Ion Trap Triple Quadrupole Mass Research Articles

LC-MS/MS-MS3 for determination and quantification of Δ9-tetrahydrocannabinol and metabolites in blood samples

Due to the high prevalence of cannabinoids in forensic toxicology, it is crucial to have an efficient method that allows the use of a small sample amount and that requires a minimal sample preparation, for the determination and quantification of low concentrations. A simple, highly selective and high throughput liquid chromatography tandem mass spectrometry method (LC-MS/MS-MS3) was developed for the determination and quantification of Δ9-tetrahydrocannabinol (THC), 11-hydroxy-Δ9-THC (THC-OH) and 11-nor-Δ9-THC-9-carboxylic acid (THC-COOH) in blood samples. Chromatographic analysis was preceded by a protein precipitation of 0.1 mL of blood samples with acetonitrile, then THC, THC-OH, THC-COOH and deuterated internal standards were separated on an Acquity UPLC® HSS T3 (100 mm × 2.1 mm i.d., 1.8 mm) reversed-phase column using a gradient elution with 2 mM aqueous ammonium formate (0.1% formic acid) and methanol, at a flow rate of 0.4 mL/min, and with a run time of 10 min. For MS/MS-MS3 analysis, a SCIEX QTRAP® 6500+ linear ion trap triple quadrupole mass spectrometer was used via electrospray ionization (ESI), operated in multiple reaction mode (MRM) and linear ion trap mode (MS3). The method was validated in accordance with international accepted criteria and guidelines. The method was selective and linear between 0.5–100 ng/mL (r2 > 0.995). The lower limits of quantification (LLOQ) corresponded to the lowest concentrations used for the calibration curves. The coefficients of variation obtained for accuracy and precision were less than 15%. The mean recoveries were between 88.0–101.4% for the studied concentration levels (1 ng/mL, 5 ng/mL and 50 ng/mL). No significant interfering compounds, matrix effects or carryover were observed. The validated method provides a sensitive, efficient and robust procedure for the quantitation of cannabinoids in blood using LC–MS/MS-MS3 and a sample volume of 0.1 mL. This work is also a proof of concept for using LC-MS3 technique to determine drugs in biological samples.

LC-MS3 Strategy for Quantification of Carbamazepine in Human Plasma and Its Application in Therapeutic Drug Monitoring.

This study developed a detection method based on the strategy of HPLC/MS3 and verified its suitability by quantifying carbamazepine in human plasma. The high-performance liquid chromatography–tandem mass spectrometry (HPLC/MS3) system was performed using a Shimadzu UFLC XR liquid chromatography and a SCIEX QTRAP® 5500 linear ion trap triple quadrupole mass spectrometer. The specific operation was as follows: the sample protein was firstly precipitated using methanol, then carbamazepine and carbamazepine-D2N15 were separated on an ACQUITY UPLC HSS T3 column using the gradient elution with solvent A (0.1% formic acid) and solvent B (0.1% formic acid in acetonitrile) at a flow rate of 0.25 mL/min. Each sample was run for 7 min. This method was validated for various parameters including accuracy, precision, selectivity, linearity, LLOQ, etc. Only 5 μL of sample plasma could obtain the result of LLOD 0.5 µg/mL. The intra-day and inter-day precision was <8.23%, and accuracy was between −1.74% and 2.92%. This method was successfully used for monitoring the blood concentration of epilepsy patients after carbamazepine treatment.

A LC-MS3 strategy to determine lamotrigine by Q-Q-trap tandem mass spectrometry coupled with triple stage fragmentation to enhance sensitivity and selectivity.

