阅读说明：本技术 一种检测禽组织、禽蛋或猪肉中青霉素g残留的确证分析方法 (Confirmation analysis method for detecting penicillin G residue in poultry tissue, poultry egg or pork ) 是由 谢恺舟 刘楚君 谢星 庞茂达 王波 严祖浩 王旭堂 刁志祥 张培杨 郭亚文 李洋 于 2020-06-12 设计创作，主要内容包括：本发明涉及兽药残留检测领域,具体涉及一种检测禽组织、禽蛋或猪肉中青霉素G残留的确证分析方法,包括以下步骤：将禽组织、禽蛋或猪肉样品通过加速溶剂萃取技术提取,固相萃取净化,三甲基硅烷基重氮甲烷衍生,经GC-MS/MS检测；所述禽组织为鸡肉、鸡肝、鸡肾、鹅肉或鸭肉；所述禽蛋为鸡蛋或鸭蛋。本方法分析效率更高、回收率、准确度和灵敏度高,重现性好,适合在批量样品分析中应用与推广。该方法能高效、准确的检测禽组织、禽蛋及猪肉中青霉素G残留,且满足中国农业农村部和欧盟兽药残留检测要求。(The invention relates to the field of veterinary drug residue detection, in particular to a confirmation analysis method for detecting penicillin G residue in poultry tissues, poultry eggs or pork, which comprises the following steps: extracting fowl tissue, fowl egg or pork sample by accelerated solvent extraction technology, solid phase extracting and purifying, deriving trimethylsilyl diazomethane, and detecting by GC-MS/MS; the poultry tissue is chicken, chicken liver, chicken kidney, goose meat or duck meat; the poultry eggs are eggs or duck eggs. The method has the advantages of higher analysis efficiency, high recovery rate, high accuracy and sensitivity and good reproducibility, and is suitable for application and popularization in batch sample analysis. The method can efficiently and accurately detect the penicillin G residue in the poultry tissues, the poultry eggs and the pork, and meets the requirement of veterinary drug residue detection in rural areas of China agriculture and European Union.)

1. A confirmation analysis method for detecting penicillin G residue in poultry tissues, poultry eggs or pork is characterized by comprising the following steps:

extracting fowl tissue, fowl egg or pork sample by accelerated solvent extraction technology, solid phase extracting and purifying, deriving trimethylsilyl diazomethane, and detecting by GC-MS/MS; the poultry tissue is chicken, chicken liver, chicken kidney, goose meat or duck meat; the poultry eggs are eggs or duck eggs.

2. The confirmatory analysis method for detecting penicillin G residues in poultry tissues, eggs or pork as claimed in claim 1, wherein the qualitative and quantitative reaction monitoring methods are selected from full scan and selective ion monitoring methods.

3. The confirmatory analysis method for detecting penicillin G residues in poultry tissues, eggs or pork as claimed in claim 1, wherein the GC-MS/MS detection gas chromatography conditions are as follows: TG-1MS is used as a capillary chromatographic column; high-purity helium is taken as carrier gas, and the flow rate of the carrier gas column is 1.0 mL/min; the high purity helium gas has a purity of > 99.999% and a pressure of 60 psi.

4. The confirmatory analysis method for detecting penicillin G residues in poultry tissues, eggs or pork according to claim 1, wherein the temperature programming step of GC-MS/MS detection is at an initial temperature of 100 ℃ for 1 min; raising the temperature to 220 ℃ at a speed of 30 ℃/min, and keeping the temperature for 1 min; raising the temperature to 280 ℃ at 30 ℃/min and keeping the temperature for 5 min.

5. The confirmatory analysis method for detecting penicillin G residues in avian tissues, poultry eggs or pork as claimed in claim 1, wherein the mass spectrometric conditions of GC-MS/MS detection are: electron bombardment ion source; ionization energy of 70 eV; high-purity argon is used as collision gas; the high purity argon gas has a purity of > 99.999% and a pressure of 40 psi.

6. The confirmatory assay for the detection of penicillin G residues in avian tissues, avian eggs or pork as claimed in claim 1, wherein the ion source temperature is 280 ℃; the temperature of the transmission line is 280 ℃; the solvent delay time was 5.0 min.

7. The confirmatory analysis method for detecting penicillin G residues in poultry tissues, eggs or pork as claimed in claim 1, wherein the sample inlet temperature for GC-MS/MS detection is 280 ℃.

8. The confirmatory analysis method for detecting penicillin G residues in poultry tissue, eggs or pork as claimed in claim 1, wherein the carrier gas flow rate is 1.0 mL/min; 2min later, the valve is opened, the carrier gas saving time is 2min, and the carrier gas saving flow is 20.0 mL/min; sample introduction volume: 1.0. mu.L.

