稀土螯合物的制备是均相时间分辨荧光免疫分析中的关键部分, 为了合成理想的稀土螯合物, 以2,6-二(溴甲基)吡啶-3,5-二甲酸二乙酯为原料, 首先优化合成了Li+2,6-{N,N’,N,N’-［二(2,2’-联吡啶-6,6’-二甲基)］二(氨甲基)}-吡啶-二羧酸乙酯, 使其产率明显提高。 进一步选择乙腈和甲醇两种反应体系合成铕螯合物, 并比较了不同反应体系下合成的铕螯合物的光谱性质。 研究表明, 乙腈和甲醇两种反应体系所得铕螯合物的激发光谱(最大激发波长为310 nm)、 发射光谱(最大发射波长为616 nm)、 量子产率基本相同, 荧光强度在10-8～10-5 mol·L-1范围内与Eu3+浓度均成线性, 相关系数分别为0.993 73和0.986 65, 两种铕螯合物(c=2.5×10-5 mol·L-1)的荧光强度略有差异, 荧光寿命分别为825和830 μs。 因此, 两种反应体系所得铕螯合物具有斯托克斯位移大、 荧光强度强以及荧光寿命长等优点, 并且此种穴状螯合剂结构中的吡啶-2,2-联吡啶可保护铕离子免受其他物质的干扰, 是理想的稀土螯合物, 可用于蛋白质、 核酸等生物分子的标记。 本研究不仅拓展了合成新型稀土螯合物的方法, 而且为进一步建立均相时间分辨荧光免疫分析奠定了基础。

Synthesis of rare-earth chelate is a key part of homogeneous time-resolved fluorescence immunoassay. To synthesize optimal rare-earth chelate, we select diethyl 2,6-dibromomethyl-3,5-pyridinecarboxylate as raw material and optimally synthesize Li+2,6-{N,N’,N,N’-［bis(2,2’-bipyridine-6,6’-dimethyl)］ bis(aminomethyl)}-pyridine-3,5-diethyl diester, and then the spectral difference of europium cryptate synthesized by the different reactive systems of acetonitrile and methyl alcohol are discussed. Studies show the excitation spectra(the maximum excitation wavelength is 310 nm), emission spectra(the maximum emission wavelength is 616 nm) and quantum yield of europium cryptate synthesized by the different reactive systems are consistent. Fluorescence intensity are linear with the concentration of europium in the range of 10-8～10-5 mol·L-1 (R2=0.993 73, 0.986 65), respectively. But the fluorescence intensity between them (c=2.5×10-5 mol·L-1) are slightly different. Their fluorescence lifetime are 825 and 830 μs, respectively. Therefore, europium cryptate synthesized has big Stoke’s shift, high fluorescence intensity, long fluorescence lifetime and so on. Pyridine-2,2-bipyridine in europium cryptate can protect europium from interference. It is an ideal europium cryptate used for the labeling of biomolecules such as protein, nucleic acid. This study not only expands the synthesis of new rare earth chelate, but also lays the foundation for the establishment of homogeneous time-resolved fluorescence immunoassay.