Optimized method for sequential extraction and analysis of polycyclic aromatic hydrocarbons, anhydrosugars and lipid biomarkers in speleothem carbonate and drip waters

Abstract

Organic molecules such as n-alkyl lipids, terpenoids, anhydrosugars (AHs), and polycyclic aromatic hydrocarbons (PAHs) are found in karst systems, including cave deposits and drip waters, and provide a potential record of surface climate, ecosystem, and fire. However, it is analytically challenging to separate and measure multiple classes of organic compounds in these low-abundance matrices. Here, we present a simple, solid-phase sequential extraction method for the separation and purification of organic matter in carbonates (∼0.5 g, powder) and drip waters (300 mL, freeze-dried). This method offers efficient extraction and separation of hydrocarbons (F_H) (saturated and PAHs) (hexane/dichloromethane; 70/30; v/v) and polar lipid compounds (F_P) (e.g., AHs, sterols, and diterpenoids) (acetone) with a high recovery of standards in F_H (78 ± 7 %, on average) and F_P (90 ± 6 %, on average) across multiple matrices (i.e., speleothems and waters). Acid digestion was applied to the post-initial sequential extraction speleothem samples to fully liberate any remaining molecules trapped in the calcite crystals. We found that ∼10% of n-alkanes (C₂₀-C₃₂) and the smallest PAHs (naphthalene, methylated naphthalene, and fluorene) of total extractable hydrocarbons were left in the acid digestion, indicating that modification of the sequential extraction could be applied based on the target of interests. In total, this sequential solid-phase extraction method provides reliable separation of geochemically significant compound classes across different matrices within karst systems.