CN115025139B - 藜麦多酚在调节肝细胞糖脂代谢及抑制氧化应激中的应用 - Google Patents
藜麦多酚在调节肝细胞糖脂代谢及抑制氧化应激中的应用 Download PDFInfo
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Abstract
本发明公开了藜麦多酚在调节肝细胞糖脂代谢及抑制氧化应激中的应用,属于天然产物治疗领域。本发明从藜麦中分离提取了结合态的藜麦多酚,主要含有原儿茶酸、对羟基苯甲酸、香草酸、丁香酸、咖啡酸、对香豆酸、阿魏酸和芥子酸。经实验验证,藜麦多酚能够促进葡萄糖的转运、加速糖代谢,有效降低细胞氧化应激水平,并抑制肝癌细胞的增殖及癌症相关基因的表达。
Description
技术领域
本发明涉及藜麦多酚在调节肝细胞糖脂代谢及抑制氧化应激中的应用,属于天然产物治疗领域。
背景技术
糖、脂代谢作为人体能量供给的主要来源,在生命活动中居于关键地位,但糖脂代谢紊乱会产生高脂血症、糖尿病、脂肪肝、肥胖、动脉硬化性心脑血管病等系列疾病,长期糖、脂、血压水平异常会对全身脏器的损害而导致其功能的逐渐减退,同时引起微血管和大血管损伤,是患者致残、致死的重要原因。
糖、脂代谢性疾病不仅发病率高,而且往往是合病或并病出现,即血糖异常往往伴有血脂异常或高血压等疾病。对血脂异常的调查发现,高脂血症患者合并有糖尿病、高血压等疾病占84.2%。对2.5万多例2型糖尿病(T2DM)患者的调查显示,T2DM患者合并有血脂异常、高血压中一种或两种的占72%。与单纯T2DM患者相比,合并血脂异常、高血压的T2DM患者心血管疾病风险高6倍,说明糖脂代谢紊乱同时存在极大增加血管疾病的发生风险。
氧化应激(Oxidative Stress,OS)是指体内氧化与抗氧化作用失衡的一种状态,倾向于氧化,导致中性粒细胞炎性浸润,蛋白酶分泌增加,产生大量氧化中间产物。氧化应激是由自由基在体内产生的一种负面作用,并被认为是导致衰老和疾病的一个重要因素。氧化应激影响主要的细胞成分:蛋白质、脂质和 DNA。肝细胞的蛋白、脂质和 DNA 是主要受ROS 和 RON 影响。有研究表明,肝病如慢性病毒性肝炎、非酒精性脂肪性肝炎的发生和发展就是由于氧化应激造成肝脏氧化损伤导致的。氧化应激的发生通常还伴随着慢性炎症的产生,这也是肝脏氧化损伤的主要诱因之一,是一种受氧化应激水平影响并受炎症细胞因子 NF-κB 调控的过程。
氧化应激时,会引起脂质过氧化反应,因此,氧化损伤的发生还可以由脂质过氧化产物引起。脂质过氧化产物最终分解为丙二醛(Malondialdehyde, MDA)或者其他醛或酸。ROS 反应和脂质过氧化反应在机体的新陈代谢过程中起着重要的作用,正常情况下两者处于动态平衡状态,维持着体内许多生理生化反应和免疫反应。一旦这种状态发生紊乱和失衡,就会引起一系列的新陈代谢失常和免疫功能降低,形成 ROS 连锁反应,损害生物膜及其功能。肥胖可以增加整个机体内部的氧化应激水平。过度饮食会导致机体内代谢失衡,此时的状态是能量摄入超过能量消耗,细胞氧化应激反应就此发生。氧化应激的发生可以诱导包括炎症在内的很多下游效应的发生。肥胖能促进白色脂肪组织分泌促炎因子,诱发持久的炎症状态。
