In-depth spectral characterization of antioxidative (1,3)-β-D-glucan from the mycelium of an identified tiger milk mushroom Lignosus rhinocerus strain ABI in a stirred-tank bioreactor

A rare Malaysian tiger milk mushroom Lignosus rhinocerus strain ABI (LRSA) was morphologically identified based on its pileus, stipe, and sclerotium. LRSA (515 bp) was sequenced and found to have 99% similar to L. rhinocerus strains CH31 and CH2. Phylogenetically, evolutionary distance (Knuc) and plasmid-matching software (ApE) for sequences of matching fungal species were used to verify that the isolate belonged to the L. rhinocerus species. The strain was cultured in a stirred-tank bioreactor and a mycelial β-glucan (G) was extracted for compound characterization. The structure of exopolysaccharide extract from mycelium of LRSA was studied using Fourier-transform infrared spectroscopy (FT-IR) and one-dimensional (1D) and two-dimensional (2D) Nuclear Magnetic Resonance (NMR). FT-IR spectroscopy showed that G exhibited a similar β-glycosidic structure to the standard (laminarin), and the presence of characteristic bands at 3277, 2919, 1638, 1545, 1400, 1078, and 896 cm−1 confirmed the similarities. 1H and 13C NMR, as well as 2D NMR: homonuclear correlation spectroscopy (COSY), total correlation spectroscopy (TOCSY), heteronuclear multiple quantum coherence (HMQC), and heteronuclear multiple bond coherence (HMBC) spectra, were used for structural elucidation of the β-glucan and confirmed the extracted material as (1,3)-β-D-glucan. In addition, the G compound exhibited antioxidant activities through total phenolic content (4.47 mg gallic acid equivalents/g), 2, 2-diphenyl-1-picrylhydrazyl (11.48 mg/mL), and ferric reducing antioxidant power (0.56 mg/mL) assays. These findings may facilitate the development of rare G production in a high-scale bioreactor using LRSA.