Keto-Acids dinitrophenylhydrazones

N6. Neish, W. J. P., On the separation of u-keto acid dinitrophenylhydrazones by ionophoresis and their polarographic estimation. Rec. trav. chim. 72, 105 (1953). 

T2. Tauber, H., Separation of a-keto acid dinitrophenylhydrazones by paper electrophoresis and their colorimetric determination. Anal, Chem. 27, 287 (1955). 

Menkes (1959) reported data collected from six more children. All showed symptoms similar to those described above, and died within 15 days to 20 months of birth. In one case, Menkes was able to obtain urine samples during the last months of the infant s life. When he treated the urine with 2,4-dinitrophenylhydrazone, which forms colored precipitates with keto compounds, he found three a-keto acids in unusually large amounts ... 

Dinitrophenylhydrazones (DNPHs) were applied to the GC analysis of keto acids. As with carbonyl compounds, they are prepared by reaction with 2,4-dinitrophenylhydrazine and are also used mainly for the preliminary isolation of keto acids. They can be isolated from a dilute aqueous sample by adsorption on activated carbon and selective desorption [178] hydrazones of aldehydes with a methyl formate-dichloromethane mixture and hydrazones of keto acids with a pyridine-water azeotropic mixture. Hydrazones of acids are released from their pyridine salts with methanol containing hydrogen chloride. After... 

Other workers separate by paper chromatography the amino acids obtained by catalytic hydrogenolysis of the 2,4-dinitrophenylhydrazones of the a-keto acids (H19, K19, T24). 

Neubauer B) and Knoop (Ba) demonstrated in the early part of this century the metabolic interconversion of a-amino and a-keto acids. In 1920, Thunberg (S) observed that glutamate was oxidized in the presence of frog muscle, and, in 1936, Weil-Malherbe (4) prepared an extract from brain tissue which catalyzed glutamate oxidation and identified the product, a-ketoglutarate, as its 2,4-dinitrophenylhydrazone. 

Apart from citrus oils, other essential oils have also been analyzed by means of LC, such as the blackcurrant bud essential oil [100]. The latter was fractionated into hydrocarbons and oxygenated compounds, and the two fractions were submitted for RP-HPLC analysis. Volatile carbonyls consist of some of the most important compounds for the blackcurrant avor and, hence, were analyzed in detail. The carbonyls were converted into 2,4-dinitrophenylhydrazones and the mixture of 2,4-dini-trophenylhydrazones was separated into derivatives of keto acids and monocarbonyl and dicarbonyl compounds. Each fraction was submitted to chromatographic investigation. 

For the determination of keto-acids, e.g. pyruvic, a-ketoglutaric and oxalacetic acids in mixtures and in biological materials, a method has been proposed employing paper electrophoretic separations of the 2,4-dinitrophenylhydrazones of these acids.< ) After elution, the polarographic wave corresponding to the reduction of the nitro-groups (in the 2,4-dinitrophenylhydrazone formed) was followed in 0-1 N HCl and the first wave at —0T4 V measured. 

Kallio, H. and Linko, R.R. (1973), Gas-liquid chromatographic analysis of 2,4-dinitrophenylhydrazones of keto acid methyl esters. J. Chromatogr., 76,229. 

Dinitrophenylhydrazones of carbonyl compounds may be formed as described for keto steroids (Section 4.1.1.4.1) by precipitation or extraction from aqueous perchlorate as described by Neuberg et al. [65], by the method of Houben-Weyl [66] or by the method of Shriner et al. [67] using sulfuric acid—water-ethanol as the reaction medium. 

The 2,4-dinitrophenylhydrazone derivatives of a-ketoisovaleric acid and a-keto-jS-methylvaleric acid are separated from 2,4-dinitrophenylhydrazine by chromatography at room temperature on a Zorbax Cjg column (4.6 mm x 250 mm). Solvent A was 25% acetonitrile in water containing 0.1% triethyla-mine (v/v) and adjusted to pH 4.5 with acetic acid. Solvent B was acetonitrile. A linear gradient from 20 to 50% B was made within 20 minutes. The effluent was monitored at 254 nm. 

It also has been used as a spray reagent and for quantitation in paper and thin layer chromatography Section 6.5). It should be noted that A - 4-3-keto steroids react much less readily with isonicotinic acid hydrazide than do A4-3-keto steroidsThe 2,4-dinitrophenylhydrazone has also been described4. 

The oxidation product has been isolated as its 2,4-dinitrophenylhydrazone and corresponds with -keto-6-aminovaleric acid. Putrescine, arginine, Manske s 1 (+) acetylornithine (118) and other related amino acids are oxidized much less readily if at all. The same authors have extracted an ornithine dehydrogenase from the young roots of Datura tatula. It requires the cooperation of a coenzyme not yet identified, and appears unable to oxidize putrescine and amino acids other than ornithine and to a lesser extent glutamic. Either of these systems, or the two linked into a H-transfer chain, would seem able to catalyze the oxidation of ornithine in the living tissues. No carbon dioxide was released from ornithine by the poly-phenolase system but on addition of an unwashed belladonna tissue-suspension carbon dioxide was liberated, presumably by decarboxylation of the a-keto-5-aminovaleric acid formed by the oxidation. 

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