The effect of caffeine on osteoblast cells viability measured by MTT assay. When osteoblast cells cultured with caffeine for one day, there was no statistically significant change in the formation of formazan; while in the 100 mM to 1 mM concentration of caffeine, the formation of formazan was significantly decreased in the third day's culture. At the 7th day's culture, decreased osteoblasts activities were observed in the presence of various concentrations of caffeine. We selected the 10 mM concentration of caffeine for the further biochemical study because the osteoblasts showed the highest activities during the 3rd and 7th testing period. (Shaded bars mean significant differences to that control: P < 0.05).