Determination of methamphetamine and amphetamine in abusers' plasma and hair samples with HPLC-FL

This paper describes highly sensitive HPLC methods for the determination of amphetamine (AP) and methamphetamine (MP) in abusers' plasma and hair samples. AP and MP were derivatized with the fluorescent reagent, DIB-Cl, to yield a highly fluorescent DIB-derivatives of AP and MP, which were then analyzed by HPLC with fluorescence detection at excitation and emission wavelengths of 325 and 430 nm, respectively. The separation was achieved on an ODS column with isocratic mobile phases composed of acetoniltrile and citrate buffer (55:45, v/v) for plasma samples and of acetonitrile-methanol-citrate buffer (45:20:37.5, v/v/v) for hair samples. The limits of detection were less than 0.87 ng/mL and 0.12 ng/mg in plasma and hair samples, respectively, for both AP and MP. The methods were then applied to the determination of MP and its metabolite AP in plasma obtained from two cases of illegally ingested MP and in one of the cases' hair received later. Case I was treated with dialysis; samples before and after dialysis were analyzed by the described method. After dialysis for 5 h, the total plasma levels of AP and MP decreased from 720 to 190 ng/mL. For case II, MP and AP levels were monitored for 3 days after digestion. Total plasma levels decreased from 57 ng/mL in the day of digestion to 11 ng/mL after 3 days. In hair samples, AP and MP could also be detected in very low concentrations.     

Pharmacokinetic studies of a novel trioxane antimalarial (99/411) in rats and monkeys using LC–MS/MS

The pharmacokinetic profile of 99/411, a novel anti‐malarial drug, was established in rats (12 mg/kg of body weight) and monkeys (20 mg/kg of body weight). Following oral administration, the presence of 99/411 was rapidly determined in rat plasma, tissues, urine, feces and monkey plasma using a validated LC–MS/MS method. The tissue distribution studies in rats indicated that the drug was partially distributed in all major tissues and plasma, and peak concentration levels were achieved within 0.5–4 h. Area under the curve in different rat tissues and plasma was found in order of blood > lung > intestine > heart > muscle > brain > kidney > spleen > liver. The total recoveries (within 86 h) of 99/411 were <0.0017% and <0.08% in urine and feces, respectively. The peak plasma concentration was 3499 ng/mL in rats after ~2 h of oral administration and 697–767 ng/mL in monkeys after ~6 h of oral administration. No plasma accumulation was observed in both male and female monkeys, even after multiple dosing. The preclinical pharmacokinetic profile and tissue distribution data are expected to assist in future clinical explorations of 99/411 as a promising anti‐malarial agent.     

Determination of telbivudine in the plasma of chronic hepatitis B patients in long‐term treatment by high‐performance liquid chromatographic–tandem mass spectrometry

Creatine kinase elevation is commonly reported in telbivudine‐treated patients. However, little is known about the relationship between this adverse drug reaction and plasma concentration. In this study, a sensitive, rapid and safe quantitative bioanalytical method has been established by using LC‐MS/MS for the determination of telbivudine in a clinical study of chronic hepatitis B patients. The assay was linear in a dynamic 10–10,000 ng/mL range (r²> 0.999) and total analysis time was 6 min in this method. The validated method was applied to quantitatively determine plasma concentration in chronic hepatitis B patients during long‐term telbivudine treatment. The results revealed that telbivudine concentration in the creatine kinase‐elevated group (707.92–2788.78 ng/mL) was significantly higher than those with normal creatine kinase (412.63–1108.32 ng/mL). This method was adapted for therapeutic drug monitoring.     

High-performance liquid chromatographic determination of lorazepam in monkey plasma

A simple, sensitive and selective method for the determination of lorazepam in monkey plasma has been developed using high-performance liquid chromatography in a reversed-phase mode. The limit of detection for lorazepam in plasma is about 2 ng/ml. The method has been applied to plasma samples obtained from cynomolgus monkeys after oral doses of 0.15 mg/kg and intravenous doses of 0.05 mg/kg of lorazepam. In this species, mean peak plasma concentrations of 12 ng/ml occurred at 2 h after oral dosing and declined with a half-life of 2,5 h; the mean terminal half-life after intravenous dosing was 1.4 h.     

Determination of 6-mercaptopurine and azathioprine in plasma by high-performance liquid chromatography.

