In Vitro Digestion of Pleurotus ostreatus Stem Flour and Extract: Release of Free Sugars and Organic Acids

Mushroom stems are rich in bioactive macromolecules, although their digestive fate remains poorly understood. To move from waste disposal to systematic valorization, it is essential to understand how different matrices influence metabolite bioaccessibility. This study evaluated the digestive transformation and release kinetics of free sugars and organic acids from Pleurotus ostreatus stems (POS) to assess their potential as functional ingredients. POS were processed into two forms: whole flour (POS-F) and an ultrasound-assisted water-soluble extract (POS-E). Both matrices were subjected to the standardized INFOGEST 2.0 in vitro digestion model. Oral, gastric, and intestinal phases were monitored kinetically. Following intestinal digestion, dialysis (3.5 kDa) separated the serum-available permeate (P) from the colon-available retentate (R). Free sugars and organic acids were identified and quantified throughout digestion by HPLC-RI/UV. Digestion promoted progressive metabolite release. Trehalose, the major sugar, increased from the oral to intestinal phases in both POS-F (7.96–12.13 mg/100 g) and POS-E (5.11–11.40 mg/100 g). Glucose and sorbitol also accumulated, reaching maximum levels in the intestinal phase. Sucrose was detected only in POS-F and remained stable during digestion (~0.7 mg/100 g in the retentate). Organic acid profiles changed markedly, with succinic and isobutyric acids increasing in the intestinal phase, especially in POS-F (10.81 and 6.45 mg/100 g, respectively). Dialysis revealed high bioaccessibility of lactic acid, reaching 9.58 mg/100 g in the POS-E permeate. In contrast, a substantial fraction of total sugars remained in the retentate (9.66 mg/100 g in POS-F and 5.11 mg/100 g in POS-E), indicating potential colonic delivery.These findings demonstrate that matrix structure governs metabolite release and bioaccessibility during digestion, supporting the use of POS as value-added functional ingredients and providing a basis for targeted food applications.