金银花蕾提取物及其3-咖啡酰奎宁酸抗乙肝病毒活性
|
摘要
以HepG2. 2. 15细胞为乙肝病毒(HBV)感染的体外实验模型,采用MTT法测定实验组细胞的活力,采用酶联免疫吸附试验(ELISA)测定实验组HepG2. 2. 15细胞上清液中乙肝病毒表面抗原(HBsAg)、e抗原(HBeAg)的含量,采用实时荧光定量PCR法检测HepG2. 2. 15细胞上清液HBV DNA水平,研究金银花蕾乙醇提取物(LA)、乙酸乙酯部位(LE)及活性成分3-咖啡酰奎宁酸体外抗乙肝病毒的作用。研究结果显示:LA、LE及3-咖啡酰奎宁酸对HepG2. 2. 15细胞的生长增殖均无显著抑制作用;LA和LE作用8 d可抑制HBeAg分泌;100μg/mL 3-咖啡酰奎宁酸对HepG2. 2. 15细胞上清液中HBsAg、HBeAg分泌及DNA复制均具有显著的抑制作用。上述结果表明,金银花中的活性成分3-咖啡酰奎宁酸具有显著的抗HBV活性。
HepG2.2.15 cells were used as a model for hepatitis B virus(HBV)infection in vitro. MTT assays were used to determine the viabilities of HepG2.2.15 cells in different experimental groups. In order to study the effects of ethanol extracts and 3-caffeoylquinic acid from Lonicera japonica flower buds against HBV in vitro, enzyme-linked immunesorbent assays(ELISA) were applied to quantify the contents of HBsAg and HBeAg in the supernatant of each experimental group, and real-time PCR was applied to quantify the HBV DNA levels. The results indicate that the ethanol extract(LA), ethyl acetate fraction(LE) and 3-caffeoylquinic acid have no inhibitory effects on the viabilities of HepG2.2.15 cells; LA and LE can inhibit the secretion of HBeAg after 8-day treatment; 3-caffeoylquinic acid can inhibit the secretion of HBsAg, HBeAg and the replication of HBV DNA in HepG2.2.15 cells at the concentration of 100 μg/mL. In summary, the active ingredient 3-caffeoylquinic acid exerts prominent activity against HBV in vitro.
HepG2.2.15 cells were used as a model for hepatitis B virus(HBV)infection in vitro. MTT assays were used to determine the viabilities of HepG2.2.15 cells in different experimental groups. In order to study the effects of ethanol extracts and 3-caffeoylquinic acid from Lonicera japonica flower buds against HBV in vitro, enzyme-linked immunesorbent assays(ELISA) were applied to quantify the contents of HBsAg and HBeAg in the supernatant of each experimental group, and real-time PCR was applied to quantify the HBV DNA levels. The results indicate that the ethanol extract(LA), ethyl acetate fraction(LE) and 3-caffeoylquinic acid have no inhibitory effects on the viabilities of HepG2.2.15 cells; LA and LE can inhibit the secretion of HBeAg after 8-day treatment; 3-caffeoylquinic acid can inhibit the secretion of HBsAg, HBeAg and the replication of HBV DNA in HepG2.2.15 cells at the concentration of 100 μg/mL. In summary, the active ingredient 3-caffeoylquinic acid exerts prominent activity against HBV in vitro.
引文
[1]中华医学会肝病学分会,中华医学会感染病学分会.慢性乙型肝炎防治指南(2010年版)[J].中华内科杂志,2011,3(1):66-82. DOI:10.3969/j. issn. 1001-5256.2011.01.031.Chinese Medical Association Liver Diseases Branch,Chinese Medical Association Infectious Diseases Branch. The guideline of prevention and treatment for chronic hepatitis B(2010 version)[J]. Chinese Journal of Internal Medicine,2011,3(1):66-82. DOI:10.3969/j. issn. 1001-5256. 2011.01.031(Ch).
[2] LEE E,KIM J S,KIM H P,et al. Phenolic constituents from the flower buds of Lonicera japonica and their 5-lipoxygenase inhibitory activities[J]. Food Chemistry,2010,120(1):134-139. DOI:10.1016/j.foodchem.2009.09.088.
