基于液相色谱-质谱联用技术定性定量分析黑豆种皮中可溶型和结合型花青素
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摘要
以超声波辅助有机溶剂提取法获得黑豆种皮可溶型花青素提取物,再进一步对不含可溶型花青素的黑豆种皮残渣使用酸水解和碱水解以及酸碱/碱酸轮提水解,获得黑豆种皮结合型花青素提取物。采用超高效液相色谱-电喷雾-四极杆飞行时间质谱联用仪分析鉴定各黑豆种皮提取物中所含有的共17种花青素成分,包括11种花青素糖苷类:飞燕草素-3-O-葡萄糖苷、飞燕草素-3-O-半乳糖苷、矮牵牛花素-3-O-葡萄糖苷、矮牵牛花素-3-O-半乳糖苷、天竺葵素-3-O-芸香糖苷、矢车菊素-3-O-葡萄糖苷、矢车菊素-3-O-半乳糖苷、天竺葵素-3-O-己糖苷、芍药花素-3-O-己糖苷、天竺葵素-3-O-(6"-丙二酰葡萄糖苷)和芹菜定-3-O-(6"-丙二酰葡萄糖苷);6种花青素苷元:飞燕草素、矢车菊素、矮牵牛花素、天竺葵素、芹菜定和芍药花素。采用高效液相色谱-电喷雾-三重四极杆质谱联用仪精确定量各类黑豆种皮提取物中的花青素含量,结果表明,酸性结合型花青素提取物中结合型花青素的总含量最高。此外,在黑豆种皮的可溶型花青素提取物中,花青素主要以花青素糖苷类形式存在,苷元含量相对极少;而在结合型花青素提取物中,则主要以花青素苷元为主,糖苷类化合物相对少见。
Ultrasonic-assisted organic solvent extraction was used to obtain soluble anthocyanins from black soybean seed coat, and bound anthocyanins from the extraction residue were obtained by treatment with acid, alkaline, sequential acid/alkali hydrolysis or the reverse sequence. By ultra-high performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry(UPLC-ESI-QTOF-MS), a total of 17 anthocyanin components,including 11 anthocyanin glycosides: delphinidin-3-O-glucoside, delphinidin-3-O-galactoside, petunidin-3-O-glucoside,petunidin-3-O-galactoside, pelargonidin-3-O-rutinoside, cyanidin-3-O-glucoside, cyanidin-3-O-galactoside, pelargonidin-3-O-hexoside, peonidin-3-O-hexoside, pelargonidin-3-O-(6"-malonyl glucoside) and apigeninidin-3-O-(6"-malonyl glucoside);and 6 anthocyanin aglycones: delphinidin, cyanidin, petunidin, pelargonidin, apigeninidin, and peonidin, were identi?ed from these extracts. High performance liquid chromatography coupled with electrospray ionization triple quadrupole mass spectrometry(HPLC-ESI-QQQ-MS) was used to accurately quantify the content of anthocyanins. The total content of bound anthocyanins in the acid extract was the highest. In addition, in the soluble anthocyanin extract, anthocyanins were present mainly in the form of anthocyanin glycoside with lesser amounts of aglycons, whereas in the bound anthocyanin extract,anthocyanin aglycons were the main components but glycosides were rare.
Ultrasonic-assisted organic solvent extraction was used to obtain soluble anthocyanins from black soybean seed coat, and bound anthocyanins from the extraction residue were obtained by treatment with acid, alkaline, sequential acid/alkali hydrolysis or the reverse sequence. By ultra-high performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry(UPLC-ESI-QTOF-MS), a total of 17 anthocyanin components,including 11 anthocyanin glycosides: delphinidin-3-O-glucoside, delphinidin-3-O-galactoside, petunidin-3-O-glucoside,petunidin-3-O-galactoside, pelargonidin-3-O-rutinoside, cyanidin-3-O-glucoside, cyanidin-3-O-galactoside, pelargonidin-3-O-hexoside, peonidin-3-O-hexoside, pelargonidin-3-O-(6"-malonyl glucoside) and apigeninidin-3-O-(6"-malonyl glucoside);and 6 anthocyanin aglycones: delphinidin, cyanidin, petunidin, pelargonidin, apigeninidin, and peonidin, were identi?ed from these extracts. High performance liquid chromatography coupled with electrospray ionization triple quadrupole mass spectrometry(HPLC-ESI-QQQ-MS) was used to accurately quantify the content of anthocyanins. The total content of bound anthocyanins in the acid extract was the highest. In addition, in the soluble anthocyanin extract, anthocyanins were present mainly in the form of anthocyanin glycoside with lesser amounts of aglycons, whereas in the bound anthocyanin extract,anthocyanin aglycons were the main components but glycosides were rare.