Using 1-ml plasma samples, levels of 6-mercaptopurine (6MP) as low as 5 ng/ml and azathioprine (AZA) as low as 40 ng/ml can be detected using a high-performance liquid chromatography reversed-phase column procedure following extraction. Both compounds were stable in frozen plasma for seven weeks. AZA stability in blood was temperature dependent; the half-lives of AZA breakdown to 6MP at 37 degrees were 28 and 46 min in blood drawn from two rhesus monkeys. Plasma levels of 6MP were measured in a rhesus monkey following 6MP (1.47 mg/kg) and AZA (3 mg/kg) intravenous administration. 6MP levels were also measured in three renal transplant patients on daily 50- and 100-mg AZA doses. Peak levels (45-75 ng/ml) were reached within an hour and 6MP levels were detected for up to 7 h.     

Solid phase extraction of oxcarbazepine and its metabolites from plasma for analysis by high performance liquid chromatography.

A rapid, sensitive and simple-to-operate high performance liquid chromatographic method for the simulataneous determination of oxcarbazepine, 10-hydroxycarbazepine and 10,11-dihydro-10,11-trans-dihydroxy-carbamazepine in plasma is described. The drug and its metabolites were extracted from plasma using commercially available reversed phase octadecylsilane bonded-silica columns (Bond Elut C18, 1 mL capacity). Chromatographic separation of oxcarbazepine and its metabolites was achieved using a mobile phase consisting of acetonitrile/methanol/water (13:25:62 by volume) at a flow rate of 1.2 mL/min in conjunction with a Waters Associates Nova-Pak C18 column. The analytical column, in Radial-Pak cartridge form, was used in combination with a LiChrospher 5 microns C18 guard column. By measuring the UV absorbance at 214 nm, plasma levels in the region of 50-100 ng/mL for the drug and its metabolites can be detected with only 100 microL of plasma. The method has been applied to pharmacokinetic studies of oxcarbazepine and its metabolites in children with epilepsy; preliminary pharmacokinetic findings in two patients at steady-state are presented.     

Quantitative determination of naltrexone and beta-naltrexol in human plasma using electron detection.

A gas-liquid chromatographic method is described for the determination of naltrexone and beta-naltrexol in human plasma following derivatization with pentafluoropropionic anhydride using electron capture detection. The lower sensitivity of the method for absolute standards is 5-10 pg. Following an acute 100-mg dose to a subject, peak levels of naltrexone of 15 ng/ml at 2 h and of beta-naltrexol 84 ng/ml at 4 h were observed. The levels of both compounds decreased by 24 h after the dose: naltrexone to 2.9 ng/ml and beta-naltrexol to 25 ng/ml. Following chronic administration for two weeks of 100 mg per day the peak levels of naltrexone and betanaltrexol increased to 26.9 and 131 ng/ml at 2 h, respectively, but by 24 h both compounds were at levels similar to those following a single dose. Thus no accumulation of either drug ro metabolite in the plasma was seen following chronic naltrexone administration.     

Determination of artemether in plasma and whole blood using HPLC with flow-through polarographic detection

An HPLC method with polarographic detection for the trace determination of artemether in plasma and whole blood was developed and applied to pharmacokinetic and clinical pharmacological studies. The method showed high sensitivity and selectivity because of the easy reduction of the peroxide linkage of artemether at the mercury drop electrode. The detection limit was 10 ng and the detector response was linear over the range of 10 ng to 1 microgram artemether injected onto the column. The largest relative standard deviation of 10 replicate measurements of standard solutions (concentrations of 10 ng/mL-1 microgram/mL) was 8%. The recovery from whole blood and plasma of added drug (concentrations of 15-480 ng/mL) was 71-100%.     

Head-column field-amplified sample stacking in capillary electrophoresis for the determination of cimetidine, famotidine, nizatidine, and ranitidine-HCl in plasma.

In this study, low concentrations of histamine2-receptor (H2-)antagonists were effected across a water plug, with separation taking place in a binary buffer comprising ethylene glycol and NaH2PO4 (pH 5.0), and detection at 214 nm. Liquid-liquid extraction with ethyl acetate- isopropanol is shown to provide extracts that are sufficiently clean. The calibration curves were linear over a concentration range of 0.1-2.00 microg/mL cimetidine, 0.2-5.0 microg/mL ranitidine-HCl, 0.3-5.0 microg/mL nizatidine, and 0.1-3.0 microg/mL famotidine. Mean recoveries were > 82%, while the intra- and interday relative standard deviations (RSDs) and relative errors (REs) were all < 13%. The method is sensitive with a detection limit of 3 ng/mL cimetidine, 30 ng/mL ranitidine HCl, 50 ng/mL nizatidine and 10 ng/mL famotidine (S/N = 3, electric-driven injection 90 s). This newly developed capillary electrophoresis (CE) method was applied for the determination of analytes extracted from plasma taken from a volunteer dosing a cimetidine, ranitidine, and nizatidine tablet simultaneously. These three H2-antagonists can be detected in real samples by this method, excluding the low dosing of famotidine tablet.     

