The biopersistence of fiber materials is one of the cornerstones in estimating potential risk to human health upon inhalation. To connect epidemiological and in vivo investigations with in vitro studies, reliable and robust methods of fiber biopersistence determination and understanding of fiber dissolution mechanism are required. We investigated dissolution properties of oil treated stone wool fibers with and without sugar-based binder (SBB) at 37 °C in the liquids representing macrophages intracellular conditions (pH 4.5). Conditions varied from batch to flow of different rates. Fiber morphology and surface chemistry changes caused by dissolution were monitored with scanning electron microscopy and time-of-flight secondary ion mass spectrometry mapping. Stone wool fiber dissolution rate depends on liquid composition (presence of ligands, such as citrate), pH, reaction products transport and fibers wetting properties. The dissolution rate decreases when: 1) citrate is consumed by the reaction with the released Al cations; 2) the pH increases during a reaction in poorly buffered solutions; 3) the dissolution products are accumulated; 4) fibers are not fully wetted with the fluid. Presence of SBB has no influence on dissolution rate if fiber material was wetted prior to dissolution experiment to avoid poorly wetted fiber agglomerates formation in the synthetic lung fluids.