藜麦(学名:Chenopodium quinoa Willd.)是藜科藜属植物。穗部可呈红、紫、黄,植株形状类似灰灰菜,成熟后穗部类似高粱穗。植株大小受环境及遗传因素影响较大,从0.3-3米不等,茎部质地较硬,可分枝可不分。单叶互生,叶片呈鸭掌状,叶缘分为全缘型与锯齿缘型。藜麦花两性,花序呈伞状、穗状、圆锥状,藜麦种子较小,呈小圆药片状,直径1.5-2毫米,千粒重1.4-3克。原产于南美洲安第斯山脉的哥伦比亚、厄瓜多尔、秘鲁等中高海拔山区。具有一定的耐旱、耐寒、耐盐性,生长范围约为海平面到海拔4500米左右的高原上,最适的高度为海拔3000-4000米的高原或山地地区。藜麦富含的维生素、多酚、类黄酮类、皂苷和植物甾醇类物质具有多种健康功效。藜麦中的优质蛋白和脂肪已被证明在糖脂代谢过程中发挥有益功效,但目前还不清楚藜麦多酚调节肝脏糖脂代谢、氧化应激以及抗癌方面的作用。主要原因在于,酚类物质主要存在于藜麦的麸皮中,因此会随着谷物的精制过程而造成严重浪费,且许多酚类物质结合在多糖及细胞壁上,被称为结合酚,而结合酚难以在人体消化道中释放出来,因此其对人体营养功效的研究还不足。
发明内容
本发明披露了藜麦多酚在调节肝细胞糖脂代谢、抑制肝细胞氧化应激、抑制肝癌细胞增殖中的应用。所述藜麦多酚至少含有原儿茶酸、对羟基苯甲酸、香草酸、丁香酸、咖啡酸、对香豆酸、阿魏酸和芥子酸。所述藜麦多酚的来源可以是:灰藜麦、白藜麦、黑藜麦、红藜麦或藜麦相关制品。
所述调节肝细胞糖脂代谢是指:藜麦多酚可以通过调控PI3K/Akt/GLUT-4信号通路促进葡萄糖的转运,加速糖代谢;还可以上调肝细胞中InSR的表达,增强肝细胞对胰岛素的敏感性。
所述抑制肝细胞氧化应激是指:藜麦多酚具有良好的细胞抗氧化活性,可以有效降低细胞氧化应激水平。
所述抑制肝癌细胞增殖是指:藜麦多酚能够有效抑制肝癌细胞增殖速率,同时有效抑制肝癌细胞癌症相关基因的表达,包括ALDH2基因,p53基因。
本发明披露了藜麦多酚在制备用于调节肝细胞糖脂代谢、抑制肝细胞氧化应激、抑制肝癌细胞增殖的药物。
所述药物,还包括药学上接受的辅料,包括溶剂、抛射剂、增溶剂、助溶剂、乳化剂、着色剂、黏合剂、崩解剂、填充剂、润滑剂、润湿剂、渗透压调节剂、稳定剂、助流剂、矫味剂、防腐剂、助悬剂、包衣材料、芳香剂、抗黏合剂、整合剂、渗透促进剂、pH值调节剂、缓冲剂、增塑剂、表面活性剂、发泡剂、消泡剂、增稠剂、包合剂、保湿剂、吸收剂、稀释剂、絮凝剂与反絮凝剂、助滤剂、释放阻滞剂等。
有益效果:
本发明从藜麦中分离提取了结合态的藜麦多酚,经过鉴定,藜麦多酚主要含有原儿茶酸、对羟基苯甲酸、香草酸、丁香酸、咖啡酸、对香豆酸、阿魏酸和芥子酸。
细胞毒性实验结果表明,藜麦多酚具有较低的细胞毒性。体外抗氧化实验结果表明藜麦多酚具有抗氧化作用,其中,黑藜麦多酚的抗氧化能力最强,其次为灰藜麦多酚和白藜麦多酚;细胞抗氧化实验结果表明,200 μg/mL浓度的藜麦多酚,CAA分别为:黑藜麦多酚151mg QE/ 100 mL,灰藜麦多酚 123mg QE/ 100 mL,白藜麦多酚 98mg QE/ 100 mL。
抗肿瘤细胞增殖活性实验结果表明,HepG2细胞的抑制率为50%时,三种藜麦多酚浓度分别为黑藜麦多酚 72.55 μg/mL,灰藜麦多酚 96.13 μg/mL,白藜麦多酚 102.34 μg/mL。
