CN101406537A - 柔毛水杨梅提取物用于制备治疗肥胖的药物的用途 - Google Patents
柔毛水杨梅提取物用于制备治疗肥胖的药物的用途 Download PDFInfo
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Abstract
本发明涉及柔毛水杨梅植物提取物用于制备治疗肥胖的药物的用途。柔毛水杨梅植物提取物为以活性化合物Gemin A为标记物的提取物,其中可水解鞣质的含量大于 30%。柔毛水杨梅植物提取物在体外显示了很强的抗脂肪酸合酶活性,在动物实验中也表现出了很好的减肥作用,可用于制备治疗肥胖的药物。
Description
技术领域:
本发明涉及医药、食品技术领域,特别是涉及显示具有减肥活性的植物药用提取物,更特定地涉及柔毛水杨梅提取物用于制备治疗肥胖的药物的应用。
背景技术:
近年来,随着人类生存年龄的提高,饮食生活的改变,运动不足及生活环境的变化引起的精神压力和应激反应的增加,致使患肥胖、癌症及慢性、老年性疾病的人数大幅增多,已成为一种迫切丞待解决的问题。
肥胖是一种复杂的,多因素决定的代谢失调症,如基因、调节能量平衡的激素、环境、生活方式以及饮食都是可能引起肥胖的因素。但从根本上来说,肥胖源于食物摄入与能量消耗之间的平衡的破坏。脂肪酸合酶(FAS)是人类和动物体内脂肪合成的关键酶,通过抑制脂肪酸酶的活性可以有效预防和减轻肥胖症状。2000年J.Hopkins大学的Loftus等通过给小鼠腹膜内注射FAS抑制剂C75,观测到小鼠体重显著减轻。这种体重的减轻呈剂量依赖性,并随剂量的增加而延长作用的持续时间。小鼠停药后随药物作用的消失均能恢复失去的体重,而不会持续消瘦。这种药物对小鼠来说有良好的耐受性,且未见明显的毒性通常,饥饿后机体的正常反应是能量消耗的减少,而C75的作用则不同:通过C75给药和饥饿疗法的比较表明,C75给药尽管导致摄食量的减少,但是并不影响机体正常的能量代谢。抑制FAS的活性,既能够阻滞生脂通路,减少脂肪的合成,又能够造成丙二酰辅酶A浓度的升高,抑制中枢系统调控饮食的神经肽从而达到降低食欲的目的,具有一举两得的作用。因此FAS很可能是体内调节能量消耗和储存的重要位点之一,也是一个很好的调节靶位。合适的FAS抑制剂既能调节脂肪的代谢和储存,又能降低食欲减少食物摄入,特别是其作用并不依赖于特定的基因缺陷,使得此项研究具有广泛而深远的意义,已引起极大关注。我国学者田维熙教授等人报道绿茶中的成分-表没食子儿茶素没食子酸酯(EGCG)是有效的FAS抑制剂,它和C75有不同的抑制表现及作用部位,研究表明绿茶和中药何首乌都具有抑制脂肪酸合酶和减肥的作用。(李丽春等,中国生物化学与分子生物学报,200319(3):297~304;张睿等,云南大学学报,2004,26(6A),42-47。).另有报道EGCG能减轻小鼠食欲和体重(Kao JH.,Endocrinology,2000,141(3):980~987)。大量的科学研究已经证实了脂肪酸合酶抑制剂在治疗和预防肥胖方面的潜在应用。
柔毛水杨梅(Geum japonicum Thunb.var.chinense F.Bolle)是蔷薇科路边青属植物,药用全草或根。在贵州少数民族地区人们叫它蓝布正,并广泛用来治疗头晕目眩,头痛等症。《中药大辞典》记载其具有补虚益气,祛风除湿,消肿散瘀的功效。治腹泻,痢疾,白带,崩漏,风湿腰腿痛,跌打损伤,痈肿疮疡。对柔毛水杨梅的研究报道较少,初步研究表明其提取物富含多酚(唐正平等,湖南中医药导报,2000,6(4):52-55)。香港中文大学李明等人申请了柔毛水杨梅提取物诱导新生血管形成和肌肉再生的用途专利(WO2004052381;WO2003043645)。我们在前期研究中从其甲醇提取物中分离纯化了包括Gemin A在内的几个多酚类化合物(刘宏伟等,沈阳药科大学学报,2006,23(11):694-697.)。在我们的研究中,首次发现了柔毛水杨梅提取物具有很好的抗脂肪酸合酶活性和减肥的作用。
