The Sakaguchi color reaction for monosubstituted guanidino compounds was applied to the measurement of fl-guanidinopropionate and phosphorylated flguanidinopropionate. The phosphorylated derivative was measured as an increase in B-guanidinopropionate following incubation with 0.1 N HCI in a boiling-water bath for 10 min. After feeding rats 1% of fl-guanidinopropionic acid in their diet for 69 days, skeletal muscle, heart, liver, kidney, and spleen contained 5-10 /zmoles of a monosubstituted guanidino compound per gram wet weight of tissue. No B-guanidinopropionate was detected in brain or testes. Phosphorylated B-guanidinopropionate was found only in skeletal muscle (27 /~moles/g) and in heart (7 /~moles/g). Creatine hydrate (2%) added to the diet containing /3guanidinopropionic acid inhibited the accumulation of phosphorylated /3guanidinopropionate in the heart and partially inhibited its accumulation in skeletal muscle.
Creatine and phosphocreatine in skeletal muscles of rats can be replaced experimentally with their structural analogs, fl-guanidinopropionate (fl-GPA) and phosphorylated /3-guanidinopropionate (/3-GPAP) simply by adding fl-guanidinopropionic acid to the diet (1,2). This compound apparently is absorbed well through the gastrointestinal tract and is transported to muscle where it competes with creatine for entry (3). After entering muscle,/3-GPA can be phosphorylated by creatine kinase (2). The replacement of creatine and phosphocreatine by/3-GPA and /3-GPAP causes a change in metabolism, indicated by decreases in glucose-6-phosphate and ATP concentrations (2,4), and a change in the function of muscle, indicated by the absence of the staircase phenomenon in repetitively stimulated, hypoxic muscles (4). These changes suggest that rats fed fl-guanidinopropionic acid may serve as a useful model for study of the roles of creatine and phosphocreatine in the metabolism and function of skeletal muscle. To make optimal use of this model, it is desirable to have an accurate and rapid method for the measurement of fl-GPA and fl-GPAP in tissues. In the present report we describe such a method, which in principle is similar to the method of Ennor and Rosenberg (5) for measuring creatine and phosphocreatine: 196