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The major difficulty encountered in the purification of phenolases is the relatively small quantities of active material present in comparison to totai inactive components. The best source of phenolases appears to be mushroom (Psalliotes canzpestris) where only 5 to 10 mg of enzyme per lb of wet material has been found (II), and enzyme from this source has considerably greater activity than shrimp enzyme(s) .In an effort to locate the area in shrimp and the specie of shrimp containing the highest concentration of phenolase, a study was made of enzyme activities of various anatomical areas of different species of shrimp including antennae, abdominal shell, press juice from various areas of the cephalothorax and blood. Thorough studies were made of phenolase activities in blood samples from Pertaeus aztecus, Penaeus setiferus, and the blue crab (Callinectes sapidus). Experiments were also carried out to determine natural substrates involved in shrimp melanosis and effects of certain oxidative inhibitors on the catecholase activity of shrimp protein. MATERIALS AND METHODSA. Shrimp samples analyzed. The P. setiferus exoskeletons, taken from freshly caught shrimp, were frozen until analyzed for phenolase activity. The Penaezcs duorarum exoskeletons were taken from shrimp that had been refrigerated in ice approximately one week prior to analysis. The exoskeletons were taken from the abdomen of shrimp and included the thin pellicle which separates the epidermis from the remainder of the shell.Juice was pressed from fresh P. aztecw and P. setiferzls heads, centrifuged (4,500 x g, 10 min) and frozen until analyzed. Blood samples were obtained by aspirating blood from the pericardial sinuses of live shrimp or immediately following their death. If freshly caught shrimp are chilled to approximately' 1.5" C immediately after death, their blood remains uncoagulated for 1 hr or more. Blood samples were collected from shrimp of different catches by inserting a No 20 hypodermic needle through the thin membrane which separates the carapace from the abdomen, into the pericardial area and aspirating the blood into small polyethylene bottles.Some of these blood samples were treated with various antioxidants and anticoagulants and centrifuged to separate the corpuscles from the plasma. Other samples were centrifuged after the blood had coagulated. The various samples were then fractionated as described in the results and analyzed for phenolase activity (3). B. Preparation of crab blood samples. Large crabs (c. sapidkks) were kept at 4" C for 1 hr and blood removed by severing one or two crawling legs between the first and second joints. The colorless blood was collected in a cold polyethylene beaker and stirred to remove fibrinogen.This blood turned light blue upon exposure to air as a result of oxidation of the hemocyanin.It was then centrifuged (27,000x g, 20 min) to remove residual clot and cells.A 50-ml aliquot of the serum was used to further purify hemocyanin. It was chilled to 0" C and cold 5% acetic slowly added until a slight...

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Objective quality tests are urgently needed by the shrimp industry to permit distribution and processing of individual catches of shrimp according to their expected storage life.Heretofore, organoleptic measurements of quality have been used almost exclusively for purchasing and grading shrimp products at various stages during their storage life. These tests are very useful for rapid examination of such products. Unfortunately, they are based on personal judgment and consequently result in numerous subjective differences which lead to disagreement in qualification of individual samples. The subjective differences render it desirable to supplement organoleptic testing with a more exact analysis based upon changes of certain constituents in shrimp during storage.Thorough organoleptic studies on shrimp quality (12) showed that definite taste changes occurred during ice storage, indicating that palatability and flavor change only slightly during the first few days of ice storage. The flavor is described as characteristic of "fresh" shrimp and is slightly sweet. This phase is followed by a period during which the shrimp no longer have this characteristic sweet flavor but instead are "flat" tasting. During this time, no off flavors associated with spoilage are noted. This second period is followed by another sharp change in flavor which denotes onset of spoilage. Repeated organoleptic studies with ice-stored shrimp by experienced taste panel personnel substantiated these three general flavor changes.Since these results show definite taste phases during ice storage of shrimp, useful objective tests must discern the three major taste changes.An ideal objective test would be one which would accurately and reproducibly give a numerical value of quality. The shrimp industry desires a test which can be correlated directly with the number of days shrimp have been in ice storage. It is rather doubtful that such a test can be perfected because physical and chemical changes are dependent not only on elapsed time of storage, but also upon treatment prior to storage and the environment from which the shrimp were obtained initially.Any measure of definite chemical changes occurring in shrimp during storage under various conditions can be used to determine relative quality provided appropriate comparisons are made. Except for several tests described below it is unlikely that specific chemical concentrations can be assigned as a means of qualifying a particular sample of shrimp since the initial content will differ depending on such facts as age, size, specie o r living environment. If analysis shows that a constituent such as ammonia nitrogen increases sharply after a specific storage time, and that slightly 611

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