栽培菊苣籽总黄酮的细胞毒性及清除羟自由基活性成分的识别

Cytotoxicity Evaluation of Total Flavonoids and Recognition of Hydroxyl Radical Scavenging Components from Cichorium endivia L. Seeds

采用3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐(3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide,MTT)法评价栽培菊苣籽总黄酮对小鼠巨噬细胞RAW264.7的毒性。从总抗氧化能力、清除2,2’-联氮-双-(3-乙基苯并噻唑啉-6-磺酸)二铵盐自由基(2,2’-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid)diammonium salt cation radical,ABTS+·)能力、清除O_2^-·能力以及抑制Fe^(2+)诱发的卵黄脂蛋白脂质过氧化能力4个方面评价栽培菊苣籽总黄酮的抗氧化活性,并与常见的抗氧化剂VC和叔丁基对苯二酚(tertbutylhydroquinone,TBHQ)作比较。之后采用高效液相色谱(high-performance liquid chromatography,HPLC)法对栽培菊苣籽总黄酮中清除·OH的活性成分进行了识别。MTT实验结果显示,菊苣籽总黄酮质量浓度为1 mg/mL时,对RAW264.7细胞具有一定的毒性,而当其质量浓度适当减小后,则基本无毒性。抗氧化实验结果表明:栽培菊苣籽总黄酮的总抗氧化活性和清除ABTS^+·的能力弱于VC和TBHQ,但在低质量浓度时,其总抗氧化活性与TBHQ非常接近;在清除O_2^-·方面,活性由强到弱为VC>菊苣籽总黄酮>TBHQ;在抑制卵黄脂蛋白脂质过氧化实验中,栽培菊苣籽总黄酮强于VC,但弱于TBHQ。HPLC法结果显示,绿原酸比洋蓟素具有更高的清除·OH的活性,且二者对·OH的清除率分别为66.42%和46.00%。

The cytotoxicity of total flavonoids from Cichorium endivia L.seeds towards RAW264.7cells was evaluated by3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide(MTT)assay and the antioxidant activities were investigated based on total antioxidant activity,2,2’-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid)diammonium salt cation radical(ABTS+·)scavenging capacity,superoxide anion radical scavenging capacity and inhibition on Fe2+-inducedlipid peroxidation in egg yolk.These activities were compared with those of the common antioxidants vitamin C(VC)and tert-butylhydroquinone(TBHQ).Furthermore,the total flavonoids extracted from C.endivia seeds were screened for those scavenging hydroxyl radical by a high-performance liquid chromatography(HPLC)method.MTT assay showed that the total flavonoids were toxic towards RAW264.7cells at a concentration of1mg/mL,but they were hardly toxic at a certain lower temperature.Antioxidant assays revealed that the total antioxidant activity and ABTS+·scavenging activity of these compounds were lower than those of VC and TBHQ.However,at low concentration,the total antioxidant activity was similar to that of TBHQ.The superoxide anion scavenging capacity of these samples was in the decreasing order of VC>total flavonoids from C.endivia seeds>TBHQ.The lipid peroxidation inhibitory activity of the total flavonoids was higher than that of VC,but lower than that of TBHQ.The HPLC analysis indicated that hydroxyl radical scavenging capacity of chlorogenic acid was higher than that of cynarin,which showed a scavenging percentage of66.42%and46.00%,respectively.