CN110025604A - 金丝桃素在制备抑制葡萄糖苷酶活性的相关产品中的用途 - Google Patents
金丝桃素在制备抑制葡萄糖苷酶活性的相关产品中的用途 Download PDFInfo
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- CN110025604A CN110025604A CN201910291434.4A CN201910291434A CN110025604A CN 110025604 A CN110025604 A CN 110025604A CN 201910291434 A CN201910291434 A CN 201910291434A CN 110025604 A CN110025604 A CN 110025604A
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- Prior art keywords
- hypericin
- product
- glucosidase
- alpha
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Abstract
本发明公开了金丝桃素在制备抑制葡萄糖苷酶活性的相关产品中的用途,本发明通过实验证明,金丝桃素能有效抑制α‑葡萄糖苷酶的活性,其对α‑葡萄糖苷酶的半抑制浓度IC 50值仅为0.00498mg/mL,抑制能力优于现有的α‑葡萄糖苷酶抑制剂,可用于制备调节糖化学代谢紊乱、降低餐后血糖、预防和/或治疗糖耐量受损和/或糖尿病及其并发症的产品,还可用于制备预防和/或治疗肥胖症、高血压、慢性乙肝、艾滋病、肿瘤等以α‑葡萄糖苷酶为靶点的疾病的药物,更可以用以制备α‑葡萄糖苷酶抑制剂,安全高效。
Description
技术领域
本发明涉及金丝桃素的用途领域,特别是涉及金丝桃素在制备抑制葡萄糖苷酶活性的相关产品中的用途。
背景技术
α-葡萄糖苷酶在食物吸收过程中起着重要的作用,必须与之结合后,食物才能消化和吸收。α-葡萄糖苷酶抑制剂的降糖机制是,通过抑制肠黏膜上的α-葡萄糖苷酶,使淀粉分解为葡萄糖的速度减缓,减少和延缓小肠对葡萄糖的吸收,以降低血糖,对餐后高血糖的作用比较明显。葡萄糖苷酶抑制剂不刺激胰岛素的分泌,单独使用本类药物通常不会引发低血糖,因此可帮助减少血糖的波动。可以明显降低糖尿病患者发生心血管病变的概率,对心肌梗死的改善作用最为显著。是少数可干预糖耐量受损的口服降糖药之一。α-葡萄糖苷酶抑制剂不仅对糖尿病有确切的疗效,而且对于肥胖症,慢性乙肝,艾滋病及肿瘤都有一定的治疗作用。目前用于临床的有阿卡波糖,伏格列波糖和米格列醇。
金丝桃素是贯叶连翘的提取物,人类在1957年首次从藤黄科植物贯叶连翘中分离出来,属于二蒽酮类化合物,是贯叶连翘中最具生物活性的物质,外观为灰黄色至棕褐色流动性粉末,味微苦,易溶于水,能散发出特殊的清香,具有抑制中枢神经和镇静的作用,添加到保健品中可增强免疫力,在欧洲(特别是德国)也被用作抗抑郁药物。金丝桃素也具有极强的抗病毒作用,能直接作用于猪瘟、口蹄疫等病毒,另外对高致病性禽流感病毒也有良好的杀灭效果。但对于金丝桃素对α-葡萄糖苷酶的抑制作用,还未见报道。
发明内容
本发明主要解决的技术问题是提供金丝桃素在制备抑制葡萄糖苷酶活性的相关产品中的用途,能够有效抑制α-葡萄糖苷酶的活性。
为解决上述技术问题,本发明采用的一个技术方案是:
提供金丝桃素、金丝桃素在药学上可接受的水合物、金丝桃素在药学上可接受的盐、和金丝桃素衍生物中的至少一种在制备抑制葡萄糖苷酶活性的产品中的用途。
