岗梅根和茎超临界CO_2萃取物中低极性挥发性成分比较及其对IEC-6细胞增殖活性的影响
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摘要
目的:比较岗梅根和茎的超临界CO_2萃取物中的低极性挥发性化学成分及其体外对大鼠小肠隐窝上皮细胞IEC-6增殖活性的影响,为充分利用岗梅野生资源和扩大其药用部位提供参考。方法:采用超临界CO_2萃取法提取岗梅根和茎中的低极性挥发性化学成分,通过气质联用法(GC-MS)分析其化学成分组成。以IEC-6细胞为对象,采用不同质量浓度的岗梅根或茎超临界CO_2萃取物(0、1、5、10、20、40、60、80、100μg/mL)处理细胞,以MTT法检测细胞相对活力,绘制细胞增殖曲线图并计算各萃取物的半数有效浓度(EC_(50))。结果:通过GC-MS分析技术从岗梅根和茎超临界CO2萃取物中分别鉴定出62、46个低极性挥发性化学成分,其中共有成分24个,主要为壬酸(根、茎中分别含14.18%、6.14%)、辛酸(根、茎中分别含10.59%、4.35%)、己酸(根、茎中分别含8.63%、10.86%)、丹皮酚(根、茎中分别含7.79%、6.00%)、2-甲基-3-苯基-丙醇(根、茎中分别含6.30%、0.58%)、乙酸(根、茎中分别含1.72%、33.77%)等。细胞体外试验结果显示,岗梅根和茎超临界CO_2萃取物质量浓度较低(≤60μg/mL)时,能显著促进IEC-6细胞增殖,EC_(50)分别为16.35、20.20μg/mL;而当质量浓度较高(≥80μg/mL)时,则表现出细胞毒活性,对IEC-6细胞增殖呈抑制作用。结论:岗梅根和茎中的低极性挥发性化学成分种类相似,其萃取物对IEC-6细胞的体外生物活性也相似;短链脂肪酸很可能是其促进细胞增殖的活性成分,而丹皮酚则可能是其细胞毒活性成分。
OBJECTIVE:To compare low-polarity volatile constituents in supercritical CO_2 extract from the roots and stem of Ilex asprella and its effects on the proliferation of IEC-6 in vitro,and to provide reference for making full use of wild resources of I. asprella and expanding its medicinal parts. METHODS:The low-polarity volatile constituents were extracted from the root and stem of I. asprella with supercritical fluid CO_2 extraction(SFE-CO_2). The chemical constituents were analyzed by GC-MS. IEC-6 cells were treated with different concentrations of supercritical CO_2 extracts(0,1,5,10,20,40,60,80,100 μg/mL) from roots or stems of I. asprella. MTT assay was used to detect the relative viability,and cell proliferation curve was drawn and EC50 of each extract were calculated. RESULTS:Sixty-two and forty-six low-polarity volatile constituents were identified from supercritical CO_2 extract in the roots and stem of I. asprella with GC-MS;there were 24 common constituents totally,mainly including pelargonic acid(14.18% and 6.14%),octanoic acid(10.59% and 4.35%),hexanoic acid(8.63% and 10.86%),paeonol(7.79% and6.00%),2-methyl-3-phenyl-propanal(6.3% and 0.58%),acetic acid(1.72% and 33.77%)in root and stem,respectively. The results of cell culture in vitro showed that when the concentration of supercritical CO_2 extract from the roots and stems of I. asprella was lower(≤60 μg/mL),it could significantly promote the proliferation of IEC-6 cells and their EC_(50) were 16.35,20.20 μg/mL,respectively;when the concentration of the extract was higher(≥80 μg/mL),it showed cytotoxicity and inhibited the proliferation of IEC-6 cells. CONCLUSIONS:There are similar species of volatile constituents in roots and stems of I. asprella and similar in vitro bioactivity of the supercritical CO_2 extracts to IEC-6 cells. The short-chain fatty acids may be the active ingredient to promote cell proliferation,while paeonol may be the cytotoxic active ingredient.
