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Analysis of Endogenous Semicarbazide during the Whole Growth Cycle of Litopenaeus vannamei and Its Possible Biosynthetic Pathway Wenlong Yu,† Weihua Liu,† Yaxin Sang,† and Xianghong Wang*,†,‡ †

Department of Food Science and Technology, Hebei Agricultural University, Baoding, Hebei 071001, People’s Republic of China Beijing Advanced Innovation Center for Food Nutrition and Human Health, Beijing Technology and Business University (BTBU), Fangshan, P. R. China

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S Supporting Information *

ABSTRACT: This research aims to analyze the biosynthetic pathway of endogenous semicarbazide (SEM) in shrimps using Litopenaeus vannamei as the model target. To achieve this objective, the content of SEM in L. vannamei throughout the whole growth cycle was monitored under the strict control of external environmental interference. Experimental results showed that SEM was found in the shrimp shell at all stages, with its content decreasing first and then increasing, and no SEM was detected in the shrimp muscle of each growth stage. This indicated that endogenous SEM in L. vannamei was derived from the shrimp shell. At the same time, the content of amino acids in the shrimp shell and the corresponding substances involved in the urea cycle in the entire growth cycle of shrimp were monitored. The correlation analysis between them and the changes in the SEM content in shrimp showed that arginine had the largest correlation coefficient (0.952) with the changes in the SEM content. The main substances of the urea cycle may be related to the production of SEM. In combination with the water environmental test of high ammonia nitrogen, it was presumed that the formation of endogenous SEM was related to the amidine group of arginine and amide structure of citrulline and urea. Arginine, citrulline, and urea in the urea cycle of L. vannamei eventually produced SEM via an oxaziridine intermediate under the action of hydrogen peroxide and ammonia, and a standardized reaction test was conducted to verify the hypothesis and, thus, provided a new idea for future endogenous SEM research. KEYWORDS: Litopenaeus vannamei, semicarbazide, biosynthetic pathway, amino acids, arginine, urea cycle

1. INTRODUCTION Semicarbazide (SEM), which is a kind of amine molecule compound, is a moderately accumulating toxic substance. It has reproductive toxicity and mutagenic effects, and it is generally considered to be a metabolite of the banned drug nitrofurazone.1−3 The chemical structure of SEM is shown in Figure 1. Today, many countries use the detection of

(disinfection or bleaching), and endogenous SEM (crustacean aquatic products).8−10 In recent years, several studies have reported that SEM was detected in many crustacean aquatic animals without feeding nitrofurazone.11,12 In particular, the detection rate of SEM in shrimps was higher. This high rate was due to SEM produced by the shrimp itself and not due to the illegal use of nitrofurazone.13 The existence of endogenous SEM nevertheless brings serious false-positive problems to the detection of nitrofurazone-derived drugs. Besides, the safety of shrimp products has been questioned, a situation that has resulted in huge economic losses to the shrimp-farming industry.14 Therefore, it is of great significance to deeply explore the mechanism and formation of endogenous SEM in shrimps. Currently, the research on the biosynthetic pathway of endogenous SEM is still in the preliminary inference stage. Many researchers have speculated that amino acids may be related to the production of endogenous SEM in aquatic products. Hoenicke et al.6 assumed that SEM was formed by the degradation of nitrogen-containing substances, such as arginine, histidine, citrulline, urea, and creatinine, with amidine or urea groups. Crews et al.8 also showed that, when the

Figure 1. Molecular structure of nitrofurazone and SEM.

nitrofurazone metabolite to determine whether nitrofurazone is illegally added in the breeding process to monitor the original drug of nitrofurazone.4,5 However, many researchers have argued that SEM cannot be used as evidence for the use of nitrofurazone because it cannot be derived only from this organic substance.6,7 That is, it can be obtained from other sources, such as flour quality improver (azocarbamide thermal degradation), contamination of resin raw materials (azocarbamide), food processing, protein treated with hypochlorite © XXXX American Chemical Society

Received: Revised: Accepted: Published: A

March 21, 2019 June 7, 2019 July 1, 2019 July 1, 2019 DOI: 10.1021/acs.jafc.9b01779 J. Agric. Food Chem. XXXX, XXX, XXX−XXX

