酶电极法快速测定甘油含量的研究

利用酶固定化技术，以甘油激酶（GK）、甘油-3-磷酸氧化酶（GPO）为反应酶，研究GK、GPO的固定化方法及固定化模式，制备甘油酶膜、甘油酶电极，并利用其测定甘油含量。结果表明，GK、GPO按1：1比例固定化时，酶电极电流信号最高；最高效固定模式为：GK固定于核微孔膜，共价偶联GPO固定于Biodyne膜，形成共价双酶膜，进而组装为甘油酶电极。性能研究表明，甘油酶电极最适pH值为7.0，最佳温度为28~32℃；最佳实验条件下，线性范围为0.05~9.00 g/L；回收率为98.4%~102.4%，稳定性高，相对标准偏差（RSD）<5%；测定结果与高效液相色谱法、高碘酸氧化法比较，无明显差异（P>0.05），且该方法操作简单，专一性强，检测快速，适于实际生产中甘油的实时定量及监控。

Study of Rapid Determination for Glycerol Content by Enzyme Electrode

Glycerokinase (GK) and glycerol-3-phosphate oxidase (GPO) were immobilized to develop glycerol enzyme membrane and enzyme electrode for determination of glycerol concentration. Results showed that the higher current output for glycerol was obtained using a 1: 1 (GK: GPO) ratio for the random immobilization. The efficient two-enzymatic membrane was prepared by immobilizing GK in nuclear pore film coupled with GPO on Biodyne B. Glycerol enzyme electrodes were realized by assembling the GK-GPO membranes previously prepared. In this way, the optimum pH was 7.0, and the fit range of temperature was 28~32℃. In the best experimental conditions, the linear scope was 0.05~9.00 g/L with excellent linearity and high stability. The recovery was 98.4%~102.4%, and its relative standard deviation (RSD) was <5%. There was no significant difference (P>0.05) in measurement results among glycerol enzyme electrode, Periodate oxidation method and HPLC.