过氧化氢的测定方法.docVIP

过氧化氢的测定方法 Three, catalase activity - ultraviolet absorption method [principle] H202 has a strong absorption at 240nm wavelengths, and hydrogen peroxide decomposes hydrogen peroxide, reducing the absorbance (A240) of the reaction solution with reaction time. The activity of catalase can be measured according to the changing rate of light absorption. Instruments and appliances [] ultraviolet spectrophotometer; centrifuge; mortar; volumetric flask 250ml1; scale Straw 0.5ml2; 2ML1; 10ml tube 3; constant temperature water bath. [reagent] 0.2mol? L-1pH7.8 phosphate buffer (containing 1% polyvinylpyrrolidone (L-1H202); 0.1mol? 0.1mol? L-1 Potassium Permanganate calibration). [Methods] 1. enzymes extracted weighed fresh wheat leaves or other plant tissue 0.5g, with a mortar, adding 2-3ml at 4 DEG C pH7.0 phosphate buffer pre cooling and a small amount of quartz sand grinding homogenized into 25ml volumetric flask, and washed with buffer mortar with several times, washing liquid, and set the volume to scale. The flask mixing, 5 DEG C in the refrigerator standing 10min, take the upper clear liquid in 4000r.min-1 centrifugal 15min, supernatant for catalase in crude extracts, standby preserved at the temperature of 5 DEG C. 2. determination of 10ml test tube 3, of which 2 samples for the test tube, 1 for blank tubes, in accordance with the order 42-2 reagents. 25 C after preheating, by adding 0.3ml0.1mol H2O2 tubes, each tube immediately after the 1 plus time, and quickly poured into the quartz cuvette, determination of absorbance of 240nm, every 1min 1 readings, were measured for 3 4min, all branch after the determination of the enzyme activity was calculated according to the type of 42-3. The 3. result was calculated to reduce the enzyme amount by 0.1 in 1min to 1 enzyme activity units (A240) (U). Catalase activity (u.g-1min-1) =? In the formula, Aso-- is added to the boiled dead enzyme solution to take care of the absorbance value; As1, As2- sample tube light absorption valu