The dissociation equilibrium between uncharged local anesthetic lidocaine (LC) and charged local anesthetic LC (LC•H+) in a surface-adsorbed film was investigated by measuring the surface tension and pH of aqueous solutions of a mixture of hydrochloric acid and LC. The surface tension values decreased slightly with increasing total molality m t at 0≤X 2≤0.5, where X 2 is the mole fraction of LC in the mixture, while they decreased rapidly with increasing m t at 0.5<X 2≤1. It was shown from the pH measurements that almost all LC molecules were changed into LC•H+ ions by protonation at 0≤X 2≤0.5 and both forms coexisted only at 0.5<X 2≤1. The quantities of the respective LC and LC•H+ transferred from the aqueous solution to the adsorbed film, i.e., their surface densities, were calculated by applying the thermodynamic equations derived to the surface tension and pH data. A greater quantity of LC than LC•H+ existed in the adsorbed film at the coexisting composition. The partitioning behavior of LC and LC•H+ in the adsorbed film was characterized by three composition regions: (1) slight partitioning of low surface-active LC•H+ in the region at 0≤X 2≤0.5, (2) preferential partitioning of LC at 0.5<X 2<around 0.7, and (3) negative partitioning of LC•H+ at around 0.7≤X 2≤1. The present results clearly indicate that uncharged local anesthetics transfer into hydrophobic environments such as cell membranes more than charged ones.