CN114062551A

CN114062551A - Method for detecting content of triflumizole amine and metabolites thereof on vegetables and fruits

Info

Publication number: CN114062551A
Application number: CN202111356435.6A
Authority: CN
Inventors: 王伟民; 韩薇; 唐红霞; 董茂锋
Original assignee: Shanghai Academy of Agricultural Sciences
Current assignee: Shanghai Academy of Agricultural Sciences
Priority date: 2021-11-16
Filing date: 2021-11-16
Publication date: 2022-02-18
Anticipated expiration: 2041-11-16
Also published as: CN114062551B

Abstract

The invention discloses a method for detecting the content of fluoroandenazole bacteria amine and metabolites thereof on vegetables and fruits, and relates to the technical field of pesticide detection. The method for detecting the content of the triflumizole amine and the metabolite thereof on the vegetables and fruits comprises the following steps: s10, crushing vegetables and fruits, adding an acetonitrile water solution, stirring, performing ultrasonic treatment, and centrifuging to obtain a first supernatant and a precipitate; s20, adding water into the precipitate, stirring and centrifuging to obtain a second supernatant, and combining and uniformly mixing the second supernatant and the first supernatant to obtain a mixed solution; s30, after the mixed solution flows through the solid phase extraction column, the solid phase extraction column is eluted by methanol to obtain eluent; s40, drying the eluent under protective gas to obtain a residue, dissolving the residue in methanol to obtain a solution, and filtering the solution to obtain a solution to be detected; and S50, measuring the solution to be measured by using a liquid chromatography-mass spectrometer to obtain the content of the fluoroandenazole amine and the metabolite thereof on the vegetables and fruits. The method is environment-friendly and has high accuracy.

Description

Method for detecting content of triflumizole amine and metabolites thereof on vegetables and fruits

Technical Field

The invention relates to the technical field of pesticide detection, and particularly relates to a method for detecting the content of fluoroandenazole hydrochloride and metabolites thereof on vegetables and fruits.

Background

Fluindacamide (IR9792) is a novel pyrazole amide fungicide developed by Isagro, Italy and FMC corporation, and can control various plant diseases such as botrytis, Alternaria, Blastomyces nigra and Coryneba, etc. It is predicted that diflufenican can be applied to various crops such as vegetables, fruits and oilseeds, and has been registered in china, the european union and the united states. The fluindanconazole amine is used as a newly developed pesticide, the reported literature on the fluindanconazole amine is less at present, but as the pesticide evaluation and technical support, the detection of the fluindanconazole amine and the metabolite thereof is crucial to the reasonable use, the residue evaluation and the food safety.

At present, the domestic detection method of the temporary fluoride-free indoxacarb and the metabolite thereof cannot simultaneously and accurately analyze the content of the fluoride-free indoxacarb and the metabolite thereof by using the most widely-applied national standard analysis method.

Disclosure of Invention

The invention mainly aims to provide a method for detecting the content of the fluoroandenazole amine and the metabolites thereof on vegetables and fruits, and aims to provide a method for detecting the content of the fluoroandenazole amine and the metabolites thereof on vegetables and fruits, which is green, environment-friendly and accurate.

In order to achieve the purpose, the invention provides a method for detecting the content of the fluoroandenazole hydrochloride and the metabolites thereof on vegetables and fruits, which comprises the following steps:

s10, crushing vegetables and fruits, adding an acetonitrile water solution, stirring, performing ultrasonic treatment, and centrifuging to obtain a first supernatant and a precipitate;

s20, adding water into the precipitate, stirring and centrifuging to obtain a second supernatant, and combining and uniformly mixing the second supernatant and the first supernatant to obtain a mixed solution;

s30, after the mixed solution flows through the solid phase extraction column, the solid phase extraction column is eluted by methanol to obtain eluent;

s40, drying the eluent under protective gas to obtain a residue, dissolving the residue in methanol to obtain a solution, and filtering the solution to obtain a solution to be detected;

and S50, measuring the solution to be measured by using a liquid chromatography-mass spectrometer to obtain the content of the fluoroandenazole amine and the metabolite thereof on the vegetables and fruits.

Optionally, in step S10, the volume ratio of acetonitrile to water in the acetonitrile water solution is 2 (3-4).

Optionally, the volume of the acetonitrile water solution added per gram of the vegetables and fruits is 1-2 mL.

