脱脂麦胚提取物抗氧化、抗肿瘤活性的研究
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摘要
小麦胚芽是小麦面粉加工的主要副产物之一,含有多种生理活性成分,极具开发前景。本论文主要对脱脂麦胚提取物(DWGE)的抗氧化和抗肿瘤活性进行研究,以期为深度开发和利用小麦胚芽资源提供理论支持。本文主要研究内容包括以下3个方面:
研究了脱脂麦胚水提物、30%醇提物、50%醇提物、70%醇提物以及无水乙醇提取物体外抗氧化活性,主要从对DPPH和ABTS自由基清除作用、在亚油酸体系中的抗氧化活性以及还原能力这四个方面展开。结果表明,DWGE对DPPH和ABTS自由基有显著的清除作用,随着提取物浓度增加,清除作用增强。从二者的IC50可以看出,70%醇提物清除DPPH自由基能力最强,无水乙醇提取物清除ABTS自由基的能力最强,水提物清除自由基的能力较弱。在亚油酸自动氧化体系中,五种提取物都具有一定的抗氧化活性,其中30%,50%和70%醇提物能够明显的抑制亚油酸的自动氧化,而水提物和无水乙醇提取物对亚油酸自动氧化的抑制作用较弱。五种DWGE中,无水乙醇提取物和70%醇提物的还原能力较强,水提物较弱。综合这四个抗氧化指标可以看出,50%和70%醇提物体外抗氧化活性较好。
研究了DWGE对高脂日粮饲喂小鼠血脂代谢、抗氧化能力的影响。选用4周龄雄性健康C57BL/6小鼠60只,按体重随机分成6组,即正常组(正常日粮)、高脂组(高脂日粮)、低剂量50%醇提组(高脂日粮+2%的50%醇提物)、高剂量50%醇提组(高脂日粮+6%的50%醇提物)、低剂量70%醇提组(高脂日粮+2%的70%醇提物)和高剂量70%醇提组(高脂日粮+6%的70%醇提物)。饲养8周后测定各组小鼠血脂指标、各组织中的总抗氧化能力、抗氧化酶活性和脂质过氧化物丙二醛(MDA)含量。实验结果表明,在高脂日粮中添加脱脂麦胚50%醇提物和70%醇提物均能够有效提高各组织中总抗氧化能力以及超氧化物歧化酶(SOD)、过氧化氢酶(CAT),谷胱甘肽过氧化物酶(GSH-PX)等抗氧化酶的活性,降低MDA含量,同时提取物的添加可以有效改善高脂日粮引起的机体血浆高密度脂蛋白胆固醇(HDL-C)含量降低,甘油三酯(TG)、总胆固醇(TC)、低密度脂蛋白胆固醇(LDL-C)的含量升高。由此判断,脱脂麦胚50%醇提物和70%醇提物可以改善高脂日粮对机体的不良影响,能够有效的缓解氧化应激和调节血脂代谢,呈现了降脂和抗氧化活性。
研究了DWGE对乳腺癌细胞(MDA-MB231)增殖抑制活性。实验表明DWGE对MDA-MB231细胞生长有明显的抑制效应,水提物作用于MDA-MB231细胞24h、48h、72h的IC50分别为1755.88ug/mL、721.07ug/mL、650.32ug/mL,30%醇提物作用于MDA-MB231细胞24h、48h、72h的IC50分别为3210.2ug/mL、1446.05ug/mL、1084.72ug/mL,呈现了一定的时间剂量效应,并且水提物的抗肿瘤活性优于30%乙醇提取物。光学显微镜和扫描电子显微镜观察到提取物作用后的细胞,细胞数明显变少,细胞贴壁异常,簇集成团,细胞纤维状消失,逐渐变圆,折光率降低,细胞表面的微绒毛消失,细胞膜皱缩,形成大量的凋亡小泡。荧光显微镜下,提取物作用后的细胞细胞核变小,皱缩,呈现致密浓染强荧光,部分细胞的细胞核呈新月状,高浓度提取物作用后细胞细胞核破裂,呈片絮状分散于细胞中。以上都是细胞发生凋亡的明显特征。
本研究证实了DWGE有良好的调节血脂、抗氧化和抑制乳腺癌MDA-MB231细胞生长增殖作用,可有效防止血脂代谢异常、心血管疾病和癌症的发生,有助于延缓衰老,为开发以脱脂麦胚为成分的抗癌、降脂、抗氧化保健品奠定基础。
Wheat germ is one of the main byproducts of wheat-milling industry, and it is a rich source of many bioactive compounds, so study on wheat germ is meaningful. In this paper, antioxidant and anticancer activities of defatted wheat germ extracts (DWGE) were studied to provide theoretical support for the development and exploitation of wheat germ.
