CN116617290A - 一种治疗糖尿病足病的中药组合物 - Google Patents
一种治疗糖尿病足病的中药组合物 Download PDFInfo
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- CN116617290A CN116617290A CN202310490806.2A CN202310490806A CN116617290A CN 116617290 A CN116617290 A CN 116617290A CN 202310490806 A CN202310490806 A CN 202310490806A CN 116617290 A CN116617290 A CN 116617290A
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Abstract
本申请涉及一种治疗糖尿病足病的中药组合物。本申请首次公开苓桂术甘汤提取物用于制备治疗糖尿病足病的药物的用途。本申请通过动物实验证实,苓桂术甘汤提取物可以显著改善糖尿病足病模型大鼠糖尿病足病引起的溃疡性创伤,并且可以有效抑制糖尿病足耐药菌感染,并促进溃疡愈合。
Description
技术领域
本发明属于中医药技术领域,更特别地,涉及一种治疗糖尿病足病的中药组合物及其制备方法。
背景技术
张仲景在《金匮要略》中首先提出“痰饮”病名,认为该病的形成虽与肺、脾、肾三焦功能失调有关,但与脾胃的关系尤为密切。在“病痰饮者,当以温药和之”的大法下,创制了从脾治饮的苓桂术甘汤。本方见于《伤寒》一处,《金匮》二处,《伤寒·太阳》篇67条:“伤寒若吐若下后,心下逆满,气上冲胸,起则头眩,脉沉紧。发汗则动经,身为振振摇者,茯苓桂枝白术甘草汤主之”。《金匮·痰饮咳嗽病脉证治》云:“心下有痰饮,胸胁支满,目眩,苓桂术甘汤主之”。又云:“夫短气有微饮,当从小便去之,苓桂术甘汤主之”,方由茯苓、桂枝、白术和甘草四味常用中药组成。
其中茯苓健脾祛湿以利水;桂枝通阳化气,温化水饮;白术健脾燥湿以祛生痰之源;甘草补脾益气,调和诸药。四药合用,而成为治疗痰饮的常用方剂。
现代医学认为苓桂术甘汤治疗证的水饮停聚与组织器官的水肿及炎症有关,该方的药理作用可归纳为:利尿、镇痛、祛痰、止咳、增强血液循环、促进机体消化及免疫功能。故现代医学中的某些眩晕、支气管炎、心血管疾病、关节炎、泌尿系统、消化系统、神经系统及内分泌系统疾病中所出现的痰饮病证候,只要中医辨证属脾胃阳虚、痰饮为患者,均可用苓桂术甘汤加减治疗。
苓桂术甘汤的仲景处方:茯苓四两,桂枝三两去皮,白术二两,甘草二两炙。本方历来以汤剂使用,其煎法为:上四味,以水六升,煮取三升,去滓,分温三服。
2020年11月,国家中医药管理局发布了古代经典名方关键信息表,其中关于苓桂术甘汤的记载如下:
茯苓,为多孔菌科真菌茯苓Poria cocos(Schw.)Wolf的干燥菌核,始载于《神农本草经》,列为上品,历代本草皆有收载。茯苓味甘、淡,性平,归心、肺、脾、肾经。《中国药典》2000年版记载:茯苓有“利水渗湿,健脾宁心”之功能,主治水肿尿少、痰饮眩悸、脾虚食少、便溏泄泻、心神不安及惊悸失眠等症。
桂枝,为樟科植物肉桂Cinnamomum cassia Presl的干燥嫩枝,始载于《神农本草经》,列为上品。桂枝味辛、甘,性温,归心、肺、膀胱经。桂枝发汗通肌,温通经脉,助阳化气,平冲降气,用于风寒感冒,脘腹冷痛,血寒经闭,关节痹痛,痰饮,水肿,心悸,奔豚。