引文
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[7]秦琦,张英蕾,张守文.黑豆的营养保健价值及研究进展[J].中国食品添加剂,2015(7):145-150. DOI:10.3969/j.issn.1006-2513.2015.07.016.
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[9] GUAJARDOFLORES D,SERNASALDI'VAR S O,GUTIERREZURIBE J A. Evaluation of the antioxidant and antiproliferative activities of extracted saponins and flavonols from germinated black beans(Phaseolus vulgaris L.)[J]. Food Chemistry,2013, 141(2):1497-1503. DOI:10.1016/j.foodchem:2013.04.010.
[10] ZHANG C, MONK J M, LU J T, et al. Cooked navy and black bean diets improve biomarkers of colon health and reduce inflammation during colitis[J]. British Journal of Nutrition, 2014, 111(9):1549-1563. DOI:10.1017/S0007114513004352.
[11] LIU R H. Whole grain phytochemicals and health[J]. Journal of Cereal Science, 2007, 46(3):207-219. DOI:10.1016/j.jcs.2007.06.010.
[12] ANDREASEN M F, KROON P A, WILLIAMSON G, et al. Esterase activity able to hydrolyze dietary antioxidant hydroxycinnamates is distributed along the intestine of mammals[J]. Journal of Agricultural&Food Chemistry, 2001, 49(11):5679-5684. DOI:10.1021/jf010668c.
[13] ACOSTA-ESTRADA B A, GUTIERREZ-URIBE J A,SERNA-SALDIVAR S O. Bound phenolics in foods, a review[J]. Food Chemistry, 2014, 152(6):46-55. DOI:10.1016/j.foodchem.2013.11,093.
[14] SHAHIDI F, YEO J D. Insoluble-bound phenolics in food[J].Molecules, 2016, 21(9):1216. DOI:10.3390/molecules21091216.
[15] TANG Y, ZHANG B, LI X, et al. Bound phenolics of quinoa seeds released by acid, alkaline and enzymatic treatments and their antioxidant and a-glucosidase and pancreatic lipase inhibitory effects[J]. Journal of Agricultural&Food Chemistry, 2016, 64(8):1712-1719. DOI:10.1021/acs.jafc.5b05761.
[16] CUEVAS M E,HILLEBRAND S,ANTEZANA A,et al. Soluble and bound phenolic compounds in different Bolivian purple corn(Zea mays L.)cultivars[J]. Journal of Agricultural and Food Chemistry,2011, 59(13):7068-7074. DOI:10.1021/jf201061x.
[17] SU D, ZHANG R, HOU F, et al. Comparison of the free and bound phenolic profiles and cellular antioxidant activities of litchi pulp extracts from different solvents[J]. BMC Complementary and Alternative Medicine, 2014, 14(1):9. DOI:10.1186/1472-6882-14-9.
[18] DE CAMARGO A C,REGITANOD'ARCE M A,BIASOTO A C,et al. Enzyme-assisted extraction of phenolics from winemaking byproducts:antioxidant potential and inhibition of alpha-glucosidase and lipase activities[J]. Food Chemistry, 2016, 212:395-402.DOI:10.1016/j.foodchem.2016.05.047.
[19] PENG H.LI W,LI H,et al. Extractable and non-extractable bound phenolic compositions and their antioxidant properties in seed coat and cotyledon of black soybean(Glycine max(L.)Merr.)[J]. Journal of Functional Foods, 2017, 32:296-312. DOI:10.1016/j.jff.2017.03.003.
[20] KWANGOH K,JUNGEUN Y,HEESEON K. Identification of anthocyanins in black soybean(Glycine max(L.)Merr.)varieties[J].Journal of Food Science&Technology, 2014, 51(2):377-381.DOI:10.1007/s 13197-011-0493-y.