藜麦多酚通过调控PI3K/Akt/GLUT-4信号通路促进葡萄糖的转运,加速糖代谢;还可以上调InSR的表达,增强胰岛素的敏感性,起到促进脂代谢,降低细胞内总胆固醇的作用。糖脂代谢实验结果表明,相比于对照组,藜麦多酚干预组可以将胞内葡萄糖含量降低6.37%~25.3%,将胞内总胆固醇含量降低 52.34%~66.17%、将胞内总甘油三酯含量降低46.98~63.37%。
附图说明
图1.藜麦总酚含量。
图2.藜麦黄酮含量。
图3.藜麦多酚体外抗氧化。
图4.藜麦酚类标准品色谱图。
图5.白藜麦酚类提取物色谱图。
图6.灰藜麦酚类提取物色谱图。
图7.黑藜麦酚类提取物色谱图。
图8.肿瘤抑制及细胞抗氧化,BQ:黑藜麦,GQ:灰藜麦,WQ:白藜麦。
图9.细胞葡萄糖摄取、总胆固醇含量、总甘油三酯含量。
图10.细胞InSR,PI3K 以及GLUT-4 mRNA 表达量的影响。
图11.细胞转录组测序KEGG分析。
图12.细胞转录组测序DO分析。
具体实施方式
实施例1 藜麦多酚的提取
(1)藜麦多酚提取物的制备
将不同品种藜麦(白、灰和黑)磨粉过60目筛制成藜麦粉,取100g藜麦粉置于2L大烧杯中,用1L正己烷搅拌脱脂4h,然后过滤掉正己烷,重复脱脂三次,脱脂完的藜麦粉置于通风橱内挥发至干燥。脱脂藜麦粉用1L的80%乙醇水溶液搅拌2h以除糖和除脂,然后过滤掉乙醇,重复两次。往脱脂和除糖的藜麦粉中加入600mL的4M NaOH,常温下振荡提取4h,然后用400mL盐酸将其pH调至2以下。用3L乙酸乙酯重复萃取5次,4000r/min离心10min并收集上清液。用旋转蒸发器将乙酸乙酯提取液蒸干,用300mL纯水复溶。然后将水提液分装至2个平皿,置于冷冻干燥机中冷冻干燥3天,对干燥后的藜麦多酚提取物(只含藜麦多酚)称量并置于-20℃冰箱保存。
(2)藜麦多酚提取物的定性及定量
取2mg藜麦多酚提取物并溶于2mL的色谱级无水甲醇中,过0.22μm滤膜,进行高效液相色谱分析。采用岛津APD-20A高效液相色谱分析,色谱柱为C18柱(4.6×250mm,5μm),流动相A为含0.1%乙酸的水溶液,流动相B为含0.1%乙酸的甲醇溶液。初始流速为1mL/min,进样体积5μL,检测波长为280nm,色谱进样程序为:0~11min,9%~14%B相;11~14min,14%~15%B相;14~17min,15%~15%B相;17~24min,15%~16.5%B相;24~28min,16.5%~19%B相;28~30min,19%~25%B相;30~36min,25%~26%B相;36~38min,26%~28%B相;38~41min,28%~35%B相;41~46min,35%~40%B相;46~48min,40%~48%B相;48~53min,48%~53%B相;53~71min,53%~70%B相;71~80min,9%B相。藜麦酚类标准品、白藜麦酚类提取物、灰藜麦酚类提取物、黑藜麦酚类提取物的色谱图如图4~7所示。
如图1所示,不同品种的藜麦在多酚含量方面有显著差异,由低到高依次为白藜麦、灰藜麦、黑藜麦。
如图2所示,不同品种的藜麦在总黄酮含量方面也有显著差异,由低到高依次为白藜麦、灰藜麦、黑藜麦,整体趋势与总酚含量相同。黄酮是酚类物质的一种,黑藜麦中也富含花色素类物质,它们大多是黄酮类衍生物。