白术,为菊科植物白术Atractylodes macrocephala Koidz.的干燥根茎,始载于《神农本草经》,列为上品。白术味苦、甘,性温,归脾、胃经。白术健脾益气,燥湿利水,止汗安胎。用于脾虚食少,腹胀泄泻,痰饮眩悸,水肿,自汗,胎动不安。
甘草,为豆科植物甘草Glycyrrhiza uralensis Fisch.,胀果甘草Glycyrrhizainflata Batal.或光果甘草Glycyrrhizaglabra L的干燥根及根茎,始载于《神农本草经》,列为上品。甘草味甘,性平,归心、肺、脾、肾经。甘草补脾益气,清热解毒,祛痰止咳,缓急止痛,调和诸药。用于脾胃虚弱,倦怠乏力,心悸气短,咳嗽痰多,脘腹、四肢挛急疼痛,痈肿疮毒,缓解药物毒性、烈性。
苓桂术甘汤方中以茯苓为君,健脾渗湿,祛痰化饮。以桂枝为臣,温阳化气,既可温阳以化饮,又能化气以利水,且兼平冲降逆;与茯苓相伍,一利一温,对于水饮停留而偏寒者,实有温化渗利之妙用。湿源于脾,脾虚则生湿,故佐以白术健脾躁湿,助脾化运,脾阳健旺,水湿自除。使以甘草益气和中,共收饮去脾和,湿不复聚之功。药虽四味,配伍严谨,湿而不热,利而不峻,确为痰饮之和剂。
茯苓中β-茯苓聚糖为主成分,约占干燥品的93%。一类四环三萜类的有机酸,如茯苓酸、土莫酸、齿孔酸、松苓酸等,还有脂肪酸,如月桂酸、棕榈酸、十二酸等和麦角甾醇、卵磷脂、无机成分等。其药理研究表明:茯苓有利尿作用,关于茯苓利尿作用原理尚未阐明,有报道认为是与茯苓素激活细胞膜上的Na+,K+-ATP酶有关。茯苓素可增强心肌收缩力,抑制毛细血管通透性,能明显降低小鼠的自发活动等,此外,茯苓素和茯苓多糖均有抗肿瘤和免疫增强作用,与环磷酰胺等抗肿瘤药物合用,有明显的增效作用。
桂枝的主要成分有桂皮醛、桂皮酸、香豆精、β-谷甾醇、原儿茶酸等,其药理研究表明:桂枝对中枢神经系统有镇静、镇痛、解热和抗惊厥作用,而其中的桂皮醛和桂皮酸为有效成分。此外桂枝能增加冠状动脉血流量,有抗菌、抗炎、抗过敏等作用。
白术中含苍术醇、苍术酮、芹子烯、倍半萜内酯化合物白术内酯和一些炔类化合物等,其药理研究表明:白术煎剂对大鼠、兔和狗均有显著而持久的利尿作用,且促进电解质特别是钠的排泄。白术的乙醇提取物具有剂量依赖性的抑制肾脏Na+,K+-ATP酶的活性,可能是白术利尿作用的机制之一。此外,白术还有血管扩张、抗凝血、抑菌、抗肿瘤等活性。
甘草中含三萜皂苷甘草酸,其钾、钙盐为甘草甜素,是甘草中的甜味成分,水解后产生甘草次酸:还有黄酮类成分,如甘草素、甘草黄酮、甘草查尔酮、甘草香豆素和四氢喹啉类生物碱、多糖等。其药理研究表明:甘草对心血管系统有降脂、抑制血小板凝集、抗心律失常作用,有肾上腺皮质激素样作用、抗炎、抗癌、抗病毒、止咳平喘和祛痰作用等。
苓桂术甘汤是《伤寒论》中苓桂剂的代表方,主要治疗水饮上逆冲心等症。随着人们对该方的研究和认识的加深,该方在临床上的应用也变得更为广泛,应用范围也不断扩大,其中以慢性心力衰竭、高血压、慢性支气管炎、美尼尔氏综合征较为常见。现代药理学实验研究表明,苓桂术甘汤主要有利尿、强心、抗眩晕、抗过敏、抗炎、改善代谢、调节肠道功能等作用。
