Comparison of HPLC Methods for the Determination of Amino Sugars in Soil Hydrolysates

Abstract

A study on the suitability of chromatographic techniques such as high performance anion exchange chromatography (HPAEC) with fluorescence detection (FL) and pulsed amperometric detection (PAD) and reversed phase (RP) chromatography for the determination of galactosamine, glucosamine, mannosamine, and muramic acid in soil hydrolysates was carried out. The reversed phase fluorescence method was rapid, provided good validation parameters, and employed relatively inexpensive instrumentation. The HPAEC methods had slightly higher limits of quantification, 0.6–5.0 µmol L⁻¹ (HPAEC-FL) and 1.0–10.0 µmol L⁻¹ (HPAEC-PAD), compared to the reversed phase fluorescence method (0.5–5.0 µmol L⁻¹). Various sample pretreatment methods and chromatographic methods were investigated and the advantages and disadvantages of the HPLC methods are discussed.

Keywords:

• Amino sugars
• Derivatization
• HPAEC
• HPLC
• Microbial residues
• OPA
• PAD
• Soil analysis

Acknowledgments

We gratefully acknowledge Gabriele Dormann, Reinhard Langel, and Marion Wacht for their technical assistance. Caroline Indorf was funded by the German Research Foundation (DFG).

Notes

LOQ = limit of quantification, defined as a signal ten times higher than noise. Limit of detection (LOD) defined as a signal three times higher than noise.

Note: intra-day precision values are mean CV of six measurements of a 90 µmol L⁻¹ (MurN: 9 µmol L⁻¹) standard mixture analyzed on the same day; inter-day precision values are mean CV of six measurements of 90 µmol L⁻¹ (MurN: 9 µmol L⁻¹) standard mixtures analyzed on three days. SD = standard deviation.

CV = mean coefficient of variation between replicate samples (n = 3); LOQ = limit of quantification; samples < LOQ were excluded from the ANOVA.