盐酸青藤碱关节腔注射微球的研究
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摘要
青藤碱(SM)是从青风藤中提取得生物碱单体，具有抗炎镇痛、抗风湿及免疫抑制作用，临床主要用于类风湿性关节炎的治疗。本文以SM模型药物，明胶为载体材料，采用乳化-交联法制备可用于关节腔注射的微球。
建立了紫外分光光度法用于SM-GMS的含量测定、体外释放度考察、及稳定性的研究。在处方前研究中，对药物的性质进行了考察，为处方及制备工艺的摸索提供了依据。以外观、粒径、载药量等性质为考察指标，通过单因素考察和正交设计确定了最佳处方工艺。
采用扫描电子显微镜、激光力度测定仪和差示扫描量热仪等仪器等分析检测方法，对微球的各方面性质进行考察，结果表明微球的平均粒径为6．40μm，粒度分布为正态分布；微球表面光滑圆整，粘连较少；载药量为10．4％，包封率为73．3％；影响因素实验表明，微球稳定性良好，温度和湿度对外观有一定影响。
对微球的体外释放结果进行拟合，结果表明研制的注射用盐酸青藤碱微球体外释放符合Higuchi方程模型，经模拟后Higuchi方程为：Q=13．248 t~(1／2)+9．0044，其中R=0．9707，t_(50)=9．58 h；由体外释放曲线可以看出，SM-GMS具有明显的缓释作用。
建立了兔血浆中药物的HPLC分析方法。SM-GMS皮下注射后血药浓度达峰时间为24h，SM-GMS可维持一定的血药浓度达到50小时以上，体内药动学研究表明，微球制剂与普通注射剂相比具有明显的缓释作用。
局部组织刺激性的考察结果表明，空白微球组和生理盐水组相比没有明显的病理生理变化，说明该制剂的组织相容性良好，长期用药不会给关节各组织造成损害。
Sinomenine Hydrochloride(SM), a pure alkaloid extracted from traditional Chinese medicine-Caulis Sinomenine, which has the Effectiveness of anti-inflammatory analgesic, anti-rheumatic and immune inhibition, has been mainly used for rheumatoid arthritis in clinical therapeutics. In this study, Sinomenine Hydrochloride gelatin microspheres (SM-GMS) were prepared by emulsification linkage for intra-articular injection.
The method of UV was set up to determine the concentration of SM in the preparation and in vitro release. The formulation and process of the SM-GMS were studied according to investigating the basic physical characters of the drug. Applying appearance, particle size and drug-loading rate of microspheres to evaluate the quality of microspheres, the best formulation and process was screened by signal factor studies and orthogonal test.
The physical and chemical characters of microspheres were evaluated by scanning electron microscope, laser size tester. The appearance was smooth without much adherence, the mean diameter was 6.40μm ,the loading amount was10.4%, the encapsulation efficiency was 73.3%. The stability study indicated that only the appearance of microspheres could be slight influenced by high temperature and high humidity.
The release kinetics of microspheres in vitro could be characterized by Higuchi equation, the equation is Q=13.248 t~(1/2)+9.0044, R=0.9707, t_(50)=9.58 h. From the in vitro release curve it is obvious that the SM-GMS is a delayed-release preparation.
The method of HPLC was developed for the analyzing of drug in rabbits plasma. The T_(max) of SM-GMS and control group were 24h and 4h respectively. The pharmacokinetics characters showed that the preparation of SM-GMS can keep the blood concentration on certain level longer than 50h, so it obviously delayed release.
According to the Local biopsy identified by experts, there is no significantly Pathological and Physiological change between blank microspheres group and the saline group .That means the biocompatibility of the preparation is good, the organizations of the joint can not be damaged under long-term using .
