Regardless of major advances in mass spectrometry imaging (MSI), there are three intrinsic limitations associated with MSI, including intricate molecular identification, low molecular coverage, and incapability to obtain "true" spatial distribution due to isobaric and particularly isomeric ions interference. We developed a novel approach that integrates in vivo dual isotope labeling of precursor metabolites with MSI (DLEMMA-MS-Imaging) for identification of spatially localized metabolite and metabolic network map reconstruction. In a proof-of-concept study, we identified 59 and 6 novel metabolites in lemna and tomato fruit, respectively. Significantly, 20-30% of the identified metabolites were found to contain at least one structural isomer that displays a different distribution pattern. The notable feature of this approach is the ability to differentiate localization of structural isomers, hence, providing the "true" distribution of molecules of interest.