蚕丝蛋白及某些染料的电化学研究与应用
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摘要
天然蛋白质材料蚕丝蛋白(丝素、丝胶)及丝素的水解产品丝素肽,以其优异的生物相容性、良好的抗氧化性和富含氨基酸而在生物医学材料、食品、化状品、医药工业、纺织工业等行业有广泛的应用。此外,印染工艺中所使用的染料除了染色纤维外,在电化学等领域也有相当多的应用。本论文围绕蚕丝蛋白产品和某些功能染料,采用电化学方法,测定了生丝液体样品中丝素蛋白含量、脱胶废水中丝胶蛋白浓度,构筑了蚕丝蛋白产品生物相容性功能界面,探索了功能界面的应用性能;通过研究某些功能染料的电化学行为,建立了测定染料染色纤维上染率的电化学方法。这些研究对开发生物相容性功能界面,拓宽电化学技术应用范围,搭建电化学与纺织印染工业的联系具有重要的意义。全文共分4章,主要研究内容如下:
1.采用循环伏安法,以丝素肽(SP)磷酸缓冲液(pH7.4)为电聚合前体,在碳糊电极(CPE)表面制备了“聚”丝素肽(“P-SP”)。与采用自组装方法将9,修饰在CPE表面上的自组装丝素肽(“S-SP”)稀疏颗粒相比,“P-SP”以大量粒子团簇形式存在;在0.1 mol·L~(-1)硫酸(pH 1.1)溶液中,P-SP-CPE界面被质子化,显示稳定的质子氧化还原反应电极过程,而S-SP-CPE却无任何电化学响应。
2.采用循环伏安法,在0.1mol·L~(-1)硫酸(pH1.1)溶液中,研究P-SP-CPE界面与荷正电荷染料孔雀石绿MG、荷负电表面活性剂十二烷基苯磺酸钠SDBS组成的电化学开关。由于P-SP-CPE界面质子化程度受到荷正电物质的减弱,使界面的质子浓度下降,导致质子氧化还原峰电流逐渐变小,并在80 min后氧化还原峰消失,即MG“关闭”P-SP-CPE界面质子的氧化还原反应;随着与MG作用后的P-SP-CPE浸在SDBS溶液中时间增加,P-SP-CPE界面质子原有的一对氧化还原峰又逐步重新出现,即SDBS“开启”质子的氧化还原反应。因此,MG、P-SP-CPE界面、SDBS组成了简单的电化学开关系统。此外,根据MG与P-SP-CPE界面作用时间与峰电流的下降值,计算出阳离子物质孔雀石绿与P-SP-CPE界面质子作用动力学方程呈现一级反应动力学方程:
ln C_(H+,0)/C_(H+,t)=1.19+9.67×10~(-2)t,其中作用速率常数=9.67×10~(-2)min~(-1),半衰期=7.17 min。
3.采用循环伏安法,在0.1 mol·L~(-1)硫酸(pH1.1)溶液中,研究β-环糊精对P-SP-CPE界面上“P-SP”的包合识别。β-环糊精对“P-SP”的包合,导致P-SP-CPE界面质子浓度降低,氧化峰电流下降,峰电位负移。由包合反应时间和P-SP-CPE界面质子氧化峰电流降低值,计算出该包合反应的动力学方程为一级反应速率方程:ln C_(H+,0)/C_(H+,t)=0.952+2.88×10~(-2)t,包合反应速率常数为2.88×10~(-2)min~(-1),半衰期=24.1 min。根据“直线法”,计算出β-环糊精包合“P-SP”的包合比为1:1,包合常数是2.9×10~5L·mol~(-1),25℃时包合反应吉布斯自由能△G_1~0=-31.136 kJ·mol~(-1)。
4.采用循环伏安法,以丝胶0.02 mol·L~(-1)磷酸缓冲液(pH 7.0)为电聚合前体,在纳米TiO_2掺杂碳糊电极表面(TCPE)制备了“聚”丝胶(“P-SE”)。“P-SE”在TCPE表面形成了网络结构。在pH1.81B-R缓冲液中,P-SE-TCPE具有一对稳定的氧化还原峰,阳离子表面活性剂对其有阻止作用,阴离子表面活性剂对其有增敏作用。此外,在0.02 mol·L~(-1)磷酸缓冲液(pH 7.0)溶液中,P-SE-TCPE可将亚铁氰化钾在TCPE上的电极反应速率常数由0.03s~(-1)提高到0.26s~(-1)。
5.采用循环伏安、方波伏安法,研究了丝素蛋白在碳糊电极上的伏安行为。丝素蛋白在0.01 mol·L~(-1)盐酸(pH 2.0)溶液中,丝素蛋白的循环伏安图中出现一个不可逆氧化峰和一对峰型较差的氧化还原峰。根据丝素蛋白的不可逆氧化峰电流与其浓度在5.8×10~(-8)-1.1×10~(-6)mol·L~(-1)范围内成正比,建立了电化学测定生丝液体样品中丝素蛋白含量的方法,丝胶及其他无机离子、氨基酸等并不干扰测定,加标平均回收率在97.0-103.0%。
