Although arsenic (As) is a common pollutant worldwide, many questions about As metabolism in nonhyperaccumulator plants remain. Concentration-and tissue-dependent speciation and distribution of As was analyzed in the aquatic plant Ceratophyllum demersum to understand As metabolism in nonhyperaccumulator plants. Speciation was analyzed chromatographically (highperformance liquid chromatography-[inductively coupled plasma-mass spectrometry]-[electrospray ionization-mass spectrometry]) in whole-plant extracts and by tissue-resolution confocal x-ray absorption near-edge spectroscopy in intact shock-frozen hydrated leaves, which were also used for analyzing cellular element distribution through x-ray fluorescence. Chromatography revealed up to 20 As-containing species binding more than 60% of accumulated As. Of these, eight were identified as thiol-bound (phytochelatins [PCs], glutathione, and cysteine) species, including three newly identified complexes: Cys-As(III)-PC 2 , Cys-As-(GS) 2 , and GS-As(III)-desgly-PC 2 . Confocal x-ray absorption near-edge spectroscopy showed arsenate, arsenite, As-(GS) 3 , and As-PCs with varying ratios in various tissues. The epidermis of mature leaves contained the highest proportion of thiol (mostly PC)-bound As, while in younger leaves, a lower proportion of As was thiol bound. At higher As concentrations, the percentage of unbound arsenite increased in the vein and mesophyll of young mature leaves. At the same time, x-ray fluorescence showed an increase of total As in the vein and mesophyll but not in the epidermis of young mature leaves, while this was reversed for zinc distribution. Thus, As toxicity was correlated with a change in As distribution pattern and As species rather than a general increase in many tissues.Arsenic (As) is ubiquitously present, considered a nonessential metalloid for plants and animals, and poses serious health hazards to humans. High levels of As in soil and drinking water have been reported around the world, with the worst situations in south and southeast Asia, where millions of people are at risk of As poisoning through drinking water and food (Chowdhury et al., 2000;Smedley and Kinniburgh, 2002;Ohno et al., 2007). Recently, As induced yield loss; thus, a threat to the sustainability of food production has been recognized as the other side of the As calamity (Brammer and Ravenscroft, 2009;Panaullah et al., 2009). Considering the enormity of As contamination, phytoremediation or the development of crops that can be grown in contaminated environments without suffering from and accumulating As in edible parts seem to be the most appropriate strategies to counter the detrimental impacts of As. These strategies demand an understanding of the mechanistic details of As uptake, toxicity, and detoxification . The in planta distribution and speciation of As are important aspects in this direction.Inorganic arsenate [HAsO 4 22 or As(V)] and arsenite [H 2 AsO 3 2 or As(III)] are the most common forms of As in aquatic and terrestrial environments. Plants take up...