显色剂加入方式及结果计算方法对土壤磷酸酶测定结果的影响

Effects of Color Development Reagent Adding and Result Calculation Methods on Determination of Soil Phosphatase Activity

为探讨有色显色剂的加液方式及加入量对显色后溶液吸收值的影响、标准曲线的绘制及结果计算方法对土壤磷酸酶测定值的影响,取广东橡胶园0~20 cm土样为材料,以磷酸苯二钠为基质,酶解土壤释放出酚,与显色剂氯代二溴对苯醌亚胺反应,以标准曲线法求出样品测试液的酚浓度,比较以不同加液方式加入不同量的显色剂对溶液吸收值的影响,同时比较用直线或一元二次曲线等两种标准曲线回归方程对样品测试液酚浓度的影响。结果表明:以胶头滴管滴加或以移液枪用多次重复使用过的吸嘴加入显色剂,显色后溶液吸收值变异系数达3.33%~6.44%,而以活塞式定量加液器加入,其变异系数在1.28%以下,且随着加入量增大变异系数降低至0.51%,重现性好;当配制的标准曲线浓度范围较宽且相应吸收值较高(接近0.9),以曲线法求得的标准曲线回归方程决定系数(R2=0.999 8)大于直线回归的(R2=0.999 7),且曲线法求出的土壤无基质对照及试剂空白酚浓度比直线法更有实际意义。

Effects of color development reagent dosing methods and dosing volumes on absorbance were investigated in this study, and effects of calculation methods on final values were also discussed in this study. Soil samples were collected in Guangdong rubber plantation from 0 ~20 cm layer, benzene disodium phosphate as the reactant, and chlorinated dibromo benzoquinoneimine as the color development reagent, the phenol concentration of tested sample solution be calculated using the standard curve methods.The effects of different dosing methods and dosing volumes of color development reagent on absorbance were compared and the phenol concentrations of tested sample solution were compared with linear or quadratic curve regression equation. The results showed that the coefficient of variation(CV) of the absorbance value ranged from 3.33% to 6.44% after adding different amounts of color development reagent using rubber dropper or pipette with continuous used tips, and CV was less than 1.28% using a piston dispenser, and CV was reduced to 0.51% with the amounts of the color reagent increasing. When standards of desired concentration range were wide, and the highest point of the standard series had a higher absorbance value, for example, the absorbance value of the highest standard closed to 0.9 in this experiment, the coefficient of determination of the standard curve regression equation based on a quadratic fit(R2=0.999 8) was higher than on a linear fit(R2=0.999 7), and the phenol concentration of tested blank without disodium phenyl phosphate and the reagent blank were more meaningful using the regression equation based on a quadratic fit than based on a linear fit.