The sulfur cycle is an important part of constructed wetland biogeochemistry because it is intimately intertwined with the carbon, nitrogen, and iron cycles. However, to date, no quantitative investigation has been conducted on the sulfur cycle in constructed wetlands because of the complexity of wetland systems and the deficiencies in experimental methodology. In this study, 34 S-stable isotope analysis was extended in terms of the calculation for the enrichment factor and the kinetic analysis for bacterial sulfate reduction. With this extended method, we attempted for the first time to assess the true rate of bacterial sulfate reduction when sulfide oxidation co-occurs. The joint application of the extended 34 S-stable isotope and mass balance analyses made it possible to quantitatively investigate the primary sulfur transformation in a wetland microcosm. Accordingly, a sulfur cycle model for constructed wetlands was quantified and validated. Approximately 75% of the input sulfur was discharged. The remainder was mainly removed through deposition as acid volatile sulfide, pyrite, and elemental sulfur. Plant uptake was negligible. These findings improve our understanding of the physical, chemical, and biological transformations of sulfur among plants, sediments, and microorganisms, and their interactions with carbon, nitrogen, and iron cycles, in constructed wetlands and similar systems.