Development and validation of an LC-MS/MS analysis for simultaneous determination of delphinidin-3-glucoside, cyanidin-3-glucoside and cyanidin-3-(6-malonylglucoside) in human plasma and urine after blood orange juice administration

Blood orange juice has a high content in anthocyanins, especially represented by delphinidin-3-glucoside (D3G), cyanidin-3-glucoside (C3G) and cyanidin-3-(6-malonylglucoside) (CMG). An LC-MS/MS method for the simultaneous determination of D3G and C3G in human plasma and urine was developed and validated. After sample preparation by SPE, chromatographic separation was performed with an RP-C(18) column, using a water/methanol linear gradient. The quantitation of target compounds was determined by multiple reaction monitoring (MRM) mode, using ESI. The method showed good selectivity, sensitivity (LOD = 0.05 and 0.10 ng/mL for C3G in plasma and urine, respectively; LOD = 0.10 ng/mL for D3G in plasma and urine), linearity (0.20-200 ng/mL; r >or= 0.998), intra- and interday precision and accuracy (相似文献   

Rapid high-performance liquid chromatographic measurement of buspirone in human plasma after overdose

For toxicological purposes, a rapid reversed-phase high-performance liquid chromatographic method was developed for the determination of the anxiolytic drug, buspirone, in human plasma. A liquid-liquid procedure was used to extract this compound from plasma in the presence of an internal standard, quinupramine. The analysis was performed on a Spherisorb S5 C8 analytical column with UV detection at 240 nm. No endogenous compounds were found to interfere. A linear response was observed over the concentration range 5-100 ng/mL. A good accuracy (bias < or =7.9%) was achieved for all quality controls, with intra-day and inter-day variation coefficients equal or less than 7.6%. The limit of quantification was 5 ng/mL. Stability of buspirone in plasma stored at different temperatures was checked. This rapid method (run time <12 min) was used to manage an acute poisoning involving buspirone.     

Therapeutic Drug Monitoring of Suramin and Protein Binding

Abstract

A liquid chromatographic method of suramin has been used for the determination of the polysulfonated naphtylurea in both plasma and plasma filtrate from prostate cancer patients treated with the drug. The chromatographic system is based on the use of tetrabutylammonium bromide as an ion-pairing agent, while UV detection at 237 nm and 313 nm is applied. The sample pretreatment is a simple deproteination step by an organic solvent. The same counter-ion as used in the phase system is added in order to increase the recovery of the almost complete protein-bound suramin. The minimum detectable concentration in plasma is ca. 20 ng/mL at 237 nm, in plasma filtrate 10 ng/mL at 237 nm.

The method was routinely applied in plasma level guided treatment of prostate cancer patients with suramin, as well as in protein binding studies. The data of the study demonstrated the necessity of therapeutic drug monitoring in suramin treatment: development of severe, irreversible toxicity could be prevented owing to timed withdrawal of suramin administration when total drug levels were beyond 300 μg/mL. Data of protein binding studies explained in part the development of severe toxicity associated with plasma levels beyond 300–350 μg/mL: at that point free fraction of suramin sharply increases from 500 ng/mL at a total plasma level of 500 μg/mL to 10 μg/mL at a total plasma concentration of 1000 μg/mL, which corresponds with a twenty-fold dose increase (2000%).     

Liquid chromatographic determination of nicarbazin in feeds

A new liquid chromatographic method has been developed for determination of nicarbazin in feeds. Approximately 40 g feed is extracted with 200 mL acetonitrile-water (80 + 20, v/v). An aliquot of the extract is filtered and assayed using a reversed-phase isocratic method that measures the 4,4'-dinitrocarbanilide moiety of nicarbazin at a wavelength of 340 nm. For medicated feeds, the method uses a standard linear range of 5 to 100 microg/mL. For lower levels, a linear range of 50 to 150 ng/mL can be used. The method has a limit of detection of 250 ng/g and a limit of quantitation of 500 ng/g in a 40 g feed sample. Recovery was 99.1%, with a range of 95.2 to 101.8%. In the typical U.S. dosing range of 27 to 113.5 g/ton, the precision of the method based on one analyst, one day, and 2 weighings ranged from 2.8% (113.5 g/ton) to 4.7% (27 g/ton).     

