The effect of methylation of humic acids on their influence on proteolytic enzyme activity

Humic acids, methylated with diazomethane, no longer inhibit or stimulate proteolytic enzymes. If an exhaustively methylated humic acid sample is hydrolysed to different degrees, there is a direct relationship between the number of carboxyl groups freed and the effect of the sample on the activities of four proteolytic enzymes. When fully hydrolysed the humic acid sample has essentially the same effect as the untreated humic acid, although the phenolic groups are still methylated. These results suggest that carboxyl groups, not phenolic groups, are responsible for the influence of humic acids on enzyme activity.