Automated Solid Phase Extraction for Efficient Extraction of Fluoroquinolone Antibiotics from Fish

The global demand for fish as a natural source of fresh animal protein, essential fats, minerals, and vitamins continues to rise with the human population. It is estimated that natural fish resources will not be sustainable, creating the need to increase the available supply of fish through aquaculture. The mechanisms for keeping fish healthy and free of disease is regulated within each country, often prompting the use of antibiotics in the water; however, the misuse of large antibiotic doses has increased attention on the impact to human health and thus the testing of aquaculture according to country-specific food safety regulations.

Fluoroquinolones are a commonly used fluorine subclass group of quinolones that are effective against gram negative bacteria. Popular for their use in aquaculture, fluoroquinolones consumed in high levels have contributed to the development of antibiotic resistance in the human population. Concerns prompted the FDA to ban the use of fluoroquinolones in food producing animals in 1997. To regulate the use and level of fluoroquinolone antibiotics in fish from imports or aquaculture within the USA, the FDA routinely inspects domestic and imported aquaculture operations and recommends a standard method for the routine testing of four frequently used fluoroquinolone antibiotics in fish.

White bass filets represented low-fat fish and trout represented high-fat fish samples. Samples were extracted using the SmartPrep® Cartridge extractor (Horizon Technology) used with Strata-XL™-C and Strata-X-C cartridges (Phenomenex, Inc.). The final extract was analyzed with a Nexera XR HPLC (Shimadzu) using a biphenyl column (Restek).

Results

Results after method development show excellent recoveries as can be seen in Table I (1).

The larger pore and particle size of the Strata-XL-C SPE cartridges in combination with the SmartPrep Extractor System efficiently automated the combined LLE sample extracts to achieve consistent %RSD values across both high and low-fat fish matrices. Using new column technology with the UHPLC system improved peak resolution for increased sensitivity at the specified 20-ppb FDA monitored level. Elution of all four fluoroquinolone peaks occurred in 7 min compared to 20 min using the original column technology. The optimized method passed all FDA criteria for % recovery, % RSD, and linearity within less chromatographic time and using current automation and consumable technology. Improving on workflow by implementing automation for the SPE process with the SmartPrep Extractor enabled objective sample handling across all samples by reducing scientist bias.

Reference

T. Hofhine et. al., "Improving Data Reliability and Laboratory Efficiency Testing of Fluoroquinolones in Fatty and Non-Fatty Fish Samples with One Method Using Automated Solid Phase Extraction and New Consumable Technologies," AN095, Horizon Technology (2014).

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