COLORIMETRIC DETERMINATION OF URINARY GLYCINE CONJUGATES

Abstract

A quantitative procedure is described for determining urinary glycine conjugates. The color development was recognized after addition of pyridine and benzenesul-fonyl chloride to glycine conjugates. The dried residue of ethyl acetate extract from urine was dissolved in pyridine and benzenesulfonyl chloride was added. After addition of chloroform to the mixture, the absorbance was determined at 420 nm (extracting method). In the improved procedure of direct colorimetric method, pyridine was added to urine and mixed; then benzenesulfonyl chloride was diluted with mixture of dimethylsulfoxide and ethanol, and absorbance was deter-mined at 430 nm (improved direct method).
To separate each glycine conjugate, paper chromatographic procedure was car-ried out. Urine specimen each was directly developed on filter paper with butanol- acetic acid-water or isopropanol-ammonia-water. On the other hand, ethyl acetate extract from urine was developed with toluene-acetic acid-water. Comparing the spots with authentic sample spots, each location was determined by illuminating with ultraviolet light. Then, each spot was extracted with pyridine. Benzen-sulfonyl chloride was added to the extract; the absorbance was determined at 420 nm. Complete separation of urinary glycine conjugates could be done by two-way chromatography.
The normal values of urinary glycine conjugates were also compared by ex-tracting method and improved direct method.