A high-performance liquid chromatography tandem mass spectrometry cubed (HPLC/MS3) method was developed and validated to quantify lamotrigine in human plasma with carbamazepine as an internal standard. The HPLC/MS/MS system is composed of a Shimadzu UFLC XR high-performance liquid chromatograph coupled with a hybrid linear ion trap triple quadrupole mass spectrometer. Following simple protein precipitation with methanol, the separation of lamotrigine and carbamazepine was performed on an Agilent Poroshell 120 SB-C18 column (4.6 × 50 mm, 2.7 μm) using gradient elution with 0.1% formic acid in water (solvent I) and 0.1% formic acid in methanol (solvent II) at a flow rate of 0.8 mL min-1. The total run time for each sample was 5 min. The method was validated for accuracy, precision, linearity, lower limit of quantification (LLOQ), selectivity, and other parameters. The LC/MS3 method was linear in the concentration range of 0.50-50.0 μg mL-1 (R2 ≥ 0.995). The LLOQ was 0.5 μg mL-1, requiring only 30 μL of human plasma. Intra- and inter-day accuracies were <6.17% and precisions were <11.4% at all concentrations. The absolute recoveries (%) and matrix effect (%) for lamotrigine in human plasma were between 83.8 and 90.7. The developed and validated LC-MS3 assay was successfully applied to monitor the lamotrigine levels in human plasma after the administration of lamotrigine.

Phytochemical profiling of the stem bark of Betula utilis from different geographical regions of India using UHPLC-ESI-MS/MS.

A simple, rapid, sensitive, and reliable ultra-high-performance liquid chromatography-hybrid linear ion trap triple quadrupole mass spectrometry method was developed and validated for simultaneous determination of 10 bioactive compounds in stem bark of Betula utilis grown in high altitude of Himalaya, India. The objective of the study is to develop and validate ultra-high-performance liquid chromatography-hybrid linear ion trap triple quadrupole mass spectrometry for investigation of geographical variations of triterpenoids, phenolics, and flavonoids contents in stem bark of B. utilis. The validated method was successfully applied to investigate geographical variations of triterpenoids, phenolics, and flavonoids in stem bark of B. utilis. The contents of betulinic acid and oleanolic acid were detected higher among selected analytes. The present variation study reveals great importance for the application and overall assessment of B. utilis.

In vitro and in vivo metabolite identification of a novel benzimidazole compound ZLN005 by liquid chromatography/tandem mass spectrometry.

A novel benzimidazole compound ZLN005 was previously identified as a transcriptional activator of peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α) in certain metabolic tissues. Upregulation of PGC-1α by ZLN005 has been shown to have a beneficial effect in a diabetic mouse model and in a coronary artery disease model in vitro. ZLN005 could also have therapeutic potential in neurodegenerative diseases involving down-regulation of PGC-1α. Given the phenotypic efficacy of ZLN005 in several animal models of human disease, its metabolic profile was investigated to guide the development of novel therapeutics using ZLN005 as the lead compound. ZLN005 was incubated with both rat and human liver microsomes and S9 fractions to identify in vitro metabolites. Urine from rats dosed with ZLN005 was used to identify in vivo metabolites. Extracted metabolites were analyzed by liquid chromatography/tandem mass spectrometry (LC/MS/MS) using a hybrid linear ion trap triple quadrupole mass spectrometer in full scan, enhanced product ion scan, neutral loss scan and precursor scan modes. Metabolites in plasma and brain of ZLN005-treated rats were also profiled using multiple reaction monitoring. Identified in vitro transformations of ZLN005 include mono- and dihydroxylation, further oxidation to carboxylic acids, and mono-O-glucuronide and sulfate conjugation to hydroxy ZLN005 as well as glutathione conjugation. Identified in vivo metabolites are mainly glucuronide and sulfate conjugates of dihydroxyl, carboxyl, and hydroxy acid of the parent compound. The parent compound as well as several major phase I metabolites were found in rat plasma and brain. Using both in vitro and in vivo methods, we elucidated the metabolic pathway of ZLN005. Phase I metabolites with hydroxylation and carboxylation, as well as phase II metabolites with glucuronide, sulfate and glutathione conjugation, were identified.

