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Abstract
A simple and rapid high pressure liquid chromatographic method using an ultraviolet detector for simultaneous analysis of histidine, tyrosin and tryptophan, is presented.
Chromatographic separation is achieved on Spherisorb-5 RP-18 5μm reversed phase column and the mobile phase is the isocratic mixture of aceto-nitrile, methanol and water (5:30:65). the eluted amino acids are detected at 220 nm. the retention time is 1.55 min for histidine, 2.21 min for tyrosin and 2.80 min for tryptophan. the correlation of the integrated peak areas with the concentration of amino acids showed a linear relationship between 0.40 to 9.43 ppm for histidine, 0.24 to 22.6 ppm for tyrosin, and 0.20 to 12.8 ppm for tryptophan per 10μl injection.
Simultaneous analysis of histidine, tyrosin and tryptophan gave reproducible results with a mean coefficient of variation 1.93 pc for tryptophan, 2.29 pc for tyrosin and 3.51 pc for histidine and r2 = 0.999. the proposed technique was applied to the analysis of these amino acids in urine samples.