Research And Application Of New Separation And Enrichment Technique In Pharmaceutical Analysis

The separation and analysis of trace components in complex matrix is agreat challenge faced by pharmaceutical analyzer. For the analysis ofcomponent in complex matrix, analyte needs to be not only separated but alsoenriched, aiming to increase detection sensitivity. The goal of traditionalsample preparation technique is usually to separate or extract analyte fromsample matrix. These methods are generally time-consuming,operation-tedious, unable to clean up matrix and with low sensitivity. Thesample preparation has become the crucial problem that restricts theimprovement of efficiency, sensitivity and accuracy in sample analysis. It isessential to seek for a simple, rapid, and without reducing sensitivity oranalyte-enriched sample preparation technique, which has become the focus inthe field of pharmaceutical analysis.This article is concentrated on the research and application of newseparation and enrichment technique in pharmaceutical analysis. New samplepretreatment methods combined with modern analytical technique, aiming toaccomplish direct injection without pretreatment and high detection sensitivity.The purpose of our study is to avoid the disadvantages of traditional samplepreparation technique and provide new simple, fast, sensitive and accuratedetection method for pharmaceutical analysis. Specific studies are as follows:(1) Traditional centrifugal ultrafiltrate technique (CF-UF) has beenwidely used in the analysis of unbound drug in plasma. The method is sosimple and fast that it involved only one step of centrifugation of plasmasamples to eliminate endogenous macromolecules in plasma and the obtainedultrafiltrate could be directly injected into HPLC for analysis. However, theaccuracy of traditional CF-UF in the analysis of marcromolecular matrix hasnot been reported. Hollow fiber centrifugal ultrafiltration technique(HFCF-UF) is established in our study and the comparison of HFCF-UF and CF-UF are conducted. With metformine (MET) as a model drug, factors thatinfluence the ultrafiltrate volume are studied and the measurement accuracy iscompared, aiming to perfect the ultrafitration theory and provide a theoreticalbasis for the correct selection and use of CF-UF technique. It is showed thatthe ultrafiltrate volume is free of centrifugal time, plasma condition andfreeze-thaw cycle times. The ultrafiltrate of HFCF-UF can be well controlledby the inner capacity of HF and the measurement is more accurate. Suitableultrafiltrate volume could be obtained by traditional CF-UF with strict controlof centrifugal time when the plasma condition is similar.In the pharmacokinetic and bioequivalence study, the measurement ofplasma concentration is the key of the study. The number of plasma samples islarge and sample preparation is tedious and time-consuming. The pretreatmenttime of all the work is above80%. Moreover, the plasma conditions aredifferent, sometimes influencing the reprocibility of measurement. Therefore,it is an urgent to establish a simple and rapid sample preparation method toshorten pretreatment time and ensure the measurement accuracy. In our study,the HFCF-UF method was successfully applied in the human pharmacokineticand bioequivalent experiment and satisfactory result was received. Theproposed method is equipped with simplicity, speed, good reproducibility,perfect recovery and high accuracy, providing a new tool for fast and accurateanalysis of clinic samples.(2) Both EPH and MEPH have a strong biological activity. It is theimportant evident data that is to review and to characterize the drug history inforensic justice practice. Blood, urine and hair are usually the most commonluused biological materials. With the body’s metablisom, the concentration ofanalyte decreases rapidly. Therefoe, it is of great significance to improve themethod sensitivity for extending the insepectation cycle and tracing themedication history of special population. A rapid and sensitive method hasbeen developed for the simultaneous determination of EPH and MEPH inurine samples by solid phase extraction-ultra performance liquidchromatography coupled with electrospray ionization tandem mass spectrometry (SPE-UPLC-ESI MS/MS). The detection sensitivity has reachedin0.01ng·mL-1and the insepectation cycle has been extended to7days. Theproposed method is simple and sensitive, which provides a new means fordetermination of trace analyte in biological matrix.Amphetamine-type stimulant (ATs) is one of the most commonlyscreened and detected illicit drugs in forensic laboratory. The concentration ofanalyte in biological material is usually low due to not timely sampling.Moreover, the analyte need to be extracted and enriched since the biologicalmatrix is complex. Traditional sample preparation methods including proteinprecipitation (PPT) and liquid liquid extraction (LLE) are not enough toremove the biological matrix that the detection sensitivity is low and unable tomeet the requirement of determination of trace component in complex matrix.We successfully apply the SPE-UPLC-MS/MS method for simultaneouslydetermination of12illicit drugs in whole blood. Three steps including acidwashing, alcohol washing and base washing are used to effectilvely removethe biological matrix. The present method possesses low matrix effect,accurate measurement and sensitive detection that reaching ppt level. Theapplication scope of the present method is wide and various ATs can bedetected simultaneously. The present method is proved to be reliable, robustand sensitive in forensic toxicological analysis.(3) Headspace gas chromatography (HS-GC) is to separate volatilecomponents from sample matrix and analyze by gas injection. Since it iscarried out in confined space, the loss of volatile components is avoided andthe detection noise induced by sample matrix is reduced. The HS-GC methodis sensitive, rapid and simple. However, it often needs to face the choice ofheadspace solvent. Traditional headspace solvent, for example, waterpossesses low boiling-point, which limits the headspace temperature andcannot be used for the analysis of volatile components with high boiling point.For DMF and DMSO, the pressure, which is formed due to the volatile,prevents the volatile component to enter into the headspace and affect thedetection sensitivity. Ionic liquids (ILs) emerge as a new headspace solvent. Its unique physicochemical properties, such as high solubility, almost novolatilization, good thermal stability and strong design features, greatlyexpand the application scope of HS-GC in pharmaceutical analysis. In ourstudy, ILs-HS-GC method is established to determine camphor (CPH) andeucalyptol (ECP) in rheo-camphoradin. The proposed method overcomes thedisadvantages of the traditional headspace solvent, effectively separates thehigh-boiling volatile components from complex matrix in Chinese medicinepreparation, and protects the chromatographic system from contamination ofnon-volatile components. It provides a simple and rapid alternative for theanalysis of high-boiling volatile components in traditional Chinese medicinepreparation. At the same time, we utilize the high solubility and successfullyapply it to simultaneously determinate7of organic residual solvents in KTZwhich is insoluble. The established method provides a fast and reliablealternative for the determination of organic residual solvent with high-boilingpoint in active pharmaceutical ingredient.