RESUMEN
Injection molded poly(methylmethacrylate) (IM-PMMA), chips were evaluated as potential candidates for capillary electrophoresis disposable chip applications. Mass production and usage of plastic microchips depends on chip-to-chip reproducibility and on analysis accuracy. Several important properties of IM-PMMA chips were considered: fabrication quality evaluated by environmental scanning electron microscope imaging, surface quality measurements, selected thermal/electrical properties as indicated by measurement of the current versus applied voltage (I-V) characteristic and the influence of channel surface treatments. Electroosmotic flow was also evaluated for untreated and O2 reactive ion etching (RIE) treated surface microchips. The performance characteristics of single lane plastic microchip capillary electrophoresis (MCE) separations were evaluated using a mixture of two dyes-fluorescein (FL) and fluorescein isothiocyanate (FITC). To overcome non-wettability of the native IM-PMMA surface, a modifier, polyethylene oxide was added to the buffer as a dynamic coating. Chip performance reproducibility was studied for chips with and without surface modification via the process of RIE with O2 and by varying the hole position for the reservoir in the cover plate or on the pattern side of the chip. Additionally, the importance of reconditioning steps to achieve optimal performance reproducibility was also examined. It was found that more reproducible quantitative results were obtained when normalized values of migration time, peak area and peak height of FL and FITC were used instead of actual measured parameters.
Asunto(s)
Electroforesis Capilar/instrumentación , Dispositivos Laboratorio en un Chip , Electroósmosis , Fluoresceína/aislamiento & purificación , Fluoresceína-5-Isotiocianato/aislamiento & purificación , Polimetil Metacrilato , Reproducibilidad de los ResultadosRESUMEN
A series of 5-keto-substituted 7-tert-buty1-2,3-dihydro-3,3- dimethylbenzofurans (DHDMBFs) were prepared and evaluated as potential nonsteroidal antiinflammatory and analgesic agents. Interest in this class of compounds arose when a DHDMBF was found to be an active metabolite of the di-tert-butylphenol antiinflammatory agent tebufelone. We have now found that a variety of 5-keto-substituted DHDMBFs have good in vivo antiinflammatory and analgesic activity after oral administration. These compounds inhibit both cyclooxygenase (COX) and 5-lipoxygenase (5-LOX) in vitro. The cyclooxygenase inhibition was found to be selective for the cyclooxygenase-2 isoform, and this combination of COX-2/5-LOX inhibition may be responsible for the gastrointestinal safety of compounds such as 30.
Asunto(s)
Antiinflamatorios/síntesis química , Benzofuranos/síntesis química , Inhibidores de la Ciclooxigenasa/síntesis química , Inhibidores de la Lipooxigenasa , Animales , Antiinflamatorios/química , Antiinflamatorios/farmacología , Benzofuranos/farmacología , Plaquetas/efectos de los fármacos , Plaquetas/enzimología , Carragenina/efectos adversos , Inhibidores de la Ciclooxigenasa/química , Inhibidores de la Ciclooxigenasa/farmacología , Edema/inducido químicamente , Edema/tratamiento farmacológico , Humanos , Masculino , Ratas , Ratas Endogámicas Lew , Ratas Sprague-DawleyRESUMEN
A series of 5-keto-substituted 7-tert-buty1-2,3-dihydro-3,3- dimethylbenzofurans (DHDMBFs) were found to be nonsteroidal antiinflammatory and analgesic agents. These compounds are inhibitors of 5-lipoxygenase (5-LOX) and cyclooxygenase (COX) with selectivity for the COX-2 isoform. A series of analogues were prepared to investigate the scope of this lead. Five ketone side chains from active DHDMBFs were used to investigate the effects of changes in the DHDMBF "core": the size and identity of the heterocycle and the substituent requirements of the heterocycle and phenyl ring. Biological testing showed that a variety of structural changes can be accommodated, but no structure was clearly superior to the DHDMBF structure.
