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目的建立高灵敏荧光增强法测定硫酸软骨素的新方法,并用于实际样品测定。方法对荧光光谱行为的考察显示,在pH 7.0的介质中,具有共轭结构的阳离子头部和长碳链尾部的阳离子表面活性剂对四磺基铝酞菁(AlS4Pc)的荧光具有高效猝灭作用,二者形成无荧光离子缔合物。此离子缔合物可对带有磺酸基团的高分子黏多糖硫酸软骨素(CS)作出灵敏响应。在CS的存在下,反应体系荧光显著恢复,最大恢复倍数可达900倍。由于猝灭效率高,荧光恢复程度大,甚至可实现目视化观测。结果工作曲线响应区间0.2~6μg·mL-1,检测限0.09μg·mL-1,工作曲线logY=0.158 23ρ-0.035 97,r=0.998 7。本实验对该法用于批量样品分析的可行性进行了考察,结果表明,所建立的方法可望用于高通量分析。结论该法操作简便,灵敏度、稳定性与检测准确性好。所建立的方法成功地用于组成复杂的实际样品中CS的定量分析,解决了实际工作中的难点。
OBJECTIVE To establish a new sensitive method for the determination of chondroitin sulfate by fluorescence enhancement and to determine the actual sample. Methods The investigation of the fluorescence spectra showed that cationic surfactants with conjugated structures and long-chain carbon-tail possessed highly efficient quenching of the fluorescence of tetrasodium aluminum phthalocyanine (AlS4Pc) in pH 7.0 media Role, both of which form non-fluorescent ion association. This ionic complex responds sensitively to the sulfonic acid group-containing polysaccharide chondroitin sulfate (CS). In the presence of CS, the fluorescence of the reaction system significantly recovered, the maximum recovery fold up to 900 times. Due to the high quenching efficiency, a large degree of fluorescence recovery, visual observation can even be achieved. Results The response curve of working curve was 0.2-6μg · mL-1 with the detection limit of 0.09μg · mL-1. The logY = 0.158 23ρ-0.03597, r = 0.998 7. In this experiment, the feasibility of the method for batch sample analysis was investigated. The results show that the proposed method is expected to be used in high-throughput analysis. Conclusion The method is simple, sensitive, stable and accurate. The established method is successfully applied to the quantitative analysis of CS in the complex real samples, which solves the difficulties in practical work.