Combined extraction method for metabolomic and PLFA analysis of soil

Abstract

Characterization of the soil metabolome contributes to our understanding of how soil microbial community functions, central to sustainable agriculture, are impacted by agricultural management practices. The aim of this study was to refine the Bligh-Dyer 2-phase extraction method, commonly used in soil phospholipid fatty acid analysis (PLFA), to enhance soil metabolite detection and use this refined method to look at the impact of specific crop production factors on the soil metabolome. We modified the Bligh-Dyer 2-phase extraction method through the addition of the volatile salt, 0.25 M ammonium bicarbonate, to increase the number of metabolites extracted from soil. A greenhouse pot study with three soil types and two crop species was conducted, and soil metabolites extracted using this refined Bligh-Dyer 2-phase extraction, to determine the impact of soil type and crop species treatments on the soil metabolome. Numerous amino acids, carbohydrates, TCA cycle intermediates, and fatty acids as well as the potentially inhibitory compounds ferulic acid, caffeic acid, oxalic acid, and quinic acid differed significantly with treatment. At the onset of the greenhouse pot experiment the three soil types differed significantly in metabolite composition and at the end of the 6 week experiment both soil and crop had significant impacts on the soil metabolome. Substituting the volatile salt 0.25 M ammonium bicarbonate for phosphate buffer in the Bligh-Dyer 2-phase extraction did not significantly impact PLFA analysis of the three soils. The extract was also found to be useable in lipidomic analysis. We conclude that the addition of this volatile salt to the Bligh-Dyer 2-phase extraction provided an effective method for analysis of both the soil metabolome and soil PLFA.