本发明通过实验数据表明,金丝桃素对α-葡萄糖苷酶的活性具有很好的抑制作用,因此可用于制备具有相关作用的产品,安全有效。
所述产品包括但不限于药品、保健品、食品等。
进一步地,所述产品为调节糖化学代谢紊乱的产品。
所述糖化学代谢紊乱即指糖代谢紊乱,糖代谢紊乱(glucose metabolismdisorders)指调节葡萄糖、果糖、半乳糖等代谢的激素或酶的结构、功能、浓度异常,或组织、器官的病理生理变化,监测血糖会有血糖的升高。通常是某些疾病、肥胖、高脂饮食等原因,或者先天性因素引起人体调节糖代谢的激素或酶的结构、功能、浓度异常或组织、器官病变导致。
葡萄糖苷酶是生物体内糖代谢途径中的重要成员之一。β-葡萄糖苷酶可以参与纤维素的代谢以及多种生理生化途径,α-葡萄糖苷酶更是直接参与淀粉及糖原的代谢途径。这类酶的功能发生异常会导致出现代谢类的疾病,同时这类酶也是多种药物与抑制剂的作用靶点,用以调节人体内的糖化学代谢。因此,金丝桃素可通过调节α-葡萄糖苷酶的活性,进而调节人体内的糖化学代谢,可用于制备相关产品。
进一步地,所述产品为降低餐后血糖的产品。
金丝桃素可通过过抑制肠黏膜上的α-葡萄糖苷酶,使淀粉分解为葡萄糖的速度减缓,减少和延缓小肠对葡萄糖的吸收,以降低血糖,对餐后高血糖的作用比较明显,可用于制备降低餐后血糖的产品。
进一步地,所述产品为预防和/或治疗糖耐量受损和/或糖尿病及其并发症的产品。
糖调节受损即糖尿病前期,主要包括空腹血糖受损(impaired fasting glucose,IFG)和糖耐量减低(impaired glucose tolerance,IGT)。金丝桃素对α-葡萄糖苷酶的活性具有很好的抑制作用,可有效改善糖调节受损,预防和/或治疗糖耐量受损和/或糖尿病,同时也可预防或改善糖尿病并发症,可用于制备相关产品。
进一步地,所述产品为治疗肥胖症、高血压、慢性乙肝、艾滋病、肿瘤中的一种或几种病症的产品。
进一步地,所述产品为葡萄糖苷酶抑制剂。
进一步地,所述葡萄糖苷酶抑制剂为α-葡萄糖苷酶抑制剂。
本发明通过实验表明,金丝桃素对α-葡萄糖苷酶的半抑制浓度IC 50值仅为0.00498mg/mL,抑制能力远优于现有的α-葡萄糖苷酶抑制剂阿卡波糖,可用于制备α-葡萄糖苷酶抑制剂。
进一步地,所述产品是由金丝桃素或其在药学上可接受的水合物、盐或衍生物为活性成分,与药学上可接受的辅料或载体组成。
以上述所的金丝桃素,与药学上可接受的辅助添加性成分混合后,按相应的常规药物制剂方法,可以制备α-葡萄糖苷酶抑制剂类的药物。例如,与在口服制剂中可以被接受的如崩解剂、赋形剂、润滑剂、粘合剂、填充剂等常用的辅助添加成份混合后,按常规的操作方法和过程,可以制成为片剂、丸剂、胶囊剂或多种相应的缓释剂、控释剂等固体口服制剂形式的药物;与常规的增溶剂、乳化剂、润湿剂、起泡或消泡剂等表面活性剂、稀释剂、防腐剂、稳定剂、矫味剂、增稠剂等混合后,按相应的常规方法,可以制成为如水剂、糖浆等液体制剂形式的口服药物。
本发明还提供了金丝桃素、金丝桃素在药学上可接受的水合物、金丝桃素在药学上可接受的盐、和金丝桃素衍生物中的至少一种在制备葡萄糖苷酶抑制剂中的用途。
本发明还提供了金丝桃素、金丝桃素在药学上可接受的水合物、金丝桃素在药学上可接受的盐、和金丝桃素衍生物中的至少一种在制备α-葡萄糖苷酶抑制剂中的用途。
本发明还提供了金丝桃素、金丝桃素在药学上可接受的水合物、金丝桃素在药学上可接受的盐、和金丝桃素衍生物中的至少一种在预防和/或治疗肥胖症、慢性乙肝、艾滋病、肿瘤中的一种或几种病症的产品中的用途。
