Bradford method reagents

Total carbohydrate content of truffle fruiting bodies and extracts obtained using PLE (50 mG/mL) was quantified by the phenol–sulfuric acid method [27 (link)]. D-glucose curve standard was used for quantification. Chitin content (10 mG/mL) was quantified as described by Tejedor-Calvo et al. (2019) [28 (link)] using a glucosamine hydrochloride standard curve. Total β-glucan content (50 mg) was evaluated by a β-glucan determination kit specific for mushrooms and yeasts (Megazyme^®, Biocom, Barcelona, Spain) following the instructions of the user’s manual. Soluble protein concentration (10 mG/mL) was determined using the Bradford method reagents (Sigma-Aldrich, Madrid, Spain) according to the instruction manual. BSA was used as standard for protein quantification. Total phenolic compound levels (10 mG/mL) were evaluated by the Folin–Ciocalteu method [29 (link)]. Gallic acid was used as standard for quantification. Truffle fruiting bodies and extracts obtained using PLE were saponified and analyzed by GC-MS-FID [28 (link)]. Ergosterol was used as standard, and Hexadecane (10% v/v) was used as internal standard and ergosterol as standard for ergosterol and derivative compounds quantification. The compound determinations were carried out in triplicate.

Soluble protein concentration (10 mg/mL) was evaluated using the Bradford method reagents (Sigma–Aldrich, Madrid, Spain) and total phenolic compound levels (10 mg/mL) by the Folin–Ciocalteu method, as indicated in a previous study [27 (link)]. BSA and gallic acid were used as standards for quantification.