对叶百部抗肺炎支原体活性部位的血清药物化学研究

Preliminary study on serum pharmacochemistry of anti-Mycoplasma pneumoniae elution fraction in Stemona tuberosa Lour. extract

目的:应用血清药物化学理论分析检测对叶百部抗肺炎支原体(Mycoplasma pneumoniae,MP)活性部位的体内成分。方法:利用D101大孔树脂进行梯度洗脱,不同洗脱部位进行抗MP实验,测定其最小抑菌浓度值。采用超高效液相色谱与飞行时间质谱联用(UPLC-Q-TOF-MS/MS)及MetaboLynx数据处理技术分析鉴定抗MP活性部位的血中移行成分。分析采用Acquity UPLC BEH C18色谱柱,流动相为0.1%甲酸水溶液(A)-0.1%甲酸乙腈(B),梯度洗脱,流速为0.40mL·min^(-1),柱温为45℃。结果:比较大孔树脂富集各洗脱部位最小抑菌浓度,水洗脱部位为29.41 mg·mL^(-1),20%乙醇洗脱部位为15.62mg·mL^(-1),50%乙醇洗脱部位为10.42mg·mL^(-1),90%乙醇洗脱部位为7.81mg·mL^(-1)。检测并鉴定了90%乙醇洗脱部位灌胃大鼠血中移行成分54个,包括14个原型成分和40个代谢产物。结论:对叶百部经大孔树脂富集90%乙醇洗脱部位抗MP活性最强,为抗MP活性部位。金刚大碱、百部新碱、对叶百部碱、新对叶百部碱、异对叶百部碱、N-氧-对叶百部碱、对叶百部碱J以及对叶百部碱H等成分可能为对叶百部抗MP的部分物质基础。

OBJECTIVE To analyze the main chemical constituents and metabolites in vivo of Stemona tuberosa anti-Mycoplasma pneumoniae（MP）elution fraction in rats after oral administration by serum pharmacochemistry.METHODS The gradient elution was performed by using D101 macroporous absorbing resin,to perform the in vitro anti-MP experiment and test the minimal inhibitory concentration（MIC）.Rapid identification of transitional constituents absorbed into blood after oral administration of anti-MP elution fraction was carried out by UPLC-Q-TOF-MS/MS and MetaboLynx software.The analysis was performed on an Acquity UPLC BEH C18 column,mobile phase was 0.1%formic acid aqueous solution（A）-0.1%formic acid and acetonitrile（B）with gradient elution,flow rate was 0.40 mL·min-1,and column temperature was maintained at 45℃.RESULTS The active fraction of anti-MP of the radix of Stemona tuberosa was filtered and extracted to the attachment resin where the factions were washed by the different concentrations of alcohol（0% alcohol,20% alcohol,50% alcohol,and 90% alcohol）,MIC values were 29.41,15.62,10.42,7.81 mg·mL-1,respectively.As a result,54 compounds including 14 prototype components and 40 metabolites in rat plasma after oral administration of anti-MP elution fraction were characterized.CONCLUSION The active fraction of Stemona tuberosa,90% ethanol elution fraction,is obtained by evaluating its efficacy in treating MP.Croomine,stemoninine,tuberostemonine,neotuberostemonine,isotuberostemonine,N-oxy-tuberostemonine,tuberostemonine J and tuberostemonine H may be parts of material basis of Stemona tuberosa on anti-MP.