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目的建立柱前衍生反相高效液相色谱法(reversed-phase high-performance liquid chromatography,RP-HPLC)测定狭叶红景天中游离及水解氨基酸种类及含量。方法以异硫氰酸苯酯为柱前衍生化试剂;色谱柱采用Ultimate Amino Acid氨基酸柱(4.6 mm×250 mm,5μm);流动相A为80%(V:V)的乙腈-水溶液,流动相B为7%(V:V)的乙腈-CH_3COONa溶液(pH=6.5);检测波长为254 nm。结果 21种氨基酸在0.0019~2.700mmol/L内呈良好的线性关系(r~2>0.9990)。游离氨基酸平均回收率在97.80%~116.82%之间,RSD在1.93%~2.47%之间(n=6);水解氨基酸平均回收率在99.26%~111.23%之间,RSD在1.70%~2.23%之间(n=6)。结论柱前衍生RP-HPLC法灵敏、准确,具有良好的重复性和稳定性,适合对狭叶红景天中氨基酸含量的测定。
Objective To establish a reversed-phase high-performance liquid chromatography (RP-HPLC) method for the determination of free and hydrolyzed amino acids in Rhodiola sachalinensis. Methods Amino-acid column (4.6 mm × 250 mm, 5 μm) was used as the precursor of phenylisothiocyanate. Phase B was 7% (V: V) acetonitrile-CH_3COONa solution (pH = 6.5); the detection wavelength was 254 nm. Results 21 kinds of amino acids in 0.0019 ~ 2.700mmol / L showed a good linear relationship (r ~ 2> 0.9990). The average recoveries of free amino acids ranged from 97.80% to 116.82% with RSDs ranging from 1.93% to 2.47% (n = 6). The average recoveries of hydrolyzed amino acids ranged from 99.26% to 111.23% with RSDs ranging from 1.70% to 2.23% (N = 6). Conclusion The pre-column derivatization RP-HPLC method is sensitive, accurate, good repeatability and stability, suitable for the determination of amino acids in Rhodiola.