Spatial mapping of PUFA incorporation into phospholipids with bisallylic deuteration

MS imaging using stable isotope-labeled fatty acids provides a groundbreaking approach to precisely localizing exogenous fatty acids and their metabolites in vivo. However, challenges persist, particularly with fatty acids labeled with fixed isotopic numbers, which can lead to spectral interferences and limit the number of metabolites that can be detected. In this study, we employed a bisallylic deuteration method to synthesize dihomo-γ-linolenic acid (DGLA) isotopes with m/z values ranging from +4 to +8 Da relative to endogenous DGLA, which allowed us to meticulously dissect DGLA metabolism in mice using LC-quadrupole TOF-MS and MS imaging. Our strategy enabled the clear selection of m/z values for phospholipids enriched with deuterated DGLA (D-DGLA) and deuterated arachidonic acid derived from D-DGLA, all while maintaining low background noise. This precision facilitated the successful visualization of D-DGLA and D-arachidonic acid-containing phospholipids in lung tissue, revealing their distinct localization compared with endogenous phospholipids. Our findings highlight bisallylic deuteration as a powerful tool for elucidating the in vivo dynamics of exogenous PUFAs through MS imaging techniques.