Determination of cytidine deaminase activity in synovial fluid by HPLC.
A reversed-phase high-performance liquid chromatographic method for the determination of cytidine deaminase activity in synovial fluid is described. Diluted synovial fluid was incubated for 10 min at 56 degrees C with 0.4 mM cytidine. The protein in 5 vol of incubate was then precipitated using 1 vol of trichloroacetic acid (20% m/v) and the substrate, cytidine, and the product, uridine, were determined in the resultant supernatant. These substances were separated by reversed-phase HPLC using 0.05 M potassium dihydrogen orthophosphate (pH 6.5) containing methanol (3% v/v) and were detected at 280 nm. The enzyme activity was determined by measuring uridine formation. The effects of substrate concentration, pH and reaction temperature on uridine formation are described.
Herbert
K E
KE
Department of Rheumatology, Medical College, St. Bartholomew's Hospital, London, UK.
Scott
D L
DL
Perrett
D
D
D000641:Ammonia / Q000032:analysis; D002851:Chromatography, High Pressure Liquid; D003564:Cytidine Deaminase / Q000032:analysis*; D004795:Enzyme Stability; D006801:Humans; D015394:Molecular Structure; D009705:Nucleosides / Q000302:isolation & purification; D011506:Proteins / Q000032:analysis; D012680:Sensitivity and Specificity; D013582:Synovial Fluid / Q000201:enzymology*
D009705:Nucleosides; D011506:Proteins; D000641:Ammonia; D003564:Cytidine Deaminase