采用反复冻融细胞破裂法、硫酸铵分级盐析以及羟基磷灰石柱层析,从钝顶螺旋藻中提取出高纯度的藻蓝蛋白样品,纯度(A62a/A280)达4.1.该蛋白的紫外-可见吸收光谱表明其特征吸收峰为280、360、620nm,荧光光谱表明其最大发射波长为650 nm.以该藻蓝蛋白为荧光探针,发展了一种基于荧光猝灭法的Hg2检测新方法.并考察了缓冲体系、缓冲液pH值、反应时间、温度以及藻蓝蛋白的浓度等因素对汞离子检测的影响,在0.05 mol/L、pH7.5的磷酸二氢钾-磷酸氢二钠缓冲液中,当藻蓝蛋白浓度为3 mg/L、反应时间为30 min、反应温度为30℃时,该方法的线性范围为0.1～10μmol/L,检出限为0.056 μmol/L.该方法表现出良好的汞离子传感选择性,而且当干扰离子与汞离子的浓度比为40∶1时,多种共存离子对汞离子的检测影响较小.该方法荧光探针提取容易,价格低且环境友好,具有较高的灵敏度和较好的重现性.%Through repeated freezing and thawing of cell, ammonium sulfate precipitation and hydroxyapatite chromatography, the phycocyanin was extracted from spirulina platensis and its purity could reach up to A620/A280 = 4. 1. UV - visible spectra and fluorescence emission spectrometry were used to study the spectral characterisitic of the protein. The results showed that the absorption peaks of the purified protein were located at 280, 360, 620 nm and the maximum emission wavelength was 650 nm. Using this phycocyanin as fluorescence probe, a new approach was developed for the sensitive and selective determination of divalent mercury ions based on fluorescence quenching. Different influence factors, including pH value, reaction time, temperature and phycocyanin concentration, were investigated. In 0. 05 mol/L KH2PO4 - Na2HPO4 , at a pH value of 7. 5, phycocyanin concentration of 3 mg/L, reaction time of 30 min and reaction temperature of 30 ℃, the relative fluorescence intensity decreased linearly with mercury ions concentration in the range of 0. 1 - 10 μmol/L. The detection limit was 0. 056 μmol/L. This approach displayed a good selectivity to mercury ions. When concentration ratio of interference ions to mercury ion reached to 40 : 1, several kinds of coexistence ions had little influence on the detection of mercury ion. This new method exhibits a bright application prospect in the detection of mercury ions due to its advantages of good simplicity, sensitivity, reproducibility and pro-environment.
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