Spectrophotometrical method for determination of nitrogen in biological preparations based on thymol-hypobromite reaction

✍ Scribed by L.I. Glebko; J.I. Ulkina; V.E. Vaskovsky

• Publisher: Elsevier Science
• Year: 1967
• Tongue: English
• Weight: 387 KB
• Volume: 20
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(67)90259-x

✦ Synopsis

The determination of nitrogen is a routinous analyses used in the biochemistry of proteins, lipids, nucleic acids, and certain carbohydrates. Calorimetric procedures involving Nessler's reagent (l-3) are most commonly applied. The major disadvantage of the method is the fact that the colored complex does not form a molecular solution; moreover, the analysis cannot be performed in the presence of cations forming precipitates with alkali. Minute amounts of heavy metals produce a considerable inhibition.

Other calorimetric techniques have been used in a number of recent studies (47). Among them, the phenol-hypochlorite procedure (8-10) seems the most advantageous; the sensitivity of this method is of the order of that involving Nessler's reagent.

In 1939, Hansen and Nielsen demonstrated that the thymol-hypobromite reaction is more suitable than the phenol-hypochlorite one, and elaborated a procedure involving distillation of ammonia (11) for the determination of nitrogen in plant materials. This method, now widely used in inorganic analysis (12-14)) has not been applied since that time to biochemical analysis.

The present paper is concerned with the study of the effect of experimental conditions on the thymol-hypobromite reaction and of its applicability to the analysis of various biological substances.

REAGENTS

All the reagents were of analytical grade.

Water was twice glass-distilled in the presence of H,SO, (1 ml/liter).

Commercial thymol was purified by vacuum distillation. An alkaline solution of thymol was prepared daily by dissolving 2.265 gm of thymol in 8 ml 2 N NaOH and adjusting with water to 190 ml.