A predictive model for the thermal inactivation kinetic parameter decimal reduction time (D_T) of spoilage yeast in simulated fruit juice (SFJ) is being reported for the first time in this study. The target microorganism was a previously identified heat-resistant strain of Pichia anomala (BIOTECH 2205), which was cultured to its mid-stationary phase (MSP) to reflect a more resilient physiological state. The spoilage yeast spp. was artificially inoculated into SFJs formulated with varying combinations of pH, processing temperature (°C), and natamycin concentration (ppm) to simulate diverse juice environments. These intrinsic (pH and natamycin) and extrinsic (temperature) variables were systematically varied based on a Rotatable Central Composite Design (RCCD) to generate a statistically sound range of treatment conditions. Response Surface Methodology (RSM) was employed to model and interpret the effects of these factors and their interactions on the thermal inactivation of the yeast. Heat treatment experiments showed that microbial inactivation followed a log-linear reduction pattern, with high coefficients of determination (R² = 0.90–0.99) indicating good model fit across all treatment combinations. Observed D_T values ranged from 5.70 ± 1.19 to 39.44 ± 4.91 seconds, highlighting substantial variation in thermal resistance depending on treatment conditions. Significantly, the D_T values were found to fit a quadratic model (p < 0.002), demonstrating the appropriateness of RSM in modeling the response. Among the tested factors, the individual linear effects of pH, temperature, and natamycin concentration, the quadratic effect of temperature (T²), and the interaction between pH and temperature (pH × T) significantly influenced the D_T values. These findings provide critical insights into the complex interplay between juice composition and processing parameters on the thermal resistance of spoilage yeasts. The developed model, subject to a validation procedure, may be useful in optimizing juice pasteurization schedules. This is the first study to assess the interaction between natamycin and heat treatment for simulated fruit juices.