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Journal of Agricultural and Food Chemistry

2.3. Analysis of SEM Using Liquid Chromatography− Tandem Mass Spectrometry (LC−MS/MS). The measurement of SEM was performed according to a previously reported procedure,26,27 with some modifications. Approximately 1 g of shrimp sample (the shrimp muscle, shrimp shell, or whole shrimp) was analyzed and spiked with 100 μL of (13C,15N2)-SEM (50 μg mL−1), 10 mL of hydrochloric acid (0.2 mol L−1, pH 3.5), and 100 μL of NBA (50 mmol L−1, 378 mg in 5 mL of dimethyl sulfoxide). The reaction mixtures were protected from light and heated on a rotary shaker at 37 °C for 16 h. Next, the samples were extracted with ethyl acetate and subjected to repeated solvent washes. After that, the final extracts were refrigerated (−4 °C) before the LC−MS/MS analysis. The final extracts were separated in a Thermo C18 column (2.1 × 150 mm, 5 μm). The mobile phase consisted of acetonitrile (A) and water with 0.1% (v/v) methane acid (B) at a constant flow rate of 0.2 mL/ min, with gradient elution. The mass spectrometer was operated in electrospray ionization positive (ESI+) mode; the source temperature was set at 120 °C; the desolvation temperature was set at 450 °C; and the desolvation gas flow was set at 800 L/h. Identification and quantification were performed using multiple reaction monitoring (MRM), taking the concentration as the abscissa and the ratio of the peak area of SEM to the peak area of the internal standard as the ordinate and establishing the standard curve of the SEM content. Samples of the aquaculture water and feedstuff were analyzed for SEM using the same procedure. 2.4. Analysis of Nitrofurazone in the Water Environment and Feedstuffs. The water environment and feedstuff of different stages were extracted with ethyl acetate, following centrifugation treatment and dilution with 50% methanol/water, and the extract solution was analyzed for the presence of nitrofurazone using highperformance liquid chromatography−tandem mass spectrometry (HPLC−MS/MS) operated in negative ion mode. 2.5. Analysis of Amino Acids Using High-Performance Liquid Chromatography−Diode Array Detector (HPLC− DAD). Approximately 1 mL of sample was added to 10 mL of 6 mol L−1 HCl solution and placed in a blast oven at 110 °C for 20 h. When the hydrolysis process was completed at 80−90 °C, 1 mL of supernatant was added to the test tube, followed using 2 mL of 0.1 mol L−1 HCl solution to dissolve the samples. Then, the mixture was vortex-mixed, then filtered over a 0.45 μm inorganic membrane, and put into a small centrifuge tube to be derived. Furthermore, 100 μL of the above sample was inserted into a 1.5 mL centrifuge tube. Other items were added to the tube, such as 200 μL of buffer salt (Na2CO3 and NaHCO3), 100 μL of derivatization agent (2,4-dinitrochlorobenzene), and 50 μL of 10% acetic acid. The mixture was diluted with water to extract the supernatant and was filtered over 0.45 μm organic membrane. After that, it was put into the sample vial to be detected by HPLC at 360 nm. 2.6. Analysis of Main Compounds in the Urea Cycle. Ornithine and citrulline were analyzed in L. vannamei using a previously reported HPLC-based method.28 Urea analysis of L. vannamei was performed using HPLC−MS/MS.29 2.7. Water Environmental Test of High Ammonia Nitrogen. In view of the influence of a high-ammonia-nitrogen environment and high-salinity environment on the urea cycle of L. vannamei, the concentration of ammonia nitrogen (1.5 mmol/L) and salinity (18 and 35) in the aquatic environments was artificially changed. L. vannamei was placed in these different environments for 24 h. After that, both the corresponding substance content of the urea cycle and the SEM content were measured. 2.8. Standardized Reactions. The main substances in the urea cycle (arginine, urea, ornithine, and citrulline) and H2O2 were mixed in 16 ways, and the amount of each substance added to the mixed system was fixed. The pretreatment method was according to the previous report,14 and SEM detection was performed using HPLC−MS/MS. By comparison of the content of SEM produced, the possible production ways of SEM were further analyzed. 2.9. Statistical Analysis. All measurements were performed in triplicate, and the data were recorded as the mean value ± standard

