氨基酸类凝胶因子对脂肪油的原位胶凝作用及性质评价
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摘要
为了制备室温下为溶液状态、给药后立即在用药部位相转变形成交联半固体状态的可注射型原位有机凝胶给药系统,本文选择以组织相容性好、可生物降解且能有序自组装的氨基酸衍生物为凝胶因子,研制了体温敏感型原位有机凝胶并对其各项性质进行了评价。
合成了N-月桂酰-l-丙氨酸甲酯(LAM)和N-月桂酰-l-丙氨酸乙酯(LAE)两种氨基酸衍生物类凝胶因子,并通过DSC、IR、MS及~1H-NMR对其结构进行了确证。
为了选择合适的油相和凝胶因子以制备可注射且注射后有良好胶凝能力的原位有机凝胶,分别对LAM和LAE进行了热重分析、最小胶凝浓度的确定和凝胶因子相转变温度的研究。结果表明,LAM和LAE在制备原位有机凝胶的条件下均热稳定,不发生分解；二者最小胶凝浓度分别为43.1mg／mL和100.3mg／mL,LAM的胶凝能力强于LAE；LAM在注射用大豆油中的浓度为20%时,既可实现在给药后立即于用药部位发生从溶胶向凝胶的转变,又能保证其在用药部位维持凝胶状态,为控制药物释放提供了可能。
为了保证原位有机凝胶系统可用于注射给药并研究其性质,对LAM的用量、抗凝剂种类和用量以及凝胶系统的载药能力进行了研究。结果表明,凝胶因子的胶凝起始时间不随LAM用量的增加而缩短,30%LAM在注射用大豆油中胶凝迅速、且胶凝时间短；NMP和EtOH的抗凝效果相同,当其浓度为60%时,可保持系统在室温时为溶液状态,适于注射；油相和抗凝剂对系统的载药能力有影响,而凝胶因子LAM对系统的载药能力无显著影响。
分别评价了原位有机凝胶系统的黏度、安全性、胶凝结构、胶凝过程及其对药物释放的影响等。结果表明,凝胶系统的黏度适宜,生物相容性好,可用于皮下注射；凝胶系统的释药以扩散为主,药物释放随凝胶因子浓度的增加而变慢,随抗凝剂浓度的增加而加快。
全文研究结果表明,以LAM为凝胶因子制备的可注射型原位有机凝胶给药系统是一种极具开发潜质的新型药物控释系统。
To study the in situ forming organogel, which is solution at room temperature, and immediately translating into hemi-solid of gel at body temperature, two kinds of amino acid derivatives'gelator were sythensized in this article. The gelators could be biodegradable and have a good biocompatibility, and they could self-assembly in the solvents. To prepare the in situ forming organogel, the gelling ablity of two gelators was investigated. At last, the novel temperature-sensitive in situ forming organogel was developed and its natures were investigated.
Refer to literature, LAM and LAE were synthesized, which were belonged to amino acid derivatives of gelator. And lauroyl chloride,l-alanine methyl ester and l-alanine ethyl ester were employed as materials. The synthesis products were determined by DSC, IR, MS and 1H-NMR. The melt point of LAM and LAE were 59.0℃and 46.3℃, and the productivity was 86% and 80%, separately.
To select the suitable oil phase and temperature-stable, good gelling ability gel, thermogravity analysize (TGA), Minimum gelation concentration (MGC) and Phase transition temperature (TSG andTGS) were determined. TGA test showed that LAM and LAE would not be reduced weight under environments of organogel preparation. MGC indicated that the gelling ability of LAM was stronger than LAE, and the MGC of LAM and LAE were 43.1mg/mL and 100.3mg/mL, respectively. Phase transition temperature of in situ forming organogel stated that LAM could transit into gel phase from sol phase at body temperature and it would not back into sol phase, when its concentration reached or more than 20%. And LAE could not transit into gel phase from sol phase at body temperature, even when its concentration reached 30%.
To prepare and study the in situ forming organogel, the amount of LAM and anti-gelling solvent, kinds of anti-gelling solvent were inspected. The formulation of in situ forming organogel was:1mL injectable soybean oil,300mg LAM, and 0.6mL NMP (or 0.6 mL EtOH). 50mg FP was loaded on injectable soybean oil, and the max amount of FP loaded on in situ forming organogel was 100mg.
