CN108324757A - 胡麻多酚的提取方法及抗肿瘤应用 - Google Patents
胡麻多酚的提取方法及抗肿瘤应用 Download PDFInfo
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Abstract
本发明提供了一种胡麻多酚的提取方法及其抗肿瘤应用。胡麻多酚提取方法：将胡麻粕干燥后粉碎，进行脱胶处理，干燥备用；按质量比1:60–75将脱胶胡麻粉溶于60％的乙醇中，在42‑55℃下，400W功率超声提取50‑60min；将溶液离心处理后，收集上清，以体积比1：1‑1.3加入乙酸乙酯中进行除脂，多次重复除脂，将收集的多酚溶液进行旋转蒸发浓缩，将剩余液体进行冷冻干燥，收集干粉即为胡麻多酚。体外抗肿瘤实验表明，胡麻多酚能明显抑制多种癌细胞增殖而无细胞毒性，具有良好的抗肿瘤活性，可在制备抗肿瘤药物和保健食品中应用。
Description
技术领域
本发明涉植物有效成分提取，特别涉及及亚麻类草本植物中多酚的提取和应用，具体属于胡麻中多酚的提取方法，及其在抗肿瘤药物和保健食品中的应用。
背景技术
胡麻(Linumusitatissimum Linn.)即油用亚麻，属亚麻科亚麻属，一年生草本植物。胡麻的种子(胡麻籽)呈扁平长卵圆形，长4～6mm，宽2～3mm，厚约l mm，每粒种子平均重3～ 13mg。表面红棕色或褐色，平滑有光泽，周边渐薄，中间微凸。一端钝圆，另一端尖而略偏斜。种脐位于尖端的凹入处。种脊浅棕色，位于一侧边缘。种皮薄，除去种皮后可见棕色薄膜状的胚乳，内有子叶2片，黄白色，富油性，胚根胡向种子的尖端。气无，嚼之有豆腥味，并有粘舌感，入水中亦有粘性物质分离出。麻籽种皮富含胶质和纤维，并含有少量蛋白质和脂肪。
传统中医认为亚麻籽具有活血润燥、祛风解毒、益肝肾养护皮肤的功效。《本经》“土伤中虚赢，补五内，益气力，长肌肉，填髓脑。”《新修本草》“生嚼涂小儿头疮，煎汤浴恶疮、妇人阴疮。大效。”《日华诸家本草》“补中益气，润养五脏，补肺气，止心惊，利人小肠。”《本草纲目》“胡麻取油，以百者为胜，服食以黑者为息，取其黑者入通于肾，而能润燥也。《本草从新》“凉血解毒，服之令人滑肠，精气不固者亦勿宜食”。胡麻籽的主要成分是油脂和蛋白质，其它的成分包括非淀粉多糖、胡麻胶、酚类物质、胰岛素抑制剂、木酚素(植物雌性激素)等。胡麻籽中的多种成分都具有优良的营养价值和保健功能。许多国家将胡麻籽作为食品配料。添加于食品中以起到营养或保健的作用，或研发出保健产品。
多酚(Polyphenols)，不是某一种物质的名称，而是某一类化学物质的总称。其主要活性成分为多羟基酚类化合物，是含若干个酚性羟基结构的次生代谢物，包括黄酮类、单宁类、酚酸类以及花色苷类等，广泛分布于植物根、枝、叶、果实，具有抑菌、抗癌、抗氧化等多种生物活性。大量研究表明植物多酚具有抗菌、抗氧化、抗癌和抗心血管疾病等疗效，目前在食品、药品和化妆品等行业普遍应用。但尚未看到胡麻多酚抗肿瘤活性的研究。
发明内容
本发明的目的在于提供一种胡麻多酚的提取方法，以及胡麻多酚在制备抗肿瘤药物和保健食品中的应用。
本发明提供的一种胡麻多酚提取方法，包括如下步骤：
(1)取胡麻粕，分离成籽粒状；
(2)按质量比1:3-4加入蒸馏水，65-75℃的水浴并搅拌0.5-1h，静置，过滤，漂洗；
