光谱学与光谱分析, 2016, 36 (1): 151, 网络出版: 2016-02-02   

同步荧光光谱技术研究胶原基表面活性剂溶液中分子的聚集行为

Aggregation Behavior of Collagen-Based Surfactant Molecules in Aqueous Solutions Based on Synchronization Fluorescence Spectrum Technology
作者单位
1 四川大学皮革化学与工程教育部重点实验室, 四川 成都 610065
2 安庆师范学院生命科学学院, 安徽 安庆 246011
摘要
基于胶原基表面活性剂(collagen-based surfactant, CBS)中酪氨酸(Tyr)和苯丙氨酸(Phe)的荧光特性, 应用恒波长差(Δλ)为15 nm的同步荧光光谱技术研究CBS浓度、 溶液pH值、 NaCl浓度和温度对其在水溶液中分子的聚集行为的影响, 并以温度为外扰, 利用二维同步荧光相关分析研究CBS分子中Tyr残基和Phe残基随温度变化的响应顺序。 结果表明, CBS分子在261和282 nm处出现了分别归属于Phe和Tyr的特征吸收峰。 随着CBS浓度的升高, CBS分子中Phe残基和Tyr残基数量逐渐增多使CBS分子聚集程度增加, 并导致荧光强度增强; CBS溶液pH值(pH 5.0)在等电点附近时, 由于CBS分子的疏水作用和氢键形成能力加强, 导致CBS分子聚集程度增强; CBS溶液中NaCl浓度的升高, 则使CBS分子间排斥力减弱, 从而导致CBS分子的聚集; 然而温度升高, CBS由聚集状态逐渐变为单分子状态, 因猝灭、 变性以及氢键形成能力降低, 荧光强度逐渐降低。 以温度为外扰的二维同步荧光相关光谱分析可知: 低温下(10~40 ℃ ), CBS聚集体随温度升高逐渐松散, 位于聚集体内部的Phe残基比Tyr残基优先响应; 而在45~70 ℃时, CBS由单分子状态逐步水解为无规卷曲构造, Tyr残基的间距变大, 氢键形成能力大大降低, Tyr残基比Phe残基优先响应。
Abstract
Due to the intrinsic fluorescence characteristic of tyrosine (Tyr) and phenylalanine (Phe), synchronization fluorescence spectrum technology which adopted the constant wavelength difference (Δλ=15 nm) was selected to investigate the effects of collagen-based surfactant (CBS) concentration, pH, NaCl concentration and temperature on the aggregation state of CBS molecules in aqueous solutions. Meanwhile, temperature-dependent two-dimensional (2D) synchronization fluorescence correlation analyses was used to investigate the variation order of Tyr and Phe residues in CBS molecules with the change of temperature. The results showed that the characteristic absorption peaks located at 261 and 282 nm were attributed to Phe and Tyr, respectively. With the increase of CBS concentration, the amount of Phe and Tyr residues increased gradually which resulted in the increase of aggregate degree of CBS molecules and then led to the increase of fluorescence intensity. When the pH value (pH 5.0) of CBS solutions was close to the isoelectric point of CBS, the aggregate degree of CBS molecules increased due to the increase of the hydrophobic interaction and the formation ability of hydrogen bond. Additionally, with the increase of NaCl concentration, the repulsion force for inter/intra-molecules of CBS decreased, which helped to improve the aggregation behavior of CBS molecules. However, with the increase of temperature, the aggregation state of CBS was changed to be monomolecular state, and then resulted in the decrease of the fluorescence intensity gradually due to the quenching, the denaturation and the decrease of hydrogen bond formation ability. Furthermore, temperature-dependent 2D synchronization fluorescence correlation spectroscopy demonstrated that at lower temperature (10~40 ℃), the aggregate state of CBS changed to be loose state and then Phe residues located in the inside of the aggregate varied before Tyr residues; while in the heating process of 45~70 ℃, the monomolecular state of CBS changed to be random coil conformation, the separation distance between Tyr residues increased and the hydrogen bond formation ability reduced strongly, which led to Tyr residues changed before Phe residues.

李从虎, 田振华, 刘文涛, 李国英. 同步荧光光谱技术研究胶原基表面活性剂溶液中分子的聚集行为[J]. 光谱学与光谱分析, 2016, 36(1): 151. LI Cong-hu, TIAN Zhen-hua, LIU Wen-tao, LI Guo-ying. Aggregation Behavior of Collagen-Based Surfactant Molecules in Aqueous Solutions Based on Synchronization Fluorescence Spectrum Technology[J]. Spectroscopy and Spectral Analysis, 2016, 36(1): 151.

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