The visicosity of in situ forming organogel were stated that it was suited to inject. FP of in situ forming organogel was released as diffusion pattern at the whole stage of release. Hemolytic study showed that NMP had hemolytic phenomenon, EtOH had not hemolytic phenomenon. Pathology study showed that in situ forming organogel has a good biocompatibility; it could be used for subcutaneous injection.
合成了N-月桂酰-l-丙氨酸甲酯(LAM)和N-月桂酰-l-丙氨酸乙酯(LAE)两种氨基酸衍生物类凝胶因子,并通过DSC、IR、MS及~1H-NMR对其结构进行了确证。
为了选择合适的油相和凝胶因子以制备可注射且注射后有良好胶凝能力的原位有机凝胶,分别对LAM和LAE进行了热重分析、最小胶凝浓度的确定和凝胶因子相转变温度的研究。结果表明,LAM和LAE在制备原位有机凝胶的条件下均热稳定,不发生分解；二者最小胶凝浓度分别为43.1mg／mL和100.3mg／mL,LAM的胶凝能力强于LAE；LAM在注射用大豆油中的浓度为20%时,既可实现在给药后立即于用药部位发生从溶胶向凝胶的转变,又能保证其在用药部位维持凝胶状态,为控制药物释放提供了可能。
为了保证原位有机凝胶系统可用于注射给药并研究其性质,对LAM的用量、抗凝剂种类和用量以及凝胶系统的载药能力进行了研究。结果表明,凝胶因子的胶凝起始时间不随LAM用量的增加而缩短,30%LAM在注射用大豆油中胶凝迅速、且胶凝时间短；NMP和EtOH的抗凝效果相同,当其浓度为60%时,可保持系统在室温时为溶液状态,适于注射；油相和抗凝剂对系统的载药能力有影响,而凝胶因子LAM对系统的载药能力无显著影响。
分别评价了原位有机凝胶系统的黏度、安全性、胶凝结构、胶凝过程及其对药物释放的影响等。结果表明,凝胶系统的黏度适宜,生物相容性好,可用于皮下注射；凝胶系统的释药以扩散为主,药物释放随凝胶因子浓度的增加而变慢,随抗凝剂浓度的增加而加快。
全文研究结果表明,以LAM为凝胶因子制备的可注射型原位有机凝胶给药系统是一种极具开发潜质的新型药物控释系统。
To study the in situ forming organogel, which is solution at room temperature, and immediately translating into hemi-solid of gel at body temperature, two kinds of amino acid derivatives'gelator were sythensized in this article. The gelators could be biodegradable and have a good biocompatibility, and they could self-assembly in the solvents. To prepare the in situ forming organogel, the gelling ablity of two gelators was investigated. At last, the novel temperature-sensitive in situ forming organogel was developed and its natures were investigated.
Refer to literature, LAM and LAE were synthesized, which were belonged to amino acid derivatives of gelator. And lauroyl chloride,l-alanine methyl ester and l-alanine ethyl ester were employed as materials. The synthesis products were determined by DSC, IR, MS and 1H-NMR. The melt point of LAM and LAE were 59.0℃and 46.3℃, and the productivity was 86% and 80%, separately.
To select the suitable oil phase and temperature-stable, good gelling ability gel, thermogravity analysize (TGA), Minimum gelation concentration (MGC) and Phase transition temperature (TSG andTGS) were determined. TGA test showed that LAM and LAE would not be reduced weight under environments of organogel preparation. MGC indicated that the gelling ability of LAM was stronger than LAE, and the MGC of LAM and LAE were 43.1mg/mL and 100.3mg/mL, respectively. Phase transition temperature of in situ forming organogel stated that LAM could transit into gel phase from sol phase at body temperature and it would not back into sol phase, when its concentration reached or more than 20%. And LAE could not transit into gel phase from sol phase at body temperature, even when its concentration reached 30%.
To prepare and study the in situ forming organogel, the amount of LAM and anti-gelling solvent, kinds of anti-gelling solvent were inspected. The formulation of in situ forming organogel was:1mL injectable soybean oil,300mg LAM, and 0.6mL NMP (or 0.6 mL EtOH). 50mg FP was loaded on injectable soybean oil, and the max amount of FP loaded on in situ forming organogel was 100mg.
