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Background: Time since death (TSD) is an important parameter to the forensic expert. Only a few studies are available in the literature to determine the relationship between the vitreous potassium (K + ) concentration and TSD especially among natural death cases in the state of Haryana and, thus, information on the same is patchy and scanty. Objective: To determine the relationship between changes in the vitreous K + concentration and increasing TSD among natural death cases. An additional objective was to derive a formula for determining TSD from changes in levels of vitreous K + in the study subjects. Materials and Methods: In this study, autopsy cases brought to the mortuary of the Department of Forensic Medicine and Toxicology of a tertiary care center, were the subjects for collection of vitreous humor for the determination of vitreous K + . Data of 90 such cases where autopsy confirmed the cause of death as natural death were included in this study. Determination of K + level in the collected sample was done by ion-selective electrode method (Biolyte 2000 autoanalyzer, Biocare, Lujhu Township, Taiwan). Results: A linear relationship was observed between vitreous K + concentration and TSD in both the eyes. External factors like seasonal variations and temperature, gender difference, and age had no appreciable effect on the concentration of vitreous K + in either eye. In this study, the linear regression equations obtained from K + concentration (y) versus time (x) for the two groups among injury (trauma) cases were as follows: For the right eye: Regression line (y) = -4.303 x + 2.148 and for the left eye: Regression line (y) = -5.260 x + 2.209. The K + levels are found to increase up to 40 h after death. Conclusion: The study highlighted the usefulness of the relationship between vitreous K + concentration and TSD in both the eyes. The formula calculated from the current study could prove to be more appropriate as it is based on a more recent study and more reliable tests.