Time-dependent variation in the biodistribution of C₆₀ in rats determined by liquid chromatography-tandem mass spectrometry

We examined the biodistribution of C₆₀ in rats after tail vein administration using LC-MS/MS. C₆₀ was detected in various tissues, such as brain, kidneys, liver, lungs, and spleen of rats. On the other hand, no C₆₀ was found in blood. The highest C₆₀ concentration was observed in the lungs, followed by spleen, liver, kidneys, and brain. These results suggested that C₆₀ injected in the tail vein could be filtered by lung capillary vessels and accumulate in the lungs prior to being distributed to other tissues. Moreover, C₆₀ not being detected in the blood indicates that clearance of C₆₀ from the blood by filtration might effectively occur in the lungs. The time-dependent variation in the biodistribution of C₆₀ was evaluated. A time-dependent decrease in C₆₀ concentrations was observed in all tissues, except spleen. Moreover, a decreasing trend of C₆₀ levels differed among tissues, which could be due to differences in accumulation. These results suggest that unmodified C₆₀ and/or C₆₀ metabolites by metabolic enzymes could be excreted into feces and/or urine. In further studies, the metabolic and excretion pathways of C₆₀ should be evaluated to understand the toxicokinetics of C₆₀.     

Bioanalysis of an oligonucleotide and its metabolites by liquid chromatography–tandem mass spectrometry

An ion-pair reversed phase liquid chromatography–tandem mass spectrometry (LC–MS/MS) method has been developed for the quantification of a phosphorothioate oligonucleotide (PS-OGN) PF-ODN and its metabolites 5′N-1/3′N-1, 5′N-2 and 5′N-3 in rat plasma. Plasma samples were prepared with an initial phenol/dichloromethane liquid–liquid extraction followed by a solid phase extraction. Chromatographic separation was performed with a gradient system on a Phenomenex Gemini C₁₈ column using hexafluoro-2-propanol/triethylamine buffer and methanol as the mobile phase at a flow rate of 0.5 mL/min. Except for 5′N-1 and 3′N-1, which were coeluted and could not be differentiated by mass spectrometer, the other analytes were separated chromatographically and mass spectrometrically. The detection was carried out in multiple reaction monitoring (MRM) mode using a negative electrospray ionization (ESI) interface. The lower limit of quantification (LLOQ) achieved was 4.0 ng/mL for PF-ODN and its four metabolites with acceptable precision and accuracy. Inter-day and intra-day RSD for three quality control (QC) levels across validation runs were less than 12.0% and the accuracy ranged from −9.6% to 6.0% for the analytes. This validated LC–MS/MS method was applied to a preliminary pharmacokinetic study of PF-ODN in rats.     

Simultaneous determination of methotrexate and its polyglutamate metabolites in Caco-2 cells by liquid chromatography–tandem mass spectrometry

In normal and malignant human cells, the folate antagonist methotrexate (MTX) is converted to a series of polyglutamates (MTXGlu_n, n = 2–5) which play a role in its therapeutic efficacy. Here we report an assay to determine MTX and MTXGlu_n in Caco-2 cells exposed to MTX. After a simple protein precipitation step, cell homogenates (2 × 10⁶ cells) were analyzed by liquid chromatography–tandem mass spectrometry (LC–MS/MS) using aminopterin as internal standard. Separation was by reversed phase HPLC on a C8 column using gradient elution with 0.1% formic acid and acetonitrile. Detection was by electrospray ionization in the positive ion mode followed by multiple reaction monitoring of the transitions of the [M+H]⁺ ions of MTX and MTXGlu_n to their common product ion at m/z 308.2 and of aminopterin at m/z 441.3 → 294.2. Calibration curves for all analytes were linear in the range 2–250 nM (r² > 0.996). Intra- and inter-day precisions (as coefficient of variation) were 3.4–15.1% and 4.3–18.4%, respectively with corresponding accuracies (as relative error) of −3.6 to +6.6% and −5.5 to +7.5%, respectively. Recoveries were in the range 60 ± 4 to 108 ± 13%. It was found that MTX undergoes only limited polyglutamation in Caco-2 cells exposed to MTX over 24 h.     

