Peroxides (H2O2, ROOR, and ROOH) are an important reaction intermediate involved in a number of natural processes, including atmospheric autoxidation and lipid peroxidation in oils and animal tissues. Iodometry is an established spectroscopic technique that has been widely used to quantify total peroxide concentration in food, indoor, and outdoor samples. Iodometry provides selectivity toward peroxides through a quantitative reaction between I- and peroxides to form I3- via a molecular iodine (I2) intermediate. However, equilibrium changes caused by a potential interaction between olefinic species and I2 can suppress I3- formation, thereby underestimating peroxide concentration. For the first time in the current study, this unrecognized interference posed by olefins (OEs) is systematically investigated to gauge its effects on the accuracy of iodometry. A number of model molecules were investigated. The interference was observed to be unique to OEs, but universally affecting different peroxide species such as H2O2, tert-butyl hydroperoxide, and aerosol-bound peroxides. A simple kinetic box model was built to explain this chemistry. The measured rate constant for 3-octenoic acid was found to be 0.84 ± 0.02 M-1 s-1. Overall, our results show matrix effects induced by OEs can underestimate peroxide concentration determined by iodometry for edible oils, indoor environments, and animal fat, but absent in most of the atmospheric samples. Nonetheless, our results point out the importance of this interfering chemistry in matrices enriched with OEs.