[21] JINHWAN L, KANG N S, SANGOUK S, et al. Characterisation of anthocyanins in the black soybean(Glycine max L.)by HPLCDAD-ESI/MS analysis[J]. Food Chemistry, 2009, 112(1):226-231.DOI:10.1016/j.foodchem.2008.05.056.
[22]洪迪清,王世清.黑豆的研究进展[J].中国药业,2008, 17(10):80.DOI:10.3969/j.issn.1006-4931.2008.10.066.
[23]孙海红,钱叶苗,宋相丽,等.固相萃取技术的应用与研究新进展[J].现代化工, 2011, 31(增刊2):21-24. DOI:10.16606/j.cnki.issn0253-4320.2011.s2.003.
[24]LIHY,DENGZ,ZHUH,etal.Highlypigmentedvegetables:anthocyanincompositionsandtheirroleinantioxidantactivities[J].FoodResearchInternational,2012,46(1):250-259.DOI:10.1016/j.foodres.2011.12.014.
[25]SUNY,LIH,HUJ,etal.Qualitativeandquantitativeanalysisof phenolicsintetrastigmahemsleyanumandtheirantioxidantand antiproliferativeactivities[J].JournalofAgriculturalandFood Chemistry, 2013, 61(44):10507-10515. DOI:10.1021/jf4037547.
[26]ALBERTSP,STANDERMA,DEVA.Advancedultrahigh pressure liquid chromatography-tandem mass spectrometric methods for the screening of red wine anthocyanins and derived pigments[J].Journal of Chromatography A, 2012, 1235(8):92-102. DOI:10.1016/j.chroma.2012.02.058.
[27]李媛,谢岩黎.制备型色谱柱对黑豆皮花青素分离纯度的影响[J].河南工业大学学报(自然科学版), 2017(6):21-25. DOI:10.16433/j.cnki.issn1673-2383.2017.06.004.
[28]WANG Y K, ZHANG X, CHEN G L, et al. Antioxidant property and theirfree,solubleconjugateandinsoluble-boundphenoliccontents in selected beans[J]. Journal of Functional Foods, 2016, 24:359-372.DOI:10.1016/j.jff.2016.04.026.
[29]SCORRANOS,LAZZOIMR,MERGOLAL,etal.Anthocyanins profile by Q-TOF LC/MS in Myrtus communis, berries from Salento area[J]. Journal of Food Analytical Methods, 2017, 10(7):2404-2411.DOI:10.1007/s12161-017-0813-6.
[30]BARNES J S, SCHUG K A. Structural characterization of cyanidin-3,5-diglucoside and pelargonidin-3,5-diglucoside anthocyanins:multidimensionalfragmentationpathwaysusinghighperformanceliquid chromatography-electrospray ionization-ion trap-time of flight mass spectrometry[J].InternationalJournalofMassSpectrometry,2015,308(1):71-80. DOI:10.1016/j.ijms.2011.07.026.
[31]SHIH C H, SIU S O, NG R, et al. Quantitative analysis of anticancer3-deoxyanthocyanidinsininfectedsorghumseedlings[J].Journalof Agricultural&Food Chemistry, 2007, 55(2):254-259. DOI:10.1021/jf062516t.
[32]DINCHEVA I, BADJAKOV I, KONDAKOVA V, et al. Identification of the phenolic components in Bulgarian raspberry cultivars by LCESI-MSn[J]. International Journal of Agricultural Science&Research,2013, 3:127-138.
[33]SDPT,SANTOSBUELGAC,RIVASGONZALOJC.LC-MS analysisofanthocyaninsfrompurplecorncob[J].Journalofthe Science of Food&Agriculture, 2002, 82(9):1003-1006. DOI:10.1002/jsfa.1143.
[2] NISHIDA C, UAUY R, KUMANYIKA S, et al. The joint WHO/FAO expert consultation on diet, nutrition and the prevention of chronic diseases:process, product and policy implications[J]. Public Health Nutrition, 2004, 7(1a):245-250. DOI:10.1079/PHN2003592.
[3] HUNG H C, JOSHIPURA K J, JIANG R, et al. Fruit and vegetable intake and risk of major chronic disease[J]. Journal of the National Cancer Institute, 2004, 96(21):1577-1584. DOI:10.1093/jnci/djh296.
