Method for determining pesticide residues in durian by gas chromatography-mass spectrometry

Abstract

The application discloses a method for determining pesticide residues in durians by gas chromatography-mass spectrometry, and the method comprises the following steps: weighing durian pulp as a sample, adding acetonitrile to extract, adding salt to salting-out, centrifuging to obtain supernatant, adding deionized water to mix, adding into a through type column to perform elution, centrifuging the collected eluent, mixing with anhydrous magnesium sulfate, centrifuging, sucking the supernatant, blowing nitrogen under water bath condition until nearly dry, adding ethyl acetate to redissolve, centrifuging to obtain the supernatant, passing the supernatant through an organic filter membrane to obtain a to-be-tested solution; meanwhile, according to the characteristics of the durian sample, the working parameters of the gas chromatography and the mass spectrometry are set, a mixed standard working solution is configured by using a pesticide mixed standard, a standard curve is drawn, finally, the to-be-tested solution is detected, and the pesticide residue amount in the durian is calculated according to the determination result. The method has the advantages of simple pretreatment, effective removal of oil and pigment in the durian, high efficiency and stability, and provides a reliable detection technical means for pesticide residue detection of the durian.

Description

Technical Field

[0001] This invention belongs to the field of pesticide residue detection technology, specifically relating to a method for determining pesticide residues in durian using gas chromatography-mass spectrometry. Background Technology

[0002] Durian is beloved by many for its rich nutritional value, unique aroma, and health benefits such as aiding digestion and promoting blood circulation. However, as a tropical fruit, durian is only suitable for cultivation in a few coastal areas of China, such as Guangdong and Hainan. Given the high demand, importing from ASEAN countries has become the primary solution. With the opening of ports and the influx of large quantities of goods into the country, strict controls are needed on imported products, especially fruits, to prevent unsafe products from entering the market. Therefore, port sample testing must handle large quantities of durian. As a representative fruit with high sugar and oil content and a complex matrix, there is currently no efficient, rapid, and high-recovery testing method specifically for durian, hindering the monitoring of pesticide residues in durian fruit products.

[0003] Currently, most pesticide residue detection methods employ chromatographic techniques, including high-performance liquid chromatography (HPLC), gas chromatography (GC), gas chromatography-tandem mass spectrometry (GC-MS), HPLC-MS, enzyme-linked immunosorbent assay (ELISA), and colloidal gold rapid test strips. HPLC (suitable for non-volatile, thermally unstable, highly polar, and large-molecule pesticides) and GC (suitable for volatile, thermally stable, and low-polarity pesticides) lack the discriminative power of mass spectrometry. They are generally suitable for samples with clean matrices and relatively low matrix interference, but are prone to interference peaks that affect accurate substance identification. Furthermore, high-throughput detection methods for hundreds of pesticides are lacking. HPLC-MS is suitable for detecting non-volatile or thermally unstable pesticide residues, offering high sensitivity and a wide detection range. However, due to the high cost of the equipment, it is often unsuitable for ordinary laboratories and is not ideal for matrices with high sugar content. Summary of the Invention

[0004] To address the aforementioned shortcomings, this invention discloses a method for determining pesticide residues in durian using gas chromatography-mass spectrometry (GC-MS). This method has the advantages of simple pretreatment, effective removal of oils and pigments from durian, high efficiency and stability, improving the accuracy of durian fruit product testing. At the same time, it can effectively shorten the instrument testing cycle and reduce reagent and consumable consumption, providing a practical and reliable testing technology for port samples.

