新型壳聚糖/抗坏血酸复合物的制备与应用性能研究
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摘要
本论文针对L-抗坏血酸(VC)在应用和处理中容易被氧化的缺点,利用壳聚糖在生物领域的特性,制备了两种新型的壳聚糖/VC复合物——壳聚糖L-抗坏血酸盐、2-O-壳聚糖基抗坏血酸。同时针对长期服用抗坏血酸容易在胃液中转化为草酸导致结石的问题,制备了乙基纤维素包衣的壳聚糖包裹抗坏血酸微胶囊,以期达到既能提高VC的稳定性,又能实现VC在肠液中的缓释效果。
本论文系统地研究了这些壳聚糖/VC复合物的合成条件,通过抗坏血酸含量、产率、抗氧化性等分析测试确定了性能最优的制备条件,利用核磁共振光谱、红外光谱、扫描电镜考察产物的结构。结果表明:壳聚糖/L-抗坏血酸盐和2-O-壳聚糖基抗坏血酸具有较好的水溶性,壳聚糖大分子结构的存在,降低了各种氧化剂对VC的破坏作用,使VC在贮存和使用过程中不容易被氧化。同时,由于VC本身有着优良的抗菌性能,所以它也进一步增强了壳聚糖的抗菌能力和对血脂的吸附能力。
本论文还利用反相乳化交联法,制备了壳聚糖/L-抗坏血酸微球,并在其外包覆乙基纤维素制成微胶囊。系统研究了壳聚糖分子量、水油比、反应时间、交联剂用量等因素对微球制备和VC释放行为的影响。结果表明:所制备的微胶囊不仅在肠液中具有显著的缓释效果,并且大大降低了VC在胃中的释放,达到肠溶的目的。
For the disadvantage of L-ascorbic acid (vitamin C, VC) that it is easily to be oxidized during the treatment and process, two new kinds of derivative---chitosan ascorbate and 2-O-chitosan-ascorbic acid were studied. For the oxalic acid is formed in high yield from VC in gastric juice, the chitosan-VC micro sphere was prepared and then was microencapsulated into ethylic cellulose (EC).
By designing appropriate experiments, the conditions of synthesizing chitosan-ascorbic acid complex were studied. The analysis of VC content, output rate, solubility helped us to get the optimal condition of synthesizing. The 13C NMR assignments, infrared spectrum permitted the identification of the structure of these complex. After the function test, it is found that chitosan ascorbate and 2-O-chitosan-ascorbic acid have good water-soluable ability. The existent of chitosan macro-molecular structure prevents the oxygen from destroying VC, so VC is to be protected. Since VC has good anti-bacteria ability and absorbability to serum lipoprotein, it enhances the same ability of chitosan.
At the same time, L-ascorbic acid, used as a model drug, was efficiently entrapped within chitosan micro spheres using reversed-phase emulsion cross linking reaction and then was microencapsulated into ethylic cellulose. The morphology and release properties of microcapsules were tested. The influential factors of preparation conditions included molecular weight of chitosan, ratio of water and oil, reaction time, concentration of cross linker were discussed. The results of in vitro release studies show that the microcapsules prepared in this article are stable in gastric juice and can realize sustained release in intestine juice.
本论文系统地研究了这些壳聚糖/VC复合物的合成条件,通过抗坏血酸含量、产率、抗氧化性等分析测试确定了性能最优的制备条件,利用核磁共振光谱、红外光谱、扫描电镜考察产物的结构。结果表明:壳聚糖/L-抗坏血酸盐和2-O-壳聚糖基抗坏血酸具有较好的水溶性,壳聚糖大分子结构的存在,降低了各种氧化剂对VC的破坏作用,使VC在贮存和使用过程中不容易被氧化。同时,由于VC本身有着优良的抗菌性能,所以它也进一步增强了壳聚糖的抗菌能力和对血脂的吸附能力。
本论文还利用反相乳化交联法,制备了壳聚糖/L-抗坏血酸微球,并在其外包覆乙基纤维素制成微胶囊。系统研究了壳聚糖分子量、水油比、反应时间、交联剂用量等因素对微球制备和VC释放行为的影响。结果表明:所制备的微胶囊不仅在肠液中具有显著的缓释效果,并且大大降低了VC在胃中的释放,达到肠溶的目的。
For the disadvantage of L-ascorbic acid (vitamin C, VC) that it is easily to be oxidized during the treatment and process, two new kinds of derivative---chitosan ascorbate and 2-O-chitosan-ascorbic acid were studied. For the oxalic acid is formed in high yield from VC in gastric juice, the chitosan-VC micro sphere was prepared and then was microencapsulated into ethylic cellulose (EC).
