Xiao-Pen Lee

Solid-phase microextraction and liquid chromatography/mass spectrometry in drug analysis

Drug monitoring is highly dependent on the development of new analytical instruments or techniques. Solid-phase microextraction (SPME) is a new solvent-free sample preparation technique. There are two distinct techniques for the SPME method. Fiber SPME is a simple technique that integrates sampling, extraction, concentration and sample introduction into a single procedure. Fiber SPME has number of advantages such as simplicity, low cost and compatibility with gas chromatography (GC), GC/mass spectrometry (MS), liquid chromatography (LC) and LC/MS. Another technique known as in-tube SPME is an automated version of SPME that can be easily coupled to a LC autosampler for on-line sample preparation, separation and quantitation. Through these advantages, SPME has now gained a lot of interest in a broad field of analysis including drug, food, biological and pharmaceutical samples. This review is focused on the use of SPME and LC/MS for the analysis of drugs and metabolites from biological samples, including urine, blood and hair.

Detection of organophosphate pesticides in human body fluids by headspace solid-phase microextraction (SPME) and capillary gas chromatography with nitrogen-phosphorus detection

SummaryOrganosphosphate pesticides have been found extractable by headspace solid-phase microextraction (SPME), and the best conditions of their extraction from human whole blood and urine samples have been investigated. The body fluid samples containing nine pesticides (IBP, methyl parathion, fenitrothion, malathion, fenthion, isoxathion, ethion, EPN and phosalone) were heated at 100°C in a septum-capped vial in the presence of various combinations of acid and salts, and SPME fiber was exposed to the headspace of the vial to allow adsorption of the pesticides before capillary gas chromatography (GC) with nitrogen-phosphorus detection. The heating with distilled water/HCl/(NH4)2SO4/NaCl and with distilled water/HCl gave the best results for urine and whole blood, respectively. Recoveries of the nine pesticides were 0.8–10.6% except for phosalone (0.03%) for whole blood, and 3.8–40.2% for urine. The calibration curves for the pesticides showed linearity in the range of 50–400 ng/0.5 mL for whole blood except for malathion (100–400 ng/0.5 mL whole blood) and 7.5–120 ng/0.5 mL for urine except for phosalone (15–120 ng/0.5 mL urine) with detection limits of 2.2–40 ng/0.5 mL for whole blood and 0.8–12 ng/0.5 mL for urine.

A simple analysis of 5 thinner components in human body fluids by headspace solid-phase microextraction (SPME)

A simple method for the extraction of 5 thinner components from human whole blood and urine, using the headspace solid-phase microextraction (SPME) method is presented. After heating a vial containing the samples with 5 compounds (toluene, benzene,n-butyl acetate,n-butanol andn-isoamyl acetate) at 80°C, a polydimethylsiloxane-coated SPME fiber was exposed to the headspace of the vial to allow adsorption of the compounds. The fiber needle was then injected into a capillary gas chromatography (GC) port. The headspace SPME-GC gave intense peaks for each compound and a low level of background noise was seen only for whole blood. Recovery rates o the 5 compounds by use of the headspace SPME-GC were 50–70%. Reproducibility for headspace SPME-GC data were excellent for both body fluids. The calibration curves showed linearity in the range 2–100 ng/0.5 ml whole blood or urine. The detection limits of each compound were 1.1–2.4 ng/0.5 ml sample. The present results on the analysis of 5 thinner components by headspace SPME-GC suggest its applicability to a number of other volatile compounds in forensic toxicology.

