Decades of live-fire training exercises have left millions of acres of military training lands contaminated with various munitions constituents such as dinitrotoluene. Those that pose a threat to higher organisms due to their toxicity and mobility in the soil are of particular concern. Plants aid in the biodegradation and phytoextraction of contaminants, and site-specific ecotoxicity determinations are critical to inform effective remediation strategy. These ecotoxicity determinations are lacking in cold-adapted plants and would be very informative for contaminated training lands in cold regions. Therefore, we conducted a phytotoxicity study to determine the median effective concentration (EC50) of 2,4-dinitrotoluene (2,4-DNT) to four native Alaskan plant species in a sub-Arctic soil at two different temperatures. Plant species investigated were white spruce (Picea glauca), field locoweed (Oxytropis campestris), bluejoint grass (Calamagrostis canadensis), and Jacob’s ladder (Polemonium pulcherrimum). Seedling emergence, fresh plant mass, and dry plant mass were used to model plant response to 2,4-DNT contamination. White spruce was most tolerant to 2,4-DNT contamination (EC50 = 130.8 mg kg−1) and field locoweed was least tolerant (EC50 = 0.38 mg kg−1). In general, Arctic plant species were more vulnerable to 2,4-DNT when compared to plant types native to temperate or tropical regions.