刺梨及其近缘种质叶片主要活性物质含量及抗氧化性分析
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摘要
为明确刺梨及其2个近缘种质无籽刺梨和无刺刺梨叶片中活性物质含量变化及其对抗氧化能力的贡献,本研究以刺梨、无籽刺梨和无刺刺梨为试验材料,测定其不同叶龄叶片(幼叶、成熟叶和老叶)中维生素C(Vc)、总三萜、总酚、总黄酮含量及超氧化物歧化酶(SOD)活性变化,并用铁离子还原(FRAP)、2,2'-联氨-二(3-乙基-苯并噻唑-6-磺酸)二铵盐(ABTS)和1,1-二苯基-2-苦肼基(DPPH)3种体系分析其体外抗氧化能力,同时对5种抗氧化活性物质进行主成分分析。结果表明,3个材料的不同叶龄叶片中,总酚、总黄酮、Vc含量和SOD活性变化以及3种抗氧化活性均依次表现为成熟叶>幼叶>老叶,而总三萜含量均在老叶中为最高; 3个试验材料成熟叶中,无刺刺梨的总酚含量及其对ABTS自由基的清除能力、刺梨的Vc含量及其对DPPH自由基的清除能力,以及无籽刺梨的SOD活性及其对FRAP抗氧化能力均高于其他2个材料,刺梨老叶中的总三萜含量显著高于其他2个材料。相关性分析表明,总酚、总黄酮、Vc与3种抗氧化方法测定结果呈显著正相关;主成分分析表明,成熟叶中具有抗氧化能力的5种活性物质对抗氧化能力贡献依次表现为总酚、Vc>总三萜、SOD>总黄酮,且3种主成分的累计贡献率达90%以上。本研究结果为了解和开发利用上述3个蔷薇属种质叶片功能成分提供了理论依据。
This study aimed to understand the changes of the active substance contents and their contributions to the antioxidant properties in leaves of Rosa roxburghii Tratt. and its two related germplasm R. sterilis S. D. Shi and R.roxburghii f. Eseiosa Ku. The contents of vitamin C( Vc) total triterpenoids,total phenols,total flavonoids,and superoxide dismutase( SOD) were determined in leaves of three developmental stages( young leaves,mature leaves and old leaves) of the three germplasms. The iron ion reduction( FRAP),2,2 '-amino-bis( 3-ethyl-benzothiazoline-6-sulfonate) ammonium salt( ABTS) and1,1-,two phenyl,-2-hydrazine group( DPPH) were used to evaluate the antioxidant activity in vitro and the principal component analysis of five kinds of antioxidant active substances were also made.The results showed that the changes of total phenolics,total flavonoids,Vc contents and SOD activities and the three antioxidant activities presented mature leaves> young leaves> old leave in 3 different leaf ages of 3 materials,while the total triterpenoids have the highest content in the old leaves. In mature leaves of 3 materials,the total phenolic contents and ABTS·+ free radical scavenging capacity of R. roxburghii f. Eseiosa,the Vc contents and DPPH·+ free radical scavenging capacity of R. roxburghii and the SOD activity and FRAP free radical capacity of R. sterilis were higher than the other two materials,whereas the total triterpene contents in old leaves of R. roxburghii were significantly higher than the other two materials. Correlation analysis showed that total phenol,total flavonoids and Vc were significantly positively correlated with the value of three antioxidant methods. The principal component analysisindicated that the five active compounds with antioxidant capacity in mature leaves contributed to the antioxidant properties showed total phenols and Vc > total triterpenes and SOD > total flavones,and the cumulative contribution rate of the three components were more than 90%. The above results provide important theoretical basis for understanding and developing the functional components of three Rosa germplasms leaves.