苓桂术甘汤对慢性心力衰竭的治疗作用主要是来自于其对NF-κB的抑制作用,同时还能改善血流动力学功能障碍,并且抑制心脏结构的变化,对提高慢性心力衰竭患者的存活率有显著的提高作用。除此之外,该汤剂还能通过清除细胞内氧自由基从而达到抑制心肌细胞凋亡的作用,对心肌细胞氧化应激损伤具有保护作用。
苓桂术甘汤能够激活动物脑内血脑屏障上的特定转运体蛋白RAGE,降低脑内Aβ淀粉样蛋白的沉积;同时也控制了脑部海马区与皮质区的炎症因子TNF-α、IL-6、IL-1β的表达,降低脑内炎症程度,对阿尔兹海默症具有较好的干预效果。
苓桂术甘汤近年来也常用于治疗肥胖和高脂血症,表明苓桂术甘汤具有调节代谢的作用。苓桂术甘汤能通过提高血清甲状腺激素水平从而减轻高脂饮食导致的脂肪性肝病。同时苓桂术甘汤还能激活过氧化物酶体受体(PPAR)和Hedgehog信号通路,从而降低炎症因子AHR、Irf2BP2和RASGEF1B的表达,提高抗氧化剂OslGN1的表达从而降低胆固醇的分泌,减轻高脂血症。
苓桂术甘汤对免疫系统也有影响。研究表明苓桂术甘汤对正常小鼠的免疫系统有影响,能明显增加小鼠免疫器官质量,提高小鼠碳粒廓清指数、血清溶血素抗体水平和淋巴细胞转化率,除此之外还能明显改善环磷酰胺诱导的免疫功能低下的模型小鼠的免疫能力。对其机制进行研究时发现苓桂术甘汤能明显提高环磷酰胺所致的免疫抑制的小鼠模型体内的T细胞总数提高,纠正T细胞亚群紊乱,同时还能促进TH细胞的功能,从而改善免疫状况。
苓桂术甘汤能改善梅尼埃病内耳膜迷路积水,减轻患者眩晕等反应。这种药理作用是由于苓桂术甘汤能增加AQP8在小肠近段和远端-十二指肠和空肠绒毛上皮细胞胞浆内的表达,从而减轻内耳膜迷路积水的程度。目前苓桂术甘汤在临床上也常用于眩晕症、梅尼埃病等疾病的治疗。
糖尿病足是糖尿病一种严重的并发症,是糖尿病患者致残的主要原因之一。糖尿病足发病机制尚不完全清楚,目前认为血脂、血糖代谢紊乱与其发生密切相关,糖尿病足的发生与慢性周围血管病变和周围神经病变密切相关。首先,糖尿病患者由于神经病变使下肢保护功能减退。其次,糖尿病患者由于长期高血糖导致动脉硬化发生微循环障碍,局部组织缺血和抵抗力下降,微小的创伤均可引起感染而形成溃疡加大。
此外,组织内含糖量高,适宜细菌生长,且白细胞吞噬能力降低,使感染率增高,感染伤口难以愈合,故病程迁延不愈,给患者及家庭带来极大的痛苦和经济负担。所以强调糖尿病足的早期治疗及预防,以防坏疽的发生,这对保存患肢,减少费用,提高生活质量极其重要。
随着抗生素的广泛应用,微生物耐药性形成的速度与抗菌素灭菌能力成比例上升,以致抗生素的继续使用加快多重耐药菌株的形成。糖尿病足部导致皮肤软组织破坏,造成外源细菌侵入,提高了糖尿病足细菌感染,尤其提高了糖尿病足耐药细菌感染的治疗难度,严重的可导致局部软组织感染,甚至骨髓炎的形成。目前,抗生素耐药性已经成为全球严重的公共卫生问题。
目前尚没有任何应用苓桂术甘汤或其加减方治疗糖尿病足病的报道。
发明内容
本申请首次公开苓桂术甘汤提取物对于糖尿病足病的治疗作用。所述苓桂术甘汤由茯苓、桂枝、白术和甘草组成。
所述苓桂术甘汤中茯苓、桂枝、白术和甘草均为生品饮片。
所述苓桂术甘汤中各个药材的质量比为茯苓:桂枝:白术:甘草=2:1.5:1.5:1。
所述苓桂术甘汤中各个药材的质量比为茯苓:桂枝:白术:甘草=55.2:41.4:41.4:27.6。
所述苓桂术甘汤的制备方法如下:
步骤1)蒸馏:取桂枝,与白术混合,加水润透,水蒸气蒸馏,收集挥发油、芳香水以及混合药渣;
