CN115219643A - Detection method for confirming furacilin source and non-furacilin source semicarbazide in aquatic product - Google Patents

Abstract

The invention discloses a detection method for confirming furacilin source and non-furacilin source semicarbazide in aquatic products, which comprises the following steps: detecting residual quantity of semicarbazide in a sample; (II) detecting the residual quantity of the 5-nitro-2-furfural derivative in the sample; and (III) confirming. The invention takes 5-nitro-2-furfural as a specific biological marker, uses 2, 4-dinitrophenylhydrazine to carry out a derivatization reaction to generate a 5-nitro-2-furfural derivative (NF-DNPH), carries out identification and detection by an UPLC-MS/MS method, and finally confirms the source of semicarbazide (non-furacilin-source semicarbazide and furacilin-source semicarbazide) by combining the detection results of semicarbazide and the 5-nitro-2-furfural derivative, thereby effectively solving the false positive problem caused by the non-furacilin-source semicarbazide in aquatic products and providing an effective method for the detection and identification of the furacilin and the semicarbazide in the aquatic products.

Description

A method for confirming nitrofurazone source and non-nitrofurazone source semicarbazide in aquatic products Detection method

technical field

The invention relates to a semicarbazide detection method, in particular to a detection method for confirming the semicarbazide source and non-nitrocarbazone source in aquatic products.

Background technique

Semicarbazide (SEM), also known as carbamoyl hydrazide, has a chemical formula of CH ₅ N ₃ O. It is generally believed that semicarbazide is a typical metabolite of nitrofurazone, a commonly used aquaculture drug, in animals. Due to its stable nature, the EU, the United States and my country all use SEM as the marked residue for residue detection and monitoring of nitrofurazone. As the metabolite of semicarbazide, Nitrofurazone (NFZ) is a typical representative of nitrofuran drugs, because of its strong bactericidal ability, broad antibacterial spectrum, resistance to drug resistance, low price, and good efficacy ^{[2-3] ]} and is widely used in aquaculture. Studies in recent years have shown that the residues of nitrofurazone and its metabolites in animal-derived foods can be transmitted to humans through the food chain, and long-term intake can cause hemolytic anemia, polyneuritis, eye damage and acute liver necrosis and other diseases. It has carcinogenic and genotoxic effects and seriously endangers human health. At present, most countries such as the United States, Japan, the European Union, and China are prohibited from using it for aquaculture. Corresponding residue limits have been formulated successively and strict residue monitoring of aquatic products has been carried out.

It has always been believed that nitrofurazone is the main source of semicarbazide production, and many studies have carried out a lot of research on the metabolic process and production mechanism of nitrofurazone. However, since 2003, studies have continuously found that semicarbazide produced by the metabolism of non-nitrofuracil in food and organisms, collectively referred to as semicarbazide derived from non-nitrofuran. After the European Union released a research report that found semicarbazide from non-furan sources and assessed the risk of exposure, it aroused widespread concern among researchers.

In recent years, some studies have reported the discovery of non-nitrofurazone-derived semicarbazide in crustacean aquatic products. Hoenicke's study found that SEM appeared in shrimp products. Becalsk, Stadler, and Christof also successively discovered that non-nitrofurazone-derived semicarbazide appeared in aquatic products. Phenomenon. Since semicarbazide is the labeled residue in the detection of nitrofurazone in aquatic products, the current detection method cannot distinguish the semicarbazide formed by the metabolism of nitrofurazone from the semicarbazide formed by other pathways. Therefore, the existence of semicarbazide derived from non-nitrofuran in aquatic products has brought serious false-positive problems for nitrofurazone drug testing, resulting in the inability of testing institutions and regulatory agencies to effectively supervise the abuse of nitrofurazone.