[0005] This invention is achieved using the following technical solution: A method for determining pesticide residues in durian using gas chromatography-mass spectrometry (GC-MS) includes the following steps: (1) Weigh 10 g of durian pulp as a sample into a 50 mL centrifuge tube, add 10 mL of acetonitrile and shake to extract for 1 min, then add 4 g of anhydrous magnesium sulfate, 1 g of sodium chloride, 1 g of sodium citrate and 0.5 g of disodium hydrogen citrate and shake for 5 min. Then put the centrifuge tube into a high-speed centrifuge and centrifuge for 5 min. Then take 2.7 mL of supernatant into a 15 mL centrifuge tube, add 0.3 mL of deionized water and vortex to mix, then add it into a permeable column and let it drip naturally to elute. Collect the eluent and add 0.2 g of anhydrous magnesium sulfate, vortex to mix, and centrifuge. Then take 2 mL of supernatant and blow it to near dryness under nitrogen in a water bath at 40 °C. Then add 1 mL of ethyl acetate and vortex to redissolve. Then centrifuge and take the supernatant and filter it through an organic filter membrane to obtain the test solution. (2) Setting the working parameters of gas chromatography: The chromatographic column is a 30 m × 0.25 mm and 0.25 μm HP-5MS UI capillary column; the carrier gas is helium; the constant flow mode is adopted and the flow rate is 1.0 mL / min; the injection volume is 1 μL; the injection method is splitless injection and the splitless time is 1.0 min; the injection port temperature is 280 ℃ and the pressure is 5.8 pai; the transfer line temperature is 280 ℃; the column oven temperature program is: the initial temperature is set at 60 ℃ and held for 1 min, then run for 1 min and increase the temperature to 170 ℃ at a program of 40 ℃ / min, then run for 3.75 min, then increase the temperature to 310 ℃ at a program of 10 ℃ / min, and continue to run for 20.75 min; (3) Set the working parameters of the mass spectrometer: the ion source is an electronic ignition (EI) source; the electron energy is 70 eV; the ion source temperature is 230 ℃; the electrode temperature is 150 ℃; the collision gas is argon; the quenching gas is He; the solvent residence time is 2.95~99.13 ms; the solvent delay is 3 min; and the dynamic multiple reaction monitoring (dMRM) mode is used for scanning and quantification. (4) To plot the standard curve, dilute the 100 mg / L pesticide mixed standard stock solution with acetone by 10 times to obtain a mixed standard stock solution with a mass concentration of 10 mg / L, and store it at -20 ℃; dilute the mixed standard stock solution with ethyl acetate to obtain an intermediate solution with a concentration of 1 mg / L, and then use the intermediate solution to prepare mixed standard working solutions with mass concentrations of 5, 10, 20, 50, 100, 150, and 200 ng / mL, and then test them on the instrument and plot the standard curve based on the test results. (5) Take the test liquid obtained in step (1) and test it on the instrument, and calculate the pesticide residue in durian based on the test results.

[0006] Further optimizations are made in step (1), where 10 g of durian pulp is weighed as a sample and the weight is accurate to 0.05 g; the centrifugation speed is 4200 r / min; and the nominal pore size of the organic filter membrane is 0.22 μm.

[0007] Further optimization, in step (1), the permeable column is an Agilent Captiva EMR-LPD permeable column, which is filled with EMR-Lipid / PSA / EC-C18 / Carbon S and has a specification size of 6 mL.

[0008] Further optimization: in step (2), the purity of helium is 99.999%.

[0009] For further optimization, in step (3), the purity of argon gas is not less than 99.999%.

[0010] Further optimization involves step (4) where the pesticide mixture standard is tested to determine the retention time and parent ion corresponding to the pesticide. Then, the parent ion is scanned for product ions to obtain the daughter ion fragments and characteristic fragment ion pairs of the target compound. The pair with the highest response value is selected as the quantitative ion pair, while the other two or three daughter ions are used as qualitative ion pairs. An ion fragment database is then established, and dMRM is used to perform time-segmented detection based on the pesticide's retention time and the fragment ion mass-to-charge ratio (m / z), optimizing and adjusting the residence time. This ensures that the ion peaks completely emerge within the corresponding time, resulting in optimal sensitivity of the quantitative ion pairs and more reliable and accurate detection results.

[0011] Further optimization involves using QQQ quantitative software for data analysis in step (5), and external standard method for quantitative analysis. Six repeated spike experiments are conducted at concentrations of 1, 2, and 10 times the limit of quantitation. This avoids random errors in single-sample testing.

