Hemp (Cannabis sativa) is a multi-use crop garnering newfound attention from researchers and consumers. While interest has emerged, a lack of substantiated research still exists regarding effects of adverse weather events on physiological health and secondary metabolite production of hemp. The aim of this experiment was to assess cold tolerance of hemp using the cultivars ‘FINOLA’ and ‘AutoCBD’. Effects of cultivar, plant age, cold acclimation, frequency of cold treatments, and intensity of cold treatments were all considered in regard to their influence on physiological stress, biomass, and cannabinoid profile. Few effects of sequential cold treatments were noted, and they were not moderated by cold acclimation, which tended to have negative effects across many responses. This detrimental effect of cold acclimation conditions was further observed in decreased total CBD % and total THC % compared to non-acclimated plants. These findings bear consideration when assessing the unpredictability of a changing climate’s effects on the heath and cannabinoid profile of hemp.