如表1所示,三种藜麦的多酚中均鉴定出了8种酚酸类物质,包括4种羟基苯甲酸(原儿茶酸、对羟基苯甲酸、香草酸和丁香酸)和4种羟基肉桂酸(咖啡酸、对香豆酸、阿魏酸和芥子酸)。8种酚类物质的保留时间分别为:原儿茶酸:14.85 min;对羟基苯甲酸:24.18min;香草酸:33.23 min;咖啡酸35.15min;丁香酸:38.61 min;对香豆酸:45.82 min;阿魏酸:48.92 min;芥子酸:50.01 min。酚酸具有很强的抗氧化能力,是很好的生理活性物质,小麦、大麦和玉米等谷物中的阿魏酸占比较高,而其他酚酸的占比很低。相比其它谷物,藜麦的酚酸组成更加均匀,白藜麦和灰藜麦的对香豆酸含量较高,黑藜麦中各个酚酸含量均很高,其中,对羟基苯甲酸含量极高,远高于其他谷物中的对羟基苯甲酸含量。三种藜麦中,黑藜麦的酚酸总量最高,其次是灰藜麦和白藜麦,具体含量如下表1。
表1 藜麦多酚组成及含量(μg/g)
实施例2 藜麦多酚调节糖脂代谢、抑制氧化应激、抑制癌细胞增殖作用的验证
本实施例所用藜麦多酚提取物的制备方法同实施例1。
本实施例所用完全培养基:William’s Medium E (WME),含5% FBS、10 mM Hepes、2 mM L-谷氨酰胺、5 μg/mL胰岛素、0.05 μg/mL氢化可的松、50 units/mL盘尼西林、50 μg/mL链霉素和100 μg/mL庆大霉素。
本实施例所用氧化培养基:Hanks’平衡盐溶液(不含酚红),含10 mM Hepes。
本实施例所用20 mM DCFH-DA储备液:0.0390 g DCFH-DA用甲醇溶解并定容至4mL。本实施例所用200 mM ABAP储备液,0.5423g ABAP,用去离子水溶解并定容至10 mL。
本实施例所用细胞培养、传代及计数方法:采用HepG2细胞模型,用完全培养基培养(37℃,5% CO2)细胞,待细胞完全覆盖T-75培养瓶后,向瓶中加入2 mL胰蛋白酶,2-3 min后用手轻轻敲打瓶边缘,待60%左右细胞脱落时,用冲洗培养基(6-8 mL/瓶)冲洗细胞,并将细胞悬浮液转移至离心管中,4℃离心(134 g)4 min;移去上层液,用手指轻弹离心管壁使细胞混匀,并向其中加入适当量的完全培养基,然后用台盼蓝染色液对细胞进行染色,之后在显微镜下用血球计数板对细胞进行计数,并按实验要求用于铺板或其他细胞实验。
(1)藜麦多酚提取物对细胞抗氧化活性的影响
体外抗氧化活性采用DPPH自由基清除试验来测定。称取DPPH粉8mg,用甲醇定容至250mL。取10mg多酚提取物,用10mL甲醇溶解。吸取100μL多酚提取物的甲醇溶液,加入3.9mL的DPPH溶液,在室温下避光孵育30min后在517nm波长下测定混合物的吸光度。根据公式计算DPPH自由基清除率:DPPH自由基清除活性(%)=1-(Ax-A1)/A0×100,Ax为样品吸光度,A1为阴性对照(0.1mL样品+3.9mL甲醇)吸光度,A0为空白对照吸光度,结果以每克样品的mgTrolox(TE)/100g当量表示。
结果如图3所示,不同品种藜麦多酚的抗氧化能力有显著差异,由低到高依次为白藜麦多酚、灰藜麦多酚、黑藜麦多酚。