Detection of ethanol in human body fluids by headspace solid-phase micro extraction (SPME)/capillary gas chromatography

SummaryEthanol has been found extractable from human whole blood and urine samples by headspace solid-phase micro extraction (SPME) with a Carbowax/divinylbenzene-coated fiber. After heating a vial containing the body fluid sample with ethanol, and isobutanol as internal standard (IS) at 70°C in the presence of (NH4)2SO4, a Carbowax/divinylbenzene-coated SPME fiber was exposed in the headspace of the vial to allow adsorption of the compounds. The fiber needle was then injected into a middle-bore capillary gas chromatography (GC) port. The headspace SPME-GC gave intense peaks for both compounds; a small amount of background noises appeared, but did not interfere with the detection of the compounds. Recoveries of ethanol and IS were 0.049 and 0.026% for whole blood, respectively, and 0.054 and 0.085% for urine, respectively. The calibration curves for ethanol showed excellent linearity in the range of 80–5000 mg L−1 for whole blood and 40–5000 mg L−1 for urine; the detection limits for both samples were 20 and 10 mg L−1, respectively. The data on actual determination of ethanol after the drinking of beer are also presented for two subjects.

Analysis of methanol or formic acid in body fluids by headspace solid-phase microextraction and capillary gas chromatography

Methanol and its metabolite formic acid have been found extractable from human whole blood and urine by headspace solid-phase microextraction (SPME) with a Carboxen/polydimethylsiloxane fiber. The headspace SPME for formic acid was carried out after derivatization to methyl formate under acidic conditions. The determinations of both compounds were made by using acetonitrile as internal standard (IS) and capillary gas chromatography (GC) with flame ionization detection. The headspace SPME-GC gave sharp peaks for methanol, methyl formate and I.S.; and low background noises for whole blood and urine samples. Extraction efficiencies were 0.25-1.05% of methanol and 0.38-0.84% formic acid for whole blood and urine. The calibration curves for methanol and formic acid showed excellent linearity in the range of 1.56 to 800 and 1.56 to 500 microg/0.5 ml of whole blood or urine, respectively. The detection limits were 0.1-0.5 microg/0.5 ml for methanol and 0.6 microg/0.5 ml for formic acid for both body fluids. The within-day relative standard deviations in terms of extraction efficiency for both compounds in whole blood and urine samples were not greater than 9.8%. By using the established SPME method, methanol and formic acid were successfully separated and determined in rat blood after oral administration of methanol.

Determination of tricyclic antidepressants in human plasma using pipette tip solid‐phase extraction and gas chromatography–mass spectrometry

A method for the simultaneous extraction of four tricyclic antidepressants from human plasma samples using pipette tip SPE with MonoTip C(18) tips is presented. Human plasma (0.1 mL) containing four tricyclic antidepressants (amitriptyline, amoxapine, imipramine, and trimipramine) and an internal standard (IS), protriptyline, was mixed with 0.4 mL of distilled water and 100 microL 1 M NaOH solution. After centrifugation of the mixture, the supernatant was extracted to the C(18) phase of the tip by 20 repeated aspirating/dispensing cycles using a manual micropipettor. The analytes retained in the tip were eluted with methanol by five repeated aspirating/dispensing cycles. Without evaporation and reconstitution, the eluate was directly injected into a gas chromatograph injector and detected by a mass spectrometer with SIM in the positive-ion electron impact mode. Recovery of the four antidepressants and IS spiked into human plasma was 80.2-92.1%. The regression equations for the four antidepressants showed excellent linearity in the range of 0.2-40 ng/0.1 mL. LODs and LOQs for the four drugs were 0.05-0.2 ng/0.1 mL and 0.2-0.5 ng/0.1 mL, respectively. Intra- and interday CVs for the four drugs in plasma were no greater than 9.5%.

Quantitative determination of phenothiazine derivatives in human plasma using monolithic silica solid-phase extraction tips and gas chromatography–mass spectrometry

Solid-phase extraction (SPE) using micropipette tips is a useful technique to prepare samples prior to mass spectrometry. However, most commercial SPE tips have loading capacities that are insufficient for quantitative determination. In this paper, we describe a rapid method for quantitative microanalysis of five phenothiazine derivatives, chlorpromazine, levomepromazine, promazine, promethazine and trimeprazine, using a recently introduced C(18) monolithic silica SPE tip, the MonoTip C(18), for extraction from human plasma. The drugs could be extracted within 5 min from 0.1-mL plasma samples, eluted with methanol, and the eluate injected directly into a gas chromatograph prior to mass spectrometry analysis. Only 0.7 mL of solvent was required for each step of the extraction process. The recoveries of the five phenothiazines spiked into plasma were 91-95% and the limits of quantification for each drug were between 0.25 and 2.0 ng/0.1 mL. The maximum intra- and inter-day coefficient of variation was 11%. The validated method was successfully used to quantify the plasma concentration of levemepromazine in a human subject after oral administration of the drug. This new method is expected to have wide applications as a pretreatment for the rapid, quantitative determination of drug concentrations in plasma samples.

