基于氨基酸偶合显色的丝胶蛋白定量分析
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摘要
为了解决在蚕丝精练实际生产中监测练减率的问题,利用丝胶蛋白中氨基酸残基与重氮盐偶合显色的特征反应来推算练减率。文章首先绘制了氨基酸模拟丝胶蛋白偶合显色的吸光度—氨基酸浓度标准工作曲线,并且研究了重氮盐的稳定性、偶合反应pH值对偶合吸光度的影响,以及丝胶蛋白碱水解后的沉淀物成分。结果表明,在0℃时重氮盐能稳定保存,但温度升高会导致其不稳定;偶合反应pH值控制在10左右时,偶合效率较高。由于精练残液碱水解后与重氮盐偶合显色推导的练减率较实际练减率低3%～4%,丝胶蛋白碱水解沉淀物不含丝胶成分,因此加上实验误差,能够反映实际生产中蚕丝精练的脱胶情况。
In order to realize monitoring degumming loss percentage in real production of silk refining,degumming loss percentage was reckoned based on the characteristic reaction of coupling coloration of amino acid residues in sericin with diazonium salt. In this study,the standard curve of absorbance-amino acid concentration of amino acid mimicking protein-coupled coloration was drawn first,and researches were conducted on stability of diazonium salt,effect of coupling reaction pH on coupled absorbance,and precipitation composition produced via sericin basic hydrolysis. The results show that diazonium salt can be stably stored at 0 ℃,but it will become instable as the temperature rises; coupling efficiency remains high as long as coupling reaction p H is controlled around 10. The degumming loss percentage reckoned after basic hydrolysis of bast liquor and coupling coloration of diazonium salt is lower than the real value by 3% to 4%,basic hydrolysis precipitation of sericin contains no glue-like component,and there exist experimental errors. In view of this,the results can reflect degumming of silk refining in actual production reliably.
In order to realize monitoring degumming loss percentage in real production of silk refining,degumming loss percentage was reckoned based on the characteristic reaction of coupling coloration of amino acid residues in sericin with diazonium salt. In this study,the standard curve of absorbance-amino acid concentration of amino acid mimicking protein-coupled coloration was drawn first,and researches were conducted on stability of diazonium salt,effect of coupling reaction pH on coupled absorbance,and precipitation composition produced via sericin basic hydrolysis. The results show that diazonium salt can be stably stored at 0 ℃,but it will become instable as the temperature rises; coupling efficiency remains high as long as coupling reaction p H is controlled around 10. The degumming loss percentage reckoned after basic hydrolysis of bast liquor and coupling coloration of diazonium salt is lower than the real value by 3% to 4%,basic hydrolysis precipitation of sericin contains no glue-like component,and there exist experimental errors. In view of this,the results can reflect degumming of silk refining in actual production reliably.
引文
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[2]李绍峰,虞晓华.桑蚕雌雄蚕丝氨基酸成分分析[J].江苏蚕业,2012,34(2):12-15.LI Shaofeng,YU Xiaohua.Analysis of amino acid composition of silkworm male silkworm[J].Jiangsu Sericulture,2012,34(2):12-15.
[3]SLUYTERMAN L A.The effect of oxygen upon the micro determination of histidine with the aid of the pauly reaction[J].Biochimica Et Biophysica Acta,1960,38(2):218-221.
[4]SENGUPTA S,BHATTACHARYYA S.Aqueous heck reaction of amino acid derived arenediazonium salts:semisynthetic modification of phenylalanine and tyrosine[J].Tetrahedron Letters,1995,36(25):4475-4478.
[5]郭旭,翁连进,甘林火,等.Pauly试剂显色测定L-酪氨酸的研究[J].化工科技,2006,14(5):4-7.GUO Xu,WENG Lianjin,GAN Linhuo,et al.Study on determination of the L-tyrosine with pauly reagent[J].Science&Technology in Chemical Industry,2006,14(5):4-7.
[6]郑今欢,邵建中,刘今强.蚕丝丝素蛋白中酪氨酸分布的初探[J].纺织学报,2001,22(6):2,7-9.ZHENG Jinhuan,SHAO Jianzhong,LIU Jinqiang.Studies on the distribution of tyrosine in silk fibroin[J].Journal of Textile Research,2001,22(6):2,7-9.
[7]林宇燕.组氨酸测定方法的建立[J].职业与健康,2008,24(8):734-736.LIN Yuyan.Establishment of a method for histidine determination[J].Occupation and Health,2008,24(8):734-736.
[8]何俊锋,张伟国.发酵液中L-组氨酸的定性定量测定[J].食品与生物技术学报,2006(4):56-58,112.HE Junfeng,ZHANG Weiguo.Quantitative and quantitative determination of L-histidine in fermentation broth[J].Journal of Food Science and Biotechnology,2006(4):56-58,112.
[9]郑今欢,周岚,邵建中.蚕丝丝素中色氨酸含量及其在丝素纤维中的径向分布研究[J].高分子学报,2005(2):161-166.ZHENG Jinhuan,ZHOU Lan,SHAO Jianzhong.Studies on content and distribution of tryptophan in silk fiber[J].Acta Polymerica Sinica,2005(2):161-166.
[10]何铠君,王宗乾,陈维国,等.三氟乙酸法制备邻硝基苯胺重氮盐及其稳定性分析[J].浙江理工大学学报,2015,33(1):16-19.HE Kaijun,WANG Zongqian,CHEN Weiguo,et al.Preparation of O-nitroaniline diazonium salt with trifluoroacetic acid and its stability analysis[J].Journal of Zhejiang SciTech University,2015,33(1):16-19.
[11]苏彦雷,刘建芳,林克江,等.量子化学计算法对合成中4个芳香重氮盐稳定性的分析[J].海峡药学,2013,25(8):186-188.SU Yanlei,LIU Jianfang,LIN Kejiang,et al.The analysis of stability of four aromatic diazonium salts in synthetic process with Quantum chemistry calculation method[J].Strait Pharmaceutical Journal,2013,25(8):186-188.
[12]叶曦雯,李引,牛增元,等.23种致癌芳香胺的显色法测定[J].纺织学报,2012,33(12):58-63.YE Xiwen,LI Yin,NIU Zengyuan,et al.Chromogenic method determination of 23 kinds of carcinogenic aromatic amines[J].Journal of Textile Research,2012,33(12):58-63.