Optionally, after step S20 and before step S30, the method further includes:

and S31, activating the solid phase extraction column by using methanol and water sequentially.

Alternatively, in step S30, the solid phase extraction column is an MCX solid phase extraction column; and/or the presence of a gas in the gas,

the vegetable and fruit comprises at least one of tomato, cucumber, eggplant, pepper, spinach, peach, jujube and orange.

Optionally, in step S40, the shielding gas includes nitrogen.

Optionally, in step S40, the temperature during drying is 35-45 ℃.

Optionally, in step S40, the dissolution solution is filtered with a nylon filter membrane.

Optionally, step S50 includes:

s51, preparing a plurality of standard solutions of the fluoroandene myclobutanil and metabolites thereof, respectively performing liquid chromatography-mass spectrometry, and performing a standard curve according to the liquid chromatography-mass spectrometry result and the standard solution concentration;

and S52, obtaining the content of the thiabendazole amine and the metabolite content thereof on the vegetables and fruits according to the liquid chromatogram-mass spectrometer determination result of the solution to be determined and the standard curve.

Alternatively, in step S50,

when the detection is carried out by a liquid chromatography-mass spectrometer, the used chromatographic column is an InertSustain swift C18 chromatographic column; and/or the presence of a gas in the gas,

when the detection is carried out by a liquid chromatogram-mass spectrometer, the detection mode is multiple reaction monitoring.

The invention provides a method for detecting the content of fluoroandenazole amine and metabolites thereof on vegetables and fruits, which adopts a solid-phase extraction treatment method, combines liquid chromatography-mass spectrometry, and simultaneously determines the fluoroandenazole amine and 5 metabolites thereof in vegetables and fruits.

Drawings

In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, it is obvious that the drawings in the following description are only some embodiments of the present invention, and for those skilled in the art, other drawings can be obtained according to the structures shown in the drawings without creative efforts.

Fig. 1 is a schematic flow chart of an embodiment of a method for detecting content of fluoroandenazole amine and metabolites thereof on vegetables and fruits according to the present invention.

The implementation, functional features and advantages of the objects of the present invention will be further explained with reference to the accompanying drawings.

Detailed Description

The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.

It should be noted that, if directional indications (such as upper, lower, left, right, front, rear, outer and inner … …) are involved in the embodiment of the present invention, the directional indications are only used to explain the relative position relationship between the components, the motion situation, etc. in a specific posture (as shown in the drawing), and if the specific posture is changed, the directional indications are changed accordingly.

In addition, if there is a description of "first", "second", etc. in an embodiment of the present invention, the description of "first", "second", etc. is for descriptive purposes only and is not to be construed as indicating or implying relative importance or implicitly indicating the number of technical features indicated. Thus, a feature defined as "first" or "second" may explicitly or implicitly include at least one such feature. In addition, the meaning of "and/or" appearing throughout includes three juxtapositions, exemplified by "A and/or B" including either A or B or both A and B. In addition, technical solutions between various embodiments may be combined with each other, but must be realized by a person skilled in the art, and when the technical solutions are contradictory or cannot be realized, such a combination should not be considered to exist, and is not within the protection scope of the present invention.

In view of the above, the invention provides a method for detecting the content of the fluoroandenazole amine and the metabolite thereof on the vegetables and fruits, and aims to provide a method for detecting the content of the fluoroandenazole amine and the metabolite thereof on the vegetables and fruits, which is green, environment-friendly and accurate. In the attached drawings, fig. 1 is a schematic flow chart of an embodiment of the method for detecting the content of the fluoroandenazole amine and the metabolites thereof on the vegetables and fruits provided by the invention.

The invention provides a method for detecting the content of fluoroandenazole bacteria amine and metabolites thereof on vegetables and fruits, which comprises the following steps:

s10, crushing the vegetables and fruits, adding an acetonitrile water solution, stirring, performing ultrasonic treatment, and centrifuging to obtain a first supernatant and a precipitate.

In the step, the fluoroandenazole amine and metabolites thereof are extracted by using an acetonitrile aqueous solution, preferably, the volume ratio of acetonitrile to water in the acetonitrile aqueous solution is 2 (3-4). For example, 2:3, 2:4, etc., at the above concentrations, the dissolution of the fluoroandenazole amine and its metabolites therein is favored.

Furthermore, the volume of the acetonitrile water solution added into each gram of vegetables and fruits is 1-2 mL, and researches show that under the mixture ratio, the thiabendazole amine and metabolites thereof can be basically dissolved, and the concentration is moderate.

