结构蛋白—猪皮胶原的提取及其理化特性的表征
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摘要
以猪皮为原料,采用醋酸和胃蛋白酶相结合的方法提取胶原,以提取率为指标,进行提取条件优化,并通过紫外光谱、红外光谱、黏度法和荧光光谱等分析方法对提取物的化学结构、变性温度、特征黏度、等电点等理化特性进行了表征。结果表明:猪皮胶原的最佳提取工艺为:料液比1:4、m胃蛋白酶:m猪皮=1:80、p H值2~3、温度4℃、时间60h,此时胶原提取率为18.87%。提取物的结构为猪皮I型胶原,变性温度为37.5℃,特性黏度为4.0230d L/g,等电点p I=6.0。
In this paper, collagen was extracted from porcine skin with the combination method of acetic acid and pepsin, using extraction yield of collagen as an index to optimize the extracting conditions, the chemical structure, the denaturation temperature, the characteristics of the viscosity, the physical and chemical properties such as isoelectric point of the extract were characterized by ultraviolet spectrum, infrared spectrum, viscosity method and fluorescence spectrum analysis method. The results indicated that the optimum conditions of extracting collagen as follow: the mass ratio of pepsin to porcine skin with a ratio of material to liquid of 1:4(m/V) under the condition of 4℃ for 60 hours, the mass ratio of pepsin to porcine skin with a ratio of 1:80(mg/g), p H value was 2 to 3, under the condition of the process, collagen yield was about 18.87%. The structure of the extract was type I collagen and denaturation temperature, isoelectric point and intrinsic viscosity were 37.5℃, 6.0 and 4.0230 d L/g, respectively.
In this paper, collagen was extracted from porcine skin with the combination method of acetic acid and pepsin, using extraction yield of collagen as an index to optimize the extracting conditions, the chemical structure, the denaturation temperature, the characteristics of the viscosity, the physical and chemical properties such as isoelectric point of the extract were characterized by ultraviolet spectrum, infrared spectrum, viscosity method and fluorescence spectrum analysis method. The results indicated that the optimum conditions of extracting collagen as follow: the mass ratio of pepsin to porcine skin with a ratio of material to liquid of 1:4(m/V) under the condition of 4℃ for 60 hours, the mass ratio of pepsin to porcine skin with a ratio of 1:80(mg/g), p H value was 2 to 3, under the condition of the process, collagen yield was about 18.87%. The structure of the extract was type I collagen and denaturation temperature, isoelectric point and intrinsic viscosity were 37.5℃, 6.0 and 4.0230 d L/g, respectively.
引文
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[24]陈培基,李来好,李刘冬.小杂鱼脱脂技术的研究[J].湛江海洋大学学报,2003,23(4):21-22.
[25]Duan Rui,Zhang JJ Konno K,et al.Analysis and improvement of stability of pepsin-solubilized collagen from skin of carp(Cyprinus carpio)[J].Chemical Papers,2012,66(7):636-641.
[26]唐世华,张宁.荧光光度法研究明胶等电点[J].明胶科学与技术,2000,20(2):69-73.
[27]Jia YJ,Wang HB,Wang HY,et al.Biochemical properties of skin collagens isolated from black carp(Mylopharyngodon piceus)[J].Food Sci and Biotechnol,2012,21:1585-1592.
[28]Manning MC.Use of infrared spectroscopy to monitor protein structure and stability[J].Expert Rev Proteomics,2005,2(5):731-743.
[29]Chang MC,Tanaka J.FT-IR study for hydroxyapatite/collagen nanocomposite cross-linked by glutaraldehyde[J].Biomaterials,2002,23(24):4811-4818.
[30]Pati F,Datta P,Adhikkari B,et al.Collagen scaffolds derived from fresh water fish origin and their biocompatibility[J].Biomed.Mater Res.Part A,2012,100A:1068-1079.
[31]Guzzi Plepis AMD,Goissis G,DAS-GUPTA D K.Dielectric and pyroelectric characterization of anionic and native collagen[J].Polym Eng Sci,1996,36:2932-2938.
[32]钱曼,武贤壮,邱承光,等.热力法和酶解法提取鱼鳞胶原蛋白的工艺及性质研究[J].食品工业科技,2007,10(28):70-72.
[2]刘龙天.胶原蛋白三螺旋结构及热稳定性的研究[D].北京:北京协和医学院&中国医学科学院,2009.
[3]Mario Hiram UM,Joe Luis AM,Maribel PJ,et al.Jumbo squid(Dosidicus gigas)mantle collagen:extration,characterization,and potential application in the prepation of chitosan-collagen biofilms[J].Bioresourece Technol,2010,101:4212-4219.
[4]Leah CA,Erin Z,Bernardo PR,et al.Review:Guide to collagen characterization for biomaterial studies[J].J Biomed Materials Res.Part B:Applied Biomaterials,2008,87(B):264-285.
[5]Chi HL,Anuj S,Yugyung L.Biomedical applications of collagen[J].International Journal of Pharmaceutics,2001,221(1-2):1-22.
