Fluorescence and Phosphorescence Assay of β-D-Glucans from Basidiomycete Medicinal Mushrooms

Basidiomycete mushrooms contain complex β-D-glucans which act as immunomodulator, immune stimulants and anti-cancer agents, which can be either free or bound to proteins. The present report consists of a novel and intrinsic synchronous fluorescence and phosphorescence assay method for β-D-glucans. This analytical technique was carried out by a spectrofluorometer in the range of 250 to 750 nm with a Δλ range of 5–30 nm which exhibited peaks at 492, 540 and 550 nm by using β-D-glucan from Euglena gracilis as a standard. A micro and high-throughput method based on a microplate fluorescence reader was devised with a excitation and emissions λ of 420 nm and 528 nm, respectively. This assay method revealed some advantages over the reported colorimetric methods, since it is a non-destructive assay method of β-D-glucans in samples with a linearity range of 0–14 μg/well, correlation coefficient (r2) of 0.9961, LOD of 0.973 μg/well, LOQ of 2.919 μg/well, greater sensitivity, fast, a high-throughput method and very economical. β-D-glucans of several mushrooms (i.e., Poria coccus, Auricularia auricula, Ganoderma lucidium, Pleurotus ostreatus, Cordyceps sinensis, Agaricus blazei, Polyporus umbellatus, Inonotus obliquee) were purified by using a sequence of various solvent extractions, quantified by either spectrofluorometer or fluorescence microtiter plate reader assay and compared with Congo red assay method. Three-dimensional spectra measurements were carried out on β-D-glucans from commercial sources and medicinal mushroom strains. FTIR spectroscopy was selected to investigate the structural properties of β-D-glucans in these mushroom samples. Therefore, the present assay method is simple, fast, cheap and non-destructive for β-D-glucans from medicinal mushrooms as well as from commercial sources.