Background This study aims to detect alterations in the spatio-chromatic pseudophakic vision produced by multifocal diffractive intraocular lenses (IOLs) and provides a physical interpretation. Methods In vitro characterization of the imaging performance of two diffractive IOLs: AT LISA Tri (Zeiss) and FineVision (PhysIOL) in on-bench model eye illuminated with red (R, 625 nm), green (G, 530 nm) and blue (B, 455 nm) lights. We used the metrics: energy efficiency (EE), area under the modulation transfer function, longitudinal chromatic aberration (LCA), and halo intensity. Through-focus (TF) analysis and calculation of the expected defocus curve under white (W) daylight were included. In vivo visual acuity (VA) of 50 pseudophakics (60 eyes) was assessed under W, R, G, B lights at far and near. Two clinical experiments evaluated LCA and R, G, B TF-EE effects on pseudophakic vision and their relative importance. Results Clinical mean VA values under W light agreed with the predicted values at far and near for both IOLs. LCA measurements and R, G, B TF-EE curves were consistent with their lens design based on the 0th and 1st diffraction orders operative for far and near vision, respectively. LCA effects were compensated at near but noticed at far (-¿0.75 D under B light). We detected strong asymmetry in visual resolution depending on the object distance and the illuminating wavelength—red predominance at far, blue predominance at near—in consistency with the TF-EE measurements. Conclusions Diffractive multifocal IOL designs produce asymmetries in the spatio-chromatic vision of pseudophakics beyond the alterations strictly due to LCA. VA asymmetry for far/near object distance under R and B illumination is clinically detectable in subjects implanted with IOLs with 0th and 1st diffraction orders for far and near vision, respectively. Such VA asymmetry cannot be explained solely from the influence of defocus, as would be derived from a chromatic difference of power, but mainly from the wavelength dependence of the EE.