Effect of process parameters on the purity of chitin isolated from black soldier fly larvae biomass using a subcritical methanol–water mixture

• Solvolytic fractionation of black soldier fly larvae with subcritical methanol–water. • Design of experiments to assess impact of process parameters on chitin purity. • 3-hour treatment of black soldier fly larvae produces chitin with 75 % purity. • Nearly all proteins are removed from the chitin, leaving only < 0.3 % amino acids. • Glutamic acid/ glutamine resisted removal most, suggesting a link to chitin-binding. Black soldier fly (BSF, Hermetia illucens ) larvae efficiently convert low-value waste streams into high-value larval biomass, containing substantial amounts of lipids, proteins, and chitin (the latter 4.73 ± 0.25 % dry matter). Although these larvae are mainly used as animal feed, isolating substances like chitin significantly boosts their economic value. However, the presence of chitin-binding proteins presents a major challenge in the isolation. Traditional chemical processes for isolating chitin typically produce large amounts of chemical waste, degrade chitin quality, and are time-consuming. This study introduces an innovative method for separating chitin from larval biomass using a subcritical methanol–water mixture, which is more sustainable and less time-consuming. Given the novelty of this process with the methanol–water mixture, the effect of process parameters on the purification was previously unknown. To address this, a design of experiments (DoE) was employed to assess the influence of reaction temperature, time, and methanol-to-water ratio on the chitin and amino acid content of the purified chitin. Following model fitting, optimal settings for maximal purity were predicted and experimentally verified. As it stands, a treatment with a 7:1 w:w methanol–water mixture at 250 °C for 158 min resulted in chitin with (74.7 ± 2.0 %) purity, and 77 – 85 % acetylation. Furthermore, the purified chitin contained very few amino acids ( i.e. , 0.27 ± 0.04 %, primarily Glx), indicating that the covalently bound proteins are almost entirely removed. Thus, the proposed process presents a significant advancement over current methods for chitin purification from insect biomass.