BackgroundThe by‐product of insect larval production, frass, can be applied to soil as an organic fertiliser. Its three main organic N fractions are assumed to be ureic, protein and chitin. The significance of the latter is unknown, and it is not known if lignaceous sources have been overlooked.AimsThis study sought to gauge the activities of the (respectively, lignolytic and chitinolytic) enzymes peroxidase and N‐acetyl‐ß‐D‐glucosaminidase following frass application to soil. Their activities were monitored under conditions of urease inhibition, with a particular focus on the fungal domain.MethodsMealworm or buffalo worm frass was applied, with or without inhibitors, to a sand/soil substrate at 3% (w/w). After 16 weeks, concentrations of the fungal biomarker ergosterol and enzyme activities were determined.ResultsSoil amendment with frass had no significant effect on peroxidase activity. Fungal biomass was stimulated in particular by application of mealworm frass, which was further improved by urease inhibition. Chitinase activity was positively correlated with fungal biomass, and was increased under urease inhibition when applied with mealworm frass.ConclusionsThere were no appreciable quantities of lignaceous compounds in the frass used in this study. Importantly, the use of urease inhibitors co‐applied with frass has demonstrated that when its ureic N breakdown is prevented, chitin becomes a significant organic N source to soil fungi. The superior fungal response to mealworm frass indicates a larger chitin content than in buffalo worm frass.