[3] YU Y,JIANG Z B,SONG W X,et al. Glucosylated caffeoylquinic acid derivatives from the Flower buds of Lonicera japonica[J]. Acta Pharmaceutica Sinica B,2015,5(3):210-214. DOI:10.1016/j.apsb.2015.01.012.
[4] YU Y,ZHU C G,WANG S J,et al. Homosecoiridoid alkaloids with amino acid units from the flower buds of Lonicera japonica[J]. Journal of Natural Products,2013,76(12):2226-2233. DOI:10.1021/np4005773.
[5] KURODA M,SHIZUME T,MIMAKI Y. Triterpene glycosides from the stems and leaves of Lonicera japonica[J]. Chemical and Pharmaceutical Bulletin,2014,62(1):92-96. DOI:10.1248/cpb.c13-00592.
[6] GE L L,LI J M,WAN H Q,et al. Novel flavonoids from Lonicera japonica flower buds and validation of their anti-hepatoma and hepatoprotective activity in vitro studies[J]. Industrial Crops and Products,2018,125:114-122. DOI:10.1016/j.indcrop.2018.08.073.
[7] XIONG J H,LI S C,WANG W J,et al. Screening and identification of the antibacterial bioactive compounds from Lonicera japonica Thunb. leaves[J]. Food Chemistry,2013,138(1):327-333. DOI:10.1016/j.foodchem.2012.10.127.
[8] SEO O N,KIM G S,PARK S,et al. Determination of polyphenol components of Lonicera japonica flower buds.using liquid chromatography-tandem mass spectrometry:Contribution to the overall antioxidant activity[J]. Food Chemistry,2012,134(1):572-577. DOI:10.1016/j.foodchem.2012.02.124.
[9] WU Y H. Naturally derived anti-hepatitis B virus agents and their mechanism of action[J]. World Journal of Gastroenterology,2016,22(1):188-204. DOI:10.3748/wjg.v22.i1.188.
[10] WU Y H,HAO B J,CAO H C,et al. Anti-hepatitis B virus effect and possible mechanism of action of 3,4-O-dicaffeoylquinic acid in vitro and in vivo[J]. EvidenceBased Complementary and Alternative Medicine,2012,2012:1-9. DOI:10.1155/2012/356806.
[11] GE L L,WAN H Q,TANG S M,et al. Novel caffeoylquinic acid derivatives from Lonicera japonica Thunb. flower buds exert pronounced anti-HBV activities[J]. RSC Advances,2018,8(62):35374-35385. DOI:10.1016/j.indcrop.2018.08.073.
[12] SELLS M A,CHEN M L,ACS G. Production with cloned hepatitis B virus particles in HepG2 cells transfected with cloned hepatitis B virus DNA[J]. Proceedings of the National Academy of Sciences,1987,84(4):1005-1009. DOI:10.1073/pnas.84.4.1005.
[13] CHEN J X,SHEN H H,NIU M,et al. Anti-hepatitis B virus effect of matrine-type alkaloid and involvement of p38mitogen-activated protein kinase and tumor necrosis factor receptor-associated factor 6[J]. Virus Research,2016,215:104-113. DOI:10.1016/j.virusres.2015.12.005.
[14] FARAG M M S,MANSOUR M T M. Characterization of subviral particles of hepatitis B virus produced by HepG2.2.15 cell line—in vitro study[J]. International Journal of Virology and Molecular Biology,2016,5(1):1-7. DOI:10.5923/j.ijvmb.20160501.01.
[15] WEN D D,WANG J N,YAN H W,et al. Effect of radix trichosanthis and trichosanthin on hepatitis B virus in HepG2. 2.15 cells[J]. Journal of Nanoscience and Nanotechnology,2015,15(3):2094-2098. DOI:10.1166/jnn.2015.9271.
[16] JI D,ZHANG C F,LI J Z,et al. A new iridoid glycoside from the roots of Dipsacus asper[J]. Molecules,2012,17(2):1419-1424. DOI:10.3390/molecules17021419.
