Observations on the mechanism of the tyrosine-tyrosinase reaction.

Raper and Wormall 1 in a recent paper conclude as follows:

“Tyrosinase first produces from tyrosine a red substance. This process requires the presence of the enzyme and only takes place in the presence of oxygen. This red substance becomes colorless spontaneously and is then oxidized to form melanin. These last two processes take place in the absence of tyrosinase, but may be accelerated by it or by other oxidases found in potato juice.”

While this statement may be correct in so far as potato tyrosinase is concerned, it is apparently incorrect as a generalization for all tyrosinase preparations. I have shown 2 that the meal worm, Tenebrio molitor, contains a very active tyrosinase which can be prepared in an insoluble form. When this insoluble tyrosinase is used, it is possible to remove the enzyme completely from a system by the simple process of filtration. Consequently it is possible to initiate a reaction, carry it to a definite stage, and then stop the action of the enzyme by filtering the reaction mixture.

Tyrosine and the insoluble tyrosinase were allowed to react until a deep rose color had developed. One-half of the mixture was now removed and filtered twice through “barium” filter papers to remove the enzyme. The other half of the mixture served as a check. Both tubes were set aside in the dark, but open to the oxygen of the air. In a few hours, the contents of the tube containing the unsoluble tyrosinase and tyrosine solution became changed, first becoming violet color and later depositing the black pigment in the usual manner. The filtered portion, on the other hand, remained an unchanged rose color for eighteen days and was then discarded.