Dual-column cation-exchange chromatographic method for β-aminoisobutyric acid and β-alanine in biological samples

A rapid automated chromatographic method has been developed for the quantitation of the nucleic acid catabolites β-aminoisobutyric acid and β-alanine in urine, serum, and other physiological fluids. The analyses were performed on a modified Beckman 121M amino acid analyzer with dual ion-exchange columns and the use of a single sodium citrate buffer (pH 4.38, 0.20 mol/liter). By carefully matching the elution pattern for the two ion-exchange columns and alternating use of these columns, analyses are completed every 40 min. The chromatography, regeneration, and equilibration of the two columns are precisely programmed, thus the detector sees only the elution of β-aminoisobutyric acid and β-alanine alternately from each column. Long-term precision and analytical recovery for the two metabolites in urine were 1.9 and 102%, and 3.3 and 10%, respectively. Their normal physiological values were determined in human serum and urine. Their excretion in the urine was also studied as a function of collection time, to validate a more convenient, less costly method of sampling. This study shows that randomly collected samples are acceptable when the concentration of the two metabolites are expressed in terms of creatinine excretion. In addition, the distribution of the free and conjugated forms of the two metabolites in urine and serum was studied. A preparative method was also developed for the quantitative isolation of β-aminoisobutyric acid from urine samples. The alternating dual-column technique may be applied to any ion-exchange chromatographic method where many analyses must be performed. This method is currently used in our laboratories for measuring these β-amino acids in urine and serum of patients with various types of cancers.