[0012] Compared with existing technologies, this technical solution has the following advantages: This invention provides a high-throughput method for pesticide residue detection in durian products using gas chromatography-tandem mass spectrometry combined with QuEChERS. The method employs a targeted per-pass column for purification in the pretreatment stage, effectively removing pigments and oils from the durian, further extending instrument lifespan, and achieving good recovery rates while reducing reagent and consumable consumption. Combined with suitable extraction reagents, extraction salts, and instrument conditions, the accuracy and efficiency of the detection are improved. This method effectively solves the pain points of the complex durian matrix (high in fat, pigments, and sulfur compounds) and the difficulty of purification, providing practical technical data support for the accurate detection of durian samples and ensuring the safety of products reaching the market. Attached Figure Description

[0013] Figure 1 This is the total ion chromatogram of the mixed standard of 208 pesticides and their metabolites in durian described in Example 1.

[0014] Figure 2 This is a comparison chart of the extraction efficiency of the target compound using different extractants in Experiment Example 1.

[0015] Figure 3 This is a comparison chart of recovery rates under different extractants in Experiment Example 1.

[0016] Figure 4 This is a comparison chart of the effects of each extractant after column chromatography in Experiment Example 1.

[0017] Figure 5 This is a comparison chart of pigment removal by different extractants after nitrogen blowing in Experiment Example 1.

[0018] Figure 6 This is a comparison chart of the extraction efficiency of the target compound in different extraction salts in Experiment Example 2.

[0019] Figure 7 This is a comparison chart of recovery rates under different extraction salts in Experiment Example 2.

[0020] Figure 8 This is a matrix effect diagram of compound 115 in durian from Experiment Example 3.

[0021] Figure 9 This is a comparison chart of nitrogen blowing before different methods in Experiment Example 4.

[0022] Figure 10 This is a comparison chart of nitrogen blowing results using different methods in Experiment Example 4.

[0023] Figure 11 This is a comparison of the liner after approximately 100 injections on the instrument in different methods of Experiment Example 4.

[0024] Figure 12 These are the relevant mass spectrometry parameters and method performance parameters of some pesticide residues in durian in Example 1.

[0025] Figure 13 These are the relevant mass spectrometry parameters and method performance parameters of some pesticide residues in durian in Example 1.

[0026] Figure 14 These are the relevant mass spectrometry parameters and method performance parameters of some pesticide residues in durian in Example 1.

[0027] Figure 15 These are the relevant mass spectrometry parameters and method performance parameters of some pesticide residues in durian in Example 1.

[0028] Figure 16These are the relevant mass spectrometry parameters and method performance parameters of some pesticide residues in durian in Example 1.

[0029] Figure 17 These are the relevant mass spectrometry parameters and method performance parameters of some pesticide residues in durian in Example 1.

[0030] Figure 18 These are the relevant mass spectrometry parameters and method performance parameters of some pesticide residues in durian in Example 1. Detailed Implementation

[0031] The following examples further illustrate the present invention, but are not intended to limit the invention. Specific experimental conditions and methods not specified in the following examples are generally conventional methods well-known to those skilled in the art. For example, vortex mixing can be performed using a VORTEX GENIE2 vortex mixer (Scientific Industries, USA), measurements are performed using an Agilent 8890B-7000D GC-MS / MS gas chromatograph-tandem mass spectrometer (Agilent Technologies, USA), and homogenization is performed using an FA25 homogenizer (FLUKO, Shanghai).