Asunto(s)
Analgésicos/síntesis química , Antiinflamatorios/síntesis química , Benzofuranos/síntesis química , Inhibidores de la Ciclooxigenasa/síntesis química , Inhibidores de la Lipooxigenasa , Analgésicos/química , Analgésicos/farmacología , Animales , Antiinflamatorios/química , Antiinflamatorios/farmacología , Benzofuranos/química , Benzofuranos/farmacología , Plaquetas/efectos de los fármacos , Plaquetas/enzimología , Carragenina/efectos adversos , Inhibidores de la Ciclooxigenasa/química , Inhibidores de la Ciclooxigenasa/farmacología , Edema/inducido químicamente , Edema/tratamiento farmacológico , Humanos , Ratas , Ratas Endogámicas Lew , Ratas Sprague-DawleyRESUMEN
We report an expansion of the scope of our initial discovery that 5-keto-substituted 7-tert-butyl-2,3-dihydro-3,3-dimethylbenzofurans (DHDMBFs) are antiinflammatory and analgesic agents. Several other functional groups have been introduced at the 5 position: amides, amidines, ureas, guanidines, amines, heterocycles, heteroaromatics, and heteroaryl ethenyl substituents in the 5 position all provide active compounds. These compounds are dual cyclooxygenase (COX) and 5-lipoxygenase (5-LOX) inhibitors. They inhibit both COX-1 and COX-2 with up to 33-fold selectivity for COX-2.
Asunto(s)
Analgésicos , Antiinflamatorios no Esteroideos , Benzofuranos , Inhibidores de la Ciclooxigenasa , Inhibidores Enzimáticos , Isoenzimas/metabolismo , Inhibidores de la Lipooxigenasa , Prostaglandina-Endoperóxido Sintasas/metabolismo , Analgésicos/síntesis química , Analgésicos/química , Analgésicos/farmacología , Animales , Antiinflamatorios no Esteroideos/síntesis química , Antiinflamatorios no Esteroideos/química , Antiinflamatorios no Esteroideos/farmacología , Benzofuranos/síntesis química , Benzofuranos/química , Benzofuranos/farmacología , Carragenina/toxicidad , Ciclooxigenasa 1 , Ciclooxigenasa 2 , Inhibidores de la Ciclooxigenasa 2 , Inhibidores de la Ciclooxigenasa/síntesis química , Inhibidores de la Ciclooxigenasa/química , Inhibidores de la Ciclooxigenasa/farmacología , Edema/inducido químicamente , Edema/tratamiento farmacológico , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Humanos , Proteínas de la Membrana , Ratas , Relación Estructura-ActividadRESUMEN
Rapid, sensitive and selective methods were developed for the determination of dextromethorphan and its major metabolite, dextrorphan, in human plasma using liquid chromatography/tandem mass spectrometry (LC/MS/MS). Plasma samples spiked with stable-isotope internal standards were prepared for analysis by a liquid-liquid back-extraction procedure. Dextromethorphan and dextrorphan were chromatographed on a short reversed-phase column, using separate isocratic mobile phase conditions optimized to elute each compound in approximately 1.1 min. For both analytes, calibration curves were obtained over four orders of magnitude and the limit of quantitation was 5 pg ml-1 using a 1 ml plasma sample volume. The accuracy across the entire range of spiked DEX and DOR concentrations was, in general, within 10% of the spiked value. The precision was generally better than 6% for replicate sample preparations at levels of 50 pg ml-1 or higher and typically better than 12% at levels below 50 pg ml-1. The method was applied for the evaluation of the pharmacokinetic profiles of dextromethorphan and dextrorphan in a human volunteer following peroral administration of a commercially available cough formulation.