OBJECTIVE:To compare low-polarity volatile constituents in supercritical CO_2 extract from the roots and stem of Ilex asprella and its effects on the proliferation of IEC-6 in vitro,and to provide reference for making full use of wild resources of I. asprella and expanding its medicinal parts. METHODS:The low-polarity volatile constituents were extracted from the root and stem of I. asprella with supercritical fluid CO_2 extraction(SFE-CO_2). The chemical constituents were analyzed by GC-MS. IEC-6 cells were treated with different concentrations of supercritical CO_2 extracts(0,1,5,10,20,40,60,80,100 μg/mL) from roots or stems of I. asprella. MTT assay was used to detect the relative viability,and cell proliferation curve was drawn and EC50 of each extract were calculated. RESULTS:Sixty-two and forty-six low-polarity volatile constituents were identified from supercritical CO_2 extract in the roots and stem of I. asprella with GC-MS;there were 24 common constituents totally,mainly including pelargonic acid(14.18% and 6.14%),octanoic acid(10.59% and 4.35%),hexanoic acid(8.63% and 10.86%),paeonol(7.79% and6.00%),2-methyl-3-phenyl-propanal(6.3% and 0.58%),acetic acid(1.72% and 33.77%)in root and stem,respectively. The results of cell culture in vitro showed that when the concentration of supercritical CO_2 extract from the roots and stems of I. asprella was lower(≤60 μg/mL),it could significantly promote the proliferation of IEC-6 cells and their EC_(50) were 16.35,20.20 μg/mL,respectively;when the concentration of the extract was higher(≥80 μg/mL),it showed cytotoxicity and inhibited the proliferation of IEC-6 cells. CONCLUSIONS:There are similar species of volatile constituents in roots and stems of I. asprella and similar in vitro bioactivity of the supercritical CO_2 extracts to IEC-6 cells. The short-chain fatty acids may be the active ingredient to promote cell proliferation,while paeonol may be the cytotoxic active ingredient.
引文
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[12]KANATA Y,SHIRAGA H,TAI A,et al.Induction of neurite outgrowth in PC12 cells by the medium-chain fatty acid octanoic acid[J].Neuroscience,2007,146(3):1073-1081.
[13]LIU C,CHU I,HWANG S.Pentanoic acid,a novel protein synthesis stimulant for Chinese hamster ovary(CHO)cells[J].J Biosci Bioeng,2001,91(1):71-75.
[14]FU J,YU L,LOU J,et al.Paeonol induces the apoptosis of the SGC-7901 gastric cancer cell line by downregulating ERBB2 and inhibiting the NF-κB signaling pathway[J].Int J Mol Med,2018,42(3):1473-1483.
[15]LYU ZK,LI CL,JIN Y,et al.Paeonol exerts potential activities to inhibit the growth,migration and invasion of human gastric cancer BGC823 cells via downregulating MMP-2 and MMP-9[J].Mol Med Rep,2017,16(5):7513-7519.
[2]陈蔚文,徐鸿华.岭南道地药材研究[M].广州:广东科技出版社,2007:291.
[3]广东省食品药品监督管理局.广东省中药材标准:第一册[S].广州:广东科技出版社,2004:111.
[4]YOSHIKI K,DE CZ.Antitumor agents,cytotoxic aspellic acid A and C and asperllic acid B,new p-coumaroyl triterpenes,from Ilex asprella[J].J Nat Prod,1993,56(12):2077-2082.
[5]HU XY,SHU XC,GUO Y,et al.Effect of an Ilex asprella root decoction on the related genes of lipid metabolism from chronic stress and hyperlipidemic fatty liver in rats[J].Chin Med J:Engl,2012,125(19):3539-3542.
[6]ZHOU M,XU M,MA XX,et al.Antiviral triterpenoid saponins from the roots of Ilex asprella[J].Planta Medica,2012,78(15):1702-1705.
[7]WANG L,CAI Y,ZHANG XQ,et al.New triterpenoid glycosides from roots of Ilex asprella[J].Carbohydr Res,2012.DOI:10.1016/j.carres.2011.12.003.
[8]蔡艳,张庆文,李旨君,等.岗梅根化学成分的研究[J].中草药,2010,41(9):1426-1429.
[9]ZHANG ZX,FU Q,ZHENG KYZ,et al.Three new triterpene glycosides from Ilex asprella[J].J Asian Nat Prod Res,2013,15(5):453-458.
[10]李敏华,俞世杰,杜上鑑.岗梅根化学成分的研究[J].中草药,1997,28(8):454-456.
[11]王璐璇,刘玥宏,朱继开,等.短链脂肪酸在疾病治疗中的研究进展[J].世界华人消化杂志,2017,25(13):1179-1186.
[12]KANATA Y,SHIRAGA H,TAI A,et al.Induction of neurite outgrowth in PC12 cells by the medium-chain fatty acid octanoic acid[J].Neuroscience,2007,146(3):1073-1081.
[13]LIU C,CHU I,HWANG S.Pentanoic acid,a novel protein synthesis stimulant for Chinese hamster ovary(CHO)cells[J].J Biosci Bioeng,2001,91(1):71-75.
[14]FU J,YU L,LOU J,et al.Paeonol induces the apoptosis of the SGC-7901 gastric cancer cell line by downregulating ERBB2 and inhibiting the NF-κB signaling pathway[J].Int J Mol Med,2018,42(3):1473-1483.
[15]LYU ZK,LI CL,JIN Y,et al.Paeonol exerts potential activities to inhibit the growth,migration and invasion of human gastric cancer BGC823 cells via downregulating MMP-2 and MMP-9[J].Mol Med Rep,2017,16(5):7513-7519.