[0032] Example 1: A method for determining pesticide residues in durian using gas chromatography-mass spectrometry (GC-MS), comprising the following steps: (1) Weigh 10 g of durian pulp as a sample (accurate to 0.05 g) into a 50 mL centrifuge tube, add 10 mL of acetonitrile and shake to extract for 1 min, then add 4 g of anhydrous magnesium sulfate, 1 g of sodium chloride, 1 g of sodium citrate and 0.5 g of disodium hydrogen citrate and shake for 5 min. Then, place the centrifuge tube in a high-speed centrifuge and centrifuge at 4200 r / min for 5 min. Then, take 2.7 mL of the supernatant into a 15 mL centrifuge tube, add 0.3 mL of deionized water and vortex to mix, then add it to a permeable column for elution by natural dripping. Collect the eluent and add 0.2 g of anhydrous magnesium sulfate, vortex to mix, and centrifuge at 4200 r / min for 5 min. Then, take 2 mL of the supernatant and blow it to near dryness under nitrogen in a water bath at 40℃, then add 1 mL of ethyl acetate and vortex to reconstitute. Then, centrifuge and take the supernatant through a nominal pore size of 0.22. The test solution was obtained after filtering through an organic filter membrane of μm; the permeable column was an Agilent Captiva EMR-LPD permeable column, which was filled with EMR-Lipid / PSA / EC-C18 / Carbon S and had a size of 6 mL. (2) Setting the working parameters of gas chromatography: The chromatographic column is a 30 m × 0.25 mm and 0.25 μm HP-5MS UI capillary column; the carrier gas is helium with a purity of 99.999%; constant flow mode is adopted, the flow rate is 1.0 mL / min; the injection volume is 1 μL; the injection method is splitless injection, the splitless time is 1.0 min; the injection port temperature is 280 ℃, the pressure is 5.8 pai; the transfer line temperature is 280 ℃; the column oven temperature program is: the initial temperature is set at 60 ℃ and held for 1 min, then run for 1 min and increase the temperature to 170 ℃ at a program of 40 ℃ / min, then run for 3.75 min, then increase the temperature to 310 ℃ at a program of 10 ℃ / min, and continue to run for 20.75 min; (3) Set the working parameters of the mass spectrometer: the ion source is an electronic ignition (EI) source; the electron energy is 70 eV; the ion source temperature is 230 ℃; the electrode temperature is 150 ℃; the collision gas is argon with a purity of not less than 99.999%; the quenching gas is He; the solvent residence time is 2.95~99.13 ms; the solvent delay is 3 min; the dynamic multiple reaction monitoring (dMRM) mode is used for scanning and quantification. (4) To plot the standard curve, dilute the 100 mg / L pesticide mixed standard stock solution with acetone by 10 times to obtain a mixed standard stock solution with a mass concentration of 10 mg / L, and store it at -20 ℃; dilute the mixed standard stock solution with ethyl acetate to obtain an intermediate solution with a concentration of 1 mg / L, and then use the intermediate solution to prepare mixed standard working solutions with mass concentrations of 5, 10, 20, 50, 100, 150, and 200 ng / mL, and then test them on the instrument and plot the standard curve based on the test results. The pesticide mixed standard stock solution is obtained by mixing 208 pesticides and their metabolites mixed standard (Tianjin Alta Technology Co., Ltd.) with acetone. Furthermore, the pesticide mixed standard was tested to determine the retention time and parent ion of the pesticide. Then, the parent ion was scanned for product ions to obtain the daughter ion fragments and characteristic fragment ion pairs of the target compound. The pair with the highest response value was selected as the quantitative ion pair, and the other two or three daughter ions were used as qualitative ion pairs. Then, an ion fragment database was established and dMRM was used to perform time-segmented detection based on the pesticide retention time and fragment ion mass-to-charge ratio (m / z) to optimize and adjust the residence time. In this embodiment, a mass spectrometry database of 208 pesticides and their metabolites was established and analyzed. Ultimately, 115 commonly used pesticide compounds were selected as the target compounds for this study based on their classification. A time-segmented dMRM was used to detect a mixed standard solution of the 208 pesticides and their metabolites. A single point at 200 ng / mL was selected to obtain the total ion chromatogram (see figure). Figure 1 The retention times, ion pair information, collision energies, and other mass spectrometry parameters of the 115 target compounds are shown in [reference needed]. Figures 12-18 ; (5) Take the test liquid obtained in step (1) and test it on the instrument. Calculate the pesticide residue in durian based on the test results. QQQ quantitative software is used for data analysis and external standard method is used for quantitative analysis. Six repeated spiked experiments are carried out at concentrations of 1, 2 and 10 times the limit of quantitation. This can avoid the random error of single sample testing.