SUMMARY OF THE INVENTION

The present invention is to solve the problem that the existence of non-nitrofurazone-derived semicarbazide in the existing aquatic products brings serious false positives to the drug detection of nitrofurazone, resulting in the inability of testing institutions and regulatory agencies to effectively supervise the abuse of nitrofurazone, and provides a The detection method used to confirm the source of nitrofurazone and semicarbazide from non-nitroxide in aquatic products is simple and easy to operate, has high sensitivity, satisfactory recovery rate and reproducibility, and can effectively solve the problem of semicarbazide caused by non-nitrocarbazone in aquatic products. It provides an effective method for the detection and identification of aquatic products nitrofurazone and semicarbazide.

In order to achieve the above object, the present invention adopts the following technical solutions:

A detection method of the present invention for confirming nitrofurazone source and non-nitrofurazone source semicarbazide in aquatic products comprises the following steps: (1) detecting the residual amount of semicarbazide in the sample. The detection of the residual amount of semicarbazide (SEM) is the prior art in the art, and reference may be made to the detection method in GB-31656.13-2021.

(2) Detect the residual amount of 5-nitro-2-furfural derivatives in the sample

(1) Take an appropriate amount of aquatic product tissue, add hydrochloric acid solution and 2,4-dinitrophenylhydrazine solution, mix by vortex, ultrasonically shake, adjust pH to 7.5 with dipotassium hydrogen phosphate, add ethyl acetate, and mix by vortex Centrifuge, take the upper layer of the extraction solution, spin and evaporate to dryness, add acetonitrile aqueous solution to dissolve, and get the machine solution after the solution passes through the PTFE membrane. Non-nitrofuran source semicarbazide, as an endogenous semicarbazide in crustacean aquatic products, can lead to false positives in the detection of nitrofuran, which has been troubled by the detection of aquatic products; A series of compounds will be produced, including 3 intermediates similar to nitrofurazone, cyano metabolites, 5-nitro-2-furfural, and other protein and DNA conjugates. The inventors confirmed through mass spectrometry and found that arginine, Histidine, citrulline, inositol, urea, and creatinine are all precursors or intermediates of semicarbazide (SEM). Since these substances are ubiquitous in aquatic products, they are used for the identification of semicarbazide from non-nitrofurazone sources. There is no specificity for identification by markers; among them, cyano metabolites have a better spectral ionization effect, and the mass spectral precursor ion is m/z169, which can be used for detection, but because it is difficult to separate pure products, it is specific as semicarbazide derived from nitrofurazone. It is very unlikely that sex markers can be used in a generalized detection method; 5-nitro-2-furfural is a metabolite of nitrofural, which co-exists with semicarbazide in nitrofurazone metabolites, but 5-nitro- 2-Furfural does not exist in nature. Therefore, in the present invention, it is determined that 5-nitro-2-furfural is used as a specific biomarker to identify semicarbazide derived from nitrofurazone and semicarbazide other than nitrofural; however, due to structural problems, 5-nitro-2-furfural is difficult to ionize directly. , its mass spectrometry response is very weak, can only reach the mg/kg level, and the sensitivity is not enough as a specific marker of nitrofurazone source semicarbazide directly, but it has a commercial standard, so it is derivatized in the present invention (using 2, 4-dinitrophenylhydrazine) to improve sensitivity and as a specific marker for distinguishing nitrofuracil-derived SEM from non-nitrofuracil-derived SEM.

The reaction derivation process of semicarbazide with 2-nitrobenzaldehyde (2-NBA), 5-nitro-2-furfural and 2,4-dinitrophenylhydrazine (DNPH) is as follows:

The derivatization process of 5-nitro-2-furfural is as follows:

(2) Extract the solution on the machine with a syringe, and use an ultra-high performance liquid chromatography-tandem mass spectrometer to detect according to the set conditions, and obtain the mass ion chromatogram of the detected sample. In the present invention, ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) is used for detection.

