-^^■ 



24 

 otherwise specified, experiments were carried out three times in 

 duplicate. For this study, the enzyme activities of potato, apple, spiny 

 lobster, Australian lobster, white shrimp, and grass prawn PPOs were 

 determined to be 10,900, 97,400, 7,000, 3,000, 5,400, and 900 units/mg 

 protein, respectively. 



pH Optima and Stability of Lobster PPQ 



The modified method of Gormori (1955) was followed for preparation 

 of various buffer solutions including 0.1 M sodium citrate-0.1 M HCl , pH 

 2.0, 3.0, and 4.0; 0.05 M sodium phosphate, pH 5.0, 6.0, 6.5, 7.0, 7.5, 

 and 8.0; 0.1 M glycine-0.1 M NaOH, pH 9.0, and 10.0; and 0.1 M sodium 

 phosphate-0.1 M NaOH, pH 11.0 and 12.0. The assay was performed at 25*'C 

 by adding 40 /xL enzyme solution to a mixture containing 280 nl of buffer 

 solution and an equal volume of 10 mM DL-DOPA in distilled water. The 

 mean velocity for dopachrome formation was determined at 475 nm using a 

 DU-7 spectrophotometer (Beckman Instruments, Inc., Irvine, CA) . 



After enzyme mixtures containing 40 juL enzyme preparation and 120 /iL 

 of each of the previously described buffer systems were incubated at 25°C 

 for 30 min, a 40 ni aliquot was removed and added to 560 /iL of 10 mM DL- ' 

 DOPA solution in distilled water. Dopachrome formation was monitored 

 spectrophotometrical ly . 



Activation Energy and Thermostability of Lobster PPQ 



Reaction mixtures containing 40 /zL enzyme extract and 560 /iL 10 mM 

 DL-DOPA solution were incubated at various temperatures ranging from 20° 

 to 60°C. Activation energy (EJ was determined according to the Arrhenius 



