Membrane bioreactors (MBRs) are increasingly employed in municipal wastewater treatment, with membranes inevitably reaching their end-of-life (EOL) primarily due to the accumulation of irrecoverable fouling. Nevertheless, the composition and properties of irrecoverable fouling remain ambiguous due to the lack of appropriate extraction and analytical methods. Herein, we developed a temperature-regulated organic solvent extraction method to extract and analyze irrecoverable foulants in EOL ultrafiltration membranes at various locations within a large-scale MBR for municipal wastewater treatment. The irrecoverable foulants encompassed a high molecular weight fraction not observed in the influent, potentially induced by aggregation from organic-inorganic foulant interaction. The terrestrial humic acid (HA)-like substance dominated the organic components of irrecoverable foulants, and the chelating cation Ca emerged as a dominant inorganic compound. Unexpectedly, we noticed the enrichment of Al, Fe, and Si in the irrecoverable fouling, despite their relatively low rejections by the EOL membrane. The potential complex interactions among HA-like substances, Ca, Al, Fe, and Si were revealed by correlation and cluster analysis. The strong bridging potentials of [Al(OH)4]-, Fe(OH)3 at neutral pH, coupled with the colloidal property of Si, may facilitate the aggregation of organic and inorganic compounds, forming complex irrecoverable foulants with strong affinity for the membrane matrix. Besides, the irrecoverable fouling exhibited an increasing trend along the water flow direction. The mystery of irrecoverable fouling of EOL membrane was, for the first time, systematically unraveled, establishing a foundation for the development of effective membrane cleaning and regeneration technology.