FISHERY BULLETIN: VOL. 80, NO. 2 



muscle and on ground (minced) muscle. Condi- 

 tions for testing were kept close to those under 

 which we knew the parasitic enzyme functioned. 

 The pH was maintained at that of the fish (6.8), 

 the substrate was the fish muscle, and the tem- 

 perature was moderate (45°C). 



Blended Fish 



Blended fish muscle was prepared by blending 

 two parts 0.1 M NaCl with one part ground fish 

 in a Lourdes Blender 3 in a quantity large enough 

 to serve for several tests. The pH (6.8) of the 

 solutions of the various potential inhibitors was 

 maintained by the addition of dilute NaOH or 

 HC1. In a 50 ml polycarbonate tube, 2 ml of the 

 blended fish was mixed with 1 ml 0.1 M NaCl, as 

 a control, or with 1 ml of the potential inhibitor. 

 The tubes, covered with parafilm, were incu- 

 bated for 90 min at 45°C. Duplicate samples of 

 the control and test material were kept at 0°C in 

 order to know the soluble protein level before in- 

 cubation. This figure was subtracted from the 

 quantity of soluble protein that was the result of 

 increased proteolysis in the incubated sample. 

 The reaction was stopped by the addition of 3 ml 

 of 10% trichloroacetic acid. After 30 min at room 

 temperature, the tubes were centrifuged at 9,750 

 g for 10 min. Protein determinations by the 

 Lowry method (Lowry et al. 1951) were done on 1 

 ml of the supernatant. The effectiveness of the 

 inhibitor was gauged by comparison of the 

 proteolysis of the control (0.1 M NaCl) with that 

 of the potential inhibitor. Since over a period of 

 time the amount of proteolysis was bound to vary, 

 a control was run with each experiment. In order 

 to calculate the amount of inhibition, an arbi- 

 trary figure of 100% was assigned to the control 

 and the effectiveness of the inhibitor was expres- 

 sed as percent inhibition by the following formula: 



g protein/ml of test 



X 100 = % proteolysis 



g protein/ml of control 



100 — % proteolysis = % inhibition. 



Ground Fish 



Ground fish was prepared by putting partially 

 frozen fillets through a 4mm die. Ten parts of 

 ground fish were thoroughly mixed with 1 part 



'Reference to trade names does not imply endorsement by 

 the National Marine Fisheries Service, NOAA. 



of 0.1 M NaCl or the inhibitor solution. Three 

 grams of this material was incubated in a 50 ml 

 covered polycarbonate tube for 30 min at 45°C. 

 The reaction was stopped by the addition of 3 ml 

 of 10% trichloroacetic acid. The remaining 

 treatment was the same as with the blended fish. 



Preparation of Ground Fish Blocks 

 for Storage 



A quantity (about 200 g) of the ground para- 

 sitized Pacific whiting was mixed with 0.1 M 

 NaCl (approximately the ionic strength of 

 muscle) as a control or an inhibitor in the ratio of 

 10 parts fish to 1 part solution. Before the blocks 

 were placed in storage, aliquots were taken to 

 test for inhibition and inhibitor residues. The 

 blocks (3" X 1" X 8") were stored at -20°C for 1 

 mo. At the end of the month, aliquots were 

 retested for inhibition and inhibitor residues. 



Effect of Proteolytic Inhibition 

 on Texture 



The blocks of parasitized whiting made for the 

 storage study and a similar block made from 

 nonparasitized Pacific whiting were used to test 

 the effectiveness of maintaining texture by 

 inhibiting proteolysis. Duplicate portions (3" X 

 1" X Y 2 ") were cut from each block and baked in 

 a covered dish (3 1 / 2 " X 2" X l 1 //'). The baked por- 

 tions were randomly mixed before presenting 

 them to an experienced panel for texture and 

 organoleptic evaluation. In order to express the 

 results in numerical values, numbers were 

 assigned to the texture categories: firm (1); soft 

 (2); mushy (3). Aliquots were taken at the same 

 time to test for percent inhibition. 



Oxidative Effect on Amino Acids 



Amino acid analyses, using the Beckman 118 

 CL Amino Acid Analyzer (Spackman et al. 

 1958), were done on acid hydrolysates of non- 

 parasitized fish, parasitized fish with no 

 treatment, and parasitized fish which had been 

 treated with either 0.5% disodium phosphate 

 peroxide plus 0.025% potassium bromate or 0.5% 

 dipotassium phosphate peroxide plus 0.025% 

 potassium bromate. 



Enzyme Inhibitors 



All chemicals were of reagent grade. Trypsin 



282 



