CN105748615A - 一种三***的制备方法及应用 - Google Patents
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Abstract
本发明属于中药制剂领域,具体提供一种三***的制备方法,由大黄300g、盐酸小檗碱5g、黄芩浸膏21g作为原料药制成,采用超临界萃取制备而成,使得含量有很大提高,用量减少,本发明还提供了三***在制备抑制小鼠软骨ATDC5细胞增殖药物中的应用。
Description
技术领域
本发明涉及中药制剂技术领域,具体涉及一种三***的制备方法及应用。
背景技术
三***记载于药典标准,处方为大黄300g、盐酸小檗碱5g、黄芩浸膏21g,以上三味,黄芩浸膏系取黄芩,加水煎煮三次,第一次1.5小时,第二次1小时,第三次40分钟,合并煎液,滤过,滤液用盐酸调节pH值至1~2,静置1小时,取沉淀,用水洗涤使pH值至5~7,烘干,粉碎成细粉。取大黄150g,粉碎成细粉,剩余大黄粉碎成粗粉,用30%乙醇回流提取三次,滤过,合并滤液,回收乙酵并减压浓缩成稠裔,加人大黄细粉、盐酸小檗碱细粉、黄芩浸膏细粉及适量辅料,混匀,制成颗粒,干燥,压制成1000片,包糖衣或薄膜衣;或压制成500片,包薄膜衣,即得。清热解毒、泻火通便。用于三焦热盛所致的目赤肿痛、口鼻生疮、咽喉肿痛、牙龈肿痛、心烦口渴、尿黄便秘。口服,一次4片,一日2次,每粒装0.3g。
现有技术中,尚未有三***在提取制备方面采用超临界技术的报道,而采用打粉等方法,工艺粗糙、复杂、落后,杂质多,导致患者用量过大,不方便服用,严重影响了本品在临床上应用。
发明内容
发明目的:为了解决上述问题,本发明的目的在于提供一种三***的制备方法及三***在制备抑制小鼠软骨ATDC5细胞增殖药物中的应用。
技术方案:本发明的目的是通过如下的方案实现的:
一种三***的制备方法,由大黄300g、盐酸小檗碱5g、黄芩浸膏21g作为原料药制成,所述的方法由下列步骤组成:以上三味,黄芩浸膏系取黄芩,加水煎煮三次,第一次1.5小时,第二次1小时,第三次40分钟,合并煎液,滤过,滤液用盐酸调节pH值至1~2,静置1小时,取沉淀,用水洗涤使pH值至5~7,烘干,粉碎成细粉,取黄芩浸膏21g,另取大黄300g,粉碎成细粉,与黄芩浸膏细粉合并,加入到CO2超临界萃取器中,乙醇作为夹带剂,夹带剂占总萃取溶剂的体积百分比为4-6%,萃取压力15-30MPa,温度30-50℃,CO2流量1-3m1/g生药·min,萃取时间150-180min,得超临界萃取物,粉碎成细粉,添加盐酸小檗碱5g,制粒,压片,每片重0.3g。
所述三***的制备方法中CO2超临界萃取夹带剂占总萃取溶剂的体积百分比为5%。
所述三***的制备方法中CO2超临界萃取的萃取压力20MPa,温度40℃,CO2流量2ml/g生药·min,萃取时间160min。
所述三***在制备抑制小鼠软骨ATDC5细胞增殖药物中的应用,三***由大黄300g、盐酸小檗碱5g、黄芩浸膏21g作为原料药制成,制备方法由下列步骤组成:以上三味,黄芩浸膏系取黄芩,加水煎煮三次,第一次1.5小时,第二次1小时,第三次40分钟,合并煎液,滤过,滤液用盐酸调节pH值至1~2,静置1小时,取沉淀,用水洗涤使pH值至5~7,烘干,粉碎成细粉,取黄芩浸膏21g,另取大黄300g,粉碎成细粉,与黄芩浸膏细粉合并,加入到CO2超临界萃取器中,乙醇作为夹带剂,夹带剂占总萃取溶剂的体积百分比为4-6%,萃取压力15-30MPa,温度30-50℃,CO2流量1-3m1/g生药·min,萃取时间150-180min,得超临界萃取物,粉碎成细粉,添加盐酸小檗碱5g,制粒,压片,每片重0.3g。
所述三***在制备抑制小鼠软骨ATDC5细胞增殖药物中的应用,制备方法中所述CO2超临界萃取夹带剂占总萃取溶剂的体积百分比为5%。
所述三***在制备抑制小鼠软骨ATDC5细胞增殖药物中的应用,制备方法中所述CO2超临界萃取的萃取压力20MPa,温度40℃,CO2流量2ml/g生药·min,萃取时间160min。
现有技术中,三***一次3片,一日3次,每片0.3g,采用本发明制备成的三***每片0.3g,但含有的药材量比原来多,在同样服用量的情况下具有更多活性成分。
具体实施方式
