Semi-automated system for analysis of vitamin B6 complex by ion-exchange column chromatography

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  • 安本 教傳
    Department of Food Science & Technology, Faculty of Agriculture, Kyoto University
  • 田寺 謙次郎
    Department of Food Science & Technology, Faculty of Agriculture, Kyoto University Department of Agricultural Chemistry, Faculty of Agriculture, Kagoishima Univeristy
  • 辻 英明
    Department of Food Science & Technology, Faculty of Agriculture, Kyoto University
  • 満田 久輝
    Department of Food Science & Technology, Faculty of Agriculture, Kyoto University

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  • Semi-automated system for analysis of vitamin B6 complex by ion-exchange columin chromatography.
  • Semi automated system for analysis of v

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A simple system for semi-automatic analysis of pyridoxal, pyridoxine and pyridoxamine has been developed using diazide of 5-chloroaniline 2, 4-disulfonyl chloride as a color-producing reagent in combination with desoxypyridoxine as an internal standard. It employs Aminex A-5 column, 10×0.9∅cm, as an adsorbent. Adsorbates were eluted successively with 3 discrete phosphate buffers (0.4N Nat). The effluent is mixed continuously with capillary streams of the diazide reagent and of sodium acetate. The mixture, maintained at 65°C in a heating bath, then passes through a spiral of Teflon tubing with a residence time of 2min. Characteristic orange colored products formed by a diazo coupling reaction are continuously monitored at 440nm in a flow photometer. The individual peaks on the recorded chromatogram are manually integrated by a conventional HW method. The elution position and recovery of desoxypyridoxine permits correction for sensitivity changes or mechanical losses which might occur during a series of analyses. The analyzer system described allow quantitation from 2 to 25μg of pyridoxal, pyridoxine and pyridoxamine in a single sample within 2 hr and with a precision of 100±4%. It is also found suitable as a routine procedure for the analysis of varied biological samples.

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