Miriam Rahat

Oxidation of methyl derivatives of pteridin-4-one, lumazine and related pteridines by bovine milk xanthine oxidase

1. Pteridin-4-ones, methylated at nitrogen or carbon, N-methylated lumazines and related oxopteridines were studied as substrates of a highly purified bovine milk xanthine oxidase (xanthine : oxygen oxidoreductase, EC 1.2.3.2). 2. The enzyme can oxidise at high rates both uncharged and anionic substrates. Variation of enzymic activity with pH is mainly due to pH-dependent changes in the active enzymic center. 3. Milk xanthine oxidases at different stages of purification convert pteridin-4-one into the 4,7-dione (compound 13 in this article). 4. Methylation at C-6 in the pyrazine moiety enhances enzymic attack at C-2 in the pyrimidine ring. N-Methylation may increase or reduce rates of oxidation. 5. For oxidation at C-2, the most favorable form of the substrate bears a double bond at C(2) = N(3). Attack at C-7 is enhanced strongly in structures bearing a double bond at C(6) = C(7). 6. In general, pteridines react with xanthine oxidase as non-hydrated molecules. However, oxidation of 8-methyllumazine at C-7 may take place by dehydrogenation of the 7-CHOH group of the covalently hydrated molecule.

Reactivity of 6-methylthiopurin-8-ones. Properties of 6-methylsulphonylpurin-8-ones

All 6-methylthiopurin-8-ones (except those bearing a 1-methyl substituent, which decompose) are converted by chlorine in methanol at 0° into the corresponding sulphones. In a series of N-methyl 6-methylthiopurin-8-ones, only the 1-methyl, the 1,9- and 3,9-dimethyl, and the 3,7,9-trimethyl derivatives undergo thiohydrolysis. In contrast, all the corresponding sulphones are attacked by hydrogen sulphide anion. A 6-methylsulphonyl substituent weakens basicity and increases the acid strength of purines. In 6-methylsulphonylpurin-8-one, anion formation follows the order N-9 → N-7. A 3-methyl substituent, by virtue of reduction of the ring current in the pyrimidine system, causes a diamagnetic shift of the 2-H signal.

Torsional barriers in 6-amino-5-formamidopyrimidin-4(3H)-ones

The chemical shifts of 6-amino-5-formamidopyrimidin-4(3H)-ones show that two rotamers are present in solution, their formyl signals being of unequal intensity. The two forms are stable both as neutral molecules and as cations, in contrast to the behaviour of N-alkylformamides. Exchange rates and barriers to rotation have been calculated. The reasons for the unequal distribution of the rotamers are discussed.

Thermal decomposition of quaternary hypoxanthinium salts and related purines

Thermal degradation of quaternary hypoxanthinium salts was achieved by heating their solutions in dimethylformamide. 1,3-Dialkylhypoxanthinium bromides or iodides lose the 3-substituent as alkyl halide. The latter then attacks the imidazole ring at N-7 or N-9. Thermolysis of 1,3-dimethyl-6,8-dioxotetrahydropurinium iodide proceeds along either of two pathways: (i) loss of the 3-methyl group as methyl iodide, leading to 7,9-dihydro-1-methylpurine-6,8-(1H)-dione; or removal of hydrogen iodide to give the corresponding betaine, which is methylated at N-9 to give the 1,3,9-trimethyl-6,8-dioxotetrahydropurinium salt. The latter in turn decomposes to 7,9-dihydro-1,9-dimethylpurine-6,8(1H)-dione. Similarly 7,9-dimethylhypoxanthinium iodide (16) is degraded mainly, by loss of methyl iodide, to 9-methylhypoxanthine, accompanied by a small amount of the 7-methyl isomer. Compound (16) can also lose hydrogen iodide to give the corresponding betaine, which suffers methylation at N-1 to give the 1,7,9-trimethylhypoxanthinium salt. The latter again undergoes thermolysis to give a mixture of 1,7- and 1,9-dimethylhypoxanthine.

Physical properties and chemical reactivity of 8-(methylthio)hypoxanthines

N-Methyl derivatives of 8-(methylthio)hypoxanthine(1) were synthesised by N-methylation of various 8-(methylthio)-compounds or by S-methylation of appropriate 8-mercaptopurin-6-ones. In 8-(methylthio)hypoxanthines, protonation takes place at the same positions as in the corresponding hypoxanthines. However, the neutral forms of compound (1), its 1- and 7-methyl derivatives and the 1,7-dimethyl homologue are methylated preferentially at N-3, in contrast with the alkylation of the corresponding hypoxanthines which takes place in the imidazole ring. This difference is ascribed to the steric effect of the 8-SMe substituent.The chemical shift of the S-methyl group is influenced little by protonation of the molecule, unless mesomerism can involve the thioether group. Participation of sulphonium-like resonance forms is, however, strongly hindered in the 7,9-dimethyl derivatives of (1). The latter compounds also undergo ready demethylation, hydrolysis, and thiolysis of the 8-methylthio-group.

Purine-6,8-diones: a study of their ionisation and their methylation reactions

Purine-6,8-diones are divided by their physical properties into three classes: those of class (a) carry a hydrogen atom or a methyl group at position 1, and those of class (b) bear a 3-methyl substituent. The members of class (c), bearing both 1- and 3-methyl groups, exist either as betaine or, after protonation, as the conjugate acids, with the positive charge confined to the pyrimidine ring. Anions are formed predominantly by proton loss from position 9. Consistent with this, methylation of monoanions proceeds at position 9, except in the case of members of class (b), where alkylation at N-9 is sterically hindered by the 3-methyl substituent. Methylation of dianions follows in general the sequence of proton attachment, unless steric factors prevent this.Protonation and alkylation of neutral molecules proceeds predominantly in the pyrimidine ring. However in the case of 9-methyl derivatives, methylation at N-3 is sterically hindered.