In this paper, a comprehensive spatio-spectral and temporal analysis for Chromobacterium violaceum colonies is reported. A hyperspectral imaging (HSI) system is used to recover the spectral signatures of pigment production in a non-homogeneous media with high spectral resolution and high sensitivity in vivo, without destructing the sample. This non-contact sensing technique opens avenues to study the temporal growing of a specific section in the bacterial colony. Further, from a 580 [nm] and 764 [nm] spatio-spectral time series, a wild-type and mutant Chromobacterium violaceum strains are characterized. Such study provides quantitative information about kinetic parameters of pigment production and bacterial growing.