And S20, adding water into the precipitate, stirring, centrifuging to obtain a second supernatant, and mixing and uniformly mixing the second supernatant and the first supernatant to obtain a mixed solution.

In order to fully extract the fluoroandenazole amine and metabolites thereof, the precipitate obtained by the first centrifugation is preferably added into water again for the second extraction, so that the accuracy of content detection is ensured.

And S30, flowing the mixed solution through a solid phase extraction column, and eluting the solid phase extraction column with methanol to obtain an eluent.

The method comprises the following steps of extracting the fluoroandenazole bacteria amine and metabolites thereof in the mixed solution into a solid phase extraction column by using the solid phase extraction column, wherein the solid phase extraction column preferably adopts an MCX solid phase extraction column, the MCX solid phase extraction column is a mixed strong cation exchange adsorbent obtained by bonding sulfonic groups on the surface of highly cross-linked PS/DVB (polyethylene/divinylbenzene copolymer), and the mixed strong cation exchange adsorbent has reversed phase and cation exchange dual retention performances and has good retention capacity on alkaline compounds; the specific surface area is large, and the ion exchange capacity is high; the pH tolerance range is wide (the pH is 1-14), and the pH is stable in an organic solution. In addition, before the extraction, it is preferable that the solid phase extraction column is activated, and specifically, after step S20 and before step S30, the method further includes:

Specifically, the solid phase extraction column is activated by using methanol and water, namely the methanol and the water sequentially flow through the solid phase extraction column, so that the solid phase extraction column is favorable for fully extracting the thiabendazole and metabolites thereof in the mixed solution.

S40, drying the eluent under protective gas to obtain a residue, dissolving the residue in methanol to obtain a solution, and filtering the solution to obtain a solution to be detected.

In the step, the eluent is dried by blowing, methanol is adopted as the eluent, so that the eluent is beneficial to complete volatilization and is easy to dry, preferably, nitrogen is adopted as protective gas, the price is low, the eluent is easy to obtain, the temperature for drying is 35-45 ℃, for example, 35 ℃, 36 ℃, 37 ℃, 38 ℃, 39 ℃, 40 ℃, 41 ℃, 42 ℃, 43 ℃, 44 ℃, 45 ℃ and the like, and preferably 40 ℃, so that the thiabendazole amine and metabolites thereof can not volatilize or decompose on the premise of ensuring that the methanol is quickly volatilized.

In the embodiment of the invention, the filtration adopts a nylon filter membrane, the material is aliphatic nylon, the nylon filter membrane has good hydrophilicity and is resistant to acid and alkali with proper concentration, the nylon filter membrane is not only suitable for aqueous solution containing acid and alkali, but also suitable for organic solvent, such as alcohols, hydrocarbons, ethers, esters, ketones, homologues of benzene and benzene, dimethylformamide, dimethyl sulfoxide and the like, and the nylon filter membrane is one of microporous filter membranes with the widest application range. In the present example, a 0.22 μm nylon filter was used.

The method comprises the following steps of determining the content of the fluoroandenazole bacteria amine and metabolites thereof in a solution to be determined, and specifically comprises the following steps:

Preferably, when a liquid chromatography-mass spectrometer is used, the used chromatographic column is an InertSustain swift C18 chromatographic column, the InertSustain swift C18 chromatographic column matrix adopts high-purity spherical silica gel, the inerting treatment is carried out on the matrix, the surface inertness is high, a sharp peak shape can be obtained for the drug with strong adsorbability, and in addition, the column loss of the chromatographic column is low, so that the peak is quicker to appear, and the peak shape is better.

In addition, during measurement, the detection mode is preferably multiple reaction monitoring, and the Multiple Reaction Monitoring (MRM) can be used for carrying out double mass screening on the structure specificity of organic molecules and parent ions and daughter ions, so that the noise interference of signals can be obviously reduced, and the detection sensitivity and the repeatability of target molecules can be improved.

The technical scheme of the invention is that a solid-phase extraction treatment method is adopted, liquid chromatography-mass spectrometry is combined, and meanwhile, the fluoroandenazole amine and 5 metabolites thereof in vegetables and fruits are measured.

