Allantoin and Allantoic acidIn nodulating legumes, ureides are synthesized from the products of nitrogen fixation via purine degradation. Roots are considered the main site of ureide synthesis from where they are translocated to different parts of the plant through xylem sap. Their synthesis in other plants also appears to proceed via purine degradation but is dependent on the inorganic nitrogen source fed to the plant; the greatest ureide production is associated with ammonia assimilation. Ureides serve as a source of nitrogen for amino acid and protein synthesis.Principle Allantoin in the presence of alkali forms allantoic acid. In turn allantoic acid in 'acidic medium forms glyoxylic acid and urea. Glyoxylic acid reacts with phenylhydrazine hydrochloride to form phenylhydrazone. This hydrazone on reaction with potassium ferricyanide produces a red color which is measured at 520nm. Materials» Sodium Hydroxide 0.5NDissolve 20g NaOH in one liter distilled water. » Hydrochloric Acid 0.65N Dilute 6.5mL of concentrated hydrochloric acid to 100mL with distilled water. » Phenylhydrazine Hydrochloride, 0.33% Dissolve 0.05g in 15mL of distilled water. Only use fresh solution. » Concentrated Hydrochloric Acid, 10N. » Allantoin Stock Standard Dissolve 50mg allantoin in 100mL distilled water. » Working Standard Dilute 1mL to 10mL (50mg/mL) » Phosphate Buffer, 0.05M, pH 7.5 Procedure
CalculationCalculate the amount of allantoin in the unknown samples using the standard curve.Note In this procedure allantoic acid present in the tissue is also estimated along with allantoin. Hence, the final value includes both allantoin and allantoic acid References Richard, J T and Larry, E S (1981) Phytochem 20 361. Young, E G and Conway, G F (1942) Jbiol Chem 142 839. |
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