[0033] Experimental Example 1: A comparative experiment was conducted using acetone, acetonitrile, and n-hexane as extraction solvents. The specific steps are as follows: First, the spiking level was determined to be 10 times the limit of quantitation (0.005 mg / kg). Then, as described in step (1) of Example 1, 10 g of the blank durian sample (accurate to 0.05 g) was weighed into a 50 mL plastic centrifuge tube. 10 mL of extraction solvent (acetone, acetonitrile, n-hexane) was added and shaken for 1 min. Then, 4 g of anhydrous magnesium sulfate, 1 g of sodium chloride, 1 g of sodium citrate, and 0.5 g of disodium hydrogen citrate were added and shaken for 5 min. The mixture was then placed in a high-speed centrifuge and centrifuged at 4200 r / min for 5 min. 2.7 mL of the supernatant was taken into a 15 mL centrifuge tube, and 0.3 mL of deionized water was added and mixed (finally 3 mL). The mixture was then added to a permeable column and allowed to drip naturally. The eluent was collected in a 15 mL centrifuge tube. After collection, 0.2 g of anhydrous magnesium sulfate (to remove moisture) was added and vortexed. The centrifuge tube was then placed in a high-speed centrifuge and centrifuged at 4200 r / min for 5 min. 2 The supernatant was collected in a 15 mL centrifuge tube and purged with nitrogen in a 40 °C water bath until nearly dry. 1 mL of ethyl acetate solution was added, and the mixture was vortexed to redissolve the supernatant before centrifugation. After completion, the solution was filtered through a 0.22 μm organic filter membrane and bottled for analysis.

[0034] Based on GB / T 27404-2026 "Laboratory Quality Control Standard for Physicochemical Testing of Food", and according to the correct method requirements for different concentrations, the proportions of compounds with spiked recoveries ranging from 60% to 120% in three different extractants were statistically analyzed. Figure 2As shown, different extractants have varying efficiencies in extracting pesticides from durian samples, thus affecting the actual recovery rate. When using hexane as an extractant, slight emulsification occurs after shaking, further impacting extraction efficiency. Due to the miscibility of acetone with water, water and oil are extracted from the durian during centrifugation with shaking; however, a faint oil layer appears after high-speed centrifugation, which is difficult to remove during subsequent nitrogen blowing. Therefore, this study selects a representative compound from different types of pesticides and compares its recovery rate with different extractants. Figure 3 As shown, acetonitrile was ultimately chosen as the extractant, considering both extraction efficiency and stability as well as the difficulty of nitrogen blowing.

[0035] Experiment Example 2: Following GB 23200.113-2026 "Determination of Residues of 242 Pesticides and Their Metabolites in Plant-Derived Foods by Gas Chromatography-Mass Spectrometry," the following salt components were extracted: 4 g anhydrous magnesium sulfate (a desiccant, acting as a salting-out agent to remove moisture), 1 g sodium chloride (a salting-out agent to precipitate moisture and promote sample stratification), 1 g sodium citrate, and 0.5 g disodium hydrogen citrate (a buffer to control pH within a specific range). The focus was on changing the type of salting-out agent while keeping the other two salts constant to screen for the most suitable extraction salt type for this experiment. Therefore, four purified salts were selected: sodium chloride, anhydrous sodium sulfate, potassium chloride, and anhydrous sodium acetate. The specific operation was as follows: Anhydrous magnesium sulfate, sodium citrate, and disodium hydrogen citrate of the same mass as the standard were weighed and placed in a container at 15°C. In a centrifuge tube, keeping the mass of the salting-out agent constant, weigh out four different types of salting-out agents: sodium chloride, anhydrous sodium sulfate, potassium chloride, and anhydrous sodium acetate, and add them to the centrifuge tube. Then, prepare the sample to be tested according to the method described in Experimental Example 1, and calculate the spiked recovery rate of different salting-out agents.