9. The confirmatory analysis method for detecting penicillin G residues in poultry tissue, eggs or pork as claimed in claim 1, wherein the poultry tissue or pork sample is extracted with 0.2M phosphate buffer solution, the eggs are extracted with 80% acetonitrile; the pH of the 0.2M phosphate buffer solution was 8.0.

10. The confirmatory analysis method for detecting penicillin G residues in poultry tissues, eggs or pork as claimed in claim 1, wherein the extraction conditions of the accelerated solvent extractor are 1500psi, 30 ℃.

Technical Field

The invention relates to the field of veterinary drug residue detection, in particular to a confirmation analysis method for detecting penicillin G residue in poultry tissues, poultry eggs or pork.

Background

At home and abroad, the detection methods of penicillin G residue mainly comprise a microbiological method, an immunoassay method, a thin-layer chromatography, a liquid chromatography-tandem mass spectrometry, a gas chromatography-mass spectrometry and the like, and the detection of penicillin G in animal food by combining a GC (gas chromatography) -external standard method and a GC-MS (gas chromatography-mass spectrometry) combined with an internal standard method has been reported in documents, so that the pretreatment process of a sample is complex, time and labor are consumed, and an internal standard substance is expensive and has a low recovery rate. And the method for detecting penicillin G residue in poultry tissues and pork by combining the GC-MS/MS with the external standard method is not reported, and compared with GC and GC-MS reported in the literature, the method has the advantages of simple sample preparation process, time and labor saving and test cost saving. The method has the advantages of more accurate qualitative and quantitative determination, good accuracy and sensitivity, high recovery rate, good repeatability and reproducibility, and fills up the blank of GC-MS/MS detection methods for penicillin G residue in poultry tissues, poultry eggs or pork at home and abroad.

Disclosure of Invention

In order to accurately and efficiently detect the penicillin G residue in poultry tissues (chicken, chicken liver, chicken kidney, goose meat and duck meat), eggs, duck eggs (whole eggs, egg white and egg yolk) and pork, the invention provides a confirmation analysis method for detecting the penicillin G residue in the poultry tissues, the eggs or the pork, and the method has the advantages of accurate determination and quantification, higher recovery rate and good sensitivity.

The technical scheme of the invention is as follows:

a confirmation analysis method for detecting penicillin G residue in poultry tissues, poultry eggs or pork is characterized by comprising the following steps:

extracting fowl tissue, fowl egg or pork sample by accelerated solvent extraction technology, solid phase extracting and purifying, deriving trimethylsilyl diazomethane, and detecting by GC-MS/MS; the poultry tissue is chicken, chicken liver, chicken kidney, goose meat or duck meat; the poultry eggs are eggs or duck eggs.

Furthermore, the method is characterized by full scanning and selective ion monitoring, and the reaction monitoring is selected for quantification.

Further, the GC-MS/MS detection gas chromatography conditions are as follows: TG-1MS is used as a capillary chromatographic column; high-purity helium is taken as carrier gas, and the flow rate of the carrier gas column is 1.0 mL/min; the high purity helium gas has a purity of > 99.999% and a pressure of 60 psi.

Further, the temperature programming step of GC-MS/MS detection is that the initial temperature is 100 ℃, and the temperature is kept for 1 min; raising the temperature to 220 ℃ at a speed of 30 ℃/min, and keeping the temperature for 1 min; raising the temperature to 280 ℃ at 30 ℃/min and keeping the temperature for 5 min.

Further, the mass spectrum conditions of GC-MS/MS detection are as follows: electron bombardment ion source; ionization energy of 70 eV; high-purity argon is used as collision gas; the high purity argon gas has a purity of > 99.999% and a pressure of 40 psi.

Further, the ion source temperature is 280 ℃; the temperature of the transmission line is 280 ℃; the solvent delay time was 5.0 min.

Further, the injection port temperature for GC-MS/MS detection was 280 ℃.

Further, the flow rate of the carrier gas is 1.0 mL/min; 2min later, the valve is opened, the carrier gas saving time is 2min, and the carrier gas saving flow is 20.0 mL/min; sample introduction volume: 1.0. mu.L.

Further, poultry tissue or pork samples are extracted by 0.2M phosphate buffer solution, and poultry eggs are extracted by 80% acetonitrile; the pH of the 0.2M phosphate buffer solution was 8.0.

Further, the extraction conditions of the accelerated solvent extractor were 1500psi, 30 ℃.

Advantageous effects

In the invention, TG-1MS (30.0m multiplied by 0.25 mu m multiplied by 0.25mm i.d.) capillary chromatographic columns are adopted, the qualitative mode of full Scanning (SCAN) and Selective Ion Monitoring (SIM) and the quantitative mode of selective reaction monitoring (Auto SRM) are adopted, and finally the chromatographic peak shape of the penicillin G derivative product is good (namely the sensitivity is high), the retention time of an analyte is moderate, and no interference of other impurity peaks exists.