细胞抗氧化活性采用CAA实验来检测。CAA实验采用12至35代的细胞(HepG2),以6.0×104个/孔的密度接种在96孔黑板中,培养24 h后,移去培养基并向每个孔中加入100μL含不同浓度的藜麦多酚提取物或标准品的氧化培养基, 100μL氧化培养基中还含有50 μM DCFH-DA,之后将细胞在37℃,5% CO2条件下培养1 h,培养结束后移去培养基,一部分细胞用PBS冲洗,另一部分细胞不做PBS冲洗处理,随后向细胞中加入100 μL、600 μM ABAP(用氧化处理培养基稀释储备液),之后迅速在酶标仪(37℃)上检测96孔黑板中各反应液在激发波长485 nm,发射波长535 nm 条件下的荧光强度,每隔5 min记录1次,共记录1 h。以时间为横坐标,荧光强度为纵坐标绘制出各样品和标准品的荧光强度随时间变化的曲线,通过样品(或标准品)曲线下面积计算样品(标准品)的 EC50值。CAA实验以槲皮素(quercetin)作为标准品,通过标准品EC50与样品EC50来计算样品的CAA值,最终结果表述为μmolquercetin equivalents (QE)/100 g(DW)。
如图8所示,抗氧化强度由低到高依次为白藜麦多酚、灰藜麦多酚、黑藜麦多酚,整体趋势与总酚和总黄酮趋势相同,因为酚类物质具有较强的抗氧化能力,是抗氧化作用的主要贡献者。我们利用HepG2细胞为模型,测定了三种藜麦提取物的细胞干预浓度,而细胞抗氧化的结果也与体外抗氧化结果相一致,结果表明黑藜麦多酚具有更好的抗氧化效果,能有效缓解肝细胞氧化应激。
(2)藜麦多酚提取物抗肿瘤细胞增殖活性和细胞毒性分析
对藜麦多酚提取物的抗肿瘤细胞增殖活性和细胞毒性进行分析比较,细胞培养方法与CAA法相似。主要步骤包括:a.将培养适宜的HepG2细胞以1.5×104cells/孔的浓度接种于96孔白色孔板上进行抗增殖活性分析,细胞毒性分析采用的细胞浓度则为2.5×104cells/孔;b.两者于37℃,5% CO2培养箱中分别孵育4 h和24 h;c.弃去培养基,用PBS洗涤孔板中的细胞;d. 然后向细胞孔中加入100 μL一系列浓度的藜麦多酚(溶解在完全培养基中),并以96孔板中与样品孔处在同一垂直位置且以100 μL完全培养基替代含藜麦多酚的完全培养基的孔作为该样品的对照,将细胞在37℃,5% CO2条件下分别培养72 h和24 h;e. 培养结束后移去培养基,用PBS冲洗细胞,然后向每个孔中加入50 μL亚甲基蓝染色液(HBSS +1.25%戊二醛+ 0.6%亚甲基蓝)并孵育1 h。孵育结束后,移去染色液并用清水清洗96孔板6次至水清澈,并在室温条件下使96孔板中的水分挥干,之后向每个细胞孔中加入100 μL 洗脱缓冲液(49% PBS + 50%乙醇+1%乙酸,v/v)并将96孔板置于振荡器中振荡20min,以使染色液溶解并混匀。在57 nm下检测各孔中反应液的吸光度,通过比较样品孔和对应对照孔的吸光度来计算不同浓度样品对细胞的抑制率,以样品浓度为横坐标,抑制率为纵坐标绘制样品对细胞的抑制率随样品浓度的变化曲线。如果某浓度的样品对细胞的抑制率(相对于对照)≥10%,则认为此样品在大于和等于该浓度时具有细胞毒性,最终结果以样品对细胞抑制率50%时的浓度表示(μg /mL)。样品的抗肿瘤细胞增殖活性和细胞毒性分析分别以EC50(mg/mL)和CC50(mg/mL)表示。
结果如表2所示。三种藜麦多酚的EC50值分别为黑藜麦多酚72.55±1.32 mg/mL,灰藜麦多酚96.13±2.11 mg/mL,白藜麦多酚102.34±1.77 mg/mL。其中,除灰藜麦多酚CC50大于800 mg/mL,其余均大于900 mg/mL。结果表明黑藜麦多酚具有更好的抗肿瘤活性与较低的细胞毒性。
表2 抗肿瘤增值及细胞毒性