[0036] Based on the standard statistical analysis of the four different extraction salt curves described in Experimental Example 1, the extraction efficiency ranged from 60% to 120%. However, due to the different properties of the extraction salts, the extraction efficiency of pesticides from durian samples may vary slightly. Figure 6As shown, sodium chloride (NaCl) is a strong electrolyte, which increases the ionic strength of the aqueous phase, promotes rapid separation of acetonitrile and the aqueous phase, and inhibits the dissolution of pesticides in the aqueous phase, thus helping to improve the recovery rate. Anhydrous sodium sulfate (Na2SO4) has weak hygroscopicity and is suitable for pesticides sensitive to magnesium ions in most cases; however, its salting-out effect is slightly weaker and the separation speed is slower. Potassium chloride (KCl) has a salting-out strength comparable to NaCl but is more specialized for low-temperature salting-out, avoiding the volatilization / degradation of the target substance at high temperatures. Sodium acetate (CH3COO Na) is a weakly alkaline buffer salt that quickly adjusts the pH to 5.5–6.5, has a mild buffering capacity, does not require the use of acid agents, simplifies the operation, and is suitable for most pesticides that are not sensitive to strong acids or bases; however, its salting-out effect and hygroscopicity are poor, making it suitable for rapid detection. Representative pesticides from different types were selected for comparison of their recovery rates under different extraction salts, such as... Figure 7 As shown, sodium chloride is chosen for salt extraction after comprehensive consideration.

[0037] Experimental Example 3: A matrix-matched standard curve and a solvent external standard curve were prepared, and the slopes of the two curves were compared to investigate the influence of the matrix. In Example 1, step (4) was used to prepare blank tubes of durian sample matrix, and mixed standard stock solutions were prepared by blowing nitrogen to near dryness and then quantitatively preparing them. Matrix-matched standard curves with mass concentrations of 5, 10, 20, 50, 100, 150, and 200 ng / mL were prepared by diluting with ethyl acetate. At the same time, solvent external standard curves with the same concentration gradient were prepared. The matrix effect was evaluated by using ME / %=(BA) / A×100% (A and B are the slopes of the linear equations of the solvent and matrix standard curves, respectively). |ME|<10% indicates a weak matrix effect, and no measures are needed. The solvent external standard curve can be used for quantification. 10%≤|ME|≤50% indicates a moderate matrix effect, which requires correction by using a matrix-matched standard curve or adding an internal standard to eliminate the influence of the matrix effect. ME|>50% indicates a strong matrix effect, which requires optimization of the pretreatment method.

[0038] Based on the matrix effect analysis results, a plot of ME (Y) was obtained using the serial numbers (X) of 115 compounds. Figure 8As shown in the figure, NO.38, NO.48, and NO.75 (flufenoxam, phorate, and pyrimethanil) exhibited matrix inhibition effects (-7.96, -27.26, and -15.53 respectively), but |ME| < 20%. The remaining target compounds showed matrix enhancement effects, with |ME| < 50%. This indicates that all 115 compounds in this method can be quantified using matrix-matched standard curves to eliminate the influence of ME. ME is a common interference phenomenon in food detection, referring to matrix components in the sample that, besides the target compounds, can affect the detection results. Durian is a fruit high in sugar and oil with a complex matrix composition. Furthermore, slight pigment residues may remain during extraction; if not properly purified during pretreatment, this can affect the qualitative and quantitative results.

[0039] Experimental Example 4: Based on the method described in this invention and the method in GB 23200.113-2026 "Determination of Residues of 242 Pesticides and Their Metabolites in Plant-Derived Foods by Gas Chromatography-Mass Spectrometry", a comparative experiment was conducted on the purification effect and recovery rate of durian samples. The specific procedure is as follows: First, the same spiking levels were applied at 1, 2, and 10 times the limit of quantitation (i.e., 0.005, 0.01, and 0.05) according to the methods described in GB 23200.113-2026 standard method experiment and Example 1. (mg / kg), with 6 parallel tests performed for each spiking level. The recovery rates of the control method at low, medium, and high concentrations ranged from 60.21% to 99.95%, 60.83% to 105.77%, and 60.46% to 106.86%, respectively, with average recoveries of 70.38%, 78.71%, and 75.59%, respectively. The method described in Example 1 had recovery rates of 79.25% to 119.95%, 74.45% to 14.64%, and 70.66% to 102.61% at low, medium, and high concentrations, respectively, with average recoveries of 104.82%, 93.31%, and 89.61%, respectively, as shown in Table 1. The comparison shows that the method of the present invention has a better recovery rate.