Quantification of coumarin and related phenolics in cinnamon samples from south India using UHPLC-ESI-QqQLIT-MS/MS method

ABSTRACTThe presence of coumarin, a hepatotoxic phenyl propanoid, in samples of the spice, cinnamon, procured from the market has raised food safety concerns world over. A validated, ultra-performance liquid chromatography- linear ion trap triple quadrupole mass spectrometry method was developed in the present study for the estimation of coumarin and other phenolics in authentic Cinnamomum verum bark samples and market samples from south India. Coumarin contents in authentic bark samples were in the range of 12.3 to 143.0 mg/kg, whereas the coumarin contents in samples procured from the market were up to 3462.0 mg/kg. The high content of coumarin and cinnamaldehyde in cinnamon samples procured from the market suggested possible blending with substitutes like Cinnamomum cassia barks, which possess substantial amounts of coumarin and cinnamaldehyde.

Major bioactive phenolics in Bergenia species from the Indian Himalayan region: Method development, validation and quantitative estimation using UHPLC-QqQLIT-MS/MS.

Bergenia species are important medicinal plants used in indigenous systems of medicine for their antilithiatic and diuretic properties. An ultra-high performance liquid chromatography coupled to hybrid linear ion trap triple quadrupole mass spectrometry (UHPLC-QqQLIT-MS/MS) method has been developed and validated for the estimation of quantitative variation of eight major bioactive phenolics in the rhizomes (150 samples) of four species of this herb, Bergenia (B. ciliata, B. ligulata, B. purpurascens and B. stracheyi). Chromatographic separation was obtained on a Waters ACQUITY UPLCTM BEH (ethylene bridged hybrid) C18 column with a mobile phase consisting of 0.1% (v/v) formic acid aqueous solution and acetonitrile under a gradient elution manner. A hybrid linear ion trap triple quadrupole mass spectrometer was operated in negative electrospray ionization mode with multiple reactions monitoring for detection and quantification of the eight compounds. The validated method demonstrated good linearity (r2 ≥ 0.9991), precision (RSD ≤ 1.87%) and accuracy (95.16–102.11%, RSD ≤ 1.83%) for all reference analytes. The quantitative results revealed that B. ligulata contains the highest amount of the major active marker-bergenin. The results also suggest that sensitive UHPLC-QqQLIT-MS/MS method, a sensitive, accurate and convenient one, could be helpful in identification of potential accession(s), rapid quality control and establishing authenticity of Bergenia species as raw material for pharmaceutical industries.

An Ultra-High Performance Liquid Chromatography-Tandem Mass Spectrometry Method for the Quantification of Vancomycin Requiring Only 2 &amp;#181;L of Rabbit Serum

A highly sensitive ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was developed for the quantification of vancomycin (VAN) in low volumes of rabbit serum. For each analysis, 2 μL rabbit serum was precipitated with methanol that contained the internal standard teicoplanin (TEI). The supernatant was transferred into a 384 well-plate, diluted with water, covered with a pierceable silicone mat and 5 μL was analyzed in positive ionization mode. The UHPLC-MS/MS consisted of an Agilent 1290 Infinity UHPLC system connected to an AB Sciex QTrap® 5500 hybrid linear ion-trap triple quadrupole mass spectrometer equipped with a Turbo Spray source. Chromatographic separation was achieved using a Waters Acquity UPLC BEH C18 (1.7 μm, 2.1 mm × 100 mm) column, a VanGuard (1.7 μm, 2.1 × 5 mm) guard column and a mobile phase of water and methanol both containing 5 mM ammonium acetate with 0.1% formic acid. VAN was quantified with multiple reaction monitoring using the transitions of m/z 725.5/144.2, and TEI was monitored at m/z 940.6/316.4. The accuracy, precision, linearity, range and lower limit of quantification (LLOQ) were determined. The accuracy was ≤9.93% and the precision was ≤10.6%. The range was established as 0.1 to 40 μg·mL-1. The LLOQ was 0.1 μg·mL-1 VAN requiring 2 μL of sample with an accuracy of -20.2% and precision of 8.39%. The method was applied successfully to determine the VAN concentrations in rabbit serum after the i.v. administration of VAN via implanted ear catheters.