6.采用电化学方法和光谱法,研究了胭脂红与丝胶蛋白的结合反应。在无丝胶的pH1.81 B-R缓冲液中,胭脂红有一对峰型好的氧化还原峰。随着丝胶蛋白的加入,尽管无新峰出现,但胭脂红原有的这对氧化还原峰却有所变化:氧化还原峰电流下降:氧化还原峰电位正移。这种现象是由于丝胶蛋白与胭脂红结合造成胭脂红的电极反应速率常数和扩散系数的下降而形成的。丝胶蛋白与胭脂红主要以静电引力方式结合。结合反应的结合常数为2.32×10~6L·mol~(-1),结合比为1:1。基于胭脂红氧化峰电流的降低值与丝胶蛋白浓度在32.0-800.0μg·mL~(-1)范围内成正比,测定了脱胶废水中丝胶蛋白含量,加标平均回收率在96.7-103.3%,与经典考马斯亮蓝G-250光谱测定法分析结果一致。
7.采用循环伏安、方波伏安法研究了三芳甲烷染料孔雀石绿、酸性染料酸性大红3R、弱酸性染料弱酸性深蓝GR、活性染料活性嫩黄K-4G在碳糊电极上的伏安行为。根据染料的伏安峰电流与其浓度在一定范围内成正比,建立了测定孔雀石绿染色腈纶、酸性大红3R染色羊毛和丝绸、弱酸性深蓝GR染色丝绸、活性嫩黄K-4G染色丝绸上染率的电化学方法,并与分光光度法测定方法比较,结果一致。
Silk protein(silk fibroin,sericin) and silk peptide(the hydrolysis product of silk fibroin) are widely used in biomedical material,food,cosmetic,pharmacy and textile industries because of their excellent biocompatibility,anti-oxidation and enough amino acids contains.Also,dyes used in textiles can be applied in other domain such as electrochemistry.In this paper,silk and some dyes are investigated with electrochemical methods.Two kinds of biocompatible functional interface are set up and their properties are investigated.The determination of silk fibroin in raw liquid silk samples and sericin in degumming waste water are performed with SWV method,respectively.The dye-uptakes of 4 kinds of dyes dyeing fiber are carried out using SWV,respectively. These studies are significant to opening the biocompatible functional interface, broadening the application range of electrochemistry and building the link between electrochemistry and textiles.The paper consists of four chapters.The main contributions are summarized and presented as follows:
1."P-SP" is obtained by electrochemical polymerization of silk peptide on carbon paste electrode surface in 0.04 mol·L~(-1) phosphate buffer(pH7.4).Compared with the few particles of the self-assembled SP on the surface of carbon paste electrode(S-SP-CPE), "P-SP" is formed with a large number of particle clusters.The P-SP-CPE is characterized by cyclic voltammetry and shows a pair of well-defined redox peaks in 0.1 mol·L~(-1) H_2SO_4 solution(pH 1.1),which is ascribed to the redox reaction of proton combined with "P-SP",while S-SP-CPE has no voltammetric response.
