Stable-labeled analogues and reliable quantification of nonprotein biomarkers by LC-MS/MS

Background: The aim was to develop, and establish as suitable to begin assessment by full validation, a quantitative LC-MS/MS method for asparagine in human plasma. Therein, to utilize a stable-labeled analogue of asparagine to act as surrogate analyte, producing complete calibration curves and corresponding QC samples and another m/z distinct stable-labeled analogue to act as internal standard. Results: From two candidates, the surrogate analyte was selected through statistical comparisons of concentration response data and the resultant method employed protein precipitation and LC on an unmodified silica column with multiple reaction monitoring detection mode. The calibration range was 50-10,000 ng/ml. Conclusion: This method was successfully proven to meet the accuracy and precision acceptance criteria of current bioanalytical method validation guidelines.