The main sections in this study are as follows:
The antioxidant potential of DWGE in vitro was measured firstly by the radical scavenging activity, the inhibition on linoleic acid peroxidation and reducing power. All the extracts showed the scavenging activities of DPPH and ABTS in a concentration-dependent manner significantly. Among the DWGE, 70% ethanol extract was excellent in scavenging DPPH radical while 100 % ethanol extract showed the highest ABTS radical scavenging activity. All the extracts could inhibit the linoleic acid peroxidation, 30% ethanol extract, 50% ethanol extract and 70% ethanol extract could prolong the induction period of autoxidation of linoleic acid significantly compared with aqueous extract and ethanol extract. All the extracts showed some degree of electron-donating capacity in a linear concentration-dependent manner. The reducing power of ethanol extract and 70% ethanol extract were superior to other three extracts. It was summarized from four assays that both 50% ethanol extract and 70% ethanol extract exhibited high antioxidant activities.
The effect of DWGE on lipid metabolism and antioxidative capacity of high-fat diet fed mice was studied. 60 male C57BL/6 mice were divided into 6 groups randomly. The control group was fed with basal diet, and the other five groups were fed with high-fat diet, high-fat diet and 2% 50% ethanol extract, high-fat diet and 6% 50% ethanol extract, high-fat diet and 2% 70% ethanol extract and high-fat diet and 6% 70% ethanol extract, respectively. After 8 weeks feeding, the plasma lipid status, the activities of antioxidases and the contents of MDA in organs were examined. The results were as follows: compared with the control, high levels of dietary fat increased fat-mediated oxidative stress, the content of MDA and the levels of plasma lipid in high-fat diet groups. The supplement of DWGE could modulate lipid metabolism, decrease the effect of high-fat diet, increase the antioxidative capacity of high-fat diet fed mice.
The inhibitory effect of DWGE on the growth and proliferation of human breast carcinoma cell line MDA-MB231 in vitro was observed. MDA-MB231 cell was incubated in vitro by cell biology technique. By the method of MTT, the inhibition of DWGE on the proliferation of MDA-MB231 cell were remarkable and existed in dose- and time-dependent manners. Aqueous extract showed higher antitumor activity than 30% ethanol extract.
The morphological changes of MDA-MB231 cell treated with DWGE was investigated by light microscope and electron microscope. After treated with DWGE, MDA-MB231 cell showed typical apoptotic characteristics, such as cell shrinkage, cell volume reduction, the villus on the surface of MDA-MB231 cell disappeared, and the membrane was shrinkage to form apoptotic bodies, the typical apoptotic features was observed from the cell nucleus of MDA-MB231 cell by fluorescent microscopy, such as chromatin condensation, nuclear fragmentation.
These results demonstrated DWGE possessed antioxidant activity, antilipemic action and antitumor capacity, and it is effective in the prevention of dyslipidemia, cardiovascular disease, aging and cancer. DWG could serve as a new source of natural antioxidants or nutraceuticals with potential applications to reducing the level of oxidative stress and related health benefits.
研究了脱脂麦胚水提物、30%醇提物、50%醇提物、70%醇提物以及无水乙醇提取物体外抗氧化活性,主要从对DPPH和ABTS自由基清除作用、在亚油酸体系中的抗氧化活性以及还原能力这四个方面展开。结果表明,DWGE对DPPH和ABTS自由基有显著的清除作用,随着提取物浓度增加,清除作用增强。从二者的IC50可以看出,70%醇提物清除DPPH自由基能力最强,无水乙醇提取物清除ABTS自由基的能力最强,水提物清除自由基的能力较弱。在亚油酸自动氧化体系中,五种提取物都具有一定的抗氧化活性,其中30%,50%和70%醇提物能够明显的抑制亚油酸的自动氧化,而水提物和无水乙醇提取物对亚油酸自动氧化的抑制作用较弱。五种DWGE中,无水乙醇提取物和70%醇提物的还原能力较强,水提物较弱。综合这四个抗氧化指标可以看出,50%和70%醇提物体外抗氧化活性较好。
研究了DWGE对高脂日粮饲喂小鼠血脂代谢、抗氧化能力的影响。选用4周龄雄性健康C57BL/6小鼠60只,按体重随机分成6组,即正常组(正常日粮)、高脂组(高脂日粮)、低剂量50%醇提组(高脂日粮+2%的50%醇提物)、高剂量50%醇提组(高脂日粮+6%的50%醇提物)、低剂量70%醇提组(高脂日粮+2%的70%醇提物)和高剂量70%醇提组(高脂日粮+6%的70%醇提物)。饲养8周后测定各组小鼠血脂指标、各组织中的总抗氧化能力、抗氧化酶活性和脂质过氧化物丙二醛(MDA)含量。实验结果表明,在高脂日粮中添加脱脂麦胚50%醇提物和70%醇提物均能够有效提高各组织中总抗氧化能力以及超氧化物歧化酶(SOD)、过氧化氢酶(CAT),谷胱甘肽过氧化物酶(GSH-PX)等抗氧化酶的活性,降低MDA含量,同时提取物的添加可以有效改善高脂日粮引起的机体血浆高密度脂蛋白胆固醇(HDL-C)含量降低,甘油三酯(TG)、总胆固醇(TC)、低密度脂蛋白胆固醇(LDL-C)的含量升高。由此判断,脱脂麦胚50%醇提物和70%醇提物可以改善高脂日粮对机体的不良影响,能够有效的缓解氧化应激和调节血脂代谢,呈现了降脂和抗氧化活性。