The present invention is not limited to the vegetables and fruits, and preferably, the vegetables and fruits include at least one of tomato, cucumber, eggplant, pepper, spinach, peach, jujube and citrus. The disease caused by the triflumizole amine mainly relates to tomato, cucumber, eggplant, hot pepper, spinach, peach, jujube and orange, so that the 8 kinds of vegetables and fruits are selected, and the pertinence is strong.

Specifically, when data processing is performed, the result measurement is calculated by adopting a standard curve or a single-point method quantitative result according to the following formula:

X＝C×n×V/m

in the formula:

x: the residual amount of the component to be measured in milligrams per kilogram (mg/kg) of the sample;

c: measured constituent solution concentrations in milligrams per liter (mg/L) obtained from a standard working curve or single point method;

n: dilution times in the pretreatment process;

v: volume fixed in milliliters (mL);

m: the sample is weighed in grams (g).

The main metabolites of the fluoroandene ipconazole amine are 5, namely 3-hydroxy-IR9792, Desmethyl-IR9792-N1-Glucoside, 1-hydroxymethyl-IR9792, 1-hydroxymethyl-N-Desmethyl-IR9792 and 1-carboxy-IR9792 respectively.

The following provides an embodiment of the method for detecting the content of the fluoroandenazole bacteria amine and the metabolites thereof on the vegetables and fruits, provided by the invention:

(1) crushing 5g of vegetables and fruits, placing the crushed vegetables and fruits into a 50mL centrifuge tube, adding 5-10 mL of acetonitrile water solution (the volume ratio of acetonitrile to water is 2 (3-4)), carrying out vortex stirring for 20min, then carrying out ultrasonic treatment for 15min, and carrying out centrifugation at 5000rpm for 5min to obtain a first supernatant and a precipitate, wherein the first supernatant is transferred into another centrifuge tube;

(2) adding 5mL of water into the precipitate, stirring in a vortex for 5min, centrifuging at 5000rpm for 5min to obtain a second supernatant, and mixing and uniformly mixing the second supernatant and the first supernatant to obtain a mixed solution;

(3) activating the MCX solid-phase extraction column by using 5mL of methanol and 5mL of water in sequence, and eluting the MCX solid-phase extraction column by using 5mL of methanol after the mixed solution flows through the MCX solid-phase extraction column to obtain eluent;

(4) drying the eluent under the protection of nitrogen at 35-45 ℃ to obtain a residue, dissolving the residue in 1mL of methanol to obtain a dissolved solution, and filtering the dissolved solution by using a 0.22-micron nylon filter membrane to obtain a solution to be detected;

(5) preparing a plurality of standard solutions of the fluoroandene myclobutanil and metabolites thereof, respectively performing liquid chromatography-mass spectrometry, and performing a standard curve according to the liquid chromatography-mass spectrometry result and the concentration of the standard solutions; and obtaining the content of the fluoroandenazole amine and the metabolite thereof on the vegetables and fruits according to the determination result of the liquid to be determined by the liquid chromatography-mass spectrometer and the standard curve, wherein when the determination is performed by the liquid chromatography-mass spectrometer, the used chromatographic column is an InertSustain swift C18 chromatographic column, and the detection mode is multiple reaction monitoring.

The technical solutions of the present invention are further described in detail with reference to the following specific examples, which should be understood as merely illustrative and not limitative.

Example 1

(1) Crushing 5g of vegetables and fruits, placing the crushed vegetables and fruits into a 50mL centrifuge tube, adding 5mL of acetonitrile water solution (the volume ratio of acetonitrile to water is 2:3), carrying out vortex stirring for 20min, then carrying out ultrasonic treatment for 15min, carrying out centrifugation at 5000rpm for 5min to obtain a first supernatant and a precipitate, and transferring the first supernatant into another centrifuge tube;

(4) drying the eluent under the protection of nitrogen at 40 ℃ to obtain a residue, dissolving the residue in 1mL of methanol to obtain a dissolved solution, and filtering the dissolved solution by using a 0.22-micron nylon filter membrane to obtain a solution to be detected;

Example 2

(1) Crushing 5g of vegetables and fruits, placing the crushed vegetables and fruits into a 50mL centrifuge tube, adding 10mL of acetonitrile water solution (the volume ratio of acetonitrile to water is 2:4), carrying out vortex stirring for 20min, then carrying out ultrasonic treatment for 15min, carrying out centrifugation at 5000rpm for 5min to obtain a first supernatant and a precipitate, and transferring the first supernatant into another centrifuge tube;

