A simple method for differentiation of carbonic anhydrase (CA) I, from CA II, is described based on using the nicotinates as specific CA I inhibitors. Nicotinates inhibit the activity of CA I activity; at a concentration of 5x10−1M 100% inhibition is complete. But no effect on CA II activity is observed.In vivoadministration of xanthinol nicotinate in doses of 20 mg/kg b.w. completely inhibited erythrocyte CA I activity. Association inex vivoof methyl-nicotinate at a concentration of 5x10−1M did not further modify erythocyte CA activity that had already been reduced by i.v. administration of xanthinol nicotinate. The nicotinate class could be used as a test for an accurate differentiation of CA I from CA II activityin vitro, in vivoand inex vivo; by subtraction of I from total CA activity, one could also find the erythrocyte CA II activity. Theex vivoassays, using the test with nicotinates we call Nicosilvanil, would easily allow monitoring of CA I and II activity changes under physiological, pathological and experimental conditions, in response to various endogenous or therapeutical stimuli with either activating or inhibitory effects. Considering convenience, simplicity, and cost effectiveness of the reagents for theNicosilvanil Test, this assay has the potential of being useful in routine analysis and differentiation of CA I from CA II activity in erythrocyte samples, as well as in other tissues and organs.