Deuterated zearalenone derivative compound, preparation method of deuterated zearalenone derivative compound, detection reagent and method for measuring content of zearalenone

Abstract

The invention relates to the technical field of compound synthesis, in particular to a deuterated zearalenone derivative compound, a preparation method of the deuterated zearalenone derivative compound, a detection reagent and a method for measuring the content of zearalenone. The deuterated zearalenone derivative compound is selected from compounds shown in the following structural formula: # imgabs0 #, wherein R is selected from any one of functional groups formed by C1-C5 alkyl, C1-C5 alkyl substituted phenyl and C1-C5 alkyl substituted biphenyl; n is any numerical value between 1 and 7. The method can be used for detecting the content of zearalenone, and overcomes the defects of serious matrix interference, influence on the accuracy of a measurement result and low detection efficiency caused by isotope zearalenone in the prior art.

Description

Technical Field

[0001] The present invention relates to the technical field of compound synthesis, and in particular to a deuterated zearalenone derivative compound and a preparation method thereof, a detection reagent and a method for determining the content of zearalenone. Background Art

[0002] Mycotoxins are toxic secondary metabolites produced by fungi on crops or other food sources, which can contaminate food and feed products and pose a major threat to human and animal health. Zearalenone (ZEN) is a naturally occurring non-steroidal estrogenic fungal toxin that is commonly found in crops such as corn, wheat, barley and sorghum and their products, and easily accumulates in foods such as cereals, meat, milk and eggs. ZEN can also accumulate in water bodies and soil, polluting the ecological environment and causing health problems for animals and humans. At the same time, some studies have shown that ZEN has neurotoxicity, immunotoxicity, carcinogenicity, hepatotoxicity, cytotoxicity and reproductive toxicity. Therefore, it is necessary to monitor the contamination of zearalenone, including qualitative or quantitative analysis and testing.

[0003] At present, the detection method of zearalenone is generally carried out by the derivative method, but this detection method is inaccurate. Therefore, its isotope is used for detection. However, the stable isotope internal standard of zearalenone on the market is mainly full carbon labeled zearalenone- 13 C 18 , which is difficult to obtain by synthesis and is generally obtained by biosynthesis, resulting in high synthesis costs. At the same time, compared with unlabeled zearalenone, zearalenone with all 18 carbons labeled by 13C has a mass number of 18 more, which may cause mass discrimination effect in mass spectrometry testing. Therefore, it is necessary to provide a new detection reagent that can effectively reduce the matrix interference effect and provide accurate detection results.

[0004] In view of this, the present invention is proposed. Summary of the invention

[0005] The purpose of the present invention is to provide a deuterated zearalenone derivative compound and a preparation method thereof, a detection reagent and a method for determining the content of zearalenone. The embodiment of the present invention provides a new deuterated zearalenone derivative compound that overcomes the defects of the prior art that the isotope zearalenone causes serious matrix interference, affects the accuracy of the measurement results, and has low detection efficiency.

[0006] The present invention is achieved in that:

[0007] In a first aspect, the present invention provides a deuterated zearalenone derivative compound, which is used to detect the content of zearalenone, and is selected from the compounds represented by the following structural formula: Wherein, R is selected from any one of the functional groups formed by C1-C5 alkyl, C1-C5 alkyl substituted phenyl and C1-C5 alkyl substituted biphenyl; and n is any value between 1-7.

[0008] In a second aspect, the present invention provides a method for preparing a deuterated zearalenone derivative compound according to the aforementioned embodiment, comprising: mixing deuterated zearalenone with a raw material containing an R group for reaction, wherein the structural formula of the raw material containing an R group is as follows: The structural formula of deuterated zearalenone is as follows:

[0009]

[0010] In a third aspect, the present invention provides a detection reagent comprising the deuterated zearalenone derivative compound described in the aforementioned embodiment.

[0011] In a fourth aspect, the present invention provides a method for determining the content of zearalenone, comprising: pre-treating a sample to be tested containing zearalenone and mixing it with a derivatization reagent to perform derivatization to form a derivatized sample to be tested containing a zearalenone derivative;

[0012] Mixing the derivative sample to be tested with the deuterated zearalenone derivative compound described in the aforementioned embodiment or the detection reagent for detecting the content of zearalenone described in the aforementioned embodiment;

[0013] Then, the mass spectrum is detected, and the content of the zearalenone derivative is determined according to the peak area of ​​the mass spectrum result;

[0014] Wherein, the added amount of the deuterated zearalenone derivative compound is not less than 100 μg / L;

[0015] Preferably, the added amount of the deuterated zearalenone derivative compound is 200 μg / L-1000 μg / L.

