The modification of cell wall pectin is a major contributing factor to apple softening, associated with intensive variations of firmness and microstructure. Thus, a better understanding of the relationship between fruit optical properties, firmness, 3D microstructure and pectin is important for improving the optical detection of apple firmness. This work explored the optical properties in the wavelength range of 430 – 1650 nm related to cell wall pectin during apple softening, in order to better determine their firmness and 3D microstructural changes. An automatic single integrating sphere system was used to quantify absorption coefficient (μa) and reduced scattering coefficient (μs′). Micro-CT imaging, SEM and TEM were used to quantify the porosity, cell numbers, cell volume, volume fraction and microstructure of Fuji apples during storage. Firmness, pectin fractions, soluble solids content (SSC), titratable acid content (TAC), and moisture content were also measured. Results showed that depolymerization of pectin substances induces an increase in porosity (r = 0.97) and modification of cellular structure, leading to a decrease in firmness (r = 0.90). The reduction in firmness and cell numbers exhibited a strong positive correlation with both μa in the range of 1000 – 1650 nm and μs′ between 430 and 1000 nm with mean r of 0.88–0.96, whereas increases in covalent-soluble pectin (CSP) and porosity displayed a negatively correlated with these bands. The linear relationship indicates that Vis-NIRS at 980 nm could better predict apple firmness than μa or μs′ alone, with a coefficient of determination R2 of 0.97. Our findings suggest that the mechanism of optical technology for detecting apple firmness may be the interaction between tissue structure and pectin on μs′ and μa.