Determination of vitamin E and its metabolites in equine urine using liquid chromatography–mass spectrometry

Equine neuroaxonal dystrophy/degenerative myeloencephalopathy (eNAD/EDM) is a hereditary, deteriorating central nervous disease in horses. Currently, the only way to confirm eNAD/EDM is through a postmortem histological evaluation of the central nervous system. Vitamin E, specifically the isoform alpha-tocopherol (α-TP), is known to protect eNAD/EDM susceptible horses from developing the clinical phenotype. While vitamin E is an essential nutrient in the diet of horses, there are no diagnostic tests able to quantitate vitamin E and its metabolites in urine. An ultra-performance liquid chromatography-atmospheric-pressure chemical ionization mass spectrometry (UPLC-APCI-MS/MS) method was developed and validated following acidic hydrolysis and solid phase extraction to quantitate vitamin E and its metabolites in equine urine. A blank control horse urine matrix was used and spiked with different concentrations of analytes to form a standard curve using either alpha-tocopherol-d6 or chlorpropamide as the internal standard. Inter-day and intra-day statistics were performed to evaluate the method for accuracy (90% to 116%) and precision (0.75% to 14%). Matrix effects, percent recovery, and stability were also assessed. The method successfully analyzed alpha-carboxyethyl hydroxychroman (α-CEHC), alpha-carboxymethylbutyl hydroxychromans (α-CMBHC), gamma-carboxyethyl hydroxychroman γ-CEHC, and α-TP concentrations in urine to determine a baseline levels of analytes in healthy horses, and can be used to determine concentrations of vitamin E metabolites in equine urine allowing for its evaluation as a diagnostic approach in the treatment of eNAD/EDM.     

Determination of repaglinide in human plasma by high-performance liquid chromatography–tandem mass spectrometry

A rapid and sensitive method based on high-performance liquid chromatography–tandem mass spectrometry (LC–MS/MS) has been developed for the determination of repaglinide in human plasma. The analyte and internal standard (I.S.), diazepam, were extracted from plasma (25 μL) by liquid–liquid extraction with diethyl ether–dichloromethane (60:40, v/v) and separated on a XDB-C₁₈ column using acetonitrile–ammonium acetate buffer (pH 6.8, 0.01 mol/L) as mobile phase. The retention times of repaglinide and I.S. were 1.95 and 2.35 min, respectively. Detection was carried out using API 4000 mass spectrometer with an ESI interface operating in the multiple reaction monitoring (MRM) mode. The assay was linear over the concentration range 0.050–50 ng/mL with a limit of detection (LOD) of 0.010 ng/mL. Intra- and inter-day precisions (as relative standard deviation, R.S.D.) were ≤5.07% and ≤11.2%, respectively, and accuracy (as relative error, R.E.) was from ?0.593% to ?1.26%. The assay was successfully applied to a pharmacokinetic study involving a single oral administration of a tablet containing 2 mg repaglinide to each of 10 healthy volunteers.     

Quantification of sunitinib in mouse plasma,brain tumor and normal brain using liquid chromatography–electrospray ionization-tandem mass spectrometry and pharmacokinetic application

The primary challenge associated with the development of an assay method for the determination of drug concentrations in relatively small amount of mouse plasma and tissue samples is to improve extraction efficiency and detection sensitivity. In this work, a liquid chromatography–tandem mass spectrometry (LC–MS/MS)-based method combined with protein precipitation, liquid–liquid extraction and solid-phase extraction techniques was developed for the determination of sunitinib in mouse plasma, brain tumor and normal brain tissue, respectively. The instrument was operated under the multiple reaction monitoring (MRM) mode using electrospray ionization (ESI) in the positive ion mode. A good linear relationship with coefficients of determination ≥0.99 was achieved over the concentration ranges of 1.37–1000 ng/mL for plasma and 4.12–1000 ng/g for the normal brain and brain tumor. The limits of quantification (LOQs) for sunitinib in mouse plasma, brain tumor and normal brain tissue are 1.37 ng/mL, 4.12 ng/g and 4.12 ng/g, respectively. The reproducibility of the LC–MS/MS method is reliable, with the intra- and inter-day precision being less than 15% and accuracy within ±15%. The established method was successfully applied to the characterization of sunitinib disposition in the brain and brain tumor as well as its systemic pharmacokinetics in a murine orthotopic glioma model.     

