Patent GB-1015556-A (GB)
Title: A process for the production of 5'-ribonucleotides
IPC: C12P19/30
Filed: 1963-11-26
Abstract: A 51-ribonucleotide, is obtained by culturing a mutant strain of a bacterium of the species Bacillus subtilis, Escherichia coli or Ptoteus retgerii, capable of producing ribonucleoside and requiring at least adenine for growth, and a strain of a bacterium of the species Pseudomonas trifolii, Pseudomonas perlurida, Pseudomonas melanogenum, Serratia marcescens, Flavobacterium harisonii, Staphylococcus aureus, Alcaligenes metacaligenes, Alcaligenes viscolactis or Achromobacter butyrii capable of producing 51-ribonucleotide from the corresponding ribonucleoside by biochemical phosphorylation, in a medium containing an assimilable carbon source, an assimilable nitrogen source, and nutrients including a source of adenine for fermentation of the said bacteria, adding to the cultured broth thereby obtained a phosphate donor, and allowing biochemical phosphorylation of the nucleoside to take place at a pH value of from 3.0 to 5.5 and a temperature of from 30 DEG  to 37 DEG  C., to accumulate 51-ribonucleotide. The culture medium is inocculated with a nucleoside producing bacterium and a phosphate group transferring bacterium, and cultured at from 30 to 37 DEG  C. for from 60 to 70 hours under aerobic conditions. Although the nucleoside producing bacterium and the phosphate group transferring bacterium may be inoculated upon the culture medium at the commencement of cultivation and cultured simultaneously, it is more practicable for the phosphate group transferring bacterium to be inoculated upon the medium in which a nucleoside producing bacterium has been previously cultured at about 30 DEG  C. for about 45 to 48 hours under aerobic conditions, and the medium thereafter cultured at about 30 DEG  C. for a further 16 to 24 hours.  In either case, the cultured broth will contain a substantial amount of nucleoside and cells of the phosphate group transferring bacterium.  A phosphate donor is then mixed with the cultured broth, the pH of the mixed solution is adjusted to from 3.0 to 5.5, and the mixture permitted to react at about 30 DEG  C. for from 24 to 48 hours, thereby to accumulate 51-ribonucleotide in the reacted solution in a high concentration.  When 10-5 to 10-1 mole of a copper salt and/or a zinc salt is added to the reaction mixture, the production of 51-ribonucleotide can be greatly increased. Specified assimilable carbon sources include glucose, starch hydrolysate and other carbohydrates. Suitable assimilable nitrogen sources include inorganic and organic ammonium salts, urea, nitrates and ammonia, these may be used alone or in admixture.  Nutrients for the fermentation of the bacteria include essential mineral salts required for growth of the bacteria, such as potassium salts, magnesium salts, manganese salts, or iron salts, and phosphates and sulphates.  In addition, certain specific growth nutrients are required, such as adenine, amino acids, and nutrients having the effect of increasing the nucleoside production or the nucleoside phosphorylating source such as corn steep liquor, yeast extract, sodium ribonucleate, casein hydrolysate, polypeptone, or meat extract.  The ribonucleoside producing strains are mutant strains which may be obtained from the normal strains by the action of ultra-violet light, X-rays or gamma radiation or contact with sodium nitrite solution.  Examples are given for the preparation of sodium 51-inosinate, 51-guanylic acid, 51-adenylic acid, 51-cytidylic acid and 51-xanthylic acid.