[0040] In addition, comparative images of centrifuge tubes after purification and after nitrogen blowing were collected from the two experiments, such as... Figure 9 and Figure 10 As shown in the figures, the left side of both images represents the method of this invention. The state before and after nitrogen blowing is cleaner and clearer than the right side, indicating that the method of this invention is more effective than the standard method GB 23200.113-2026 in removing pigments and grease. Simultaneously, the liner tubes were collected after approximately 100 injections into the instrument for both experiments. This comparison shows that, with the same number of samples injected, the degree of liner contamination was significantly improved in both experiments. Figure 11As shown, the liner after sample injection using the method of the present invention is significantly cleaner than that after sample injection using the standard method GB23200.113-2026. This indicates that the purification effect of the method of the present invention is comparable to that of the standard method GB 23200.113-2026, representing a breakthrough in reducing reagent and consumable consumption, extending instrument maintenance cycles, and improving instrument lifespan.

[0041] Table 1. Recovery rate range of the method of this invention and the standard method GB 23200.113-2026 Example 5: Quantitative analysis was performed using matrix-matched standard curves (5, 10, 20, 50, 100, 150, 200 ng / mL); the minimum spiked concentration that satisfies the signal-to-noise ratio ≥10 was defined as the limit of quantitation for this method. Referring to GB / T 27404-2026 "Laboratory Quality Control Standard for Food Physicochemical Testing", three levels of concentration (low, medium, and high) were selected for spiking tests. The spiking levels were set at 1, 2, and 10 times the limit of quantitation (i.e., 0.005, 0.01, and 0.05 mg / kg). Six parallel tests were performed for each spiking level. The remaining steps were performed using the pretreatment method described in step (4) of Example 1. The average spiked recovery rate of the sample was calculated using the formula: (average sample concentration / spiking concentration) × 100%. The method precision was calculated using the results of six parallel experiments: (standard deviation / average sample concentration) × 100%.

[0042] The experimental results were plotted with the peak area of ​​the target analyte on the ordinate (Y) and the mass concentration on the abscissa (X, ng / mL). A standard curve was then drawn to obtain the linear correlation coefficient (r). 2 The average spiked recovery rate and relative standard deviation of the samples were calculated using the spiked experimental data, and the results are shown in Table 2. All target compounds exhibited good linearity within the linear range of 10–200 ng / mL. 2 Between 0.9951 and 0.9999, the spiked recoveries ranged from 65.86% to 113.0%, with relative standard deviations of 0.58% to 12.74%. These results indicate that the accuracy and precision of this method meet the requirements of the national standard GB / T 27404-2026 "Laboratory Quality Control Standard for Physicochemical Testing of Food," demonstrating good reproducibility and stability. The limit of quantitation (LOQ) of the method was investigated using a blank matrix spiked method. In the spiked recovery experiment, the minimum spiked concentration that achieved a signal-to-noise ratio (S / N) ≥ 10 was defined as the LQ of this method. Data from the spiked experiments showed that the S / N for all target compounds at a spiked level of 0.005 mg / kg was greater than 10. Therefore, the LQ for all 115 target compounds in this method is 0.005 mg / kg.

[0043] Furthermore, it should be understood that although this specification describes embodiments, not every embodiment contains only one independent technical solution. This narrative style is merely for clarity. Those skilled in the art should consider the specification as a whole, and the technical solutions in each embodiment can also be appropriately combined to form other embodiments that can be understood by those skilled in the art.

