135 

 exceeds 0.28 mM (Figure 30), On the basis of molar extinction coefficient 

 for Cu'^-bathocuprione disulfonate complex (Blair and Diehl, 1961), all the 

 Cu^"^ present in the reaction mixture was reduced to Cu* when kojic acid was 

 added at a concentration greater than 0.28 mM. 



Effect of Ko.iic Acid on Quinone Products 



Spectrophotometric scanning of the product generated from the 

 reaction of DL-DOPA and mushroom PPG revealed two distinct absorption 

 peaks at 316 and 480 nm (Figure 31a). The addition of 5.63 mM kojic acid 

 to this solution caused the color to change from red-brown to violet, and 

 the subsequent disappearance of the 480 nm peak which represents 

 dopaquinone (Figure 31b) (Fling et al., 1963). Similar phenomena occurred 

 when kojic acid (5.63 mM) was added to the reaction mixture containing 

 lobster PPG and DL-DGPA. Thus, the formation of dopaquinone from DL-DGPA 

 through the action of PPG was affected by the presence of kojic acid. 



This finding was further verified by the TLC analysis of the DL-DOPA 

 standard and the reaction mixture (dopaquinone) containing 3.5 mM of kojic 

 acid. A reddish-purple spot with a R^ value of 0.76 was detected for the 

 DL-DOPA of these two samples on TLC plate following spraying with 

 ninhydrin reagent. However, no such reddish-purple spot was detected for 

 control sample that contained only dopaquinone. 



The kojic acid effect on the quinone products formed by the action 

 of PPG on DL-DGPA is attributed to the reduction of dopaquinones back to 

 diphenols (DL-DGPA). Many reagents including cysteine, bisulfite, and 

 ascorbic acid are known to retard enzymatic browning through this 



