RRLC-Q-TOF MS法分析鲜人参与仙人掌果配伍发酵前后人参皂苷成分的变化
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摘要
采用高分离度快速液相色谱-四极杆飞行时间质谱(RRLC-Q-TOF MS)法对鲜人参与仙人掌果配伍发酵过程中的人参皂苷进行定性和定量分析。采用Agilent Zorbax SB-C18柱(2.1m×150mm×3.5μm),以0.1%甲酸和乙腈作为流动相进行梯度洗脱,在电喷雾离子源负离子模式下进行质谱检测。结果表明,在发酵液中共鉴别了27种人参皂苷,通过其总离子流图比较了发酵前后人参皂苷成分的差异,对发酵后含量明显增加的人参皂苷Rh1、Rg2、F2、Rg3、Rh2、CK进行了定量分析,确定其含量分别为6.345 2、20.452 2、6.255 9、27.452 8、55.384 6、30.472 9mg/L。利用该方法能够快速发现人参配伍后产生的新化合物,并依据人参皂苷类化合物的质谱裂解规律对其进行鉴定。
Ginseng and Radix Acanthopanacis(Panax ginseng C.A.Mey.)are dry roots,which are widely used in the field of medicine and food.The cactus fruit flesh contain polysaccharides and flavonoids,showing that it can activate blood,eliminate dampness cooling,improve immunity.Therefore,this study focused on the quantitative analysis of rare ginsenoside Rh1,Rg2,F2,Rg3,Rh2 and CK in the fermentation broth.This research was designed to evaluate the effect of ginseng and cactus fruit as raw material and fermented health wine on health functions such as improving immuni-ty,anti fatigue and preventing senile dementia.In order to discover new compounds produced by the compatibility of ginseng,rapid resolution liquid chromatography coupled with quadruple-time-of-flight tandem mass spectrometry(RRLC-Q-TOF MS)was used for the qualitative and quantitative analysis of the content of ginsenosides in the fermentation liquid of fresh ginseng with cactus pear fruit.The analysis was carried out on an Agilent Zorbax SB-C18 column(2.1 m×150 mm×3.5μm)by gradient elution with acetonitrile-water(0.1% formic acid)as mobile phase,and was detected by RRLC-Q-TOF MS with negative electrospray ionization mode.Twenty-seven ginsenosides were identified,and the contents of ginsenoside Rh1,Rg2,F2,Rg3,Rh2 and CK increased obviously,which reached to 6.345 2,20.452 2,6.255 9,27.452 8,55.384 6,30.472 9 mg/L in the fermentation liquid,respectively.The findings suggested that 19 kinds of ginsenosides are detected before fermentation,while 27 kinds of ginsenosides are detected after fermentation.The method will be applied to find new compounds after being compatibility and to identify ginsenosides based on fragment pathway.
Ginseng and Radix Acanthopanacis(Panax ginseng C.A.Mey.)are dry roots,which are widely used in the field of medicine and food.The cactus fruit flesh contain polysaccharides and flavonoids,showing that it can activate blood,eliminate dampness cooling,improve immunity.Therefore,this study focused on the quantitative analysis of rare ginsenoside Rh1,Rg2,F2,Rg3,Rh2 and CK in the fermentation broth.This research was designed to evaluate the effect of ginseng and cactus fruit as raw material and fermented health wine on health functions such as improving immuni-ty,anti fatigue and preventing senile dementia.In order to discover new compounds produced by the compatibility of ginseng,rapid resolution liquid chromatography coupled with quadruple-time-of-flight tandem mass spectrometry(RRLC-Q-TOF MS)was used for the qualitative and quantitative analysis of the content of ginsenosides in the fermentation liquid of fresh ginseng with cactus pear fruit.The analysis was carried out on an Agilent Zorbax SB-C18 column(2.1 m×150 mm×3.5μm)by gradient elution with acetonitrile-water(0.1% formic acid)as mobile phase,and was detected by RRLC-Q-TOF MS with negative electrospray ionization mode.Twenty-seven ginsenosides were identified,and the contents of ginsenoside Rh1,Rg2,F2,Rg3,Rh2 and CK increased obviously,which reached to 6.345 2,20.452 2,6.255 9,27.452 8,55.384 6,30.472 9 mg/L in the fermentation liquid,respectively.The findings suggested that 19 kinds of ginsenosides are detected before fermentation,while 27 kinds of ginsenosides are detected after fermentation.The method will be applied to find new compounds after being compatibility and to identify ginsenosides based on fragment pathway.