(3) Standard curve drawing:

(i) Preparation of standard stock solution: add 250 mL of acetonitrile aqueous solution to 10 mg of 5-nitro-2-furfural derivative standard product, and the volume ratio of acetonitrile to water in the acetonitrile aqueous solution is 7:3 to prepare 5-nitro-2- Standard stock solution of furfural derivatives;

(ii) Preparation of standard working solution: The standard stock solution of 5-nitro-2-furfural derivatives was gradually prepared into 0.4, 1.0, 4.0, 10 and 20 μg/L of 5-nitro-2-furfural derivatives with acetonitrile aqueous solution Standard working solution, the volume ratio of acetonitrile to water in acetonitrile aqueous solution is 7:3;

(iii) Preparation of standard curve: use external standard method for quantification, inject the standard working solution of 5-nitro-2-furfural derivatives according to step (2), and analyze each time three times to measure the characteristic ion mass spectrum peaks. The peak area of is the ordinate, the corresponding standard working solution concentration is the abscissa, and the standard curve is established;

(4) Measurement

Substitute the peak area of the 5-nitro-2-furfural derivative in the ion chromatogram of the detected sample mass spectrum in step (2) into the standard curve, and the actual content of the 5-nitro-2-furfural derivative can be obtained by calculation;

(3) Confirmation

If the 5-nitro-2-furfural derivative is not detected in step (2), it can be determined that the semicarbazide detected in step (1) is semicarbazide from non-nitrofurazone; if it is detected in step (2) that contains 5-nitrocarbazone -nitro-2-furfural derivative, then it can be determined that the semicarbazide detected in step (1) is nitrofurazone source semicarbazide.

Preferably, in step (1), the method for detecting the residual amount of semicarbazide in the sample adopts ultra-high performance liquid chromatography-tandem mass spectrometry.

Preferably, in step (1), take 1.0±0.1g of aquatic product tissue, add 10mL, 0.2mol/L hydrochloric acid solution, then add 100μL, 1.0mg/L 2,4-dinitrophenylhydrazine solution, vortex mixing 1min, ultrasonically shake for 30min, adjust the pH to 7.5 with 0.1mol/L dipotassium hydrogen phosphate, add 8mL of ethyl acetate, mix by vortex, centrifuge at 5000r/min for 5min, take the upper layer of the extraction solution, spin and evaporate to dryness at 40°C , add 1 mL of acetonitrile aqueous solution, the volume ratio of acetonitrile to water in the acetonitrile aqueous solution is 7:3, ultrasonically dissolve for 1 min, and after passing through the PTFE membrane, the machine solution is obtained.

Preferably, in step (2), the chromatographic conditions are: chromatographic column: ACQUITY UPLC BEH C18 column, 2.1 mmi.d.×50 mm, particle size 1.7 μm; column temperature 40°C; sample temperature 10°C; injection volume: 10 μL ; Flow rate: 0.2 mL/min; Mobile phase A is an aqueous solution containing 2 mM ammonium acetate and 0.1% formic acid, mobile phase B is acetonitrile, eluted with 70% A and 30% B isocratic.

Preferably, in step (2), mass spectrometry conditions: ion source: electrospray ion source, positive ion scanning; detection mode: multiple reaction monitoring; capillary voltage: 3.5kV; ion source temperature: 120 °C; desolvation gas temperature: 380 ℃; Cone gas flow: 50L/h; Desolvation gas flow: 600L/h. The reaction monitoring conditions are:

Note: * is the quantitative product ion.

Preferably, in step (i), the 5-nitro-2-furfural derivative standard product is prepared by the following method: dissolve 100 mg of 5-nitro-2-furfural in 10 mL of acetonitrile, add dropwise to acidify 2 within 30 min , 4-dinitrophenylhydrazine; after the dropwise addition, the mixture was shaken overnight in the dark, the obtained reddish-brown precipitate was washed with hydrochloric acid, and recrystallized from methanol-acetonitrile to obtain the crude product; the crude product was redissolved in 20 mL of acetonitrile aqueous solution In the acetonitrile aqueous solution, the volume ratio of acetonitrile to water in the acetonitrile aqueous solution was 9:1; after filtration by ultrasonic-0.2 micron filter membrane, it was purified by semi-preparative chromatography; all the collected materials were evaporated to dryness with a rotary evaporator; The purity of the 5-nitro-2-furfural derivative standard is greater than 99%. At present, there is no commercially available standard product of 5-nitro-2-furfural derivatives, so the inventor synthesized a standard product of 5-nitro-2-furfural derivatives (NF-DNPH) by himself; 5-nitro-2-furfural derivatives synthesized by this method - The standard product of nitro-2-furfural derivatives has high purity and is stored in a brown bottle and stored in a refrigerator at 4 °C; within two months, the standard solution of 5-nitro-2-furfural derivatives is Analyzed every two weeks, the relative standard deviation of chromatographic peak areas is less than 5%, which indicates that 5-nitro-2-furfural derivatives have sufficient stability to be stored for several months; different kinds of blank samples (fish, The specificity of the derivatives was examined by the method of crab and shrimp); the results showed that there was no interference from the matrix effect near the detection target.

