将连续变化的碰撞能量与串联质谱相结合,建立了能量分辨质谱(Energy-resolved mass spectrometry,ERMS)分析方法用以区分和检测人参皂苷同分异构体20(S)-Rf和Rg_(1).ERMS将子离子与母离子强度的比值定义为强度分数(Intensity fraction,I...将连续变化的碰撞能量与串联质谱相结合,建立了能量分辨质谱(Energy-resolved mass spectrometry,ERMS)分析方法用以区分和检测人参皂苷同分异构体20(S)-Rf和Rg_(1).ERMS将子离子与母离子强度的比值定义为强度分数(Intensity fraction,IF),两个子离子强度的比值定义为强度比(Intensity ratio,IR),并通过IF和IR分别对连续变化的碰撞能量作图建立特征碎片离子的IF和IR曲线.虽然20(S)-Rf和Rg_(1)的串联质谱图无明显差异,但是多元统计分析结果显示其特征碎片离子[M-Glc-H]-,[M-2Glc-H]-和[M-2Glc-C_(6)H_(12)-H]-的IF和IR曲线差异显著,有明显的区分边界,可以直接区分20(S)-Rf和Rg_(1).进一步利用ERMS实时分析了原人参三醇型皂苷生物转化产物中20(S)-Rf和Rg_(1)的相对摩尔含量.结果表明,ERMS不需液相色谱分离即可实现20(S)-Rf和Rg_(1)的快速区分和相对含量检测.展开更多
Objective To explore the synergic mechanism of ginsenoside Rg_(1)(Rg_(1))and aconitine(AC)by acting on normal neonatal rat cardiomyocytes(NRCMs)and pentobarbital sodium(PS)-induced damaged NRCMs.Methods The toxic,non-...Objective To explore the synergic mechanism of ginsenoside Rg_(1)(Rg_(1))and aconitine(AC)by acting on normal neonatal rat cardiomyocytes(NRCMs)and pentobarbital sodium(PS)-induced damaged NRCMs.Methods The toxic,non-toxic,and effective doses of AC and the most suitable compatibility concentration of Rg_(1) for both normal and damaged NRCMs exposed for 1 h were filtered out by 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-diphenytetrazoliumromide,respectively.Then,normal NRCMs or impaired NRCMs were treated with chosen concentrations of AC alone or in combination with Rg_(1) for 1 h,and the cellular activity,cellular ultrastructure,apoptosis,leakage of acid phosphatase(ACP)and lactate dehydrogenase(LDH),intracellular sodium ions[Na^(+)],potassium ions[K^(+)]and calcium ions[Ca^(2+)]levels,and Nav1.5,Kv4.2,and RyR_(2) genes expressions in each group were examined.Results For normal NRCMs,3000µmol/L AC significantly inhibited cell viability(P<0.01),promoted cell apoptosis,and damaged cell structures(P<0.05),while other doses of AC lower than 3000µmol/L and the combinations of AC and Rg_(1) had little toxicity on NRCMs.Compared with AC acting on NRCMs alone,the co-treatment of 3000 and 10µmol/L AC with 1µmol/L Rg_(1) significantly decreased the level of intracellular Ca^(2+)(P<0.01 or P<0.05),and the co-treatment of 3000µmol/L AC with 1µmol/L Rg_(1) significantly decreased the level of intracellular Ca^(2+)via regulating