The main purpose of this study was to identify the inert fractions in olive mill wastewater (OMW) before and after chemical treatment by employing size distribution and structural resin fractionation procedures. Thermal acid cracking was selected as the chemical treatment process due to its superior performance in terms of organic matter removal from the OMW sample. Size fractionation of OMW before and after thermal acid cracking treatment was carried out using different molecular weight cut-off membranes ranging from >1600 nm to <1 nm. The original and chemical treated OMW was also subjected to structural fractionation into hydrophobic and hydrophilic (acid-base-neutral) structures using XAD-8, XAD-4, AG-MP-50 and Duolite A-7 resins. Fourier transform infrared (FTIR) spectroscopy and gas chromatography-mass spectrometry (GC-MS) analyses were also undertaken to support the size distribution analysis and structural assessment of the original and chemically treated OMW samples. The thermal acid cracking process resulted in 58%, 45%, 39%, 96% and 95% COD, TOC, total phenols, oil-grease and suspended solids removals, respectively. According to the mass balances that were established from the size distribution analysis before and after thermal acid cracking-, the chemically "inert" OMW fractions were identified as soluble (<2 nm) and colloidal (especially in the 5-8 nm size interval). Mass balances gathered from the structural resin fractionation analyses indicated that the recalcitrance of the OMW sample originated from its hydrophobic-neutral components. GC-MS and FTIR analyses confirmed that around 80% of the polyphenolic organic matter was composed of relatively high molecular weight compounds featuring lignin-like structures that also contributed the inert fraction of the OMW, whereas the remaining 20% was composed of relatively low molecular weight compounds including catechol and hydroxytyrosol. Size distribution and structural fractionation results also revealed a positive correlation between the recalcitrant nature and inhibitory effect of OMW. (C) 2015 Elsevier B.V. All rights reserved.