Claims

1. A method for determining pesticide residues in durian using gas chromatography-mass spectrometry (GC-MS), characterized in that: Includes the following steps: (1) Weigh 10 g of durian pulp as a sample into a 50 mL centrifuge tube, add 10 mL of acetonitrile and shake to extract for 1 min, then add 4 g of anhydrous magnesium sulfate, 1 g of sodium chloride, 1 g of sodium citrate and 0.5 g of disodium hydrogen citrate and shake for 5 min. Then put the centrifuge tube into a high-speed centrifuge and centrifuge for 5 min. Then take 2.7 mL of supernatant into a 15 mL centrifuge tube, add 0.3 mL of deionized water and vortex to mix, then add it into a permeable column and let it drip naturally to elute. Collect the eluent and add 0.2 g of anhydrous magnesium sulfate, vortex to mix, and centrifuge. Then take 2 mL of supernatant and blow it to near dryness under nitrogen in a water bath at 40 °C. Then add 1 mL of ethyl acetate and vortex to redissolve. Then centrifuge and take the supernatant and filter it through an organic filter membrane to obtain the test solution. (2) Setting the working parameters of gas chromatography: The chromatographic column is a 30 m × 0.25 mm and 0.25 μm HP-5MS UI capillary column; the carrier gas is helium; the constant flow mode is adopted and the flow rate is 1.0 mL / min; the injection volume is 1 μL; the injection method is splitless injection and the splitless time is 1.0 min; the injection port temperature is 280 ℃ and the pressure is 5.8 pai; the transfer line temperature is 280 ℃; the column oven temperature program is: the initial temperature is set at 60 ℃ and held for 1 min, then run for 1 min and increase the temperature to 170 ℃ at a program of 40 ℃ / min, then run for 3.75 min, then increase the temperature to 310 ℃ at a program of 10 ℃ / min, and continue to run for 20.75 min; (3) Set the working parameters of the mass spectrometer: the ion source is an electron impact ion source; the electron energy is 70 eV; the ion source temperature is 230 ℃; the electrode temperature is 150 ℃; the collision gas is argon; the quenching gas is He; the solvent residence time is 2.95~99.13 ms; the solvent delay is 3 min; and the dynamic multiple reaction monitoring mode is used for scanning and quantification. (4) To plot the standard curve, dilute the 100 mg / L pesticide mixed standard stock solution with acetone by 10 times to obtain a mixed standard stock solution with a mass concentration of 10 mg / L, and store it at -20℃; dilute the mixed standard stock solution with ethyl acetate to obtain an intermediate solution with a concentration of 1 mg / L, and then use the intermediate solution to prepare mixed standard working solutions with mass concentrations of 5, 10, 20, 50, 100, 150, and 200 ng / mL, and then test them on the instrument and plot the standard curve based on the test results. (5) Take the test liquid obtained in step (1) and test it on the instrument, and calculate the pesticide residue in durian based on the test results.

2. The method for determining pesticide residues in durian using gas chromatography-mass spectrometry according to claim 1, characterized in that: In step (1), 10 g of durian pulp was weighed as a sample, accurate to 0.05 g; the centrifugation speed was 4200 r / min; and the nominal pore size of the organic filter membrane was 0.22 μm.

3. The method for determining pesticide residues in durian using gas chromatography-mass spectrometry according to claim 1, characterized in that: In step (1), the permeable column is an Agilent Captiva EMR-LPD permeable column, which is filled with EMR-Lipid / PSA / EC-C18 / Carbon S and has a specification size of 6 mL.

4. The method for determining pesticide residues in durian using gas chromatography-mass spectrometry according to claim 1, characterized in that: In step (2), the purity of helium is 99.999%.

5. The method for determining pesticide residues in durian using gas chromatography-mass spectrometry according to claim 1, characterized in that: In step (3), the purity of argon gas is not less than 99.999%.

6. The method for determining pesticide residues in durian using gas chromatography-mass spectrometry according to claim 1, characterized in that: In step (4), the pesticide mixed standard is tested to determine the retention time and parent ion corresponding to the pesticide. Then, the parent ion is scanned for product ions to obtain the daughter ion fragments and characteristic fragment ion pairs of the target compound. The pair with the highest response value is selected as the quantitative ion pair, and the other two or three daughter ions are used as qualitative ion pairs. Then, an ion fragment database is established and dMRM is used to perform time-segmented detection based on the pesticide retention time and fragment ion mass-to-charge ratio, and the residence time is optimized and adjusted.

7. The method for determining pesticide residues in durian using gas chromatography-mass spectrometry according to claim 1, characterized in that: In step (5), QQQ quantitative software was used for data analysis, and the external standard method was used for quantitative analysis. Six repeated spiked experiments were conducted at concentrations of 1, 2, and 10 times the limit of quantitation.

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