本发明的有益效果是:本发明通过实验证明,金丝桃素能有效抑制α-葡萄糖苷酶的活性,其对α-葡萄糖苷酶的半抑制浓度IC 50值仅为0.00498mg/mL,抑制能力优于现有的α-葡萄糖苷酶抑制剂,可用于制备α-葡萄糖苷酶抑制剂类产品,也可用于制备预防和/或治疗以α-葡萄糖苷酶为靶点的疾病的药物,如治疗糖耐量受损、糖尿病、高血压、肥胖症、慢性乙肝、艾滋病、肿瘤等疾病的药物。
附图说明
图1是本发明不同浓度金丝桃素的抑制剂的抑制率;
图2是不同浓度的阿卡波糖抑制率曲线。
具体实施方式
下面结合附图对本发明的技术方案进行清楚、完整地描述,显然,所描述的实施例是本发明一部分实施例,而不是全部的实施例。基于本发明中的实施例,本领域普通技术人员在没有做出创造性劳动前提下所获得的所有其他实施例,都属于本发明保护的范围。
实施例1
一、仪器与试剂
NaH2PO4·2H2O(天津市大茂化学试剂厂);
Na2HPO4·12H2O(天津市大茂化学试剂厂);
无水Na2CO3(天津市河东区红岩试剂厂);
酵母α-葡萄糖苷酶(美国sigma公司,100UN);
pNPG(阿拉丁试剂(上海)有限公司);
阿卡波糖(上海源叶科技有限公司);
金丝桃素(上海源叶科技有限公司);
酶标仪(BioTek公司,型号:EPOCH2);
恒温振荡器(THOMMO SHAKER,型号:BE-9010)。
二、实验方法
1、试剂的配制
(1)甲液的配置:NaH2PO4·2H2O称取15.603g,定容至500mL,4℃,棕色瓶保存,备用。
(2)乙液的配置:Na2HPO4·12H2O称取35.822g,定容至500mL,4℃,棕色瓶保存,备用。
(3)0.1M磷酸缓冲液的配置:量取甲液51mL,乙液49mL,加入100mL水,混匀,得pH值6.8的磷酸缓冲液,4℃,棕色瓶保存,备用。
(4)酵母α-葡萄糖苷酶:将100U/ml酶原液用磷酸缓冲液(pH 6.8)稀释为20U/mL的酶溶液,冷冻备用,使用前用磷酸缓冲液(pH 6.8)稀释为1U/mL,备用。
(5)底物pNPG配制:精密称取0.3766g pNPG,加入适量的磷酸钠缓冲液中溶解,再定容至50mL,配制成25mmol/L母液,使用前用磷酸钠缓冲液配制成0.5mmol/L,备用。
(6)阿卡波糖抑制剂配制:精密称取13.9mg阿卡波糖,用DMSO定容至1mL,配制成13.9mg/mL,备用。
(7)0.1mol/L的Na2CO3配制:称取1.06g Na2CO3于烧杯中,加入适量蒸馏水溶解,并定容到100mL,4℃下保存,备用。
2、抑制剂的制备
精密称取3.6mg金丝桃素至10mL容量瓶中,用少量DMSO溶解后定容至刻度,并用DMSO稀释4倍浓度为0.09mg/mL作为母液,需要使用时再稀释至适当的浓度,即得系列不同浓度金丝桃素的抑制剂。
3、金丝桃素对α-葡萄糖苷酶的活性抑制效果实验
原理:对硝基苯-α-D-葡萄糖苷(pNPG)经α-葡萄糖苷酶水解可产生对硝基苯酚,其在405nm呈特异性吸收,因此可以通过检测对硝基苯酚的生成量检测α-葡萄糖苷酶的活性。
实验分为空白组、对照组、样品空白组和样品组,制备方法:各反应物按表1中剂量在96孔板中进行加样,每组3个平行,将抑制剂、DMSO、缓冲液和酶溶液混合均匀,在恒温振荡器中37℃保温10min,结束后,取出,加入50μL 0.5mmol/L pNPG溶液,充分混匀,于37℃水浴反应20min,结束后加入50μL 0.1mol/L的Na2CO3溶液中止反应,即得各试验组(空白组、对照组、样品空白组和样品组)。
由于PNPG在α-葡萄糖苷酶的作用下能水解产生葡萄糖和PNP,PNP在405nm处有最大吸收,使用酶标仪测定其吸光度,根据公式便可计算出各样品α-葡萄糖苷酶的抑制率及IC 50值。
公式:其中,Ac为空白组吸光值,AB为对照组吸光值,As为样品组吸光值,ASB为样品空白组吸光值。
