X-intrinsic proteins (XIPs) are a novel class of major intrinsic proteins found in diverse organisms. Recently, XIP genes have been reported to be involved in the transport of a wide range of hydrophobic solutes; however, the evolutionary forces driving their structural and functional divergence in plants are poorly understood. In the present study, comprehensive bioinformatics analyses were performed to gain insight into the molecular and evolutionary mechanisms driving this structural and functional diversification. Phylogenetic analyses have revealed the major lineage-specific expansions of XIP genes in plants. Within the eudicots, XIP genes have diverged into Asterid and Rosid-specific phylogenetic lineages and have also undergone several independent duplications during the course of evolution. Investigation of functional divergence at the protein level showed evidence for shifting evolutionary rate and/or altered constraints on the physiochemical properties of specific amino acid sites following gene duplication. Selection pressure analyses suggest that purifying selection is the predominant evolutionary force acting on the XIP gene subfamily, along with episodic positive selection. However, only a few amino acid sites were found to be subjected to such episodic positive selection. Furthermore, protein functional divergence analysis has identified critical amino acid residues, which must be validated by future experimental studies, that could provide new insights into the role of XIPs in transport of a wide range solutes of physiological importance.