LCMS-IT-TOF Analysis of Amoxicillin

Importance of the Topic

Analytical methods for antibiotic detection play a critical role in food safety and public health due to concerns about drug residues and antibiotic resistance. The ability to confirm the presence and structure of ß-lactam antibiotics such as amoxicillin in animal products demands robust techniques combining high resolution and accurate mass measurements.

Objectives and Study Overview

This study demonstrates the application of the Shimadzu LCMS-IT-TOF for detailed analysis of amoxicillin. The goals include achieving high mass accuracy for parent and fragment ions up to MS10, showcasing MSn capabilities, and comparing performance to other TOF-based instruments.

Methodology and Instrumentation

• Liquid Chromatography
  • Shimadzu Prominence Series LC with dual pumps and autosampler
  • Shimpack VP-ODS column (4.6 x 150 mm, 4.6 µm)
  • Gradient elution from 0 to 50% acetonitrile over 25 minutes
  • Flow rate 0.3 mL/min, injection volume 0.5 µL
• Mass Spectrometry
  • Shimadzu LCMS-IT-TOF with ion trap and time-of-flight analyzer
  • Compressed ion injection for efficient ion accumulation
  • Ballistic ion extraction for simultaneous ion ejection
  • Argon cooling gas for enhanced damping and resolution
  • Fast polarity switching in 100 ms
  • ESI positive mode; scan ranges MS1 100–1000 m/z, MS2/MS3 up to 400 m/z

Main Results and Discussion

• Chromatographic separation yielded a clear total ion chromatogram and extracted ion trace of amoxicillin.
• MS, MS2 and MS3 spectra revealed characteristic cleavages such as loss of NH3 and β-lactam ring opening at m/z 160, 208 and 349.
• Mass accuracy for the protonated molecule (m/z 366.1114) and fragments was consistently below 3.5 ppm without using an internal standard.
• Comparison with reported data from other TOF systems showed comparable or superior performance of the LCMS-IT-TOF.

Benefits and Practical Application of the Method

• High resolution and accurate mass enable unambiguous structural confirmation.
• MSn capability allows detailed fragmentation analysis for screening multiple β-lactam antibiotics.
• Fast polarity switching supports comprehensive analysis in a single run.
• No external calibrants required for routine high-accuracy measurements.

Future Trends and Potential Applications

• Expansion to other antibiotic classes and complex matrices such as tissues and environmental samples.
• Integration with automated workflows for high-throughput screening in QA/QC laboratories.
• Development of targeted quantitation methods using high-resolution MS for regulatory monitoring.
• Advancements in ion trapping and extraction techniques to further boost sensitivity and resolution.

Conclusion

The Shimadzu LCMS-IT-TOF demonstrates robust performance for amoxicillin analysis, combining MSn fragmentation with high mass accuracy. Its unique ion injection, extraction and cooling strategies enable detailed structural studies and reliable screening without internal standards.

References

• Heller DN, Ngoh MA. Rapid Commun Mass Spectrom 1998;12:2031-2040
• Fagerquist CK, Lightfield AR. Rapid Commun Mass Spectrom 2003;17:660-671
• Baere SD et al. Anal Chem 2002;74:1393-1401
• Fagerquist CK et al. Anal Chem 2005;77:1473-1482
• Nägele E, Moritz R. J Am Soc Mass Spectrom 2005;16:1670-1676