Sterol extraction

To study sterol synthesis, Candida albicans CAAL93, C. krusei CAKR7 and C. glabrata CAGL2 blastopores were incubated in 50 ml Sabouraud broth medium (Sigma) during 18 h at 35 °C with stirring. Cells were collected by centrifugation at 1500 g. Pellet was suspended in 3 ml of fresh ethanolic potassium hydroxide solution (25 g of KOH, 36 ml of distilled water and brought to 100 ml with 100% ethanol). Saponification was performed at 80 °C for 60 min. Sterols were extracted by addition of 3 ml of hexane and the organic phase was then washed by adding 1 ml of sterile water. Final organic phase was transferred to a new collection tube after 5 min centrifugation step at 2000 rpm.