Nav1.5,RyR_(2) expression(P<0.01).For damaged NRCMs,1500µmol/L AC aggravated cell damage(P<0.01),and 0.1 and 0.001µmol/L AC showed moderate protective effect.Compared with AC used alone,the co-treatment of Rg_(1) with AC reduced the cell damage,0.1µmol/L AC with 1µmol/L Rg_(1) significantly inhibited the level of intracellular Na+(P<0.05),1500µmol/L AC with 1µmol/L Rg_(1) significantly inhibited the level of intracellular K+(P<0.01)via regulating Nav1.5,Kv4.2,RyR_(2) expressions in impaired NRCMs.Conclusion Rg_(1) inhibited the cardiotoxicity and enhanced the cardiotonic effect of AC via regulating the ion channels pathway of[Na^(+)],[K^(+)],and[Ca^(2+)].展开更多
目的探讨人参皂苷Rg_(3)对过氧化氢诱导的人晶状体上皮细胞氧化损伤的改善作用及对核转录因子E2相关因子2(nuclear factor E2 related factor 2,Nrf2)/血红素加氧酶-1(heme oxygenase 1,HO-1)信号通路的调节作用。方法用不同浓度人参皂...目的探讨人参皂苷Rg_(3)对过氧化氢诱导的人晶状体上皮细胞氧化损伤的改善作用及对核转录因子E2相关因子2(nuclear factor E2 related factor 2,Nrf2)/血红素加氧酶-1(heme oxygenase 1,HO-1)信号通路的调节作用。方法用不同浓度人参皂苷Rg_(3)处理过氧化氢诱导的SRA01/04细胞,用噻唑盐(methyl thiazolyl tetrazolium,MTT)法检测细胞存活率。将第3代对数生长期SRA01/04细胞随机分为正常组、氧化损伤组(用200μmol∙mL^(−1)过氧化氢处理)、人参皂苷Rg_(3)低剂量组和人参皂苷Rg_(3)高剂量组(分别用40、80μg∙mL^(−1)人参皂苷Rg_(3)处理6 h,更换培养基后用200μmol∙mL^(−1)过氧化氢处理12 h),用MTT法检测细胞存活率,用流式细胞仪检测细胞凋亡率,用试剂盒检测丙二醛(malondialdehyde,MDA)、超氧化物歧化酶(superoxidedismutase,SOD)和谷胱甘肽过氧化物酶(glutathioneperoxidase,GSH-Px)的含量,用蛋白印迹法检测Nrf2、Kelch样环氧氯丙烷相关蛋白1(Kelch like epichlorohydrin related protein 1,Keap1)和HO-1蛋白的相对表达量。结果与0μg∙mL^(−1)人参皂苷Rg_(3)组比较,10、20、40、80μg∙mL^(−1)人参皂苷Rg_(3)组的细胞存活率逐渐升高(P<0.05)。与正常组比较,氧化损伤组的细胞存活率、SOD和GSHPx含量以及Nrf2、Keap1和HO-1蛋白相对表达量降低,细胞凋亡率和MDA含量升高(P<0.05);与氧化损伤组比较,人参皂苷Rg_(3)低剂量和人参皂苷Rg_(3)高剂量组的细胞存活率、SOD和GSH-Px含量以及Nrf2、Keap1和HO-1蛋白的相对表达量升高,细胞凋亡率和MDA含量降低(P<0.05);人参皂苷Rg_(3)低剂量和人参皂苷Rg_(3)高剂量组各项指标水平变化规律相同,人参皂苷Rg_(3)高剂量组更显著(P<0.05)。结论人参皂苷Rg_(3)可抑制过氧化氢诱导的人晶状体上皮细胞凋亡,减轻氧化应激损伤,其可能是通过激活Nrf2/HO-1信号通路发挥调节作用的。展开更多
文摘将连续变化的碰撞能量与串联质谱相结合,建立了能量分辨质谱(Energy-resolved mass spectrometry,ERMS)分析方法用以区分和检测人参皂苷同分异构体20(S)-Rf和Rg_(1).ERMS将子离子与母离子强度的比值定义为强度分数(Intensity fraction,IF),两个子离子强度的比值定义为强度比(Intensity ratio,IR),并通过IF和IR分别对连续变化的碰撞能量作图建立特征碎片离子的IF和IR曲线.虽然20(S)-Rf和Rg_(1)的串联质谱图无明显差异,但是多元统计分析结果显示其特征碎片离子[M-Glc-H]-,[M-2Glc-H]-和[M-2Glc-C_(6)H_(12)-H]-的IF和IR曲线差异显著,有明显的区分边界,可以直接区分20(S)-Rf和Rg_(1).进一步利用ERMS实时分析了原人参三醇型皂苷生物转化产物中20(S)-Rf和Rg_(1)的相对摩尔含量.结果表明,ERMS不需液相色谱分离即可实现20(S)-Rf和Rg_(1)的快速区分和相对含量检测.
