The intracellular content and turnover of S-adenosyl-L-methionine (AdoMet) were measured in cultured human lymphoid cells. AdoMet levels were found to be 59 nmol/ml cell volume in exponentially growing WI-L2 lymphoblasts, 3.3-8.1 nmol/ml cell volume in unstimulated peripheral blood mononuclear cells, and 25-33 nmol/ml cell volume 24-48 h after the latter were stimulated with phytohemagglutinin. Increases in the AdoMet content of stimulated cells occurred within 2 h after addition of lectin. First order, pool turnover rates were of the same order of magnitude (0.029-0.091/min) for all three types of cultured cells, but owing to the differences in AdoMet content, absolute utilization rates differed markedly and were 4.5, 0.12-0.40, and 1.41-1.57 nmol/min/ml cell volume in WI-L2, unstimulated peripheral mononuclear cells, and lectin-stimulated peripheral mononuclear cells, respectively. Measurements of homocysteine accumulation in growth medium and of transsulfuration to cysteine indicate that a minimum of 82% of the AdoMet synthesized by WI-L2 is used for transmethylation. Remethylation of homocysteine by these cells could not be detected. AdoMet synthesis accounts for 20-23% of methionine utilization by WI-L2. Judging from the accumulation of homocysteine in the medium of phytohemagglutinin-stimulated peripheral mononuclear cells, a minimum of 38% of AdoMet synthesized must be used for transmethylation. Even though AdoMet utilization by unstimulated peripheral mononuclear cells is relatively small compared to that of stimulated cells and WI-L2, our data indicate that AdoMet turnover in such "resting" cells is three to five times that estimated for nonhepatic tissues. These findings may be relevant to the hypothesis that lymphoid cells are unusually sensitive to inhibition of transmethylation reactions.