Kwon et al. BMC Cancer 2010, 10:392 http://www.biomedcentral.com/1471-2407/10/392 Statistical analysis A two-tailed Student’s t-test was employed where P < 0.05 was considered to be statistically significant (*p < 0.05, **p < 0.005, and ***p < 0.001). Results Cinnamon extract inhibits tumor cell growth in vitro To define the optimal concentration at which cinnamon extract does not induce cell damage, the cytotoxicity test was performed in melanoma cell lines. As we reported previously, treatment of cinnamon extract up to 0.5 mg/ml did not induce growth inhibition and mor- phological changes till 24 hrs [22]. However, tumor cells showed significant decrease in cell proliferation after 48 hrs treatment of cinnamon extract (Figure 1). They showed condensed, shrank and aggregated shapes. Seventy two hours after treatment of cinnamon extract, most of the cells became floating with aggregated form detached from the plates (Figure 1A). To confirm the effects of cinnamon treatment on tumor cell survival, proliferation and cell viability were measured (Figure 1B). In the presence of cinnamon extract, the rate of tumor cell growth was significantly inhibited (Figure 1B). To validate our experimental system, we used EL4 T lymphoma cell line as a positive control that showed cell cycle arrest and growth inhibition upon treatment of polyphenolic compound from cinnamon [24]. We checked whether cinnamon extract also induced growth inhibition of normal cells. Primary mouse lymphocytes Page 4 of 10 were treated with same concentration (0.5 mg/ml) of cinnamon extracts and cell viability was measured (Additional file 1, Figure S1A). Interestingly, treatment of cinnamon extract (0.5 mg/ml) did not induce any growth inhibition in primary mouse lymphocyte. Treatment of cinnamon extract induces active cell death of melanoma cells in vitro Growth inhibition of tumor cells by the treatment of cinnamon extract could be mediated by several mechan- isms. A recent report showed that phenolic compound of cinnamon bark induces cell cycle arrest in hematolo- gic cell lines [11]. Hence, we first tested whether cinna- mon extract induced cell cycle arrest in mouse melanoma cell line, B16F10 cells. Treatment of cinna- mon extract slightly induced cell cycle arrest at S phage (Additional file 2, Table S1). We further tested whether growth inhibition by treatment of cinnamon is related with induction of apoptosis (Figure 2). Although cinna- mon treatment did not induce growth inhibition and morphological chance within 24 hrs [22], a gradual + increase in early apoptotic population (Annexin V ) was observed (Figure 2A and 2B; 12 and 24 hrs). Further enhancement of apoptotic population in late stage (Annexin V+/7-ADD+) was observed in a time-depen- dent manner (Figure 2A). To further confirm these results, we checked whether treatment of cinnamon extract induces apoptosis in another type of cancer cell line, Caco2 (human epithelial colorectal adenocarcinoma Figure 1 Treatment of cinnamon extract inhibits the growth of various cancer cells. Cinnamon (0.5 mg/ml) was treated for 0, 48 and 72 hrs to cancer cell lines (Hela; cervical cancer, Caco2; colon cancer, Clone M3; melanoma and B16F10; melanoma). After treatment of cinnamon extract, (A) morphological changes of each cancer cell lines were monitored by microscopic observation. (B) Proliferation and viability of cancer cells were measured by cell viability assay at the indicated time points. Error bars indicated SD. Data are representative of three independent experiments.