[4] SINGH R B, DUBNOV G, NIAZ M A, et al. Effect of an indomediterranean diet on progression of coronary artery disease in high risk patients(indo-mediterranean diet heart study):a randomised single-blind trial[J]. Lancet, 2003, 12(2):1455-1461. DOI:10.1016/S0140-6736(02)11472-3.
[5] WILLETT W C, SACKS F, TRICHOPOULOU A, et al.Mediterranean diet pyramid:a cultural model for healthy eating[J].American Journal of Clinical Nutrition, 1995, 61(Suppl 6):1402S.DOI:10.1093/ajcn/61.6.1402S.
[6] LLOYD B J,SIEBENMORGEN T J,BEERS K W. Effects of commercial processing on antioxidants found in rice bran[J].Cereal Chemistry, 2000, 77(5):551-555. DOI:10.1094/CCHEM.2000.77.5.551.
[7]秦琦,张英蕾,张守文.黑豆的营养保健价值及研究进展[J].中国食品添加剂,2015(7):145-150. DOI:10.3969/j.issn.1006-2513.2015.07.016.
[8] FURUTA S, TAKAHASHI M, TAKAHATA Y, et al. Radicalscavenging activities of soybean cultivars with black seed coats[J].Food Science&Technology International Tokyo, 2007, 9(1):73-75.DOI:10.3136/fstr.9.73.
[9] GUAJARDOFLORES D,SERNASALDI'VAR S O,GUTIERREZURIBE J A. Evaluation of the antioxidant and antiproliferative activities of extracted saponins and flavonols from germinated black beans(Phaseolus vulgaris L.)[J]. Food Chemistry,2013, 141(2):1497-1503. DOI:10.1016/j.foodchem:2013.04.010.
[10] ZHANG C, MONK J M, LU J T, et al. Cooked navy and black bean diets improve biomarkers of colon health and reduce inflammation during colitis[J]. British Journal of Nutrition, 2014, 111(9):1549-1563. DOI:10.1017/S0007114513004352.
[11] LIU R H. Whole grain phytochemicals and health[J]. Journal of Cereal Science, 2007, 46(3):207-219. DOI:10.1016/j.jcs.2007.06.010.
[12] ANDREASEN M F, KROON P A, WILLIAMSON G, et al. Esterase activity able to hydrolyze dietary antioxidant hydroxycinnamates is distributed along the intestine of mammals[J]. Journal of Agricultural&Food Chemistry, 2001, 49(11):5679-5684. DOI:10.1021/jf010668c.
[13] ACOSTA-ESTRADA B A, GUTIERREZ-URIBE J A,SERNA-SALDIVAR S O. Bound phenolics in foods, a review[J]. Food Chemistry, 2014, 152(6):46-55. DOI:10.1016/j.foodchem.2013.11,093.
[14] SHAHIDI F, YEO J D. Insoluble-bound phenolics in food[J].Molecules, 2016, 21(9):1216. DOI:10.3390/molecules21091216.
[15] TANG Y, ZHANG B, LI X, et al. Bound phenolics of quinoa seeds released by acid, alkaline and enzymatic treatments and their antioxidant and a-glucosidase and pancreatic lipase inhibitory effects[J]. Journal of Agricultural&Food Chemistry, 2016, 64(8):1712-1719. DOI:10.1021/acs.jafc.5b05761.
[16] CUEVAS M E,HILLEBRAND S,ANTEZANA A,et al. Soluble and bound phenolic compounds in different Bolivian purple corn(Zea mays L.)cultivars[J]. Journal of Agricultural and Food Chemistry,2011, 59(13):7068-7074. DOI:10.1021/jf201061x.
[17] SU D, ZHANG R, HOU F, et al. Comparison of the free and bound phenolic profiles and cellular antioxidant activities of litchi pulp extracts from different solvents[J]. BMC Complementary and Alternative Medicine, 2014, 14(1):9. DOI:10.1186/1472-6882-14-9.
[18] DE CAMARGO A C,REGITANOD'ARCE M A,BIASOTO A C,et al. Enzyme-assisted extraction of phenolics from winemaking byproducts:antioxidant potential and inhibition of alpha-glucosidase and lipase activities[J]. Food Chemistry, 2016, 212:395-402.DOI:10.1016/j.foodchem.2016.05.047.
