Analysis of Chloramphenicol in Shrimp and Chicken Egg Extracts Using Triple Quadrupole LC/MS/MS

Importance of the Topic

Chloramphenicol is a broad-spectrum veterinary antibiotic prohibited in food due to genotoxic and carcinogenic concerns. Its detection at trace levels ensures compliance with regulatory limits and protects public health.

Study Objectives and Overview

This study aimed to update and validate a triple quadrupole LC-MS/MS method for quantifying chloramphenicol and its glucuronide conjugates in shrimp and chicken eggs following Japan's revised guidelines.

Methodology and Instrumentation

• Sample pretreatment: 10 g homogenized sample extracted with methanol, centrifuged, volume adjusted to 100 mL. After solvent removal, samples were hydrolyzed with beta-glucuronidase in phosphate buffer, extracted with ethyl acetate, and purified on a divinylbenzene-N-vinylpyrrolidone copolymer column.
• Chromatography and MS/MS conditions: Shim-pack HR-ODS column (150 × 2.1 mm, 3 μm) at 40 °C; mobile phase 10 mmol/L ammonium acetate/water–acetonitrile (70:30 v/v), flow 0.35 mL/min; injection 5 μL. Negative ESI, probe voltage -1.0 kV; DL 300 °C; block heater 500 °C; interface 400 °C; nebulizing gas 3 L/min; drying gas 10 L/min; heating gas 10 L/min.
• MRM transitions: Chloramphenicol m/z 321.10 > 152.25 (quantifier), 321.10 > 257.05 (qualifier); chloramphenicol glucuronide m/z 496.90 > 193.05 (quantifier), 496.90 > 113.00 (qualifier).

Main Results and Discussion

A five-point calibration curve showed excellent linearity (R2 > 0.999) from 0.1 μg/L for chloramphenicol and 0.5 μg/L for the glucuronide. The method's limit of detection in animal products was 0.005 mg/kg. Spike recovery tests at 0.1 μg/L yielded recovery rates of approximately 88% (shrimp, RSD 7.6%) and 96% (eggs, RSD 13.5%). No analytes were detected in blank samples, confirming method specificity and complete hydrolysis of conjugates.

Benefits and Practical Applications

• High sensitivity and precision enable reliable enforcement of non-detectable limits for chloramphenicol in food.
• Streamlined sample preparation and purification reduce matrix effects and false negatives.
• Applicable to regulatory laboratories, food safety monitoring, and quality control.

Future Trends and Potential Applications

• Automation of sample preparation to increase throughput.
• Extension to other antibiotic residues and metabolites in diverse matrices.
• Integration with high-resolution mass spectrometry for non-targeted screening.

Conclusion

The validated triple quadrupole LC-MS/MS method provides robust quantification of chloramphenicol and its glucuronide in shrimp and eggs at sub-microgram levels, meeting stringent regulatory requirements and ensuring food safety.

References

• Ministry of Health, Labour and Welfare Notification No. 49, 2017.
• Ministry of Health, Labour and Welfare Notification No. 499, 2005.
• Ministry of Health and Welfare Notification No. 370, 1959.
• Shimadzu Corporation. LCMS-8050 User Manual, 2019.