This article presents a comparative gas chromatography (GC)-mass spectrometry (MS)-based metabolomic analysis of mycelia and fruiting bodies of the medicinal mushroom Ganoderma lucidum. Three aqueous extracts-mycelia, fruiting bodies, and a mixture of them-and their sequential fractions (methanolic and ethyl acetate), prepared using an accelerated solvent extractor, were characterized by GC-MS to determine volatile organic compounds and by high-performance thin-layer chromatography to quantify ascorbic acid, a potent antioxidant. In addition, these extracts and fractions were assessed against Candida albicans and C. glabrata biofilms via the XTT reduction assay, and their antioxidant potential was evaluated. Application of chemometrics (hierarchical cluster analysis and principal component analysis) to GC data revealed variability in volatile organic compound profiles among G. lucidum extracts and fractions. The mycelial aqueous extract demonstrated higher anti-Candida activity and ascorbic acid content among all the extracts and fractions. Thus, this study illustrates the preventive effect of G. lucidum against C. albicans and C. glabrata biofilms along with its nutritional value.