表1各反应物添加计量和顺序 (单位:μL)
不同浓度金丝桃素的抑制剂与阿卡波糖不同浓度的抑制率见表2、表3,在分别对两组数据绘制拟合曲线,如图1、图2所示,以浓度为横坐标,抑制率为纵坐标,不同浓度金丝桃素的抑制剂的抑制曲线见图1,不同浓度的阿卡波糖抑制曲线见图2。再由拟合的曲线得到曲线方程:
不同浓度金丝桃素的抑制剂的抑制曲线方程为:y=108.01x-0.038,R2=0.997。
不同浓度的阿卡波糖抑制曲线方程为:y=0.516x+0.034,R2=0.998。
通过上述两个曲线方程即可分别求得金丝桃素和阿卡波糖对α-葡萄糖苷酶50%的抑制率时的浓度,求得金丝桃素对α-葡萄糖苷酶的半抑制浓度IC 50值为0.00498mg/mL,阿卡波糖对α-葡萄糖苷酶的半抑制浓度IC 50值为0.902mg/mL。
表2不同浓度金丝桃素的抑制剂的抑制率(n=3)
表3阿卡波糖不同浓度的抑制率(n=3)
阿卡波糖为一种α-葡萄糖苷酶抑制剂,可抑制各种α-葡萄糖苷酶如麦芽糖酶、异麦芽糖酶、葡萄糖淀粉酶及蔗糖酶的活性,使淀粉分解成寡糖如麦芽糖(双糖)、麦芽三糖及糊精(低聚糖)进而分解成葡萄糖的速度减慢,使蔗糖分解成葡萄糖和果糖的速度减慢,因此造成肠道葡萄糖的吸收减缓,从而缓解餐后高血糖,达到降低血糖的作用。长期服用,可降低空腹血糖和糖化血红蛋白的浓度。
从上述实验结果可以看出,金丝桃素能有效抑制α-葡萄糖苷酶的活性,其对α-葡萄糖苷酶的半抑制浓度远低于阿卡波糖对α-葡萄糖苷酶的半抑制浓度,效果显著,可用于制备α-葡萄糖苷酶抑制剂,也可用于制备治疗以α-葡萄糖苷酶为靶点的疾病的药物,如治疗糖耐量受损、糖尿病、高血压、肥胖症、慢性乙肝、艾滋病、肿瘤等疾病的药物。
尽管已经示出和描述了本发明的实施例,对于本领域的普通技术人员而言,可以理解在不脱离本发明的原理和精神的情况下可以对这些实施例进行多种变化、修改、替换和变型,本发明的范围由所附权利要求及其等同物限定。
Claims (10)
1.金丝桃素、金丝桃素在药学上可接受的水合物、金丝桃素在药学上可接受的盐、和金丝桃素衍生物中的至少一种,在制备抑制葡萄糖苷酶活性的产品中的用途。
2.根据权利要求1所述的用途,其特征在于,所述产品为调节糖化学代谢紊乱的产品。
3.根据权利要求1所述的用途,其特征在于,所述产品为降低餐后血糖的产品。
4.根据权利要求1所述的用途,其特征在于,所述产品为预防和/或治疗糖耐量受损和/或糖尿病及其并发症的产品。
5.根据权利要求1所述的用途,其特征在于,所述产品为治疗肥胖症、高血压、慢性乙肝、艾滋病、肿瘤中的一种或几种病症的产品。
6.根据权利要求1所述的用途,其特征在于,所述产品为葡萄糖苷酶抑制剂;进一步地,所述葡萄糖苷酶抑制剂为α-葡萄糖苷酶抑制剂。
7.根据权利要求1~6所述的用途,其特征在于,所述产品是金丝桃素、金丝桃素在药学上可接受的水合物、金丝桃素在药学上可接受的盐、和金丝桃素衍生物中的至少一种,与药学上可接受的辅料或载体组成。
8.金丝桃素、金丝桃素在药学上可接受的水合物、金丝桃素在药学上可接受的盐、和金丝桃素衍生物中的至少一种,在制备葡萄糖苷酶抑制剂中的用途。
9.金丝桃素、金丝桃素在药学上可接受的水合物、金丝桃素在药学上可接受的盐、和金丝桃素衍生物中的至少一种,在制备α-葡萄糖苷酶抑制剂中的用途。
10.金丝桃素、金丝桃素在药学上可接受的水合物、金丝桃素在药学上可接受的盐、和金丝桃素衍生物中的至少一种,在预防和/或治疗肥胖症、慢性乙肝、艾滋病、肿瘤中的一种或几种病症的产品中的用途。
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