Improved extraction of ethanol from human body fluids by headspace solid-phase microextraction with a carboxen-polydimethylsiloxane-coated fiber

SummaryEthanol can be extracted from human whole blood and urine by headspace solid-phase microextraction (SPME) with a Carboxen-polydimenhylsiloxane-coated fiber. Whole-blood containing ethanol, with isobutanol as internal standard (IS), is heated at 60 °C in the presence of (NH4)2SO4 and sodium dithionite. A Carboxen-polydimethylsiloxane-coated SPME fiber is then exposed to the headspace in the vial for 15 min and then injected into a mdium-bore capillary GC injection port. For urine the SPME procedure is essentially the same as that for whole blood except that the sodium dithionite is omitted. Recoveries of ethanol and IS were, respectively, 1.98 and 28.1% for whole blood, and 2.91 and 35.1% for urine. The calibration curves for ethanol were linear in the range 2.5–400 mg L−1 for whole blood and 0.5–400 mg L−1 for urine; the detection limits for whole blood and urine were 0.5 mg L−1 and 0.2 mg L−1, respectively. Data obtained for determination of ethanol after the drinking of beer are presented for two subjects. This SPME method with the new fiber for ethanol and IS resulted in sensitivity 40–1080 times greater than that obtained with the previous SPME method employing a Carbowax-divinylbenzene-coated fiber.

New and unique methods of solid-phase extraction for use before instrumental analysis of xenobiotics in human specimens

For forensic toxicological analysis of human specimens, extractive pretreatments are critical for successful analysis of target compounds. Nowadays, solidphase extraction (SPE) cartridges or columns packed with silica-type or polymer-type spherical particles are most widely used. In recent years, monolithic sorbent products, which have sponge-like structures, have been developed for SPE. The most significant advantage of the monolithic sorbent is that it allows very fast flow through the sorbent structure. Utilizing this property, pipette tips and spin columns packed with the monolithic sorbent have become commercially available. In this review, we present some details of SPE techniques using the monolithic pipette tips and spin columns, and discuss their advantages. The recent development of molecularly imprinted polymers (MIPs), which can selectively trap target compounds and act as an SPE sorbent, is also discussed with consideration of their advantages and disadvantages in SPE.

Simultaneous determination of β-blockers in human plasma using liquid chromatography–tandem mass spectrometry

A detailed procedure for the analysis of four beta-blockers, acebutolol, labetalol, metoprolol and propranolol, in human plasma by high-performance liquid chromatography (LC)-tandem mass spectrometry (MS-MS) using an MSpak GF column, which enables direct injection of crude plasma samples, is presented. Protein and/or macromolecule matrix compounds were eluted first from the column, while the drugs were retained on the polymer stationary phase of the MSpak GF column. The analytes retained on the column were then eluted into an acetonitrile-rich mobile phase using a gradient separation technique. All drugs showed base peak ions due to [M + H]+ ions by LC-MS with positive ion electrospray ionization, and the product ions were produced from each [M + H]+ ion by LC-MS-MS. Quantification was performed by selected reaction monitoring. The recoveries of the four beta-blockers spiked into plasma were 73.5-89.9%. The regression equations for all compounds showed excellent linearity in the range 10-1000 ng/mL of plasma, with the exception of propranolol (10-800 ng/mL). The limits of detection and quantification for each drug were 1-3 and 10 ng/mL, respectively, of plasma. The intra- and inter-day coefficients of variation for all drugs in plasma were not greater than 10.9%.