引文
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[7]杨婧娟.三七总皂苷的发酵辅助提取及其活性评价[D].浙江:浙江大学,2013.
[8]FELKAI-HADDACHE L,DAHMOUNE F,REMINI H,et al.Microwave optimization of mucilage extraction from Opuntia ficus indica Cladodes[J].International Journal of Biological Macromolecules,2015,84(8):24.
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[10]MSADDAK L,ABDELHEDI O,KRIDENE A,et al.Opuntia ficus-indica,cladodes as a functional ingredient:bioactive compounds profile and their effect on antioxidant quality of bread[J].Lipids in Health&Disease,2017,16(1):32.
[11]张嘉麟,徐文安,杨荫康,等.三七中人参皂甙对老年鼠血液中抗氧化酶活力影响的研究[J].昆明医科大学学报,2000,21(2):63-65.ZHANG Jialin,XU Wen’an,YANG Yinkang,et al.A preliminary study on the relationship between the effect of Panax notoginsen Rb1and Rg1on the aging process[J].Academic Journal of Kunming Medical College,2000,21(2):63-65(in Chinese).
[12]郝颖,于珊珊,戴雨霖,等.RRLC-Q-TOF MS/MS法分析生晒参和大力参中的皂苷类成分[J].质谱学报,2014,35(4):311-316.HAO Ying,YU Shanshan,DAI Yulin,et al.Study on ginsenosides in white ginseng and dali ginseng by RRLC-Q-TOF MS/MS[J].Journal of Chinese Mass Spectrometry Society,2014,35(4):311-316(in Chinese).
[13]SUN J,WU W,GUO Y,et al.Pharmacokinetic study of ginsenoside Rc and simultaneous determination of its metabolites in rats using RRLC-QTOF-MS[J].Journal of Pharmaceutical&Biomedical Analysis,2014,88:16-21.
[14]张晓旭,王洪平,杨阳,等.超高效液相色谱-串联四极杆飞行时间质谱法快速鉴别生晒参中的皂苷类成分[J].中国医药导报,2015,12(9):130-136.ZHANG Xiaoxu,WANG Hongping,YANG Yang,et al.Rapid analysis of ginsenosides in dried fresh ginseng by ultra-performance liquid chromatography with quadrupole time-of-flight mass spectrometry[J].China Medical Herald,2015,12(9):130-136(in Chinese).
[15]YONG-KOOK K,SUK A M,SUK P J,et al.Discrimination of cultivation ages and cultivars of ginseng leaves using Fourier transform infrared spectroscopy combined with multivariate analysis[J].Journal of Ginseng Research,2014,38(1):52-58.
[16]郁杰,秦枫,洪伟鸣,等.HPLC-MS/MS法检测三七茎叶总皂苷中人参皂苷Rb1的含量[J].江苏农业科学,2010,(5):370-372.YU Jie,QIN Feng,HONG Weiming,et al.Simultaneous determination of ginsenoside Rb1in total saponins in leaves of Panax notoginseng by HPLC-MS/MS[J].Jiangsu Agricultural Sciences,2010,(5):370-372(in Chinese).
[17]金高娃,丁俊杰,陈雪,等.反相/亲水色谱法分析糖苷类化合物[J].分析测试学报,2014,33(2):133-137.JIN Gaowa,DING Junjie,CHEN Xue,et al.Separation of glucoside compounds by reversedphase and hydrophilic interaction liquid chromatography[J].Journal of Instrumental Analysis,2014,33(2):133-137(in Chinese).
[18]刘欣.人参皂苷Rb水解酶的分离纯化及其性质的研究[D].大连:大连医科大学,2003.
[19]杨宁,杨世林,赵余庆.稀有人参皂苷糖苷化合成方法的研究进展[J].现代药物与临床,2014,29(5):574-582.YANG Ning,YANG Shilin,ZHAO Yuqing.Research progress on glycosylation methods of rare ginsenosides[J].Drugs&Clinic,2014,29(5):574-582(in Chinese).
[20]刘莉,齐洋.降糖宁胶囊中人参皂苷Rg1、Re的含量方法学考察[J].黑龙江医药,2014,(2):304-306.LIU Li,QI Yang.Methodological study on contents of ginsenoside Rg1,ginsenoside Re in Jiangtangning capsules[J].Heilongjiang Medicine Journal,2014,(2):304-306(in Chinese).
