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The decline in hepatitis C viral load on treatment with peginterferon-α-2b is not continuous. The aim of this study was to investigate whether twice weekly dosing of peginterferon-α-2b may improve viral kinetics. Ten interferon-naïve patients with chronic hepatitis C (genotype 1a or b) were randomized to receive either 1.0 μg/kg peginterferon-α-2b once (group A) or twice weekly (group B) for 4 weeks. Viral load and serum concentrations of peginterferon-α-2b were measured. Peginterferon-α-2b reached maximal blood concentrations 24 h after the first dose, followed by a linear decline during the subsequent days. On the day before administration of the next dose, peginterferon-α-2b was undetectable in nine patients in group A (once weekly dosing). The same pattern was observed during the next 3 weeks of therapy. In group B (twice weekly dosing) peginterferon-α-2b was detectable at any given time point and higher than in group A (P between 0.01 and <0.0001). Viral load decreased in all patients within 2 days after the first dose of peginterferon-α-2b, but increased again on day 3. In group A, it further increased until day 7. A similar pattern was observed in the second week. In contrast, in group B, viral load decreased again on day 4 and remained lower until the end of the study (P < 0.001). To achieve continuous drug exposure and to improve initial viral clearance, peginterferon-α-2b has to be given at least two times weekly.