Simultaneous analysis of Δ(9)-tetrahydrocannabinol and 11-nor-9-carboxy-tetrahydrocannabinol in hair without different sample preparation and derivatization by gas chromatography-tandem mass spectrometry

The present study describes a gas chromatography/tandem mass spectrometry-negative ion chemical ionization assay (GC/MS/MS-NCI) for simultaneous analysis of Δ(9)-tetrahydrocannabinol (THC) and 11-nor-9-carboxy-tetrahydrocannabinol (THCCOOH) in hair. Each hair sample, of approximately 20mg, was weighed and the sample was dissolved in 1ml of 1M sodium hydroxide (30min at 85°C) in the presence of THC-d(3) and THCCOOH-d(3). For the analysis of THC, hair samples were extracted with n-hexane:ethyl acetate (9:1) two times; acetic acid and sodium acetate buffer were added for the analysis of THCCOOH, and hair samples were re-extracted with n-hexane:ethyl acetate (9:1) two times. The extracts were then derivatized with pentafluoropropionic anhydride (PFPA) and pentafluoropropanol (PFPOH). This method allowed the analysis of THC and THCCOOH using the GC/MS/MS-NCI assay. This method was also fully validated and applied to hair specimens (n=54) collected from known cannabis users whose urine test results were positive. The concentrations of THC and THCCOOH in hair ranged from 7.52 to 60.41ng/mg and from 0.10 to 11.68pg/mg, respectively. In this paper, we simultaneously measured THC and THCCOOH in human hair using GC/MS/MS-NCI without requiring different sample preparation and derivatization procedures. The analytical sensitivity for THCCOOH in hair was good, while that for THC in hair needs to be improved in further study.     

Determination of sulfamonomethoxine in tilapia (Oreochromis niloticus × Oreochromis mossambicus) by liquid chromatography-tandem mass spectrometry and its application pharmacokinetics study

A precise and reliable analytical method to measure trace levels of sulfamonomethoxine (SMM) and N⁴-acetyl metabolite in tilapia samples using liquid chromatography-tandem mass spectrometry was developed. Optimized chromatographic separation was performed on C18 reversed-phase columns using gradient elution with methanol and 5 mmol/L of an ammonium acetate aqueous solution (adjusted to pH 3.5 using formic acid). This study investigated the pharmacokinetic properties and tissue distribution of SMM and its major metabolite N⁴-acetyl sulfamonomethoxine (AC-SMM) in tilapia after a single dose of 100 mg kg^?1 body weight of orally administered SMM. Blood and tissues were collected between 0.5 and 192 h with 14 total sampling time points. SMM was rapidly absorbed, and extensively distributed in the bile and liver through systemic circulation. Enterohepatic circulation of SMM was observed in the tilapia body. Acetylation percentages were 45% (blood), 90% (liver), 62% (kidney), 98% (bile), and 52% (muscle). High concentrations of AC-SMM accumulated in the tilapia bile. At 192 h, AC-SMM concentration in the bile remained at 4710 μg kg^?1. The k_e value of AC-SMM (0.015 h^?1) in the blood was lower than that of SMM (0.032 h^?1). This study demonstrated effective residue monitoring and determined the pharmacokinetic properties of SMM and AC-SMM in tilapia.     

Analysis of enantiomers of sibutramine and its metabolites in rat plasma by liquid chromatography–mass spectrometry using a chiral stationary-phase column

Sibutramine, a monoamine reuptake inhibitor, is used as a racemate, for the treatment of obesity. It is converted in vivo mainly to two desmethyl active metabolites, mono-desmethylsibutramine (MDS) and di-desmethylsibutramine (DDS). In the present study, we introduced a rapid and simple chromatographic method for separating the R(+)- and S(−)-isomers of sibutramine, MDS, and DDS, respectively. The stereoisomers of the three compounds were extracted from rat plasma using diethyl ether and n-hexane under alkaline conditions. After evaporating the organic layer, the residue was reconstituted in the mobile phase (10 mM ammonium acetate buffer adjusted to pH 4.03 with acetic acid:acetonitrile, 94:6, v/v). The enantiomers in the extract were separated on a Chiral-AGP stationary-phase column and were quantified in a tandem mass spectrometry. The accuracy and precision of the assay were in accordance with FDA regulations for the validation of bioanalytical methods. This method was used to measure the concentrations of the enantiomers of sibutramine, MDS, and DDS in plasma after a single oral dose of 10 mg/kg racemic sibutramine in rats.     