(4) drying the eluent under the protection gas nitrogen of 35 ℃ to obtain a residue, dissolving the residue in 1mL of methanol to obtain a dissolved solution, and filtering the dissolved solution by using a nylon filter membrane of 0.22 mu m to obtain a solution to be detected;

Example 3

(1) Crushing 5g of vegetables and fruits, placing the crushed vegetables and fruits into a 50mL centrifuge tube, adding 7.5mL of acetonitrile water solution (the volume ratio of acetonitrile to water is 2:3.5), carrying out vortex stirring for 20min, then carrying out ultrasonic treatment for 15min, carrying out centrifugation at 5000rpm for 5min to obtain a first supernatant and a precipitate, and transferring the first supernatant into another centrifuge tube;

(4) drying the eluent under the protection of nitrogen at 45 ℃ to obtain a residue, dissolving the residue in 1mL of methanol to obtain a dissolved solution, and filtering the dissolved solution by using a 0.22-micron nylon filter membrane to obtain a solution to be detected;

Example 4

(1) Crushing 5g of vegetables and fruits, placing the crushed vegetables and fruits into a 50mL centrifuge tube, adding 10mL of acetonitrile water solution (the volume ratio of acetonitrile to water is 2:3), carrying out vortex stirring for 20min, then carrying out ultrasonic treatment for 15min, carrying out centrifugation at 5000rpm for 5min to obtain a first supernatant and a precipitate, and transferring the first supernatant into another centrifuge tube;

(4) drying the eluent under the protection gas nitrogen at 38 ℃ to obtain a residue, dissolving the residue in 1mL of methanol to obtain a dissolved solution, and filtering the dissolved solution by using a 0.22-micron nylon filter membrane to obtain a solution to be detected;

Example 5

(1) Crushing 5g of vegetables and fruits, placing the crushed vegetables and fruits into a 50mL centrifuge tube, adding 7mL of acetonitrile water solution (the volume ratio of acetonitrile to water is 2:4), carrying out vortex stirring for 20min, then carrying out ultrasonic treatment for 15min, carrying out centrifugation at 5000rpm for 5min to obtain a first supernatant and a precipitate, and transferring the first supernatant into another centrifuge tube;

(4) drying the eluent under the protection gas nitrogen at 43 ℃ to obtain a residue, dissolving the residue in 1mL of methanol to obtain a dissolved solution, and filtering the dissolved solution by using a 0.22-micron nylon filter membrane to obtain a solution to be detected;

Taking example 1 as an example, the specific process and effect of the method for detecting the content of the fluoroandenazole amine and metabolites thereof on the vegetables and fruits will be described.

During the analytical determination, the column used was InertSustain swift C18 column (2.1X 50mm, 3 μm), column temperature: 40 ℃, flow rate: 0.4mL/min, sample size: 5 mu L in a scanning mode of a negative ion mode of fluroindolizidine (IR9792), a negative ion mode of 3-hydroxy-IR9792 and a negative ion mode of Desmethyl-IR9792-N1-Glucoside, and the gradient elution conditions are as shown in the following table 1:

TABLE 1 gradient elution conditions

The scanning mode is as follows: 1-hydroxyymethyl-IR 9792 anion mode, 1-hydroxyymethyl-N-Desmethyl-IR 9792 anion mode, 1-carboxy-IR9792 anion mode;

gradient elution conditions are shown in table 2 below:

TABLE 2 gradient elution conditions

t/min	Flow rate (mL/min)	Methanol (%)	Water (%)
				0.10	0.4	10	90
4.00	0.4	70	30
				5.00	0.4	80	20
6.50	0.4	10	90
				8.50	0.4	10	90

An ion source: electrospray ion source ESI; the scanning mode is as follows: a negative ion mode; flow rate of the atomizer: 3L/min; heater flow rate: 10L/min; interface temperature: 300 ℃; DL temperature: 250 ℃; temperature of the heating block: 400 ℃; flow rate of drying gas: 10L/min; the detection mode is as follows: multiple Reaction Monitoring (MRM), MS/MS parameters of fluoroandenazole amine and metabolites thereof are shown in Table 3.

TABLE 3 MS/MS parameters of Fluindacazole amine and its metabolites

a collision energy of the primary product ions; b collision energy of secondary product ions

After the instrument is stabilized, the retention time, chromatographic peak shape, fragment ion pair of the compound in the sample are consistent with the standard solution of flurindoxacarb, 3-hydroxy-IR9792, Desmethyl-IR9792-N1-Glucoside, 1-hydroxymethyl-IR9792, 1-hydroxymethyl-N-Desmethyl-IR9792 and 1-carboxy-IR9792 under the chromatographic conditions.

