CN114949023A - 显脉旋覆花提取物的新用途 - Google Patents
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Abstract
本发明公开显脉旋覆花提取物的新用途,属于药物、天然药物技术领域。该应用是显脉旋覆花提取物在制备抗酒精性肝病药物、保肝护肝功能食品或保健品中的应用。所述提取物的制备方法如下:将显脉旋覆花的根茎和/或花瓣粉碎至0.1~50mm,按花瓣和/或根茎与水溶液的重量体积比为1:1~50的比例加入水,进行超声搅拌提取,离心收集上清液,浓缩得到显脉旋覆花水提取物,通过冷冻干燥得到显脉旋覆花提取物。本发明所提供的显脉旋覆花提取物对酒精性肝病具有明显的抑制肝脏脂肪变性及肝细胞损伤的作用。
Description
技术领域
本发明涉及显脉旋覆花提取物的新用途,属于药物、天然药物技术领域。
背景技术
自古以来,中国的酒文化源远流长,早已成为生活中不可或缺的一部分,大多数人普遍认为,少量饮酒有益健康,过量饮酒才是导致死亡的主要因素。40%的酒精滥用者会发展为酒精性肝炎,30%至 60%的酒精性肝炎患者会导致严重的并发症,例如肝衰竭和门静脉高压,短期死亡率非常高,并且大约 40%的酒精性肝炎患者发展为坏死性炎症和纤维化。目前酒精性肝病的发病机制较为复杂,可能是由于细胞损伤、氧化应激、菌群失调等多种驱动因素交叉作用所致。近年来,对于酒精性肝病的营养治疗、药物治疗、肝移植治疗等研究手段虽取得一定进展,但对于终末期的酒精肝患者而言,有效的治疗方法仍然少之又少。因此,酒精性脂肪肝作为酒精性肝病的早期阶段,也是酒精性肝病的发生发展过程中的可逆阶段,是治疗和缓解酒精性肝病的重要阶段。
目前酒精性肝病尚无有效的治疗药物,酒精性肝病治疗的第一步是停止酒精使用,酒精摄入严重影响大量和微量营养元素的代谢,降低了氨基酸的吸收,降低了肝脏中蛋白质(脂蛋白,白蛋白和纤维蛋白原)的合成和分泌。在不可避免的情况下,使用皮质类固醇(Corticosteroids)能够短期治疗严重的酒精性肝炎。然而,这些药物有明显的副作用,这刺激了对治疗酒精性肝病安全有效药物的需求。从我国传统药食同源资源中寻找安全和对酒精性肝病具有良好干预作用的物质是一个重要的方向。
显脉旋覆花,俗称小黑药,又名云威灵、威灵草等,主要分布云南北部、四川西南部、贵州北部和广西西部,是云南贵州等地常用的民族药,它主要用于治疗咳嗽,跌打损伤、风湿骨痛、消化不良,神经衰弱等症状。2010年,显脉旋覆花被***批准作为新食品原料。有研究表明,显脉旋覆花提取物具有良好的抗氧化的作用。但是目前尚未有关于显脉旋覆花缓解酒精性肝病的报道。
发明内容
本发明的目的是提供显脉旋覆花提取物在制备抗酒精性肝病药物中的应用。
或者显脉旋覆花提取物在制备保肝护肝功能食品或保健品中的应用。
优选的,本发明所述显脉旋覆花提取物为水提物。
优选的,本发明所述显脉旋覆花提取物的制备方法具体包括以下步骤:将显脉旋覆花的根茎和/或花瓣粉碎至0.1~50mm,按花瓣和/或根茎与水溶液的重量体积比为1: 1~50的比例加入水,进行超声搅拌提取,离心收集上清液,浓缩得到显脉旋覆花水提取物,通过冷冻干燥得到显脉旋覆花提取物。
优选的,本发明所述超声提取的温度为50~80℃,功率为200W;搅拌速率为2000-5000rpm /min。
本发明的有益效果:
(1)本发明首次探索研究显脉旋覆花提取物在抗酒精性肝病药物上的应用。
