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Method for extracting nitric oxide reductase in activated sludge and determining activity

A technology of activated sludge and nitric oxide, which is applied in the direction of oxidoreductase, biochemical equipment and methods, and the determination/inspection of microorganisms, to achieve the effect of maintaining enzyme activity, maintaining stability, and simple and fast operation steps

Active Publication Date: 2020-08-21
XI'AN UNIVERSITY OF ARCHITECTURE AND TECHNOLOGY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, there is no relevant report on the extraction of nitric oxide reductase from activated sludge of sewage biological treatment

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] This embodiment 1 provides a method for extracting nitric oxide reductase in activated sludge and measuring its activity, comprising the following steps:

[0046] Take 20mL of the activated sludge sample from the anoxic tank of A / O SBR, centrifuge at 4000r / min for 5min to separate the muddy water in the activated sludge sample, pour the supernatant, collect the solid sample, and dilute the solid sample with buffer to 20mL; among them, the buffer solution is 20mM Tris-HCL buffer solution with a pH of 7.5; use a shaker to re-mix the solution after constant volume, and centrifuge to obtain a solid mud sample; repeat the above washing operation three times; after repeated washing , use the buffer solution to re-dilute the solid sample to 20mL, and mix well to obtain the activated sludge sample.

[0047] Wherein, the buffer solution is prepared by adding KCl to the Tris-HCL buffer solution, wherein the concentration of the Tris-HCL buffer solution is 5 mM, the concentration ...

Embodiment 2

[0055] This embodiment 2 provides a method for extracting nitric oxide reductase in activated sludge and measuring its activity, comprising the following steps:

[0056] Take 20mL of the activated sludge sample from the anoxic tank of A / O SBR, centrifuge at 5000r / min for 15min to separate the muddy water in the activated sludge sample, pour the supernatant, collect the solid sample, and dilute the solid sample with buffer to 20mL; among them, the buffer is 200mM Tris-HCL buffer, pH 8.0; use the shaker to re-mix the solution after constant volume, and centrifuge to obtain a solid mud sample; repeat the above washing operation three times; after repeated washing , use the buffer solution to re-dilute the solid sample to 20mL, and mix well to obtain the activated sludge sample.

[0057] Wherein, the buffer solution is prepared by adding KCl to the Tris-HCL buffer solution, wherein the concentration of the Tris-HCL buffer solution is 50 mM, the capacity of KCl is 150 mM; the pH of...

Embodiment 3

[0065] This embodiment 3 provides a method for extracting nitric oxide reductase in activated sludge and measuring its activity, comprising the following steps:

[0066] Take 20mL of the activated sludge sample from the aerobic tank of A / O SBR, centrifuge at 4000r / min for 10min to separate the muddy water in the activated sludge sample, pour the supernatant, collect the solid sample, and dilute the solid sample with buffer to 20mL; among them, the buffer is 200mM Tris-HCL buffer, pH 8.0; use the shaker to re-mix the solution after constant volume, and centrifuge to obtain a solid mud sample; repeat the above washing operation three times; after repeated washing , use the buffer solution to re-dilute the solid sample to 20mL, and mix well to obtain the activated sludge sample.

[0067] Wherein, the buffer solution is prepared by adding KCl to the Tris-HCL buffer solution, wherein the concentration of the Tris-HCL buffer solution is 50 mM, the concentration of KCl is 150 mM; the...

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Abstract

The invention discloses an extraction and activity determination method of nitric oxide reductase in activated sludge, which comprises the following steps: crushing and centrifuging an activated sludge sample to obtain a supernatant; adding phenethyl alcohol and dodecyl-beta-D-maltoside into the supernatant for a reaction to obtain a nitric oxide reductase sample to-be-detected; in an anaerobic state, adding a buffer solution, a sodium ascorbate solution, a sodium acetate solution and phenazine methyl sulfate into a closed container, and introducing nitric oxide gas to obtain an enzyme activity reaction solution; oscillating a nitric oxide reductase sample to-be-detected in the enzymatic activity reaction solution, and obtaining the activity of nitric oxide reductase by using a gas chromatography method. The efficient extraction of nitric oxide reductase is realized by crushing and centrifuging an activated sludge sample, and the operation steps of the extraction process are simple andrapid; the activity of nitric oxide reductase is determined by gas chromatography, and the determination result is accurate, efficient and stable.

Description

technical field [0001] The invention belongs to the technical field of sewage biological treatment, in particular to a method for extracting and measuring activity of nitric oxide reductase in activated sludge. Background technique [0002] Greenhouse gas nitrous oxide emissions and sludge bulking are two key issues that sewage treatment plants have been facing and urgently need to be solved; there are three ways of generating nitrous oxide in sewage treatment plants, namely incomplete oxidation of hydroxylamine, heterotrophic reaction Nitrification process and denitrification process of nitrifying bacteria; Among them, the incomplete oxidation of hydroxylamine refers to the process of ammonia oxidizing bacteria in the nitrification process, ammonia nitrogen in the periplasm of the cell under the action of transmembrane protein ammonia monooxygenase Generate hydroxylamine; in the process of hydroxylamine being catalyzed and oxidized by amine oxidase to generate nitrite nitro...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/06C12Q1/26
CPCC12N9/0044C12Q1/26C12Y107/01014G01N2333/90688Y02W10/10
Inventor 袁林江杨睿
Owner XI'AN UNIVERSITY OF ARCHITECTURE AND TECHNOLOGY
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