Procyanidins are one of the main polyphenols in apple fruit. In this study, we aimed to increase the amount of extractable procyanidins by micro-wet milling (MWM), a novel milling process that can wet-mill foods to micrometer scale, in addition to commonly applied apple juice manufacturing processes. The effects of milling, pasteurization, centrifugation, and enzymatic treatment on extractable procyanidin concentration were investigated, and MWM was shown to increase the procyanidin concentration by 16.7% compared with mixer milling. Conversely, other processes such as pasteurization, centrifugation, and enzymatic treatment decreased the procyanidin concentration in apple juice. Since procyanidin concentrations in apple juice are also affected by the variability between individual apples, we attempted to nondestructively estimate the procyanidin concentration at each process of apple juice manufacturing by obtaining the fluorescence fingerprint (FF). The FFs are a set of fluorescence spectra acquired at consecutive excitation wavelengths. Partial least-squares regression was used to estimate the procyanidin concentrations of apple juice from the FFs, and the most accurate model was able to estimate procyanidin concentration at the quality control analysis level.