An investigation into the performance of luminescence-based hyperspectral imaging (LHSI) for denim fiber bundle discrimination has been conducted. We also explore the potential of nitromethane (CH₃NO₂) -based quenching to improve discrimination, and we determine the quenching mechanism. The luminescence spectra (450–850 nm) and images from the denim fiber bundles were obtained with excitation at 325 or 405 nm. LHSI data were recorded in less than 5 s and subsequently assessed by principal component analysis or rendered as red, green, blue (RGB) component histograms. The results show that LHSI data can be used to rapidly and uniquely discriminate between all the fiber bundle types studied in this research. These non-destructive techniques eliminate extensive sample preparation and allow for rapid hyperspectral image collection, analysis, and assessment. The quenching data also revealed that the dye molecules within the individual fiber bundles exhibit dramatically different accessibilities to CH₃NO₂.