Post mortem interval (PMI) determination is a particularly crucial detail often needed in medico-legal cases. Though, today, several methods of estimating PMI exist, most of these methods lack practical relevance, which brings about a need to find new, cost-efficient, and more reliable methods. One such temperature independent method is the evaluation of post-mortem pupillary reactivity to various pharmacological substances. The objective of the present work is to test and provide evidence of the effectiveness of pupillary reactivity to pilocarpine eye drop as an indicator to determine PMI. A total of 133 deceased with known PMI were instilled with 2% pilocarpine eye drops and measure change in pupil diameter using ImageJ freeware. A negative correlation between the changes of pupil diameter under the influence of the pilocarpine eye drop showed less change with increase PMI. Pearson correlation showed the negative correlation between the pupillary changes with PMI with p value 0.0001, 0.00017, 0.0341 and 0.0021 where á=0.05 for natural death, hanging, burn and poisoning respectively. Observations showed postmortem chemical excitability could be used as an indicator in PMI using pilocarpine eye drop.