基金Supported by the National Natural Science Foundation of China(Nos.81630101 and 81891012)Department of Science and Technology of Sichuan Province(No.2018JZ0081)+1 种基金the Open Research Fund of Chengdu University of Traditional Chinese Medicine Key Laboratory of Systematic Research of Distinctive Chinese Medicine Resources in Southwest China(No.2020XSGG001)Special Project for the Central Government to Guide the Development of Local Science and Technology in Sichuan Province(No.20ZYKJCX0006)。
文摘Objective To explore the synergic mechanism of ginsenoside Rg_(1)(Rg_(1))and aconitine(AC)by acting on normal neonatal rat cardiomyocytes(NRCMs)and pentobarbital sodium(PS)-induced damaged NRCMs.Methods The toxic,non-toxic,and effective doses of AC and the most suitable compatibility concentration of Rg_(1) for both normal and damaged NRCMs exposed for 1 h were filtered out by 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-diphenytetrazoliumromide,respectively.Then,normal NRCMs or impaired NRCMs were treated with chosen concentrations of AC alone or in combination with Rg_(1) for 1 h,and the cellular activity,cellular ultrastructure,apoptosis,leakage of acid phosphatase(ACP)and lactate dehydrogenase(LDH),intracellular sodium ions[Na^(+)],potassium ions[K^(+)]and calcium ions[Ca^(2+)]levels,and Nav1.5,Kv4.2,and RyR_(2) genes expressions in each group were examined.Results For normal NRCMs,3000µmol/L AC significantly inhibited cell viability(P<0.01),promoted cell apoptosis,and damaged cell structures(P<0.05),while other doses of AC lower than 3000µmol/L and the combinations of AC and Rg_(1) had little toxicity on NRCMs.Compared with AC acting on NRCMs alone,the co-treatment of 3000 and 10µmol/L AC with 1µmol/L Rg_(1) significantly decreased the level of intracellular Ca^(2+)(P<0.01 or P<0.05),and the co-treatment of 3000µmol/L AC with 1µmol/L Rg_(1) significantly decreased the level of intracellular Ca^(2+)via regulating Nav1.5,RyR_(2) expression(P<0.01).For damaged NRCMs,1500µmol/L AC aggravated cell damage(P<0.01),and 0.1 and 0.001µmol/L AC showed moderate protective effect.Compared with AC used alone,the co-treatment of Rg_(1) with AC reduced the cell damage,0.1µmol/L AC with 1µmol/L Rg_(1) significantly inhibited the level of intracellular Na+(P<0.05),1500µmol/L AC with 1µmol/L Rg_(1) significantly inhibited the level of intracellular K+(P<0.01)via regulating Nav1.5,Kv4.2,RyR_(2) expressions in impaired NRCMs.Conclusion Rg_(1) inhibited the cardiotoxicity and enhanced the cardiotonic effect of AC via regulating the ion channels pathway of[Na^(+)],[K^(+)],and[Ca^(2+)].
文摘目的探讨人参皂苷Rg_(3)对过氧化氢诱导的人晶状体上皮细胞氧化损伤的改善作用及对核转录因子E2相关因子2(nuclear factor E2 related factor 2,Nrf2)/血红素加氧酶-1(heme oxygenase 1,HO-1)信号通路的调节作用。方法用不同浓度人参皂苷Rg_(3)处理过氧化氢诱导的SRA01/04细胞,用噻唑盐(methyl thiazolyl tetrazolium,MTT)法检测细胞存活率。将第3代对数生长期SRA01/04细胞随机分为正常组、氧化损伤组(用200μmol∙mL^(−1)过氧化氢处理)、人参皂苷Rg_(3)低剂量组和人参皂苷Rg_(3)高剂量组(分别用40、80μg∙mL^(−1)人参皂苷Rg_(3)处理6 h,更换培养基后用200μmol∙mL^(−1)过氧化氢处理12 h),用MTT法检测细胞存活率,用流式细胞仪检测细胞凋亡率,用试剂盒检测丙二醛(malondialdehyde,MDA)、超氧化物歧化酶(superoxidedismutase,SOD)和谷胱甘肽过氧化物酶(glutathioneperoxidase,GSH-Px)的含量,用蛋白印迹法检测Nrf2、Kelch样环氧氯丙烷相关蛋白1(Kelch like epichlorohydrin related protein 1,Keap1)和HO-1蛋白的相对表达量。结果与0μg∙mL^(−1)人参皂苷Rg_(3)组比较,10、20、40、80μg∙mL^(−1)人参皂苷Rg_(3)组的细胞存活率逐渐升高(P<0.05)。与正常组比较,氧化损伤组的细胞存活率、SOD和GSHPx含量以及Nrf2、Keap1和HO-1蛋白相对表达量降低,细胞凋亡率和MDA含量升高(P<0.05);与氧化损伤组比较,人参皂苷Rg_(3)低剂量和人参皂苷Rg_(3)高剂量组的细胞存活率、SOD和GSH-Px含量以及Nrf2、Keap1和HO-1蛋白的相对表达量升高,细胞凋亡率和MDA含量降低(P<0.05);人参皂苷Rg_(3)低剂量和人参皂苷Rg_(3)高剂量组各项指标水平变化规律相同,人参皂苷Rg_(3)高剂量组更显著(P<0.05)。结论人参皂苷Rg_(3)可抑制过氧化氢诱导的人晶状体上皮细胞凋亡,减轻氧化应激损伤,其可能是通过激活Nrf2/HO-1信号通路发挥调节作用的。
