Objective To investigate the relationship between K+ concentration in rabbit vitreous humor and postmortem interval (PMI) under different ambient temperature. Methods Rabbit corpses were stored at 5℃ , 15℃ , 25℃ , and 35℃ after execution, and 80~100μL vitreous humor was extracted from each eye of the rabbit in turn every 12 hours. The concentrations of K+ were examined by Modular DPPI automatic biochemistry analyzer. The Interpolation Functions were used to analyze the statistical relationship between PMI and K+ concentration under different temperature. Results In each animal group, K+ concentration increased with PMI. Equation was obtained after interpolation analysis on range of temperature 5℃ ~30℃ . The three-variable quintic surface equation was f(x,y)=-1.998e14+1.345e12x+5.902e13y+0.005585x2-4.509e11xy-3.876e12y2-0.0002868x3+0.003545x2y+4.406e10xy2-1.746e10y3+2.669e-6x4-1.568e-5x3y-0.0001771x2y2-1.64e9xy3+6.669e9y4-8.672e-9x5+4.467e-8x4y+2.354e-7x3y2+2.459e-6x2y3+2.05e7xy4-1.214e8y5(R2=0.9956), x stands for temperature, y stands for K+ concentration, f(x,y) stands for PMI. Conclusion The rule of K+ concentration changes at ambient temperature complied with three-variable quintic surface equation distribution. Measurement of interpolation function may be used for PMI estimation at different ambient temperature.