Nicotine content in honey is currently the major issue of concern to honey quality in Tanzania. This study was carried out to determine nicotine content in honey from tobacco and non-tobacco growing areas in Kigoma Region, Tanzania. Specifically, the study determined nicotine contents in honey and bee fodder samples. Fresh honey samples were collected from beehives within tobacco and non-tobacco growing areas and from vendors for laboratory tests. Results showed that nicotine content in the fresh honey which was collected from beehives located within tobacco growing areas is significantly higher than that collected from non-tobacco growing areas (0.46µg/g vs. 0.26µg/g, p <0.05). The honey samples from vendors in town and villages had nicotine contents of 0.41µg/g and 2.98µg/g, respectively. It was observed that most of the bee fodder tree species in Miombo woodland had traces of nicotine. Among the most important bee fodder tree species with the highest amount of nicotine were Brachystegia spiciformis (20.966µg/g), and Julbernardia globiflora (13.168µg/g). Meanwhile Vernonia colata, Combretum collinum and Combretum mole had the lowest nicotine contents of 0.836µg/g, 0.006µg/g, and 0.004µg/g, respectively. Generally, honey samples which were collected from all sources contained different amounts of nicotine. However, these contents were perceived to be tolerable for human health. The sampled bee forage tree species were also observed to have nicotine, which is associated with nicotine in the honey. The study recommends for the establishment of the national standards indicating the allowable amount of nicotine in honey for human consumption. Further research needs to be done in other areas and vegetation types in order to address this concern country- wise