Rapid and sensitive determination of major polyphenolic components in Euphoria longana Lam. seeds using matrix solid-phase dispersion extraction and UHPLC with hybrid linear ion trap triple quadrupole mass spectrometry.

A rapid and sensitive method for the extraction and determination of four major polyphenolic components in Euphoria longana Lam. seeds is presented for the first time based on matrix solid-phase dispersion extraction followed by ultra high performance liquid chromatography with hybrid triple quadrupole linear ion trap mass spectrometry. Matrix solid-phase dispersion method was designed for the extraction of Euphoria longana seed constituents and compared with microwave-assisted extraction and ultrasonic-assisted extraction methods. An Ultra high performance liquid chromatography with hybrid triple quadrupole linear ion-trap mass spectrometry method was developed for quantitative analysis in multiple-reaction monitoring mode in negative electrospray ionization. The chromatographic separation was accomplished using an ACQUITY UPLC BEH C18 (2.1 mm × 50 mm, 1.7 μm) column with gradient elution of 0.1% aqueous formic acid and 0.1% formic acid in acetonitrile. The developed method was validated with acceptable linearity (r2 > 0.999), precision (RSD ≤ 2.22%) and recovery (RSD ≤ 2.35%). The results indicated that matrix solid-phase dispersion produced comparable extraction efficiency compared with other methods nevertheless was more convenient and time-saving with reduced requirements on sample and solvent volumes. The proposed method is rapid and sensitive in providing a promising alternative for extraction and comprehensive determination of active components for quality control of Euphoria longana products.

A rapid and highly sensitive method for simultaneous determination of bioactive constituents in leaf extracts of six Ocimum species using ultra high performance liquid chromatography-hybrid linear ion trap triple quadrupole mass spectrometry

Ocimum species have tremendous value in pharmaceutical, perfumery, food processing and cosmetic industries, also in traditional rituals and medicines.

Simultaneous determination of multi-class bioactive constituents for quality assessment of Garcinia species using UHPLC–QqQLIT–MS/MS

Plants of genus Garcinia are therapeutically important and widely used in ayurvedic and traditional systems of medicine for the treatment of an array of ailments. In present study, a highly sensitive and efficient analytical method was developed and validated for rapid determination of twenty-six multi-class bioactive constituents in the leaf extracts of eleven Garcinia species using ultra high performance liquid chromatography-hybrid linear ion trap triple quadrupole mass spectrometry (UHPLC–QqQLIT–MS/MS). Chromatographic separation was accomplished on an Aquity UPLC BEH C18 column (50mm×2.1mm id, 1.7μm) using gradient elution of 0.1% formic acid in water and acetonitrile within 7.5min. Quantitative analysis was carried out by multiple-reaction monitoring mode (MRM) in negative electrospray ionization. The developed method was validated according to International Conference on Harmonization (ICH, Q2R1) guidelines. All standard calibration curves expressed good linear relationship (r2≥0.9991) over the concentration range of 0.1–300ng/mL. The precision was assessed by intra- and inter-day study which revealed RSD≤1.93%. The recoveries of quantified compounds were between 95.45 and 104.43% with RSD≤1.89%. This method was successfully applied to investigate quantitative variation among eleven Garcinia species. Moreover, principal component analysis (PCA) and hierarchical clustering analysis (HCA) were performed to compare and evaluate the quality of studied Garcinia species based on their quantitative data. The results indicated significant variation among eleven Garcinia species on the basis of twenty-six bioactive constituents. The developed method could be used as a tool for quality control and authenticity establishment of Garcinia species.

