Scaling up a microbore separation for semipreparative analysis: differential recoveries of radiolabeled amino acids.
Application of a high-sensitivity microbore system designed to separate and quantify nonderivatized amino acids by anion exchange chromatography and amperometric detection for determination of amino acid-specific activities in biological samples requires high capacity to recover sufficient labeled material for adequate count statistics. Scale up from a low (25-1000 pmol) to a high (500-15,000 pmol) working range was achieved by use of a thick working electrode gasket to reduce sensitivity and eliminate peak splitting and tailing and by modification of the wash procedure to eliminate carryover. Analysis of recoveries of labeled amino acids revealed that specific amino acids are either selectively retained on the column or partially degraded during analysis and that assessment of purities of labeled compounds and metabolic labeling patterns requires careful analysis of recoveries of labeled compounds in the appropriate eluate fraction.
Lasater
Amelia
A
Department of Neurology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, USA.
Cruz
Nancy F
NF
Cheng
Jun
J
Jandik
Petr
P
Dienel
Gerald A
GA
D000596:Amino Acids / Q000302:isolation & purification*; D002250:Carbon Radioisotopes; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation / Q000379:methods*; D007553:Isotope Labeling; D012680:Sensitivity and Specificity
D000596:Amino Acids; D002250:Carbon Radioisotopes