The visicosity of in situ forming organogel were stated that it was suited to inject. FP of in situ forming organogel was released as diffusion pattern at the whole stage of release. Hemolytic study showed that NMP had hemolytic phenomenon, EtOH had not hemolytic phenomenon. Pathology study showed that in situ forming organogel has a good biocompatibility; it could be used for subcutaneous injection.
引文
[1]魏刚,徐晖,郑俊民.原位凝胶的形成机制及在药物控制释放领域的应用[J].中国药学杂志,2003,38(8)：564-568.
[2]刘志东,赵诤.原委凝胶传递系统的研究进展[J].中外健康文摘临床医药版,2008,5(3)：18-20.
[3]魏培,邓树海,李凌冰,徐丽洒,宋燕青.原位凝胶缓释给药系统的研究进展[J].中国医药工业杂志,2007,38(12)：890-894.
[4]刘志东,赵诤.原委凝胶传递系统的研究进展[J].2008,5(3)：18-20.
[5]魏培,邓树海,李凌冰,徐丽洒,宋燕青,张四喜.原位凝胶缓释给药系统的研究进展[J].中国生化药物杂志,2007,28(5)：356-359.
[6]董吉,蒋曙光,平其能.注射型原位凝胶植入剂的研究进展[J].药学进展,2007,31：110-113.
[7]Francois P, Aude M, Anne-claude C, et al. First report on the efficacy of L-alanine-based in situ-forming implants for the long-term parenteral delivery of drugs [J]. Journal of controlled release, 2005,108:433-441.
[8]杨亚江,崔文瑾.能使有机溶剂凝胶化的凝胶因子研究进展[J].有机化学,2001,9：632-639.
[9]Anda V, Jean-christophe L. Organogels and their use in drug delivery-A review [J]. Journal of controlled release,2008,125:179-192.
[10]杨亚江,崔文瑾.能使有机溶剂凝胶化的凝胶因子研究进展[J]．有机化学,2001,9：632-639.
[11]侯仲轲,陈立功,薛福华,宋健.有机低分子凝胶因子[J].化学通报,2005,9：643-649.
[12]MADER K, BITTNER B, LI Y, et al. Monitoring microviscosity and microacidity of the albumin microenvironment inside degrading microparticles from poly(lactide-co-glycolide) (PLG) or ABA-triblock polymers containing hydrophobic poly(lactide-co-glycolide) A blocks and hydrophilic poly(ethyleneoxide) B blocks[J]. Pharm.Res,1998,15:787-793.
[13]Kang G D, Cheon S H, Khang G, et al. Thermosensitive poly (organophosphazene) hydrogels for a controlled drug delivery [J]. Eur J Pharm Biopharm,2006,63(3):340-346.
[14]董吉,蒋曙光,平其能.注射型原位凝胶植入剂的研究进展[J].药学进展,2007,31：110-113．
[15]Anda V, Jean-christophe L. Organogels and their use in drug delivery-A review [J]. Journal of controlled release,2008,125:179-192.
[16]倪沛洲.有机化学[M].人民卫生出版社,第五版.P387.
[17]倪沛洲.有机化学[M].人民卫生出版社,第五版.P387.
[18]Aude M, Michel L, Anne-claude C, et al Characterization and biocompatibility of organogels based on L-alanine for parenteral drug delivery implants [J]. Biomaterials,2005,26:6242-6253.
[19]Roman L, Zbigniew P, Bozena S. Glucofuranose derivatives as a library for designing and investigating low molecular mass organogelators[J]. Tetrahedron,2005,61:10122-10128.
[20]KAZUHIRO Y, YUSUKE T, NORIHIRO M, et al. self-assembly of carbazole-containing gelators: alignment of the chromophore in fibrous aggregates [J]. Tetrahedron,2007,63:7358-7365.
[21]彭瑾,张倩,苏鹏.PLA／PLGA及其嵌段共聚物就地成形植入装置的研究进展[J].上海生物医学工程,2005,26(1)：51-55.
[22]DONG W Y, KORBER M, et al. Stability of poly(D,L-lactide-co-glycolide) and leuprolide acetate in in-situ forming drug delivery systems[J]. Journal of controlled release,2006,115:158-167.
