We performed a pooled analysis with trial sequential analysis (TSA) to evaluate the efficacy and safety of chitosan supplementation on serum lipids in humans. Medline, EMBASE, and CENTRAL databases were queried. Impact was expressed as a weighted mean difference (WMD) and 95% confidence interval (CI). Sensitivity analysis was conducted using the leave-one-out method. Statistical heterogeneity, publication bias, TSA, and subgroup analyses were also assessed. Fourteen trials (21 treatment arms) encompassing 1108 participants were suitable for statistical pooling. Chitosan supplementation significantly improved the total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) concentrations in all patients. The WMDs were -0.20 mmol L-1 (95% CI, -0.35 to -0.05; p = 0.009) for TC, and -0.20 mol L-1 (95% CI, -0.26 to -0.15; p = 0.0001) for LDL-C, respectively. TSA demonstrated that the cumulative Z-curve crossed the trial sequential monitoring boundary for benefit providing conclusive evidence for the benefit of chitosan. However, no significant changes were seen with high-density lipoprotein cholesterol (HDL-C) and triglycerides. Our findings were robust after sensitivity analyses, and no serious adverse events were reported with chitosan intake. Supplementation with chitosan effectively reduces plasma concentrations of TC and LDL-C. Current evidence indicates daily chitosan supplementation as a candidate for therapeutic lipid management strategies. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.