The effects of a single voluntary contraction of the quadriceps muscle group on phosphate incorporation into the phosphorylatable light chains (P-light chains) of fast and slow myosin isolated from the vastus lateralis muscle and potentiation of the electrically stimulated twitch tension was studied in intact human muscle. Twitch potentiation was maximal 20 s after the voluntary contraction. Thereafter, twitch potentiation declined, but was still significantly higher than pre-contraction values 2 min after the voluntary contraction. Phosphate incorporation into the P-light chain of fast myosin followed a similar time course to twitch potentiation, but no phosphate was incorporated into slow myosin P-light chains. These observations suggest that myosin light chain kinase activity is mainly associated with fast-twitch muscle fibers and, in agreement with previous studies, suggests that twitch potentiation associated with P-light chain phosphorylation is confined to the fast-twitch fibers of human muscle.