Vacuum impregnation is used in the food industry to facilitate the impregnation of porous products with, e.g. firming, antioxidant, antimicrobial or cryoprotective agents. X-ray micro-tomography (mu CT) was used to study the process of vacuum impregnation in spinach leaves. Low (300 mbar absolute pressure) and mild vacuum (150 mbar absolute pressure) impregnation protocols were used to impregnate an isotonic solution of trehalose in the leaves and mu CT was used to make observations of the cross section of the impregnated samples and quantify their porosity. Results revealed that the free volume in the spongy mesophyll is easier to impregnate than the spaces around the palisade mesophyll. The low vacuum impregnation protocol provoked less impregnation close to the edge of the leaf than in its centre, probably accounting for an influence of the tissue structure on impregnation. The vacuum impregnation protocols tested in this investigation drastically decreased the proportion of large pores (>100 mu m) and increased the proportion of small pores (<50 mu m). The mild vacuum impregnation protocol, which was designed on the basis of measured apparent porosity, did not achieve full impregnation of the tissue. (C) 2016 Elsevier Ltd. All rights reserved.