建立了紫外分光光度法用于SM-GMS的含量测定、体外释放度考察、及稳定性的研究。在处方前研究中，对药物的性质进行了考察，为处方及制备工艺的摸索提供了依据。以外观、粒径、载药量等性质为考察指标，通过单因素考察和正交设计确定了最佳处方工艺。
采用扫描电子显微镜、激光力度测定仪和差示扫描量热仪等仪器等分析检测方法，对微球的各方面性质进行考察，结果表明微球的平均粒径为6．40μm，粒度分布为正态分布；微球表面光滑圆整，粘连较少；载药量为10．4％，包封率为73．3％；影响因素实验表明，微球稳定性良好，温度和湿度对外观有一定影响。
对微球的体外释放结果进行拟合，结果表明研制的注射用盐酸青藤碱微球体外释放符合Higuchi方程模型，经模拟后Higuchi方程为：Q=13．248 t~(1／2)+9．0044，其中R=0．9707，t_(50)=9．58 h；由体外释放曲线可以看出，SM-GMS具有明显的缓释作用。
建立了兔血浆中药物的HPLC分析方法。SM-GMS皮下注射后血药浓度达峰时间为24h，SM-GMS可维持一定的血药浓度达到50小时以上，体内药动学研究表明，微球制剂与普通注射剂相比具有明显的缓释作用。
局部组织刺激性的考察结果表明，空白微球组和生理盐水组相比没有明显的病理生理变化，说明该制剂的组织相容性良好，长期用药不会给关节各组织造成损害。
Sinomenine Hydrochloride(SM), a pure alkaloid extracted from traditional Chinese medicine-Caulis Sinomenine, which has the Effectiveness of anti-inflammatory analgesic, anti-rheumatic and immune inhibition, has been mainly used for rheumatoid arthritis in clinical therapeutics. In this study, Sinomenine Hydrochloride gelatin microspheres (SM-GMS) were prepared by emulsification linkage for intra-articular injection.
The method of UV was set up to determine the concentration of SM in the preparation and in vitro release. The formulation and process of the SM-GMS were studied according to investigating the basic physical characters of the drug. Applying appearance, particle size and drug-loading rate of microspheres to evaluate the quality of microspheres, the best formulation and process was screened by signal factor studies and orthogonal test.
The physical and chemical characters of microspheres were evaluated by scanning electron microscope, laser size tester. The appearance was smooth without much adherence, the mean diameter was 6.40μm ,the loading amount was10.4%, the encapsulation efficiency was 73.3%. The stability study indicated that only the appearance of microspheres could be slight influenced by high temperature and high humidity.
The release kinetics of microspheres in vitro could be characterized by Higuchi equation, the equation is Q=13.248 t~(1/2)+9.0044, R=0.9707, t_(50)=9.58 h. From the in vitro release curve it is obvious that the SM-GMS is a delayed-release preparation.
The method of HPLC was developed for the analyzing of drug in rabbits plasma. The T_(max) of SM-GMS and control group were 24h and 4h respectively. The pharmacokinetics characters showed that the preparation of SM-GMS can keep the blood concentration on certain level longer than 50h, so it obviously delayed release.
According to the Local biopsy identified by experts, there is no significantly Pathological and Physiological change between blank microspheres group and the saline group .That means the biocompatibility of the preparation is good, the organizations of the joint can not be damaged under long-term using .
