5- Azauracil preparation

The common method of preparation of 6-alkyl-2,4-dioxotetrahydro-triazines is the cyclization of acyl-biurets by aqueous hydroxide. Formyl biuret which should by analogy 3deld 5-azauracil had not been known until recently. Its transient formation can be expected during further synthesis of 5-azauracil. Piskala and GuP achieved... 

Other derivatives of s-triazine, in particular the 2,4-disubstituted ones, are usually prepared by total synthesis and are therefore not closely linked with the chemistry of 5-azauracil unlike the analogous derivatives of 1,2,4-triazine. 2,4-Dimethoxy-l,3,5-triazine was mentioned earlier (e.g., Section II,A,2,a), the other substances are not related to the present subject. 

The chemistry of the 6-aza analogs of pyrimidine bases which has been developed from the biochemical aspect since about 1956 was based on work reported in relatively numerous older papers. In spite of the fact that 6-azauracil was prepared only in 1947 and suitable syntheses were described only quite recently, substances of this type and methods of their preparation had been known for a long time. The chemistry of 6-aza analogs of pyrimidine bases is therefore relatively closely linked with the chemistry of the 1,2,4-triazine derivatives. 

In this connection the possibility of oxidation of these substances to the tetrahydro derivatives should be mentioned. It was made use of by Thiele and Bailey for the preparation of 6-methyl-3,5-dioxo-2,3,4,5-tetrahydro-l,2,4-triazine (6-azathymine) (46) and only recently by Grundman et al. for that of 6-azauracil (42). 

With semicarbazones of lower a-keto acids the reaction proceeds with some difficulty or not at all. Thus, the semicarbazones of pyruvic acid cannot be cyclized and that of glyoxylic acid is predominantly hydrolyzed so that the yield of the cyclization product is only 20-25%. ° This reaction was used in work with a different object, for preparing 6-azauracil, for the first time. 

For unsubstitUted or lower alkylated dioxotriazines, it is advantageous to cyclize semicarbazones by sodium ethylate in ethylene glycol as described by Chang and XJlbricht. In this reaction 6-aza-uracil is obtained in 66% yield. The procedure was used for the preparation of labeled 6-azauracil ° and later for the synthesis of a number of 6-alkyl derivatives including 6-azathymine. °... 

Using a single-step process, 6-azauracil can be prepared from chloral 3-methylisothiosemicarbazone (59), The apparent intermedi-... 

The alkylation of 6-azauracil will be treated later. The first, but not exactly identified dimethyl derivative was prepared by Grundmann." The course of alkylation was studied in greater detail by Gut et al. These authors found that in aqueous alkaline solution and on using alkyl halides or dialkyl sulfates, the main alkylation product is the 1,3-dialkyl derivative (64). Since, however, the alkylation is to some... 

The sodium or potassium salt of 6-azauracil in aqueous ethanol, anhydrous ethanol, or ethylene glycol reacted with methyl iodide practically exclusively to give the 3-methyl derivative (63). In toluene the sodium, potassium, and mercuric salts produced no methylated derivatives whereas the silver salt also yielded the 3-methyl derivative, Similarly, the 3-methyl derivative was prepared from the mercuric salt of 6-azathymine, and its structure was established by hydrolysis to pyruvic acid 4-methylthiosemicarbazone. ... 

The preparation of W-alkyl derivatives of 6-benzyl-3,5-dioxo-l,2,4-triazine by hydrolysis of the corresponding alkylmercapto derivatives was systematically studied by Cattelain. The conversion to known alkyl derivatives of dioxotriazines was used to determine the structure of alkylated methylmercapto derivatives. As will be shown later (e.g., Section H,B,4,b) this procedure has a general preparative significance for 1-alkyl derivatives of 6-azauracil. ... 

Some of them were obtained for the first time by an enzymatic procedure which, of course, can result only in the aza analogs of natural nucleosides, i.e., ribofuranosyl-6-azauracil (6-azauridine) (75) and 2 -deoxyribofuranosyl-6-azathymine (6-azathymidine). The first of these was prepared by Skoda et ai. and a modification of their procedure was used by Handschumacher, In this way it is possible to obtain the crystalline nucleoside on the large scale. 

This synthesis appears to be quite general for the preparation of 1-substituted nucleosides and was used with small modifications for the synthesis of l-ribofuranosyl-6-azathymine and 2 -deoxyribo-furanosyl-6-azauracil and -6-azathymine. In the case of 2 -deoxy-ribofuranosyl a mixture of a- and )8-anomers is produced, their ratio depending on the reaction conditions. In the preparation of 2 -deoxy-ribofuranosyl-6-azathymine only one anomer was obtained having probably the )8-configuration, ... 

Azauridine was also synthesized using the knowledge of the course of alkylation of 6-azauracil 2-methylmercapto derivatives (e.g., Section II,B,4,b). The 1-ribofuranosyl derivative obtained by reaction of the mercury salt of the 2-methylmercapto derivative with tri-O-benzoyl-jS-D-ribofuranosyl chloride on removal of the methyl-mercapto and then benzoyl groups yielded crystalline 6-azauridine, The main difference between uracil and 6-azauracil nucleosides consists in the preparation of cyclic nucleosides. It is known that uridine can be readily converted to cyclic nucleosides by the reaction of 2 (50-O-mesyl derivatives with nucleophilic agents, Analogous... 