发明内容:
本发明提供了可从柔毛水杨梅的植物全草获得的植物提取物在生产治疗和预防减肥药物或膳食补充剂中的用途。
柔毛水杨梅提取物可选用水及甲醇、乙醇、丙醇、丁醇等有机溶剂或上述任何两种或多种互溶的混合溶剂,在室温下浸渍提取或超声提取或加热回流提取获得,提取次数可为一次或多次,减压干燥,称重。提取物分散到适量的水中,然后利用等量氯仿、正己烷、环己烷或石油醚等有机溶剂中的任何一种萃取除去脂溶性杂质,再利用等量正丁醇萃取获得活性提取物。
参照2005版中国药典的方法(皮粉法)测定了柔毛水杨梅正丁醇提取物的鞣质含量,含量大于30%。利用高效液相的方法表征了提取物的指纹图谱(C18(RP)分析型色谱柱;流动相:A 0.1%醋酸-水溶液;B甲醇;0-10min,10%B;20-50min,10%B-30%B;50-70min,30%B-100%B),流速为1.0mL/min,可监测到10-16个色谱峰,活性化合物Gemin A的保留时间是31分钟。
本发明的优点是:我们采用体外抗脂肪酸合酶活性检测方法,对柔毛水杨梅正丁醇提取物的生物活性进行了研究,证实了柔毛水杨梅正丁醇提取物具有很的强抗脂肪酸合酶活性,并通过体内试验证实了柔毛水杨梅正丁醇提取物的减肥作用。柔毛水杨梅正丁醇提取物,配以适当的赋形剂,可制备治疗和预防肥胖的各种药物剂型或膳食补充剂。
附图说明:
图1为柔毛水杨梅正丁醇提取物的HPLC分析图谱。
具体实施方式:
实施例1:柔毛水杨梅正丁醇提取物的制备:取柔毛水杨梅干燥药材100克,加入500毫升甲醇室温浸提48小时,提取3次,过滤合并提取液,40摄氏度喷雾干燥获得柔毛水杨梅提取物9.3克,收率9.3%。将甲醇提取物分散到100毫升的水中,利用等体积的氯仿萃取3次,然后用等体积正丁醇萃取3次,合并正丁醇萃取部分,减压干燥得到正丁醇提取物3克。
实施例2:柔毛水杨梅正丁醇提取物可水解鞣质含量测定:精密称取实施例1正丁醇提取物1.0克,置于100毫升容量瓶,加水溶解,并定容至100毫升作为供试液。精密吸取1mol/L醋酸锌20mL置500mL容量瓶中,加入氨水14mL,摇匀,使生成的白色沉淀溶解,精密加入供试品溶液30mL,振摇1min,置(35±2)℃水浴温热30min,不断振摇。取出,冷却至室温,加蒸馏水至刻度,摇匀,滤过,精密吸取续滤液50mL置500mL锥形瓶中,分别加入蒸馏水300mL、氨-氯化氨缓冲液25mL和铬黑T指示剂1mL,摇匀,用EDTA(010508mol/L)缓缓滴定,至紫红色转变为蓝绿色,结果用空白校正。按公式计算鞣质含量,鞣质含量(%)=011556×[20×CZn 2+-10(V空-V样)CEDTA]/V×100%。其中0.155 6为1mol·L-1Zn2+标准溶液消耗量与药材中鞣质含量的比例常数,V为供试品溶液的体积。结果表明鞣质含量在35%。
实施例3:柔毛水杨梅正丁醇提取物可水解鞣质定性分析:将1中得到的柔毛水杨梅正丁醇提取物真空干燥至恒重,精密称取100mg,加少量甲醇溶解,转移到10毫升容量瓶,定容。吸取20微升,注入Agilent Series 1100高效液相系统,记录HPLC色谱图(如图-1)。色谱柱及洗脱条件等如下:
色谱柱:C18(RP)分析型色谱柱(5mm,4.6mm-250mm;Shimazu,Japan)
流速:1.0mL/min
检测波长:254nm
流动相:(A)0.1%醋酸-水溶液;(B)甲醇:0-10min,10%B;20-50min,10%B-30%B;50-70min,30%B-100%B
