Determination of 32 Low Molecular Mass Organic Acids in Biomass by Ion Chromatography Mass Spectrometry

Importance of the Topic

LMMOAs formed during biomass pretreatment are potent inhibitors of microbial fermentation, limiting the yield and efficiency of biofuel production. Accurate profiling of these acids is critical for optimizing conversion processes, improving strain performance and ensuring process reliability.

Objectives and Study Overview

This study aimed to develop a single-run IC/MS method capable of separating and quantifying over 30 low molecular mass organic acids in complex biomass hydrolysates. The goal was to achieve high selectivity and sensitivity, even for closely eluting compounds, to support biofuel research and quality control.

Methodology

An anion exchange separation used a hydroxide gradient (1–60 mM KOH over 38 min) on an AS11-HC column with AG11-HC guard. Suppressed conductivity detection provided broad coverage, while negative-mode ESI-MS in Selected Ion Monitoring mode enabled specific detection of individual acids. Internal standards and calibration curves supported quantitation from sub-ppm to hundreds of ppm levels.

Used Instrumentation

• Ion Chromatograph: Thermo Scientific Dionex ICS-2000
• Analytical Column: IonPac AS11-HC with AG11-HC guard (2 mm)
• Eluent Generation: EGC II KOH module
• Conductivity Detector: ASRS 300 suppressor (2 mm), deionized water regenerant
• Mass Spectrometer: Thermo Scientific MSQ Plus, negative ESI, SIM mode, nitrogen nebulizer at 80 psi, 450 °C probe, 3000 V needle

Main Results and Discussion

The method successfully resolved and quantified 32 organic acids, including formate, acetate, lactate, and TCA cycle intermediates, with retention times spanning 6.9 to 36.7 min and m/z values from 45.0 to 209.0. Concentration ranges from 0.14 ppm to over 200 ppm were measured with good repeatability. MS-SIM overcame co-elution challenges, while conductivity detection offered complementary screening.

Benefits and Practical Applications

The IC/MS approach provides a robust analytical platform for fermentation monitoring, allowing rapid profiling of inhibitory acids in real-world samples. It aids process development, troubleshooting and quality assurance in biofuel and bioprocess industries.

Future Trends and Potential Applications

Integration with online sampling and automated data analysis could enable real-time monitoring of fermentation streams. Coupling with high-resolution MS or miniaturized IC systems may further enhance sensitivity and throughput. The approach can be adapted to other biorefining processes and environmental matrices.

Conclusion

The developed IC/MS method offers high selectivity, sensitivity and throughput for comprehensive profiling of low molecular mass organic acids in biomass. Its application supports process optimization and quality control in biofuel production without extensive sample preparation.