Liquid chromatographic/mass spectrometric assay of rabeprazole in dog plasma for a pharmacokinetic study

In order to evaluate the pharmacokinetic (PK) profile of rabeprazole (RA) sterile powder for injection, a rapid, sensitive and specific assay for quantitative determination of RA in dog plasma was developed and validated. After a liquid-liquid extraction procedure, samples were analyzed by liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS) using omepazole as the internal standard (IS). The analyte and IS was chromatographed on a ZORBAX Extend-C(18) analytical column (50 x 2 mm i.d, 5 microm, Agilent Technologies, USA). The assay was linear in the range 1-2000 ng/mL. The lower limit of quantification of RA was 1 ng/mL. The recovery of RA was greater than 70%. The within- and between-batch accuracy was 102.7-107.4% and 103.5-105.7%, respectively. The plasma samples for the PK study were collected at defined time points during and after an intravenous injection (1 mg/kg) to beagle dogs and analyzed by LC-ESI-MS method. The PK parameters, such as half-life, volume of distribution, total clearance and elimination rate constant, were determined. The PK profile of RA gave insights into the application in the clinics.     

Use of 18O3-clodronate as an internal standard for the quantitative analysis of clodronate in human plasma by gas chromatography/electron ionisation mass spectrometry

A sensitive and specific method for the quantitative determination of clodronate in human plasma is presented. The drug was extracted from plasma by anion-exchange chromatography and derivatised to the tetra-tert-butyldimethylsilyl derivative. 18O3-Clodronate was prepared from unlabeled clodronate and used as an internal standard. Gas chromatography/mass spectrometry (GC/MS) under electron ionisation conditions was used for quantitative measurement of the drug, using m/z 643.16 and 651.17 for target and internal standard, respectively. Calibration graphs were linear within the range of 10-1280 ng/mL plasma. Intra-day precision was 1.8% (10 ng/mL), 0.5% (40 ng/mL), 1.0% (120 ng/mL), 0.5% (200 ng/mL), 0.5% (400 ng/mL) and 2.7% (800 ng/mL) and inter-day variability was found to be 0.7% (10 ng/mL), 1.6% (40 ng/mL), 1.3% (120 ng/mL), 2.3% (200 ng/mL), 2.5% (400 ng/mL) and 1.2% (800 ng/mL). Intra-day accuracy showed deviations of 0.8% (10 ng/mL), 0.8% (40 ng/mL), 0.9% (120 ng/mL), 0.9% (200 ng/mL), 1.9% (400 ng/mL) and 0.3% (800 ng/mL) and intra-day accuracy was of -1.4% (10 ng/mL), 0.0% (40 ng/mL), -0.7% (120 ng/mL), -0.4% (200 ng/mL), -1.2% (400 ng/mL) and -3.3% (800 ng/mL). The stable isotope labeled standard was found to be stable under analysis conditions.     

Rapid and sensitive LC-MS/MS method for the determination of metoprolol in Beagle dog plasma with a simple protein precipitation treatment and its pharmacokinetic applications

A rapid LC-MS/MS method with good accuracy and sensitivity was developed and validated for the pharmacokinetics study of metoprolol (MP) in beagle dogs. The plasma samples were simply precipitated by methanol and then analyzed by LC-MS/MS. An Ultimate XB-C?? column (150 × 2.1 mm ID, 5 μm) was used for separation, with methanol-water containing 0.2% formic acid (65:35, v/v) as the mobile phase at a flow rate of 0.2 mL/min. Monitoring ions of MP and internal standard (hydroxypioglitazone) were m/z 268.1/115.6 and m/z 373.1/150.2, respectively. The linear range was 3.03-416.35 ng/mL with an average correlation coefficient of 0.9996, and the limit of quantification was 3.03 ng/mL. The intra- and inter-day precision was less than 15%. At low, middle and high concentrations, the recovery, the matrix effect and the accuracy was in the range of 76.06%-95.25%, 93.67%-104.19% and 95.20%-99.96% respectively. The method was applied for the pharmacokinetics study of MP tartrate tablets (50 mg). The AUC(0-t), T(max) and C(max) were respectively 919.88 ± 195.67 μg/L·h, 0.96 ± 0.33 h, 349.12 ± 78.04 ng/mL.     

Quantitative determination of a new anticonvulsant (CGS 18416A) in human plasma using capillary gas chromatography/mass spectrometry.

