324 GEORGE \V. CROSBIE 



or homogenates, or a study of growth factor requirements of microbial 

 mutants. 



The work of Barnes and Schoenheimer 3 using N 15 -ammonium citrate 

 clearly showed that mammalian and avian polynucleotides could be derived 

 from small molecule precursors. N-l has been shown 4 to be derived from 

 ammonia, C-2 from C0 2 , 6 while the remainder of the pyrimidine ring (N-3, 

 C-4, C-5, and C-6) is derived from aspartic acid. 6 ' 7 



The implication of the Lactobacillus bulgaricus 09 growth factors, car- 

 bamylaspartic acid (ureidosuccinic acid), and orotic acid (uracil-4-carbox- 

 ylic acid) in pyrimidine synthesis was indicated by their incorporation into 

 the polynucleotides of that organism. 8 Confirmation of the role of orotic 

 acid as a pyrimidine precursor has been provided by the observation of 

 its incorporation into the polynucleotides of Escherichia coli B, 9 yeast, 10 

 mammalian liver slices, 11 • 12 and tumors 13 and by its incorporation into the 

 uridine-5'-phosphate of the acid-soluble fraction of rat liver. 1416 



The synthesis of orotic acid in liver slices has been studied by Rei- 

 chard and Lagerkvist. 7 In an extensive investigation they have studied the 

 incorporation of N 15 -labeled ammonium chloride, bicarbonate-C 13 , L-as- 

 partate-N 15 , L-aspartate-1 , 4-C 13 , L-aspartate-2 , 3-C 14 and L-carbamylas- 

 partate-N 15 into an added pool of orotic acid. The results obtained confirm 

 and amplify those of previous investigations and together with other iso- 

 typic and microbiological evidence clearly implicate aspartic acid, car- 

 bamylaspartic acid, and orotic acid (or close derivatives thereof) as bio- 

 synthetic precursors of uridine-5'-phosphate (Fig. 1). The implication of 

 dihydroorotic acid in this pathway is indicated by its conversion to orotic 

 acid by rat liver homogenates 17 and by its ability to support the growth 

 of L. bulgaricus 09. 18 



3 F. W. Barnes and R. Schoenheimer, J. Biol. Chem. 151, 123 (1943). 

 « U. Lagerkvist, Arkiv Kemi 5, 569 (1953). 



B M. R. Heinrich and D. W. Wilson, J. Biol. Chem. 186, 447 (1950). 

 6 U. Lagerkvist, P. Reichard, and G. Ehrensvard, Acta Chem. Scand. 5, 1212 (1951). 

 » P. Reichard and U. Lagerkvist, Acta Chem. Scand. 7, 1207 (1953). 

 s L. D. Wright, C. S. Miller, H. R. Skeggs, J. W. Huff, L. L. Weed, and D. W. Wil- 

 son, J. Am. Chem. Soc. 73, 1898 (1951). 

 » L. L. Weed and S. S. Cohen, J. Biol. Chem. 192, 693 (1951). 



10 M. Edmonds, A. M. Delluva, and D. W. Wilson, /. Biol. Chem. 197, 251 (1952). 



11 L. L. Weed and D. W. Wilson, J. Biol. Chem. 189, 435 (1951). 



12 P. Reichard and S. Bergstrom, Acta Chem. Scand. 5, 190 (1951). 



13 L. L. Weed, Cancer Research 11, 470 (1951). 



i" R. B. Hurlbert and V. R. Potter, J. Biol. Chem. 195, 257 (1952). 

 " R. B. Hurlbert, Federation Proc. 11, 234 (1952); 12, 222 (1953). 



16 E. Herbert, V. R. Potter, and L. I. Hecht, J. Biol. Chem. 225, 659 (1957). 



17 C. Cooper, R. Wu, and D. W. Wilson, J. Biol. Chem. 216, 37 (1955). 



'8 C. S. Miller, J. T. Gordon, and E. L. Engelhardt, J. Am. Chem. Soc. 75, 6086 (1953). 