(3)重复步骤(2)多次，直到胡麻籽粒表面无粘胶为止，然后将脱胶的胡麻籽粒在40℃烘箱中完全干燥，粉碎，得到脱胶胡麻粉；
(4)按质量比1:60-75将脱胶胡麻粉溶于60％的乙醇中，在42-55℃下，在400W的超声功率下超声提取50-60分钟；
(5)将超声提取的混合料液离心，弃去沉淀，上清在相同条件下再离心一次，收集上清；
(6)将步骤(5)得到的上清进行去脂处理，按1:1-1.3的体积比加入乙酸乙酯，离心，室温静置，分层，将最下层多酚溶液吸出；吸出的多酚溶液重复去脂处理步骤3-4次，得到脱脂的多酚溶液；
(7)将步骤(6)得到的脱脂的多酚溶液进行旋转蒸发，冷冻干燥，得到的粉末即为胡麻多酚。
步骤(4)中脱胶胡麻粉和60％的乙醇的质量比1:65-72。
步骤(4)中提取温度50℃，在400W的超声功率下超声提取50分钟。
步骤(6)中上清和乙酸乙酯体积比为1:1。
步骤(7)中所述的冷冻干燥的条件：置于-80℃，冷冻过夜，再置于冷冻干燥机中冷冻 1-3天。
所述胡麻粕又叫亚麻粕，是亚麻籽榨油后的副产品。
上述制备方法制得的胡麻多酚可在制备新型抗癌药物中应用，也可在制备保健食品中应用。
与现有技术相比本发明的有益效果：
上述方法制得的胡麻多酚可明显抑制多种癌细胞增殖，且具有良好的抗癌活性。经体外抗肿瘤活性检测结果显示，用不同浓度的该胡麻多酚处理多种人结肠癌细胞以及宫颈癌细胞等(DLD1，HCT-116，SW480，SW620，HT-29，A549，HeLa,HepG2,MCF-7)24小时后，均具有抑制作用，所以该胡麻多酚具有广谱的抗肿瘤作用。
该方法制备的多酚，因其来源于纯天然的植物，加之纯化工艺简单，因此有较大的开发前景和应用价值。如：在医药方面，可作为其他药物的辅料或制备具有抗肿瘤活性的癌症辅助药物；在保健品方面，可制备具有抗肿瘤和抗氧化功能的保健口服液。
本发明以胡麻粕为材料，提取出多酚成分，有利于促进胡麻粕资源的深度开发和有效利用。
附图说明
图1胡麻多酚样品
图2胡麻多酚影响DLD1，HCT-116癌细胞和FHC正常细胞增殖的柱状图
图3胡麻多酚抑制SW480，SW620和HT-29细胞的增殖的柱状图
图4胡麻多酚抑制A549，HeLa，HepG2和MCF-7癌细胞的增殖的柱状图
具体实施方式
实施例1
胡麻多酚的制备：
(1)取胡麻粕500g，分离成籽粒状；
(2)以质量比1:3加入蒸馏水，70℃的水浴并搅拌1小时，静置，用双层纱布过滤，漂洗；
(3)重复步骤(2)多次，直到胡麻籽粒表面无粘胶为止，然后将脱胶的胡麻籽粒在40℃烘箱中完全干燥，粉碎，得到脱胶胡麻粉；
(4)取20g脱胶胡麻粉，溶于1600mL的60％的乙醇中，在50℃下，在400W的超声功率下超声提取50分钟；
(5)将步骤(4)获得的混合料液8000×rpm离心5分钟，弃去沉淀，相同条件再离心一次，收集上清；
(6)将步骤(5)得到的上清进行去脂处理，按1:1的体积比加入乙酸乙酯，11000×rpm 离心10分钟，室温静置5分钟，分层，将最下层多酚溶液吸出；吸出的多酚溶液重复去脂处理步骤4次，得到脱脂的多酚溶液；
(5)将步骤(6)得到的脱脂的多酚溶液进行旋转蒸发，然后置于-80℃，冷冻过夜，再置于冷冻干燥机中冷冻2天，得到的浅黄色粉末即为胡麻多酚(如图1所示)。
