The bioactive polysaccharides (named ZPF1) from yam ( Dioscorea batatas) were chemically determined, suggesting repeating beta-1,4-mannan as mainly having a feature of acetylation on C2-OH and C3-OH, around 28%. The ZPF1 participated in the stimulation of murine wild-type macrophages predominantly in tumor necrosis factor-alpha (TNFalpha). Toll-like receptor 4 is proved to be one of the cellular receptors in ZPF1-mediated TNFalpha secretion. Reactive oxygen species transmission and PI3-kinase are found necessary for regulating TNFalpha secretion by ZPF1 stimulation. Moreover, we found that extracellular signal-regulated kinase 1/2, Jun N-terminal kinase 1/2, and p38 mitogen-activated protein kinase play important roles in the regulation of TNFalpha secretion in ZPF1-stimulated macrophages.