Determination of tapentadol (Nucynta®) and N-desmethyltapentadol in authentic urine specimens by ultra-performance liquid chromatography-tandem mass spectrometry

Urine specimens from pain management patients dosed with Nucynta (Tapentadol) were confirmed for the presence of tapentadol and N-desmethyltapentadol using ultra-performance liquid chromatography-tandem mass spectrometry to minimize sample preparation and urine volume requirements. The linearity of the method for both tapentadol and N-desmethyltapentadol demonstrated correlation coefficients (R2) above 0.99 and linear ranges from 50 to 500,000 ng/mL for tapentadol and 100 to 500,000 ng/mL for N-desmethyltapentadol. The intraday precision of the assay for both analytes ranged from 2.2 to 6.9% over three concentrations; the interday precision for both analytes ranged from 1.2 to 8.4%. The limits of quantitation were 50 and 100 ng/mL for tapentadol and N-desmethyltapentadol, respectively, and the upper limit of linearity for both analytes was determined to be 500,000 ng/mL. Urine samples were collected within 24 h of dosing with tapentadol and shipped overnight to the laboratory. Samples were hydrolyzed with acid prior to analysis to measure total (unconjugated and conjugated) tapentadol and N-desmethyltapentadol. Further investigation into characterization of metabolites was performed by using a hybrid quadrupole-time-of-flight mass spectrometer in lieu of suitable analytical reference standards. The presence of significant N-desmethyltapentadol glucuronide was demonstrated for the first time.     

Simultaneous quantification of enalapril and enalaprilat in human plasma by high-performance liquid chromatography–tandem mass spectrometry and its application in a pharmacokinetic study

A rapid, selective and sensitive high-performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS) method was developed to simultaneously determine enalapril and enalaprilat in human plasma. With benazepril as internal standard, sample pretreatment involved in a one-step protein precipitation (PPT) with methanol of 0.2 ml plasma. Analysis was performed on an Ultimate™ XB-C₁₈ column (50 mm × 2.1 mm, i.d., 3 μm) with mobile phase consisting of methanol–water–formic acid (62:38:0.2, v/v/v). The detection was performed on a triple quadrupole tandem mass spectrometer by multiple reaction-monitoring (MRM) mode via electrospray ionization (ESI) source. Each plasma sample was chromatographed within 2.5 min. The linear calibration curves for enalapril and enalaprilat were both obtained in the concentration range of 0.638–255 ng/ml (r² ≥ 0.99) with the lower limit of quantification (LLOQ) of 0.638 ng/ml. The intra-day precision (R.S.D.) was below 7.2% and inter-day R.S.D. was less than 14%, while accuracy (relative error R.E.) was within ±8.7 and ±5.5%, determined from QC samples for enalapril and enalaprilat which corresponded to requirement of the guidance of FDA. The HPLC–MS/MS method herein described was fully validated and successfully applied to the pharmacokinetic study of enalapril maleate capsules in 20 healthy male volunteers after oral administration.     

Determination of drugs in exhumed liver and brain tissue after over 9 years of burial by liquid chromatography-tandem mass spectrometry—Part 1: Cardiovascular drugs

This paper presents analytical results of cardiovascular drugs that were determined in exhumed liver and brain tissue in a total of 115 cases. Exhumations were executed after 9.5–16.5 years of burial. Analysis was performed by solid phase extraction and liquid chromatography–electrospray mass spectrometry. Bisoprolol, verapamil, and xipamide could be proven in all cases in which these were supposedly administered within a defined time frame. Amiodarone was detectable in over 90%, amlodipine in over 80%, and cafedrine, clonidine, phenprocoumon, and torsemide in over 50% of relevant cases. By contrast, atropine, esmolol, furosemide, hydrochlorothiazide, and lisinopril were found in less than 50% of relevant cases, but nifedipine and nitrendipine were not detectable at all. The percentage of positive results for liver and brain tissue for the relevant administered drugs and corresponding postmortem periods are presented per analyte. Neither time since death nor stage of degradation was a reliable predictor of the success rate, as exemplarily shown for furosemide. The presented data may serve as a reference when deciding whether to exhume a corpse for forensic-toxicological examinations in comparable cases.     

