Automated in-tube solid-phase microextraction coupled with liquid chromatography/electrospray ionization mass spectrometry for the determination of beta-blockers and metabolites in urine and serum samples.
ABSTRACT The technique of automated in-tube solid-phase microextraction (SPME) coupled with liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) was evaluated for the determination of beta-blockers in urine and serum samples. In-tube SPME is an extraction technique for organic compounds in aqueous samples, in which analytes are extracted from the sample directly into an open tubular capillary by repeated draw/eject cycles of sample solution. LC/MS analyses of beta-blockers were initially performed by liquid injection onto a LC column. Nine beta-blockers tested in this study gave very simple ESI mass spectra, and strong signals corresponding to [M + H]+ were observed for all beta-blockers. The beta-blockers were separated with a Hypersil BDS C18 column using acetonitrile/methanol/water/acetic acid (15:15:70:1) as a mobile phase. To optimize the extraction of beta-blockers, several in-tube SPME parameters were examined. The optimum extraction conditions were 15 draw/eject cycles of 30 microL of sample in 100 mM Tris-HCl (pH 8.5) at a flow rate of 100 microL/min using an Omegawax 250 capillary (Supelco, Bellefonte, PA). The beta-blockers extracted by the capillary were easily desorbed by mobile-phase flow, and carryover of beta-blockers was not observed. Using in-tube SPME/LC/ESI-MS with selected ion monitoring, the calibration curves of beta-blockers were linear in the range from 2 to 100 ng/mL with correlation coefficients above 0.9982 (n = 18) and detection limits (S/N = 3) of 0.1-1.2 ng/mL. This method was successfully applied to the analysis of biological samples without interference peaks. The recoveries of beta-blockers spiked into human urine and serum samples were above 84 and 71%, respectively. A serum sample from a patient administrated propranolol was analyzed using this method and both propranolol and its metabolites were detected.
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ABSTRACT: In drug discovery and development, the quantification of drugs in biological samples is an important task for the determination of the physiological performance of the investigated drugs. After sampling, the next step in the analytical process is sample preparation. Because of the low concentration levels of drug in plasma and the variety of the metabolites, the selected extraction technique should be virtually exhaustive. Recent developments of sample handling techniques are directed, from one side, toward automatization and online coupling of sample preparation units. The primary objective of this review is to present the recent developments in microextraction sample preparation methods for analysis of drugs in biological fluids. Microextraction techniques allow for less consumption of solvent, reagents, and packing materials, and small sample volumes can be used. In this review the use of solid phase microextraction (SPME), microextraction in packed sorbent (MEPS), and stir-bar sorbtive extraction (SBSE) in drug analysis will be discussed. In addition, the use of new sorbents such as monoliths and molecularly imprinted polymers will be presented.01/2014; 2014:921350. DOI:10.1155/2014/921350
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ABSTRACT: This paper reviews the recent developments in bioanalysis sample preparation techniques and gives an update on basic principles, theory, applications and possibilities for automation, and a comparative discussion on the advantages and limitation of each technique. Conventional liquid-liquid extraction (LLE), protein precipitation (PP) and solid-phase extraction (SPE) techniques are now been considered as methods of the past. The last decade has witnessed a rapid development of novel sample preparation techniques in bioanalysis. Developments in SPE techniques such as selective sorbents and in the overall approach to SPE, such as hybrid SPE and molecularly imprinted polymer SPE, have been addressed. Considerable literature has been published in the area of solid-phase micro-extraction and its different versions, e.g. stir bar sorptive extraction, and their application in the development of selective and sensitive bioanalytical methods. Techniques such as dispersive solid-phase extraction, disposable pipette extraction and micro-extraction by packed sorbent offer a variety of extraction phases and provide unique advantages to bioanalytical methods. On-line SPE utilizing column-switching techniques is rapidly gaining acceptance in bioanalytical applications. PP sample preparation techniques such as PP filter plates/tubes offer many advantages like removal of phospholipids and proteins in plasma/serum. Newer approaches to conventional LLE techniques (salting-out LLE) are also covered in this review article.Biomedical Chromatography 01/2011; 25(1-2):199-217. DOI:10.1002/bmc.1560 · 1.66 Impact Factor
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ABSTRACT: The majority of human drugs enter aquatic systems after ingestion and subsequent excretion in the form of parent compounds and metabolites. Environmental exposure to drug metabolites has not been reported so far. The goal of the present study was to apply the in vitro method of biotransformation of compounds with S9 fraction to the ecotoxicological analysis. beta-adrenoceptor antagonist propranolol was metabolized with S9 rat liver fraction. The parent compound was quantified with HPLC, and the metabolites were identified with QToF MS. Propranolol was metabolized rapidly, during the first hour its level decreased by 80 and 50% of the initial 20 and 100 mg L(-1), respectively. Ten peaks were observed on the HPLC-RF chromatogram. Four peaks were identified with QToF MS/MS propranolol (m/z = 260), N-desisopropylpropranolol (m/z = 218), hydroxypropranolol (m/z = 276), and hydroxy N-desisopropranolol glycol (m/z = 235). Then the ecotoxicity of the reaction mixture was studied with two bioassays Spirotox with the protozoan Spirostomum ambiguum and Thamnotoxkit F with the anostracean crustacean Thamnocephalus platyurus. Propranolol is twofolds more toxic to Spirotox than to Thamnotoxkit F with 24 h-EC50 = 1.77 mg L(-1) and 24 h-LC50 = 3.86 mg L(-1), respectively. No statistically significant differences were found between the toxicity of the reaction mixtures after S9 biotransformation and the propranolol solution. These results indicate that the biological activity of the metabolites is similar to that of the parent drug. The presented method of in vitro biotransformation of drugs with S9 fraction followed by HPLC and ecotoxicity tests, may be used as screening method for evaluation of the toxicity of drug metabolites.Environmental Toxicology 02/2008; 23(1):52-8. DOI:10.1002/tox.20310 · 2.56 Impact Factor