Analysis of Amino Acids

Significance of the Topic

Amino acid profiling plays a pivotal role in biochemistry, clinical diagnostics, food quality control, and pharmaceutical research. Reliable quantification of amino acids underpins metabolic studies, nutritional analysis, and ensures product consistency across industries.

Objectives and Study Overview

This application note demonstrates a reversed-phase liquid chromatography method for the separation and detection of key amino acids using the Shim-pack GISS C18 column. The aim is to achieve efficient resolution of acidic, neutral, and amide-containing amino acids under mild conditions suitable for routine analysis.

Methodology and Used Instrumentation

The protocol employs a Shim-pack GISS C18 column (250 mm × 4.6 mm I.D., 5 μm) maintained at 40 °C. The mobile phase consists of:

• A: 5 mM sodium 1-octanesulfonate in 0.1 % phosphoric acid aqueous solution
• B: Acetonitrile

An isocratic ratio of A/B = 95/5 (v/v) is delivered at 1.0 mL/min. A 10 µL sample injection is monitored by UV absorbance at 210 nm. Key instrumentation details include the HPLC system coupled to UV detection and temperature-controlled column compartment.

Main Results and Discussion

The method efficiently separated four representative amino acids— aspartic acid, glutamine, threonine, and proline—within a single analytical run. The ion-pairing agent sodium 1-octanesulfonate enhanced retention of polar analytes, delivering sharp peak shapes and reproducible retention times. UV detection at 210 nm provided adequate sensitivity for routine concentration ranges.

Benefits and Practical Applications

The described protocol offers:

• Robust separation of acidic and neutral amino acids without derivatization
• Short analysis time compatible with high-throughput laboratories
• Reproducible performance suited for QA/QC and research workflows

This approach is directly applicable to food composition analysis, clinical sample screening, and formulation studies in pharmaceutical development.

Future Trends and Potential Applications

Advances in ultra-high-performance liquid chromatography (UHPLC) and novel stationary phases may further reduce analysis time and solvent consumption. Coupling with mass spectrometry would expand detection capabilities to trace-level quantification and complex sample matrices. Automated sample preparation and data processing are expected to drive higher throughput in industrial and clinical settings.

Conclusion

The Shim-pack GISS C18 reversed-phase method described here provides a straightforward, reliable route for routine amino acid analysis. Its ease of implementation and robust performance make it a valuable tool for diverse applications in analytical chemistry.

Reference

Shimadzu Corporation. Analysis of Amino Acids Using Shim-pack GISS C18 Column. First Edition, December 2021.