基于多组分分析的百合地黄汤古、今组方化学成分差异性研究

Study on chemical composition difference between ancient and modern Baihe Dihuang decoction based on multi-component analysis

目的:采用多组分分析法研究百合地黄汤古代与现代临床组方化学成分差异。方法:采用蒽酮-硫酸-紫外分光光度计法在580 nm紫外光下对百合地黄汤多糖提取液进行测定。采用高效液相色谱串联电喷雾检测器(UPLC-CAD),使用Agilent Infinity Lab Poroshell 120 HILIC-Z(3.0 mm×100 mm,2.7μm)色谱柱,流动相A为0.2%氨水-乙腈,流动相B为0.2%氨水-纯水,梯度洗脱,流速0.8 mL·min^(-1),柱温为35℃,CAD检测器雾化器温度70℃,分别对百合地黄汤古代方和现代方中6个单、寡糖成分进行定量分析。采用高效液相色谱-四极杆串联飞行时间质谱(UPLC-Q TOF MS/MS)及高效液相色谱串联三重四极杆质谱(UPLC-QQQ MS),分析百合地黄汤古代方与现代方的差异性成分,并对9个差异性成分进行定量分析。结果:百合地黄汤多糖与6个单寡糖及9个差异性成分在各自的测定范围内线性关系良好(r>0.999),平均加样回收率在97.7%~104.9%,RSD≤3.1%。精密度、重复性、稳定性均符合要求。结果表明百合地黄汤古代方与现代方的多糖、单寡糖含量以及9个差异性成分含量均具有显著性差异(P<0.05)。结论:本实验从多种化学成分角度明确了百合地黄汤古、今组方的差异。所采用方法均准确灵敏,稳定性和重复性好,可为百合地黄汤的临床应用与高品质中药制剂的开发提供参考。

Objective:To study the difference of chemical composition between ancient and modern clinical prescriptions of Baihe Dihuang decoction by multi-component analysis.Methods:The polysaccharide extract of Baihe Dihuang decoction was determined by anthrone-sulfuric acid-ultraviolet spectrophotometer under 580 nm ultraviolet light.Six monosaccharides and oligosaccharides in the ancient and modern Baihe Dihuang decoction were determined by UPLC-CAD.An Agilent InfinityLab Poroshell 120 HILIC-Z(3.0×100 mm,2.7μm)column was used.The mobile phase was consisted of A 0.2%ammonia in acetonitrile and 0.2%ammonia in water with gradient elution.The flow rate was 0.8 mL·min^(-1),the column temperature 35℃and the CAD detector nebulizer temperature was 70℃.Data acquisition frequency was 10 Hz and the filter was 1.05 s.The injection volumn was 2μL.The non-polar components were qualitatively and quantitatively analyzed by high performance liquid-quadrupole tandem time-of-flight mass spectrometry(UPLC-Q TOF MS/MS)and high performance liquid-phase tandem triple quadrupole mass spectrometry(UPLC-QQQ MS),respectively.The differential components of ancient and modern Baihe Dihuang Decoction were analyzed and 9 components were determined.Results:The linear relationship of 6 mono-oligosaccharides/polysaccharides and 9 differential components was good(r>0.999).The average recoveries ranged from 97.7%to 104.9%with RSDs≤3.1%.Precision,repeatability and stability met the requirements.The results showed that the conents of all the analytes in ancient and modern Baihe Dihuang decoction were significantly different(P<0.05).Conclusion:The methods used in this experiment are accurate,sensitive,stable and reproducible,which can provide reference for the quality control and pharmacodynamic material basic research of Baihe Dihuang Decoction.