Extraction of .BETA.-Glucan from the Water-insoluble Residue of Hericium erinaceum with Combined Treatments of Enzyme and Microwave Irradiation

Abstract

Recently, it was found that Yamabushitake (the fruiting body of Hericium erinaceum) had a variety of bioactive compounds. 1) Since most of the bioactive compounds in H. erinaceum are included in its lowmolecularweight compounds, they are extracted with solvents such as ace-tone, ethanol and so on. 2,3) In addition, H. erinaceum has been known to contain several polysaccharides including βglucans. 4) Within βglucans, (13;16)βDglucans are important because of their antitumor and immunopo-tentiating activities. 5) For example, such βglucans known as Krestin from Trametes versicolor, Lentinan from Len-tinus edodes and Schizophyllan from Schizophyllum commune were used medicinally as hostmediated activity drugs. 6) Various βglucans of this type are found in many kinds of resources; prokaryotic organisms such as bacteria , fungi and yeast. 7) Among these resources fruiting bodies of fungi (mushrooms) are the most attractive starting materials for getting (13;16)βDglucan because of commercial availability as food stuffs. It is important to extract βglucans included in Yamabushitake for their application to medicinal and diet usages. In a previous paper , we indicated that polysaccharides, especially β glucans, remained in the residue given after removal of the lowmolecularweight compounds are extractable from the fruiting body of H. erinaceum by irradiation of microwave in water. 8,9) The duration of time necessary to obtain waterextractable (13;16)βDglucan could be extremely reduced by applying microwave irradiation in water. However, it was found that 75.9% of βglucan still remained in the residue. Therefore, development of new methods effective for production of βglucans from Yamabushitake is important to use this mushroom more extensively. Some (13;16)βDglucans have been known to form complex networks with protein and chitin in a cell wall of fungi. 10,11) Based on these previous observations, similar complex networks may be present in the cell wall of H. erinaceum. Degradation of protein andor chitin by specific enzymes is, therefore, expected to improve extraction of βglucan remained in the residue. In this study, we examined the effects of treatments of proteolytic and chitin degrading enzymes followed by microwave irradiation on extractability of (13;16)βD glucan from the waterinsoluble residue of H. erinaceum. MATERIALS AND METHODS Materials and microwave irradiation. A column of YMCPack Diol300 (8×500 mm) was supplied by YMC Co., Ltd. Pullulans (Shodex Standard P82) used for estimation of molecular weight as the standard were obtained from Showa Denko, Co., Ltd. All reagents used were of analytical reagent grade. Pulverized fruiting bodies of Hericium erinaceum were supplied from Saishin, Co., Ltd. in Nagano Prefecture, Ja-pan. Mushrooms of H. erinaceum (FERMP19600) were fruited on sawdustbased medium by cultivation at 18 20 C for 2030 days, followed by heating at 1012 C under relative humidity of >90%. Fruiting bodies developed were harvested before development of coloration. Harvested fruiting bodies were dried by three steps; keeping for 68 h at 3842 C, then heating up to 65 C and keeping at this temperature for about 2 h, and finally drying for 23 h at 82 C to make moisture content below 7%. Before use, the dried raw materials were treated with ace-tone and 80% ethanol to remove lipids and low molecular weight constituents, as described previously. 8) The residue (R1) was irradiated by microwave in water to remove watersoluble constituents. Microwave irradiation was car-Abstract: We attempted to extract (13;16)-β-D-glucan from the water-insoluble residue of Hericium eri-naceum with proteolytic and chitin degrading enzyme treatments followed by microwave irradiation. Two sets of four commercially available enzymes were used in this study; Actinase E, Proteinase K, Chitinase-RS and Chitinase. The results showed that the enzyme treatment using Proteinase K and Chitinase simultaneously had the most significant synergistic effects on the amount of glucan extracted; 95.5% of the amount of glucan extractable with 5% sodium hydroxide. The structure of (13;16)-β-D-glucan obtained by this enzyme treatment was different from that obtained by extraction with alkaline solution in that the former was rich in (16) linkages, whereas the latter was rich in (13) linkages.