采用福林酚法，测定胡麻多酚含量，结果发现100g的脱胶胡麻粉中多酚含量约为23.83g。
实施例2
分别选取对数生长期的DLD1，HCT-116，SW480，SW620，HT-29，A549，HeLa, HepG2,MCF-7癌细胞和FHC正常细胞，将细胞制成细胞悬液，通过细胞计数，每孔3×103个接种到96孔板中，37℃、5％CO2培养箱培养24小时待细胞贴壁后，弃掉旧的培养基，加入不同胡麻多酚终浓度(0、0.05、0.1、0.3、0.5、0.7、0.9mg/mL)的培养基，设6 个平行孔，将培养板继续培养24小时，每孔加入20μL 5.0mg/mL MTT，再孵育4小时。移去板中培养液，每孔加入150μL的DMSO溶解活细胞产生的甲瓒结晶，置摇床上振荡10 min，在酶标仪于波长570nm处测定各孔中的吸光度。
本发明所得胡麻多酚，对人多种癌细胞株以及正常细胞进行上述体外抗肿瘤活性检测。结果表明：用不同浓度的胡麻多酚(0、0.05、0.1、0.3、0.5、0.7、0.9mg/mL)处理细胞24小时后，胡麻多酚对DLD1，HCT-116，SW480，SW620，HT-29，A549，HeLa，HepG2， MCF-7癌细胞有明显抑制作用，IC50值分别为0.62mg/mL，0.31mg/mL，0.48mg/mL，0.71 mg/mL，0.53mg/mL，0.36mg/mL，0.76mg/mL，0.69mg/mL，0.9mg/mL。而对正常细胞FHC 没有抑制作用。(如图2,3,4所示)。
Claims (7)
1.一种胡麻多酚的提取方法，其特征在于，包括如下步骤：
(1)取胡麻粕，分离成籽粒状；
(2)按质量比1:3-4加入蒸馏水，65-75℃的水浴并搅拌0.5-1小时，静置，过滤，漂洗；
(3)重复步骤(2)多次，直到胡麻籽粒表面无粘胶为止，然后将脱胶的胡麻籽粒在40℃烘箱中完全干燥，粉碎，得到脱胶胡麻粉；
(4)按质量比1:60-75将脱胶胡麻粉溶于60％的乙醇中，在42-55℃下，在400W的超声功率下超声提取50-60分钟；
(5)将超声提取的混合料液离心，弃去沉淀，上清在相同条件下再离心一次，收集上清；
(6)将步骤(5)得到的上清进行去脂处理，按1:1-1.3的体积比加入乙酸乙酯，离心，室温静置，分层，将最下层多酚溶液吸出；吸出的多酚溶液重复去脂处理步骤3-4次，得到脱脂的多酚溶液；
(7)将步骤(6)得到的脱脂的多酚溶液进行旋转蒸发，冷冻干燥，得到的粉末即为胡麻多酚。
2.如权利要求1所述的胡麻多酚的制备方法，其特征在于，步骤(4)中脱胶胡麻粉和60％的乙醇的质量比为1:65-72。
3.如权利要求1所述的胡麻多酚的制备方法，其特征在于，步骤(4)中提取温度为50℃，在400W的超声功率下超声提取50分钟。
4.如权利要求1所述的胡麻多酚的制备方法，其特征在于，步骤(6)中上清和乙酸乙酯体积比为1:1。
5.如权利要求1所述的胡麻多酚的制备方法，其特征在于，步骤(7)中所述的冷冻干燥的条件：置于-80℃，冷冻过夜，再置于冷冻干燥机中冷冻1-3天。
6.如权利要求1-5任一所述方法制备的胡麻多酚在制备抗肿瘤药物中的应用。
7.如权利要求1-5任一所述方法制备的胡麻多酚在制备保健食品中应用。
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