Propionyl coenzyme A carboxylase and

The method described is suitable for the assay of four biotin-containing carboxylases pyruvate carboxylase, acetyl-coenzyme A carboxylase, propionyl-coenzyme A carboxylase, and 3-methylcrotonyl-coenzyme A carboxylase. The assays do not require radioisotopes and are suitable for use in clinical laboratories. 

Substrates and products are separated by reversed-phase chromatography at 45°C on a Nucleosil Qs column (4.6 mm X 250 mm). For assay of acetyl-coenzyme A carboxylase, propionyl-coenzyme A carboxylase, and 3-methylcrotonyl-coenzyme A carboxylase, a linear gradient from solvent A (0.1 M sodium phosphate buffer, pH 2.1) to solvent B (methanol-solvent A, 80 20, v/v) was applied in 15 minutes at a flow rate of 1.5 mL/min. Quantitation was based on the absorbance of the product (malonyl-CoA, methylmalonyl-CoA, and 3-methylglutaconyl-CoA, respectively) at 260 nm. For assay of pyruvate carboxylase, pyruvate was separated by isocratic elution using 0.1 M sodium phosphate buffer (pH 2.1) containing 0.1 M sodium sulfate. Quantitation was based on the disappearance of pyruvate as followed at 210 nm. 

---