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This study aimed to optimize polysaccharide extraction conditions to maximize yield and glucan content, and to evaluate the biological activity of the obtained fractions in an experimental model of intestinal mucositis. Methods: Polysaccharides were extracted using a combination of hot-water extraction and ethanol precipitation, optimized by response surface methodology. Optimal conditions (121 \u00b0C for 120 min followed by 90% ethanol precipitation) yielded a crude polysaccharide fraction (Poli-GL). A subsequent freeze\u2013thaw process generated a soluble fraction (S-Poli-GL). Structural and compositional characterization was performed using enzymatic assays, monosaccharide profiling, and NMR spectroscopy. The biological effects of Poli-GL and S-Poli-GL were evaluated in a 5-fluorouracil-induced intestinal mucositis model following oral administration at doses of 30, 100, and 300 mg\/kg. Results: The optimized extraction protocol enabled efficient recovery of polysaccharides enriched in glucans. S-Poli-GL exhibited a high total glucan content, including 43.3% \u03b2-glucans and 3.45% \u03b1-glucans, along with minor amounts of galactose and mannose. Structural analysis confirmed the predominance of branched \u03b2-(1\u21923),(1\u21926)-D-glucans. While Poli-GL did not prevent mucositis development, S-Poli-GL significantly reduced the disease activity index and attenuated intestinal inflammation, indicating enhanced biological activity associated with the soluble glucan-rich fraction. Conclusions: Optimization of extraction and fractionation improves the functional properties of G. lucidum polysaccharides. The soluble glucan-enriched fraction (S-Poli-GL) demonstrated significant protective effects in intestinal mucositis, supporting its potential as a therapeutic candidate and warranting further investigation for clinical application.<\/jats:p>","DOI":"10.3390\/ph19060946","type":"journal-article","created":{"date-parts":[[2026,6,18]],"date-time":"2026-06-18T00:49:42Z","timestamp":1781743782000},"page":"946","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Optimized Extraction of Medicinal Mushroom Polysaccharides and Their Protective Effects Against 5-Fluorouracil-Induced Gastrointestinal Mucositis"],"prefix":"10.3390","volume":"19","author":[{"given":"Jean Felipe","family":"dos Santos","sequence":"first","affiliation":[{"name":"Faculdades Pequeno Pr\u00edncipe, Av. Igua\u00e7u, 333, Rebou\u00e7as, Curitiba CEP 80230-020, PR, Brazil"},{"name":"Instituto de Pesquisa Pel\u00e9 Pequeno Pr\u00edncipe, Av. Silva Jardim, 1632, \u00c1gua Verde, Curitiba CEP 80250-060, PR, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1539-225X","authenticated-orcid":false,"given":"Karien","family":"Sauruk da Silva","sequence":"additional","affiliation":[{"name":"Faculdades Pequeno Pr\u00edncipe, Av. Igua\u00e7u, 333, Rebou\u00e7as, Curitiba CEP 80230-020, PR, Brazil"},{"name":"Instituto de Pesquisa Pel\u00e9 Pequeno Pr\u00edncipe, Av. Silva Jardim, 1632, \u00c1gua Verde, Curitiba CEP 80250-060, PR, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marcello","family":"Iacomini","sequence":"additional","affiliation":[{"name":"Departamento de Bioqu\u00edmica e Biologia Molecular, Universidade Federal do Paran\u00e1, Curitiba CEP 81531-980, PR, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9094-2258","authenticated-orcid":false,"given":"Fhernanda Ribeiro","family":"Smiderle","sequence":"additional","affiliation":[{"name":"Faculdades Pequeno Pr\u00edncipe, Av. Igua\u00e7u, 333, Rebou\u00e7as, Curitiba CEP 80230-020, PR, Brazil"},{"name":"Instituto de Pesquisa Pel\u00e9 Pequeno Pr\u00edncipe, Av. Silva Jardim, 1632, \u00c1gua Verde, Curitiba CEP 80250-060, PR, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7992-510X","authenticated-orcid":false,"given":"Daniele","family":"Maria-Ferreira","sequence":"additional","affiliation":[{"name":"Faculdades Pequeno Pr\u00edncipe, Av. Igua\u00e7u, 333, Rebou\u00e7as, Curitiba CEP 80230-020, PR, Brazil"},{"name":"Instituto de Pesquisa Pel\u00e9 Pequeno Pr\u00edncipe, Av. Silva Jardim, 1632, \u00c1gua Verde, Curitiba CEP 80250-060, PR, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,6,16]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"219","DOI":"10.1080\/11263504.2012.753133","article-title":"Macrofungi as ecosystem resources: Conservation versus exploitation","volume":"147","author":"Zotti","year":"2013","journal-title":"Plant Biosyst."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"e1700434","DOI":"10.1002\/cbdv.201700434","article-title":"Comparative Study of Fatty Acids Profile in Eleven Wild Mushrooms of Boletacea and Russulaceae Families","volume":"15","author":"Dimitrijevic","year":"2018","journal-title":"Chem. 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