[2] LEE E,KIM J S,KIM H P,et al. Phenolic constituents from the flower buds of Lonicera japonica and their 5-lipoxygenase inhibitory activities[J]. Food Chemistry,2010,120(1):134-139. DOI:10.1016/j.foodchem.2009.09.088.
[3] YU Y,JIANG Z B,SONG W X,et al. Glucosylated caffeoylquinic acid derivatives from the Flower buds of Lonicera japonica[J]. Acta Pharmaceutica Sinica B,2015,5(3):210-214. DOI:10.1016/j.apsb.2015.01.012.
[4] YU Y,ZHU C G,WANG S J,et al. Homosecoiridoid alkaloids with amino acid units from the flower buds of Lonicera japonica[J]. Journal of Natural Products,2013,76(12):2226-2233. DOI:10.1021/np4005773.
[5] KURODA M,SHIZUME T,MIMAKI Y. Triterpene glycosides from the stems and leaves of Lonicera japonica[J]. Chemical and Pharmaceutical Bulletin,2014,62(1):92-96. DOI:10.1248/cpb.c13-00592.
[6] GE L L,LI J M,WAN H Q,et al. Novel flavonoids from Lonicera japonica flower buds and validation of their anti-hepatoma and hepatoprotective activity in vitro studies[J]. Industrial Crops and Products,2018,125:114-122. DOI:10.1016/j.indcrop.2018.08.073.
[7] XIONG J H,LI S C,WANG W J,et al. Screening and identification of the antibacterial bioactive compounds from Lonicera japonica Thunb. leaves[J]. Food Chemistry,2013,138(1):327-333. DOI:10.1016/j.foodchem.2012.10.127.
[8] SEO O N,KIM G S,PARK S,et al. Determination of polyphenol components of Lonicera japonica flower buds.using liquid chromatography-tandem mass spectrometry:Contribution to the overall antioxidant activity[J]. Food Chemistry,2012,134(1):572-577. DOI:10.1016/j.foodchem.2012.02.124.
[9] WU Y H. Naturally derived anti-hepatitis B virus agents and their mechanism of action[J]. World Journal of Gastroenterology,2016,22(1):188-204. DOI:10.3748/wjg.v22.i1.188.
[10] WU Y H,HAO B J,CAO H C,et al. Anti-hepatitis B virus effect and possible mechanism of action of 3,4-O-dicaffeoylquinic acid in vitro and in vivo[J]. EvidenceBased Complementary and Alternative Medicine,2012,2012:1-9. DOI:10.1155/2012/356806.
[11] GE L L,WAN H Q,TANG S M,et al. Novel caffeoylquinic acid derivatives from Lonicera japonica Thunb. flower buds exert pronounced anti-HBV activities[J]. RSC Advances,2018,8(62):35374-35385. DOI:10.1016/j.indcrop.2018.08.073.
[12] SELLS M A,CHEN M L,ACS G. Production with cloned hepatitis B virus particles in HepG2 cells transfected with cloned hepatitis B virus DNA[J]. Proceedings of the National Academy of Sciences,1987,84(4):1005-1009. DOI:10.1073/pnas.84.4.1005.
[13] CHEN J X,SHEN H H,NIU M,et al. Anti-hepatitis B virus effect of matrine-type alkaloid and involvement of p38mitogen-activated protein kinase and tumor necrosis factor receptor-associated factor 6[J]. Virus Research,2016,215:104-113. DOI:10.1016/j.virusres.2015.12.005.
[14] FARAG M M S,MANSOUR M T M. Characterization of subviral particles of hepatitis B virus produced by HepG2.2.15 cell line—in vitro study[J]. International Journal of Virology and Molecular Biology,2016,5(1):1-7. DOI:10.5923/j.ijvmb.20160501.01.
[15] WEN D D,WANG J N,YAN H W,et al. Effect of radix trichosanthis and trichosanthin on hepatitis B virus in HepG2. 2.15 cells[J]. Journal of Nanoscience and Nanotechnology,2015,15(3):2094-2098. DOI:10.1166/jnn.2015.9271.
[16] JI D,ZHANG C F,LI J Z,et al. A new iridoid glycoside from the roots of Dipsacus asper[J]. Molecules,2012,17(2):1419-1424. DOI:10.3390/molecules17021419.