Beiträge zur enzymatischen Histochemie. VIII. Eine Mikromethode zur Bestimmung der Aktivität lipolytischer Enzyme.

Abstract

Using the apparatus developed by K. Linderst0m-Lang and Heinz Holter, a micro method for the determination of lipolytic enzymes was worked out, on the principle that the acid formed during enzymatic action lowers the alkalinity of an alkaline buffer so that less mineral acid is needed to reduce the pH to some definite value. 7 cu. mm. of buffer-substrate mixture, composed of 1% methyl buty-rate in 0.4 M glycine and 0.1 M NaOH, is added to 7 cu. mm. of enzyme preparation (usually in 30% glycerine) and the whole incubated at 40[degree] for 5 hrs. The enzyme action is stopped by addition of 50 cu. mm. of 2% phenol solution, containing about 0.004% brom-thymol blue, and the solution titrated with M/20 HCl, the end-point being adjusted to pH 6.5, by comparing with a standard color tube. The titrations are reproducible to 0.14 cu. mm. The titration value obtained is subtracted from the blank value at zero time. This method is applicable to studies on microtome sections of tissue or tiny amounts of liquid.