Solubilization of yeast cell-wall β-(1→3)-D-glucan by sodium hypochlorite oxidation and dimethyl sulfoxide extraction

The limulus test is a well-established method for the diagnosis of both Gram-negative sepsis and invasive fungal infection. To diagnose fungal infections, a beta-(1 --> 3)-D-glucan-specific chromogenic kit (Fungitec G test MK) has been developed and applied clinically. We are concentrating our main efforts on developing a better standard to improve the precision of this method. To this end, we have successfully developed a protocol to obtain a soluble Candida spp. beta-(1-->3)-D-glucan (CSBG) by sodium hypochlorite (NaClO) oxidation and subsequent dimethyl sulfoxide (Me2SO) extraction (yield of 9.6 +/- 4.1%) of acetone-dried whole-cell preparations. The beta-glucan fraction is free from the cell-wall mannan, gives a symmetrical peak by gel filtration, and is soluble in dilute NaOH. The product is composed mainly of beta-(1 --> 3)- and beta-(1 --> 6)-D-glucosidic linkages. The specific activity of the beta-glucan is comparable with pachyman when combined with the Fungitec G test as the standard glucan and reacted as low as 10(-11) g/mL. (C) 1999 Elsevier Science Ltd. All rights reserved.