Metabolic Responses of Pleurotus djamor to Lignocellulosic Substrate Gradients

Pleurotus djamor is cultivated on diverse lignocellulosic residues, but the relationship between progressive substrate formulation, mushroom productivity, and metabolomic responses remains insufficiently characterized. This study evaluated the effects of progressive replacement of barley straw with coconut fiber or coffee husk on mushroom productivity and GC–MS-based metabolomic profiles. Productivity differed markedly among substrate formulations, with intermediate inclusion levels producing the highest yield and biological efficiency. In contrast, PERMANOVA did not detect significant global differences in metabolomic composition across either substrate gradient, and no individual metabolite remained significant after multiple-testing correction. PCA and heatmap analyses revealed exploratory variation in amino acids, carbohydrates, lipids, and related metabolite groups. Pathway analysis identified statistically supported enrichment of alanine, aspartate and glutamate metabolism and cyanoamino acid metabolism in the coffee husk gradient, whereas the remaining pathway associations were exploratory after FDR correction. Overall, the results indicate that substrate composition strongly influenced productivity, while the global metabolomic organization of P. djamor remained comparatively stable, with localized metabolite and pathway patterns that require further validation.