A method for the direct injection and analysis of small volume human blood spots and plasma extracts containing high concentrations of organic solvents using revered-phase 2D UPLC/MS.

The emergence of micro sampling techniques holds great potential to improve pharmacokinetic data quality, reduce animal usage, and save costs in safety assessment studies. The analysis of these samples presents new challenges for bioanalytical scientists, both in terms of sample processing and analytical sensitivity. The use of two dimensional LC/MS with, at-column-dilution for the direct analysis of highly organic extracts prepared from biological fluids such as dried blood spots and plasma is demonstrated. This technique negated the need to dry down and reconstitute, or dilute samples with water/aqueous buffer solutions, prior to injection onto a reversed-phase LC system. A mixture of model drugs, including bromhexine, triprolidine, enrofloxacin, and procaine were used to test the feasibility of the method. Finally an LC/MS assay for the probe pharmaceutical rosuvastatin was developed from dried blood spots and protein-precipitated plasma. The assays showed acceptable recovery, accuracy and precision according to US FDA guidelines. The resulting analytical method showed an increase in assay sensitivity of up to forty fold as compared to conventional methods by maximizing the amount loaded onto the system and the MS response for the probe pharmaceutical rosuvastatin from small volume samples.

Investigation of microbore UPLC and nontraditional mobile phase compositions for bioanalytical LC-MS/MS.

BACKGROUND: The movement towards environmentally friendly or green chemistry solutions has gained more prominence recently in the scientific community. One way in which scientists can address this issue is to limit the use of hazardous chemicals in their everyday processes. Therefore, the focus of this study was on the utilization of microbore-scale chromatography and nontraditional alcoholic mobile phases as an alternative approach to traditional bioanalytical LC-MS/MS assay parameters. RESULTS: Replacement of the traditional narrowbore LC column with a microbore format reduced solvent consumption and produced a greater than threefold increase in S/N. The nontraditional alcoholic mobile phases, ethanol or isopropanol, produced either greater peak area counts, or S/N, for over half of the compounds evaluated, compared with the traditional organic mobile phases of acetonitrile and methanol. These nontraditional alcoholic mobile phases also showed improved capability in the removal of plasma phospholipid components from the chromatographic column. The ionizable background detected in each of the organic mobile phases utilized in this study produced a unique background that may or may not interfere with compounds undergoing analysis. CONCLUSION: The combination of microbore columns and nontraditional alcoholic mobile phases has been shown to produce effective, alternative method conditions to traditional bioanalytical LC-MS/MS method parameters.