(3)肝脏糖脂代谢实验
利用HepG2细胞构建肝脏糖脂代谢模型,实验分为对照组(Con):25 mmol /L 葡萄糖+ 10 g /L(质量浓度) BSA,模型组(Mod):20 mmol /L 葡萄糖 + 15mmol/L 果糖 + 1mmol /L 油酸钠(OANa) + 1 mmol /L 棕榈酸钠(PANa) +10 g /L BSA,以及藜麦多酚干预组,根据之前藜麦多酚肿瘤抑制及细胞毒性实验结果,我们选择200 μg/mL的浓度进行干预,即,在模型组(Mod)的基础上添加200 μg/mL藜麦多酚,干预组又分为黑藜麦多酚组(BQ)、灰藜麦多酚组(GQ)以及白藜麦多酚干预组(WQ)。各组共同孵育24 h小心弃去培养液,然后准备测定胞内葡萄糖含量等。
测定胞内葡萄糖含量:将细胞用PBS洗1次,换为葡萄糖浓度为12.5 mmol/L DMEM培养基,培养24 h 后取5 μL上清液,用葡萄糖试剂盒测定葡萄糖胞内含量。
测定胞内糖原含量:将细胞用PBS洗1次,细胞用RIPA裂解液裂解,用糖原试剂盒测定胞内糖原含量。
测定细胞TG、TC含量:将细胞用刮刀刮下后与培养基一同混匀,并超声破碎1 min,然后用TG试剂盒测定TG含量,用TC试剂盒测定TC含量。
如图9所示,细胞葡萄糖含量的测定结果显示,模型组细胞上清葡萄糖含量极显著高于对照组。不同藜麦中的多酚(BQ、GQ和WQ)与造模液共处理24h后,与模型组相比,WQ组细胞上清葡萄糖水平显著降低,其他各实验组细胞上清葡萄糖水平极显著降低。结果表明,油酸钠以及棕榈酸钠诱导后,HepG2 细胞吸收葡萄糖的能力下降,三种藜麦提取物可促进细胞摄取葡萄糖,从而改善细胞的胰岛素抵抗水平。此外模型组细胞内总胆固醇、总甘油三酯含量显著上升,脂代谢出现异常,而藜麦多酚具有促进脂代谢,降低细胞内总胆固醇的作用。
(4)藜麦多酚提取物对InSR,PI3K 以及 GLUT-4 mRNA 的表达
利用HepG2细胞构建肝脏糖脂代谢模型,实验分为对照组(Con):25 mmol /L 葡萄糖+ 10 g /L(质量浓度) BSA,模型组(Mod):20 mmol /L 葡萄糖 + 15mmol/L 果糖 + 1mmol /L 油酸钠(OANa) + 1 mmol /L 棕榈酸钠(PANa) +10 g /L BSA,以及藜麦多酚干预组,根据之前藜麦多酚肿瘤抑制及细胞毒性实验结果,我们选择200 μg/mL的浓度进行干预,即,在模型组(Mod)的基础上添加200 μg/mL藜麦多酚,干预组又分为黑藜麦多酚组(BQ)、灰藜麦多酚组(GQ)以及白藜麦多酚干预组(WQ)。各组共同孵育24 h小心弃去培养液。
采用TransZol Up Plus RNA Kit试剂盒提取各组HepG2细胞中的总RNA,检测其浓度和纯度。用TransScriptR Green One-Step qPCR SuperMix 试剂盒检测 InSR,PI3K以及GLUT-4 mRNA的表达,以GAPDH为内参。各引物序列如下表3。
结果如图10所示,与对照组相比,模型组细胞中InSR,PI3K 以及 GLUT-4 mRNA表达量显著下降(P<0.05),FFAs 会导致胰岛素受体InSR的丝氨酸/苏氨酸磷酸化,从而降低活化PI3K的能力,并削弱下游胰岛素受体 GLUT-4的信号传导。说明藜麦提取物可以通过调控PI3K/Akt/GLUT-4信号通路促进葡萄糖的转运,加速糖代谢;还可以上调InSR的表达,增强胰岛素的敏感性。
