Separation of Organic Acids on an Agilent InfinityLab Poroshell 120 Aq-C18 Column

Significance of the Topic

Organic acids play a central role in determining the flavor, stability, nutritional value, and authenticity of food and beverage products. Reliable quantitation of these compounds supports quality control, regulatory compliance, and product development across industries such as winemaking, soft drink production, and juice processing.

Study Objectives and Overview

This application study evaluates the performance of an Agilent InfinityLab Poroshell 120 Aq-C18 column for the isocratic separation of nine common organic acids under highly aqueous mobile-phase conditions. Two additives—phosphoric acid and phosphate buffer—are compared to optimize retention, peak shape, and reproducibility.

Methodology and Instrumentation

The following instrumentation and materials were employed:

• Agilent 1290 Infinity LC system with binary pump, autosampler, thermostatted column compartment, and diode array detector
• InfinityLab Poroshell 120 Aq-C18 column (4.6 × 100 mm, 2.7 µm)
• Mobile Phase A: 97.5 % (0.1 % H₃PO₄) / 2.5 % methanol
• Mobile Phase B: 99 % 50 mM NaH₂PO₄ buffer (pH 2.5) / 1 % methanol
• Flow rate of 1.0 mL/min, column temperature 25 °C, UV detection at 210 nm, injection volume 1 µL
• Stock solutions of oxalic, tartaric, malic, lactic, acetic, maleic, citric, fumaric, and succinic acids prepared in water

Main Results and Discussion

Both mobile-phase additives achieved baseline separation of all nine acids within 4.5 minutes. Key findings include:

• Phosphate buffer at pH 2.5 significantly improved peak shapes for late-eluting acids such as citric and fumaric.
• High aqueous content enabled by the Aq-C18 phase prevented pore dewetting and maintained strong retention of polar analytes.
• Six consecutive injections under buffer conditions showed consistent retention times and peak areas, confirming excellent method reproducibility.

Benefits and Practical Applications

This method offers:

• Rapid, isocratic analysis with minimal organic solvent consumption.
• High resolution and robustness for routine quality control of food and beverage matrices.
• Direct injection of aqueous samples without peak distortion.
• Compatibility with standard UV detection and potential extension to MS detection.

Future Trends and Opportunities

Emerging directions include:

• Expansion of column chemistry to further enhance retention of highly polar analytes.
• Integration with high-resolution mass spectrometry for structural confirmation.
• Development of even greener protocols using fully aqueous or bio-based solvents.
• Application to a broader range of food matrices and complex biological samples.

Conclusion

The Agilent InfinityLab Poroshell 120 Aq-C18 column enables fast, reproducible separation of nine organic acids under highly aqueous isocratic conditions. Its specialized superficially porous particles and optimized ligands deliver strong retention and excellent peak shape for polar analytes, making it a valuable tool for food and beverage analysis.

Reference

1. Mack A. Analysis of Organic Acids on an Agilent InfinityLab Poroshell 120 HILIC-Z Column. Agilent Technologies Application Note, 2018.
2. Fu R-J, Wei T-C. Analysis of Polar Compounds Using an Agilent InfinityLab Poroshell 120 Aq-C18 Column with Improved and Reliable Performance. Agilent Technologies Application Note, 2022.