2.Electrochemical switch formed by positive-charged dye malachite green(MG) and negative-charged anionic surfactant sodium dodecyl benzene sulfonate(SDBS) on the interface of "P-SP" is studied in 0.1 mol·L~(-1) H_2SO_4 solution(pH1.1) using cyclic voltammetry.The protonated degree of "P-SP" is reduced by the interaction of MG, which induce the concentration of proton decrease and then the peak current of proton redox peaks become low.After 80 min,no response is nearly observed,suggested that "MG" shuts off the redox of proton on P-SP interface.This interface is named P-SP-MG-CPE.The redox response of proton is found again with the interacting time increase of P-SP-MG-CPE with SDBS solution,indicating that "SDBS" opens the redox of proton.Therefore,MG,SDBS and P-SP form a simple "electrochemical switch".In addition,the interaction between MG and P-SP obeys the 1st order rate equation:In C_(H+,0)/C_(H+,t)=1.19 + 9.67×10~(-2) t;interaction rate constant = 9.67×10~(-2) min~(-1),t_(1/2)= 7.17 min.
3.The inclusion and recognition ofβ-CD with "P-SP" is investigated in 0.1 mol·L~(-1) H_2SO_4 solution(pH 1.1) with cyclic voltammetry.The oxidative peak currents decrease with the peak potential shifting negatively as a result ofβ-CD inclusion "P-SP".The inclusion reaction obeys the 1st order rate rate equation:In C_(H+,0)/ C_(H+,t)= 0.952 +2.88×10~(-2) t,inclusion rate constant = 2.88×10~(-2) min~(-1),t_(1/2) = 24.1 min.Based on the "Linear method",the inclusion constant and ratio are calculated as 2.9×10~5 L·mol~(-1), 1:1,respectively.The△G_1~0 of inclusion reaction is -31.136 kJ·mol~(-1).
4."P-SE" is prepared by electrochemical polymerization of sericin in 0.02 mol·L~(-1) phosphate buffer(pH 7.0) on nanometer TiO_2 modified carbon paste electrode surface. The net structure is formed on this interface.A pair of stable redox peaks has been found in pH 1.81B-R buffer for P-SE-TCPE.This redox reaction can be hindered by cationic surfactants while enhanced by anionic surfactants.Also,the electrode reaction rate constant of potassium ferrocyanide in 0.02 mol·L~(-1) PBS(pH 7.0) on P-SE-TCPE is 0.26 s~(-1),which is higher than that of 0.03s~(-1) on TCPE.
5.The electrochemical behavior of protein material silk fibroin is studied in pH 2.0 of hydrochloric acid solution with cyclic voltammetry and square wave voltammetry at a CPE.Silk fibroin has a clearly irreversible oxidation peak(P_(a,1)) and a pair of weak-defined reversible peak(P_(a,2) and P_c).The peak currents of peak P_(a,1) are proportional to the concentration of silk fibroin in the range of 5.8×10~(-8) to 1.1×10~(-6) mol·L~(-1).Sericin,amino acids and inorganic ions do not interfere with silk fibroin determination.This method is applied to the determination of SF in raw silk liquid samples without any pre-separation and pre-purification.The average recoveries are 97.0-103.0%.
6.The interaction between carmine and sericin is investigated with electrochemical methods and the UV-Vis spectrophotometry.In the absence of sericin,carmine has a pair of well-defined redox peaks in a pH 1.81 B-R buffer solution.Although no new redox peaks appear upon the addition of sericin into a carmine solution,the peak currents of the old peaks reduce while the peak potentials shift positively.This observation is attributed to the decrease in the diffusion coefficient and electrode reaction rate constant of carmine in the presence of sericin.The main binding mode between carmine and sericin is electrostatic attraction.The binding constant and binding ratio are calculated as 2.32×10~6 L·mol~(-1) and 1:1,respectively.Furthermore,the decrease in the peak currents is found proportional to the sericin concentration in the range of 32.0-800.0μg·mL~(-1).The method is further applied to the determination of sericin in a degumming wastewater with satisfied average recoveries from 96.7 to 103.3%.The results are in good agreement with those obtained by the conventional Coomassie brilliant blue G-250 spectrophotometric method.