arginine content in Photinia pollen increased suddenly, the content of SEM in honey also rose sharply. Similarly, Ngoan et al.15 confirmed that arginine was the most abundant amino acid in some crustacean species. They also speculated that arginine might be related to the formation of endogenous SEM in the crustacean. With guanidine and amidine groups, arginine participates in the urea cycle, and this substance can be hydrolyzed into ornithine and urea by arginase. While urea and SEM have certain structural similarities.16−18 Abernethy19 suggested that SEM might in part be formed during the acid conditions of analysis in foods containing urea compounds. Hence, it is speculated that the process of the urea cycle has a potential relationship with the generation of SEM. However, many factors affect the urea cycle of shrimps.20 Some studies have confirmed that high-saline and high-ammonia-nitrogen environments could cause the rapid increase of urea in shrimps, leading to the accumulation of ammonia and urea in the hemolymph and catalyzing the decomposition of hemocyanin and protein in the hemolymph to form free amino acids. The salinity and ammonia nitrogen concentration also had an effect on the activity of arginase and catalase in shrimps.21−25 Therefore, the ammonia nitrogen concentration and salinity in the water growth environment may also affect the production of SEM in shrimps. However, these speculations cannot be taken seriously because they cannot be substantiated with sufficient experimental data. The purpose of this study is to analyze the biosynthetic pathway of endogenous SEM in shrimps using Litopenaeus vannamei as the research object. To achieve this goal, its SEM content, amino acid content, and urea-cycle-related substance content will be monitored during the complete growth cycle. The possible biosynthetic pathway of endogenous SEM in L. vannamei will also be analyzed using correlation analysis, water environmental test of high ammonia nitrogen, and standardized reaction test. Finally, a new theory for the generation of SEM in L. vannamei will be presented. However, this research will presume that endogenous SEM in shrimp will be obtained by the reaction of the amidine group of arginine with citrulline and/or urea containing the urea structure.

2. MATERIALS AND METHODS 2.1. Materials and Instruments. The Waters TQD with a MassLynx version 4.1 data system (Waters, Milford, MA, U.S.A.), SHA-B constant temperature oscillator (Shanghai, China), nitrogen blower (EYELA, Japan), rotary evaporator (EYELA, Japan), samples of SEM (99%), and isotopically labeled SEM (13C,15N2, 99%) were purchased from Yuanye Co., Ltd. (Shanghai, China). The samples of sodium hydroxide, sodium chloride, and triethylamine were purchased from Tianli Chemical Reagent Co., Ltd. (Tianjin, China). Methanol, ethyl acetate, hexane, and acetonitrile were bought from Merck (Germany), while high-performance liquid chromatography (HPLC)-grade 2-nitrobenzaldehyde (NBA, 98%) was acquired at Yuanye Co., Ltd. (Shanghai, China). Both 2,4-dinitrochlorobenzene and urea were provided by Sigma-Aldrich (St. Louis, MO, U.S.A.). 2.2. Sample Collection and Preparation. L. vannamei were bred in the ecological shrimp breeding center of Huanghua, Hebei, China. During the complete growth stage of L. vannamei, samples were taken every 7 days for a total of 91 days and the addition of nitrofuran antibiotics was strictly prohibited. Also, the aquaculture water environment, feed safety, and disinfection method (avoiding sodium hypochlorite) were monitored throughout the process. To minimize interference from any possible external factors, the aquaculture water and feedstuff were taken every 7 days for a total of 91 days to detect the presence of nitrofurazone and SEM. B

DOI: 10.1021/acs.jafc.9b01779 J. Agric. Food Chem. XXXX, XXX, XXX−XXX

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Figure 2. Content of SEM in different components of the whole growth cycle of shrimp.

