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Method for quantitatively detecting antibiotics

A technology for antibiotics and penicillin, which is applied in biochemical equipment and methods, microbiological determination/inspection, and measuring devices, and can solve problems such as increased effective dose, high instrument cost, and uneconomical and practical

Active Publication Date: 2020-10-27
INST OF AGRO FOOD SCI & TECH CHINESE ACADEMY OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the use of antibiotics will lead to drug resistance of pathogenic microorganisms, so that the effective dose of antibiotics to kill bacteria will continue to increase, posing a potential threat to human health
With the continuous improvement of living standards, people's requirements for food safety are also increasing. Among food safety problems, the problem of antibiotic pollution is more prominent. The existing methods for detecting antibiotics are mostly high-performance liquid chromatography, gas chromatography and Mass spectrometry, these methods have high instrument costs and are not economical and practical

Method used

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  • Method for quantitatively detecting antibiotics
  • Method for quantitatively detecting antibiotics
  • Method for quantitatively detecting antibiotics

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preparation example Construction

[0026] Optionally, in step S1, the method for preparing the dilution of the penicillin-binding protein crude protein supernatant includes: ultrasonically breaking the penicillin-binding protein for 2-4 minutes; centrifuging to obtain the supernatant; centrifuging at 6500-9500rpm for 3-7min , take the supernatant; the preparation method of the diluent of the synthetic peptide comprises: first diluting the synthetic peptide into a mother solution of 0.8-1.2mg / mL with 0.08-0.12M PBS buffer solution, diluting the mother solution with distilled water for 900- 1100 times.

[0027] Optionally, the penicillin-binding protein is PBP1, PBP2 or PBP3, preferably PBP3; the synthetic peptide is pentameric D-Ala or D-Cys-D-Ala-D-Ala.

[0028] Optionally, in step S2 and step S3, the centrifugation method includes: the centrifugation speed is 7500-8500rpm, and the centrifugation time is 3-7min.

[0029] Optionally, in step S2, the drawing method of the antibiotic concentration gradient standa...

Embodiment 1

[0033] Prepare the supernatant of the crude protein of penicillin-binding protein: after the penicillin-binding protein is ultrasonically disrupted for 3 minutes; centrifuge to take the supernatant; centrifuge at 8000rpm for 5 minutes to take the supernatant.

[0034] Firstly, the synthetic peptide was diluted with 0.1 M PBS buffer solution to a 1 mg / mL stock solution, and the stock solution was diluted 1000 times with distilled water.

[0035] Preparation of PBP carboxypeptidase hydrolyzed peptide chain system: dilute penicillin-binding protein crude protein supernatant 5 times to obtain penicillin-binding protein crude protein supernatant dilution and synthetic peptide dilution and mix well.

[0036] Draw the concentration gradient standard calibration curve of penicillin: dilute the penicillin standard solution with water into solutions with 8 concentration gradients of 0.15mM, 0.75mM, 1.5mM, 3.0mM, 4.5mM, 6.0mM, 7.5mM and 9.0mM, add Penicillin-binding protein carboxypeptid...

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Abstract

The invention relates to a method for quantitatively detecting antibiotics. The method comprises the following steps of: uniformly mixing a diluent of penicillin-binding protein crude extraction protein supernatant and a diluent of synthetic peptide to obtain a penicillin-binding protein carboxypeptidase hydrolysis peptide chain system solution; respectively adding antibiotic aqueous solutions with different concentrations into the penicillin-binding protein carboxypeptidase hydrolysis peptide chain system solution to obtain a reaction system solution, centrifuging the reaction system solutionto obtain a supernatant, adding a ninhydrin color developing agent into the supernatant, determining an absorbance value, and establishing a concentration gradient standard correction curve of antibiotics according to the absorbance value; and adding a to-be-detected solution into the penicillin-binding protein carboxypeptidase hydrolysis peptide chain system, carrying out centrifuging, adding the ninhydrin color developing agent, determining the absorbance value of the to-be-detected solution, and calculating the concentration of the antibiotics in the to-be-detected solution according to the concentration gradient standard calibration curve of the antibiotics.

Description

technical field [0001] The present application relates to the field of food testing, in particular to a method for quantitatively detecting antibiotics. Background technique [0002] As a drug that can inhibit and kill bacteria, antibiotics have been widely used in industries such as medical and health care, livestock and poultry breeding, and agricultural production, and have made great contributions to the economic development of society. However, the use of antibiotics will lead to drug resistance of pathogenic microorganisms, which will increase the effective dose of antibiotics to kill bacteria, posing a potential threat to human health. With the continuous improvement of living standards, people's requirements for food safety are also increasing. Among food safety problems, the problem of antibiotic pollution is more prominent. The existing methods for detecting antibiotics are mostly high-performance liquid chromatography, gas chromatography and Mass spectrometry, th...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/78C12Q1/37
CPCG01N21/78C12Q1/37G01N2333/948
Inventor 范蓓黄亚涛金诺王凤忠次顿单吉浩卢嘉李敏敏刘佳萌孙玉凤孙晶邓凯琳
Owner INST OF AGRO FOOD SCI & TECH CHINESE ACADEMY OF AGRI SCI