[0016] The present invention has the following beneficial effects: The embodiment of the present invention provides a new deuterated zearalenone derivative compound, which can be used to detect the content of zearalenone in complex samples, and can effectively eliminate matrix interference during the detection process, thereby improving the accuracy of the measurement results and the detection efficiency. At the same time, the synthesis method of the deuterated zearalenone derivative compound does not require the use of expensive transition metal catalysts, has mild reaction conditions, simple synthesis process, good selectivity, green and environmental protection, low cost, high conversion rate, high product purity, etc., and is suitable for large-scale production. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings required for use in the embodiments are briefly introduced below. It should be understood that the following drawings only show certain embodiments of the present invention and therefore should not be regarded as limiting the scope. For ordinary technicians in this field, other related drawings can be obtained based on these drawings without creative work.

[0018] Figure 1 NMR spectra provided for characterization of embodiments of the present invention;

[0019] Figure 2 High-resolution time-of-flight mass spectrometry (Q-TOF) provided for characterization of embodiments of the present invention;

[0020] Figure 3 High-liquid spectra provided for characterization of embodiments of the present invention. DETAILED DESCRIPTION

[0021] In order to make the purpose, technical scheme and advantages of the embodiments of the present invention clearer, the technical scheme in the embodiments of the present invention will be described clearly and completely below. If the specific conditions are not specified in the embodiments, they are carried out according to conventional conditions or conditions recommended by the manufacturer. If the manufacturer of the reagents or instruments used is not specified, they are all conventional products that can be purchased commercially.

[0022] In a first aspect, an embodiment of the present invention provides a deuterated zearalenone derivative compound, which is used to detect the content of zearalenone, and is selected from the compounds represented by the following structural formula:

[0023] Wherein, R is selected from any one of the functional groups formed by C1-C5 alkyl, C1-C5 alkyl substituted phenyl and C1-C5 alkyl substituted biphenyl; and n is any value between 1-7.

[0024] For example, R can be a C1-C5 alkyl-substituted phenyl group, preferably a C1-C5 alkyl-multiple-substituted phenyl group; more preferably a C1-C5 alkyl-disubstituted phenyl group or a C1-C5 alkyl-trisubstituted phenyl group.

[0025] For example, it is selected from the compounds represented by the following structural formula:

[0026] R 1 and R 2 They are independently selected from C1-C5 alkyl, and n is any value between 1-7.

[0027] The above-mentioned alkyl group refers to C1-C5 alkyl groups such as methyl, ethyl, propyl, butyl, etc., and n is any value between 1 and 7 such as 1, 2, 3, 4, 5, 6 and 7.

[0028] Specifically, the deuterated zearalenone derivative compound is selected from any one of the compounds represented by the following structural formulas:

[0029]

[0030] Herein, n is any value between 1-7, such as 1, 2, 3, 4, 5, 6 and 7.

[0031] In a second aspect, the present invention provides a method for preparing the deuterated zearalenone derivative compound described in the aforementioned embodiment, comprising:

[0032] S1, synthesis of deuterated zearalenone;

[0033] Zearalenone, heavy water and a strong base and a weak acid salt are mixed and reacted to form deuterated zearalenone. The structural formula of the deuterated zearalenone is shown below:

[0034]

[0035] Specifically, during the reaction, the strong base weak acid salt is selected from any one or more of potassium carbonate, sodium carbonate and dipotassium hydrogen phosphate; preferably potassium carbonate; the mass ratio of zearalenone to the strong base weak acid salt is 1: (5-10), for example, 1: 5, 1: 6, 1: 7, 1: 8, 1: 9 and 1: 10, etc., any value between 1: (5-10). The mass ratio of zearalenone to the heavy water is 1- (20: 100); for example, 1: 20, 1: 30, 1: 40, 1: 50, 1: 60, 1: 70, 1: 80, 1: 90 and 1: 100, etc., any value between 1- (20: 100). The temperature is room temperature, for example, 10-35 ° C, more preferably 20-30 ° C. The organic solvent used in the reaction process includes, but is not limited to, any one or more of tetrahydrofuran, N, N-dimethylformamide, and dimethyl sulfoxide, preferably tetrahydrofuran.

[0036] The temperature and base provided in the embodiment of the present invention can not only improve the purity of the product deuterated zearalenone, but also improve its yield.

