Date of Award
Doctor of Philosophy (PhD)
Arthur G. Rand, Jr.
The glucose oxidase and catalase (GOX/CAT) enzyme system was studied as an inhibitor of polyphenol oxidase (PPO) in extracts of whole pink shrimp (Pandalus borealis).
PPO was isolated from shrimp heads and purified by acetone extraction, ultrafiltration, and gel filtration chromatography. A purification of 7.74-fold was obtained with a recovery of 54.3% activity. The shrimp PPO activity was compared to that of mushroom PPO activity. The shrimp PPO was found to have an optimum pH of 6.0-6.5 and to be stable at 25-35 °C. Below freezing, the shrimp PPO was damaged completely and no activity was obtained.
Several phenolic compounds were tested as the substrate of shrimp PPO. L-DOPA and DL-DOPA yielded the highest activity. The Km of shrimp and mushroom PPO were 4.54 and 0.5 mM, respectively. Varying the concentration of D-glucose, a substrate of GOX/CAT, over the range of 0.5 to 4% had no significant effect on the activity of both mushroom and shrimp PPO. GOX/CAT at a concentration of 2 units/ml inhibited 95.2% and 97% of shrimp and mushroom PPO, respectively, with 0.5% D-glucose. The PPO was completely inhibited by 6 units/ml of GOX/CAT.
The substrates and end products of GOX/CAT were studied for their potential to inhibit mushroom and shrimp PPO. Mushroom and shrimp PPO activity were inhibited completely by the GOX/CAT end products hydrogen peroxide and gluconic acid at concentrations of 2.0 mg/ml, while glucono-lactone showed a weak inhibition of less than 7.6% and 9.0%, respectively.
Oxygen consumption by mushroom and shrimp PPO was measured during the 2 min duration of the experiment. Oxygen removal reached 2.05 ppm with mushroom PPO and 1.0 ppm with shrimp PPO. However, when GOX/CAT was added, oxygen removal increased rapidly to 7.5 and 7.48 ppm, with 96.2% and 95.9% of the oxygen consumed during the 2 min period for mushroom and shrimp PPO, respectively. D-glucose showed no inhibitory effect on the activity of mushroom and shrimp PPO even at a concentration of 5.0 mg/ml.
The potential of the glucose oxidase/catalase (GOX/CAT) enzyme system for delaying melanosis in iced shrimp was studied. Fresh, pink shrimp (Pandalus borealis) with heads on were treated with various dip solutions. The dip solutions were: firstly, GOX/CAT at concentrations of 2, 4, 6, and 10 units/ml, each with 0.5% D-glucose as a substrate; secondly, sodium tripolyphosphate (STPP) at a concentration of 9% alone and at concentrations of 3%, 6%, and 9% in combination with GOX/CAT at a concentration of 6 units/ml. Finally, sodium metabisulfite at a concentration of 1.25% alone and at concentrations of 0.32%, 0.625%, and 1.25% in combination of GOX/CAT at a concentration of 6 units/ml.
In general, all treated shrimp showed a delay in melanosis development compared to untreated shrimp. Based on the melanosis scores, GOX/CAT at concentrations of 2 and 4 units/ml delayed the melanosis formation by 2 days compared to controls, and by 4 days with GOX/CAT at concentration of 6 and 10 units/ml. The dip solution of 9% STPP delayed melanosis by 2 days compared to the controls, and by 4 days with 3% and 6% STPP, each with GOX/CAT, and by 6 days with the combination of 9% STPP and GOX/CAT. Shrimp treated with sodium metabisulfite, at all concentrations, exhibited no melanosis for the duration of the study compared to the control shrimp. GOX/CAT in combination with sodium metabisulfite did not increase the melanosis inhibition compared to sulfite alone. This was due to the sulfite inhibition effect on GOX/CAT activity.
Finally, the metabisulfite dip treatment was most effective in preventing melanosis development. The combination dip of 9% STPP and GOX/CAT at concentrations of 6 units/ml appeared to be a promising alternative as a second choice. The GOX/CAT concentration of 6 units/ml and 0.5% 0-glucose as a dip had a slight effect on preventing black spot in iced shrimp.
Al-Jassir, Mohammed H. S., "Polyphenol Oxidase Inhibition by Glucose Oxidase in Pink Shrimp (Pandalus borialis)" (1987). Open Access Dissertations. Paper 554.