Figure 3. Heatmap of the amino acid content in shrimp shell throughout the whole growth cycle. deviation. Statistical significance (p value of intestinal glands and internal organs > muscle; Table S2 of the Supporting Information), which was consistent with the distribution of arginase activity, indirectly confirming that SEM had a certain relationship with D

DOI: 10.1021/acs.jafc.9b01779 J. Agric. Food Chem. XXXX, XXX, XXX−XXX

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circulation in ammonia excretion increased; therefore, the urea content of the shrimp skyrocketed, and the content of the corresponding substance in the urea cycle also soared. The results showed the SEM content of L. vannamei in different water environment systems (salinity of 35 + 1.5 mmol/L ammonia nitrogen > salinity of 35 > salinity of 18 + 1.5 mmol/ L ammonia nitrogen > salinity of 18). This indicated that the increase of salinity and ammonia nitrogen in the water environment can not only change the urea cycle in shrimps but also affect the formation of SEM in shrimps. The main substances of the urea cycle may be related to the production of SEM. 3.7. Possible Biosynthetic Pathway of Endogenous SEM in L. vannamei. On the basis of the experimental results of this study, in combination with previous research results,19 we propose a possible pathway for the natural generation of SEM in L. vannamei; the proposed mechanism is illustrated in Figure 6. In the high ammonia nitrogen environment, the proportion of urea circulation in ammonia excretion increases. Thus, there is an increase in both the urea content and the corresponding substance in the urea cycle. Additionally, the shrimp body contains hydrogen peroxide, and the content of hydrogen peroxide expands sharply with the increase of the ammonia nitrogen concentration in the water environment. Citrulline and/or urea are oxidized by hydrogen peroxide, and they react with the amidine group of arginine to form oxaziridine. The action of ammonia or amines on oxaziridine consequently generates hydrazones. Under hydrolysis in the presence of urea, hydrazine may be released from hydrazones to react with urea, forming SEM. Some hydrazones will react with urea to form azine, which includes a semicarbazone substructure. The reaction of urea with oxaziridine may also generate a semicarbazone directly. Semicarbazone undergoes hydrolysis to form SEM. During HPLC−MS/MS analysis, any semicarbazone present will release SEM and form NBA−SEM for analysis. 3.8. Standardized Reactions. To verify the conjecture about the biosynthetic pathway of endogenous SEM in L. vannamei. The SEM detection of standardized compounds with different combinations was carried out based on the analysis of the possible precursors produced by SEM derived from the experiment. The results are shown in Table 3. A certain amount of SEM was detected in the arginine + H2O2, arginine + urea + H2O2, arginine + citrulline + H2O2, arginine + urea + ornithine + H2O2, arginine + urea + citrulline + H2O2, arginine + ornithine + citrulline + H2O2, and arginine + urea + ornithine + citrulline + H2O2. Arginine and H2O2 were found

Figure 4. RDA between amino acid and SEM in the whole growth cycle.

the decomposition of arginine. Given that arginine enzymes were important substances involved in the urea cycle of shrimps, the production of SEM may be related to the urea cycle. 3.5. Determination of the Main Compounds in the Urea Cycle. The contents of citrulline, ornithine, and urea in the shrimp shells were detected using HPLC and HPLC−MS/ MS in the entire growth cycle of L. vannamei (data not shown). In combination with the changes in SEM and arginine content in shrimp shells, the Spearman correlation analysis and RDA were performed on the whole data with SPSS Statistics and CANOCO software. In Spearman correlation analysis (see Figure 5A), the contents of arginine and citrulline showed a strong correlation with the changes in the SEM content. The correlation coefficients were greater than 0.9. In RDA (see Figure 5B), the contents of citrulline, arginine, and urea showed a strong correlation with the changes in the SEM content. The results suggested that the changes in the SEM content in the total growth cycle of L. vannamei might have a certain correlation with the main substances in the urea cycle. 3.6. Effects of Ammonia Nitrogen in the Water Environment on the Formation of SEM in L. vannamei. As shown in Table 2, the shrimp survived 24 h in a highammonia-nitrogen environment. The proportion of urea

Figure 5. (A) Spearman correlation analysis and (B) RDA of the changes in the SEM content with the main substances in the urea cycle. E

