To compare the efficacy of 14- and 7-d bismuth-based quadruple therapies as second-line eradication treatment for Helicobacter pylori (H. pylori) infection. Between 2004 and 2014, the medical records of 790 patients who had experienced failure of first-line proton pump inhibitor (PPI)-based eradication therapy and were then treated with bismuth-based quadruple therapy were retrospectively reviewed. Those who received bismuth-based quadruple therapy [PPI, bismuth, metronidazole, and tetracycline (PBMT)] for either 7 d or 14 d were assigned to a PBMT-7 group (n = 543) or a PBMT-14 group (n = 247), respectively. The eradication rates for both groups were determined by intention-to-treat (ITT) and per-protocol (PP) analyses. ITT analysis compared the treatment groups as originally allocated while the PP analysis including only those patients who had completed the treatment as originally allocated. Successful eradication therapy for H. pylori infection was defined as a negative (13)C-urea breath test 4 wk after the end of eradication treatment. The overall ITT eradication rate was 69.1% (546/790). Final ITT eradication rates were 67.4% (366/543; 95%CI: 63.1%-71.7%) in the PBMT-7 group and 72.8% (180/247; 95%CI: 67.4%-78.2%) in the PBMT-14 group (P = 0.028). The overall PP eradication rate was 80.0% (546/682), and the final PP eradication rates were 78.2% (366/468; 95%CI: 72.1%-84.0%) in the PBMT-7 group and 84.1% (180/214; 95%CI: 76.8%-90.8%) in the PBMT-14 group (P = 0.009). The H. pylori eradication rates in the PBMT-14 group were significantly higher than in the PBMT-7 group according to both ITT (P = 0.028) and PP analysis (P = 0.009). Compliance was similar in both groups (PBMT-7 group: 97.9%; PBMT-14 group: 96.4%). Adverse event rates were 10.7% (51/478) and 17.1% (38/222) in the PBMT-7 and PBMT-14 groups, respectively (P = 0.487). The 14-d bismuth-based quadruple therapy is a significantly more effective second-line eradication treatment for H. pylori infection than the 7-d alternative.