Role of hemoglobin in the differential biotransformation of glyceryl trinitrate and isosorbide dinitrate by human erythrocytes.

Incubation of 2 X 10(-7) M glyceryl trinitrate (GTN) at 37 degrees C with human red blood cells resuspended in saline resulted in a 73.4 +/- 3.5% (S.D.) elimination of GTN after 10 min. The elimination of GTN was accompanied by the appearance of an equimolar amount of the GTN metabolites. The biotransformation of GTN and another organic nitrate, isosorbide dinitrate (ISDN), was examined in more detail using the 25,000 X g supernatant fraction of human red blood cells (RBC-SF). Incubation of 2 X 10(-7) M GTN or ISDN at 37 degrees C with RBC-SF resulted in a 46.3 +/- 7.3% (S.D.) elimination of GTN after 40 min and a 51.8 +/- 5.9% (S.D.) elimination of ISDN after 240 min. The elimination of the parent organic nitrate was accompanied by the appearance of an equimolar amount of metabolites. The biotransformation of ISDN was inhibited completely by pretreatment of the RBC-SF with trypsin, N-ethylmaleimide or heating at 65 degrees C, whereas GTN biotransformation was only inhibited partially by these treatments. Biotransformation of GTN was inhibited partially by pretreatment of the RBC-SF with CO or potassium ferricyanide; these treatments had no effect on ISDN biotransformation. Treatment of the RBC-SF with the combination of N-ethylmaleimide plus CO or trypsin plus CO resulted in complete inhibition of GTN biotransformation. We conclude that ISDN biotransformation by erythrocytes is a sulfhydryl-dependent enzymatic process, whereas the biotransformation of GTN is due to a combination of a sulfhydryl-dependent enzymatic process and an interaction with reduced hemoglobin.