The numerous 6-substituted 3-thioxo-5-oxo derivatives prepared by earlier authors are reviewed by Erickson et ak Subsequently a number of alkyl, and further 4-acetaminophenyl, 2-thienyl, 4-pyridyl," 4-antipyryl, " and 2-aminoethyP derivatives were prepared plus some others mentioned in the section on the synthesis of 6-azauracil. 

Thioxo-3-oxo-2,3,4,5-tetrahydro-l,2,4-triazine (4-thio-6-azauracil) (88) and 3,5-dithioxo-2,3,4,5-tetrahydro-l,2,4-triazine (2,4-dithio-6-azauracil) (89) w ere prepared by Hitchings et al. treating 6-aza-... 

In agreement with the results of Cattelain, further methylation of the 3-methylmercapto derivative (96) results practically exclusively in 2-methyl-3-methylmercapto-5-oxo-2,5-dihydro-l,2,4-triazine (97). Further methylation of 5-methylmercapto derivative (90) yields 2-methyl-5-methylmercapto-3-oxo-2,3-dihydro-l,2,4-triazine (100). Their structure was confirmed by acid hydrolysis leading to 2-methyl-3,5-dioxo derivatives (62), As was already mentioned, this reaction is a suitable general procedure for preparing the 1-alkyl derivatives of 6-azauracil. ... 

Direct bromination readily yields the 6-bromo derivative (111), just as with uracil. Analogous chlorination and iodination requires the presence of alkalies and even then proceeds in low yield. The 6-chloro derivative (113) was also obtained by partial hydrolysis of the postulated 3,5,6-trichloro-l,2,4-triazine (e.g.. Section II,B,6). The 6-bromo derivative (5-bromo-6-azauracil) served as the starting substance for several other derivatives. It was converted to the amino derivative (114) by ammonium acetate which, by means of sodium nitrite in hydrochloric acid, yielded a mixture of 6-chloro and 6-hydroxy derivatives. A modified Schiemann reaction was not suitable for preparing the 6-fluoro derivative. The 6-hydroxy derivative (115) (an isomer of cyanuric acid and the most acidic substance of this group, pKa — 2.95) was more conveniently prepared by alkaline hydrolysis of the 6-amino derivative. Further the bromo derivative was reacted with ethanolamine to prepare the 6-(2-hydroxyethyl) derivative however, this could not be converted to the corresponding 2-chloroethyl derivative. Similarly, the dimethylamino, morpholino, and hydrazino derivatives were prepared from the 6-bromo com-pound. ... 

It may be said in conclusion that the reactivity of position 5 (i.e., 6 of the triazine ring) is similar to that of uracil. The only difference seems to be in the failure to prepare 5-nitro-6-azauracil although this reaction proceeds readily with uracil. 

Trichloro-l,2,4-triazine was reacted with menthanol and subsequently crystallized from w ater to yield 5-chloro-6-azauracil. A chloro-dimethoxy derivative appears to be an intermediate product, this being further cleaved by hydrogen chloride. No 2,4-dimethoxy derivatives have been prepared so far. 

Sorm et a/. prepared azacytidine and some of its derivatives in a similar way. The 4-thio derivative was obtained from 2, 3, 5 -tri-0-acetyl- or 2, 3, 5 -tri-0-benzoyl-6-azauridine by treatment with phosphorus pentasulfide this liberated 4-thio-6-azauridine (126) which was identified with 4-thio-6-azauracil on comparing the UV spectra. Treatment with ammonia produced 6-azacytidine (127) treatment with hydrazine, hydroxylamine, and n-butylamine yielded the corresponding derivatives. 

A more convenient method for the preparation of this ring system started with thiosemicarbazone 123, whose alkaline cyclization to 2-thio-5-(o-nitrobenzyl)-6-azauracil 125 proceeds smoothly (84CCC2628). In contrast to the above method, it is not necessary to isolate the Z-form of thiosemi-... 

The deoxy analogs of 2.1 were prepared by coupling l-benzyloxy-2-chloromethoxypropane with silylated thymine, 6-azathymine, uracil, and 6-azauracil to give the anticipated nucleosides 1194 in addition to minor quantities of benzyloxymethylated produce 1195 of type 5.1 (94MI15). None of the deprotected nucleosides exhibited significant activity against HIV. 

Azauracil [1,2,4-triazine-3,5(2,4)-dione] inhibits the growth of various micro-organisms. When grown in the presence of 6-azauracil- -C, Streptococcus jaecalis accumulates radioactive metabolites in the acid-soluble fraction of the cells. A major metabolite is D-ribofuranosyl-6-aza-uracil. This material is identical with material prepared by condensing tri-O-benzoyl-D-ribofuranosyl chloride with the mercuric derivative of 6-azauracil, followed by debenzoylation. A second major metaboUte was tentatively shown to be D-ribosyl-6-azauracil 5-phosphate. Bacteria develop resistance against 6-azauracil and its D-ribosyl derivative. Resistant Streptococcus faecalis will not convert 6-azauracil to its D-ribosyl derivative or to other bound forms, and the bacterium has also lost the ability to incorporate uracil into the nucleic acids of its cells. 

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