Antidepressant-like effects of Δ⁹-tetrahydrocannabinol and rimonabant in the olfactory bulbectomised rat model of depression

The endocannabinoid signalling system is widely accepted to play a role in controlling the affective state. Plant cannabinoids are well known to have behavioural effects in animals and humans and the cannabinoid CB₁ receptor antagonist rimonabant has recently been shown to precipitate depression-like symptoms in clinical trial subjects. The aim of the present study was to investigate the behavioural and neurochemical effects of chronic administration of Δ⁹-tetrahydrocannabinol (THC) and rimonabant on intact and olfactory bulbectomised (OB) rats used as a model of depression.As expected, OB rats were hyperactive in the open field. Repeated THC (2 mg/kg, i.p. once every 48 h for 21 days) and rimonabant (5 mg/kg, i.p. once every 48 h for 21 days) reduced this hyperactivity, which is typical of clinically effective antidepressant drugs. In intact animals, chronic THC increased brain derived neurotrophic factor (BDNF) expression levels in the hippocampus and frontal cortex but rimonabant had no effect. Rimonabant increased the levels of phosphorylated extracellular signal regulated kinases (p-ERKs_1/2) in the hippocampus and prefrontal cortex and THC also increased expression in frontal cortex. OB did not affect BDNF or p-ERK_1/2 expression in the hippocampus or frontal cortex and in, contrast to the intact animals, neither THC nor rimonabant altered expression in the OB rats.These findings indicate antidepressant-like behavioural properties of both THC and rimonabant in OB rats although additional studies are required to clarify the relationship between the chronic effects of cannabinoids in other pre-clinical models and in human depression.     

Direct quantitative analysis of benzodiazepines,metabolites, and analogs in diluted human urine by rapid resolution liquid chromatography–tandem mass spectrometry

Rapid resolution liquid chromatography (RRLC) coupled with triple quadrupole mass spectrometry (QqQ-MS) was developed for direct quantitative analysis of benzodiazepines (BDZs) and their metabolites, as well as BDZ analogs (zopiclone, zolpidem, and zaleplon) in diluted human urine. A perfluorophenyl column packed with sub-2-μm particles and a C8 precolumn were applied to accomplish RRLC analysis with symmetric peak shapes. Urine samples were diluted 10-fold with Milli-Q water prior to autoinjection for direct quantification of 34 target analytes with negligible matrix effect. Gradient elution and dynamic multiple reaction monitoring were used for simultaneous quantitation of 34 BDZs, metabolites, and BDZ analogs. Good recovery was obtained in the range of 80.2–98.5% and the limit of detection ranged from 0.01 ng/mL to 0.5 ng/mL for all 34 target analytes in spiked urine. Moreover, good precision and accuracy were obtained for quantitative determination in diluted urine samples by the proposed RRLC/QqQ-MS method for intra-day/inter-day assays in the ranges of 0.1–8.8%/0.1–8.9% and 91.2–106.1%/89.6–104.6%, respectively. The applicability of this newly developed RRLC/QqQ-MS method was demonstrated by quantitative determination of BDZs, metabolites, and BDZ analogs in various clinical urine samples.     

Analysis of tetrahydrocannabinol and its metabolite, 11-nor-Δ⁹-tetrahydrocannabinol-9-carboxylic acid, in oral fluid using liquid chromatography with tandem mass spectrometry

This paper describes the determination of tetrahydrocannabinol (THC) and its metabolite, 11-nor-Δ?-tetrahydrocannabinol-9-carboxylic acid (THC-COOH) in oral fluid using solid-phase extraction and liquid chromatography with tandem mass spectral detection (LC-MS-MS) and its application to proficiency specimens. The method employs collection of oral fluid with the Quantisal? device, base hydrolysis, solid-phase extraction and LC-MS-MS in positive ion electrospray mode. Because the concentration of the metabolite in oral fluid is quite low, extremely sensitive analytical methods are necessary. The requisite sensitivity was achieved by a simple, rapid derivatization of the compound after extraction. The derivatization conditions did not affect parent THC. The method was fully validated using standard parameters including linearity, sensitivity, accuracy, intra-day and inter-day imprecision, drug recovery from the collection pad, limit of quantitation, limit of detection and matrix effects. The procedure was applied to oral fluid proficiency specimens previously analyzed to assess the stability of THC-COOH.     

