Pretreatment of anaerobic digester samples by hydrochloric acid (HCl) resulted in removal of Fe-based mineral and coordination compounds, attenuating their interferences with solution-state nuclear magnetic resonance (NMR) spectroscopic characterization of the solid phase organic matter. Substrate (influent) and digestate (effluent) samples from two full-scale anaerobic digesters, designated CD (co-digester) and SSD (sewage sludge digester), were investigated. Pretreatment of CD samples with 0.2-2.0 mol l(-1) HCl and pretreatment of SSD samples with 1.0-3.0 mol l(-1) HCl removed 96-100% and 76-80% of total Fe, respectively. Pretreatment declined overall paramagnetic characteristics of digestate samples, manifested by 50% (CD) and 70% (SSD) decrease in electron paramagnetic resonance signal intensities. As a result, meaningful solution-state H-1,C-13 heteronuclear single quantum coherence and H-1 NMR spectra of DMSO-d(6) soluble organic matter could be acquired. Sample pretreatment with the lowest concentration of HCl resulted in alteration of C:N ratios in solid phase, likely due to removal of labile organic and inorganic C- and N-containing compounds, while elevating the HCl concentration did not further change the C:N ratios. Furthermore, sample pretreatment increased the solubility of carbohydrates and proteins in DMSO-d(6), enabling the detection of NMR resonances from certain structural units of carbohydrates (e.g. anomeric O2CH) and proteins (e.g. CH alpha in amino acids). Both attenuation of the paramagnetic matrix as well as art enhanced solubility of carbohydrate and protein fractions of the samples in DMSO-d(6) solvent contributed to an improved molecular characterization of anaerobic digester samples by solution-state NMR analysis.