引文
[1]王华燕，胡文祥，刘接卿等．药物治疗风湿性关节炎的研究近况[J]．中国医药导刊，2007，9，50(3) 237-24．
[2]朱德湘．正清风痛宁抗风湿的作用机理浅谈[J]．湖南中医杂志，1994，10(4)：54-55．
[3]李乐，张彩玲，宋必卫等．青藤碱的药理研究与临床应用[J]．Traditional Chinese Drug Research & Clinical Pharmacology，2006，17(4)，310-313．
[4]王岩，周莉玲，李锐青风藤的研究进展[J]．中药材，2002；25(s)：209-211
[5]张欣．毛青藤治疗类风湿关节炎1524例临床分析甘肃中医．1996；9(3)：22-23．
[6]付绍首，张士善，李蕴山等．青藤碱的药理作用11．毒性及一般药理作用[J]．药学学报，1963， 10(11)：673-676．
[7]刘启德，梁美蓉，欧卫平等．青藤碱时辰药代动力学研究[J]．中药新药与临床药理，1995，6(1)： 23-26．
[8]催福德主编．药剂学[M]．中国医药科技出版社，2002，8．
[9]任海霞，朱家壁，汤王月．微球制剂的应用研究进展[J]．西北药学杂志，2007．31(2)96-98
[10]宋群亮，张平．中药微球制剂的应用研究进展[J]．安徽医药2004，9(2)87-88
[11]陆扬．明胶微球的研究进展[J]．明胶科学与技术,2006,26(2)
[12]钟延强．关节腔注射用氟比洛芬明胶微球的研究．药学学报．1999．34(7)：543-546
[13]李志平，梅兴国．关节腔注射用醋酸地塞米松微球的体内外评价及组织相容性考察[J]．中国药学杂 志，2006，41(17)
[14]刘迎春，赵兵，王国清．电位滴定法测定盐酸青藤碱的电离常数．沈阳药科大学学 报，1999，16(3)：217-218．
[15]陆扬．明胶等电点对明胶微球分散度影响的研究．明胶科学与技术[J]．2005，25(4)：203-207：58-74
[16]杨永新，李岩，孙殿甲等．交联固化对明胶微球质量的影响．西北药学杂志[J]．2001，16(1)．40-41
[17]付绍萱，张士善，李蕴山等．青藤碱的药理作用Ⅱ毒性及一般药理作用[J]．药学学报，1963，10(11)： 673-676．
[18]张丽峰，张淑秋．盐酸青藤碱控释微丸的制备与含量测定．中国药物与临床[J]．2006，6(10)：748-750．
[19]马廷升，李高，刘志华．盐酸青藤碱不同晶形在人体生物利用度的研究．中国药业[J]．2006，15(12)：3-4．
[20]施杰明．高效液相色谱法测定盐酸青藤碱缓释微丸的含量．广东药学院学报．2007，23(2)：164-165．
[21]赵铁，李媛，姜清华．盐酸青藤碱缓释胶囊的家犬体内药动学和相对生物利用度研究．中国药学杂??志．2007，42(5)：371-373
[22]国家药典委员会．中国人民共和国药典[S]．2005年版．二部．北京：化学工业出版社．2005：120．
[23]郑俊民．可注射缓释制剂[M]．化学工业出版社，2005，6．
[24]李岩．盐酸去氢骆驼碱动脉栓塞微球明胶微球的研究．沈阳药科大学博士学位论文．2003，3
[25]陈维．盐酸青藤碱脉冲释放片的研究．沈阳药科大学硕士学位论文．2003，5
[26] Elisabetta Esposito, Rita Cortesi and Claudio Nastruzzi.Gelatin microspheres: influence of preparationparameters and thermal treatment on chemico-physical and biopharmaceutical properties[J]. Biomaterals1996. 17(20): 2009-2020.
[27] Hao-Ji Wei, Hang-Hsing Yang. Gelatin microspheres encapsulated with a anonpeptide angiogenicagent,ginsenoside Rgl. for intramyocardial injection in a rat model with infarcted myocardium[J].Journal of Controlled Release, 2007, 120: 27-34.
[28] R. Narayani, K. Panduranga Rao. Gelatinmicrosphere cocktails of different sizes controlled release ofanticancer drugs forthe[J]. Int J of Pharmaceutics. 1996, 143: 255-258.
[29] Suzuki,Makoto,Ishigakiete.Inira-artieularpr, arationforthetreatmentofarthxoPathy.UnitedStatesPatent, 6,428, 804.August6, 2002.
[29] Lu yang Studies on the influence of gelatins'IEP on the dispersivity of gelatin microspheres[J]. TheScience and Technology of Gelatin 2005, 25(4).