By designing appropriate experiments, the conditions of synthesizing chitosan-ascorbic acid complex were studied. The analysis of VC content, output rate, solubility helped us to get the optimal condition of synthesizing. The 13C NMR assignments, infrared spectrum permitted the identification of the structure of these complex. After the function test, it is found that chitosan ascorbate and 2-O-chitosan-ascorbic acid have good water-soluable ability. The existent of chitosan macro-molecular structure prevents the oxygen from destroying VC, so VC is to be protected. Since VC has good anti-bacteria ability and absorbability to serum lipoprotein, it enhances the same ability of chitosan.
At the same time, L-ascorbic acid, used as a model drug, was efficiently entrapped within chitosan micro spheres using reversed-phase emulsion cross linking reaction and then was microencapsulated into ethylic cellulose. The morphology and release properties of microcapsules were tested. The influential factors of preparation conditions included molecular weight of chitosan, ratio of water and oil, reaction time, concentration of cross linker were discussed. The results of in vitro release studies show that the microcapsules prepared in this article are stable in gastric juice and can realize sustained release in intestine juice.
引文
[1]黎观红,翟明任,晏向华等,VC的营养和应用研究进展,粮食与饲料工业,2001,4:28~32
[2]张骁,束梅英,郭建新,VC产销现状和市场前景展望,中国药房,1997,1:10~12
[3]彭司勋,药物化学,北京:化学工业出版社,1988:365~368
[4]王境岩,朱圣庚,徐长法,生物化学(第三版),高等教育出版社,2002:461~463
[5]闻芝梅,陈君石,现代营养学,北京:人民卫生出版社,1988:140~153
[6]丁峰,施雪枫,顾勇等,抗坏血酸透析液改善维持性血液透析患者氧化应激的初步研究,复旦学报(医学版),2006,33(2):161~166
[7]孔宝华,冯一冰,抗坏血酸在食品中的应用,食品工业,1995,2:51~52
[8]马姓,张道波,王克行,饲料添加包膜VC和VC磷酸醋镁对中国对虾幼体的影响,海洋与湖沼,1999,3:275~276
[9]张华峰,冯惠勇,VC产业现状与发展,中国食品添加剂,2004,2:47~48
[10]张辉,刘晓平,VC钙盐合成工艺的改进,辽宁化工,2003,32(5):188~189
[11]王三永,李晓光,李春荣等,VC磷酸醋镁的合成研究,食品与机械,2001,2:34~35
[12] David F, Plainfield NJ, Process for preparing ascorbyl-3-phosphate and salts, US, 3671549, 1972
[13]金显春,王锦堂,维生察C多聚磷酸酯的合成研究,精细石油化工,2003,4:27~29
[14]蔡力创,吕积国,高立贞,L-抗坏血酸-6棕榈酸酯的合成研究,江西科学,1999,3(17):11~17
[15]陶剑虹,VC系列产品开发概况及发展趋势,化学医药工业信息,1997,13(1):11~14
[16]韦开昕,维C银翘薄膜包衣工艺的研究,医学文选,1999,1:56~60
[17]张裕中,原料配方与操作参数对挤压饲料品质的影响,粮食与饲料工业, 2002,1:11~14
[18] Sudarshan NR, Hoover DG., Knorr D, Antibacterial action of chitosan, Food Biotechnology, 1992, 6(3): 257~261
[19]张静,杨洪强,几丁质及其衍生物的生物活性与在农业中的应用,植物学通报, 2003,20(2):178~180
[20] Muzzarelli RA, Biochemical significance of exogenous chitins and chitosans in animals and patients, Carbohydr Polym, 1993, (20): 7~16
[21] Nishimura K, Minami S, Application of chitin and chitosan for biomaterials, Biotechnol Genet Eng Rev, 1995, 13: 383~420
[22]张建湘,汤健,徐斌等,壳聚糖棒材的组织相容性和安全性评价,生物医学工程学杂志,1996,13(4):293~297
[23] Roller S, Covill N, The antimicrobial properties of chitosan in mayonnaise and mayonnaise-based shrimp salads, Food Prot, 2000, 63(2): 202~209
