Florfenicol in Nile tilapia (Oreochromis niloticus) : an understanding of the metabolic profile in tropical waters

Author(s)

Anna Paula Rocha de Queiroga

• Document type: Doctoral Thesis
• Press: Campinas, SP.
• Institution: Universidade Estadual de Campinas (UNICAMP). Instituto de Química
• Defense date: 2025-03-14

Examining board members

Susanne Rath; Jonas Augusto Rizzato Paschoal; André Tadeu Gotardo; Adriana Pavesi Arisseto Bragotto; Jarbas José Rodrigues Rohwedder

• Advisor: Susanne Rath

Abstract

The increasing need for antimicrobial use in aquaculture is directly related to the expansion of intensive farming systems, which favor the spread of diseases, compromise animal health, and negatively impact productivity. Florfenicol (FF), a widely used antimicrobial in the fisheries industry, lacks detailed studies on its metabolic profile in Nile tilapia (Oreochromis niloticus) raised in tropical and subtropical waters. Therefore, this study aimed to identify FF metabolites in Nile tilapia and evaluate their depletion after oral treatment with medicated feed containing FF. To achieve these objectives, potential FF metabolites—florfenicol amine (FFA), florfenicol alcohol (FFOH), and monochloro-florfenicol (FFCl)—were synthesized in the laboratory and used as analytical standards, with purities exceeding 94%. These products were characterized by ¹H and ¹³C nuclear magnetic resonance (NMR), high-resolution mass spectrometry, and quantitative NMR. An analytical method using two-dimensional liquid chromatography coupled with tandem mass spectrometry (LC-LC-MS/MS) was developed to quantify FF and its metabolites (FFOH, FFCl, and FFA) in Nile tilapia fillet, liver, and kidney. The method was validated and showed quantification limits (LOQ) =50 µg kg?¹. In the field trial, 120 male Nile tilapia with an average weight of 372 ± 57 g were housed in eight tanks (15 fish per tank). The water temperature was 28.1 ± 1.4 ºC. The fish were treated with FF via medicated feed at a dose of 10 mg kg?¹ body weight for 10 consecutive days. Samples of fillet, liver, and kidney were collected on days 1, 5, and 10 (during the treatment period) and on days 11, 12, 13, and 15 (corresponding to days 1, 2, 3, and 5 of the post-treatment period, respectively). The highest concentrations of FF and its metabolites were observed on day 5, with specific distribution profiles for each tissue: FF predominated in the kidney, followed by the liver and fillet, while FFOH and FFA showed higher concentrations in the liver, followed by the kidney and fillet. FFCl was identified as a minor metabolite in all tissues. During the depletion phase, FFOH was the primary and most persistent metabolite in the fillet and liver, while FFA predominated in the kidney, indicating that hepatobiliary and renal mechanisms are the main excretion pathways for FF. Using different sample preparation methods (acid hydrolysis and the QuEChERS approach), it was found that there are bound and non-extractable residues in the fillet. The results suggest that FFOH could replace the currently used marker residue (FF and the sum of metabolites expressed as FFA), simplifying monitoring methods. Unlike acid hydrolysis, which is not compatible with multi-residue approaches, FFOH allows the application of the QuEChERS methodology, facilitating quantification by LC-MS/MS. Finally, it was found that FF metabolism is faster in tilapia raised in tropical climates compared to fish from temperate regions, which may allow for a reduction in the withdrawal period (degree-day). This characteristic represents an advantage for producers, contributing to more efficient and sustainable management of antimicrobial use (AU)

• FAPESP's process: 22/08804-0 - Consequences of antimicrobial use in aquaculture on human health and the aquatic ecosystem
• Grantee: Anna Paula Rocha de Queiroga
• Support Opportunities: Scholarships in Brazil - Doctorate