Reactions with Phenylhydrazine Osazone Formation

Phenylhydrazones are best prepared in dilute acetic acid solution use of more concentrated acetic acid can lead to formation of 1-acetyl-1-phenyl-hydrazine, m.p. 130-131° (corr.) if there is a neighboring hydroxyl group, which can lead to osazone formation, and if this is to be avoided, then any acidity is unfavorable and the reaction should be carried out as near the neutral point as possible. Condensation usually occurs in the cold, and almost always on short warming on the water-bath. A solvent is often unnecessary. <%,/ -Unsaturated aldehydes and ketones may cyclize to pyrazole derivatives on reaction with phenylhydrazine. 

Derivatives of the Carbonyl Group. Not many reactions of the aldehyde or keto group of sugars are observed, because the carbonyl forms of sugars comprise only a fraction of one percent in the equilibrium mixture. They can be trapped, however, in a few reactions, e.g. by oximfe formation. Important for the identification of sugars is the reaction with phenylhydrazine or with substituted phenylhydrazines (E. Fischer), which involves two carbon atoms and results in osazones ... 

Another reaction that is characteristic of a-hydroxy aldehydes or ketones, which has been found of value for the characterisation of sugars, is the formation of osazones with phenylhydrazine. This reagent reacts with either an aldose... 

Historically, techniques such as the formation of osazones and the demonstration of fermentation have contributed significantly to the separation and identification of carbohydrates. Observation of the characteristic crystalline structure and melting point of the osazone derivative, prepared by reaction of the monosaccharide with phenylhydrazine, was used in identification. This method is not completely specific, however, because the reaction involves both carbon atoms 1 and 2 with the result that the three hexoses, glucose, fructose and mannose (Figure 9.19), will yield identical osazones owing to their common enediol form. 

Since the discovery of the reaction of phenylhydrazine with sugars to form osazones by Fischer,35 many investigations have utilized this reaction in the identification and separation of sugars, but the precise structures of osazones and their mode of formation are still in doubt.34 There is no doubt, however, that the initial reaction involves the formation of a hydrazone, which then reacts further to yield eventually a f)u-l,2-hydrazone or osazone. 

Several mechanisms34 have been proposed for this reaction. That of Weygand,36 in which an Amadori rearrangement is proposed, has considerable merit.37 Illustrations of unusual osazone formation are described by Bonner and Drisko.38 When phenyl /S-D-xylopyranosyl sulfone (XXII) or /J-D-glucopyranosyl sulfone (XXIV) is oxidized by periodic acid, a dialdehyde oxidation product (XXIII or XXV), which is susceptible toward further oxidation, is obtained. The reaction of XXIII or XXV with phenylhydrazine yields glyoxal phenylosazone and benzenesulfinic acid. Surprisingly, both XXII and XXIII react with phenylhydrazine to form D-xylosazone and D-glucosazone, respectively. 

Fischer phenylhydrazone and osazone reaction. Formation of phenylhydrazones and osazones by heating sugars with phenylhydrazine in dilute acid. 

R. D. Guthrie, Arylazo-steroids Part IV. The reaction of phenylhydrazine and 1-methyl-l-phenylhydrazine with bromo- and dibromo-5-a-cholestanones. Osazone formation from a-halogcno-ketones, Chem. Commun., (1966) 781-782 J. Chem. Soc., (C), (1968) 3079-3084. 

A study of the scope of this reaction has been performed which shows that excess phenylhydrazine will convert the disubstituted quinoxalines 19 into the pyrazolo[3,4-h]quinoxaline 20. Analogously hydroxylamine gave the isoxazolo compound 21 from these starting materials. The compounds 19 (R = Me or Ph) did not undergo cyclization with phenylhydrazine. A reaction mechanism has been suggested (see Scheme 1) which is analogous to osazone formation in sugar chemistry. 

Due to the presence of carbonyl, aldehyde or ketone radicals, sugars are capable of addition reactions with nucleophilic reagents such as phenylhydrazine (C6H5-NH-NH2). The addition of three phenylhydrazine molecules to an aldose (Figure 3.12) leads to the formation of osazone, a crystallized product with specific physicochemical characteristics, especially its melting point. This makes it possible to identify the corresponding sugar. 

Sugar Osazones, By treatment of sugars with an excess of phenylhydrazine at 100°C., two phenylhydrazine residues are introduced into the molecule, and sugar osazones, difficultly soluble in water, are formed 216), Optimal conditions for the preparation of glucosazone have been determined 217), The reaction proceeds most rapidly in the presence of acetate buffers at a pH of about 4 to 6 in more acid solution (particularly in the absence of air), and with the free base, only the hydrazone is formed 208, 218), The presence of sodium bisulfite in the reaction mixture inhibits the formation of colored by-products 219),... 

The reaction of glucose with an excess of phenylhydrazine (phenyl-diazane) is particularly noteworthy because two phenylhydrazine molecules are incorporated into one of glucose. Subsequent to the expected phenyl-hydrazone formation, and in a manner that is not entirely clear, the —CHOH— group adjacent to the original aldehyde function is oxidized to a carbonyl group, which then consumes more phenylhydrazine to form a crystalline derivative called an osazone, or specifically glucose phenylosazone ... 

In contrast to the formation of an osazone from phenylhydrazine and the 2-bromo-2-deoxy-hexoses derived from the adduct of tri-O-acetylglucal with bromine, two isomeric products [jwhich were assigned the D-threo-3,4-diacetoxytetrahydro-2-furaldehyde (p-nitrophenyl)hydrazone structures (21)] were isolated from the reaction between 3,4-di-0-acetyl-2-bromo-2-deoxy-D-tAreo-pentose (20) and (p-nitrophenyl)hydrazine. The... 

Aldehydes and ketones react with one equivalent of phenylhydrazine, forming phenylhydrazones (Section 18.6). Aldoses and ketoses, in contrast, react with three equivalents of phenylhydrazine, forming osazones. One equivalent functions as an oxidizing agent and is reduced to aniline and ammonia. Two equivalents form imines with carbonyl groups. The reaction stops at this point, regardless of how much phenylhydrazine is present. (Recall that the pH at which imine formation is carried out must be carefully controlled Section 18.6.)... 

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