7.The electrochemical behaviors of malachite green(MG),brilliant acid scarlet red 3R,C.I. Acid Blue 120(WAGR) and reactive yellow K-4G are studied with cyclic voltammetry and square wave voltammetry at a CPE.Based on the linear correlation between peak currents and dye concentrations,SWV methods are set up to the determination of dye-uptakes of these four dyes dyeing different fibers(MG dyeing acrylic fiber,brilliant acid scarlet red 3R dyeing silk and wool,C.I.ACID BLUE 120 dyeing silk and reactive yellow K-4G dyeing silk).The results are in accordance with the traditional spectrophotometric method.
1.采用循环伏安法,以丝素肽(SP)磷酸缓冲液(pH7.4)为电聚合前体,在碳糊电极(CPE)表面制备了“聚”丝素肽(“P-SP”)。与采用自组装方法将9,修饰在CPE表面上的自组装丝素肽(“S-SP”)稀疏颗粒相比,“P-SP”以大量粒子团簇形式存在;在0.1 mol·L~(-1)硫酸(pH 1.1)溶液中,P-SP-CPE界面被质子化,显示稳定的质子氧化还原反应电极过程,而S-SP-CPE却无任何电化学响应。
2.采用循环伏安法,在0.1mol·L~(-1)硫酸(pH1.1)溶液中,研究P-SP-CPE界面与荷正电荷染料孔雀石绿MG、荷负电表面活性剂十二烷基苯磺酸钠SDBS组成的电化学开关。由于P-SP-CPE界面质子化程度受到荷正电物质的减弱,使界面的质子浓度下降,导致质子氧化还原峰电流逐渐变小,并在80 min后氧化还原峰消失,即MG“关闭”P-SP-CPE界面质子的氧化还原反应;随着与MG作用后的P-SP-CPE浸在SDBS溶液中时间增加,P-SP-CPE界面质子原有的一对氧化还原峰又逐步重新出现,即SDBS“开启”质子的氧化还原反应。因此,MG、P-SP-CPE界面、SDBS组成了简单的电化学开关系统。此外,根据MG与P-SP-CPE界面作用时间与峰电流的下降值,计算出阳离子物质孔雀石绿与P-SP-CPE界面质子作用动力学方程呈现一级反应动力学方程:
ln C_(H+,0)/C_(H+,t)=1.19+9.67×10~(-2)t,其中作用速率常数=9.67×10~(-2)min~(-1),半衰期=7.17 min。
3.采用循环伏安法,在0.1 mol·L~(-1)硫酸(pH1.1)溶液中,研究β-环糊精对P-SP-CPE界面上“P-SP”的包合识别。β-环糊精对“P-SP”的包合,导致P-SP-CPE界面质子浓度降低,氧化峰电流下降,峰电位负移。由包合反应时间和P-SP-CPE界面质子氧化峰电流降低值,计算出该包合反应的动力学方程为一级反应速率方程:ln C_(H+,0)/C_(H+,t)=0.952+2.88×10~(-2)t,包合反应速率常数为2.88×10~(-2)min~(-1),半衰期=24.1 min。根据“直线法”,计算出β-环糊精包合“P-SP”的包合比为1:1,包合常数是2.9×10~5L·mol~(-1),25℃时包合反应吉布斯自由能△G_1~0=-31.136 kJ·mol~(-1)。
4.采用循环伏安法,以丝胶0.02 mol·L~(-1)磷酸缓冲液(pH 7.0)为电聚合前体,在纳米TiO_2掺杂碳糊电极表面(TCPE)制备了“聚”丝胶(“P-SE”)。“P-SE”在TCPE表面形成了网络结构。在pH1.81B-R缓冲液中,P-SE-TCPE具有一对稳定的氧化还原峰,阳离子表面活性剂对其有阻止作用,阴离子表面活性剂对其有增敏作用。此外,在0.02 mol·L~(-1)磷酸缓冲液(pH 7.0)溶液中,P-SE-TCPE可将亚铁氰化钾在TCPE上的电极反应速率常数由0.03s~(-1)提高到0.26s~(-1)。
5.采用循环伏安、方波伏安法,研究了丝素蛋白在碳糊电极上的伏安行为。丝素蛋白在0.01 mol·L~(-1)盐酸(pH 2.0)溶液中,丝素蛋白的循环伏安图中出现一个不可逆氧化峰和一对峰型较差的氧化还原峰。根据丝素蛋白的不可逆氧化峰电流与其浓度在5.8×10~(-8)-1.1×10~(-6)mol·L~(-1)范围内成正比,建立了电化学测定生丝液体样品中丝素蛋白含量的方法,丝胶及其他无机离子、氨基酸等并不干扰测定,加标平均回收率在97.0-103.0%。
6.采用电化学方法和光谱法,研究了胭脂红与丝胶蛋白的结合反应。在无丝胶的pH1.81 B-R缓冲液中,胭脂红有一对峰型好的氧化还原峰。随着丝胶蛋白的加入,尽管无新峰出现,但胭脂红原有的这对氧化还原峰却有所变化:氧化还原峰电流下降:氧化还原峰电位正移。这种现象是由于丝胶蛋白与胭脂红结合造成胭脂红的电极反应速率常数和扩散系数的下降而形成的。丝胶蛋白与胭脂红主要以静电引力方式结合。结合反应的结合常数为2.32×10~6L·mol~(-1),结合比为1:1。基于胭脂红氧化峰电流的降低值与丝胶蛋白浓度在32.0-800.0μg·mL~(-1)范围内成正比,测定了脱胶废水中丝胶蛋白含量,加标平均回收率在96.7-103.3%,与经典考马斯亮蓝G-250光谱测定法分析结果一致。
7.采用循环伏安、方波伏安法研究了三芳甲烷染料孔雀石绿、酸性染料酸性大红3R、弱酸性染料弱酸性深蓝GR、活性染料活性嫩黄K-4G在碳糊电极上的伏安行为。根据染料的伏安峰电流与其浓度在一定范围内成正比,建立了测定孔雀石绿染色腈纶、酸性大红3R染色羊毛和丝绸、弱酸性深蓝GR染色丝绸、活性嫩黄K-4G染色丝绸上染率的电化学方法,并与分光光度法测定方法比较,结果一致。
