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应用HPLC反相色谱法对维生素B_6(VB_6)的三种不同同系物(吡哆醇、吡哆胺、吡哆醛)进行分离测定是一种新的VB_6测定法,经充分粉碎的样品,用0.1M硫酸溶液,在120℃下水解30分钟,冷却后经离心、过滤,用ODS-C_(18)反相柱、pH2.0～2.5的硫酸溶液作流动相,在1.0ml/min的流速下,能将吡哆胺、呲哆醛、吡哆醇完全分离,并在激发波长293nm,发射波长395nm时,应用外标法定量,其回收率在99～103％,变异系数在5.0％～5.3％。实验结果表明,HPLC反相色谱法测定VB_6(吡哆胺、吡哆醛、吡哆醇)是一种快速,简便、灵敏度高,重现性好的方法。
Determination of three different homologues (pyridoxine, pyridoxamine, pyridoxal) of vitamin B_6 (VB_6) by RP-HPLC was a new VB_6 assay. After thoroughly pulverized, 0.1M sulfuric acid solution, hydrolysis at 120 ℃ for 30 minutes, cooled and centrifuged, filtered, with ODS-C_ (18) reversed phase column, pH 2.0 ~ 2.5 sulfuric acid solution as the mobile phase at a flow rate of 1.0ml / min , Pyridoxamine, pyridoxal and pyridoxine could be completely separated, and the external standard method was used to quantify the fluorescence at the excitation wavelength of 293nm and the emission wavelength of 395nm. The recoveries ranged from 99% to 103% and the coefficients of variation varied from 5.0% 5.3%. The experimental results showed that HPLC reversed-phase chromatography was a rapid, simple, sensitive and reproducible method for the determination of VB_6 (pyridoxamine, pyridoxal, pyridoxine).