[24] Rabea EI, Badawy ME, Stevens CV, et al, Chitosan as antimicrobial agent: applications and mode of action, Biomacromolecules, 2003, 4(6): 1457~1465
[25]宋琳,壳聚糖及其衍生物与大肠杆菌DNA、蛋白质的作用关系研究:[硕士学位论文],天津;天津大学,2007
[26]顾云,王绿娅,张颖,几丁聚糖降低血清脂质的临床观察,中国海洋药物,2000,19(30):45~47
[27]吕朋,李八方,夏兰,壳聚糖在医药保健中的应用,中国海洋药物,2001,20(5):30~34
[28]魏菁,甲壳素及其衍生物在眼科的应用,洛阳医专学报,2000,18(3):258~260
[29]黄俊明,吴丽明,陈瑞仪等,甲壳素和壳聚糖对小鼠免疫调节的研究,广东卫生免疫,1999,25(1):4~6
[30]阳元娥,罗发兴,壳聚糖及其衍生物在医药中的应用,中国医药工业杂志,2002,33(8): 408~411
[31] Ikinci G, Senel S, Akincibay H, et al, Effect of chitosan on a periodontal pathogen Porphyromonas gingivalis, Int J Pharm, 2002, 20 (1): 121~127
[32] Choi BK, Kim KY, Yoo YJ, et al, In Vitro Antimicrobial Activity of a Chitooligosaccharide Mixture Against Actinobacillus Actinomycetemcomitans and Streptococcus Mutans, Int J Antimicrob Agents, 2001, 18(6): 553~557
[33] Muzzarelli R, Biagini G, Pugnaloni A, et al, Reconstruction of parodontal tissue with Chitosan, Biomaterials, 1989, 10(9): 598~603
[34] Uomalainen K, Sorsa T, Lindy O, et al, Hypochlorous acid induced activation of human neutrophil and gingival crevicular fluid collagenase can be inhibited by ascorbate, Scand J Dent Res, 1991, 99(5): 397~405
[35] Yamamoto I, Muto N, Murakami K, et al, L-ascorbic acid alpha-glucoside formed by regioselective transglucosylation with rat intestinal and rice seed alpha-glucosidases: its improved stability and structure determination, Chem Pharm Bull(Tokyo), 1990, 38(11): 3020~3023
[36] Mandai T, Yoneyama M, Sakai S, et al, The crystal structure and physicochemical properties of L-ascorbic acid 2-glucoside, Carbohydr Res, 1992, 232(2): 197~205
[37] Wakamiya H, Suzuki E, Yamamoto I, et al, Vitamin C activity of 2-O-alpha-D- glucopyranosyl-L-ascorbic acid in guinea Pigs, J Nutr Sci Vitaminol(Tokyo), 1992, 38(3): 235~245
[38] Yamamoto I, Suga S, Mitoh Y, et al, Antiscorbutic activity of L-ascorbic acid 2-glucoside and its availability as a vitamin C supplement in normal rats and guinea pigs, J Pharmacobiodyn, 1990, 13(11): 688~695
[39] Yamamoto I, Muto N, Murakami K, et al, Collagen synthesis in human skin fibroblasts is stimulated by a stable form of ascorbate, 2-O-alpha-D- glucopyranosyl-L-ascorbic acid, J Nutr, 1992, l22(4): 871~877
[40] Fujinami Y, Tai A, Yamamoto I, Radical scavenging activity against 1,1-diphenyl-2-picrylhydrazyl of ascorbic acid 2-glucoside(AA-2G) and 6-acyl- AA-2G, Chem Pharm Bull(Tokyo), 2001, 49(5): 642~644
[41] Kumano Y, Sakamoto T, Egawa M, et al, In vitro and in vivo prolonged biological activities of novel vitamin C derivative, 2-O-alpha-D-glucopyranosyl- L-ascorbic acid(AA-2G), in cosmetic fields, J Nutr, 1998, 44(3): 345 ~359
[42] Kunihiko Nakata,Atsushi Yamamoto, Yasunori Natta, X-ray Studay of Chitosan L- and D-Ascorbates, Chem. Mater, 1996, 8: 2349~2351