步骤2)提取:取甘草,与茯苓以及步骤1)所述混合药渣混合,加水提取,所得提取液经过滤以及浓缩,制备水提物清膏;
步骤3)包合:取步骤1)所得芳香水和挥发油混合物,加入羟丙基-β-环糊精,充分溶解后,研磨包合,制备包合液;
步骤4)干燥:分别取步骤3)所述包合液与步骤2)所述水提物清膏,干燥,得浸膏粉,即为苓桂术甘汤提取物。
所述蒸馏的步骤中:所述水的质量是所述桂枝的质量与所述桂枝和白术的质量之和的1~3倍。
所述蒸馏的步骤中:所述水蒸气蒸馏的时长为1.0~3h。
所述提取的步骤中:所述水的质量是所述桂枝和白术药渣的质量、所述甘草与茯苓的质量之和的10~20倍;
所述提取的步骤中:所述水提物清膏的相对密度为1.03~1.15。
所述提取的步骤中:所述浓缩采用的温度为50~70℃。
所述包合的步骤中:所述羟丙基-β-环糊精的质量是所述挥发油质量的10~20倍。
所述包合的步骤中:所述包合的方式采用研磨包合。
所述干燥的步骤中:所述干燥的方式采用喷雾干燥。进风温度为140~195℃,所述喷雾干燥的出风温度为60~95℃。
所述苓桂术甘汤提取物通过口服起效。
所述苓桂术甘汤提取物可以提高糖尿病足病伤口闭合率促进伤口闭合。
所述苓桂术甘汤提取物具有显著的抗铜绿假单胞菌等耐药菌感染的作用。
所述苓桂术甘汤提取物可以单独应用,也可以和其他药物联合使用。
具体实施方式
通过以下实验进一步说明本发明的有益效果。但并不局限于下述实施例,本领域的技术人员在本发明基础上所作的,不脱离本发明实质内容的等同替代、变换或处方加减,亦均在本发明的保护范围之内。
实施例1苓桂术甘汤提取物的制备
(1)蒸馏:取桂枝饮片60g、白术饮片60g,合并,加水330ml,浸泡60分钟,通入水蒸汽蒸馏(1.0小时),收取挥发油及芳香水混合液共90ml以及桂枝和白术药渣,其中挥发油约0.5ml;
(2)提取:向蒸馏后所得药渣加入茯苓饮片90g、甘草饮片40g,加水3300ml,煎煮1.0小时,滤取药液;
(3)步骤2)所得药液在60±5℃下减压浓缩成相对密度为1.06(65℃)的清膏;
(4)向步骤1)所得挥发油及芳香水混合液中加入羟丙基-β-环糊精8.5g,溶解,研磨包合60分钟,所得包合液加入到浓缩清膏中,搅拌均匀,在进风温度185℃、出风温度75℃下喷雾干燥,得干燥粉48.6g,为苓桂术甘汤提取物。
对比例1:缺味药材的对比配方药物提取物的制备
按实施例1的药材配伍、配比和提取方法,分别制备缺少茯苓、桂枝、白术和甘草的含有三味药材的提取物。即对比配方1-4,药材配伍如下:
实施例2糖尿病足治疗效果实验
2.1糖尿病足大鼠模型的建立
受试动物为SPF级SD雄性大鼠63只,体重220~240g,自由饮水饮食,饲养间室温保持在25±1℃,明暗周期为12小时。
SD大鼠适应性喂养一周后,测量大鼠血糖,腹腔注射脲佐菌素,注射量为75mg/kg,继续高脂饲料喂养一周后测量大鼠血糖,血糖水平高于16.7mmol/L,且伴随有多食、多饮、多尿和体重减轻的情况,则视为糖尿病大鼠造模成功。将大鼠随机分为8组,每组7只。
2.2给药
在给予脲佐菌素2周后,在每只大鼠的右后爪(足部)产生伤口,并将这一天定义为第0天。方法如下:腹腔注射10%的水合氯醛(0.3ml/100g)将每只大鼠麻醉。使用柔性透明塑料模板在右后爪(足部)的背部表面标出矩形图案,然后除去标准面积约2mm×5mm的全厚皮层。在诱导糖尿病伤口之后,向大鼠给予蒸馏水10mg/kg,记为糖尿病创面组,其余组给予苓桂术甘汤提取物和对比配方,给药情况见表1,进行连续灌胃给药21天,每日两次。没有糖尿病诱导的单独的大鼠组也保持伤口产生,定义为正常创面组(7只),向大鼠给予蒸馏水,剂量为10mg/kg。
