目的 基于HPLC法探讨炮制辅料黄酒对九蒸九晒制备得到的熟地黄的化学成分动态变化的影响差异性。方法 色谱柱为Hypersil GOLD aQ C₁₈色谱柱(250 mm×4.6 mm,5μm),体积流量为1 mL/min,柱温为25℃。梓醇定量测定:流动相为乙腈-0.1%磷酸水(1:99),检测波长为210 nm;地黄苷A定量测定:流动相为乙腈-0.1%磷酸水(3:97),检测波长为203 nm;地黄苷D和益母草苷定量测定:流动相为乙腈-0.1%磷酸水(4:96),检测波长为203 nm;毛蕊花糖苷定量测定:流动相为乙腈-0.1%磷酸水(16:84),检测波长为334 nm;5-羟甲基糠醛(5-HMF)定量测定:流动相为乙腈-0.1%磷酸水(11:89),检测波长为284 nm。糖类成分的测定:色谱柱为Accurasil NH₂柱(250 mm×4.6 mm,5μm),流动相为乙腈-水(72:28),体积流量为1 mL/min,柱温25℃,检测器为ELSD,Alltech2000ES,漂移管温度93℃,气体体积流量2.6 L/min。采用SPSS 20.0进行统计分析,将清蒸-九蒸九晒与酒蒸-九蒸九晒制得的不同蒸晒次数的熟地黄中12种化学成分定量测定数据进行配对样本t-检验。结果 随蒸晒次数的增加,清蒸法与酒蒸法制得熟地黄中梓醇及益母草苷、毛蕊花糖苷、水苏糖、蔗糖、棉子糖均减少;地黄苷A及地黄苷D略为增加;5-HMF、果糖、葡萄糖、甘露三糖均增加。结论 建立的方法符合方法学验证要求。在古法熟地黄九蒸九晒的炮制过程中,炮制辅料黄酒对炮制品的质量存在显著影响,且清蒸法与酒蒸法均以第3次、第4次及第6次蒸晒所得熟地黄的相关物质的量呈较大的波动性。

Objective By HPLC, to explore the differences of adjuvants for the changes of chemical components of Rehmanniae Radix Praeparata during process by steamed for nine times and shined for nine times.Methods HPLC was conducted on a Hypersil GOLD aQ C₁₈ column(250 mm×4.6 mm, 5μm), flow rate was 1 mL/min, and the column temperature was 25℃.With the mobile phase of acetonitrile and 0.1% phosphoric acid water(1:99), the detection wavelength was 210 nm(catalpol), 203 nm(rehmannioside A, rehmannioside D, and leonuride), acetonitrile and 0.1% phosphoric acid water(3:97 and 4:96);334 nm(actecosode), acetonitrile and 0.1% phosphoric acid water(16:84), and 284 nm(5-HMF), acetonitrile and 0.1% phosphoric acid water(11:89).Sugars was conducted on an Accurasil NH₂ column(250 mm×4.6 mm, 5μm) with the mobile phase of acetonitrile and 0.1% phosphoric acid water(72:28), flow rate was 1 mL/min, the column temperature was 25℃.The detector was ELSD, Alltech2000ES, drift tube temperature was 93℃, and the gas flow rate was 2.6 L/min.The statistic analysis was carried out by SPSS 20.0 and the paired sample t-test was carried out for 12 kinds of chemical composition in Rehmanniae Radix Praeparata obtained by steamed for nine times and shined for nine times and wine-steamed for nine times and shined for nine times.Results With the increase of steamed times, the contents of catalpol, leonuride, actecosode, stachyose, sucrose, and raffinose in Rehmanniae Radix Praeparata obtained by two ways are all decreased;While the contents of rehmannioside A and rehmannioside D were slightly increased;the contents of 5-HMF, fructose, glucose, and mannose trisaccharide were increased.Conclusion The established method conforms to the validation requirement of the methodology.In the process of Rehmanniae Radix Praeparata with steamed for nine times and shined for nine times, processing adjuvant has a marked impact on the quality of the processed product.And the related substances of Rehmanniae Radix Praeparata obtained by the two ways have a larger fluctuation in times 3, 4, and 6 of steamed and shined processes, and the reason for this still needs to be further studied.

山西省中药现代化关键技术研究振东专项(2014ZD0302)