(2)试验证明,本发明的显脉旋覆花提取物对长期酒精摄入和短期大量酒精摄入造成的肝脏脂肪变性与肝细胞损伤具有一定程度的缓解作用,随着提取物作用浓度的增加,对酒精性肝损伤的缓解作用呈现出明显的剂量-时间依赖效应,组织病理学切片结果表明显脉旋覆花提取物可以减少酒精摄入导致的肝细胞变性和炎性细胞因子浸润的情况;并且显脉旋覆花提取物还可以显著减轻的肝脏毒性的水平;因此本发明对于制备抗酒精性肝病药物具有重要意义。
附图说明
图1为显脉旋覆花提取物对小鼠的肝脏中甘油三酯和总胆固醇含量的调节情况;
图2为显脉旋覆花提取物对小鼠的肝脏组织H&E染色切片的影响;
图3为显脉旋覆花提取物对小鼠血浆中肝毒性物质含量的调节情况;
图4为显脉旋覆花提取物对大鼠的肝脏中甘油三酯和总胆固醇含量的调节情况;
图5为显脉旋覆花提取物对大鼠的肝脏组织H&E染色切片的影响;
图6为显脉旋覆花提取物对大鼠血浆中肝毒性物质含量的调节情况。
具体实施方式
下面结合附图和具体实施方式对本发明作进一步详细说明,但本发明的保护范围并不限于所述内容。
实施例1
显脉旋覆花提取物的制备,具体包括以下步骤:
(1)对采集得到的全株显脉旋覆花进行风干,得到干燥的显脉旋覆花植250g。将显脉旋覆花植粉碎至50mm后与蒸馏水混合(1:1)超声搅拌提取3 h,超声提取的温度为50℃,功率为200W,搅拌速率 2000rpm /min,然后将超声提取液过滤得到滤液,再以30℃旋转蒸发浓度后,冷冻干燥得到显脉旋覆花水提取物19.46 g。
实施例2
(1)对采集得到的全株显脉旋覆花进行风干,得到干燥的显脉旋覆花植250g,将显脉旋覆花植粉碎至500mm后与蒸馏水混合(1:50)超声搅拌提取3 h,超声提取的温度为80℃,功率为200W,搅拌速率 5000rpm /min,然后将超声提取液过滤得到滤液,再以50℃旋转蒸发浓度,冷冻干燥得到显脉旋覆花水提取物26.32 g。
验证实验1
下面通过动物实验进一步验证显脉旋覆花提取物在缓解急性酒精性肝病中的应用效果。
1、分组与建模
将40只雄性(6周龄,20±2g) C57BL/6小鼠随机分为正常组,模型组,干预组1(实施例1所得提取物)、干预组2(实施例2所得提取物),每组十只。小鼠适应性喂养一周后,IRL、IRH组分别灌胃对应的提取物。在此期间对照组使用等量生理盐水灌胃,持续三周。在喂养期间,每两天记录小鼠体重,并调整灌胃剂量。最后一次灌胃12 h后,对照给予蒸馏水12 mL/kg·体重,其余各组灌胃50%乙醇溶液12 mL/kg·体重。最后 1 次酒精灌胃后禁食12小时后处死, 使用含有抗凝剂的离心管收集血液并分离出小鼠肝脏。
2、肝脏甘油三酯与总胆固醇含量测定
将摘取的小鼠肝脏用生理盐水漂洗,并用滤纸吸干水分后,切取部分组织,进行匀浆处理后,使用试剂盒测定甘油三酯(TG)和总胆固醇(TC)含量;结果如图1所示,相比正常组,ALD组小鼠肝脏组织中TC和TG含量明显增加;显脉旋覆花提取物给药后,小鼠肝脏组织中的TC和TG含量明显降低;说明脉显脉旋覆花提取物处理缓解了急性酒精摄入导致的肝脏脂肪变性。
3、肝脏组织病理学情况
对收集的肝脏组织进行病理组织形态学观察。结果如图2所示, ALD组小鼠肝细胞有较多脂肪滴的出现并伴有炎性细胞浸润;显脉旋覆花提取物干预后,小鼠肝脏组织中未见到明显的脂肪滴和炎性细胞浸润的情况,小鼠急性酒精摄入导致的肝脏脂肪堆积和肝细胞损伤情况得到明显改善。
4、血清生化指标水平检测
将采集的血液样品经过低速离心机3500g离心15min得到血浆,血浆中AST、ALT水平;结果如图3所示,ALD组小鼠血清中AST和ALT水平明显高于对照组;给予显脉旋覆花提取物后,血清中AST和ALT水平明显降低。
5、肝脏生化指标水平检测