Respectively weighing a proper amount of fluoroandenazole bacteria amine (IR9792), 3-hydroxy-IR9792, Desmethyl-IR9792-N1-Glucoside, 1-hydroxymethyl-IR9792, 1-hydroxymethyl-N-Desmethyl-IR9792 and 1-carboxy-IR9792 to prepare a 10mg/L mixed standard working solution. And then diluting the mixed standard working solution into 0.001, 0.01, 0.05, 0.1 and 0.5mg/L series of matrix mixed standard solutions by using the blank matrix solution, and making a standard curve by using the concentration of the target compound matrix standard solution and the area of the monitored ion peak. Quantitative determination is carried out by adopting an external standard method, and the matrix standard solution is prepared for use.

1. Minimum limit of quantitation

Under the above chromatographic conditions, the minimum limit of quantitation (LOQ; minimum detectable concentration) of the fluoroandenazole amine, 3-hydroxy-IR9792, Desmethyl-IR9792-N1-Glucoside, 1-hydroxymethyl-IR9792, 1-hydroxymethyl-N-Desmethyl-IR9792 and 1-carboxy-IR9792 was 0.001 mg/kg.

2. Addition recovery test

The method is verified according to Chinese pesticide analysis method guidelines (NY/T788-2018,2018) and SANTE standards (Commission, 2019), three different addition levels are set according to 0.001mg/kg (minimum limit of quantitation), 0.01mg/kg and 0.1mg/kg, 3-level concentration of fluroindolizine, 3-hydroxy-IR9792, Desmethyl-IR9792-N1-Glucoside, 1-hydroxy-IR 9792, 1-hydroxy-N-Desethyl-IR 9792 and 1-carboxy-IR9792 are respectively added into blank brown rice, rice husks and straws, the method is repeated for 5 times per level, the recovery rate is determined by the analysis method, 3 addition levels (0.001, 0.01 and 0.1mg/kg) and 5 times of repeated sulfenamide and metabolic substances thereof are added into a sample of a homogenized food after being homogenized for 15 minutes, and sealed at-20 ℃ until analyzed. The analytical methods were evaluated for linearity, detection limit, precision (RSD) and Matrix Effect (ME), and the results are shown in Table 4.

TABLE 4 Standard Curve, linearity, detection Limit, precision (RSD) and Matrix Effect (ME) of Fluindacarboxamine and its metabolites in 8 matrices

Wherein, the calibration curves in the solvent are respectively as follows: ninhydrin iparidinamine y ═ 95548100x +20712.8(0.9992), 3-hydroxy-IR9792 y ═ 236324000x +673765(0.9993), Desmethyl-IR 9792-N1-glucose y ═ 21104100x +134807(0.9990), 1-hydroxymethyl-IR9792 y ═ 244732000x +1721230(0.9988), 1-hydroxymethyl-N-Desmethyl-IR9792 y ═ 98749400x-583249(0.9992), and 1-carboxy-IR9792 y ═ 92073900x-725988 (0.9981).

As can be seen from Table 4, the standard curve, linearity, detection limit, precision (RSD) and Matrix Effect (ME) of the fluoroandenazole amine and its metabolites in 8 matrices are all better.

The results of five repeated addition recovery tests through linear curve determination and three addition levels show that the concentrations of standard solutions of the fluoroandenazole amine, the 3-hydroxy-IR9792, the Desmethyl-IR9792-N1-Glucoside, the 1-hydroxymethyl-IR9792, the 1-hydroxymethyl-N-Desmethyl-IR9792 and the 1-carboxy-IR9792 are in good linear relation with the instrument response, and the correlation coefficient R is²0.995 to 1.000; the average addition recovery rate (accuracy) and precision in tomato, cucumber, eggplant, pepper, spinach, peach, jujube and orange are respectively 70-111% and 0.2-15%, and the result shows that the residue analysis method meets the requirement of the pesticide residue experimental criteria in NY/T788-one 2018 crops (see Table 5). After the pretreatment conditions, extraction and solid-phase extraction column are optimized, the sample is analyzed, and the fluoroandenazole amine and metabolites thereof are detected. The method fully utilizes the advantages of solid phase extraction, and realizes a simple, safe, effective and reliable analysis method.