A validated method for the simultaneous quantification of cannabidiol, Δ9-tetrahydrocannabinol,and their metabolites in human plasma and application to plasma samples from an oral cannabidiol open-label trial

Cannabidiol (CBD) and Δ⁹-tetrahydrocannabinol (THC) are the two best known and most extensively studied phytocannabinoids within Cannabis sativa. An increasing number of preclinical studies and clinical trials have been conducted with one or both compounds, often probing their therapeutic effects in conditions such as paediatric epilepsy, anxiety disorders or chronic pain. Accurate monitoring of THC and CBD and their metabolites is essential for tracking treatment adherence and pharmacokinetics. However, fully validated methods for the comprehensive analysis of major Phase I CBD metabolites are yet to be developed due to a historical lack of commercially available reference material. In the present study, we developed, optimised and validated a method for the simultaneous quantification of CBD, THC and their major Phase I metabolites 6-hydroxy-CBD (6-OH-CBD), 7-hydroxy-CBD (7-OH-CBD), 7-carboxy-CBD (7-COOH-CBD), 11-hydroxy-tetrahydrocannabinol (11-OH-THC) and 11-carboxy-tetrahydrocannabinol (11-COOH-THC) as per Food and Drug Administration (FDA) guidelines for bioanalytical method validation. The method is accurate, reproducible, sensitive and can be carried out in high-throughput 96-well formats, ideal for larger scale clinical trials. Deuterated internal standards for each analyte were crucial to account for variable matrix effects between plasma lots. The application of the method to plasma samples, taken from people who had been administered oral CBD as part of an open-label trial of CBD effects in anxiety disorders, demonstrated its immediate utility in ongoing and upcoming clinical trials. The method will prove useful for future studies involving CBD and/or THC and can likely accommodate the inclusion of additional metabolites as analytical reference materials become commercially available.     

Tetrodotoxin and paralytic shellfish poisons in gastropod species from Vietnam analyzed by high-performance liquid chromatography and liquid chromatography–tandem mass spectrometry

Among marine toxins, tetrodotoxin (TTX) and paralytic shellfish poisons (PSPs) are known as notorious neurotoxins that induce serious food poisoning incidents in the Southeast Asia region. The aim of this study was to investigate whether TTX and PSP toxins are important issues of seafood safety. Paralytic toxicity was observed in mice exposed to 34 specimens from five species of gastropods using a PSP bioassay. Five species of gastropods, Natica vitellus, Natica tumidus, Oliva hirasei, Oliva lignaria, and Oliva annulata, were collected from the coastal seawaters in Nha Trang City, Vietnam, between August 2007 and October 2007. The average lethal potency of gastropod specimens was 90 ± 40 (mean ± standard deviation) mouse units (MU) for N. vitellus, 64 ± 19 MU for N. tumidus, 42 ± 28 MU for O. hirasei, 51 ± 17 MU for O. lignaria, and 39 ± 18 MU for O. annulata. All toxic extracts from the sample species were clarified using a C18 Sep-Pak solid-phase extraction column and a microcentrifuge filter prior to analysis. High-performance liquid chromatography coupled with fluorescence detection indicated that the toxins of the olive shell (O. hirasei, O. lignaria, and O. annulata) were mainly composed of saxitoxin (STX) (73–82%), gonyautoxin (GTX) 2, 3 (12–22%), and minor levels of TTX (5–6%). The toxins of N. vitellus and N. tumidus were mainly composed of STX (76–81%) and GTX 1, 4 (19–24%). Furthermore, liquid chromatography–tandem mass spectrometry analysis was used to verify the identity of the PSPs and TTX. Our evidence shows that these gastropods have novel toxin profiles.     