[43] Zoldners J, Kiseleva T, Kaiminsh I, Influence of ascorbic acid on the stability of chitosan solutions, Carbohydrate Polymers, 2005, 60: 215~218
[44] Mullan BA, Youns LS, Fee H, et al, Ascorbic acid reduces blood pressure and arterial stiffness in type 2 diabetes Hyperte-nsion, J Nutr, 2002, 40: 804
[45]孟祥光,李建梅等,CTAB对H2O2氧化抗坏血酸反应动力学的影响,物理化学学报,2005,21(3):283~286
[46]黄熙,李方,朱孔营,H2O2氧化降解壳聚糖的动力学,天津理工大学学报,2005,21(4):71~73
[47]卢玲,何炳林,袁直,LDL的结构研究进展,离子交换与吸附,2001,17(5):363~368
[48]董庆年,红外光谱法,北京:化学工业出版社,1979
[49] Dolphin D, Wick AE, Tabulation of Infrared Spectral Data, New York: John Wiley & sons, 1977
[50]西尔弗斯坦,巴斯勒,莫里尔,有机化合物光谱鉴定,北京:科学出版社,1988:232~236
[51]蒋大挺,甲壳素,北京:化学工业出版社, 2003
[52]刘程,表面活性剂应用大全,北京:化学工业出版社,1992:50～51
[2]张骁,束梅英,郭建新,VC产销现状和市场前景展望,中国药房,1997,1:10~12
[3]彭司勋,药物化学,北京:化学工业出版社,1988:365~368
[4]王境岩,朱圣庚,徐长法,生物化学(第三版),高等教育出版社,2002:461~463
[5]闻芝梅,陈君石,现代营养学,北京:人民卫生出版社,1988:140~153
[6]丁峰,施雪枫,顾勇等,抗坏血酸透析液改善维持性血液透析患者氧化应激的初步研究,复旦学报(医学版),2006,33(2):161~166
[7]孔宝华,冯一冰,抗坏血酸在食品中的应用,食品工业,1995,2:51~52
[8]马姓,张道波,王克行,饲料添加包膜VC和VC磷酸醋镁对中国对虾幼体的影响,海洋与湖沼,1999,3:275~276
[9]张华峰,冯惠勇,VC产业现状与发展,中国食品添加剂,2004,2:47~48
[10]张辉,刘晓平,VC钙盐合成工艺的改进,辽宁化工,2003,32(5):188~189
[11]王三永,李晓光,李春荣等,VC磷酸醋镁的合成研究,食品与机械,2001,2:34~35
[12] David F, Plainfield NJ, Process for preparing ascorbyl-3-phosphate and salts, US, 3671549, 1972
[13]金显春,王锦堂,维生察C多聚磷酸酯的合成研究,精细石油化工,2003,4:27~29
[14]蔡力创,吕积国,高立贞,L-抗坏血酸-6棕榈酸酯的合成研究,江西科学,1999,3(17):11~17
[15]陶剑虹,VC系列产品开发概况及发展趋势,化学医药工业信息,1997,13(1):11~14
[16]韦开昕,维C银翘薄膜包衣工艺的研究,医学文选,1999,1:56~60
[17]张裕中,原料配方与操作参数对挤压饲料品质的影响,粮食与饲料工业, 2002,1:11~14
[18] Sudarshan NR, Hoover DG., Knorr D, Antibacterial action of chitosan, Food Biotechnology, 1992, 6(3): 257~261
[19]张静,杨洪强,几丁质及其衍生物的生物活性与在农业中的应用,植物学通报, 2003,20(2):178~180
[20] Muzzarelli RA, Biochemical significance of exogenous chitins and chitosans in animals and patients, Carbohydr Polym, 1993, (20): 7~16
[21] Nishimura K, Minami S, Application of chitin and chitosan for biomaterials, Biotechnol Genet Eng Rev, 1995, 13: 383~420
[22]张建湘,汤健,徐斌等,壳聚糖棒材的组织相容性和安全性评价,生物医学工程学杂志,1996,13(4):293~297
[23] Roller S, Covill N, The antimicrobial properties of chitosan in mayonnaise and mayonnaise-based shrimp salads, Food Prot, 2000, 63(2): 202~209
[24] Rabea EI, Badawy ME, Stevens CV, et al, Chitosan as antimicrobial agent: applications and mode of action, Biomacromolecules, 2003, 4(6): 1457~1465
[25]宋琳,壳聚糖及其衍生物与大肠杆菌DNA、蛋白质的作用关系研究:[硕士学位论文],天津;天津大学,2007
[26]顾云,王绿娅,张颖,几丁聚糖降低血清脂质的临床观察,中国海洋药物,2000,19(30):45~47
[27]吕朋,李八方,夏兰,壳聚糖在医药保健中的应用,中国海洋药物,2001,20(5):30~34
[28]魏菁,甲壳素及其衍生物在眼科的应用,洛阳医专学报,2000,18(3):258~260
[29]黄俊明,吴丽明,陈瑞仪等,甲壳素和壳聚糖对小鼠免疫调节的研究,广东卫生免疫,1999,25(1):4~6
[30]阳元娥,罗发兴,壳聚糖及其衍生物在医药中的应用,中国医药工业杂志,2002,33(8): 408~411
[31] Ikinci G, Senel S, Akincibay H, et al, Effect of chitosan on a periodontal pathogen Porphyromonas gingivalis, Int J Pharm, 2002, 20 (1): 121~127
