Development of methods for analyzing and monitoring antibiotic and pesticide residues in fish and shrimp

Abstract

Background: Aquaculture has seen significant growth in recent years, especially in fish and shrimp production. This expansion is driven by increasing global demand for seafood, prompting substantial advancements in the fish and shrimp industries and higher production levels for both species. Bangladesh boasts an expansive wetland region that supports a wide array of aquaculture practices. The aquaculture industry contributes substantially to the economy of Bangladesh with regards to employment, foreign exchange earnings, nutrition and income. In aquaculture, antibiotics are widely used from the larval stage through to the growth stage for disease prevention and treatment purposes. The extensive use of antibiotics in aquaculture can result in residues remaining in shrimp products, which are consumed globally and may lead to various toxic effects. Similarly, pesticides are widely used in modern agriculture worldwide, albeit in varying amounts. While pesticides can boost crop yields, they pose significant environmental, health, and safety concerns for both terrestrial and aquatic ecosystems. Aquatic creatures, including fish, are exposed to pesticides and their residues through multiple pathways, such as run-off and spray drift from agricultural fields. Bangladesh exports shrimp and fish to various countries around the world. To ensure the safety and quality of fish and fish products, it is mandatory to submit a pre-export test report and health certification before shipping the consignment. The objective of the research was to develop sensitive, economical, and straightforward methods for determining and quantifying antibiotic and pesticide residues in shrimp and fish. Methods: Three methods were developed for the analysis of antibiotic and pesticide residues. In the first experiment, a modified QuEChERS technique was developed to simultaneously analyze tylosin (Tyl) and metronidazole (MNZ) residues in shrimp samples using LC-ESI-MS/MS. In the second experiment, A sensitive and effective method was developed and validated using QuEChERS extraction for detecting 19 organochlorine pesticides in tilapia fish. In the third experiment, a novel method was developed and validated for the first time to quantify three pesticide residues (chlorpyrifos, cypermethrin, and acetamiprid) in tilapia fish using QuEChERS extraction combined with GC-ECD. xi Result: The first method demonstrated excellent linearity for both analytes over the concentration range of 0.5–50 µg/L. The coefficients of determination (R²) for Tyl and MNZ were 0.9997 and 0.9998, respectively. The recovery rates were 81-85% for Tyl with a relative standard deviation (RSD) ≤ ± 4.9%, and 85-88% for MNZ with RSD ≤ ± 4.07%. The limits of quantification (LOQ) were determined to be 1 µg/kg for Tyl and 0.9 µg/kg for MNZ. The limits of detection (LOD) were found to be 0.4 µg/kg for Tyl and 0.3 µg/kg for MNZ. The second method employed linearity, accuracy, precision, limit of detection (LOD), and limit of quantification (LOQ) to validate the procedure. The recoveries of the 19 organochlorine pesticides ranged from 81.03% to 102.02%, with relative standard deviations (RSD) ≤ 6% across three different levels of fortification. The third method demonstrated excellent linearity, accuracy, precision, limit of detection (LOD), and limit of quantification (LOQ) during method validation. The recoveries for chlorpyrifos, cypermethrin, and acetamiprid at fortification levels of 0.01, 0.02, and 0.05 mg/kg ranged from 85% to 95%, with relative standard deviations (RSDr) ≤ 4.5%. The matrix-matched calibration curve exhibited good linearity (r² ≥ 0.998). The LOD ranged from 2.9 µg/kg to 3.1 µg/kg, and the LOQ was 10.00 µg/kg for all analytes. Conclusion: To our knowledge, this is the first time in Bangladesh that the modified QuEChERS extraction method has been developed for the determination of Tyl and MNZ antibiotic residues in shrimp muscles. The second method allows for the simultaneous detection of nineteen organochlorine residues in tilapia fish. Additionally, the third method, developed for the first time in Bangladesh, utilizes GC coupled with an Electron Capture Detector (ECD) for the determination of chlorpyrifos, cypermethrin, and acetamiprid residues. These three methods can be employed for the routine analysis of antibiotic and pesticide residues in shrimp and tilapia fish, contributing to enhanced food safety standards in Bangladesh. This improvement in food safety will also bolster the country's exports.