Preclinical pharmacokinetic evaluation of praziquantel loaded in poly (methyl methacrylate) nanoparticle using a HPLC–MS/MS

Praziquantel (PZQ) is the drug recommended by the World Health Organization for treatment of schistosomiasis. However, the treatment of children with PZQ tablets is complicated due to difficulties to adapt the dose and the extremely bitter taste of PZQ. For this reason, poly (methyl methacrylate) nanoparticles loaded with Praziquantel (PZQ-NP) were developed for preparation of a new formulation to be used in the suspension form. For this reason, the main aim of the present study was to evaluate the pharmacokinetic (PK) profile of PZQ-NP, through HPLC–MS/MS assays.Analyses were performed with an Omnisphere C18 column (5.0μm×4.6mm×150.0mm), using a mixture of an aqueous solution containing 0.1wt% of formic acid and methanol (15:85—v/v) as the mobile phase at a flow rate of 0.800mL/min. Detection was performed with a hybrid linear ion-trap triple quadrupole mass spectrometer with multiple reactions monitoring in positive ion mode via electrospray ionization. The monitored transitions were m/z 313.18>203.10 for PZQ and m/z 285.31>193.00 for the Internal Standard. The method was validated with the quantification limit of 1.00ng/mL, requiring samples of 25μL for analyses. Analytic responses were calibrated with known concentration data, leading to correlation coefficients (r) higher than 0.99. Validation performed with rat plasma showed that PZQ was stable for at least 10 months when stored below −70°C (long-term stability), for at least 17h when stored at room temperature (RT, 22°C) (short-term stability), for at least 47h when stored at room temperature in auto-sampler vials (post-preparative stability) and for at least 8 successive freeze/thaw cycles at −70°C.For PK assays, Wistar rats, weighing between 200 and 300g were used. Blood samples were collected from 0 to 24h after oral administration of single doses of 60mg/kg of PZQ-NP or raw PZQ (for the control group). PZQ was extracted from plasma by liquid–liquid extraction with terc-butyl methyl ether. The values obtained for maximum concentration (Cmax) and area under curve (AUC) for the PZQ-NP group were about 3 times smaller than the respective values obtained for the control group. However, the time for achieving maximum concentration (Tmax), the elimination constant (Ke) and the half-life time of elimination (T½β) were not statistically different. These results suggest that PZQ absorption is probably the rate-limiting step for obtainment of better PK parameters for PZQ-NP. Thus, further studies are needed to understand both the PZQ-NP absorption mechanisms and the drug diffusion process through the polymer matrix in vivo, in order to improve the PZQ-NP release profile.

P13 Distribution patterns of frequently prescribed veterinary drugs in horse hair

Horse hair analysis has proved to be a useful tool in testing for doping agents alternatively to urine due to its long window of detection. The aim of this study was to develop and validate an LC-MS/MS method for the detection and quantification of 13 frequently prescribed veterinary drugs. The method should be applied to 12 mane and tail hair samples from seven horses. Based on a hair growth rate of about 2 cm per month time of drug administration should be compared with the drug concentration found in the corresponding hair segment. Mane and/or tail hair were obtained from horses having received medical treatment. Time of drug administration was reported by the horse holders. Hair samples were washed with water and acetone, dried and cut in 2 or 4-cm-segments depending on total hair length (16 cm to 70 cm). Segments were cut into snippets and extracted for four hours with methanol under sonication. The extracts were analyzed by LC-ESI-MS/MS using a Dionex Ulti- Mate 3000 HPLC system and an Applied Biosystems 5500 QTrap linear ion trap triple quadrupole mass spectrometer operated in multiple reaction monitoring mode. Analytes were separated on a Kinetex C18-column (2,6 μm, 50 mm x 2,1 mm). The method was validated for the detection and quantification of tramadol, butorphanol, morphin, buprenorphine, acepromazine, chlorpromazine, fluphenazine, detomidine, flunixin, phenylbutazone, firocoxib, meloxicam and ketoprofen according to international guidelines. All analytes except fluphenazine and acepromazine fulfilled the main validation criteria. Two and 4-cm segments allowed a monitoring of one to two months period of drug administration. In fair mane and tail hair samples of horses no drugs could be detected. Concentrations in dark hair samples ranged up to 39 pg/mg for flunixin, 1.8 pg/mg for detomidine, 4 pg/mg for butorphanol, 1.1 pg/mg for tramadol, 1.4 pg/mg for morphine, 1.2 pg/mg for acepromazine, 1050 pg/mg for phenylbutazone and below the limit of quantification for meloxicam. In some cases, drugs were detected in the hair segment corresponding to the time of medical treatment. Drugs could be also detected in the vicinal segments. This drug distribution patterns suggest transport of drugs along the hair shaft. The study presents a sensitive method for the validated quantification of frequently prescribed veterinary drugs in horse hair. Hair analysis for these drugs could be used to test for medical treatment of horses to obtain indication of the horse's fitness, e.g. before purchase.