DOI: 10.1021/acs.jafc.9b01779 J. Agric. Food Chem. XXXX, XXX, XXX−XXX

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Journal of Agricultural and Food Chemistry Table 2. Water Environmental Test of High Ammonia Nitrogen SEM (ng/g) salinity salinity salinity salinity

of of of of

18 18 + 1.5 mmol/L ammonia nitrogen 35 35 + 1.5 mmol/L ammonia nitrogen

2.1 3.6 7.4 16.9

± ± ± ±

0.1 0.2 0.2 0.7

arginine (μg/g) 423.1 573.2 689.8 845.1

± ± ± ±

3.1 3.9 8.1 9.1

urea (mg/kg) 13.1 14.7 13.6 18.7

± ± ± ±

0.5 0.8 0.7 1.3

ornithine (μg/g) 482.4 511.2 425.7 492.6

± ± ± ±

7.1 8.7 7.6 3.1

citrulline (μg/g) 609.5 632.3 500.9 511.6

± ± ± ±

9.3 8.1 6.3 6.1

Figure 6. Possible biosynthetic pathway of endogenous SEM in L. vannamei.

detection. Arginine contains guanidine, which is easy to hydrolyze into urea under acidic conditions (see Figure 7). Urea then reacts with the amidine group of arginine to form SEM. The standardized reaction results shown above verified the speculation of the biosynthetic pathway of endogenous SEM in L. vannamei and further indicated the relationship between the production of endogenous SEM and the main substance of the urea cycle in L. vannamei. In conclusion, in the case of the water environment and feedstuff monitoring, the addition of the original antibiotic was not detected. However, SEM was still detected in L. vannamei, and the production of endogenous SEM was demonstrated. Clearly, the validity of SEM as a marker of nitrofurazone use is further scrutinized. At the same time, the detection of SEM in different parts of shrimps proved that endogenous SEM existed only in the shrimp shell. This indicates the irrationality of using the whole shrimp as a research object in the quarantine detection of SEM. While the origins of SEM in shrimps remain unclear, this study provides further evidence that it can occur in shrimps for reasons unrelated to nitrofurans. Through the detection of SEM and amino acid content changes in the whole growth cycle, arginine may be the potential precursor of endogenous SEM. In combination with the analysis of the changes in the main substances involved in the urea cycle, the formation of endogenous SEM is closely related to the amidine group of arginine and amide structure of citrulline and urea. This

Table 3. SEM Content of Standardized Compounds with Different Combinations sample

content of SEM (μg/L)

arginine + urea + ornithine + citrulline arginine + H2O2 urea + H2O2 ornithine + H2O2 citrulline + H2O2 arginine + urea + H2O2 arginine + ornithine + H2O2 arginine + citrulline + H2O2 urea + ornithine + H2O2 urea + citrulline + H2O2 ornithine + citrulline + H2O2 arginine + urea + ornithine + H2O2 arginine + urea + citrulline + H2O2 arginine + ornithine + citrulline + H2O2 urea + ornithine + citrulline + H2O2 arginine + urea + ornithine + citrulline + H2O2

NDa 0.18 ± 0.01 ND ND ND 3.88 ± 0.02 ND 3.47 ± 0.01 ND ND ND 3.43 ± 0.01 3.49 ± 0.03 3.38 ± 0.02 ND 3.36 ± 0.02

a

ND = not detected.

in each detection group. At the same time, urea and/or citrulline were present in all of the detection groups, except the group of arginine + H2O2. SEM was also detected in the group of arginine + H2O2; this may be due to the need for hydrochloric acid hydrolysis in the pretreatment of SEM F

