Medicinal plant Rehmanniae Radix has long been used as traditional herbal medicine in Korea. In this study, we investigated quantitative changes of marker substance 5-hydroxymethyl-2-furaldehyde (5-HMF) in Rehmanniae Radix Preparata fermented with Ganoderma lucidum, Paecilomyces japonica, honey, and Nuruk, using HPLC according to the Korean Pharmacopoeia. As a result, 5-HMF was decreased in SDT and SYT, and increased in SST and SNT, comparing with control group. Quantitative increasing of 5-HMF is not exactly in direct proportion with fermentation, and it is need further studies to elucidate mechanism of quantitative changes by converted and newly produced substances with fermentation.