Astragaloside IV (ASI), a pure compound derived from Radix Astragali, is commonly used in degenerative bone diseases such as osteoporosis. Our previous study identified in vivo the osteogenetic effect of Fu Fang Qi She Pills (FFQSP), a Chinese herbal formula containing Radix Astragali from which ASI was extracted. In this study, we investigated the osteogenetic effects of ASI under the conditions of centrifugating pressure on OCT-1 cells. These preosteoblasts were grown in 3D-culture, and treated with ASI at 50 micromol/l with centrifugation at 200 rpm, 500 rpm for 3 and 5 days. Morphocytological examination, morphometry of alkaline phosphatases (ALP) staining was performed. Expression of type I collagen (Col I) was detected by immunocytochemistry assays. ALP, Col1a2, Osteocalcin (OC), and runt-related transcription factor-2 (Runx2) mRNA expression were determined via real-time PCR. The results showed ASI plus 500 rpm for 3 days and ASI plus 200 rpm for 5 days significantly induced osteogenesis related protein and gene expression. We concluded that ASI would promote osteogenesis when cells of preosteoblast OCT-1 were subjected to proper centrifugating pressure and a pertinent period of time.