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United States Department of Agriculture

Agricultural Research Service

Research Project: NOVEL TECHNOLOGIES AND TECHNIQUES FOR THE DETECTION OF RESIDUES, TOXINS, AND OTHER CHEMICALS IN FOODS

Location: Residue Chemistry and Predictive Microbiology

Title: Determination of oxytetracycline residue in shrimp using a portable time-resolved analyzer and HPLC-MS/MS validation

Author
item CHEN, GUOYING

Submitted to: Sensing and Instrumentation for Food Quality and Safety
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: February 13, 2012
Publication Date: February 29, 2012
Citation: Chen, G. 2012. Determination of oxytetracycline residue in shrimp using a portable time-resolved analyzer and HPLC-MS/MS validation. Sensing and Instrumentation for Food Quality and Safety. 5:165-171.

Interpretive Summary: In the last two decades, shrimp farming has expanded rapidly in the U.S. and abroad. Oxytetracycline (OTC) is the most prominent antibiotic in shrimp aquaculture. In this work, OTC residue in shrimp muscle is determined using a portable analyzer built in this laboratory based on time-resolved luminescence (TRL). First, OTC is extracted and cleaned up. Then reagents are added to allow sensitive and specific OTC detection. The long lasting luminescence signal is then detected. A brief time delay effectively rejects stray light from the light source as well as interfering fluorescence from matrix components that coexist with OTC. The signal intensity depends linearly on OTC concentration in the 10-10,000 parts-per-billion range. The method reproducibility is 90% and the limit of detection is 4.6 parts per billion. This method achieved a low background that corresponds to <20 parts per billion for shrimp of seven different origins. Shrimp producers can monitor OTC in shrimp on site during medication, regulatory agencies will benefit from simplified assay protocol without slow chromatographic separation, and consumers will benefit from safe OTC-free seafood.

Technical Abstract: Oxytetracycline (OTC) is the most prominent antibiotic in shrimp aquaculture. In this work, OTC residue in shrimp muscle is determined using a portable analyzer built in this laboratory based on europium-sensitized luminescence (ESL). First, OTC is extracted in McIlvaine buffer at pH 4.0 with 0.1 M ethylenediaminetetraacetic acid and cleaned up using Oasis HLB cartridges. To the eluate, Eu(III) is added to form a Eu:OTC chelate at pH 8.5. In a micellar environment, when OTC is excited by 385 nm pulses from a light emitting diode (LED), the absorbed energy is transferred efficiently to Eu(III) which emits luminescence at 614 nm. This long lasting ESL signal is detected by a gated photomultiplier tube and integrated over a 25-1000 us interval. The 25-us delay effectively rejects stray light from the light source as well as fluorescence and scattering from matrix components that coelute with OTC. The signal intensity depends linearly (r2 = 0.9977) on the OTC concentration over >3 orders of magnitude (10-10000 ng/g). The average relative standard deviation is 3.75% and the limit of detection is 4.6 ng/g. Average background corresponds to 10.4 ng/g with 18.0% relative standard deviation for seven shrimp samples of different origins. This combination of a portable analyzer and a simplified protocol provides high specificity for detection without chromatographic separation, high sensitivity for trace-level determination, and the possibility for field deployment.

Last Modified: 9/10/2014
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