This study aimed to understand the changes of the active substance contents and their contributions to the antioxidant properties in leaves of Rosa roxburghii Tratt. and its two related germplasm R. sterilis S. D. Shi and R.roxburghii f. Eseiosa Ku. The contents of vitamin C( Vc) total triterpenoids,total phenols,total flavonoids,and superoxide dismutase( SOD) were determined in leaves of three developmental stages( young leaves,mature leaves and old leaves) of the three germplasms. The iron ion reduction( FRAP),2,2 '-amino-bis( 3-ethyl-benzothiazoline-6-sulfonate) ammonium salt( ABTS) and1,1-,two phenyl,-2-hydrazine group( DPPH) were used to evaluate the antioxidant activity in vitro and the principal component analysis of five kinds of antioxidant active substances were also made.The results showed that the changes of total phenolics,total flavonoids,Vc contents and SOD activities and the three antioxidant activities presented mature leaves> young leaves> old leave in 3 different leaf ages of 3 materials,while the total triterpenoids have the highest content in the old leaves. In mature leaves of 3 materials,the total phenolic contents and ABTS·+ free radical scavenging capacity of R. roxburghii f. Eseiosa,the Vc contents and DPPH·+ free radical scavenging capacity of R. roxburghii and the SOD activity and FRAP free radical capacity of R. sterilis were higher than the other two materials,whereas the total triterpene contents in old leaves of R. roxburghii were significantly higher than the other two materials. Correlation analysis showed that total phenol,total flavonoids and Vc were significantly positively correlated with the value of three antioxidant methods. The principal component analysisindicated that the five active compounds with antioxidant capacity in mature leaves contributed to the antioxidant properties showed total phenols and Vc > total triterpenes and SOD > total flavones,and the cumulative contribution rate of the three components were more than 90%. The above results provide important theoretical basis for understanding and developing the functional components of three Rosa germplasms leaves.
引文
[1]李福明,汪洋,韦敏.刺梨叶醇提物体外抗氧化活性和α-葡萄糖苷酶抑制活性研究[J].中国现代应用药学,2015,32(6):685-688
[2]孙兆峰,张霞,夏作理.刺梨叶对2型糖尿病大鼠脂代谢的影响[J].社区医学杂志,2015,13(10):53-55
[3]郭建军,陆锦锐,罗俊,孙晓惠,周艺.刺梨茶对糖尿病小鼠血糖的影响[J].中国民族民间医药,2017,26(14):50-53
[4]周艺,郭建军,郁建平.刺梨茶总黄酮的提取工艺的建立及其抗氧化活性[J].氨基酸和生物资源,2017,39(2):135-140
[5]向显衡,何照凡,牛爱珍.刺梨叶果中维生素C含量的变化[J].中国果树,1984(1):50-54
[6] Li L L,Lu M,An H. Expression profiles of the genes involved in lascorbic acid biosynthesis and recycling in Rosa roxburghii,leaves of various ages[J]. Acta Physiologiae Plantarum,2017,39(2):44-51
