Myoglobin content is considered as a crucial index to evaluate the quality of frozen pork. In this study, a portable visible and near-infrared (Vis-NIR) spectrometer combined with chemometrics was used to detect myoglobin content in frozen pork. Metmyoglobin, deoxymyoglobin, oxymyoglobin, and total myoglobin were assessed spectrophotometrically. The raw Vis-NIR spectra of frozen pork samples were pre-processed using 1st derivatives (FD). Afterward, Synergy Interval Partial Least Square (Si-PLS) coupled Competitive Adaptive Reweighted Sampling algorithm (Si-CARS-PLS) was applied to select characteristic variables. The Si-CARS-PLS models revealed the probability of estimating myoglobin content in frozen pork, with predictive correlation coefficients (Rp) for metmyoglobin, deoxymyoglobin, oxymyoglobin, and total myoglobin as 0.9095, 0.9004, 0.8578, and 0.9133, respectively. The findings of this study showed that Vis-NIR spectroscopy coupled with Si-CARS-PLS is a promising method and offered a way forward for determining the myoglobin content in frozen pork.