A validated assay to quantitate serotonin in lamb plasma using ultrahigh-performance liquid chromatography-tandem mass spectrometry: applications with LC/MS3.

An ultrahigh-performance liquid chromatography-tandem mass spectrometry (UHPLC/MS/MS) method was developed and validated for the quantification of serotonin (5-HT) in lamb plasma using [(2)d(4)]-serotonin ([(2)d(4)]-5-HT) as an internal standard. Charcoal-stripped human plasma was used as the blank matrix during validation, and 5-HT was quantitated using selected reaction monitoring. The UHPLC/MS/MS system consisted of an Agilent 1290 Infinity ultrahigh-performance liquid chromatograph coupled with an AB SCIEX QTRAP(®) 5500 hybrid linear ion trap triple quadrupole mass spectrometer. The method was validated for accuracy, precision, linearity, lower limit of quantification (LLOQ), selectivity, and other parameters. The LLOQ was 1.0 ng/mL, requiring 100 μL of sample. The method was applied to monitor the 5-HT levels in lamb plasma after the administration of fluoxetine. Tandem mass spectrometry cubed (MS(3)) experiments were also performed to investigate the fragmentation pattern of 5-HT and [(2)d(4)]-5-HT. A liquid chromatography-MS(3) (LC/MS(3)) method was developed, and the UHPLC/MS/MS and the LC/MS(3) methods were compared for performance.

Validated assay for the simultaneous determination of cortisol and budesonide in human plasma using ultra high performance liquid chromatography–tandem mass spectrometry

An ultra high performance liquid chromatography–tandem mass spectrometry (UHPLC/MS/MS) method was developed and validated for the quantification of cortisol and budesonide in human plasma. Charcoal stripped human plasma was used as the blank matrix during validation. Cortisol, budesonide, and dexamethasone (internal standard) were extracted from human plasma with methyl-tert-butyl ether, and the chromatographic separation of the peaks was achieved using a Waters Acquity UPLC BEH C18, 1.7μm, 2.1mm×50mm column with a run time of 4.0min. Cortisol, budesonide, and dexamethasone were monitored at the total ion current of their respective multiple reaction monitoring transition signals. The UHPLC/MS/MS system consisted of an Agilent 1290 Infinity ultra high performance liquid chromatograph coupled with an AB Sciex Qtrap® 5500 hybrid linear ion-trap triple quadrupole mass spectrometer. The method was validated for accuracy, precision, linearity, range, selectivity, lower limit of quantification (LLOQ), recovery, matrix effect, dilution integrity, and evaluation of carry-over. All validation parameters met the acceptance criteria according to regulatory guidelines. The LLOQ was 1.0ng/mL for both compounds requiring 100μL of sample. To our knowledge, this is the first validated LC/MS/MS method for the simultaneous quantitative analysis of cortisol and budesonide in human plasma. The method was applied successfully in a clinical investigation of the impact of nasally administered Pulmicort (budesonide) on the hypothalamic-pituitary-adrenal axis of patients with chronic rhinosinusitis.

