Hepatitis C virus (HCV) is known to infect and replicate within peripheral blood mononuclear cells (PBMC), thereby enabling the direct evaluation of antiviral mechanisms by analyzing HCV dynamics in PBMC. To address potential molecular differences associated with distinct antiviral regimens, we studied HCV dynamics in both serum and PBMC in 44 patients with HCV genotype 1b and high viral load who were randomly assigned to the following 4 different treatment groups: 1) combination therapy with 6 MU daily of interferon alfa 2b (IFN-α2b) plus 800 mg of ribavirin; 2) monotherapy with 6 MU daily of IFN-α2b; 3) monotherapy with twice-daily intravenous administration with 3MU of IFN-β; and 4) monotherapy with daily intravenous administration with 6 MU of IFN-β. HCV-RNA levels were measured serially using highly sensitive real-time detection polymerase chain reaction (PCR). HCV dynamics in both the serum and PBMC showed a “biphasic” pattern. The exponential decay slopes of the second phase were significantly higher in the combination or twice-daily dosing regimen groups compared with groups 2 or 4 (0.10 ± 0.08 vs. 0.02 ± 0.09 or 0.16 ± 0.09 vs. 0.02 ± 0.04 day−1; P < .05 or P < .0005, respectively). Moreover, the viral half-lives in the second phase were significantly shorter in these groups (73.2 ± 42.5 vs. 240.1 ± 120.7 or 56.0 ± 44.6 vs. 361.6 ± 293.5 hours; P < .05 or P < .05, respectively). Additionally, the slope of HCV decline in PBMC tended to be higher in the combination regimens, as compared with monotherapy. Taken together, our data on HCV dynamics provide molecular insight into utilization of combination or twice-daily dosing regimens to increase rates of sustained viral eradication of HCV.