[30] E. Esposito, C. Pastesini, R. Cortesi, Controlled release of 1-/3-D- arabinofuranosylcytosine(ara-C)from hydrophilic gelatin microspheres: in vitro studies. International Journal of Pharmaceutics117(1995) 151-158
[31] Elisabetta Esposito, Rita Cortesi and Claudio Nastruzzi. Gelatin microspheres: influence ofpreparationparameters and thermaltreatment on chemico-physical andbiopharmaceutical properties. Biomaterials199617(20):2009-2020
[32] Norihisa Nitta. Shinichi Ohta, Toyohiko Tanaka. Gelatin microspheres:Initial clinical experience forthetranscatheter arterial embolization. European Journal of Radiology.2007,12(9):21-26
[33] Jian Wang. Yasuhiko Tabata. Kazuhiro Morimoto. Aminated gelatin microspheres as a nasal deliverysystem for peptide drugs:Evaluation of in vitro release and in vivo insulin absorption in rats. Journal ofControlled Release 2006 113:31-37
[34] Yi-You Huang. Tze-Wen Chung, Tzeng-Wen Tzeng. A method using biodegradablepolylactides:polyethylene glycolfor drug release with reduced initial burst. International Journal ofPharmaceutics .1999 182(94):93-100
[35] Katsuya Kawai, Shigehiko Suzuki, Yasuhiko Tabata, Accelerated tissue regeneration through incorporation of basic fibroblast growth factor-impregnated gelatin microspheres into artificial dermis Biomaterials 21(2000) 489-499
[36] R. Cortesi.C. Nastruzzi~*. S.S. Davis. Sugar cross-linked gelatin for controlled release microspheres and disks. Biomaterials 19 (1998) 1641(?)1649
[2]朱德湘．正清风痛宁抗风湿的作用机理浅谈[J]．湖南中医杂志，1994，10(4)：54-55．
[3]李乐，张彩玲，宋必卫等．青藤碱的药理研究与临床应用[J]．Traditional Chinese Drug Research & Clinical Pharmacology，2006，17(4)，310-313．
[4]王岩，周莉玲，李锐青风藤的研究进展[J]．中药材，2002；25(s)：209-211
[5]张欣．毛青藤治疗类风湿关节炎1524例临床分析甘肃中医．1996；9(3)：22-23．
[6]付绍首，张士善，李蕴山等．青藤碱的药理作用11．毒性及一般药理作用[J]．药学学报，1963， 10(11)：673-676．
[7]刘启德，梁美蓉，欧卫平等．青藤碱时辰药代动力学研究[J]．中药新药与临床药理，1995，6(1)： 23-26．
[8]催福德主编．药剂学[M]．中国医药科技出版社，2002，8．
[9]任海霞，朱家壁，汤王月．微球制剂的应用研究进展[J]．西北药学杂志，2007．31(2)96-98
[10]宋群亮，张平．中药微球制剂的应用研究进展[J]．安徽医药2004，9(2)87-88
[11]陆扬．明胶微球的研究进展[J]．明胶科学与技术,2006,26(2)
[12]钟延强．关节腔注射用氟比洛芬明胶微球的研究．药学学报．1999．34(7)：543-546
[13]李志平，梅兴国．关节腔注射用醋酸地塞米松微球的体内外评价及组织相容性考察[J]．中国药学杂 志，2006，41(17)
[14]刘迎春，赵兵，王国清．电位滴定法测定盐酸青藤碱的电离常数．沈阳药科大学学 报，1999，16(3)：217-218．
[15]陆扬．明胶等电点对明胶微球分散度影响的研究．明胶科学与技术[J]．2005，25(4)：203-207：58-74
[16]杨永新，李岩，孙殿甲等．交联固化对明胶微球质量的影响．西北药学杂志[J]．2001，16(1)．40-41
[17]付绍萱，张士善，李蕴山等．青藤碱的药理作用Ⅱ毒性及一般药理作用[J]．药学学报，1963，10(11)： 673-676．
[18]张丽峰，张淑秋．盐酸青藤碱控释微丸的制备与含量测定．中国药物与临床[J]．2006，6(10)：748-750．
[19]马廷升，李高，刘志华．盐酸青藤碱不同晶形在人体生物利用度的研究．中国药业[J]．2006，15(12)：3-4．
[20]施杰明．高效液相色谱法测定盐酸青藤碱缓释微丸的含量．广东药学院学报．2007，23(2)：164-165．
[21]赵铁，李媛，姜清华．盐酸青藤碱缓释胶囊的家犬体内药动学和相对生物利用度研究．中国药学杂??志．2007，42(5)：371-373