DOI: 10.1021/acs.jafc.9b01779 J. Agric. Food Chem. XXXX, XXX, XXX−XXX

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Figure 7. Schematic diagram of acid hydrolysis of arginine. (6) Hoenicke, K.; Gatermann, R.; Hartig, L.; Mandix, M.; Otte, S. Formation of semicarbazide (SEM) in food by hypochlorite treatment: Is SEM a specific marker for nitrofurazone abuse? Food Addit. Contam. 2004, 21, 526−537. (7) Park, M. S.; Kim, K. T.; Kang, J. S. Development of an analytical method for detecting nitrofurans in bee pollen by liquid chromatography-electrospray ionization tandem mass spectrometry. J. Chromatogr. B: Anal. Technol. Biomed. Life Sci. 2017, 1046, 172− 176. (8) Ye, J.; Wang, X. H.; Sang, Y. X.; Liu, Q. Assessment of the determination of azodicarbonamide and its decomposition product semicarbazide: Investigation of variation in flour and flour products. J. Agric. Food Chem. 2011, 59, 9313−9318. (9) Crews, C. Potential natural sources of semicarbazide in honey. J. Apic. Res. 2014, 53, 129−140. (10) Saari, L.; Peltonen, K. Novel source of semicarbazide: Levels of semicarbazide in cooked crayfish samples determined by LC/MS/MS. Food Addit. Contam. 2004, 21, 825−832. (11) Van Poucke, C.; Detavernier, C.; Wille, M.; Kwakman, J.; Sorgeloos, P.; Van Peteghem, C. Investigation into the possible natural occurence of semicarbazide in Macrobrachium rosenbergii prawns. J. Agric. Food Chem. 2011, 59, 2107−2112. (12) Kwon, J.-W. Semicarbazide (SEM): Natural occurrence and uncertain evidence for its formation from food processing. Food Control 2017, 72, 268−275. (13) Wang, D. N.; Zhou, F.; Li, S. Y.; Xu, W. G.; Wang, Y. Background value survey and source analysis of semicarbazide in shellfish. Chin. Fish. Qual. Stand. 2016, 6, 6−11. (14) Yu, H. J.; Li, B.; Cai, Y. Q.; Ye, F. T.; Tai, J. M.; Hui, Y. H.; Xu, J.; Feng, B. Determination of Semicarbazide Content in Crustaceans by Liquid Chromatography Tandem Mass Spectrometry. Fenxi Huaxue 2012, 40, 1530−1535. (15) Noonan, G. O.; Begley, T. H.; Diachenko, G. W. Semicarbazide Formation in Flour and Bread. J. Agric. Food Chem. 2008, 56, 2064− 2067. (16) Huong, D. T. T.; Yang, W.-J.; Okuno, A.; Wilder, M. N. Changes in free amino acids in the hemolymph of giant freshwater prawn Macrobrachium rosenbergii exposed to varying salinities: Relationship to osmoregulatory ability. Comp. Biochem. Physiol., Part A: Mol. Integr. Physiol. 2001, 128, 317−326. (17) Lee, D. L.; Her, B. Y.; Cheng, S. J.; Liao, I. C. Seasonal variations of extractive components of wild grass prawn Penaeus monodon in Tungkang Taiwan area. J. Fish. Soc. Taiwan 1989, 16, 281−292. (18) Greenway, P. Nitrogenous excretion in aquatic and terrestrial Crustacea. Mem. Queensl. Mus. 1991, 31, 215−227. (19) Abernethy, G. A. Generation of semicarbazide from natural azine development in foods, followed by reaction with urea compounds. Food Addit. Contam., Part A 2015, 32, 1416−1430. (20) Liu, C. H.; Chen, J. C. Effect of ammonia on the immune response of white shrimp Litopenaeus vannamei and its susceptibility to vibrio alginolyticus. Fish Shellfish Immunol. 2004, 16, 321−334. (21) Chen, J. C.; Cheng, S. Y. Hemolymph oxygen content, oxyhemocyanin, protein levels and ammonia excretion in the shrimp Penaeus monodon exposed to ambient nitrite. J. Comp. Physiol., B 1995, 164, 530−535. (22) Liu, X. Y. The peroxisome in the tissue cell of Penaeus chinensis. Mar. Sci. 2003, 27, 43−46.

research proposes that SEM may be generated from arginine and citrulline and urea via an oxaziridine intermediate. This study also provides a new theory for future research in the field of endogenous SEM.



ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jafc.9b01779. Chromatograms for the detection of SEM in standards, water environment, and feedstuff of nitrofurazone detection (Figure S1), chromatograms for the detection of SEM in the water environment and feedstuff (Figure S2), chromatograms for the detection of SEM in L. vannamei (Figure S3), content of SEM in shrimp shell and gonad of maternal shrimp (Table S1), and SEM content of each part of L. vannamei (Table S2) (PDF)



AUTHOR INFORMATION

Corresponding Author

*Telephone: 86-312-7528195. E-mail: wangshipin2017@163. com. ORCID

Xianghong Wang: 0000-0002-0833-334X Funding

This work was supported by the National Key R&D Program of China, the Ministry of Science and Technology of the People’s Republic of China (2016YFD0401101) and Beijing Advanced Innovation Center for Food Nutrition and Human Health, Beijing Technology and Business University (BTBU) (20181014). Notes

The authors declare no competing financial interest.