将本发明实施例1苓桂术甘汤提取物及对比例1对比配方1-4所得提取物,以0.1%羧甲基纤维素钠水溶液配制为1mg/mL的混悬液。按剂量要求给予试验动物。
2.3.实验结果如表1所示:
按如下公式计算伤口闭合率:
伤口闭合%=[(W0-WN)/W0]×100%;
W0=第0天的伤口面积;
WN=第3、7、14、21天的伤口面积。
对照组的伤口闭合是负数,表明伤口创伤面积变大,伤口恶化。
正常组和本发明苓桂术甘汤提取物组和对比配方组的伤口闭合是正数,表明伤口创伤面积变小,伤口愈合。
表1试验分组及伤口闭合率统计数据(n=15)
结果显示:在21天内,正常伤口闭合为约85.6%,未治疗的糖尿病伤口(糖尿病创面组)显著恶化,伤口尺寸增加至约131.2%。本发明所述的苓桂术甘汤提取物治疗组中糖尿病伤口显示,每日两次分别在相同时段灌胃5mg/kg、10mg/kg和约20mg/kg,21天后的伤口闭合率分别为约43.5%、约56.7%和约73.6%。而四个对比配方组虽然给药量和苓桂术甘汤提取物高剂量组相同,但是21天后伤口闭合率均不及苓桂术甘汤低剂量组。但是仍然较糖尿病创面组为优。
综上数据所述,本发明所述的苓桂术甘汤提取物治疗显示在治疗约14天之后糖尿病伤口显著闭合,表明苓桂术甘汤提取物对于糖尿病足病引起的溃疡性创伤具有显著的改善作用。尤其是高剂量组。四个对比配方,从药材组成上看,虽然仅比苓桂术甘汤少一味药材,但是其治疗效果显著低于苓桂术甘汤三个剂量组。
实施例3糖尿病足耐药菌感染溃疡的治疗效果实验
3.1测试化合物:
以呋喃酮化合物(Z)-4-溴-5-(溴亚甲基)-2(5H)-呋喃酮为阳性对照,以生理盐水为阴性对照,将阳性药物、本发明实施例1苓桂术甘汤提取物及对比例1对比配方1-4所得提取物,以0.1%羧甲基纤维素钠水溶液配制为1mg/mL的混悬液。
3.2动物模型建立及分组
90只SD大鼠适应性喂养一周后,测量大鼠血糖,腹腔注射脲佐菌素,注射量为75mg/kg,继续高脂饲料喂养一周后测量大鼠血糖,血糖水平高于16.7mmol/L,且伴随有多食、多饮、多尿和体重减轻的情况,则视为糖尿病大鼠造模成功。
在给予脲佐菌素2周后,在每只大鼠的右后爪(足部)产生伤口,并将这一天定义为第0天。伤口产生方式为腹腔注射10%的水合氯醛(0.3ml/100g)将每只大鼠麻醉后用烫伤仪烫伤(1000kg,90℃,10S),达到深度II烫伤。
三天后,在所有大鼠溃疡部位皮下注射pH7.4磷酸盐缓冲液和1×108ufc/ml铜绿假单胞耐药菌各0.1ml。
SD大鼠随机分为正常耐药菌感染溃疡组,糖尿病耐药菌感染对照组,阳性对照组,苓桂术甘汤提取物低、中、高剂量组和对比配方1-4组,每组9只。
正常耐药菌感染溃疡组与糖尿病耐药菌感染对照组每天生理盐水处理,阳性对照组每天采用(Z)-4-溴-5-(溴亚甲基)-2(5H)-呋喃酮处理,给药测试组给予不同剂量苓桂术甘汤提取物,测量各组愈合时间,具体结果参见表2。
表2糖尿病足铜绿假单胞耐药菌感染愈合时间(n=9)