Silk protein(silk fibroin,sericin) and silk peptide(the hydrolysis product of silk fibroin) are widely used in biomedical material,food,cosmetic,pharmacy and textile industries because of their excellent biocompatibility,anti-oxidation and enough amino acids contains.Also,dyes used in textiles can be applied in other domain such as electrochemistry.In this paper,silk and some dyes are investigated with electrochemical methods.Two kinds of biocompatible functional interface are set up and their properties are investigated.The determination of silk fibroin in raw liquid silk samples and sericin in degumming waste water are performed with SWV method,respectively.The dye-uptakes of 4 kinds of dyes dyeing fiber are carried out using SWV,respectively. These studies are significant to opening the biocompatible functional interface, broadening the application range of electrochemistry and building the link between electrochemistry and textiles.The paper consists of four chapters.The main contributions are summarized and presented as follows:
1."P-SP" is obtained by electrochemical polymerization of silk peptide on carbon paste electrode surface in 0.04 mol·L~(-1) phosphate buffer(pH7.4).Compared with the few particles of the self-assembled SP on the surface of carbon paste electrode(S-SP-CPE), "P-SP" is formed with a large number of particle clusters.The P-SP-CPE is characterized by cyclic voltammetry and shows a pair of well-defined redox peaks in 0.1 mol·L~(-1) H_2SO_4 solution(pH 1.1),which is ascribed to the redox reaction of proton combined with "P-SP",while S-SP-CPE has no voltammetric response.
2.Electrochemical switch formed by positive-charged dye malachite green(MG) and negative-charged anionic surfactant sodium dodecyl benzene sulfonate(SDBS) on the interface of "P-SP" is studied in 0.1 mol·L~(-1) H_2SO_4 solution(pH1.1) using cyclic voltammetry.The protonated degree of "P-SP" is reduced by the interaction of MG, which induce the concentration of proton decrease and then the peak current of proton redox peaks become low.After 80 min,no response is nearly observed,suggested that "MG" shuts off the redox of proton on P-SP interface.This interface is named P-SP-MG-CPE.The redox response of proton is found again with the interacting time increase of P-SP-MG-CPE with SDBS solution,indicating that "SDBS" opens the redox of proton.Therefore,MG,SDBS and P-SP form a simple "electrochemical switch".In addition,the interaction between MG and P-SP obeys the 1st order rate equation:In C_(H+,0)/C_(H+,t)=1.19 + 9.67×10~(-2) t;interaction rate constant = 9.67×10~(-2) min~(-1),t_(1/2)= 7.17 min.
