采用荧光光谱技术考察了浓度、温度和pH对天然胶原和酰化胶原分子聚集行为的影响。结果表明,天然胶原与酰化胶原分子在277和293 nm处分别出现了归属于苯丙氨酸(phenylalanine,Phe) 和酪氨酸(tyrosine, Tyr)的特征荧光峰。与天然胶原相比,酰化胶原分子在360~400 nm处出现了归属于Tyr激发态的宽峰,表明其聚集程度强于天然胶原。随着浓度的增加,天然胶原与酰化胶原分子的聚集程度增加,导致荧光强度逐渐增大;然而随着温度的升高,Phe和Tyr的荧光量子产率逐渐降低,同时天然胶原与酰化胶原分子不断解聚集直至出现无规卷曲构造,导致荧光强度不断降低;随pH升高,天然胶原与酰化胶原分子聚集程度先增加后降低,在等电点附近,聚集程度最大。二维荧光光谱数据表明,浓度影响下,天然胶原分子的Phe和酰化胶原分子的Tyr激发态优先响应;温度影响下,天然胶原分子的Tyr和酰化胶原分子的Tyr聚集态优先响应;pH影响下,天然胶原分子的Tyr和酰化胶原分子的Tyr激发态优先响应。
The effects of concentration, temperature, and pH on the aggregation behavior of native and acylated collagen molecules were investigated by fluorescence spectrum technology. Results showed that the characteristic fluorescence peaks of native and acylated collagen molecules presented at 277 and 293 nm, which belonging to the typical peak of phenylalanine (Phe) and tyrosine (Tyr), respectively. Compared with native collagen, acylated collagen molecules peak had a wide range at 360-400 nm, belonging to the excited state of Tyr, indicating that the aggregation degree of acylated collagen molecules was larger than that of native one. With the increase of concentration, the aggregation degree of native and acylated collagen molecules increased which resulted in the gradual increase of fluorescence intensity. However, with the increase of temperature, the fluorescence quantum yield of Phe and Tyr decreased, and both native and acylated collagen molecules depolymerized until the random curl structure appeared. Therefore, the fluorescence intensity of native and acylated collagen molecules decreased with the increase of temperature. Moreover, with the increase of pH, the aggregation degree of native and acylated collagen molecules first increased and then decreased gradually. The aggregation degree of native and acylated collagen molecules was the largest near the isoelectric point. Two-dimensional fluorescence spectrum data showed that under the influence of concentration, Phe of native collagen molecules and the excited state of Tyr of acylated collagen molecules responded preferentially. Under the influence of temperature, Tyr of native collagen molecules and the aggregation state of Tyr of acylated collagen molecules responded firstly. Under the influence of pH, Tyr of native collagen molecules and the excited state of Tyr of acylated collagen molecules responded preferentially.
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