[7]樊卫国,刘进平,向灵.刺梨花粉和叶的营养成分分析[J].营养学报,1998,20(1):107-110
[8]安华明,樊卫国,刘庆林,陈力耕.刺梨果实和叶片发育过程中抗坏血酸和抗氧化酶的协同变化[J].园艺学报,2007,34(5):1293-1296
[9]蒋辉,龙远德. 4种蔷薇属植物叶片黄酮含量的季节性变化[J].天然产物研究与开发,2000,12(3):58-63
[10] Cruz-Rus E,Amaya I,Sánchez-Sevilla J F,Botella M A,Valpuesta V. Regulation of L-ascorbic acid content in strawberry fruits[J].Journal of Experimental Botany,2011,62(12):4191-4201
[11] Gomez M L P,Lajolo F M. Ascorbic acid metabolism in fruits:activity of enzymes involved in synthesis and degradation during ripening in mango and guava[J]. Journal of the Science of Food&Agriculture,2010,88(5):756-762
[12] Imai T,Yusuke B,Terakami S. l-Ascorbate biosynthesis in peach:cloning of six l-galactose pathway-related genes and their expression during peach fruit development[J]. Physiologia Plantarum,2009,136(2):139-149
[13] Xu Y,Zhu X,Chen Y,Gong Y,Liu L. Expression profiling of genes involved in ascorbate biosynthesis and recycling during fleshy root development in radish[J]. Plant Physiology&Biochemistry,2013,70(1):269-277
[14] Ioannidi E,Kalamaki M S,PaterakiⅠ,MellidouⅠ,Kanellis A K. Expression profiling of ascorbic acid-related genes during tomato fruit development and ripening and in response to stress conditions[J]. Journal of Experimental Botany,2009,60(2):663-678
[15]戴支凯,余丽梅,杨小生,石京山.刺梨三萜化合物CL1体外抗人子宫内膜腺癌作用[J].时珍国医国药,2011,22(7):1656-1658
[16]黄姣娥,江晋渝,罗勇,戴支凯.刺梨三萜对人肝癌SMMC-7721细胞增殖的影响[J].食品科学,2013,34(13):275-279
[17] Zeng F F,Ge Z W,Limwachiranon J,Li L,Feng S M,Wang Y S,Luo Z S. Antioxidant and tyrosinase inhibitory activity of Rosa roxburghii fruit and identification of main bioactive phytochemicals by UPLC-Triple-TOF/MS[J]. International Journal of Food Science&Technology,2017,52(4):897-905
[18] He J Y,Zhang Y H,Ma N,Zhang X L,Liu M H,Fu W M.Comparative analysis of multiple ingredients in Rosa roxburghii,and R. sterilis,fruits and their antioxidant activities[J]. Journal of Functional Foods,2016,27:29-41
[19] Xiang Q S,Xiao H F,Gao S B,Wen M T,Jia G L,Ren X M. In vitro antioxidant properties of rosa roxburghii aqueous extracts[J].Australian Journal of Crop Science,2012,6(5):854-860
[20]谢国芳,徐小燕,王瑞,刘志刚.金刺梨果实和叶中酚类、Vc含量及其抗氧化能力分析[J].植物科学学报,2017,35(1):122-127
[21]王乐乐,安华明. HPLC测定刺梨果实中维生素C含量方法的优化[J].现代食品科技,2013,29(2):397-400
[22]周巧霞,张经硕,顾明,顾振纶.测定山楂及提取物总三萜酸的含量[J].中国野生植物资源,2004,23(5):43-44
[23]张雁冰,王克让,刘宏民.马桑叶中总三萜酸的含量测定[J].时珍国医国药,2006,17(4):529-530
[24]蔡文国,吴卫,邵金凤,陈鹊,王元彪. Folin-Ciocalteu法测定鱼腥草多酚的含量[J].食品科学,2010,31(14):201-204
[25]罗锋,汪河滨,杨玲,周忠波.超声-微波协同萃取法提取甘草黄酮的研究[J].食品研究与开发,2006,27(8):127-128
[26]吴媛琳,王世军,张社奇,李丙智.修剪对苹果枝(梢)皮层总黄酮含量的影响[J].西北林学院学报,2013,28(6):103-107
[27]王学奎.植物生理生化实验原理和技术[M].北京:高等教育出版社,2000
[28]杜国荣.猕猴桃、柿和苹果果实的抗氧化能力及其抗氧化活性成分的分析[D].杨凌:西北农林科技大学,2009