[0037] After the reaction is completed, post-treatment is performed. For example, after the reaction is completed, the reaction system is poured into a beaker, a certain volume of sulfuric acid solution (for example, 0.2M) is added and stirred thoroughly, and then transferred to a separatory funnel, an extractant is added for extraction, and the organic phase is separated and collected, dried, filtered and concentrated to obtain the deuterated gearalenone product.

[0038] The volume ratio of the reaction solution to the sulfuric acid solution is 1:30-1:60. The base used in the reaction system is neutralized with sulfuric acid to terminate the reaction, thereby increasing the yield of the reaction product and maintaining the stability of deuterated zearalenone under acidic conditions.

[0039] The extractant is one or more of dichloromethane, n-hexane and ethyl acetate. Preferably, the extractant is dichloromethane.

[0040] S2, synthesis of deuterated zearalenone derivative compounds;

[0041] Deuterated zearalenone is mixed with a raw material containing an R group to react, wherein the structural formula of the raw material containing an R group is as follows: Specifically, the deuterated zearalenone, the R-group-containing raw material and the reducing agent are mixed to react.

[0042] The molar ratio of deuterated zearalenone to the R-containing raw material is 1:(1-5); for example, any value between 1:1, 1:2, 1:3, 1:4 and 1:5, etc. The molar ratio of the reducing agent to the deuterated zearalenone is 1:(5-1); for example, any value between 1:1:2, 1:3, 1:4 and 1:5, etc. The reaction temperature is 10-50°C, and the reaction time is 10-20h.

[0043] If the reaction temperature is too high or too low, it may lead to excessive energy consumption or reduce the yield or purity.

[0044] The reducing agent is selected from sodium cyanoborohydride, sodium triacetoxyborohydride, and lithium aluminum tetrahydride. Preferably, the reducing agent is sodium cyanoborohydride.

[0045] Further, after the reaction is completed, the organic phase is evaporated under reduced pressure to obtain a crude product of the derivatizing agent, the crude product is chromatographed on a silica gel column, gradient eluted with an eluent, the target product components are collected in a glass test tube, and the solvent is removed by vacuum filtration to obtain the target product. The eluent is a mixed solution of ethyl acetate and petroleum ether in a volume ratio of 1:3-5.

[0046] In summary, the embodiment of the present invention uses the fungal toxin gerarenone as a raw material, and realizes the efficient preparation of the deuterated gerarenone derivative agent by a simple reaction process of hydrogen-deuterium exchange and dehydration condensation reaction under alkaline conditions in a room temperature water bath, and after column chromatography separation, the product purity can reach 99%. The synthesis method provided by the present invention does not require the use of expensive transition metal catalysts, has mild reaction conditions, simple synthesis process, good selectivity, green environmental protection and low cost, high conversion rate, high product purity and other advantages, is suitable for large-scale production, can realize accurate quantification of gerarenone in different matrices, and has broad application prospects.

[0047] In a third aspect, the present invention provides a detection reagent, which includes the deuterated zearalenone derivative compound described in the above embodiment. The detection reagent may also include other auxiliary materials or solvents.

[0048] In a fourth aspect, the present invention provides a method for determining the content of zearalenone, comprising: pre-treating a sample containing zearalenone and mixing it with a derivatization reagent for derivatization, wherein the derivatization reagent can be For example, 2-[(3,4-dimethylphenyl)amino]acetohydrazide is selected, and the conditions for the derivatization can refer to the existing reaction, and the present embodiment will not be described in detail. The selection of n and R is also the same as described above.

[0049] It should be noted that the sample containing zearalenone to be tested may be a pre-treated sample, and the pre-treatment steps include: adding an organic solvent such as methanol for shaking extraction, centrifuging to take the supernatant, adding n-hexane to further remove impurities such as protein, and centrifuging for purification; transferring the above solution to a medium containing C 18 , PSA and anhydrous magnesium sulfate in a centrifuge tube, shake to purify, and centrifuge to obtain the supernatant.

[0050] Next, a derivatizing agent and sodium cyanoborocyanide are added for derivatization.

[0051] Then, the derivative sample to be tested is mixed with the deuterated zearalenone derivative compound or the detection reagent described in the above embodiment, and then the mass spectrum is detected, and the content of the zearalenone derivative is determined according to the peak area of ​​the mass spectrum result. At this time, the amount of the deuterated zearalenone derivative compound added is known and clear, so the content of the derivative compound derived from zearalenone can be determined according to the ratio of the peak area of ​​the deuterated zearalenone derivative compound to the peak area of ​​the derivative compound derived from zearalenone, and then the content of zearalenone can be inferred.

