Media and fermentation

All media were made using distilled water and the pH was adjusted to 7 using concentrated potassium hydroxide. The stock culture medium contained 30 g l⁻¹ Difco Trypticase Soy Broth (Difco Laboratories, Detroit, MI, USA). The seed medium developed by Merck (Rahway, NJ, USA) and employed in this study contained (g l⁻¹): 20 soluble starch, 10 dextrose, 5 NZ Amine type A, 3 beef extract, 5 Bacto-Peptone, 5 yeast extract and 1 CaCO₃. All flasks were autoclaved at 121 °C for 30 min prior to use. The stock culture medium was inoculated with a sample of S. platensis MA7327 and incubated on the gyratory shaker at 220 r.p.m. at 28 °C for 2 days followed by storage in the refrigerator until use. The seed medium was inoculated with 0.5 ml of S. platensis stock culture and shaken at 220 r.p.m. and 28 °C for 5 days. The 5-day seed culture was spun down in a sterile 50-ml conical tube using a clinical one-speed centrifuge for 10 min. The supernatant fluid was discarded, replaced with an equal volume of sterile water and the pellet was resuspended. This washing process was repeated two more times. The culture was stored as a liquid in the refrigerator after the third addition of water. Fermentations were carried out in 125-ml Erlenmeyer flask containing 25 ml of fermentation broths on a rotary shaker at 28 °C and 250 r.p.m.