Preferably, the chromatographic conditions of the semi-preparative chromatography are: mobile phase: acetonitrile-water, the volume ratio of acetonitrile and water is 9:1; column temperature: 40°C; detector wavelength 360nm; thing.

Preferably, the acidified 2,4-dinitrophenylhydrazine is prepared by the following method: 150 mg of 2,4-dinitrophenylhydrazine is dissolved in 20 mL of 12M concentrated hydrochloric acid.

Therefore, the present invention has the following beneficial effects:

(1) Using 5-nitro-2-furfural as a specific biomarker, 2,4-dinitrophenylhydrazine was used for derivatization to generate 5-nitro-2-furfural derivative (NF-DNPH), which was obtained by UPLC-MS/MS method was used to identify and detect, and finally, the source of semicarbazide (non-nitrofurazone and nitrofurazone-derived semicarbazide) was confirmed by combining the test results of semicarbazide and 5-nitro-2-furfural derivatives, which can effectively solve the problem. The false positive problem caused by semicarbazide from non-nitrofurazone in aquatic products provides an effective method for the detection and identification of nitrofurazone and semicarbazide in aquatic products;

(2) The detection method for the residual amount of 5-nitro-2-furfural derivatives in the present invention is in the linear range of 0.4-20 ng/g, the linearity is good, the recovery rate is 90.6-104.7%, and the relative standard deviation is less than 5%; method The limit of detection is 0.1ng/g, and the limit of quantification is 0.2ng/g, which can meet the limit standard of 1.0ng/g in the European Union, with high sensitivity and satisfactory recovery and reproducibility.

Detailed ways

The present invention will be further described below through specific embodiments.

The standard 5-nitro-2-furfural derivatives in the following examples were prepared by the following method: 100 mg of 5-nitro-2-furfural was dissolved in 10 mL of acetonitrile, and added dropwise to acidified 2,4- In dinitrophenylhydrazine, acidified 2,4-dinitrophenylhydrazine was prepared by the following method: 150mg 2,4-dinitrophenylhydrazine was dissolved in 20mL, 12M concentrated hydrochloric acid; after the dropwise addition, the The mixture was shaken overnight in the dark, the obtained reddish-brown precipitate was washed with hydrochloric acid, and the crude product was obtained after recrystallization from methanol-acetonitrile; the crude product was redissolved in 20 mL of acetonitrile aqueous solution, and the volume ratio of acetonitrile to water in the acetonitrile aqueous solution was 9:1; After ultrasonic-0.2 micron membrane filtration, it was purified by semi-preparative chromatography (600 analytical/semi-preparative high performance liquid chromatograph equipped with UV detector, American waters company), and the chromatographic conditions of semi-preparative chromatography were: mobile phase: acetonitrile-water, The volume ratio of acetonitrile and water is 9:1; the column temperature: 40°C; the detector wavelength is 360nm; the effluent from the effluent end is collected for 14 to 16 minutes; all the collected materials are evaporated to dryness with a rotary evaporator; , The purity of 5-nitro-2-furfural derivative standard product is greater than 99%; it is stored in a brown bottle and stored in a refrigerator at 4°C.

Example 1

(1) The residual amount of semicarbazide in the test sample, the test sample is the crab shell of wild swimming crab, the test method is the same as the semicarbazide test method in GB-31656.13-2021, and the test results are shown in Table 1.

