In mammalian skeletal muscle there are four carboxylases involved in several biochemical processes like gluconeogenesis, tricarboxylic acid cycle anaplerosis, metabolism of fatty acids and metabolism of various amino acids. It has been shown that biotin deficiency reduces body weight at the expense of muscular mass. When necessary, the liver uses skeletal muscle protein to provide glucose and amino acids to organs in need of such compounds. In this paper we analyzed carboxylase specific activities in hind limb skeletal muscle of 3 weeks old BALB/c male mice, at 0, 1, 4, 7, and 14 days of a specific diet with different biotin concentrations. Biotin was used at 0.0, 1.8 or 98.2 mg per kg of food; and was referred to as biotin deficient, sufficient and supplemented, respectively. Water and food supply and consumption by the three groups of mice were the same. Therefore, the observed effects were directly related to biotin ingestion. The body weight of biotin supplemented mice was the same as the body weight of mice in the biotin sufficient group, while biotin deficiency caused body weight reduction after 7 days of biotin depletion. We found that the total protein concentration in the vastus lateralis muscle is associated with the biotin content in the diet. After 7 days, the muscle total protein content was lower in mice of the biotin deficient group while it was higher in the mice from the biotin supplemented group (P < 0.001). Of the four analyzed enzymes, only pyruvate carboxylase specific activity was reduced in both cases: by consuming the supplemented diet and by the lack of this vitamin. Our data show that PC and muscle metabolism are differentially altered by both, biotin excess and biotin deficiency. The mechanisms of these effects are currently under investigation.