This study was conducted to investigate the fermentation characteristics and anti-obesity effects of Cheonnyuncho (Oputia Humifusa) fruit fermented with Lactobacillus plantarum SRCM 100320 (FC). The pH gradually decreased from 4.77 to 3.63 at 72 hours during fermentation. Counts of lactic acid bacteria, total polyphenol and flavonoid contents and DPPH scavenging activity were the highest at 48 hours during fermentation. Evaluation of the composition of polyphenols and flavonoids of FC fermented at 48 hours by HPLC revealed hyperoside (quercetin 3-galactoside), luteolin and kaempferol were the major components. The hyperoside content of FC was decreased, while the luteolin and kaempferol contents of FC were increased compared to unfermented Cheonnyuncho (NFC). Evaluation of the anti-obesity effects of FC in 3T3L-1 cells revealed that the accumulation of triglyceride was inhibited by about 27.3% in cells treated with FC at 150 μg/mL compared to NFC. These findings indicate FC has the potential for use as an anti-obesity material.