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Nicotinic acid, at a concentration of 2.5 µm, was a more effective precursor of NAD than was nicotinamide in perfused rat kidney. NAD synthesis from nicotinic acid occurs via nicotinic acid ribonucleotide and desamido-NAD. Nicotinamide deamidase was not active but the nicotinamide, released by NAD glycohydrolase action or added to the perfusate, was incorporated into NAD via NMN. Mono- and dinucleotides as well as quinolinate entered the kidney cells slowly. The latter was converted to nicotinic acid ribonucleotide and thence to NAD. Nicotinic acid ribonucleoside served as a precursor of NAD. Urinary products from nicotinic acid included nicotinic acid, nicotinuric acid, nicotinamide, and a trace of 6-hydroxynicotinic acid. From nicotinamide no products other than the unchanged amide were detected. Nicotinamide was the preferred precursor of NAD in the testis as shown by intratesticular injection in the whole animal or by homogenate experiments. Quinolinic acid did not serve as a source of the pyridine moiety of NAD in the testis.

In contrast to human erythrocytes, rat erythrocytes synthesize NAD directly from nicotinamide via NMN. 14C from nicotinamide appeared in NMN and NAD and to lesser extent in nicotinamide ribonucleoside and NADP. Mono- and dinucleotides formed were present solely within the cells, whereas nucleosides and the unchanged substrates readily passed through the cell membrane. NMN pyrophosphorylase, the first enzyme of this pathway, has been purified about 400-fold from rat erythrocytes and was shown to require ATP and Mg++, to have a pH optimum of 8.5 to 9.0 and to have Km values of 0.067 and 3.8 µm for nicotinamide and PP-ribose-P, respectively. The enzyme activity was increased 50% in the presence of 1 mm EDTA. Although NMN pyrophosphorylase cannot be measured in a buffer system of pH 7.4, due to the hydrolysis of the product by NAD glycohydrolase in the hemolysate, it can be readily measured at pH 8.8 where NAD glycohydrolase is not active. From radioactive nicotinic acid the major labeled products were nicotinic acid ribonucleotide, nicotinic acid ribonucleoside, and NAD with lesser amounts of label in nicotinic acid adenine dinucleotide and nicotinamide. The results suggest that nicotinic acid is converted to NAD by the Preiss-Handler pathway while nicotinamide is metabolized via the parallel pathway through NMN.