Asunto(s)
Antitusígenos/sangre , Dextrometorfano/sangre , Dextrorfano/sangre , Antagonistas de Aminoácidos Excitadores/sangre , Adulto , Antitusígenos/farmacocinética , Calibración , Cromatografía Líquida de Alta Presión , Dextrometorfano/farmacocinética , Dextrorfano/farmacocinética , Antagonistas de Aminoácidos Excitadores/farmacocinética , Congelación , Humanos , Masculino , Espectrometría de Masas , SolucionesRESUMEN
Two clinical studies were conducted to determine the relative amounts of ketorolac detectable locally in the gingival crevicular fluid (GCF) and systemically in plasma after oral, topical drug administration. The rinse study compared topical administration of three concentrations of ketorolac tromethamine (0.1%, 0.5%, and 0.01%) in oral rinse formulations administered topically and a perorally administered capsule (10 mg), and the dentifrice study compared two concentrations of ketorolac in dentifrice formulations (0.15% and 1.0%) with a 0.1% oral rinse, all treatments administered topically. The dose-corrected systemic availability of the three oral rinses evaluated in the rinse study relative to the peroral capsule was about 15%. However, the ratios of the observed maximum GCF ketorolac concentration to maximum plasma ketorolac concentration ranged from 22 to 49, compared to less than 1 for the peroral ketorolac capsule. Using this ratio as an estimate of the ability of a treatment to target the drug to the gingival tissue, these data indicate that the ketorolac oral rinses achieved greater delivery of drug to the gingival tissue (presumed site of action for periodontitis) with a lower systemic drug load than peroral administration of a ketorolac capsule. The dose-corrected relative systemic bioavailabilities for the dentifrice treatments with respect to the 0.1% rinse in the dentifrice study were 59.2% and 86.4% for the 1.0% and 0.15% dentifrices, respectively, indicating that significantly less ketorolac was systemically available from the two dentifrices relative to the oral rinse. The relative bioavailabilities of ketorolac in the GCF after dosing with the dentifrice formulations with respect to the rinse were 89.1% for the 1.0% dentifrice and 19.7% for the 0.15% dentifrice. Thus, the 1.0% dentifrice appears to provide statistically equivalent levels of ketorolac to the gingival tissue as the 0.1% oral rinse with significantly less systemic exposure. The T1/2 of ketorolac in the GCF was about 0.5 h for all three treatments, which is significantly less than the plasma half-life of about 5.3 h. These data suggest that GCF levels of ketorolac should remain above the IC50 for PGE2-stimulated IL-1 bone resorption for about 7 h following treatment, assuming continuation of the first-order elimination observed over the first two postdosing hours. We conclude that oral rinses and dentifrices are effective and preferred vehicles for administration of ketorolac for use in treatment of periodontitis.
Asunto(s)
Dentífricos , Líquido del Surco Gingival/química , Antisépticos Bucales , Tolmetina/análogos & derivados , Administración Tópica , Adulto , Anciano , Método Doble Ciego , Semivida , Humanos , Ketorolaco , Persona de Mediana Edad , Placebos , Tolmetina/sangre , Tolmetina/farmacocinética , Tolmetina/uso terapéuticoRESUMEN
Systemic non-steroidal anti-inflammatory drugs (NSAIDs) have been shown to reduce alveolar bone loss in periodontitis. This study assesses the efficacy of a topical NSAID rinse, containing ketorolac tromethamine as the active agent. Adult periodontitis patients (n = 55) were studied in this 6-month randomized, double blind, parallel, placebo and positive-controlled study. Each patient had a least 3 sites at high risk for bone loss as assessed by low dose bone scan. Groups, balanced for gender, were assigned to one of three regimens: bid ketorolac rinse (0.1%) with placebo capsule; 50 mg bid flurbiprofen capsule (positive control) with placebo rinse; or bid placebo rinse and capsule. Prophylaxes were provided every 3 months. Monthly examinations assessed safety, gingival condition, and gingival crevicular fluid PGE2. Standardized radiographs were taken at baseline and at 3 and 6 months for digital subtraction radiography. A significant loss in bone height was observed during the study period in the placebo group (-0.63 +/- 0.11; P < 0.001), but not in the flurbiprofen (-0.10 +/- 0.12; P = 0.40) or ketorolac rinse (+0.20 +/- 0.11 mm; P = 0.07) groups. Nested ANOVA revealed that ketorolac and flurbiprofen groups had less bone loss (P < 0.01) and reduced gingival crevicular fluid PGE2 levels (P < 0.03) compared to placebo. ANOVA suggests (P = 0.06) that ketorolac rinse preserved more alveolar bone than systemic flurbiprofen at the dose regimens utilized. These data indicate that ketorolac rinse may be beneficial in the treatment of adult periodontitis.