3.The inclusion and recognition ofβ-CD with "P-SP" is investigated in 0.1 mol·L~(-1) H_2SO_4 solution(pH 1.1) with cyclic voltammetry.The oxidative peak currents decrease with the peak potential shifting negatively as a result ofβ-CD inclusion "P-SP".The inclusion reaction obeys the 1st order rate rate equation:In C_(H+,0)/ C_(H+,t)= 0.952 +2.88×10~(-2) t,inclusion rate constant = 2.88×10~(-2) min~(-1),t_(1/2) = 24.1 min.Based on the "Linear method",the inclusion constant and ratio are calculated as 2.9×10~5 L·mol~(-1), 1:1,respectively.The△G_1~0 of inclusion reaction is -31.136 kJ·mol~(-1).
4."P-SE" is prepared by electrochemical polymerization of sericin in 0.02 mol·L~(-1) phosphate buffer(pH 7.0) on nanometer TiO_2 modified carbon paste electrode surface. The net structure is formed on this interface.A pair of stable redox peaks has been found in pH 1.81B-R buffer for P-SE-TCPE.This redox reaction can be hindered by cationic surfactants while enhanced by anionic surfactants.Also,the electrode reaction rate constant of potassium ferrocyanide in 0.02 mol·L~(-1) PBS(pH 7.0) on P-SE-TCPE is 0.26 s~(-1),which is higher than that of 0.03s~(-1) on TCPE.
5.The electrochemical behavior of protein material silk fibroin is studied in pH 2.0 of hydrochloric acid solution with cyclic voltammetry and square wave voltammetry at a CPE.Silk fibroin has a clearly irreversible oxidation peak(P_(a,1)) and a pair of weak-defined reversible peak(P_(a,2) and P_c).The peak currents of peak P_(a,1) are proportional to the concentration of silk fibroin in the range of 5.8×10~(-8) to 1.1×10~(-6) mol·L~(-1).Sericin,amino acids and inorganic ions do not interfere with silk fibroin determination.This method is applied to the determination of SF in raw silk liquid samples without any pre-separation and pre-purification.The average recoveries are 97.0-103.0%.
6.The interaction between carmine and sericin is investigated with electrochemical methods and the UV-Vis spectrophotometry.In the absence of sericin,carmine has a pair of well-defined redox peaks in a pH 1.81 B-R buffer solution.Although no new redox peaks appear upon the addition of sericin into a carmine solution,the peak currents of the old peaks reduce while the peak potentials shift positively.This observation is attributed to the decrease in the diffusion coefficient and electrode reaction rate constant of carmine in the presence of sericin.The main binding mode between carmine and sericin is electrostatic attraction.The binding constant and binding ratio are calculated as 2.32×10~6 L·mol~(-1) and 1:1,respectively.Furthermore,the decrease in the peak currents is found proportional to the sericin concentration in the range of 32.0-800.0μg·mL~(-1).The method is further applied to the determination of sericin in a degumming wastewater with satisfied average recoveries from 96.7 to 103.3%.The results are in good agreement with those obtained by the conventional Coomassie brilliant blue G-250 spectrophotometric method.
7.The electrochemical behaviors of malachite green(MG),brilliant acid scarlet red 3R,C.I. Acid Blue 120(WAGR) and reactive yellow K-4G are studied with cyclic voltammetry and square wave voltammetry at a CPE.Based on the linear correlation between peak currents and dye concentrations,SWV methods are set up to the determination of dye-uptakes of these four dyes dyeing different fibers(MG dyeing acrylic fiber,brilliant acid scarlet red 3R dyeing silk and wool,C.I.ACID BLUE 120 dyeing silk and reactive yellow K-4G dyeing silk).The results are in accordance with the traditional spectrophotometric method.
引文
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