动态光散射和透射电镜法研究pH和NaCl对丝胶蛋白微观结构的影响

Analysis of the Effects of pH and Salt on the Conformation of the Sericin Particles by DLS and TEM Measurements

应用近红外光谱法、动态光散射法及透射电镜扫描法,研究了pH和NaCl浓度对丝胶蛋白聚集微观结构的影响。近红外光谱显示丝胶蛋白在酰氨Ⅰ带1700～1600cm-1附近有较强吸收峰。通过zeta电势法测定丝胶蛋白的表观等电点为pH3.7。应用动态光散射法测定了在不同pH和NaCl浓度下丝胶蛋白颗粒的分散粒径,pH4及高NaCl浓度时,丝胶蛋白聚集颗粒粒径大且分散系数大;pH3或pH8及低NaCl浓度时丝胶蛋白聚集颗粒粒径和分散系数相对较小。采用透射扫描电镜观察丝胶蛋白在pH3或pH8条件下聚集形成松散的松针状微观结构,在pH4或者高NaCl浓度下会聚集形成相对紧密的微观结构。pH4时可以观察到丝胶蛋白的椭圆形单聚体大小约为(60±6)nm(n=10)。并探讨了静电斥力、氢键和范德华吸引力对丝胶蛋白微观结构形成的影响,为丝胶蛋白作为生物材料的应用提供了理论依据。

The particles conformation of the sericin protein extracted from silkworm Bombyx mori was studied under the conditions of different pH and salt concentrations by infrared spectroscopy （IR）, dynamic light scattering （DLS） and transmission electron microscopy （TEM） measurements. The IR spectrum of sericin protein arises predominantly from CO stretching vibration around the amide I region of 1 700-1 600 cm^-1. A strong trend of aggregation of the protein could be observed under specified experimental conditions. The apparent isoelectric point of the sericin protein was about 3.7. The DLS method was used to investigate the effects of pH and NaCl on the size distribution, where a large polydispersity of the system could be observed. Compared to pH 4 or high NaCl concentration, at pH 3, 8 or low NaCl concentration the sericin aggregation shows a relatively smaller size but larger polydispersity. TEM was used to investigate the microstructure of the aggregated sericin protein, where a loose and pine-like branched form could be observed at pH 3 or 8; however, a relatively compact structure was observed near pH 4 or at high salt concentration. At pH 4 the spherical monomer size can be calculated at around （60±6） nm （n=10） by TEM measurement. These phenomena could be explained by the effects of the electrostatic repulsion, hydrogen bonding and Van der Waals attractive force, which provide a basic theory for the application of sericin as biomaterial.