[0052] Wherein, the added amount of the deuterated zearalenone derivative compound is not less than 100 μg / L; the added amount of the deuterated zearalenone derivative compound is 200 μg / L-1000 μg / L.

[0053] It should be noted that the derivatizing agent for derivatizing zearalenone is determined based on the R-group-containing raw material of deuterated zearalenone.

[0054] The features and performance of the present invention are further described in detail below in conjunction with the embodiments.

[0055] Example 1

[0056] The present invention provides a structural formula of a deuterated zearalenone derivative compound:

[0057]

[0058] The embodiment of the present invention provides a method for preparing a deuterated zearalenone derivative compound, comprising:

[0059] S1, synthesis of deuterated zearalenone;

[0060] The synthesis was carried out according to the following synthesis route:

[0061] Specifically, 500 mg of zearalenone, 30 mL of heavy water, 6.2 mL of THF and 3.9 g of potassium carbonate were added to a 50 mL round-bottom flask in sequence, stirred at room temperature (25 ° C), and reacted overnight. After the reaction was completed, the reaction system was poured into a beaker, 1800 mL of 0.2 M sulfuric acid was added and stirred, the solution was poured into a separatory funnel, extracted three times with 600 mL of dichloromethane, and the organic phase was collected. The organic phase was dried with anhydrous magnesium sulfate, filtered, and dichloromethane was removed under reduced pressure to obtain 456 mg of white crystalline deuterated zearalenone, with a yield of 90% and a purity of 99%.

[0062] S2, synthesis of deuterated zearalenone derivative compounds;

[0063] The synthesis was carried out according to the following synthesis route:

[0064]

[0065] Specifically, the above 500 mg of deuterated zearalenone and 305 mg of 2-[(3,4-dimethylphenyl)amino]acetylhydrazine were dissolved in 10 mL of methanol (pH adjusted by 0.1 M hydrochloric acid standard solution) at pH = 6.5, 97 mg of sodium cyanoborohydride was added, and the reaction was carried out at room temperature (25°C) overnight. After the reaction, the methanol was evaporated. The crude product was chromatographed on a 100 mesh silica gel column, wet column loaded, wet sample loaded, and purified by gradient elution with ethyl acetate / petroleum ether (1 / 5-1 / 3), and the target product components were collected. After the solvent was removed under reduced pressure, 462 mg of a white deuterated zearalenone derivative compound was obtained, with a yield of 60% and a purity of 99%.

[0066] Characterization

[0067] The zearalenone, deuterated zearalenone and deuterated zearalenone derivatives of Example 1 were characterized by nuclear magnetic resonance hydrogen spectrum, high-resolution time-of-flight mass spectrometry and high-performance liquid chromatography mass spectrometry. The results are shown in Figure 1-Figure 3 .

[0068] The reaction product was subjected to ion scanning analysis by liquid chromatography-mass spectrometry, and the determination steps were as follows: 1 mg of zearalenone, deuterated zearalenone of Example 1, and deuterated zearalenone derivative were weighed and dissolved in 1 ml of chromatography-grade methanol to prepare 1000 mg / L zearalenone and deuterated zearalenone stock solutions. Methanol was used to dilute the working solution to 1 mg / L for liquid chromatography-mass spectrometry analysis.

[0069] in, Figure 1 The H-NMR spectra of zearalenone and deuterated zearalenone are shown. Figure 2 A high-resolution time-of-flight mass spectrum (Q-TOF) of deuterated zearalenone is shown; Figure 3 Represents the HPLC spectrum of deuterated zearalenone derivative compounds.

[0070] according to Figure 1-Figure 3 It can be seen that after the deuteration reaction, the hydrogen ion content of deuterated zearalenone at the chemical shift value of 6.41 is significantly reduced, and its position is replaced by a deuterium atom, and hydrogen-deuterium exchange occurs. Figure 1 As shown, the main chemical shift peaks are assigned as follows: the doublet of 1.38 is generated by hydrogen at 11', the multiplet of 1.49 is generated by hydrogen at 4', the multiplet of 1.65 is generated by hydrogen at 9', the multiplet of 1.80 is generated by hydrogen at 8', the multiplets of 2.05-2.28 are generated by hydrogen at 3', 4', 5' and 7', the multiplet of 2.40 is generated by hydrogen at 3', the broad doublet of 2.61 is generated by hydrogen at 7', the broad doublet of 2.86 is generated by hydrogen at 5', the multiplet of 5.0 is generated by hydrogen at 10', the multiplet of 5.68 is generated by hydrogen at 2', the singlet of 6.35 is generated by hydrogen at 3, and the two doublets of 7.02 are generated by hydrogen at 1'. The above results show that the chemical shift values ​​correspond to zearalenone and its deuterated products, proving that deuterated zearalenone was successfully synthesized.