[21]南国,张鹏,王萌,等.HPLC法测定稳心颗粒中三七皂苷R1、人参皂苷Rg1、Rb1、Rd与党参炔苷的含量[J].天津中医药,2016,33(7):434-436.NAN Guo,ZHANG Peng,WANG Meng,et al.Determination of notoginsenoside R1,ginsenoside Rg1,Rb1,Rd and lobetyolin in Wenxin Keli by HPLC[J].Tianjin Journal of Traditional Chinese Medicine,2016,33(7):434-436(in Chinese).
[22]唐艳,闫述模,汪静静,等.基于UPLC及多成分分析的西洋参质量评价[J].中国中药杂志,2016,41(9):1 678-1 684.TANG Yan,YAN Shumo,WANG Jingjing,et al.Quality evaluation of American ginseng using UPLC coupled with multivariate analysis[J].China Journal of Chinese Materia Medica,2016,41(9):1 678-1 684(in Chinese).
[23]马帅,刘新瑶,林柯.人参皂苷在人参发酵酒中变化规律的研究[J].农产品加工,2016,(11):9-13.MA Shuai,LIU Xinyao,LIN Ke.Change law of ginseng saponins in Panax ginseng extraction research[J].Academic Periodical of Farm Products Processing,2016,(11):9-13(in Chinese).
[2]XIAO D,YUE H,XIU Y,et al.Accumulation characteristics and correlation analysis of five ginsenosides with different cultivation ages from different regions[J].Journal of Ginseng Research,2015,39(4):338-344.
[3]李慧,许亮,温美佳,等.不同产地人参皂苷成分含量UPLC法测定及质量评价[J].中华中医药杂志,2015,(6):1 963-1 968.LI Hui,XU Liang,WEN Meijia,et al.Study on the content measurement and quality evaluation of ginsenoside constituent from different habitats with UPLC analysis method[J].China Journal of Traditional Chinese Medicine and Pharmacy,2015,(6):1 963-1 968(in Chinese).
[4]CHEN Y J,ZHAO Z Z,CHEN H B,et al.Determination of ginsenosides in Asian and American ginsengs by liquid chromatography quadrupole time of flight MS:assessing variations based on morphological characteristics[J].Journal of Ginseng Research,2015,41(1):10-22.
[5]刘丹,濮社班,朱玲英,等.RP-HPLC法同时测定不同产地红参中6种人参皂苷的含量[J].药学进展,2011,35(6):278-281.LIU Dan,PU Sheban,ZHU Lingying,et al.Simultaneous determination of six ginsenosides in red ginseng of different origins by RP-HPLC[J].Progress in Pharmaceutical Sciences,2011,35(6):278-281(in Chinese).
[6]郑志忠,明艳林.人参皂苷体内代谢及其产物的生物活性[J].海峡药学,2012,24(1):9-13.ZHENG Zhizhong,MING Yanlin.Metabolism of ginsenoside in vivo and biological activities of metabolites[J].Strait Pharmaceutical Journal,2012,24(1):9-13(in Chinese).
[7]杨婧娟.三七总皂苷的发酵辅助提取及其活性评价[D].浙江:浙江大学,2013.
[8]FELKAI-HADDACHE L,DAHMOUNE F,REMINI H,et al.Microwave optimization of mucilage extraction from Opuntia ficus indica Cladodes[J].International Journal of Biological Macromolecules,2015,84(8):24.
[9]ORDAZOCHOA G,JUREZCARATACHEA A,PREZSNCHEZ R E,et al.Effect of spineless cactus intake(Opuntia ficus-indica)on blood glucose levels in lactating sows and its impact on feed intake,body weight loss,and weaning-estrus interval[J].Tropical Animal Health&Production,2017,49(5):1 025-1 033.
[10]MSADDAK L,ABDELHEDI O,KRIDENE A,et al.Opuntia ficus-indica,cladodes as a functional ingredient:bioactive compounds profile and their effect on antioxidant quality of bread[J].Lipids in Health&Disease,2017,16(1):32.
[11]张嘉麟,徐文安,杨荫康,等.三七中人参皂甙对老年鼠血液中抗氧化酶活力影响的研究[J].昆明医科大学学报,2000,21(2):63-65.ZHANG Jialin,XU Wen’an,YANG Yinkang,et al.A preliminary study on the relationship between the effect of Panax notoginsen Rb1and Rg1on the aging process[J].Academic Journal of Kunming Medical College,2000,21(2):63-65(in Chinese).