TABLE 5 requirement of different addition levels for recovery and relative standard deviation

Concentration of additive (mg/kg)	Recovery (%)	Relative standard deviation (%)
			C>1	70～110	≤10
0.1＜C≤1	70～110	≤15
			0.01＜C≤0.1	70～120	≤20
0.001＜C≤0.01	60～120	≤30
			C≤0.001	50～120	≤35

3. Compared with the prior method

Respectively measuring by using national standard GB/T20769-2008 < liquid chromatogram-tandem mass spectrometry for measuring the residual quantity of 450 pesticides and related chemicals in fruits and vegetables > and GB/T23200.121-2021 < liquid chromatogram-mass spectrometry combined use method for measuring the residual quantity of 331 pesticides and metabolites thereof in plant-derived food >, adding the standard substance into an eggplant or peach homogenate matrix at 0.1mg/kg for 5 times, and processing the sample according to the two selected methods. When the sample is pretreated according to GB/T20769-2008, all results of the eggplant and the peach are not qualified, the standard recovery rate is lower than 10 percent, and the Relative Standard Deviation (RSD) is 0.97-10.29 percent. For GB/T23200.121-2021, the overall analysis was improved, in particular 5 target compounds (72-102%); whereas the recovery of 1-carboxy-IR9792 in eggplant and peach was 25% and 28%, respectively. Therefore, the verification result shows that GB/T20769-2008 and GB/T23200.121-2021 cannot simultaneously analyze the fluoroandenazole amine and 5 metabolites thereof. Besides the adsorption effect, the physical and chemical properties of the compound are also main factors for analysis, so that the acetonitrile can be effectively extracted in the salting-out process, and particularly the compound with better water solubility can be effectively extracted.

In conclusion, the method for detecting the content of the fluoroandenazole amine and the metabolites thereof on the vegetables and fruits optimizes and improves the solid phase extraction process, reduces the use of organic reagents, and simplifies the steps according to the performance of the MCX solid phase extraction column. From the aspects of linearity, detection limit, precision (RSD) and Matrix Effect (ME), the analysis method is acceptable, completely accords with the Chinese pesticide analysis method guide (NY/T788-2018,2018) and the SANTE standard (European Commission, 2019), and has wide application prospect.

The above description is only a preferred embodiment of the present invention, and is not intended to limit the scope of the present invention, and all modifications and equivalents of the present invention, which are made by the contents of the present specification and the accompanying drawings, or directly/indirectly applied to other related technical fields, are included in the scope of the present invention.

Claims

1. A method for detecting the content of the fluoroandenazole bacteria amine and the metabolite thereof on vegetables and fruits is characterized by comprising the following steps:

2. The method for detecting the content of fluoroandenazole amine and metabolites thereof on vegetables and fruits according to claim 1, wherein in step S10, the volume ratio of acetonitrile to water in the aqueous solution of acetonitrile is 2 (3-4).

3. The method for detecting the content of the fluoroandenazole amine and metabolites thereof on the vegetable and fruit of claim 2, wherein the volume of the acetonitrile aqueous solution added per gram of vegetable and fruit is 1-2 mL.

4. The method for detecting the content of fluoroandenazole amine and metabolites thereof on vegetables and fruits of claim 1, wherein after step S20 and before step S30, the method further comprises:

5. The method for detecting the content of fluoroandenazole amine and metabolites thereof on vegetables and fruits of claim 1, wherein in step S30, the solid phase extraction column is an MCX solid phase extraction column; and/or the presence of a gas in the gas,

6. The method for detecting the content of fluoroandenazole amine and metabolites thereof on vegetables and fruits of claim 1, wherein in step S40, the shielding gas comprises nitrogen.

7. The method for detecting the content of the fluoroandenazole amine and the metabolite thereof on the vegetable or fruit according to claim 1, wherein the temperature during the blow-drying in step S40 is 35 to 45 ℃.

8. The method for detecting the content of fluoroandenazole amine and metabolites thereof on vegetables and fruits of claim 1, wherein in step S40, the dissolution solution is filtered through a nylon filter membrane.

9. The method for detecting the content of the fluoroandenazole amine and the metabolite thereof on the vegetable or fruit according to claim 1, wherein step S50 comprises:

10. The method for detecting the content of fluoroandenazole amine and metabolites thereof on vegetables and fruits of claim 1, wherein in step S50,