[23]郑爱敏,刘红梅,黄开勋.可注射原位给药体系在蛋白类药物给药中的应用[J].北京生物医学工程,2006,5：556-559.
[24]ANDERSON J M. Biological responses to materials [J]. Ann Rev Mater Res 2001,31:81-110.
[25]ANDERSON J M, SHIVE M S. Biodegradation and biocompatibility of PLA and PLGA microspheres [J]. Adv Drug Deliv Res 1997,28:5-24.
[26]ANDERSON J M. In vivo biocompatibility of implantable delivery system and biomaterials [J]. Eur J Pharm Biopharm 1994,40:1-8.
[27]Schwartz JB, Simonelli AP Higuchi WI. Drug release from wax matrics and analysis of data with first-order kinetics and with diffusion-controlled model [J].J Pharm Sci.1968,57 (2):274-277.
[28]Dredan J, Antal I, Racz I. Evaluation of mathematical models describing drug release from lipophilic matrices [J].Int J Pharm.1996,145:61-64.
[29]Higuchi T. Mechianism of sustained-action medication. Theoretical analysis of rate of release of solid drugs dispersed in solid matrices [J]. J Pharm. Sci.1963,52(12):1145-1149.
[30]Baker RW, Lonsdale HK, In A.C. Controlled release of biologically active agents. Plenum, New York,1974,15-71.
[31]Peppas N, Analysis of fickian and non-fickian drugs drug release from polymers [J].pharm acta helv, 1985,60:110-111.
[32]Oliver G, Erwin S, Seiji S. Gelators for organic liquids based on self-assembly:a new facet of supramolecular and combinatorial chemistry [J]. Current opinion in colloid and interface science, 2002,7:148-156.
[33]彭瑾,张倩,苏鹏.PLA／PLGA及其嵌段共聚物就地成形植入装置的研究进展[J].上海生物医学工程,2005,26(1)：51-55.
[34]DONG W Y, KORBER M, et al. Stability of poly(D,L-lactide-co-glycolide) and leuprolide acetate in in-situ forming drug delivery systems[J]. Journal of controlled release,2006,115:158-167.
[35]R.G. Strickley. Solubilizing excipients in oral and injectable formulations. Pharm. Res.21 (2004) 201-230.
[36]郑爱敏,刘红梅,黄开勋.可注射原位给药体系在蛋白类药物给药中的应用[J].北京生物医学工程,2006,5：556-559.
[2]刘志东,赵诤.原委凝胶传递系统的研究进展[J].中外健康文摘临床医药版,2008,5(3)：18-20.
[3]魏培,邓树海,李凌冰,徐丽洒,宋燕青.原位凝胶缓释给药系统的研究进展[J].中国医药工业杂志,2007,38(12)：890-894.
[4]刘志东,赵诤.原委凝胶传递系统的研究进展[J].2008,5(3)：18-20.
[5]魏培,邓树海,李凌冰,徐丽洒,宋燕青,张四喜.原位凝胶缓释给药系统的研究进展[J].中国生化药物杂志,2007,28(5)：356-359.
[6]董吉,蒋曙光,平其能.注射型原位凝胶植入剂的研究进展[J].药学进展,2007,31：110-113.
[7]Francois P, Aude M, Anne-claude C, et al. First report on the efficacy of L-alanine-based in situ-forming implants for the long-term parenteral delivery of drugs [J]. Journal of controlled release, 2005,108:433-441.
[8]杨亚江,崔文瑾.能使有机溶剂凝胶化的凝胶因子研究进展[J].有机化学,2001,9：632-639.
[9]Anda V, Jean-christophe L. Organogels and their use in drug delivery-A review [J]. Journal of controlled release,2008,125:179-192.
[10]杨亚江,崔文瑾.能使有机溶剂凝胶化的凝胶因子研究进展[J]．有机化学,2001,9：632-639.
[11]侯仲轲,陈立功,薛福华,宋健.有机低分子凝胶因子[J].化学通报,2005,9：643-649.
[12]MADER K, BITTNER B, LI Y, et al. Monitoring microviscosity and microacidity of the albumin microenvironment inside degrading microparticles from poly(lactide-co-glycolide) (PLG) or ABA-triblock polymers containing hydrophobic poly(lactide-co-glycolide) A blocks and hydrophilic poly(ethyleneoxide) B blocks[J]. Pharm.Res,1998,15:787-793.
