| Abstrakt: |
Nicotinate is readily converted to NAD in animal tissues. The pathway involves nicotinate mononucleotide and nicotinate adenine dinucleotide as intermediates (1, 2). Animal tissue also contains enzyme(s) having NAD glycohydrolase activity that breaks NAD down to nicotinamide and adenosine diphosphoribose (3). The administration of low levels of nicotinate to mice results in a marked increase in hepatic nicotinamide levels (4) presumably from NAD via NAD glycohydrolase. This observation that nicotinate appears to be a more efficient precursor of hepatic nicotinamide than nicotinamide itself motivated us to explore the matter further. The results presented here indicate that hyperphysiological levels of hexestrol cause a marked alteration in the means whereby nicotinate is converted to nicotinamide in mice.Materials and Methods.The C57 black male mice (25-35 g) received hexestrol or sesame oil intramuscularly as described in the tables. The 14C-nicotinate was administered by intravenous injection at the level indicated. The animals were sacrificed by cervical rupture at the time indicated, the livers immediately removed and dropped into a dry ice-acetone freezing mixture. Individual frozen livers were dropped into 3 ml of boiling 0.04 Mpotassium phosphate buffer, pH 5.4. After 1 min in a boiling water bath the samples were homogenized for 30 sec, cooled in an ice bath and centrifuged at 21,000g for 15 min. Carrier NAD, nicotinamide, and nicotinate where then added to the supernatant fluid and aliquots were applied to sheets of Whatman 3MM paper. The sheets were developed in either the isobutyrate-NH3solvent of Magasanik et al.(4), or in butanol saturated with water containing solid NH4HCO3(5). The spots corresponding to NAD, nicotinamide, and nicotinate were cut out and quantitated for radioactivity (6).Result and Discussion.The intravenous administration of tracer amount of 14C—nictinate (1.5-7.5 μg) (Table I, Fig. 1) results in the following events: (i) The intrahepatic levels of labeled nicotinate decreases rapidly during the first few minutes after intravenous injection. |