[22]国家药典委员会．中国人民共和国药典[S]．2005年版．二部．北京：化学工业出版社．2005：120．
[23]郑俊民．可注射缓释制剂[M]．化学工业出版社，2005，6．
[24]李岩．盐酸去氢骆驼碱动脉栓塞微球明胶微球的研究．沈阳药科大学博士学位论文．2003，3
[25]陈维．盐酸青藤碱脉冲释放片的研究．沈阳药科大学硕士学位论文．2003，5
[26] Elisabetta Esposito, Rita Cortesi and Claudio Nastruzzi.Gelatin microspheres: influence of preparationparameters and thermal treatment on chemico-physical and biopharmaceutical properties[J]. Biomaterals1996. 17(20): 2009-2020.
[27] Hao-Ji Wei, Hang-Hsing Yang. Gelatin microspheres encapsulated with a anonpeptide angiogenicagent,ginsenoside Rgl. for intramyocardial injection in a rat model with infarcted myocardium[J].Journal of Controlled Release, 2007, 120: 27-34.
[28] R. Narayani, K. Panduranga Rao. Gelatinmicrosphere cocktails of different sizes controlled release ofanticancer drugs forthe[J]. Int J of Pharmaceutics. 1996, 143: 255-258.
[29] Suzuki,Makoto,Ishigakiete.Inira-artieularpr, arationforthetreatmentofarthxoPathy.UnitedStatesPatent, 6,428, 804.August6, 2002.
[29] Lu yang Studies on the influence of gelatins'IEP on the dispersivity of gelatin microspheres[J]. TheScience and Technology of Gelatin 2005, 25(4).
[30] E. Esposito, C. Pastesini, R. Cortesi, Controlled release of 1-/3-D- arabinofuranosylcytosine(ara-C)from hydrophilic gelatin microspheres: in vitro studies. International Journal of Pharmaceutics117(1995) 151-158
[31] Elisabetta Esposito, Rita Cortesi and Claudio Nastruzzi. Gelatin microspheres: influence ofpreparationparameters and thermaltreatment on chemico-physical andbiopharmaceutical properties. Biomaterials199617(20):2009-2020
[32] Norihisa Nitta. Shinichi Ohta, Toyohiko Tanaka. Gelatin microspheres:Initial clinical experience forthetranscatheter arterial embolization. European Journal of Radiology.2007,12(9):21-26
[33] Jian Wang. Yasuhiko Tabata. Kazuhiro Morimoto. Aminated gelatin microspheres as a nasal deliverysystem for peptide drugs:Evaluation of in vitro release and in vivo insulin absorption in rats. Journal ofControlled Release 2006 113:31-37
[34] Yi-You Huang. Tze-Wen Chung, Tzeng-Wen Tzeng. A method using biodegradablepolylactides:polyethylene glycolfor drug release with reduced initial burst. International Journal ofPharmaceutics .1999 182(94):93-100
[35] Katsuya Kawai, Shigehiko Suzuki, Yasuhiko Tabata, Accelerated tissue regeneration through incorporation of basic fibroblast growth factor-impregnated gelatin microspheres into artificial dermis Biomaterials 21(2000) 489-499
[36] R. Cortesi.C. Nastruzzi~*. S.S. Davis. Sugar cross-linked gelatin for controlled release microspheres and disks. Biomaterials 19 (1998) 1641(?)1649