[13]Kang G D, Cheon S H, Khang G, et al. Thermosensitive poly (organophosphazene) hydrogels for a controlled drug delivery [J]. Eur J Pharm Biopharm,2006,63(3):340-346.
[14]董吉,蒋曙光,平其能.注射型原位凝胶植入剂的研究进展[J].药学进展,2007,31：110-113．
[15]Anda V, Jean-christophe L. Organogels and their use in drug delivery-A review [J]. Journal of controlled release,2008,125:179-192.
[16]倪沛洲.有机化学[M].人民卫生出版社,第五版.P387.
[17]倪沛洲.有机化学[M].人民卫生出版社,第五版.P387.
[18]Aude M, Michel L, Anne-claude C, et al Characterization and biocompatibility of organogels based on L-alanine for parenteral drug delivery implants [J]. Biomaterials,2005,26:6242-6253.
[19]Roman L, Zbigniew P, Bozena S. Glucofuranose derivatives as a library for designing and investigating low molecular mass organogelators[J]. Tetrahedron,2005,61:10122-10128.
[20]KAZUHIRO Y, YUSUKE T, NORIHIRO M, et al. self-assembly of carbazole-containing gelators: alignment of the chromophore in fibrous aggregates [J]. Tetrahedron,2007,63:7358-7365.
[21]彭瑾,张倩,苏鹏.PLA／PLGA及其嵌段共聚物就地成形植入装置的研究进展[J].上海生物医学工程,2005,26(1)：51-55.
[22]DONG W Y, KORBER M, et al. Stability of poly(D,L-lactide-co-glycolide) and leuprolide acetate in in-situ forming drug delivery systems[J]. Journal of controlled release,2006,115:158-167.
[23]郑爱敏,刘红梅,黄开勋.可注射原位给药体系在蛋白类药物给药中的应用[J].北京生物医学工程,2006,5：556-559.
[24]ANDERSON J M. Biological responses to materials [J]. Ann Rev Mater Res 2001,31:81-110.
[25]ANDERSON J M, SHIVE M S. Biodegradation and biocompatibility of PLA and PLGA microspheres [J]. Adv Drug Deliv Res 1997,28:5-24.
[26]ANDERSON J M. In vivo biocompatibility of implantable delivery system and biomaterials [J]. Eur J Pharm Biopharm 1994,40:1-8.
[27]Schwartz JB, Simonelli AP Higuchi WI. Drug release from wax matrics and analysis of data with first-order kinetics and with diffusion-controlled model [J].J Pharm Sci.1968,57 (2):274-277.
[28]Dredan J, Antal I, Racz I. Evaluation of mathematical models describing drug release from lipophilic matrices [J].Int J Pharm.1996,145:61-64.
[29]Higuchi T. Mechianism of sustained-action medication. Theoretical analysis of rate of release of solid drugs dispersed in solid matrices [J]. J Pharm. Sci.1963,52(12):1145-1149.
[30]Baker RW, Lonsdale HK, In A.C. Controlled release of biologically active agents. Plenum, New York,1974,15-71.
[31]Peppas N, Analysis of fickian and non-fickian drugs drug release from polymers [J].pharm acta helv, 1985,60:110-111.
[32]Oliver G, Erwin S, Seiji S. Gelators for organic liquids based on self-assembly:a new facet of supramolecular and combinatorial chemistry [J]. Current opinion in colloid and interface science, 2002,7:148-156.
[33]彭瑾,张倩,苏鹏.PLA／PLGA及其嵌段共聚物就地成形植入装置的研究进展[J].上海生物医学工程,2005,26(1)：51-55.
[34]DONG W Y, KORBER M, et al. Stability of poly(D,L-lactide-co-glycolide) and leuprolide acetate in in-situ forming drug delivery systems[J]. Journal of controlled release,2006,115:158-167.
[35]R.G. Strickley. Solubilizing excipients in oral and injectable formulations. Pharm. Res.21 (2004) 201-230.
[36]郑爱敏,刘红梅,黄开勋.可注射原位给药体系在蛋白类药物给药中的应用[J].北京生物医学工程,2006,5：556-559.