Long-term sustainability of intensively managed forests depends on historical management and inherent soil productivity. However, long-term effects of prescribed fire and herbicide on forest floor and soil C and N pools are poorly understood, whereas benefits to biodiversity are well supported. Therefore, we investigated forest floor and mineral soil C and N responses to factorial combinations of mid-rotation dormant season prescribed fire and imazapyr herbicide (Arsenal®) in intensively managed mid-rotation pine (Pinus species) stands in east-central Mississippi. We used a randomized complete block design of six pine stands, each containing four 10-ha treatment plots receiving one of three treatments (herbicide only, prescribed fire only, both herbicide and fire) and a control at random. We applied herbicide via skidder in fall 1999 and prescribed fires using drip torches in winter, 2000, 2003, and 2006. During winter 2009, we sampled substrate using soil cores, a hammer core, and a fixed-area sampling frame for forest floor samples. Prescribed fire, with or without herbicide, reduced forest floor total N. However, C:N was only greater in burn + herbicide plots than controls, with independent treatments intermediate to treatment extremes. Considering the minimal effects of fire and herbicide use on soil nutrients and observed biodiversity benefits from these practices, prescribed fire with or without imazapyr herbicide does not seem to negatively impact long-term sustainability within similar managed pine landscapes of the southeastern United States.