the lipase activities of the micro-organisms that we have tested have been too low to be assayed by such techniques.