Isolation of chloramphenicol from whole eggs by supercritical fluid extraction with in-line collection

Egg consumption, at more than 65 billion per year in the United States, represents a potentially significant source of exposure to drug residues, particularly if the laying hens are treated with antimicrobial compounds or fed a diet containing medicated feed. Residues resulting from the use of chloramphenicol (CAP) is especially problematic if this compound is not used in accordance with national registration, e.g., for the control of Salmonella microorganisms in poultry. The most commonly used methods for the determination of CAP in biological samples require the use of large amounts of organic solvent. As a result, a less solvent intensive supercritical fluid extraction (SFE) method was developed for CAP in whole chicken eggs, and the results were compared with those for a solvent extraction procedure. In the SFE method, the egg sample is extracted with supercritical CO2 (without a modifier) at 10,000 psi (680 bar), 80 degrees C, and an expanded gas flow rate of 3.0 L/min to a total volume of 150 L. The CAP is trapped in-line on a Florisil sorbent bed. The CAP is eluted post-SFE by using the liquid chromatographic mobile phase solvent (water-methanol), and determined on a C8 column with ultraviolet detection at 280 nm. Recovery from eggs fortified at the 10 ppb level (n = 6) was 81.2 +/- 4.3%. To obtain eggs containing incurred CAP, hens were given a single daily dose of 75 mg CAP (orally by gelatin capsule) for 2 consecutive days, and the eggs were collected over a 12-day period. The mean value for "normally incurred" CAP in the eggs (n = 17) analyzed by SFE ranged from none detected to 174.5 ppb, with an overall mean of 60.5 ppb, compared with a mean of 60.4 ppb for the solvent extraction method. No significant difference in results was found between methods. However, the SFE method is more rapid, uses less solvent, and gives recoveries similar to those for the solvent extraction method, making it ideal for regulatory monitoring.