以下通过实施例形式,对本发明的上述内容再作进一步的详细说明,但不应将此理解为本发明上述主题的范围仅限于以下的实例,凡基于本发明上述内容所实现的技术均属于本发明的范围。
实施例1:取大黄300g、盐酸小檗碱5g、黄芩浸膏21g,以上三味,黄芩浸膏系取黄芩,加水煎煮三次,第一次1.5小时,第二次1小时,第三次40分钟,合并煎液,滤过,滤液用盐酸调节pH值至1,静置1小时,取沉淀,用水洗涤使pH值至7,烘干,粉碎成细粉,取黄芩浸膏21g,另取大黄300g,粉碎成细粉,与黄芩浸膏细粉合并,加入到CO2超临界萃取器中,乙醇作为夹带剂,夹带剂占总萃取溶剂的体积百分比为4%,萃取压力30MPa,温度30℃,CO2流量3m1/g生药·min,萃取时间150min,得超临界萃取物,粉碎成细粉,添加盐酸小檗碱5g,制粒,压片,每片重0.3g。
实施例2:取大黄300g、盐酸小檗碱5g、黄芩浸膏21g,以上三味,黄芩浸膏系取黄芩,加水煎煮三次,第一次1.5小时,第二次1小时,第三次40分钟,合并煎液,滤过,滤液用盐酸调节pH值至2,静置1小时,取沉淀,用水洗涤使pH值至5,烘干,粉碎成细粉,取黄芩浸膏21g,另取大黄300g,粉碎成细粉,与黄芩浸膏细粉合并,加入到CO2超临界萃取器中,乙醇作为夹带剂,夹带剂占总萃取溶剂的体积百分比为6%,萃取压力15MPa,温度50℃,CO2流量1m1/g生药·min,萃取时间180min,得超临界萃取物,粉碎成细粉,添加盐酸小檗碱5g,制粒,压片,每片重0.3g。
实施例3:取大黄300g、盐酸小檗碱5g、黄芩浸膏21g,以上三味,黄芩浸膏系取黄芩,加水煎煮三次,第一次1.5小时,第二次1小时,第三次40分钟,合并煎液,滤过,滤液用盐酸调节pH值至1.5,静置1小时,取沉淀,用水洗涤使pH值至6,烘干,粉碎成细粉,取黄芩浸膏21g,另取大黄300g,粉碎成细粉,与黄芩浸膏细粉合并,加入到CO2超临界萃取器中,乙醇作为夹带剂,夹带剂占总萃取溶剂的体积百分比为5%,萃取压力25MPa,温度40℃,CO2流量2m1/g生药·min,萃取时间160min,得超临界萃取物,粉碎成细粉,添加盐酸小檗碱5g,制粒,压片,每片重0.3g。
实施例4:三***抑制小鼠软骨ATDC5细胞增殖的实验研究资料
1实验材料
1.1实验用细胞株:小鼠软骨ATDC5细胞,南京医科大学实验室细胞库,DMEM+10%FBS常规培养。
1.2实验药物:研究药物:本发明三***:按实施例3方法制备。药液储液:称取100mg三***内容物,混悬于5ml无水乙醇中,0.2μm滤器过滤,500μldoff管分装,-20℃存储,同时0.2μm滤器过滤无水乙醇以备对照组之用。
1.3实验试剂:DMEM(GIBCO公司Cat.No.12100-061Lot.No.758137);胎牛血清(浙江天杭生物科技有限公司Lot.No.100419);NaHCO3(上海久亿化学试剂有限公司Cat.No.11810-033Lot.No.1088387);Trypsin(AMRESCO公司批号:2010/04);EDTA(AMRESCO公司批号:2009/10);PenicillinGSodiumSalt(AMRESCO公司批号:2010242);StreptomycinSulfate(AMRESCO公司批号:2010382);无水乙醇(南京化学试剂有限公司批号:080310182);MTT(Biosharp批号:0793);PBS(实验室自配);
1.4实验器材:莱卡倒置显微镜(德国Leica型号:DM1L);可见-紫外光微孔板检测仪(美国MD公司型号:SPECTRAMAX190);CO2培养箱(FORMA型号:3111);超净台(苏净集团安泰公司制造型号:SW-CJ-ZFD);纯水仪(美国Spring公司型号:S/N020579);精密移液器(法国吉尔森公司型号:P2);电子天平(德国赛多利斯有限公司型号:BT323S);全自动高压灭菌锅(日本SANYO公司型号:MLS-3020);台式电热鼓风干燥箱(上海精密实验设备公司型号:DHG9123A);冰箱(西门子公司型号:KG18V21TI);液氮罐(CBS型号:2001);低速离心机(上海安亭科学仪器厂型号:KA-1000);0.2μm滤器(MILLIPORE型号:SLGP033RB);10cm培养皿(NEST公司)、96孔培养板(NEST公司);细胞计数板;离心管、移液管、Tips若干。