Asunto(s)
Pérdida de Hueso Alveolar/prevención & control , Antiinflamatorios no Esteroideos/uso terapéutico , Antisépticos Bucales/uso terapéutico , Periodontitis/tratamiento farmacológico , Tolmetina/análogos & derivados , Trometamina/análogos & derivados , Adulto , Pérdida de Hueso Alveolar/diagnóstico por imagen , Pérdida de Hueso Alveolar/etiología , Análisis de Varianza , Índice de Placa Dental , Dinoprostona/análisis , Método Doble Ciego , Femenino , Flurbiprofeno/uso terapéutico , Líquido del Surco Gingival/química , Humanos , Interleucina-1/análisis , Ketorolaco Trometamina , Masculino , Persona de Mediana Edad , Índice Periodontal , Periodontitis/complicaciones , Radiografía , Estadísticas no Paramétricas , Técnica de Sustracción , Tolmetina/administración & dosificación , Tolmetina/efectos adversos , Tolmetina/sangre , Tolmetina/uso terapéutico , Resultado del Tratamiento , Trometamina/administración & dosificación , Trometamina/efectos adversos , Trometamina/uso terapéuticoRESUMEN
A chemical and a stable-isotope analog, p-fluoroketorolac and [18O3]ketorolac respectively, were directly compared for applicability as internal standards for the determination of ketorolac in plasma samples using gas chromatography/mass spectrometry (GC/MS) with selective-ion-monitoring detection, following derivatization to form the methyl esters. This comparison involved analyzing ketorolac calibration standards and spiked plasma samples that contained both internal standard candidates. The response for ketorolac and each internal standard was monitored simultaneously and electronically integrated peak heights were obtained. Thus, for each analysis performed, a response ratio was obtained for each internal standard relative to an identical ketorolac response. Linearity of response for ketorolac calibration standards and accuracy for spiked plasma sample analysis were compared using each internal standard. The use of [18O3]ketorolac as the internal standard provided superior accuracy data for the analysis of ketorolac in plasma samples.
Asunto(s)
Analgésicos no Narcóticos/análisis , Antiinflamatorios no Esteroideos/análisis , Tolmetina/análogos & derivados , Cromatografía de Gases y Espectrometría de Masas/métodos , Ketorolaco , Isótopos de Oxígeno , Estándares de Referencia , Reproducibilidad de los Resultados , Tolmetina/análisis , Tolmetina/sangreRESUMEN
A method for the measurement of human parathyroid hormone fragment 1-34 (PTH1-34) in dog plasma was developed by modification of a commercially available immunodiometric assay (IRMA) designed for the determination of rat PTH1-34 in serum. Major modifications were made to the assay in order to circumvent significant problems encountered during the validation of the IRMA. PTH1-34 was found to be highly unstable in both rat serum and dog serum and plasma at room temperature, in contrast to literature reports. The addition of a protease inhibitor cocktail to serum or plasma samples was necessary to prevent in-vitro proteolytic degradation of human PTH1-34 prior to analysis. Additionally, plasma was chosen over serum as the sample matrix to expedite the separation of samples from cells, minimizing proteolytic degradation prior to the addition of cocktail. Finally, the reported 100% cross-reactivity between rat and human PTH1-34 was found to be only 65%; therefore, a human PTH1-34 standard was substituted for the rat standard. These modifications allowed the accurate measurement of human PTH1-34 in plasma obtained from dogs dosed intravenously and subcutaneously with human PTH1-34 using a commercially available kit.