REFERENCES

(1) Fernando, R.; Munasinghe, D.; Gunasena, A.; Abeynayake. Determination of nitrofuran metabolites in shrimp muscle by liquid chromatography-photo diode array detection. Food Control 2017, 72, 300−305. (2) European Commission.. Commission Regulation 37/2010 of 22 December 2009 on pharmacologically active substances and their classification regarding maximum residue limits in foodstuffs of animal origin. Off. J. Eur. Union 2010, 53, 1. (3) Vass, M.; Hruska, K.; Franek, M. Nitrofuran antibiotics: A review on the application, prohibition and residual analysis. Vet. Med. 2008, 53, 469−500. (4) Radovnikovic, A.; Moloney, M.; Byrne, P.; Danaher, M. Detection of banned nitrofuran metabolites in animal plasma samples using uhplc-ms/ms. J. Chromatogr. B: Anal. Technol. Biomed. Life Sci. 2011, 879, 159−166. (5) Maranghi, F.; Tassinari, R.; Lagatta, V.; Moracci, G.; Macri, C.; Eusepi, A.; Di Virgilio, A.; Scattoni, M. L.; Calamandrei, G. Effects of the food contaminant semicarbazide following oral administration in juvenile Sprague-Dawley rats. Food Chem. Toxicol. 2009, 47, 472− 479. G

DOI: 10.1021/acs.jafc.9b01779 J. Agric. Food Chem. XXXX, XXX, XXX−XXX

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Journal of Agricultural and Food Chemistry (23) Lee, W. C.; Chen, J. C. Hemolymph ammonia, urea and uric acid levels and nitrogenous excretion of Marsupenaeus japonicus at different salinity levels. J. Exp. Mar. Biol. Ecol. 2003, 288, 39−49. (24) Chen, J.-C.; Chen, K.-W. Hemolymph oxyhemocyanin, protein levels, acid−base balance, and ammonia and urea excretions of Penaeus japonicus exposed to saponin at different salinity levels. Aquat. Toxicol. 1996, 36, 115−128. (25) Shinji, J.; Wilder, M. N. Dynamics of free amino acids in the hemolymph of pacific whiteleg shrimp Litopenaeus vannamei exposed to different types of stress. Fish. Sci. 2012, 78, 1187−1194. (26) Mottier, P.; Khong, S.-P.; Gremaud, E.; Richoz, J.; Delatour, T.; Goldmann, T.; Guy, P. A. Quantitative determination of four nitrofuran metabolites in meat by isotope dilution liquid chromatography−electrospray ionisation−tandem mass spectrometry. J. Chromatogr. A 2005, 1067, 85−91. (27) Wilasinee, D.; Sutthivaiyakit, P.; Sutthivaiyakit, S. Determination of nitrofurans in chicken feed by high-performance liquid chromatography−tandem mass spectrometry. Anal. Lett. 2015, 48, 1979−1987. (28) Zhou, H.; Hao, N.; Yan, M.; Xu, L. Determination of Lcitrulline and L-ornithine by high performance liquid chromatography with precolumn derivation. J. Nanjing Univ. Technol. 2009, 217, 229− 232. (29) Wang, Z. X.; Jiang, J.; Sun, L.; Zhou, H. B.; Zhao, Y. X. Determination of Urea, Biruet and Dicyandiamide in Foods by High Performance Liquid Chromatography−Tandem Mass Spectrometry. J. Instrum. Anal. 2012, 31, 593−599. (30) Chen, H. W.; Lin, H. C.; Chuang, Y. H.; Sun, C. T.; Chen, W. Y.; Kao, C. Y. Effects of environmental factors on benthic species in a coastal wetland byredundancy analysis. Ocean Coastal Manage. 2019, 169, 37−49.

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DOI: 10.1021/acs.jafc.9b01779 J. Agric. Food Chem. XXXX, XXX, XXX−XXX