Online Enrichment And Retention Mechanism Of RAM Column For The Analytes In Biological Fluids

Biological sample pretreatment is a crucial step for vivo pharmaceutical analysis; traditional preparation methods are protein precipitation, off-line solid phase extraction (SPE), liquid-liquid extraction (LLE), etc. These methods are offline operation and require large volumes of organic reagents, resulting in inaccurate results due to the analyte is diluted, lost and contaminated. Moreover, the biological sample contains a lot of protein and other biological macromolecules, which will pollute the SPE column, not commonly used in chromatography. One class of biocompatible sample preparation materials that enables the direct injection of biological fluids is the restricted access material (RAM), which having two different surfaces, the outer surface simultaneous with size-exclusion and hydrophilic, the inner surface bonded with different functional groups, only small molecules could penetrate into the internal surface and be retained. Compared to protein precipitation and SPE, RAM avoids loss and contamination of the analytes in the biological sample. RAM has been applied to direct injection of biological sample.However, they could not completely meet the requirement of biological analysis due to trace amount drugs in biological fluids, although the high sensitivity of high performance liquid chromatography-mass spectrometry (HPLC-MS), there is still considered to be highly sophisticated and expensive pieces. So, conventional HPLC method is typically more robust, cheaper and easier to operate, which still plays a key role in quantitative bioanalyses. In order to improve the sensitivity of HPLC method, the paper developed and optimized a column-switching HPLC method with homemade RAM column, through the large volume direct injection (LVI) to detect the concentration of drug in a variety of biological fluids (plasma, urine, cerebrospinal fluid), and explore the enrichment performance and retention mechanism of RAM column, provides a simple, rapid and accurate on-line analytical method for the biological samples, and also provides experimental basis for the synthesis of more type of RAM.