Article

Phenol-Hypochlorite Reaction for Determination of Ammonia

Analytical Chemistry (Impact Factor: 5.64). 07/1967; 39(8). DOI: 10.1021/ac60252a045

ABSTRACT

The Berthelot color reaction has been investigated with the particular aim of presenting a simple, reliable analytical procedure. The combination of two reagents prepared readily, phenol plus nitroprusside and alkali plus hypochlorite, gave excellent reproducibility with great convenience. After the ammonium sulfate sample was mixed with the phenol reagent the hypochlorite reagent could be added up to 30 minutes later without change of absorbance, whereas if this sample were mixed with a reagent containing hypochlorite or with alkaline phenate plus nitroprusside, there was a decrease in absorbance unless the second reagent was added immediately. Satisfactory color development with minimal precautions was given by a variety of reagent concentrations and by reaction temperatures of 20°, 25°, 37°, and 75° C. Sensitivity was increased at 75° C although the time to reach maximum absorbance was longer than at 37° C. The absorbance maximum was reached quickly at 100° C but this temperature is not recommended because the absorbance decreased rapidly thereafter.

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    • "The measurement conditions were: 40 @BULLET C; a mobile phase of 0.1% phosphoric acid; and a mobile phase flow rate of 1 mL min −1 . The ammonium was measured by the phenol-hypochlorite method [21]. The -PGA concentration was measured by cetylmethylammonium bromide (CTAB) method222324. "
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    • "Extraction was carried out according to De Block et al. [11]. Ammonium nitrogen content was measured according to Weatherburn [12]. Water content was determined by drying for 24 h at 105 ∘ C. All data were subjected to analysis of variance (ANOVA). "
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    • "Urease inhibition assay. The urease inhibition activity was determined by measuring the amount of ammonia being produced using the indophenol method described somewhere (Weatherburn, 1967). The assay mixture, containing 10 μL of enzyme and 10 μL of compounds (25 μg/mL) in 40 μL buffer (100 mM urea, 0.01 M K 2 HPO 4 , 1mM EDTA, and 0.01 M LiCl 2 , pH 8.2), were incubated for 30 min at 37 °C in 96-well plates. "
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