[0071] according to Figure 2 It can be seen that due to the deuteration reaction, the molecular ion peak of the compound changes in the negative ion mode, among which the molecular weight of deuterated zearalenone is 321.1623, which is basically consistent with the theoretical predicted value of 321.1691, indicating that the conversion of zearalenone to deuterated zearalenone is achieved through the deuteration reaction.

[0072] according to Figure 3 It can be seen that the deuterated zearalenone derivative compound is in positive ion mode. At the same time, the theoretical molecular mass of the derivative product obtained by the reaction of deuterated zearalenone with 2-[(3,4-dimethylphenyl)amino]acetylhydrazine is 498.1, and its molecular mass in positive ion mode is 499.1, which is consistent with the actual measured liquid chromatography mass spectrometry. Figure 1The results show that the desired deuterated zearalenone derivative compound can be synthesized by the present invention.

[0073] Application Examples

[0074] The deuterated zearalenone derivative compound provided in Example 1 of the present invention is used for detection, and the specific process is as follows:

[0075] Take 2mL of blank milk sample and place it in a 50mL centrifuge tube, add 120uL ZON (zearalenone) (10ppm), shake and mix thoroughly, then add 10mL of methanol with pH=6.5, shake for 10min, centrifuge at 12000rpm / min for 10min, and take the supernatant in a 50mL centrifuge tube. Add 10mL of n-hexane, shake for 10min, centrifuge at 12000rpm / min for 10min, discard the upper n-hexane, and wait for purification. Transfer the above solution to a centrifuge tube containing 80mg C18, 60mg PSA and 80mg anhydrous magnesium sulfate, shake for 10min, centrifuge at 12000rpm / min for 10min, and take the supernatant. The supernatant was added with 1 mL of a derivatizing agent (5 ppm), 120 uL of the deuterated zearalenone derivative compound of Example 1 (10 ppm) and 10 mg of sodium cyanoborohydride, derivatized at 40° C. for 15 hours, dried with nitrogen at 40° C., re-dissolved in 1 mL of the initial mobile phase ratio, and passed through a membrane for determination. The recovery rate of zearalenone in the milk sample was measured to be 98%.

[0076] The national standard method is used for testing, and the specific process is as follows:

[0077] Take 2mL of blank milk sample and place it in a 50mL centrifuge tube, add 120uL ZON (10ppm), shake and mix thoroughly, add 15mL of anhydrous ether, shake and extract for 5min, centrifuge at 4000r / min for 2min, transfer the supernatant to a concentration bottle, and rotate and concentrate to near dryness below 40℃. Add 1mL of chloroform to dissolve the residue, ultrasonically dissolve for 2min, transfer to a 10mL centrifuge tube, rinse the concentration bottle with 3mL of 0.5mol / L sodium hydroxide solution, transfer to the same centrifuge tube, vortex and mix, centrifuge at 4000r / min for 2min, and absorb the upper sodium hydroxide solution. Add 1mL of phosphoric acid-water solution, mix well and wait for purification. Elute with 5mL of water and 5mL of methanol-water solution, discard the effluent; then elute with 10mL of methanol and collect the eluate. Blow dry the eluate with nitrogen below 40℃. The residue was dissolved in 1.0 mL of acetonitrile, vortexed and filtered through a 0.2 μm microporous filter membrane. Standard curves of different matrix concentrations were prepared and quantified by the external standard method. The recovery rate of zearalenone in the milk sample was measured to be 92%.

[0078] The deuterated zearalenone provided in Example 1 of the present invention is used for detection, and the specific process is as follows:

[0079] Take 2mL blank milk sample and place it in a 50mL centrifuge tube, add 10mL methanol with pH=6.5, add 120uL ZON (10ppm), 120uL deuterated corn zearalenone (10ppm) of Example 1, shake for 10min, centrifuge at 12000rpm / min for 10min, and take the supernatant in a 50mL centrifuge tube. Add 10mL of n-hexane, shake for 10min, centrifuge at 12000rpm / min for 10min, discard the upper n-hexane, and wait for purification. Transfer the above solution to a centrifuge tube containing 80mg C18, 60mg PSA and 80mg anhydrous magnesium sulfate, shake for 10min, centrifuge at 12000rpm / min for 10min, and take the supernatant. Dry with nitrogen at 40℃, redissolve in 1mL of the initial mobile phase ratio, pass the membrane for determination, and the recovery rate of zearalenone in the milk sample is 89%.

