A method for measuring corn germ lipoxygenase activity

A lipoxygenase and corn seed technology, applied in biochemical equipment and methods, microbial measurement/inspection, material analysis through observation of the influence of chemical indicators, etc., can solve the problem of corn seed lipoxygenase UV spectrophotometer The method has not been reported and other problems, and achieves the effect of easy operation, simple reaction steps and cost saving

Inactive Publication Date: 2011-12-14
CHINA AGRI UNIV
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Problems solved by technology

[0005] The UV spectrophotometer method of soybean seed and peanut seed lipoxygenase has been

Method used

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  • A method for measuring corn germ lipoxygenase activity
  • A method for measuring corn germ lipoxygenase activity
  • A method for measuring corn germ lipoxygenase activity

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Embodiment 1, investigate the optimal reaction buffer pH value and concentration of corn germ lipoxygenase activity ultraviolet assay method

[0023] 1) Extract corn seed embryo lipoxygenase solution: take 0.3 g of corn seed embryo, add 3.0 ml of disodium hydrogen phosphate-sodium dihydrogen phosphate buffer solution with a pH value of 7.0 and a concentration of 0.05 mol / L for grinding on ice, and grind After crushing, centrifuge and collect the supernatant, which is the corn germ lipoxygenase crude enzyme solution, store it at 4°C, and measure it as soon as possible. The Coomassie Brilliant Blue method was used to measure the content of the soluble protein in the crude enzyme solution, the standard curve was established from bovine serum albumin with a concentration of 0-0.2 mg, and the standard curve equation was Y=4.964X+0.0422.

[0024] 2) Prepare the reaction substrate (linoleic acid solution containing Tween 20): prepare a 10 mmol / L linoleic acid solution containi...

Embodiment 2

[0042] Embodiment 2, measure different artificial aging time corn seed 178 kinds of embryo lipoxygenase activity

[0043] Maize inbred line 178 showed good storage characteristics in both natural storage and artificial accelerated aging tests. We have obtained the 178 seeds that germination rate is 90%, 80%, 76%, 64% by the method of artificial aging, utilize the optimum method that embodiment 1 determines to carry out lipoxygenase activity measurement to the seed of above-mentioned different germination rate, result See Table 3.

[0044] Table 3 Lipoxygenase activity of corn inbred line 178

[0045]

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Abstract

The invention discloses a method for testing the activity of corn seed embryo lipoxidase. The method comprises the following steps of: reacting an enzyme activity detection system at temperature of 24 to 26 DEG C, testing outside dimension (OD) values of the system at a position of 234 nanometers after reaction for t1 and t2, and recording the values as OD(t1) and OD(t2); and calculating the activity of the lipoxidase of the corn seed embryo according to the following formula: A=60s*[OD(t2)-OD(t1)]/0.01/(t2-t1). The enzyme activity detection system consists of tween 20, linoleic acid, Tris-Hc1 buffer solution with pH value of 8.0 and concentration of 0.02 mol/ml and the corn seed embryo lipoxidase to be tested, wherein the concentration of the linoleic acid is 0.3mmol/L, the concentrationof tween 20 is 0.25mmol/L; and the concentration of the corn seed embryo lipoxidase to be tested is not more than 0.0167mg/3ml. The enzyme activity is defined that: the enzyme amount required by the enzyme activity detection system for increasing 0.01 absorbance at every minute is an enzyme activity unit.

Description

technical field [0001] The invention relates to a method for measuring the activity of corn germ lipoxygenase. Background technique [0002] An enzyme is a protein with biological catalytic activity. Its activity is affected by conditions such as temperature, pH value of reaction buffer, buffer type, concentration, and substrate concentration. Therefore, to measure the activity of an enzyme, it is necessary to know the conditions of the enzyme reaction. [0003] Lipoxygenase (LOX, EC1.13.11.12) is one of the key enzymes in lipid degradation. It uses unsaturated fatty acids as substrates to produce toxic substances such as aldehydes and ketones. All prove that lipoxygenase and related to the storability properties of seeds. Wang Jingyan, Zhu Shengkang, Xu Changfa (2002) pointed out that the reaction catalyzed by lipoxygenase requires the participation of active oxygen, and the mechanism of action is to oxidize and break the double bonds in unsaturated fatty acids into smal...

Claims

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Application Information

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IPC IPC(8): G01N21/78C12Q1/26
Inventor 王建华赵正楠王国英胡新民
Owner CHINA AGRI UNIV
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