(2) Detect the residual amount of 5-nitro-2-furfural derivatives in the sample

(1) Take 1.0g of the test sample (crab shells of wild sea swimming crabs, bought at Zhoushan Lincheng Market), add 10mL, 0.2mol/L hydrochloric acid solution, and then add 100μL, 1.0mg/L of 2,4- Dinitrophenylhydrazine solution, mixed by vortex for 1min, sonicated for 30min, adjusted to pH 7.5 with 0.1mol/L dipotassium hydrogen phosphate, added 8mL of ethyl acetate, mixed by vortex, centrifuged at 5000r/min for 5min, and the upper layer was taken. The solution was extracted and evaporated to dryness by rotary evaporation at 40°C. Then, 1 mL of acetonitrile aqueous solution was added. The volume ratio of acetonitrile to water in the acetonitrile aqueous solution was 7:3, and the solution was ultrasonically dissolved for 1 min. After passing through the PTFE membrane, the machine solution was obtained.

，美国waters公司)按照设定好的条件进行检测，得检测样品质谱离子流图；色谱条件为：色谱柱：ACQUITY UPLC BEH C18柱，2.1mm i.d.×50mm，粒径1.7μm；柱温40℃；样品温度10℃；进样量：10μL；流速：0.2mL/min；流动相A为含2mM乙酸铵和0.1％甲酸的水溶液，流动相B为乙腈，以70％A和30％B等度洗脱；质谱条件：离子源：电喷雾离子源，正离子扫描；检测方式：多反应监测；毛细管电压：3.5kV；离子源温度：120℃；脱溶剂气温度：380℃；锥孔气流量：50L/h；脱溶剂气流量：600L/h；反应监测条件为：(2) Use a syringe to extract the solution on the machine, and use an ultra-high performance liquid chromatography-tandem mass spectrometer (Quattro PremierXE ^TM) .

, U.S. waters company) according to the set conditions for detection, the mass spectrometry ion chromatogram of the detected sample is obtained; the chromatographic conditions are: chromatographic column: ACQUITY UPLC BEH C18 column, 2.1mm id × 50mm, particle size 1.7μm; column temperature 40 ℃ ; Sample temperature: 10 °C; Injection volume: 10 μL; Flow rate: 0.2 mL/min; Mobile phase A is an aqueous solution containing 2 mM ammonium acetate and 0.1% formic acid, and mobile phase B is acetonitrile, with 70% A and 30% B isocratic elution; mass spectrometry conditions: ion source: electrospray ion source, positive ion scanning; detection mode: multiple reaction monitoring; capillary voltage: 3.5kV; ion source temperature: 120 °C; desolvation gas temperature: 380 °C; cone gas flow : 50L/h; flow rate of desolventizing gas: 600L/h; reaction monitoring conditions are:

Note: * is the quantitative product ion.

(3) Drawing of standard curve (i) Preparation of standard stock solution: add 250 mL of acetonitrile aqueous solution to 10 mg of 5-nitro-2-furfural derivative standard product, and the volume ratio of acetonitrile to water in the acetonitrile aqueous solution is 7:3 to make 5 - Standard stock solution of nitro-2-furfural derivatives.

(ii) Preparation of standard working solution: The standard stock solution of 5-nitro-2-furfural derivatives was gradually prepared into 0.4, 1.0, 4.0, 10 and 20 μg/L of 5-nitro-2-furfural derivatives with acetonitrile aqueous solution Standard working solution, the volume ratio of acetonitrile to water in acetonitrile aqueous solution is 7:3.

(iii) Preparation of standard curve: use external standard method for quantification, inject the standard working solution of 5-nitro-2-furfural derivatives according to step (2), analyze three times each, and use the measured characteristic ion mass spectrum peaks The peak area of is the ordinate, the corresponding standard working solution concentration is the abscissa, and the standard curve is established. The correlation coefficients of the linear equations are: r ² =0.993.

(4) Measurement

Substitute the peak area of the 5-nitro-2-furfural derivative in the ion chromatogram of the detected sample mass spectrum in step (2) into the standard curve, and the actual content of the 5-nitro-2-furfural derivative can be obtained by calculation, The test results are shown in Table 1.