[12]郝颖,于珊珊,戴雨霖,等.RRLC-Q-TOF MS/MS法分析生晒参和大力参中的皂苷类成分[J].质谱学报,2014,35(4):311-316.HAO Ying,YU Shanshan,DAI Yulin,et al.Study on ginsenosides in white ginseng and dali ginseng by RRLC-Q-TOF MS/MS[J].Journal of Chinese Mass Spectrometry Society,2014,35(4):311-316(in Chinese).
[13]SUN J,WU W,GUO Y,et al.Pharmacokinetic study of ginsenoside Rc and simultaneous determination of its metabolites in rats using RRLC-QTOF-MS[J].Journal of Pharmaceutical&Biomedical Analysis,2014,88:16-21.
[14]张晓旭,王洪平,杨阳,等.超高效液相色谱-串联四极杆飞行时间质谱法快速鉴别生晒参中的皂苷类成分[J].中国医药导报,2015,12(9):130-136.ZHANG Xiaoxu,WANG Hongping,YANG Yang,et al.Rapid analysis of ginsenosides in dried fresh ginseng by ultra-performance liquid chromatography with quadrupole time-of-flight mass spectrometry[J].China Medical Herald,2015,12(9):130-136(in Chinese).
[15]YONG-KOOK K,SUK A M,SUK P J,et al.Discrimination of cultivation ages and cultivars of ginseng leaves using Fourier transform infrared spectroscopy combined with multivariate analysis[J].Journal of Ginseng Research,2014,38(1):52-58.
[16]郁杰,秦枫,洪伟鸣,等.HPLC-MS/MS法检测三七茎叶总皂苷中人参皂苷Rb1的含量[J].江苏农业科学,2010,(5):370-372.YU Jie,QIN Feng,HONG Weiming,et al.Simultaneous determination of ginsenoside Rb1in total saponins in leaves of Panax notoginseng by HPLC-MS/MS[J].Jiangsu Agricultural Sciences,2010,(5):370-372(in Chinese).
[17]金高娃,丁俊杰,陈雪,等.反相/亲水色谱法分析糖苷类化合物[J].分析测试学报,2014,33(2):133-137.JIN Gaowa,DING Junjie,CHEN Xue,et al.Separation of glucoside compounds by reversedphase and hydrophilic interaction liquid chromatography[J].Journal of Instrumental Analysis,2014,33(2):133-137(in Chinese).
[18]刘欣.人参皂苷Rb水解酶的分离纯化及其性质的研究[D].大连:大连医科大学,2003.
[19]杨宁,杨世林,赵余庆.稀有人参皂苷糖苷化合成方法的研究进展[J].现代药物与临床,2014,29(5):574-582.YANG Ning,YANG Shilin,ZHAO Yuqing.Research progress on glycosylation methods of rare ginsenosides[J].Drugs&Clinic,2014,29(5):574-582(in Chinese).
[20]刘莉,齐洋.降糖宁胶囊中人参皂苷Rg1、Re的含量方法学考察[J].黑龙江医药,2014,(2):304-306.LIU Li,QI Yang.Methodological study on contents of ginsenoside Rg1,ginsenoside Re in Jiangtangning capsules[J].Heilongjiang Medicine Journal,2014,(2):304-306(in Chinese).
[21]南国,张鹏,王萌,等.HPLC法测定稳心颗粒中三七皂苷R1、人参皂苷Rg1、Rb1、Rd与党参炔苷的含量[J].天津中医药,2016,33(7):434-436.NAN Guo,ZHANG Peng,WANG Meng,et al.Determination of notoginsenoside R1,ginsenoside Rg1,Rb1,Rd and lobetyolin in Wenxin Keli by HPLC[J].Tianjin Journal of Traditional Chinese Medicine,2016,33(7):434-436(in Chinese).
[22]唐艳,闫述模,汪静静,等.基于UPLC及多成分分析的西洋参质量评价[J].中国中药杂志,2016,41(9):1 678-1 684.TANG Yan,YAN Shumo,WANG Jingjing,et al.Quality evaluation of American ginseng using UPLC coupled with multivariate analysis[J].China Journal of Chinese Materia Medica,2016,41(9):1 678-1 684(in Chinese).
[23]马帅,刘新瑶,林柯.人参皂苷在人参发酵酒中变化规律的研究[J].农产品加工,2016,(11):9-13.MA Shuai,LIU Xinyao,LIN Ke.Change law of ginseng saponins in Panax ginseng extraction research[J].Academic Periodical of Farm Products Processing,2016,(11):9-13(in Chinese).