SYMPOSIUM ON PREPARATION OF SAMPLES FOR MICROBIOLOGICAL ASSAY

Abstract

The precision and reliability of most of the In samples heated to destroy the natural encommonly used microbiological assay techniques zymes, 5 mg of chicken pancreas enzyme effected have been fairly well established. Many of the maximum release of folic acid from 1 gm of problems encountered in obtaining satisfactory spinach. Samples could be autoclaved over a pH analyses are due, not to the assay procedure per range of 5 to 13 without any loss of folic acid se, but to difficulties in preparation of the sample activity. for assay. With the growing popularity of microIn the extraction of pantothenic acid from biological methods, there has been an increase foods, a double enzyme system of alkaline intesin the complexity of the materials that are subtinal phosphatase and pigeon liver enzyme was jected to analysis, and consequent emphasis on superior to mylase-P. Maximum release was obthe need for good extraction procedures. This served when samples were incubated in the pressymposium was organized to focus attention on ence of both enzymes. A hog kidney enzyme some of the problems encountered, and to discuss preparation was found to be as effective as a few of the specific methods which have been pigeon liver enzyme in the double enzyme system. extensively studied. Care must be exercised in the preparation and Toepfer and Reynolds, whose paper was preuse of either of these enzymes to prevent loss of sented by their colleague, Dr. O'Barr, discussed conjugase activity and consequent low pantothe preparation of samples for vitamin assay, thenic acid values. Evidence of the efficiency of with particular reference to folic acid, pantothe double enzyme system in releasing pantothenic acid, vitamin B6 and vitamin B12. Working thenic acid from conjugates in several foods primarily with various foods, they found that, was provided by the results of parallel microbiowith the exception of vitamin B12, enzymatic logical and bioassays. digestion of food samples generally was necessary In assaying selected food for vitamin B6 acto assure complete liberation of these vitamins in tivity with Saccharomyces carlsbergensis, autoforms available to the assay organisms used. claving samples for 15 min with 0.055 N HCl, Of several enzyme preparations tested for the followed by overnight incubation with clarase or release of folic acid from a variety of foods, intestinal phosphatase was as effective as autochicken pancreas enzyme was found to be suclaving for 5 hr in 0.055 N HCl in releasing this perior to papain, takadiastase or hog kidney vitamin. These enzymes also were effective in enzyme. Using dried spinach as a substrate, they releasing pyridoxal from pyridoxal phosphate. found naturally occurring enzymes in unheated Current evidence indicates that enzymatic disuspensions which were about as effective as gestion generally is not required in the preparachicken pancreas enzyme in releasing folic acid. tion of samples for microbiological assay of vita-