实施例3:在37℃下,在2毫升石英比色皿中加入该酶底物:0.2mM乙酰辅酶A溶液25微升,0.4mM丙二酰辅酶A溶液50微升和1.3mM NADPH溶液50微升,再加入0.1M,pH 7.0磷酸盐缓冲液1.85毫升及FAS溶液30微升混匀,启动酶促反应。用分光光度计监测单位时间内340nm波长处吸光值A的变化(ΔA/min)来表示FAS的活力。在一定浓度柔毛水杨梅正丁醇提取物存在下,FAS全酶反应的活力下降,当其活力下降一半时所需柔毛水杨梅正丁醇提取物的浓度即为IC50,用该数值表示柔毛水杨梅正丁醇提取物对FAS的抑制能力,上述测活体系中FAS的终浓度为7微克/毫升。结果如表1所示:柔毛水杨梅正丁醇萃取物具有非常强的抗脂肪酸合酶的活性,IC50是2.5μg/ml.。
实施例6:大鼠减肥实验,将雌性大鼠随机分组(10只每组,1只每笼),每只大鼠体重约200克。柔毛水杨梅正丁醇提取物加水溶解,可以适当加热,超声,配成浓度0.3g/mL的溶液。与对照组饲喂双蒸水,试验组饲喂按上述方法准备的柔毛水杨梅正丁醇提取物,每公斤400毫克。实验中饲料为标准配方。饲喂方法为用圆头灌注器插入大鼠食道向胃内灌注,每天一次,每只2ml,上午10~11时喂药.饲料和饮水为自主取食.实验时间20天,每2天测量一次每只大鼠的体重和摄食量,并观察其排泄物状况。数据表明喂柔毛水杨梅正丁醇提取物的大鼠,体重下降约10.2%,有减肥作用。
表1柔毛水杨梅正丁醇提取物的减肥作用
给药前(g) | 平均体重(g) | 平均体重变化 | |
水柔毛水杨梅正丁醇提取物 | 21.522.8 | 23.1120.47 | 7.5%-10.2% |
Claims (3)
1、柔毛水杨梅正丁醇提取物在制备用于治疗或预防肥胖的药物或膳食补充剂中的用途。
2、一种提取用于如权利要求1所述应用的柔毛水杨梅正丁醇提取物的方法,其特征在于选用水及甲醇、乙醇、丙醇、乙酸乙酯、丁醇等有机溶剂或上述任何两种或多种互溶的混合溶剂,在室温下浸渍提取或超声提取或加热回流提取获得,提取次数可为一次或多次,提取物分散到适量的水中,然后利用与分散该提取物的水等量的氯仿、正己烷、环己烷或石油醚等有机溶剂中的任何一种萃取除去脂溶性杂质,再利用与分散该提取物的水等量的正丁醇萃取获得活性提取物。
3、如权利要求1所述的用途,其中柔毛水杨梅正丁醇提取物是以活性成分GeminA作为标记物的提取物,其特征是富含鞣质,含量在30%以上。
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CN112603842A (zh) * | 2020-12-31 | 2021-04-06 | 中国科学院成都生物研究所 | 一种蔷薇花活性提取物及其应用 |
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WO2010143058A1 (en) * | 2009-06-12 | 2010-12-16 | Generex Pharmaceuticals, Inc. | Compositions and methods for the prevention and treatment of heart failure |
WO2010143065A1 (en) * | 2009-06-12 | 2010-12-16 | Generex Pharmaceuticals, Inc. | Compositions and methods for increasing lifespan and health span |
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CN112603842A (zh) * | 2020-12-31 | 2021-04-06 | 中国科学院成都生物研究所 | 一种蔷薇花活性提取物及其应用 |
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