山萸肉炮制前后11种成分的变化

Quantitative analysis of eleven active constituents in crude and processed Cornus officinalis

目的：以山萸肉中的11种化合物为指标,系统评价炮制工艺对其主要成分含量的影响。方法：以3批山萸肉为原料,分别依法进行酒蒸、酒炖或加压酒蒸炮制。采用Diamonsil（钻石）C18色谱柱（200 mm×4.6 mm,5μm）,以乙腈（A）-0.1%甲酸水（B）溶液为流动相,梯度洗脱（0～6 min,20%A;6～7 min,20%A→22%A;7～10 min,22%A;10～13 min,22%A→80%A;13～16 min,80%A;16～17 min,80%A→20%A;17～18 min,20%A）,流速1.0 m L·min-1,柱温30℃;采用电喷雾离子源（ESI）,多反应监测模式（MRM）检测,在同一周期内进行正负离子切换扫描,运用液相色谱-串联质谱（LC-MS/MS）技术同步测定并比较各样品中目标化合物成分的含量。结果：本文方法具有良好的专属性、线性（r〉0.995 3）、重复性（RSD〈5.31%）和回收率（93.2%～109.7%）。山萸肉中11种目标化合物的检测限和定量限范围分别为0.01～0.20和0.04～0.59μg·m L-1。11种化合物的日内和日间精密度分别小于7.32%和9.33%。3批山萸肉生品中没食子酸、5-羟甲基糠醛、獐芽菜苷、山茱萸苷、莫诺苷、马钱苷、7α-O-甲基莫诺苷、7β-O-甲基莫诺苷、7α-O-乙基莫诺苷、7β-O-乙基莫诺苷、山茱萸新苷的含量均值分别为0.26、0、0.54、0.28、7.26、6.67、0.048、0.068、0、0.012和2.09 mg·g-1,酒蒸品中对应的含量的均值分别为1.77、11.40、0.86、0.33、8.70、6.26、0.04、0.071、0.075、0.087和1.36 mg·g-1,酒炖品中对应含量的均值分别为1.94、11.38、0.89、0.25、10.39、9.26、0.04、0.061、0.083、0.096和1.19 mg·g-1,加压酒蒸品中对应含量的均值分别为2.10、16.87、0.90、0.22、9.37、8.31、0.039、0.068、0.052、0.056和1.08 mg·g-1。结论：与山萸肉生品相比,炮制后除了7α-O-甲基莫诺苷、山茱萸新苷含量下降外,其他活性成分含量均呈显著增加趋势;此外,7α-O-乙基莫诺苷和5-羟甲基糠醛为炮制后新增加的成分。

Objective：To systematically investigate the effect of processing on the contents of eleven active constituents of Cornus officinalis.Methods：Processed Cornus officinalis were prepared by steaming with wine,stewing with wine,pressurized steaming with wine from 3 different batches.The HPLC analysis was performed on a Diamonsil C18 column（200 mm×4.6 mm,5 ？m）with gradient elution using aqueous solution containing 0.1% formic acid and acetonitrile（A）as the mobile phase（0-6 min,20%A;6-7 min,20%A → 22%A;7-10 min,22%A;10-13 min,22%A → 80%A;13-16 min,80%A;16-17 min,80%A → 20%A;17-18 min,20%A）.The flow rate was 1.0 m L·min-1,and the column temperature was kept at 30℃.Electrospray ionization（ESI）source was adopted.Quantification was operated at multiple reaction monitoring（MRM）modes,and the switching scan was used from positive and negative ion during the same period.A liquid chromatography tandem mass spectrometry（LC-MS/MS）method was established for the simultaneous determination of eleven components in Cornus officinalis including 5-hydroxymethylfufural,gallic acid,sweroside,cornin,morroniside,loganin,7α-O-methylmorroniside,7β-O-methylmorroniside,7α-O-ethylmorroniside,7β-Oethylmorroniside,and cornuside.Results：The established method showed good specificity,linearity（r〉0.995 3）,repeatability（RSD〈5.31%）,and recovery（93.2%-109.7%）.The limits of detection and quantification of these compounds were in the range of 0.01-0.20 and 0.04-0.59 μg·m L-1,respectively.The intra- and interday precision was lower than 7.32% and 9.33%,respectively.In crude Cornus officinalis from 3 batch numbers,the average contents of gallic acid,5-hydroxymethylfufural,sweroside,cornin,morroniside,loganin,7α-Omethylmorroniside,7β-O-methylmorroniside,7α-O-ethylmorroniside,7β-O-ethylmorroniside,and cornuside were 0.26,0,0.54,0.28,7.26,6.67,0.048,0.068,0,0.012 and 2.09 mg·g-1,respectively;the average contents of eleven compounds in Jiuzhengpin were 1.77,11.40,0.86,0.33,8.70,6.26,0.04,0.071,0.075,0.087 and 1.36 mg·g-1,respectively;the average contents were 1.94,11.38,0.89,0.25,10.39,9.26,0.04,0.061,0.083,0.096 and 1.19 mg·g-1 in Jiudunpin;In Jiayajiuzhengpin,the average contents of these constituents were 2.10,16.87,0.90,0.22,9.37,8.31,0.039,0.068,0.052,0.056 and 1.08 mg·g-1,respectively.Conclusion：Compared to crude Cornus officinalis,the contents of most constituents increased after processing procedures,while the contents of 7α-O-methylmorroniside and cornuside decreased after processing.Moreover,the results indicated that 7α-O-ethylmorroniside and 5-hydroxymethylfufural are the newly generated chemical constituents during processing.This method can be used as a comprehensive quality control for Cornus officinalis.