Purine Bases

PURINE BASES (alloxuric, xanthine), derivatives of purine-C₅H₄N₄... The structural formula of purine, established by E. Fischer, represents a combination of a pyrimidine ring and an imidazole ring: When hydrogen at the carbon atom in the purine ring is replaced by an NH₂ group, aminopurine substances of basic character are obtained, hence the name "purine bases". Hydroxyl derivatives of purine have acidic properties and apparently exist in tautomeric forms, the oxo-form and the oxo-form (see Uric Acid). Purin itself is not found in the body, but its derivatives are widely distributed in the animal and plant world. N=ONH₂

NH-CO

NH-CO II II >n N-C-N⁺ Adenine NH-CO CO C-NH I

II Vh Xanthine CH C-NH II II ^CN N-----C-N⁺ Hypoxanthine N(CH₃)-CO CO NH- C-NH !| \, -C-N⁺ H₂N-C C-NH II II >n N-----C-N⁺ Guanine NH-CO I I CO C-N.CH₃ \ NH-C-N⁺ Heteroxanthin J-Methylxanthine

P. o. give with mineral acids well-crystallizing salts, which with the exception of adenine salts are decomposed by water. In alkalis P. o. are easily soluble; in ammonia with the exception of guanine and epiguanine are easily soluble. From acid solutions they precipitate upon addition of phosphotungstic acid. They can also be obtained in precipitate by the action of an ammonia solution of silver (the resulting precipitate is soluble in boiling HNO₃ sp. gr. 1.1) upon addition of Fehling's solution in the presence of reducing substances, e.g., hydroxylamine, upon boiling with CuSO₄ and NaHSO₃. Close chemical and physiological relationship to P. o. have pyrimidine bases, derivatives of pyrimidine-C₄H₄N₂. The structural formula of pyrimidine, proposed by E. Fischer, represents a ring: (1) N=CH (6) (2) CH CH (5) II II (3) N-CH (4) Pyrimidine bases can be isolated by precipitation with silver nitrate and barium. The following pyrimidine bases are known: cytosine-C₄H₆N₃O (6-amino-2-oxypyrimidine), uracil - C₄H₄N₂O₂ (2,6-dioxypyrimidine) and thymine-C₅H₆N₂O₂ (5-methyluracil). Cytosine and uracil are part of plant nucleosides: cytidine and uridine, and uridine apparently exists in plant cells in a preformed state.

In the body purine and pyrimidine bases are found partly in free form in negligible amounts (mainly xanthine and hypoxanthine) and partly in bound form, being part of nucleic acids (see Nucleoproteins and Nucleic acids), in larger quantities they are found in organs rich in cell nuclei, e.g., glands; in blood normally traces; in urine they are found in small amounts; their content in blood and urine increases with food rich in cell nuclei (meat) and especially in leukemia. Purine and pyrimidine bases in the body are products of the breakdown of nucleoproteins; they are cleaved off from nucleic acids and undergo further changes in cells. From various tissues and organs enzymes have been isolated that deaminate (adenase, guanase) and oxidize (xanthine oxidase) free purines and purines that are part of nucleic acids (nucleoside-deamidases and nucleoside-oxidases). In this process adenine first passes into hypoxanthine, then into xanthine; guanine directly into xanthine, xanthine under the action of xanthine oxidase into uric acid. The final product of P. o. metabolism in humans and anthropoid apes is uric acid, in other mammals allantoin. Plant purines introduced with food-theobromine, theophylline and caffeine undergo demethylation in the body and are excreted with urine in the form of heteroxanthine C₇H₆N₄O₂ (7-methyl-2,6-dioxypurine), paraxanthine C₇H₈N₄O₂ (1,7-dimethyl-2,6-dioxypurine) and 1-methyl-xanthine C₇H₆N₄O₂ (1-methyl-2,6-dioxypurine). In urine epiguanine C₆H₇N₅O (7-methyl-guanine) and episarkin (C₄H₆N₃O) have also been found, the amount of which increases in leukemia.

Some amount of P. o. apparently undergoes complete oxidation with ring cleavage in the body. When introduced per os, P. o. are partly broken down in the intestine with the release of ammonia. It is very difficult to determine the purine balance, since part of P. o. undergoes complete breakdown, part is not resorbed. After administration of nucleic acid to humans-the larger part of purine N is excreted in urine in the urea fraction, the smaller-in the uric acid fraction and only a very small part-in the form of P. o. The body has the ability to synthesize P. o.; histidine and arginine may serve as material. Purine metabolism has a close relationship to creatine metabolism- part of P. o. passes into creatine. Very little is known about pyrimidine metabolism. The pyrimidine ring in the body is apparently broken down; there are data indicating that upon oxidation of uracil and thymine urea is formed; it is established that cytosine passes into uracil in the body. Quantitative determination of free and bound (nucleotides) purine bodies in blood serum by the method of Thannhauser and Czoniczer. The method is based on the fact that when precipitating serum proteins with uranyl acetate, nucleotides are also precipitated; free purine bases and uric acid remaining in the filtrate are isolated as copper compounds and the N in them is determined by Kjeldahl. When precipitating proteins with sulfosalicylic acid by boiling in the filtrate remain both free and bound purine bases, the total amount of which is determined as in the first case. The amount of bound purine bases is determined by the difference.

1. Determination of the amount of free P. o. The serum diluted with an equal volume of water (40 cm³) is precipitated with 40 cm³ of a 1.55% solution of uranyl acetate. The liquid is diluted with water and filtered from the precipitate. 60 cm³ of the clear, non-biuret reaction filtrate are taken with a pipette and concentrated on a boiling water bath to a volume of 15 cm³, 0.5 g of chemically pure sodium acetate and 1 cm³ of a 40% solution of NaHSO₃ are added, heated to strong boiling and 1 cm³ of a 10% solution of CuSO₄ is added, after which strong boiling is continued for another 3-4 minutes. When the liquid has completely cooled, the precipitate is centrifuged, washed 3-4 times with water on the centrifuge and N in it is determined by micro-Kjeldahl (see Kjeldahl method).-2. Determination of the amount of bound P. o. 30 cm³ of serum are diluted with 55 cm³ of water; to the mixture heated to boiling 5 cm^{3 of a 20% solution of sulfosalicylic acid are added. The liquid with the precipitate is cooled by placing it in a vessel with ice and then filtered. 50 cm3 of the filtrate (3 cm3 of which = 1 cm3 of serum) are concentrated on a water bath to a volume of 15 cm3. The slightly turbid liquid is precipitated with CuSO₄ and bisulfite as described above and nitrogen in the precipitate is determined by micro-Kjeldahl.}