Asunto(s)
Hormona Paratiroidea/sangre , Fragmentos de Péptidos/sangre , Teriparatido/sangre , Animales , Reacciones Cruzadas , Perros , Estabilidad de Medicamentos , Congelación , Humanos , Ensayo Inmunorradiométrico/métodos , Inyecciones Intravenosas , Inyecciones Subcutáneas , Hormona Paratiroidea/farmacocinética , Fragmentos de Péptidos/farmacocinética , Ratas , Reproducibilidad de los Resultados , TemperaturaRESUMEN
Peptidyl fluoromethyl ketones (PFMK) are irreversible inhibitors of cathepsin B, a cysteine proteinase thought to be involved in the degradation of cartilage. It has been speculated that PFMK inhibitors may metabolize in rodents to form fluoroacetate (FAC), an extremely toxic poison. A highly selective and sensitive separation and detection scheme was developed to measure trace levels of FAC in rat tissues following PFMK dosing. The procedure consisted of extracting FAC from tissue and spiking the extract with [18O]2-fluoroacetate (18O-FAC) as an internal standard. FAC and 18O-FAC were further isolated from matrix components using ion-exchange, solid-phase extraction. The pentafluorobenzyl esters of FAC and 18O-FAC were formed to facilitate the chromatographic separation. Two-dimensional gas chromatography coupled with selected-ion-monitoring detection provided the final measurement. The assay had a limit of detection of 2 ng FAC per g tissue, and was capable of accurately quantitating as little as 10 ng FAC per g tissue with a S/N ratio of 40:1. Linearity was established over two orders of magnitude, from 2-500 ng ml-1, with 5 microliters injected on-column. The method was used to demonstrate that FAC was formed in rats following dosing with Z-Phe-Ala-CH2-F, a PFMK cathepsin enzyme inhibitor.
Asunto(s)
Inhibidores de Cisteína Proteinasa/farmacocinética , Dipéptidos/farmacocinética , Fluoroacetatos/análisis , Cetonas/farmacocinética , Rodenticidas/análisis , Animales , Calibración , Fluoroacetatos/metabolismo , Cromatografía de Gases y Espectrometría de Masas , Hígado/química , Hígado/metabolismo , Ratas , Rodenticidas/metabolismoRESUMEN
Vanilloids are a class of compounds structurally related to capsaicin, the pungent principle of hot peppers, which are under development as a novel class of analgesics. Vanilloids undergo extensive first-pass metabolism when dosed orally to rats and mice. These compounds, as well as capsaicin, would be anticipated to be susceptible to three major routes of metabolism: (omega, beta)-oxidation of the alkyl side chain, hydrolysis of the amide bond and conjugation of the phenolic group. Olvanil [N-(3-methoxy-4-hydroxybenzyl)oleamide], radiolabelled with either 14C at the benzylic carbon or 3H in the oleyl side chain, was studied in various in vitro, in situ and in vivo metabolism models to determine the major route(s) of intestinal and hepatic metabolism in rats for this new class of compounds. Models used in metabolism studies included isolated hydrolytic enzymes, cell-free intestinal and liver supernatants, hepatocytes, enterocytes, perfused intestine and whole animal studies. Reversed-phase liquid chromatography (LC) with on-line radiochemical detection was used to examine the metabolic profiles from the different models. The major metabolic route for olvanil in both the intestine and the liver was found to be hydrolysis of the amide bond. The benefits of selective 14C and 3H labels in conjunction with LC with on-line radiochemical detection are discussed.
Asunto(s)
Analgésicos/metabolismo , Antiinflamatorios no Esteroideos/metabolismo , Capsaicina/análogos & derivados , Animales , Capsaicina/metabolismo , Cromatografía Líquida de Alta Presión , Mucosa Intestinal/metabolismo , Hígado/metabolismo , Masculino , Perfusión , Radioquímica , Ratas , Ratas EndogámicasAsunto(s)
Estradiol/análisis , Estriol/análisis , Inmunoensayo , Inmunoglobulina G/análisis , Animales , Bovinos , Haptenos/análisis , Polarografía , PotenciometríaRESUMEN
OBJECTIVE: Micro-computed tomography (microCT) imaging has the potential to allow the three-dimensional (3D) visualization of cartilage morphology. However, cartilage intensity on a microCT image is weak because cartilage does not strongly attenuate X-rays. This work was designed to demonstrate that exposure of cartilage to charged gadolinium compounds modifies the intensity to allow an improved visualization of cartilage morphology and the determination of proteoglycan content. DESIGN: Trypsin was used to deplete proteoglycan in bovine nasal cartilage disks. Disks were then exposed to Gd(3+), gadopentetate (Gd-DTPA(2-)), or gadoteridol (Gd-HP-DO3A), and imaged with microCT. The intensities of the disks were measured from the images and compared to the actual proteoglycan content determined with a dimethylmethylene blue assay. RESULTS: Treatment of naïve disks with 200 mM Gd(3+) for 24h at room temperature produced a 2.8-fold increase in intensity on microCT images. Similar treatment with 200 mM Gd-DTPA(2-) produced a 1.4-fold increase. After 2h of trypsin treatment at room temperature, the intensities of cartilage disks exposed to 20 0mM Gd(3+) decreased by 12%. Conversely, the intensities of trypsin-treated disks exposed to 200 mM Gd-DPTA(2-) increased by 15%. Trypsin treatment caused a 4% increase in the intensities of disks exposed to neutral Gd-HP-DO3A. The correlation between proteoglycan content and the microCT intensity of cartilage treated with Gd(3+) was very good (r(2)=0.81). CONCLUSIONS: Gadolinium and microCT allow an improved 3D visualization of cartilage and quantification of its proteoglycan content.