[29] BenzieⅠF F,Strain J J. The Ferric reducing ability of plasma(FRAP)as a measure of“Antioxidant power”:The FRAP assay[J]. Analytical Biochemistry,1996,239(1):70-76
[30] Re R, Pellegrini N, Proteggente A, Pannala A, Yang M.Antioxidant activity applying an improved ABTS radical cation decolorization assay[J]. Free Radical Biology and Medicine,1999,26(9/10):1231-1237
[31] Takebayashi J, Chen J, Tai A. A method for evaluation of antioxidant activity based on inhibition of free radical-induced erythrocyte hemolysis[J]. Methods in Molecular Biology,2010,594:287-296
[32] Mayne S T. Antioxidant nutrients and chronic disease:Use of biomarkers of exposure and oxidative stress status in epidemiologic research[J]. Journal of Nutrition,2003,133:933-940
[33] Albarracin S L,Stab B,Casas Z,Sutachan J J,SamudioⅠ,Gonzalez J,Gonzalo L,Capani F,Morales L,Barreto G E. Effects of natural antioxidants in neurodegenerative disease[J]. Nutritional Neuroscience,2012,15:1-9
[34] Kawaguchi K,Matsumoto T,Kumazawa Y. Effects of antioxidant polyphenols on TNF-alpha-related diseases[J]. Current Topics in Medicinal Chemistry,2011,11:1767-1779
[35] Singh B N, Shankar S, Srivastava R K. Green tea catechin,epigallocatechin-3-gallate(EGCG):Mechanisms,perspectives and clinical applications[J]. Biochem Pharmacol,2011,82:1807-1821
[36]张秀梅,骆党委,朱祝英,刘玉革,杜丽清. 3种热带果树叶子甲醇粗提物的抗氧化活性比较[J].核农学报,2015,29(1):95-100
[37]林立,林乐静,毛阳正,祝志勇,付涛.秀丽槭叶总黄酮的提取及其抗氧化能力研究[J].核农学报,2016,30(12):2373-2381
[38] Fan Z L,Wang Z Y,Liu J R. Cold-field fruit extracts exert different antioxidant and antiproliferative activities in vitro[J]. Food Chemistry,2011,129(2):402-407
[39] Giri L,Dhyani P,Rawat S,Bhatt I D,Nandi S K,Rawal R S,Pande V. In vitro production of phenolic compounds and antioxidant activity in callus suspension cultures of Habenaria edgeworthii:A rare Himalayan medicinal orchid[J]. Industrial Crops&Products,2012,37(1):1-6
[40]王有琼,马李一,张重权,段琼芬.印楝叶多酚提取及体外抗氧化活性[J].精细化工,2015,32(1):64-68
[41] Shanmugam M K,An H N,Kumar A P,Tan B K H,Gautam S.Targeted inhibition of tumor proliferation,survival,and metastasis by pentacyclic triterpenoids:Potential role in prevention and therapy of cancer[J]. Cancer Letters,2012,320(2):158-170
[42] Fujioka T,Kashiwada Y,Kilkuskie R E,Cosentino L M,Ballas L M,Jiang J B,Janzen W P,ChenⅠS,Lee K H. Anti-AIDS agents,11. Betulinic acid and platanic acid as anti-HIV principles from Syzigium claviflorum,and the anti-HIV activity of structurally related triterpenoids[J]. Journal of Natural Products,1994,57(2):243-247
[43]陈明,黄仁华,刘仁道,王丹,丁振柱.费约果叶片提取物抗氧化活性研究[J].果树学报,2012,29(4):593-597
[44]秦晶晶,李齐激,薛琰,马琳,杨小生.刺梨总三萜提取方法及其α-葡萄糖苷酶抑制活性研究[J].食品工业科技,2014,35(10):186-189
