多维度定性定量评价电子束辐照灭菌对黄芪质量的影响

Multi-Dimensional Qualitative and Quantitative Evaluation of the Influence of Electron Beam Irradiation Sterilization on Astragalus Quality

为探究电子束辐照灭菌对黄芪质量的影响,采用不同辐照吸收剂量(2、4、6、8、10、12、14、16、18 kGy)处理黄芪样品,测定其微生物水平、多种化学成分含量、抗氧化活性及大鼠肝星状细胞(HSC-T6)的细胞存活率,比较灭菌前后表面增强拉曼光谱(SERS)指纹图谱及高效液相色谱(HPLC)指纹图谱。结果表明,吸收剂量≥10 kGy时,黄芪微生物水平符合《中华人民共和国药典2020年版四部》规定;不同吸收剂量下黄芪中毛蕊异黄酮含量、毛蕊异黄酮苷含量、刺芒柄花素含量、芒柄花苷含量、2,2-联苯基-1-苦基肼基自由基(DPPH·)清除率、3-氧代-2苯基-4,4,5,5-四甲基咪唑啉-1氧自由基(PTIO·)清除率、铁氰化钾还原力及大鼠肝星状细胞(HSC-T6)存活率较灭菌前均无显著差异(P>0.05);灭菌后黄芪SERS指纹图谱、HPLC指纹图谱与未灭菌样品图谱相比,相似度均大于0.900;液相色谱-质谱联用(LC-MS)确认HPLC指纹图谱中13个共有峰结构,灰色关联度分析显示13个共有峰与抗氧化活性关联度均大于0.600,毛蕊异黄酮、毛蕊异黄酮苷、刺芒柄花素、芒柄花苷含量与黄芪抗氧化活性的关联度均大于0.700。综上,吸收剂量18 kGy及以下的电子束辐照灭菌对黄芪质量无影响,上述4种化合物含量与黄芪抗氧化性具有较强的相关性,可作为其抗氧化活性的质量标志物。本研究可为电子束灭菌技术在黄芪灭菌加工中的应用提供理论依据和技术支持。

To investigate the effect of electron beam irradiation sterilization on the quality of Astragalus,the samples were treated with different absorption doses(2,4,6,8,10,12,14,16,18 kGy),and the microbial levels,chemical components,antioxidant activity and cell survival rate of rat hepatic stellate cells(HSC-T6)were determined.Surface-enhanced Raman spectroscopy(SERS)fingerprints and high performance liquid chromatography(HPLC)fingerprints of the samples were compared before and after sterilization.The results showed that,the level of Astragalus microbial conforms to the pharmacopoeia of the People’s Republic of China 2020 Edition of the fourth part when the absorption dose≥10 kGy.There were no significant differences among the contents of calycosin,calycosin-7-O-β-D-glucopyranoside,formononetin,ononin,2,2-diphenyl-1-picrylhydrazyl radical(DPPH·)clearance rate,2-phenyl-4,4,5,5-tetramethylimidazoline-3-oxide-1-oxyl radical(PTIO·)clearance rate,potassium ferricyanide reducing power and survival rate of rat hepatic stellate cells(HSC-T6)in Astragalus at different absorbed doses compared with those before sterilization(P>0.05).The SERS fingerprints and HPLC fingerprints of Astragalus after sterilization were compared with those of non-sterilized samples,and the similarity were both greater than 0.900.Liquid chromatograph-mass spectrometer(LC-MS)confirmed 13 common peak structures in the HPLC fingerprints,and grey correlation analysis revealed that the correlations between 13 common peaks and antioxidant activity were greater than 0.600.The correlation between the antioxidant activity of Radix Astragalus and calycosin,calycosin-7-o-β-d-glucopyranoside,formononetin and ononin were greater than 0.700.In summary,electron beam irradiation sterilization with an absorbed dose of 18 kGy or below had no effect on the quality of Astragalus.The above four compounds had strong correlation with the antioxidant activity of Astragalus,and could be used as quality markers of Astragalus’antioxidant activity.This study provides theoretical basis and technical support for the application of electron beam sterilization technology in Astragalus sterilization processing.