Asunto(s)
Cartílago/química , Proteoglicanos/análisis , Animales , Cartílago/diagnóstico por imagen , Bovinos , Medios de Contraste , Gadolinio , Gadolinio DTPA , Compuestos Heterocíclicos , Imagenología Tridimensional/métodos , Tabique Nasal/química , Tabique Nasal/diagnóstico por imagen , Compuestos Organometálicos , Tomografía Computarizada por Rayos X/métodosRESUMEN
Supercritical carbon dioxide was used for the direct extraction of drugs from plasma prior to analysis. The supercritical fluid was directly passed through plasma samples spiked with either a neutral (flavone) or an acidic (ketorolac) drug. The addition of an antifoam agent to the plasma prior to extraction was required to avoid restrictor plugging caused by denaturation of the plasma proteins by the supercritical fluid. The effluent from the extraction cell was bubbled through a small volume of methanol or into an empty tube to trap the extracted drug. The effect of extraction pressure and time on absolute recovery were examined. The absolute recovery, selectivity, precision and accuracy of the supercritical fluid extraction approach was compared to conventional liquid-liquid extraction using reversed-phase HPLC with ultraviolet detection.
Asunto(s)
Dióxido de Carbono , Flavonoides/sangre , Preparaciones Farmacéuticas/aislamiento & purificación , Tolmetina/análogos & derivados , Animales , Radioisótopos de Carbono , Fenómenos Químicos , Química Física , Cromatografía Líquida de Alta Presión , Perros , Flavonas , Flavonoides/química , Hexanos , Ketorolaco , Cloruro de Metileno , Sensibilidad y Especificidad , Tecnología Farmacéutica , Tolmetina/sangre , Tolmetina/químicaRESUMEN
A new sample preparation technique, solid-phase extraction with supercritical fluid elution, was developed for the selective isolation of ultratrace levels of drugs from plasma. Plasma samples spiked with a drug were applied to octadecylsilane cartridges and the cartridges were then washed, briefly dried and directly fitted into cells for subsequent supercritical fluid elution. The absolute recovery was studied by using a radiolabeled model compound. The extraction selectivity was examined by chromatographing the extracts with a reversed-phase high-performance liquid chromatographic method with ultraviolet detection. The effects of extraction pressure and the length of capillary restrictors on drug recovery were examined in order to determine the optimal conditions for supercritical fluid elution. The performance of the method was compared to that of conventional solid-phase extraction in terms of recovery, selectivity, precision and accuracy of analysis. Flavone was used as the model compound and dog plasma as the biological matrix for these studies.
Asunto(s)
Cromatografía Liquida/métodos , Flavonoides/sangre , Animales , Cromatografía Líquida de Alta Presión , Perros , Flavonas , Reproducibilidad de los Resultados , Espectrofotometría Ultravioleta , Oligoelementos/sangreRESUMEN
In these competitive and "sandwich"-type heterogeneous enzyme immunoassays, based on liquid chromatography with electrochemical detection, rabbit immunoglobulin G is used as a model compound. Alkaline phosphatase (EC 3.1.3.1), the labeling enzyme, catalyzes conversion of phenyl phosphate to phenol. After separation on an octyldecylsilane column, the enzyme-generated phenol is detected in a thin-layer cell at a carbon-paste working electrode. The detection limit for phenol is 5.0 nmol/L. The electrode response varies linearly with concentration over a range of three orders of magnitude. For the sandwich-type assay procedure the detection limit is 10 ng/L; the linearity ranges over four orders of magnitude. The detection limit of the competitive immunoassay is 5 micrograms/L. The dynamic range spans two orders of magnitude.
