How to trace ancient root systems and fossil rhizospheres? A method comparison between base hydrolysis and thermally assisted hydrolysis and methylation

• We compared base hydrolysis and pyrolysis to trace fossil root-derived OM. • Both approaches revealed the presence of suberin monomers in calcified roots. • Pyrolysis failed to detect suberin monomers in rhizosphere and root-free sediments. • In OM-poor and fossil systems, only base hydrolysis enables tracing of root remains. Hydrolyzable lipids including suberin-derived monomers have been used to identify and quantify input of root-derived organic matter (OM) in soils. The two most common analytical methods are base hydrolysis and thermally assisted hydrolysis and methylation (THM), each followed by gas chromatography and detection via mass spectrometry. We compared both approaches in ancient root systems to test their suitability for tracing root remains in subsoils and geological settings. We sampled a fossil rhizolith (calcified root from unknown source vegetation) from a Central European loess-paleosol sequence together with rhizosphere loess, as well as root-free loess. After removal of free lipids, hydrolyzable lipids were released in one sample aliquot via base hydrolysis, and in another sample aliquot similar lipids were investigated via online THM. For verification, also a younger (sub-recent) rhizolith and respective rhizosphere, attributed to the recent oak vegetation, were sampled in a Southeast European soil profile developed on fluvial sand. Both methods were able to reveal the presence of suberin monomers in rhizoliths, however in case of the fossil sample set THM failed to detect suberin monomers in rhizosphere and root-free materials. Based on base hydrolysis results, we could trace root-derived OM in rhizospheres both of the sub-recent and the fossil root. In both sample sets, ω-hydroxy carboxylic acids were found in root and rhizosphere but were absent or occurred only in very low amounts in root-free sediments, and a trend of decreasing chain length of ω-hydroxy carboxylic acids from root towards root-free sediment was observed. We conclude that THM is a suitable method to rapidly detect and quantify root input in case of living and recently deceased roots and has the advantage that also lignin-derived monomers are detected. However, in OM-poor and fossil samples, only base hydrolysis provides useful insights into bound lipids and specifically root-derived OM composition for potential source apportionment.