Table 1 embodiment 1 actual sample detection and analysis results

^a ND, not detected or <LOD.

(3) Confirmation

As can be seen from Table 1, SEM is detected in the sample of Example 1, but NF is not detected. According to this, it can be determined that the wild sea bream is not added with nitrofurazone, and natural endogenous SEM is produced, which is consistent with the relevant literature reports.

Example 2

The detection sample in the embodiment 2 adopts the shrimp shell of Penaeus vannamei (buy in Lincheng Market, Zhoushan), the detection method is the same as that of the embodiment 1, and the detection results are shown in Table 2.

Table 2 embodiment 2 actual sample detection and analysis results

As can be seen from Table 2, SEM and NF were simultaneously detected in the sample of Example 2, according to which it could be determined that the commercially available Penaeus vannamei was added with nitrofurazone.

In the method for detecting the residual amount of 5-nitro-2-furfural derivatives in the present invention, NF-DNPH has good linearity in the concentration range of 0.4-20ug/L, and the recovery rate is 90.6-104.7 in the linear range of 0.4-20ng/g %, relative standard deviation <5%, linearity, recovery and precision are shown in Table 3.

Table 3 Linearity, recovery and precision

The above-mentioned embodiment is only a preferred solution of the present invention, and does not limit the present invention in any form, and there are other variations and modifications under the premise of not exceeding the technical solution recorded in the claims.

Claims (8)

1. A detection method for confirming furacilin source and non-furacilin source semicarbazide in aquatic products is characterized by comprising the following steps:

detecting the residual quantity of the semicarbazide in the sample;

(II) detecting the residual quantity of the 5-nitro-2-furfural derivative in a sample

(1) Taking a proper amount of water product tissue, adding a hydrochloric acid solution and a 2, 4-dinitrophenylhydrazine solution, carrying out vortex mixing, carrying out ultrasonic oscillation, adjusting the pH to 7.5 by using dipotassium phosphate, adding ethyl acetate, carrying out vortex mixing, centrifuging, taking an upper layer extraction solution, carrying out rotary evaporation, adding an acetonitrile aqueous solution for dissolving, and passing the solution through a PTFE (polytetrafluoroethylene) membrane to obtain an upper machine solution;

(2) Extracting the solution fed into the machine by using a sample injection needle, and detecting by using an ultra-high performance liquid chromatography-tandem mass spectrometer according to set conditions to obtain a mass spectrum ion flow diagram of a detected sample;

(3) Drawing of standard curve

Preparation of standard stock solutions: adding 250mL of acetonitrile aqueous solution into 10mg of 5-nitro-2-furfural derivative standard, wherein the volume ratio of acetonitrile to water in the acetonitrile aqueous solution is 7:3, preparing a standard stock solution of the 5-nitro-2-furfural derivative;

(ii) preparing a standard working solution: using a 5-nitro-2-furfural derivative standard working solution with acetonitrile aqueous solution of 0.4, 1.0, 4.0, 10 and 20 mug/L step by step, wherein the volume ratio of acetonitrile to water in the acetonitrile aqueous solution is 7:3;

(iii) preparation of a standard curve: quantifying by adopting an external standard method, carrying out sample injection on the 5-nitro-2-furfural derivative standard working solution according to the step (2), analyzing for three times respectively, taking the peak area of the measured characteristic ion mass spectrum peak as a vertical coordinate and the corresponding standard working solution concentration as a horizontal coordinate, and establishing a standard curve;

(4) Measurement of

Substituting the peak area of the 5-nitro-2-furfural derivative in the ion flow diagram of the detected sample quality spectrum in the step (2) into a standard curve, and calculating to obtain the actual content of the 5-nitro-2-furfural derivative;

(III) confirmation

If 5-nitro-2-furfural derivatives are not detected in the step (II), determining that the semicarbazide detected in the step (I) is non-furacilin semicarbazide; and (d) if the 5-nitro-2-furfural derivative is detected in the step (II), determining that the semicarbazide detected in the step (I) is furacilin-derived semicarbazide.

