Abstract Background The recommended second-line therapy for Helicobacter pylori ( H. pylori ) eradication is a quadruple regimen that fails in up to 30% of patients. Several recent studies suggest levofloxacin-based triple therapies as an alternative rescue treatment. However, dosage and length of levofloxacin-based regimens have not been established. Aim To compare the efficacy and tolerability of four second-line levofloxacin-based schemes for H. pylori eradication. Methods One hundred and sixty patients (aged 18–70 years, 72 male patients) who were H. pylori positive after standard triple therapies were randomised to receive esomeprazole 20 mg b.d. and amoxicillin 1 g b.d. plus levofloxacin 500 mg o.d., for 7 or 10 days (Groups A and B) or levofloxacin 500 mg b.d. for 7 days or 10 days (Groups C and D). H. pylori status was assessed by 13-C Urea Breath Test or rapid urease test, before and 6 weeks after therapy. Incidence of side effects was evaluated by a questionnaire. Results No dropouts were observed. Eradication of H. pylori infection was successful in: 65% of patients in Group A; 90% in Group B; 70% in Group C; 85% in Group D. Based upon duration of treatment, eradication rates were: 67.5% in 7 days groups and 87.5% in 10 days groups ( p = 0.004). Dosage of levofloxacin did not affect the eradication rates (77.5% both in the once daily and twice daily groups). Mild adverse events were reported overall in 16% of patients (22.5% in 7 days groups; 27.5% in 10 days groups; p = 0.58; 12% in the once daily group; 32.5% in the twice daily group; p = 0.04). Conclusions 10 days levofloxacin-based second-line regimens were effective in curing H. pylori infection in more than 85% of patients with a lower incidence of adverse effects in levofloxacin single-dosage scheme. The 10 days levofloxacin-based regimens were more effective than 7 days course of treatment showing that duration of therapy is the crucial factor affecting eradication rate.