表2结果显示糖尿病耐药菌感染对照组大鼠的溃疡伤口愈合时间相对于正常组明显延长,显示了糖尿病耐药菌感染大鼠符合慢性溃疡的特征,阳性对照组,苓桂术甘汤低、中、高剂量组和对比配方1-4组均显示了明显缩短溃疡伤口愈合时间的效果,表明了苓桂术甘汤提取物具有治疗糖尿病足耐药菌感染的效果,可用于糖尿病足细菌感染,特别是糖尿病足耐药细菌感染的治疗,但是如果缺少任何一味药材,其治疗效果则大打折扣,表明苓桂术甘汤提取物为一特定组合,其中四味药材缺一不可。
上述实施例仅例示性说明本发明的原理及功效,而非用于限制本发明。任何本领域的技术人士皆可在不违背本发明的精神及范畴下,对上述实施例进行修饰或改变。因此,任何本领域的技术人士在未脱离本发明所揭示的精神与技术思想下所完成的一切等效修饰或改变,仍应由本发明的权利要求所涵盖,即以苓桂术甘汤或苓桂术甘汤加减,单独或与其他药物或辅料联合施用,治疗糖尿病足病,都落入要求保护的本发明专利申请范围内,本发明专利申请要求保护范围由所附的权利要求书及其等效物界定。
Claims (9)
1.苓桂术甘汤提取物用于制备治疗糖尿病足病药物的用途,其特征在于,所述苓桂术甘汤提取物由茯苓、桂枝、白术和甘草加工而成。
2.如权利要求1所述的用途,其特征在于,所述茯苓、桂枝、白术和甘草均为生品饮片。
3.如权利要求2所述的用途,其特征在于,所述苓桂术甘汤中各个药材的质量比为茯苓:桂枝:白术:甘草=2:1.5:1.5:1。
4.如权利要求2所述的用途,其特征在于,所述苓桂术甘汤中各个药材的质量比为茯苓:桂枝:白术:甘草=55.2:41.4:41.4:27.6。
5.如权利要求3或4所述的用途,其特征在于,所述苓桂术甘汤提取物的制备方法如下:
步骤1)蒸馏:取桂枝,与白术混合,加水润透,水蒸气蒸馏,收集挥发油、芳香水以及混合药渣;
步骤2)提取:取甘草,与茯苓以及步骤1)所述混合药渣混合,加水提取,所得提取液经过滤以及浓缩,制备水提物清膏;
步骤3)包合:取步骤1)所得芳香水和挥发油混合物,加入羟丙基-β-环糊精,充分溶解后,研磨包合,制备包合液;
步骤4)干燥:分别取步骤3)所述包合液与步骤2)所述水提物清膏,干燥,分别得浸膏粉,即为苓桂术甘汤提取物;
所述蒸馏的步骤中:所述水的质量是所述桂枝的质量与所述桂枝和白术的质量之和的1~3倍;
所述蒸馏的步骤中:所述水蒸气蒸馏的时长为1.0~3h;
所述提取的步骤中:所述水的质量是所述桂枝和白术药渣的质量、所述甘草与茯苓的质量之和的10~20倍;
所述提取的步骤中:所述水提物清膏的相对密度为1.03~1.15;
所述提取的步骤中:所述浓缩采用的温度为50~70℃;
所述包合的步骤中:所述羟丙基-β-环糊精的质量是所述挥发油质量的10~20倍;
所述包合的步骤中:所述包合的方式采用研磨包合;
所述干燥的步骤中:所述干燥的方式采用喷雾干燥。进风温度为140~195℃,所述喷雾干燥的出风温度为60~95℃。
6.如权利要求1所述的用途,其特征在于,所述苓桂术甘汤提取物通过口服起效。
7.如权利要求1所述的用途,其特征在于,所述苓桂术甘汤提取物用于制备提高糖尿病足病伤口闭合率药物的用途。
8.如权利要求1所述的用途,其特征在于,所述苓桂术甘汤提取物用于制备治疗铜绿假单胞菌感染的药物的用途。
9.如权利要求1所述的用途,其特征在于,所述苓桂术甘汤提取物可以单独应用,也可以和其他药物联合使用。
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