The flesh oil content (OC) is a crucial commercial indicator of avocado maturity and directly correlates with its nutritional quality. To meet export standards and optimize edible characteristics, avocados must be harvested at the appropriate stage of physiological maturity. The significant variability in OC during maturation, without any external morphological indicators, poses a longstanding challenge. Currently, harvesting maturity is optimized through time-consuming, destructive laboratory methods like freeze-drying and chemical extraction, which use representative samples to estimate the maturity of entire orchards. In this study, for the first time, we employed fluorescence imaging of avocado skin using 365-nm UV polarized light excitation to estimate the OC in the ‘Bacon’ avocado cultivar. We developed a surface fluorescence index that strongly correlates with OC, achieving correlation coefficients up to − 0.91. Our non-destructive and rapid approach achieved a cross-validation accuracy with an R2 value of 0.81, enabling the classification of avocados with low and high OC. This pioneering method shows considerable potential for further improvement and refinement. This study lays the groundwork for developing a portable, cost-effective, and real-time method for non-destructive in situ monitoring of avocado OC in the field and its integration into large-scale post-harvest grading systems.