2. The method for detecting the furacilin source and the semicarbazide which is not the furacilin source in the aquatic products according to claim 1, wherein the method for detecting the residual quantity of the semicarbazide in the sample in the step (1) adopts ultra high performance liquid chromatography-tandem mass spectrometry.

3. The method for detecting the furacilin source and the non-furacilin source semicarbazide in the aquatic product according to claim 1, characterized in that in the step (1), 1.0 +/-0.1 g of aquatic product tissue is taken, 10mL and 0.2mol/L of hydrochloric acid solution are added, 100 mu L and 1.0mg/L of 2, 4-dinitrophenylhydrazine solution are added, vortex mixing is carried out for 1min, ultrasonic oscillation is carried out for 30min, 0.1mol/L of dipotassium hydrogen phosphate is used for adjusting the pH value to 7.5, 8mL of ethyl acetate is added, after vortex mixing, the mixture is centrifuged at the rotating speed of 5000r/min for 5min, the upper layer extracting solution is taken, after rotary evaporation at 40 ℃, 1mL of acetonitrile aqueous solution is added, and the volume ratio of acetonitrile to water in the acetonitrile aqueous solution is 7: and 3, ultrasonically dissolving for 1min, and passing through a PTFE membrane to obtain the upper machine solution.

4. The method for detecting semicarbazide which is capable of confirming the source of nitrofural and non-nitrofural in aquatic products according to claim 1, wherein in the step (2), the chromatographic conditions are as follows: a chromatographic column: an ACQUITY UPLC BEH C18 column, 2.1mm i.d. x 50mm, particle size 1.7 mu m; the column temperature is 40 ℃; the sample temperature is 10 ℃; sample introduction amount: 10 mu L of the solution; flow rate: 0.2mL/min; mobile phase a was an aqueous solution containing 2mM ammonium acetate and 0.1% formic acid, mobile phase B was acetonitrile, isocratically eluted at 70% a and 30% B.

5. The detection method for confirming nitrofurazone-derived and non-nitrofurazone-derived semicarbazides in aquatic products as claimed in claim 1, wherein in the step (2), mass spectrum conditions are as follows: an ion source: electrospray ion source, positive ion scanning; the detection mode is as follows: monitoring multiple reactions; capillary voltage: 3.5kV; ion source temperature: 120 ℃; desolventizing gas temperature: 380 ℃; taper hole gas flow: 50L/h; desolventizing agent gas flow: 600L/h.

6. The method of claim 1, wherein in step (i), the 5-nitro-2-furfural derivative standard is prepared by the following method: dissolving 100mg of 5-nitro-2-furfural in 10mL of acetonitrile, and dropwise adding into acidified 2, 4-dinitrophenylhydrazine within 30 min; after the dropwise addition is finished, oscillating the mixture overnight in the dark, washing the obtained reddish brown precipitate with hydrochloric acid, and recrystallizing with methanol-acetonitrile to obtain a crude product; the crude product was redissolved in 20mL of an aqueous acetonitrile solution having a volume ratio of acetonitrile to water of 9:1; filtering with ultrasonic-0.2 μm filter membrane, and purifying with semi-preparative chromatography; evaporating all the collected materials to dryness by using a rotary evaporator; the purity of the 5-nitro-2-furfural derivative standard substance is more than 99 percent through HPLC-TUV detection.

7. The detection method for confirming nitrofurazone-derived and non-nitrofurazone-derived semicarbazides in aquatic products as claimed in claim 6, wherein chromatographic conditions of the semi-preparative chromatography are as follows: mobile phase: acetonitrile-water, the volume ratio of acetonitrile to water is 9:1; column temperature: 40 ℃; the wavelength of the detector is 360nm; the outflow end collected the effluent for 14 to 16 min.

8. The detection method for confirming nitrofurazone-derived and non-nitrofurazone-derived semicarbazides in aquatic products as claimed in claim 6, wherein said acidified 2, 4-dinitrophenylhydrazine is prepared by the following method: 150mg of 2, 4-dinitrophenylhydrazine was dissolved in 20mL of 12M concentrated hydrochloric acid.