GASOMETRIC METHOD OF ESTIMATING OXIDASE ACTIVITY

In the study of biological oxidations, two eriteria are used as indications of oxidase activity. One of these is the oxidation-reduction potential of the system concerned, determined as described by CLARK (3) and others. The other is the rate of oxidation measured by one of several inethods. In our particular studies on the oxidase systems of various fruits, measurement of rate is much more useful than measurement of oxidation-reduction potential. Rate measurements Four important methods of measuring the rate of enzymic oxidation have been reported in the literature. These are the (1) colorimetric, (2) titrimetric, (3) electrometric, and (4) manometric methods. If fruit oxidase is at all specific, it is very probable that the rate at which it oxidizes an added indicator is different from that at which it oxidizes the naturally occurring substrate. It is also possible that different catalysts are involved in oxidation of the natural and the added substrates. There are other objeetionable features aLso, such as interference of turbidity of the juice, and difficulty in duplicating tints in permanent color standards. Several of the more important colorimetric methods are those of DYE (6), ROHMAN and SPITZER (13). and WILLSTATTER and WEBER (17). Most colorimetric methods require the addition of H1202. This addition alters natural conditions, and CRUEss and FONG (4) have shown that its concentration greatly affects the results obtained. GUTHRIE (8) uses in a titrimnetric technique a special substrate formed by the action of NaOH on glucose solution. HAEHN and STERN (9) have reported upon a method in which unchanged added tyrosin is determined at intervals. In the electrometric method of STEARN and DAY (15), hydroquinone of the quinone-hydroquinone complex used with the quinhydrone electrode is oxidized by the oxidase, changing the ratio of quinone to hydroquinone and thus the E.M.F. of the quinhydrone electrode. The resultino drift in potential is taken as a measure of the rate of oxidation. One of the earliest applications of the manometric method was made by FoA (7). The well known BARCROFT manometric apparatus and its modified form, the WARBURG (16) apparatus, are in very general use. DIXON and ELLIOTT (5) have recently still further modified the BARCROFT apparatus. The BUNZEL (1, 2) apparatus also makes use of the mianometric principle (see also SENNHAUSER 14). Practical objections to the manometric apparatus are its high cost and 327