[0080] The above recovery rate is calculated by the formula of the actual detected zearalenone content / the actual sample added zearalenone content. The recovery rate indicates the accuracy of the corresponding detection method in the actual sample detection. According to the above results, the use of synthetic deuterated zearalenone derivatives as internal standards can effectively reduce the influence of matrix interference and improve the accuracy of actual sample detection, which provides a new idea for the detection of zearalenone in actual samples.

[0081] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. For those skilled in the art, the present invention may have various modifications and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A deuterated zearalenone derivative compound, characterized in that: It is used to detect the content of zearalenone, which is selected from the compounds shown in the following structural formula: Wherein, R is selected from any one of the functional groups formed by C1-C5 alkyl, C1-C5 alkyl substituted phenyl and C1-C5 alkyl substituted biphenyl; and n is any value between 1-7.

2. The deuterated zearalenone derivative compound according to claim 1, characterized in that: R is selected from C1-C5 alkyl substituted phenyl, preferably C1-C5 alkyl multi-substituted phenyl; more preferably C1-C5 alkyl di-substituted phenyl or C1-C5 alkyl tri-substituted phenyl.

3. The deuterated zearalenone derivative compound according to claim 1 or 2, characterized in that: It is selected from the compounds represented by the following structural formula: R1 and R2 are independently selected from C1-C5 alkyl, and n is any value between 1 and 7; Preferably, R1 and R2 are independently selected from C1-C3 alkyl; More preferably, n is any value between 1-5, and more preferably 1-3.

4. The deuterated zearalenone derivative compound according to claim 1 or 2, characterized in that: It is selected from any one of the compounds represented by the following structural formulas: Wherein, n is any value between 1 and 7.

5. A method for preparing the deuterated zearalenone derivative compound according to claim 1, characterized in that: include: Deuterated zearalenone is mixed with a raw material containing an R group to react, wherein the structural formula of the raw material containing an R group is as follows: The structural formula of deuterated zearalenone is as follows:

6. The preparation method according to claim 5, characterized in that: include: The deuterated zearalenone, the R-group-containing raw material and the reducing agent are mixed to react.

7. The preparation method according to claim 6, characterized in that: The molar ratio of the deuterated zearalenone to the R-group-containing raw material is 1:(1-5); Preferably, the molar ratio of the reducing agent to the deuterated zearalenone is 1:(5-1); Preferably, the reaction temperature is 10-50°C and the reaction time is 10-20h; Preferably, the reducing agent is selected from any one of sodium cyanoborohydride, sodium triacetoxyborohydride and lithium aluminum tetrahydride. Preferably, the reducing agent is sodium cyanoborohydride.

8. The preparation method according to claim 5, characterized in that: The preparation method of the deuterated zearalenone is as follows: zearalenone, heavy water and a strong base and a weak acid salt are mixed and reacted; Preferably, the strong base and weak acid salt is selected from any one or more of potassium carbonate, sodium carbonate and dipotassium hydrogen phosphate; preferably potassium carbonate; Preferably, the reaction temperature is 10-35°C, more preferably 20-30°C; Preferably, the mass ratio of the zearalenone to the heavy water is 1-(20:100); Preferably, the mass ratio of zearalenone to the strong base and weak acid salt is 1:(5-10).

9. A detection reagent, characterized in that: It comprises the deuterated zearalenone derivative compound according to claim 1.

10. A method for determining the content of zearalenone, characterized in that: include: The sample to be tested containing zearalenone is pre-treated and mixed with a derivatization reagent to be derivatized to form a derivatized sample to be tested containing zearalenone derivatives; The derivative sample to be tested is mixed with the deuterated zearalenone derivative compound of claim 1 or the detection reagent of claim 9, Then, the mass spectrum is detected, and the content of the zearalenone derivative is determined according to the peak area of ​​the mass spectrum result; Wherein, the added amount of the deuterated zearalenone derivative compound is not less than 100 μg / L; Preferably, the added amount of the deuterated zearalenone derivative compound is 200 μg / L-1000 μg / L.

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