2实验方法:1)小鼠软骨ATDC5细胞用DMEM+10%FBS于37℃、5%CO2进行常规培养(10cm培养皿),当细胞生长至对数期时,收集细胞,弃去培养液,PBS轻洗3遍,加入3ml0.25%胰蛋白酶-0.04%EDTA,37℃消化2min后,向其中加入5ml完全培养基中和反应,吹打细胞后将其转入离心管中,1000rpm离心5min,调整细胞悬液浓度3×104个/ml。2)将细胞种入96孔培养板中,每孔加入细胞悬液180μl,培养板放入细胞培养箱中(37℃,5%CO2)常规培养。3)根据细胞生长情况,一般长至50%-70%,加入三***溶液,继续培养24h。4)24h后加入20μlMTT溶液(5mg/ml,即0.5%MTT),继续培养4h。5)4h后扣板法倒去上清,用吸水纸轻轻拍干,每孔加入200μl二甲基亚砜,置摇床上低速振荡10min,使结晶物充分溶解。在酶联免疫检测仪490nm处测量各孔的吸光值。6)同时设置本底(不加细胞,只加培养液),对照孔(细胞、相同浓度的药物溶解介质、培养液、MTT、二甲基亚砜),每组设定6个复孔。7)结果以药物对细胞的抑制率表示:细胞增值抑制率(%)=(对照孔OD值-给药孔OD值)/对照孔OD值×100%。实验重复3次。
3实验结果:MTT法实验后统计结果显示,与对照组比较,当剂量达到5mg/ml时,对小鼠软骨ATDC5细胞增殖抑制有差异(P<0.05),剂量在10mg/ml时该差异具有显著性(P<0.01),当剂量达到15-20mg/ml时有极显著性差异(P<0.001)。
表1三***对小鼠软骨ATDC5细胞增殖抑制影响研究
注:与对照组比较,*P<0.01;**P<0.001
4实验结论:三***可以抑制小鼠软骨ATDC5细胞增殖,减少小鼠软骨ATDC5细胞的细胞生长数目,该作用呈剂量依赖性。
Claims (6)
1.一种三***的制备方法,由大黄300g、盐酸小檗碱5g、黄芩浸膏21g作为原料药制成,其特征在于所述的方法由下列步骤组成:以上三味,黄芩浸膏系取黄芩,加水煎煮三次,第一次1.5小时,第二次1小时,第三次40分钟,合并煎液,滤过,滤液用盐酸调节pH值至1~2,静置1小时,取沉淀,用水洗涤使pH值至5~7,烘干,粉碎成细粉,取黄芩浸膏21g,另取大黄300g,粉碎成细粉,与黄芩浸膏细粉合并,加入到CO2超临界萃取器中,乙醇作为夹带剂,夹带剂占总萃取溶剂的体积百分比为4-6%,萃取压力15-30MPa,温度30-50℃,CO2流量1-3m1/g生药·min,萃取时间150-180min,得超临界萃取物,粉碎成细粉,添加盐酸小檗碱5g,制粒,压片,每片重0.3g。
2.根据权利要求1所述三***的制备方法,其特征在于所述CO2超临界萃取夹带剂占总萃取溶剂的体积百分比为5%。
3.根据权利要求1所述三***的制备方法,其特征在于所述CO2超临界萃取的萃取压力20MPa,温度40℃,CO2流量2ml/g生药·min,萃取时间160min。
4.三***在制备抑制小鼠软骨ATDC5细胞增殖药物中的应用,其特征在于三***由大黄300g、盐酸小檗碱5g、黄芩浸膏21g作为原料药制成,制备方法由下列步骤组成:以上三味,黄芩浸膏系取黄芩,加水煎煮三次,第一次1.5小时,第二次1小时,第三次40分钟,合并煎液,滤过,滤液用盐酸调节pH值至1~2,静置1小时,取沉淀,用水洗涤使pH值至5~7,烘干,粉碎成细粉,取黄芩浸膏21g,另取大黄300g,粉碎成细粉,与黄芩浸膏细粉合并,加入到CO2超临界萃取器中,乙醇作为夹带剂,夹带剂占总萃取溶剂的体积百分比为4-6%,萃取压力15-30MPa,温度30-50℃,CO2流量1-3m1/g生药·min,萃取时间150-180min,得超临界萃取物,粉碎成细粉,添加盐酸小檗碱5g,制粒,压片,每片重0.3g。
5.根据权利要求4所述三***在制备抑制小鼠软骨ATDC5细胞增殖药物中的应用,其特征在于三***的制备方法中所述CO2超临界萃取夹带剂占总萃取溶剂的体积百分比为5%。
6.根据权利要求4所述三***在制备抑制小鼠软骨ATDC5细胞增殖药物中的应用,其特征在于三***的制备方法中所述CO2超临界萃取的萃取压力20MPa,温度40℃,CO2流量2ml/g生药·min,萃取时间160min。
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