Method for quickly measuring activity of lipoxidase in grain processing byproduct
A rapid detection technology of lipoxygenase, which is applied in the field of deep grain processing, can solve the problems of difficult removal of fat layer, poor light transmittance of substrate solution system, and complicated purification steps, so as to improve the detection limit and save complicated steps , to ensure a relatively stable effect
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Embodiment 1
[0026] 1. Partial degreasing of rice bran: Take 1kg of fresh rice bran, pass through a 300-mesh sieve, take the sieved rice bran, add 5 times the volume of n-hexane, place it in an air-bath shaker, shake and degrease at 150rpm at room temperature for 30min.
[0027] 2. Desolvation and air-drying: After the rice bran is partially degreased, pour out the upper layer of n-hexane, spread the rice bran and let it dry naturally at room temperature, and collect it for later use.
[0028] 3. Extract crude enzyme solution: Take 3 portions of defatted rice bran, 10g each, add 40ml of phosphate buffer solution (pH7.5) and stir at 5°C for 30min, then pass through double-layer clean gauze. The filtrate was centrifuged at 4°C and 12000 rpm for 30 min to obtain a clarified crude enzyme solution.
[0029] 4. Prepare the substrate solution: weigh 111 μL with a concentration of 2.53×10 -3 Add mol / L linoleic acid into a 10mL volumetric flask, dilute to 10mL with absolute ethanol, take 3.55m...
Embodiment 2
[0037] Take 1 kg of fresh rice bran, pass it through a 300-mesh sieve, take the sieved rice bran, add 10 times the volume of n-hexane, shake and degrease at 180 rpm at room temperature for 40 minutes. Then the upper layer of n-hexane was poured out, and the remaining defatted rice bran was paved and air-dried at room temperature.
[0038] Accurately weigh 111 μL of linoleic acid and add it to a 10 mL volumetric flask, dilute to the mark with absolute ethanol, take 3.55 mL of the above solution, and add 40 μL Tween-20. Ethanol was removed by evaporation under reduced pressure, and the remaining solid was dissolved in 50 mL of 0.05 mol / L Na 2 HPO 4 Then add 0.5 mol / L NaOH dropwise thereto to adjust the pH value to 9.0, and the clarity of the substrate solution is high at this time. The concentrations of linoleic acid and Tween-20 in the substrate solution were 2.53×10 -3 mol / L, 0.08% (w / v).
[0039] Take 3 portions of defatted rice bran, 20g each, add 80ml of phosphate buff...
Embodiment 3
[0043] Accurately weigh 111 μL of linoleic acid and add it to a 10 mL volumetric flask, dilute to the mark with absolute ethanol, take 3.55 mL of the above solution, and add 40 μL Tween-20. Ethanol was removed by evaporation under reduced pressure, and the remaining solid was dissolved in 50 mL of 0.05 mol / L Na 2 HPO 4 Then add 0.5 mol / L NaOH dropwise thereto to adjust the pH value to 9.0, and the clarity of the substrate solution is high at this time. The concentrations of linoleic acid and Tween-20 in the substrate solution were 2.53×10 -3 mol / L, 0.08% (w / v).
[0044] Take 1 kg of fresh rice bran, pass it through a 300-mesh sieve, take the sieved rice bran, add 8 times the volume of n-hexane, shake and degrease at 160 rpm at room temperature for 20 minutes. Then the upper layer of n-hexane was poured out, and the remaining defatted rice bran was paved and air-dried at room temperature.
[0045] Take 3 portions of defatted rice bran, 25g each, add 100ml of phosphate buff...
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