GC-MS method for determining faecal sterols as biomarkers of human and pastoral animal presence in freshwater sediments

Abstract

In order to determine sterols and stanols in freshwater sediments to reconstruct the past presence of humans and pastoral animals, we developed an analytical method based on pressurised liquid extraction (PLE), clean-up performed using solid phase extraction (SPE) and sterol determination using gas chromatography-mass spectrometry (GC-MS) analysis. PLE extraction conditions were optimised using dichloromethane (DCM) and DCM/methanol mixtures. Clean-up was performed with 2 g silica SPE cartridges, and the concentrated extracts were eluted with 70 mL DCM. Extraction yield was evaluated using an in-house reference material spiked with (13)C-labelled cholesterol and aged for 10 days. In comparison with pre-extraction, where the sediment is extracted and then spiked with a known analyte concentration, this approach preserves the original composition of the sediment. DCM and DCM/methanol mixtures resulted in high extraction yields ranging from 86 to 92 % with good reproducibility (relative standard deviation (RSD) 5-8 %). PLE extraction yields obtained with DCM as the extracting solvent were about 1.5 times higher than extractions using an ultrasonic bath. The solvent extraction mixture and matrix composition strongly affected the solvent extraction composition where higher overall recoveries (70-80 %) for each compound were obtained with DCM. The extraction mixture and matrix composition also affected the analyte concentrations, resulting in a method precision ranging from 1 to 18 %. Diatomaceous earth spiked with 10 to 100 ng of sterols, and environmental samples fortified with suitable amounts of sterols provided apparent recovery values ranging from 90 to 110 %. We applied the method to environmental samples both close to and upstream from sewage discharge zones, resulting in substantially higher faecal sterol (FeSt) concentrations near the sewage. In addition, we also applied the method to a 37-cm freshwater sediment core in order to evaluate its applicability for obtaining vertical sterol profiles.

Fire, vegetation, and Holocene climate in a southeastern Tibetan lake: a multi-biomarker reconstruction from Paru Co

“…A combination of innovative molecular markers were used to infer past fires, vegetation, and human interactions in sediment cores analysed from Guatemala (Schüpbach et al, 2015) and East Africa (Battistel et al, 2016). Using a similar approach, we use biomarkers that are produced under specific environmental conditions and then transported, accumulated, and stored in lacustrine sediments: monosaccharide anhydrides (MAs), fecal sterols and stanols (FeSts), polycyclic aromatic hydrocarbons (PAHs), and normal n-alkanes.…”

“…Using a similar approach, we use biomarkers that are produced under specific environmental conditions and then transported, accumulated, and stored in lacustrine sediments: monosaccharide anhydrides (MAs), fecal sterols and stanols (FeSts), polycyclic aromatic hydrocarbons (PAHs), and normal n-alkanes. Significant concentrations of these compounds are present in soil and sedimentary archives with ages older than 10 cal kyr BP (D'Anjou et al, 2012;Johnsen et al, 2005;, suggesting that degradation, if happening, is a low-kinetic process (Battistel et al, 2016) and that these compounds resist over the Holocene or longer timescales. Within the listed biomarkers, MAs are specific tracers of vegetation combustion (Simoneit, 2002;Zangrando et al, 2013).…”

“…Cellulose pyrolysis creates the molecular marker levoglucosan (1,6-anhydro-β-D-glucopyranose; Simoneit et al, 1999), while hemicellulose combustion produces the isomers mannosan (1,6-anhydro-β-D-mannopyranose) and galactosan (1,6-anhydro-β-D-galactopyranose) (Kuo et al, 2011). Several studies examine levoglucosan (L), mannosan (M), and galactosan (G) in aerosols and ice cores (Kehrwald, 2012;Simoneit, 2002;Yao et al, 2013;Zennaro et al, 2014;Zhang et al, 2008), as well as in sediment cores (Battistel et al, 2016;Kirchgeorg et al, 2014;, demonstrating the suitability of MAs as paleofire proxies. PAHs are a wide group of organic compounds made up of two or more benzene rings combined together in linear, angular, or clustered arrangements (Zakir Hossain et al, 2013).…”

Characterization of ancient lipids in prehistoric organic residues: Chemical evidence of livestock‐pens in rock‐shelters since early neolithic to bronze age

“…The extraction of these compounds from fecal samples has been commonly carried out using Soxhlet as extraction technique (24-36 h/sample) [19,[23][24][25]. However, alternative extraction techniques such as ultrasound-assisted solidliquid extraction (20-80 min) [26][27][28][29] or PLE [30,31] have been used to accelerate the extraction time and reduce the extraction solvent. Commonly, different mixtures of dichloromethane/methanol (DCM/MeOH = 1:1, 2:1, 3:1, or 9:1) and volumes (20-200 mL) [24,25,32,33] have been used for the total lipid extraction.…”

“…For that propose, a saponification step is commonly used [18,34] before LLE and a clean-up step (SPE) to eliminate interferences and separate the analytes. Sterols and bile acids can be separated by the LLE [24,33] or in the SPE [24,30,[35][36][37] and they are analyzed separately by using GC-MS after a derivatization step.…”