[6]Cliche S,Amiot J,Avezard C,et al.Extration and characterization of collagen with or without telopeptides from chicken skin[J].Poultry Sci,2003,82:503-509.
[7]蓝蔚青,李燕,周培根.猪皮胶原蛋白的提取与应用浅析[J].肉类研究,2006,(3):44-48.
[8]Singh P,Benjakul S,Maqsood S,et al.Isolation and characterisation of collagen extracted from the skin of striped catfish(Pangasianodon hypophthalmus)[J].Food Chem,2011,124:97-105.
[9]王学川,任龙芳,强涛涛,等.胶原蛋白的研究进展及其在化妆品中的应用[J].日用化学工艺,2005,35(6):388-392.
[10]Liu HY,Zhao L,Guo SD,et al.Modification of fish skin collagen film and absorption property of tannic acid[J].Food Sci Technol,2014,51(6):1102-1109.
[11]Park SH,Song TJ,Tae SB,et al.Comparative analysis of collagens extracted from different animal sources for application of cartilage tissue engineering[J].International J Precision Eng and manuf,2012,13(11):2059-2066.
[12]Maeda M,Tanis,Sano A,et al.Microstructure and release characteristics of the minipellet,a collagen based drug delivery system for controlled release of protein drugs[J].J Controlled Release,1999,62(3):313-324.
[13]Ramshsw JAM,Werkmeister JA,Glatteur V.Collagen based biomaterials[J].Biotechnol Rev,1995,13:336-382.
[14]李贺,郑庚修,王秋芬,等.生物医学材料胶原蛋白的研究进展(II)-胶原的性质及其在生物医学领域中的应用[J].中国皮革,2006,35(3):27-31.
[15]杨贤庆,张帅,郝淑贤,等.罗非鱼皮胶原蛋白的提取条件优化及性质[J].食品科学,2009,30(16):106-110.
[16]傅燕凤,沈月新,杨承刚,等.淡水鱼鱼皮胶原蛋白的提取[J].上海水产大学学报,2004,13(2):146-150.
[17]Keiji Y,Mariko T,Daiki H,et al.Prepation and dynamic viscoelasticity characterization of alkali-solubilized collagen from shark skin[J].J Agric Food Chem,2000,48(3):685-690.
[18]Ka JL,Hee YP,YEON K K,et al.Biochemical characterization of collagen from the starfish asterias amurensis[J].J Korean Soc Apply Biol Chem,2009,52(3):221-226.
[19]Peck LK,Mat DM.Isolation and characterization of collagen from the skin of malaysian catfish(hybrid clarias sp.)[J].J Korean Soc Applied Biol Chem,2013,56:441-450.
[20]Liu HY,Li D,Guo SD.Extration and properties of gelatin from channel catfish(letalurus punetaus)skin[J].LWT-Food Sci and Technol,2008,41(3):414-419.
[21]周玉惠,叶正涛,肖立芳,等.猪皮胶原蛋白的提取及其结构表征[J].武汉:湖北大学,2008.
[22]JEONG HS,Jayachandran V,KIim SK.Isolation and characterization of collagen from marine fish(Thunnus obesus)[J].Biotechnol and Bioprocess Eng.2013,18:1185-1191.
[23]Wu GP,Wang XM,Lin LP,et al.Isolation and characterization of pepsin-solubilized collagen from the skin of black carp(Mylopharyngdon piceus)[J].Advances in Biosci and Biotechnol,2014,5:642-650.
[24]陈培基,李来好,李刘冬.小杂鱼脱脂技术的研究[J].湛江海洋大学学报,2003,23(4):21-22.
[25]Duan Rui,Zhang JJ Konno K,et al.Analysis and improvement of stability of pepsin-solubilized collagen from skin of carp(Cyprinus carpio)[J].Chemical Papers,2012,66(7):636-641.
[26]唐世华,张宁.荧光光度法研究明胶等电点[J].明胶科学与技术,2000,20(2):69-73.
[27]Jia YJ,Wang HB,Wang HY,et al.Biochemical properties of skin collagens isolated from black carp(Mylopharyngodon piceus)[J].Food Sci and Biotechnol,2012,21:1585-1592.
[28]Manning MC.Use of infrared spectroscopy to monitor protein structure and stability[J].Expert Rev Proteomics,2005,2(5):731-743.
[29]Chang MC,Tanaka J.FT-IR study for hydroxyapatite/collagen nanocomposite cross-linked by glutaraldehyde[J].Biomaterials,2002,23(24):4811-4818.
[30]Pati F,Datta P,Adhikkari B,et al.Collagen scaffolds derived from fresh water fish origin and their biocompatibility[J].Biomed.Mater Res.Part A,2012,100A:1068-1079.
[31]Guzzi Plepis AMD,Goissis G,DAS-GUPTA D K.Dielectric and pyroelectric characterization of anionic and native collagen[J].Polym Eng Sci,1996,36:2932-2938.
[32]钱曼,武贤壮,邱承光,等.热力法和酶解法提取鱼鳞胶原蛋白的工艺及性质研究[J].食品工业科技,2007,10(28):70-72.
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