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We compared the the nutrient and functional contents of two immature soybean (edamame) varieties, "Tamafukura" and "Yuagarimusume" (as the control), using matrix-assisted laser desorption/ionization (MALDI) quadrupole (Q) time-of-flight (TOF) mass spectrometry imaging (MSI). From soybean cross sections, we detected vitamin B1 (VB1; m/z 265.123) and vitamin B2 (VB2; m/z 377.146), and daidzein 7-O-glycoside as an isoflavone precursor (m/z 417.118) by tandem MS analysis. Results from MALDI-Q-TOF MSI indicated VB1 and VB2, and daidzein 7-O-glycoside as functional components localized in the cotyledon and hilum, respectively. The relative amounts of VB1 and daidzein 7-O-glycoside were 1.13-1.15-fold higher in Tamafukura than in Yuagarimusume. There was no difference in the relative amount of VB2 between the two types of edamame. These findings highlight the importance of not only determining the difference between soybean varieties, but also the value of visualizing the localization of their content. [ABSTRACT FROM AUTHOR]