研究了DWGE对乳腺癌细胞(MDA-MB231)增殖抑制活性。实验表明DWGE对MDA-MB231细胞生长有明显的抑制效应,水提物作用于MDA-MB231细胞24h、48h、72h的IC50分别为1755.88ug/mL、721.07ug/mL、650.32ug/mL,30%醇提物作用于MDA-MB231细胞24h、48h、72h的IC50分别为3210.2ug/mL、1446.05ug/mL、1084.72ug/mL,呈现了一定的时间剂量效应,并且水提物的抗肿瘤活性优于30%乙醇提取物。光学显微镜和扫描电子显微镜观察到提取物作用后的细胞,细胞数明显变少,细胞贴壁异常,簇集成团,细胞纤维状消失,逐渐变圆,折光率降低,细胞表面的微绒毛消失,细胞膜皱缩,形成大量的凋亡小泡。荧光显微镜下,提取物作用后的细胞细胞核变小,皱缩,呈现致密浓染强荧光,部分细胞的细胞核呈新月状,高浓度提取物作用后细胞细胞核破裂,呈片絮状分散于细胞中。以上都是细胞发生凋亡的明显特征。
本研究证实了DWGE有良好的调节血脂、抗氧化和抑制乳腺癌MDA-MB231细胞生长增殖作用,可有效防止血脂代谢异常、心血管疾病和癌症的发生,有助于延缓衰老,为开发以脱脂麦胚为成分的抗癌、降脂、抗氧化保健品奠定基础。
Wheat germ is one of the main byproducts of wheat-milling industry, and it is a rich source of many bioactive compounds, so study on wheat germ is meaningful. In this paper, antioxidant and anticancer activities of defatted wheat germ extracts (DWGE) were studied to provide theoretical support for the development and exploitation of wheat germ.
The main sections in this study are as follows:
The antioxidant potential of DWGE in vitro was measured firstly by the radical scavenging activity, the inhibition on linoleic acid peroxidation and reducing power. All the extracts showed the scavenging activities of DPPH and ABTS in a concentration-dependent manner significantly. Among the DWGE, 70% ethanol extract was excellent in scavenging DPPH radical while 100 % ethanol extract showed the highest ABTS radical scavenging activity. All the extracts could inhibit the linoleic acid peroxidation, 30% ethanol extract, 50% ethanol extract and 70% ethanol extract could prolong the induction period of autoxidation of linoleic acid significantly compared with aqueous extract and ethanol extract. All the extracts showed some degree of electron-donating capacity in a linear concentration-dependent manner. The reducing power of ethanol extract and 70% ethanol extract were superior to other three extracts. It was summarized from four assays that both 50% ethanol extract and 70% ethanol extract exhibited high antioxidant activities.
The effect of DWGE on lipid metabolism and antioxidative capacity of high-fat diet fed mice was studied. 60 male C57BL/6 mice were divided into 6 groups randomly. The control group was fed with basal diet, and the other five groups were fed with high-fat diet, high-fat diet and 2% 50% ethanol extract, high-fat diet and 6% 50% ethanol extract, high-fat diet and 2% 70% ethanol extract and high-fat diet and 6% 70% ethanol extract, respectively. After 8 weeks feeding, the plasma lipid status, the activities of antioxidases and the contents of MDA in organs were examined. The results were as follows: compared with the control, high levels of dietary fat increased fat-mediated oxidative stress, the content of MDA and the levels of plasma lipid in high-fat diet groups. The supplement of DWGE could modulate lipid metabolism, decrease the effect of high-fat diet, increase the antioxidative capacity of high-fat diet fed mice.
The inhibitory effect of DWGE on the growth and proliferation of human breast carcinoma cell line MDA-MB231 in vitro was observed. MDA-MB231 cell was incubated in vitro by cell biology technique. By the method of MTT, the inhibition of DWGE on the proliferation of MDA-MB231 cell were remarkable and existed in dose- and time-dependent manners. Aqueous extract showed higher antitumor activity than 30% ethanol extract.
The morphological changes of MDA-MB231 cell treated with DWGE was investigated by light microscope and electron microscope. After treated with DWGE, MDA-MB231 cell showed typical apoptotic characteristics, such as cell shrinkage, cell volume reduction, the villus on the surface of MDA-MB231 cell disappeared, and the membrane was shrinkage to form apoptotic bodies, the typical apoptotic features was observed from the cell nucleus of MDA-MB231 cell by fluorescent microscopy, such as chromatin condensation, nuclear fragmentation.
These results demonstrated DWGE possessed antioxidant activity, antilipemic action and antitumor capacity, and it is effective in the prevention of dyslipidemia, cardiovascular disease, aging and cancer. DWG could serve as a new source of natural antioxidants or nutraceuticals with potential applications to reducing the level of oxidative stress and related health benefits.
引文
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