[19] PENG H.LI W,LI H,et al. Extractable and non-extractable bound phenolic compositions and their antioxidant properties in seed coat and cotyledon of black soybean(Glycine max(L.)Merr.)[J]. Journal of Functional Foods, 2017, 32:296-312. DOI:10.1016/j.jff.2017.03.003.
[20] KWANGOH K,JUNGEUN Y,HEESEON K. Identification of anthocyanins in black soybean(Glycine max(L.)Merr.)varieties[J].Journal of Food Science&Technology, 2014, 51(2):377-381.DOI:10.1007/s 13197-011-0493-y.
[21] JINHWAN L, KANG N S, SANGOUK S, et al. Characterisation of anthocyanins in the black soybean(Glycine max L.)by HPLCDAD-ESI/MS analysis[J]. Food Chemistry, 2009, 112(1):226-231.DOI:10.1016/j.foodchem.2008.05.056.
[22]洪迪清,王世清.黑豆的研究进展[J].中国药业,2008, 17(10):80.DOI:10.3969/j.issn.1006-4931.2008.10.066.
[23]孙海红,钱叶苗,宋相丽,等.固相萃取技术的应用与研究新进展[J].现代化工, 2011, 31(增刊2):21-24. DOI:10.16606/j.cnki.issn0253-4320.2011.s2.003.
[24]LIHY,DENGZ,ZHUH,etal.Highlypigmentedvegetables:anthocyanincompositionsandtheirroleinantioxidantactivities[J].FoodResearchInternational,2012,46(1):250-259.DOI:10.1016/j.foodres.2011.12.014.
[25]SUNY,LIH,HUJ,etal.Qualitativeandquantitativeanalysisof phenolicsintetrastigmahemsleyanumandtheirantioxidantand antiproliferativeactivities[J].JournalofAgriculturalandFood Chemistry, 2013, 61(44):10507-10515. DOI:10.1021/jf4037547.
[26]ALBERTSP,STANDERMA,DEVA.Advancedultrahigh pressure liquid chromatography-tandem mass spectrometric methods for the screening of red wine anthocyanins and derived pigments[J].Journal of Chromatography A, 2012, 1235(8):92-102. DOI:10.1016/j.chroma.2012.02.058.
[27]李媛,谢岩黎.制备型色谱柱对黑豆皮花青素分离纯度的影响[J].河南工业大学学报(自然科学版), 2017(6):21-25. DOI:10.16433/j.cnki.issn1673-2383.2017.06.004.
[28]WANG Y K, ZHANG X, CHEN G L, et al. Antioxidant property and theirfree,solubleconjugateandinsoluble-boundphenoliccontents in selected beans[J]. Journal of Functional Foods, 2016, 24:359-372.DOI:10.1016/j.jff.2016.04.026.
[29]SCORRANOS,LAZZOIMR,MERGOLAL,etal.Anthocyanins profile by Q-TOF LC/MS in Myrtus communis, berries from Salento area[J]. Journal of Food Analytical Methods, 2017, 10(7):2404-2411.DOI:10.1007/s12161-017-0813-6.
[30]BARNES J S, SCHUG K A. Structural characterization of cyanidin-3,5-diglucoside and pelargonidin-3,5-diglucoside anthocyanins:multidimensionalfragmentationpathwaysusinghighperformanceliquid chromatography-electrospray ionization-ion trap-time of flight mass spectrometry[J].InternationalJournalofMassSpectrometry,2015,308(1):71-80. DOI:10.1016/j.ijms.2011.07.026.
[31]SHIH C H, SIU S O, NG R, et al. Quantitative analysis of anticancer3-deoxyanthocyanidinsininfectedsorghumseedlings[J].Journalof Agricultural&Food Chemistry, 2007, 55(2):254-259. DOI:10.1021/jf062516t.
[32]DINCHEVA I, BADJAKOV I, KONDAKOVA V, et al. Identification of the phenolic components in Bulgarian raspberry cultivars by LCESI-MSn[J]. International Journal of Agricultural Science&Research,2013, 3:127-138.
[33]SDPT,SANTOSBUELGAC,RIVASGONZALOJC.LC-MS analysisofanthocyaninsfrompurplecorncob[J].Journalofthe Science of Food&Agriculture, 2002, 82(9):1003-1006. DOI:10.1002/jsfa.1143.
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