High Throughput simultaneous analysis of 39 amino acids in various foods without derivatization using LC-MS/MS

Importance of the topic

Amino acids are fundamental for nutritional quality and organoleptic properties of food. Rapid and comprehensive profiling of free amino acids supports nutritional research, quality control and flavor development in the food industry. Conventional LC‐MS/MS analysis often requires derivatization or ion‐pair reagents, limiting throughput and introducing complexity. A streamlined approach for simultaneous quantification of 39 amino acids without derivatization enables higher productivity and broader applicability.

Study objectives and overview

This study expands on previous work by developing a high-throughput method to detect 39 amino acids in diverse food matrices by LC-MS/MS without derivatization. Key aims include:

• Optimizing chromatographic separation to resolve structural isomers
• Establishing sensitive MRM transitions for each analyte
• Validating linearity, precision and recovery across complex samples

Methodology and instrumentation

Liquid chromatography employed an Imtakt Intrada Amino Acid mixed-mode column (3.0×50 mm, 3 µm), with a binary gradient of acetonitrile/formic acid and ammonium formate. A Shimadzu Nexera UHPLC system was coupled to an LCMS-8050 triple-quadrupole with electrospray ionization. Ultra-Fast Polarity Switching (5 ms) at 555 channels/sec enabled simultaneous positive/negative MRM acquisition.

Main results and discussion

The method achieved baseline separation of 39 amino acids, including critical isomer pairs (e.g., alanine/sarcosine). Calibration curves displayed excellent linearity (r2 > 0.97) over wide concentration ranges. Repeatability was robust, with RSD values generally below 10%. Sample analyses demonstrated applicability to bonito extract, pork muscle and wine, revealing characteristic amino acid profiles such as high carnosine in meat and elevated proline and GABA in wine. Spike recovery experiments yielded acceptable recoveries (70–120%).

Benefits and practical applications

• Elimination of derivatization simplifies workflow and reduces analysis time
• Mixed-mode separation enhances isomer resolution without ion-pair agents
• Simultaneous polarity switching maximizes coverage of both acidic and basic amino acids
• Applicable to routine quality control, nutritional profiling and research

Future trends and possibilities for use

The approach can be extended to plant-based and fermentation-derived products, enabling large-scale nutritional studies. Integration with automated sample preparation and high-resolution MS may further enhance throughput and identify novel bioactive peptides and metabolites.

Conclusion

This LC-MS/MS method provides a rapid, sensitive and comprehensive platform for amino acid analysis in complex food matrices without derivatization. Its robustness and high throughput support diverse applications in food science and quality assurance.

Reference

• Imtakt Technical Report T1734E, Imtakt Corporation, Kyoto.
• Matsumoto K, et al. Poster TP510, ASMS 2014, Baltimore.