The invention provides a gas chromatography-tandem mass spectrometry detection (GC-MS/MS) method. Compared with other detection methods, the method has the advantages of higher analysis efficiency, high recovery rate, high accuracy and sensitivity and good reproducibility, and is suitable for application and popularization in batch sample analysis. The method can efficiently and accurately detect the penicillin G residue in the poultry tissues, the poultry eggs and the pork, and meets the requirement of veterinary drug residue detection in rural areas of China agriculture and European Union. Drawings

FIG. 1 Total ion current chromatogram (TIC) and Mass Chromatogram (MC) of quantitative and qualitative ions of blank Chicken sample (a) and blank Chicken sample with addition of 50.0. mu.g/kg penicillin G standard (b)

FIG. 2 Total Ion Chromatogram (TIC) and Mass Chromatogram (MC) of quantitative and qualitative ions of blank chicken liver sample (a) and blank chicken liver sample with 50.0. mu.g/kg penicillin G standard (b)

FIG. 3 shows the total ion current chromatogram (TIC) and the Mass Chromatogram (MC) of quantitative and qualitative ions of a blank chicken kidney sample (a) and a blank chicken kidney sample added with 50.0. mu.g/kg penicillin G standard (b)

FIG. 4 shows the total ion current chromatogram (TIC) and the Mass Chromatogram (MC) of quantitative and qualitative ions of a goose meat sample (a) and a goose muscle sample (b) added with 50.0. mu.g/kg penicillin G standard (b)

FIG. 5 shows the total ion current chromatogram (TIC) and the Mass Chromatogram (MC) of quantitative and qualitative ions of a blank duck meat sample (a) and a blank duck muscle sample added with 50.0 μ G/kg penicillin G standard (b)

FIG. 6 shows the total ion current chromatogram (TIC) and the Mass Chromatogram (MC) of quantitative and qualitative ions of a blank whole egg sample (a) and a blank whole egg sample (b) to which a penicillin G standard (b) was added in an amount of 25.0. mu.g/kg

FIG. 7 Total Ion Chromatogram (TIC) and Mass Chromatogram (MC) of quantitative and qualitative ions of blank egg white sample (a) and blank egg white sample with addition of 25.0. mu.g/kg penicillin G standard (b)

FIG. 8 Total Ion Chromatogram (TIC) and Mass Chromatogram (MC) of quantitative and qualitative ions of blank yolk sample (a) and blank yolk sample with addition of 25.0. mu.g/kg penicillin G standard (b)

FIG. 9 shows the total ion current chromatogram (TIC) and the Mass Chromatogram (MC) of quantitative and qualitative ions of a blank duck whole egg sample (a) and a blank duck whole egg sample (b) added with 25.0 μ G/kg penicillin G standard (b)

FIG. 10 Total Ion Chromatogram (TIC) and Mass Chromatogram (MC) of quantitative and qualitative ions of blank duck egg white sample (a) and blank duck egg white sample with addition of 25.0. mu.g/kg penicillin G standard (b)

FIG. 11 Total ion current chromatogram (TIC) and Mass Chromatogram (MC) of quantitative and qualitative ions of blank duck egg yolk sample (a) and blank duck egg yolk sample with addition of 25.0. mu.g/kg penicillin G standard (b)

FIG. 12 Total ion flux chromatogram (TIC) and Mass Chromatogram (MC) of quantitative and qualitative ions of a pork blank sample (a) and a pork blank sample with 50.0. mu.g/kg penicillin G standard (b)

FIG. 13 Standard Curve for penicillin G addition to the blank Chicken substrate

FIG. 14 standard curve of penicillin G addition to chicken liver matrix blank

FIG. 15 Standard Curve for penicillin G addition to the blank Chicken Kidney substrate

FIG. 16 standard curve of penicillin G addition to a blank goose meat matrix

FIG. 17 Standard Curve for adding penicillin G to a blank Duck matrix

FIG. 18 Standard Curve of matrix addition penicillin G in blank Chicken Whole egg

FIG. 19 Standard Curve for penicillin G addition to blank egg white matrix

FIG. 20 standard curve of penicillin G addition to a blank egg yolk matrix

FIG. 21 standard curve of penicillin G addition to blank duck whole egg matrix

FIG. 22 standard curve of penicillin G addition to blank duck egg white matrix

FIG. 23 standard curve of penicillin G addition to blank duck egg yolk matrix

FIG. 24 Standard Curve of penicillin G addition to pork blank matrix

Detailed Description