表3 引物序列
(5) 细胞转录组测序分析
将上述步骤(4)中获取的RNA,通过干冰寄送至专业公司,进行转录组测序分析。实验结果利用KEGG及DO数据库进行分析。
KEGG是由日本京都大学和东京大学联合开发的数据库,是基因组测序和其他高通量实验技术生成的大规模分子数据集的整合和解读的突出参考知识库,可以用来查询代谢途径、酶(或编码酶的基因)、产物等,也可以通过BLAST比对查询未知序列的代谢途径信息。KEGG PATHWAY数据库是一个代谢通路集合的数据库,包含以下几方面的分子间相互作用和反应网络:(1)新陈代谢(Metabolism)、(2)遗传信息加工(Genetic InformationProcessing)、(3)环境信息加工(Environmental Information Processing)、(4)细胞过程方面(Cellular Processes)、(5)生物体系统方面(Organismal Systems)、(6)人类疾病方面(Human Diseases)、(7)药物开发方面(Drug Development)。
如图11所示,三种藜麦多酚均能对细胞新陈代谢细胞过程、有机系统、人类疾病方面基因产生影响,特别是人类疾病与代谢方面。
DO数据库是描述人类基因功能与疾病相关的数据库,通过参照MeSH,ICD等疾病分类标准,对人类的常见疾病与罕见病进行了归纳整理,提供了一个统一的,标准化的疾病分类系统。利用DO数据库,可以对基因和基因产物按解剖实体疾病、细胞增殖疾病、心理健康疾病、代谢疾病、遗传性疾病、精神疾病、综合征这七个方面进行分类注释。
如图12所示,三种藜麦多酚均能有效改善人体疾病相关基因,特别是癌症、氧化应激、糖脂代谢相关基因表达。
虽然本发明已以较佳实施例公开如上,但其并非用以限定本发明,任何熟悉此技术的人,在不脱离本发明的精神和范围内,都可做各种的改动与修饰,因此本发明的保护范围应该以权利要求书所界定的为准。
Claims (3)
1.藜麦多酚作为唯一活性成分在制备用于调节肝细胞糖脂代谢的产品中的应用,其特征在于:
所述藜麦多酚的制备方法:
将藜麦磨粉过60目筛制成藜麦粉,取100g藜麦粉置于2L大烧杯中,用1L正己烷搅拌脱脂4h,然后过滤掉正己烷,重复脱脂三次,脱脂完的藜麦粉置于通风橱内挥发至干燥,脱脂藜麦粉用1L的80%乙醇水溶液搅拌2h以除糖和除脂,然后过滤掉乙醇,重复两次,往脱脂和除糖的藜麦粉中加入600mL的4M NaOH,常温下振荡提取4h,然后用400mL盐酸将其pH调至2以下,用3L乙酸乙酯重复萃取5次,4000r/min离心10min并收集上清液,用旋转蒸发器将乙酸乙酯提取液蒸干,用300mL纯水复溶,然后将水提液分装至2个平皿,置于冷冻干燥机中冷冻干燥3天,对干燥后的藜麦多酚提取物称量并置于-20℃冰箱保存;
所述藜麦多酚至少含有原儿茶酸、对羟基苯甲酸、香草酸、丁香酸、咖啡酸、对香豆酸、阿魏酸和芥子酸。
2.根据权利要求1所述的应用,其特征在于,所述藜麦多酚的来源是:灰藜麦、白藜麦、黑藜麦、红藜麦。
3.根据权利要求1或2所述的应用,其特征在于,所述产品包括药物。
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