Asunto(s)
Técnicas para Inmunoenzimas , Fosfatasa Alcalina , Animales , Tampones (Química) , Cromatografía Liquida , Electroquímica , Inmunoglobulina G/análisis , ConejosRESUMEN
The performance of a benchtop GC-ion trap MS-MS instrument, the Varian Saturn 4D, was evaluated for the analysis of a model drug, tebufelone, in plasma. The sample preparation scheme was designed to provide a highly complex extract with matrix-derived interferences eluting near and at the retention time of tebufelone and its stable-isotope-labeled analog. The performance of the ion trap in the selected-reaction-monitoring mode was evaluated and also compared with results obtained on a benchtop GC-MS linear quadrupole instrument operated in the selected-ion-monitoring mode. The ion trap, operated in the selected-reaction-monitoring mode, was found to provide a higher degree of selectivity for the analysis of tebufelone. The increased selectivity obtained on the ion trap operated in the selected-reaction-monitoring mode resulted in superior accuracy and precision, as well as a lower limit of quantitation relative to that obtained by the GC-MS analysis. A linear standard curve was obtained over three orders of magnitude and the limit of quantitation for tebufelone in plasma was 100 pg/ml using the GC-ion trap MS-MS instrument.
Asunto(s)
Alquinos/sangre , Antiinflamatorios no Esteroideos/sangre , Cromatografía de Gases y Espectrometría de Masas/instrumentación , Fenoles/sangre , Animales , Isótopos de Carbono , Estudios de Evaluación como Asunto , Indicadores y Reactivos , Isótopos de Oxígeno , Conejos , Estándares de ReferenciaRESUMEN
The binding of electroactively labeled estriol with estrogen-specific antibody and its subsequent displacement by unlabeled estriol have been monitored by differential pulse polarography. Estriol was found to be electro-inactive in the potential range -200 mV to -1000 mV vs a silver/silver chloride electrode. Estriol labeled in the 2 and 4 position with nitro groups was electroactive, giving two reduction waves at -422 mV and -481 mV vs a silver/silver chloride electrode. The peak current was linear with concentration over the range 60 micrograms/L to 3.7 mg/L. The addition of aliquots of estrogen-specific antibody reduced the peak current proportionately, indicating the binding of ligand to specific antibody. Subsequent addition of unlabeled estriol produced incremental increases in peak reduction current, indicating competitive and reversible binding of the two ligands for the antibody. Separation of bound from free labeled hapten was not necessary because reduction of the antibody-bound labeled estriol is attenuated at the electrode.
Asunto(s)
Anticuerpos Monoclonales/inmunología , Haptenos/inmunología , Sitios de Unión , Reacciones Cruzadas , Electroquímica , Electrodos , Estriol/análogos & derivados , Estriol/inmunología , Inmunoglobulina G/inmunología , Polarografía/métodosRESUMEN
Supercritical fluid extraction was coupled with solid-phase extraction using octadecylsilane cartridges for the selective isolation of ultratrace levels of a drug metabolite, mebeverine alcohol, from plasma. Plasma was directly applied to the extraction cartridge, the cartridge was washed to remove protein and then extracted under supercritical conditions using CO2/5% methanol. The effluent from the extraction cell was bubbled through a small volume of 2-propanol to trap the extracted mebeverine alcohol. The effects of extraction pressure and temperature on analyte recovery were examined. The absolute recovery, selectivity, precision, and accuracy of the combined supercritical fluid extraction/solid-phase extraction approach were compared to those of conventional solid-phase extraction using gas chromatography/mass spectrometry in the selected-ion monitoring mode. Mebeverine alcohol was used as a model compound, and dog plasma was employed as the biological matrix for these studies.