[2]孙兆峰,张霞,夏作理.刺梨叶对2型糖尿病大鼠脂代谢的影响[J].社区医学杂志,2015,13(10):53-55
[3]郭建军,陆锦锐,罗俊,孙晓惠,周艺.刺梨茶对糖尿病小鼠血糖的影响[J].中国民族民间医药,2017,26(14):50-53
[4]周艺,郭建军,郁建平.刺梨茶总黄酮的提取工艺的建立及其抗氧化活性[J].氨基酸和生物资源,2017,39(2):135-140
[5]向显衡,何照凡,牛爱珍.刺梨叶果中维生素C含量的变化[J].中国果树,1984(1):50-54
[6] Li L L,Lu M,An H. Expression profiles of the genes involved in lascorbic acid biosynthesis and recycling in Rosa roxburghii,leaves of various ages[J]. Acta Physiologiae Plantarum,2017,39(2):44-51
[7]樊卫国,刘进平,向灵.刺梨花粉和叶的营养成分分析[J].营养学报,1998,20(1):107-110
[8]安华明,樊卫国,刘庆林,陈力耕.刺梨果实和叶片发育过程中抗坏血酸和抗氧化酶的协同变化[J].园艺学报,2007,34(5):1293-1296
[9]蒋辉,龙远德. 4种蔷薇属植物叶片黄酮含量的季节性变化[J].天然产物研究与开发,2000,12(3):58-63
[10] Cruz-Rus E,Amaya I,Sánchez-Sevilla J F,Botella M A,Valpuesta V. Regulation of L-ascorbic acid content in strawberry fruits[J].Journal of Experimental Botany,2011,62(12):4191-4201
[11] Gomez M L P,Lajolo F M. Ascorbic acid metabolism in fruits:activity of enzymes involved in synthesis and degradation during ripening in mango and guava[J]. Journal of the Science of Food&Agriculture,2010,88(5):756-762
[12] Imai T,Yusuke B,Terakami S. l-Ascorbate biosynthesis in peach:cloning of six l-galactose pathway-related genes and their expression during peach fruit development[J]. Physiologia Plantarum,2009,136(2):139-149
[13] Xu Y,Zhu X,Chen Y,Gong Y,Liu L. Expression profiling of genes involved in ascorbate biosynthesis and recycling during fleshy root development in radish[J]. Plant Physiology&Biochemistry,2013,70(1):269-277
[14] Ioannidi E,Kalamaki M S,PaterakiⅠ,MellidouⅠ,Kanellis A K. Expression profiling of ascorbic acid-related genes during tomato fruit development and ripening and in response to stress conditions[J]. Journal of Experimental Botany,2009,60(2):663-678
[15]戴支凯,余丽梅,杨小生,石京山.刺梨三萜化合物CL1体外抗人子宫内膜腺癌作用[J].时珍国医国药,2011,22(7):1656-1658
[16]黄姣娥,江晋渝,罗勇,戴支凯.刺梨三萜对人肝癌SMMC-7721细胞增殖的影响[J].食品科学,2013,34(13):275-279
[17] Zeng F F,Ge Z W,Limwachiranon J,Li L,Feng S M,Wang Y S,Luo Z S. Antioxidant and tyrosinase inhibitory activity of Rosa roxburghii fruit and identification of main bioactive phytochemicals by UPLC-Triple-TOF/MS[J]. International Journal of Food Science&Technology,2017,52(4):897-905
[18] He J Y,Zhang Y H,Ma N,Zhang X L,Liu M H,Fu W M.Comparative analysis of multiple ingredients in Rosa roxburghii,and R. sterilis,fruits and their antioxidant activities[J]. Journal of Functional Foods,2016,27:29-41
[19] Xiang Q S,Xiao H F,Gao S B,Wen M T,Jia G L,Ren X M. In vitro antioxidant properties of rosa roxburghii aqueous extracts[J].Australian Journal of Crop Science,2012,6(5):854-860
[20]谢国芳,徐小燕,王瑞,刘志刚.金刺梨果实和叶中酚类、Vc含量及其抗氧化能力分析[J].植物科学学报,2017,35(1):122-127
[21]王乐乐,安华明. HPLC测定刺梨果实中维生素C含量方法的优化[J].现代食品科技,2013,29(2):397-400