An analytical method has been developed for the determination of a new antiepileptic drug, CGS 18416A, in human plasma. CGS 18416A is a new anticonvulsant representative of a novel class of water-soluble agents being developed for the treatment of epilepsy. Preclinical trials indicate sustained efficacy at relatively low oral doses, indicating a need for a sensitive assay. The method is based on capillary gas chromatography/mass spectrometry and utilizes stable isotope-labelled CGS 18416A as the internal standard. Samples (1 mL) are acidified, then washed with pentane/ethyl acetate, followed by liquid/liquid extraction at pH 11 with pentane/ethyl acetate. Extracts are then concentrated and analysed directly by gas chromatography/mass spectrometry. Separation is accomplished on a thick film methylsilicone capillary column. Mass spectrometry was carried out under positive ion ammonia Cl conditions with selected ion monitoring of the protonated molecular ions (m/z = 248 and 252) for CGS 18416A and the 13CD3-CGS 18416A, respectively. Specificity was demonstrated by the lack of interfering peaks at the retention time of CGS 18416A and the internal standard. Recovery and reproducibility assessments indicate good accuracy and precision over the validated concentration range of 0.2-51 ng/mL. The limit of quantification is 0.2 ng/mL and the method has sufficient sensitivity to support clinical trials. This is illustrated with an example of quantification in a normal volunteer following oral dosing.     

High-performance liquid chromatographic determination of the calcium channel blocker nimodipine in monkey plasma.

A new high-performance liquid chromatographic (HPLC) assay was developed for the determination of nimodipine in monkey plasma. An ethyl acetate extraction procedure was employed with a reversed-phase HPLC separation for the analysis. Absolute recovery of nimodipine from plasma was over 95% with a lower limit of quantitation of 10 ng/ml. This method was applied to a preliminary pharmacokinetic study in which 0.25 mg/kg nimodipine was administered intravenously to three monkeys. Protein binding and stability of nimodipine in monkey plasma were also examined. The pharmacokinetic parameters of nimodipine in monkeys were similar to those obtained in humans and indicate that monkeys are an appropriate animal model for further pharmacokinetic investigations.     

Establishment of a HPLC method for preclinical pharmacokinetic study of the novel anti-Parkinson's disease candidate drug FLZ in rats

An HPLC method was established and validated for the determination of compound FLZ, a synthetic novel anti-Parkinson's disease candidate drug, in rat plasma. FLZ and the internal standard bicyclol were extracted from plasma by solid-phase extraction method and analyzed on a Restek C18 column (4.6 x 250 mm, 5 microm) with a mobile phase consisting of methanol and water (60:40, v/v) at a flow rate of 1.0 mL/min. The detection wavelength was set at 320 nm. The calibration curve was linear within the concentration range from 25 to 500 ng/mL (r2 > 0.999), the limit of quantitation was 25 ng/mL and the average recovery was 92.0% with the RSD less than 5.9%. The relative standard deviation for intra- and inter-day precision was less than 3.8 and 6.9%, respectively. The established HPLC method was validated to be a simple, rapid and reliable procedure and applied to study the preclinical pharmacokinetics of FLZ in rat plasma, and it was the first time that the pharmacokinetics of FLZ had been investigated.     

Quantitative analysis of the farnesyl transferase inhibitor lonafarnib (Sarasartrade mark, SCH66336) in human plasma using high-performance liquid chromatography coupled with tandem mass spectrometry

Lonafarnib is a novel anticancer drug that inhibits farnesyl transferase. To assess its pharmacokinetic properties, we developed a sensitive and quantitative assay using liquid chromatography coupled with tandem mass spectrometry for the determination of lonafarnib levels in human plasma. Sample pretreatment consisted of the addition of an isotopically labeled internal standard and protein precipitation with acetonitrile using 100 microL plasma. Chromatographic separation was performed on an Inertsil ODS-3 analytical column (50 x 2.1 mm i.d., particle size 5 microm) with acetonitrile/water/formic acid (50:50:0.05, v/v/v) as the mobile phase, at a flow rate of 0.2 mL/min. The analytical run time was 8 min. An API365 triple quadrupole mass spectrometer was used for specific and sensitive detection. It was operated in the positive ion mode and multiple reaction monitoring was used for drug quantification. The method was validated using a concentration range of 2.5 to 2500 ng/mL lonafarnib. Validation of the assay was performed according to the most recent FDA guidelines for bioanalytical method validation and all results were within the requirements. The described method was successfully applied to support a clinical phase I trial with lonafarnib.