2.5 Cell Viability Assay
The modified method of Glenner (Glenner, 1977) was used. Briefly, cells were collected from 1 mL culture broth by centrifugation (16,000 rpm for 10 min) and resuspended in 1 mL 2,3,5-triphenyl-2H-tetrazolium chloride (TTC) solution (1 g/L) by vigorously vortexing. After incubation in dark at room temperature for 30 min, cells were collected by centrifugation and mixed with 1 mL methanol to extract the pink pigment. The color intensity in the supernatant was measured at 485 nm using a spectrophotometer (UV-16-1, Shimazu, Columbia, MD). The viability of tested cells was determined using the exponential-phase cells as the control with 100% viability and boiled (dead) cells as the negative control with 0% viability.