用改進(jìn)逆流分配法分離三尖杉酯堿和高三尖杉酯堿

1、    用改進(jìn)逆流分配法分離三尖杉酯堿和高三尖杉酯堿        摘要對(duì)三尖杉酯堿和高三尖杉酯堿混合物的分離利用經(jīng)改進(jìn)的逆流分配法,用經(jīng)pH 5緩沖液飽和的氯仿作流動(dòng)相,經(jīng)氯仿飽和的pH 5磷酸氫二鈉-枸櫞酸作固定相,醋酸乙酯丙酮(62.5)作展開劑,堿性硅膠板作薄層板,經(jīng)對(duì)5批混合物進(jìn)行分離,均分別獲得三尖杉酯堿和高三尖杉酯堿的白色結(jié)晶。各項(xiàng)指標(biāo)檢查均符合標(biāo)準(zhǔn)。關(guān)鍵詞逆流分配,三尖杉酯堿,高三尖杉酯堿 The isolation of harringtonine and

2、homoharringtonine by an advanced countercurrent distribution methodLi Jianhui (Tianya Pharmaceutical Factory,No 187 Military Hospital,Haikou 571159)An advanced countercurrent distribution method was introduced for the isolation of harringtonine and homoharringtonine.In this method,pH 5 buffer satura

3、ted with CHCl3 was used as mobile phase,CHCl3 saturated disodium hydrogenphosphate-citric acid as stationary phase,ethyl acetateacetone (62.5) as spreading agent,alkaline silica gel as thin layer.Based on the former method,dosage was increased and procedure was simplified.Five kinds of compound of h

4、arringtonine and homoharringtonine were isolated successfully by this advanced method.In this way,greater amount of products can be got with better effects and shorter time.It has the practical value for the production of harringtonine and homoharringtonine from experimental production to bulk produ

5、ction.Key wordscountercurrent distribution,harringtonine,homoharringtonine三尖杉酯堿(Harringtonine)和高三尖杉酯堿(Homoharringtonine)是從粗榧科三尖杉屬植物中提取得到的生物堿,為臨床治療白血病之有效藥物1。由于三尖杉酯堿和高三尖杉酯堿為同系物,分離比較困難。以往多用硅膠分配柱層析;或逆流分配分離。前者上樣量少(1250),流程長(zhǎng)(23d),成本高。后者操作較為煩瑣。根據(jù)高三尖杉酯堿結(jié)構(gòu)多一個(gè)“-CH2-”基,親酯性比三尖杉酯堿稍強(qiáng)的性質(zhì),我們利用逆流分配的原理,對(duì)分離方法作了一些改進(jìn),取得

6、比較滿意的結(jié)果,現(xiàn)介紹如下。1材料混合酯堿樣品為本院從海南粗榧中cephalotaxus hainanepsis(紅殼粉)提得;薄層層析板(青島海洋化工廠出品薄層硅膠G0.02mol.L-1氫氧化鈉=12薄板,105烘烤30min);展開劑(醋酸乙酯丙酮=62.5),氧化鋁(上海五四廠中性Al2O3,200300目,活性為級(jí))。2方法和結(jié)果2.1流動(dòng)相(用pH 5緩沖液飽和的氯仿液);固定相(用氯仿飽和的pH 5磷酸氫二鈉-枸櫞酸液)。2.2準(zhǔn)備容量適當(dāng)?shù)姆忠郝┒?520個(gè),取混合酯堿適量溶解于流動(dòng)相中(13)置1號(hào)分液漏斗。2.3加入與流動(dòng)相等體積的固定相,振搖46min;使之混勻,靜置5m

7、in,待兩相完全分層后,放下流動(dòng)相于盛有與流動(dòng)相等體積的固定相的2號(hào)分液漏斗中。2.4將新的流動(dòng)相等體積加入1號(hào)分液漏斗。2.5同時(shí)振搖1和2號(hào)分液漏斗46min,待分層。爾后,將2號(hào)分液漏斗下層流動(dòng)相放入盛有等體積固定相的3號(hào)漏斗里,1號(hào)分液漏斗下層之流動(dòng)相則放入2號(hào)分液漏斗中。2.6將新的流動(dòng)相加入1號(hào)分液漏斗中,然后同時(shí)振搖3個(gè)分液漏斗,按前法同樣操作。如此遞增,可用薄層檢查始端分液漏斗的固定相,用改良Dragendorff試劑顯色,如若薄層還有兩個(gè)斑點(diǎn)則繼續(xù)傳遞下去。致只剩一個(gè)斑點(diǎn)時(shí),可將該分液漏斗中的固定相收集置容器中,用濃氨水堿化,氯仿萃取。萃取液用無水硫酸鈉脫水后,回收氯仿,抽干

8、,得三尖杉酯堿粗品()。當(dāng)傳遞到第10號(hào)漏斗時(shí),可用薄層檢查最前面流動(dòng)相,用高三尖杉酯堿稱品作對(duì)照。如薄層檢查只有一個(gè)斑點(diǎn)時(shí),即可脫水,回收氯仿,收集抽干,得高三尖杉酯堿粗品(),否則繼續(xù)傳遞下去。當(dāng)傳遞至1518號(hào)分液漏斗時(shí),可將薄層檢查還出現(xiàn)兩個(gè)斑點(diǎn)的固定相和流動(dòng)相回收合并抽干,供下次加入分離樣品。附改進(jìn)逆流分配法分離過程簡(jiǎn)將()和()抽干物分別稱重,混入少量層析用氧化鋁分別研勻,按130分別上氧化鋁柱層析,用無水乙醚洗脫。洗脫液大部分回收,剩余濃縮液常溫放置。()析出白色細(xì)柱狀結(jié)晶或無定形粉末,()析出白色方狀結(jié)晶或無定形粉末。用本法分離5批樣品,其中()有4批得到白色結(jié)晶,一批析出無定

9、形粉末;()全部得到高三尖杉酯堿白色結(jié)晶。得率見附表。()各項(xiàng)指標(biāo)符合海南省藥品標(biāo)準(zhǔn)(1993)2,高三尖杉酯堿符合中國(guó)藥典(1995版)標(biāo)準(zhǔn)3?？勺鳛獒槃┑脑?。附表5批混合酯堿分離結(jié)果批次混合酯堿(g)三尖杉酯堿(g)高三尖杉酯堿(g)得率(%)15819.512.354.8212345.228.559.9313246.330.658.2415053.835.359.4516560.439.660.63討論改進(jìn)逆流分配法,每次投料量可以從原來幾克,增加到幾十克甚至幾百克,大大提高了分離效率。且產(chǎn)品質(zhì)量比硅膠分配柱層析分離的質(zhì)量好(以前硅膠分配柱層析所得產(chǎn)品大部分是白色無定形粉末,很少得到結(jié)晶),分離流程只需要在12d內(nèi)就可以完成。作者單