Determination of paclitaxel and other six taxoids in Taxus species by high-performance liquid chromatography–tandem mass spectrometry

A method of high-performance liquid chromatography–tandem mass spectrometry (LC–MS-MS) has been developed for the trace analysis of paclitaxel and other six taxoids in three Taxus species including Taxus cuspidata, Taxus media and Taxus chinensis var. mairiei. Seven taxoids were separated using a gradient mode on an Eclipse XDB-C18 column (4.6 × 150 mm; i.d., 5 μm) at 20 °C. The compound separations were detected by an API 3000 mass spectrometer equipped with a TurboIonSpray™ interface. The compounds were detected using electrospray ionization (ESI) in positive-ion mode and quantified by multiple-reaction monitoring (MRM) mode using the transition mass of m/z 567.4 → 445.4, 609.3 → 549.5, 944.9 → 286.4, 812.6 → 286.1, 832.8 → 264.1, 854.4 → 286.1, and 812.6 → 286.1 for 10-DAB III, baccatin III, 7-xyl-10-DAT, 10-DAT, cephalomannine, paclitaxel, and 7-epi-10-DAT, respectively. The ranges of limit of quantitation (LOQ) for taxoids were 32, 14, 26, 14, 20, 32, and 26 ng/mL for 10-DAB III, baccatin III, 7-xyl-10-DAT, 10-DAT, cephalomannine, paclitaxel, and 7-epi-10-DAT, respectively. Linearity was confirmed over the whole calibration range (0.07–45, 0.058–37.5, 0.058–37.5, 0.056–36.3, 0.053–33.8, 0.057–37.5, and 0.06–38.8 μg/mL for 10-DAB III, baccatin III, 7-xyl-10-DAT, 10-DAT, cephalomannine, paclitaxel, and 7-epi-10-DAT, respectively) with coefficients higher than 0.9903. The inter- and intra-day precision of taxoids ranged from 2.86% to 6.31% for retention time and ranged from 3.91% to 7.33% for peak area. The recovery rates of this method were higher than 94.32% for 10-DAB III, 94.68% for baccatin III, 93.65% for 7-xyl-10-DAT, 93.29% for 10-DAT, 92.91% for cephalomannine, 93.41% for paclitaxel, and 93.06% for 7-epi-10-DAT, respectively.     

Determination of articaine in human plasma by liquid chromatography–mass spectrometry and its application in a preliminary pharmacokinetic study

A specific liquid chromatography–mass spectrometric (LC–MS) method using an ion trap spectrometer was developed for the quantitation of articaine in human plasma. Articaine and the internal standard (trazodone) were extracted in a single step with diethyl-ether from 0.5 mL of alkalinized plasma. The mobile phase consisted of acetonitrile with 0.1% formic acid (40:60, v/v). It was delivered at a flow rate of 0.3 mL/min. The effluent was monitored by MS in positive-ion mode. Ionisation was performed using an electrospray ion source operating at 200 °C. Articaine was identified and quantified in SIM mode at m/z 185. Calibration curves were linear over the concentration range of 78.1–5000 ng/mL with determination coefficients > 0.996. This method was fast (total run-time < 3 min), accurate (bias < 16%), and reproducible (intra-assay and inter-assay precision < 14%) with a quantitation limit of 78.1 ng/mL. The good specificity and sensitivity achieved by this method allowed the determination of articaine plasma levels in patients following a submucosal infiltration injection of articaine in the patients undergoing a third molar surgery.     

Quantitative determination of pimozide in human plasma by liquid chromatography–mass spectrometry and its application in a bioequivalence study

A simple, sensitive and specific LC–ESI/MS method was developed for the determination of pimozide in human plasma. Pimozide and cinnarizine (internal standard) were isolated from plasma samples by liquid–liquid extraction. The chromatographic separation was accomplished on a Thermo Hypersil-HyPURITY C18 reversed-phase column (150 mm × 2.1 mm, i.d., 5 μm) with the mobile phase consisting of 5 mM ammonium acetate (pH 3.5, adjusted with acetic acid)–methanol–acetonitrile (39:5:56, v/v/v). The lower limit of quantification was 0.02 ng/mL, and the assay exhibited a linear range of 0.025–12.800 ng/mL. The established method has been successfully applied to a bioequivalence study of 2 pimozide formulations in 32 healthy male Chinese volunteers.