[22]周巧霞,张经硕,顾明,顾振纶.测定山楂及提取物总三萜酸的含量[J].中国野生植物资源,2004,23(5):43-44
[23]张雁冰,王克让,刘宏民.马桑叶中总三萜酸的含量测定[J].时珍国医国药,2006,17(4):529-530
[24]蔡文国,吴卫,邵金凤,陈鹊,王元彪. Folin-Ciocalteu法测定鱼腥草多酚的含量[J].食品科学,2010,31(14):201-204
[25]罗锋,汪河滨,杨玲,周忠波.超声-微波协同萃取法提取甘草黄酮的研究[J].食品研究与开发,2006,27(8):127-128
[26]吴媛琳,王世军,张社奇,李丙智.修剪对苹果枝(梢)皮层总黄酮含量的影响[J].西北林学院学报,2013,28(6):103-107
[27]王学奎.植物生理生化实验原理和技术[M].北京:高等教育出版社,2000
[28]杜国荣.猕猴桃、柿和苹果果实的抗氧化能力及其抗氧化活性成分的分析[D].杨凌:西北农林科技大学,2009
[29] BenzieⅠF F,Strain J J. The Ferric reducing ability of plasma(FRAP)as a measure of“Antioxidant power”:The FRAP assay[J]. Analytical Biochemistry,1996,239(1):70-76
[30] Re R, Pellegrini N, Proteggente A, Pannala A, Yang M.Antioxidant activity applying an improved ABTS radical cation decolorization assay[J]. Free Radical Biology and Medicine,1999,26(9/10):1231-1237
[31] Takebayashi J, Chen J, Tai A. A method for evaluation of antioxidant activity based on inhibition of free radical-induced erythrocyte hemolysis[J]. Methods in Molecular Biology,2010,594:287-296
[32] Mayne S T. Antioxidant nutrients and chronic disease:Use of biomarkers of exposure and oxidative stress status in epidemiologic research[J]. Journal of Nutrition,2003,133:933-940
[33] Albarracin S L,Stab B,Casas Z,Sutachan J J,SamudioⅠ,Gonzalez J,Gonzalo L,Capani F,Morales L,Barreto G E. Effects of natural antioxidants in neurodegenerative disease[J]. Nutritional Neuroscience,2012,15:1-9
[34] Kawaguchi K,Matsumoto T,Kumazawa Y. Effects of antioxidant polyphenols on TNF-alpha-related diseases[J]. Current Topics in Medicinal Chemistry,2011,11:1767-1779
[35] Singh B N, Shankar S, Srivastava R K. Green tea catechin,epigallocatechin-3-gallate(EGCG):Mechanisms,perspectives and clinical applications[J]. Biochem Pharmacol,2011,82:1807-1821
[36]张秀梅,骆党委,朱祝英,刘玉革,杜丽清. 3种热带果树叶子甲醇粗提物的抗氧化活性比较[J].核农学报,2015,29(1):95-100
[37]林立,林乐静,毛阳正,祝志勇,付涛.秀丽槭叶总黄酮的提取及其抗氧化能力研究[J].核农学报,2016,30(12):2373-2381
[38] Fan Z L,Wang Z Y,Liu J R. Cold-field fruit extracts exert different antioxidant and antiproliferative activities in vitro[J]. Food Chemistry,2011,129(2):402-407
[39] Giri L,Dhyani P,Rawat S,Bhatt I D,Nandi S K,Rawal R S,Pande V. In vitro production of phenolic compounds and antioxidant activity in callus suspension cultures of Habenaria edgeworthii:A rare Himalayan medicinal orchid[J]. Industrial Crops&Products,2012,37(1):1-6
[40]王有琼,马李一,张重权,段琼芬.印楝叶多酚提取及体外抗氧化活性[J].精细化工,2015,32(1):64-68
[41] Shanmugam M K,An H N,Kumar A P,Tan B K H,Gautam S.Targeted inhibition of tumor proliferation,survival,and metastasis by pentacyclic triterpenoids:Potential role in prevention and therapy of cancer[J]. Cancer Letters,2012,320(2):158-170
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