[32] Choi BK, Kim KY, Yoo YJ, et al, In Vitro Antimicrobial Activity of a Chitooligosaccharide Mixture Against Actinobacillus Actinomycetemcomitans and Streptococcus Mutans, Int J Antimicrob Agents, 2001, 18(6): 553~557
[33] Muzzarelli R, Biagini G, Pugnaloni A, et al, Reconstruction of parodontal tissue with Chitosan, Biomaterials, 1989, 10(9): 598~603
[34] Uomalainen K, Sorsa T, Lindy O, et al, Hypochlorous acid induced activation of human neutrophil and gingival crevicular fluid collagenase can be inhibited by ascorbate, Scand J Dent Res, 1991, 99(5): 397~405
[35] Yamamoto I, Muto N, Murakami K, et al, L-ascorbic acid alpha-glucoside formed by regioselective transglucosylation with rat intestinal and rice seed alpha-glucosidases: its improved stability and structure determination, Chem Pharm Bull(Tokyo), 1990, 38(11): 3020~3023
[36] Mandai T, Yoneyama M, Sakai S, et al, The crystal structure and physicochemical properties of L-ascorbic acid 2-glucoside, Carbohydr Res, 1992, 232(2): 197~205
[37] Wakamiya H, Suzuki E, Yamamoto I, et al, Vitamin C activity of 2-O-alpha-D- glucopyranosyl-L-ascorbic acid in guinea Pigs, J Nutr Sci Vitaminol(Tokyo), 1992, 38(3): 235~245
[38] Yamamoto I, Suga S, Mitoh Y, et al, Antiscorbutic activity of L-ascorbic acid 2-glucoside and its availability as a vitamin C supplement in normal rats and guinea pigs, J Pharmacobiodyn, 1990, 13(11): 688~695
[39] Yamamoto I, Muto N, Murakami K, et al, Collagen synthesis in human skin fibroblasts is stimulated by a stable form of ascorbate, 2-O-alpha-D- glucopyranosyl-L-ascorbic acid, J Nutr, 1992, l22(4): 871~877
[40] Fujinami Y, Tai A, Yamamoto I, Radical scavenging activity against 1,1-diphenyl-2-picrylhydrazyl of ascorbic acid 2-glucoside(AA-2G) and 6-acyl- AA-2G, Chem Pharm Bull(Tokyo), 2001, 49(5): 642~644
[41] Kumano Y, Sakamoto T, Egawa M, et al, In vitro and in vivo prolonged biological activities of novel vitamin C derivative, 2-O-alpha-D-glucopyranosyl- L-ascorbic acid(AA-2G), in cosmetic fields, J Nutr, 1998, 44(3): 345 ~359
[42] Kunihiko Nakata,Atsushi Yamamoto, Yasunori Natta, X-ray Studay of Chitosan L- and D-Ascorbates, Chem. Mater, 1996, 8: 2349~2351
[43] Zoldners J, Kiseleva T, Kaiminsh I, Influence of ascorbic acid on the stability of chitosan solutions, Carbohydrate Polymers, 2005, 60: 215~218
[44] Mullan BA, Youns LS, Fee H, et al, Ascorbic acid reduces blood pressure and arterial stiffness in type 2 diabetes Hyperte-nsion, J Nutr, 2002, 40: 804
[45]孟祥光,李建梅等,CTAB对H2O2氧化抗坏血酸反应动力学的影响,物理化学学报,2005,21(3):283~286
[46]黄熙,李方,朱孔营,H2O2氧化降解壳聚糖的动力学,天津理工大学学报,2005,21(4):71~73
[47]卢玲,何炳林,袁直,LDL的结构研究进展,离子交换与吸附,2001,17(5):363~368
[48]董庆年,红外光谱法,北京:化学工业出版社,1979
[49] Dolphin D, Wick AE, Tabulation of Infrared Spectral Data, New York: John Wiley & sons, 1977
[50]西尔弗斯坦,巴斯勒,莫里尔,有机化合物光谱鉴定,北京:科学出版社,1988:232~236
[51]蒋大挺,甲壳素,北京:化学工业出版社, 2003
[52]刘程,表面活性剂应用大全,北京:化学工业出版社,1992:50～51