取部分小鼠肝脏组织,制备10%的肝脏组织匀浆。用试剂盒测定肝脏中IL-6和TNF-α水平;结果如表1所示,ALD组小鼠肝脏组织中炎症因子IL-6和TNF-α水平与对照组相比显著升高;给予两种显脉旋覆花提取物后小鼠肝脏组织中炎症因子IL-6、 TNF-α水平与ALD组大鼠相比显著降低,并且高剂量组效果显著优于低剂量组。
表1. 显脉旋覆花提取物对酒精性肝损伤小鼠肝脏组织炎症因子的影响
验证实验2
下面通过动物实验进一步验证显脉旋覆花提取物在缓解慢性酒精性脂肪肝中的应用效果。
1、分组与建模
将40只雄性(6周龄,200±10g) SD大鼠随机分为正常组,模型组,干预组1(实施例1所得提取物)、干预组2(实施例2所得提取物),每组十只;除正常组大鼠每天供矿泉水自由饮用外, 第一周每天灌胃 20 % 酒精1次,灌胃体积为5ml/kg,余下时间全部使用56%的酒精进行灌胃。在灌胃完酒精后,实验组分别灌胃对应的提取物,灌胃体积为5ml/kg;在此期间正常组使用对应体积的蒸馏水进行灌胃,持续12周;造模期间,每天的早中晚三次观察大鼠精神行为状态是否正常并及时做好记录,每3天对大鼠进行称重并记录;最后 1 次灌胃后禁食 12小时后处死, 使用含有抗凝剂的离心管收集血液并分离出大鼠肝脏。
2、肝脏甘油三酯与总胆固醇含量测定
将摘取的大鼠肝脏制备为10%的肝脏组织匀浆后,使用试剂盒测定甘油三酯(TG)和总胆固醇(TC)含量;结果如图4所示,相比正常组,AFLD组大鼠肝脏组织中TC和TG含量明显增加;显脉旋覆花提取物给药后,小鼠肝脏组织中的TC和TG含量明显降低;此外IWH组大鼠的肝脏组织中TC和TG含量相比IWL组均显著降低。说明脉显脉旋覆花提取物处理缓解了急性酒精摄入导致的肝脏脂肪变性,并随着添加剂量的提高,缓解效果也显著增加。
3、肝脏组织病理学情况
对收集的肝脏组织进行病理组织形态学观察;结果如图5所示,AFLD组大鼠肝脏组织脂肪变性情况严重,出现较大的脂肪滴;显脉旋覆花提取物干预后,大鼠肝脏组织中脂肪变性和炎性细胞浸润的情况得到明显减轻。
4、血清生化指标水平检测
将采集的血液样品经过低速离心机3500g离心15min得到血浆,血浆中AST、ALT水平;血浆中AST、ALT水平反应了肝毒性情况,结果如图6所示,AFLD组大鼠血清中AST和ALT水平明显高于对照组;给予显脉旋覆花提取物后,血清中AST和ALT水平明显降低。
5、肝脏生化指标水平检测
取部分大鼠肝脏组织,制备10%的肝脏组织匀浆。用试剂盒测定肝脏中IL-6和TNF-α水平;结果如表2所示,AFLD组大鼠肝脏组织中炎症因子IL-6和TNF-α水平与对照组相比显著升高;给予两种不同的显脉旋覆花提取物后,大鼠肝脏组织中炎症因子IL-6、 TNF-α水平与ALD组大鼠相比显著降低,且高剂量组效果显著优于低剂量组。
表2. 显脉旋覆花提取物对酒精性脂肪肝大鼠肝脏组织炎症因子的影响
Claims (5)
1.显脉旋覆花提取物在制备抗酒精性肝病药物中的应用。
2.显脉旋覆花提取物在制备保肝护肝功能食品或保健品中的应用。
3.根据权利要求1或2所述应用,其特征在于:所述显脉旋覆花提取物为水提物。
4.根据权利要求3所述应用,其特征在于:所述显脉旋覆花提取物的制备方法具体包括以下步骤:将显脉旋覆花的根茎和/或花瓣粉碎至0.1~50mm,按花瓣和/或根茎与水溶液的重量体积比为1: 1~50的比例加入水,进行超声搅拌提取,离心收集上清液,浓缩得到显脉旋覆花水提取物,通过冷冻干燥得到显脉旋覆花提取物。
5.根据权利要求4所述应用,其特征在于:所述超声提取的温度为50~80℃,功率为200W;搅拌速率为2000-5000rpm /min。
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