Quantitative determination of lysophosphatidic acids (LPAs) in human saliva and gingival crevicular fluid (GCF) by LC–MS/MS

Lysophosphatidic acid (LPA) is a phospholipid mediator that plays multiple cellular functions by acting through G protein-coupled LPA receptors. LPAs are known to be key mediators in inflammation, and several lines of evidence suggest a role for LPAs in inflammatory periodontal diseases. A simple and sensitive liquid chromatography coupled with tandem mass spectrometry (LC–MS/MS) method has been developed and validated to quantify LPA species (LPA 18:0, LPA 16:0, LPA 18:1 and LPA 20:4) in human saliva and gingival crevicular fluid (GCF). LPA 17:0 was used as an internal standard and the LPA species were extracted from saliva by liquid–liquid extraction using butanol. Chromatography was performed using a Macherey-Nagel NUCLEODUR ® C8 Gravity Column (125 mm × 2.0 mm ID) with a mixture of methanol/water: 75/25 (v/v) containing 0.5% formic acid and 5 mM ammonium formate (mobile phase A) and methanol/water: 99/0.5 (v/v) containing 0.5% formic acid and 5 mM ammonium formate (mobile phase B) at a flow rate of 0.5 mL/min. LPAs were detected by a linear ion trap-triple quadrupole mass spectrometer with a total run time of 8.5 min. The limit of quantification (LOQ) in saliva was 1 ng/mL for all LPA species and the method was validated over the range of 1–200 ng/mL. The method was validated in GCF over the ranges of 10–500 ng/mL for LPA 18:0 and LPA 16:0, and 5–500 ng/mL for LPA 18:1 and LPA 20:4. This sensitive LC–MS/MS assay was successfully applied to obtain quantitative data of individual LPA levels from control subjects and patients with various periodontal diseases. All four LPA species were consistently elevated in samples obtained from periodontal diseases, which supports a role of LPAs in the pathogenesis of periodontal diseases.

A hybrid liquid chromatography–mass spectrometry strategy in a forensic laboratory for opioid, cocaine and amphetamine classes in human urine using a hybrid linear ion trap-triple quadrupole mass spectrometer

A rapid method has been developed to analyse morphine, codeine, morphine-3-glucuronide, 6-monoacetylmorphine, cocaine, benzoylegonine, buprenorphine, dihydrocodeine, cocaethylene, 3,4-methylenedioxyamphetamine, ketamine, 3,4-methylenedioxymethamphetamine, pseudoephedrine, lignocaine, benzylpiperazine, methamphetamine, amphetamine, 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine and methadone in human urine. Urine samples were diluted with methanol:water (1:1, v/v) and sample aliquots were analysed by hybrid linear ion trap-triple quadrupole mass spectrometry with a runtime of 12.5 min. Multiple reaction monitoring (MRM) as survey scan and an enhanced product ion (EPI) scan as dependent scan were performed in an information-dependent acquisition (IDA) experiment. Finally, drug identification and confirmation was carried out by library search with a developed in-house MS/MS library based on EPI spectra at a collision energy spread of 35 ± 15 in positive mode and MRM ratios. The method was validated in urine, according to the criteria defined in Commission Decision 2002/657/EC. At least two MRM transitions for each substance were monitored in addition to EPI spectra and deuterated analytes were used as internal standards for quantitation. The reporting level was 0.05 μg mL −1 for the range of analytes tested. The regression coefficients ( r 2) for the calibration curves (0–4 μg mL −1) in the study were ≥0.98. The method proved to be simple and time efficient and was implemented as an analytical strategy for the illicit drug monitoring of opioids, cocaines and amphetamines in criminal samples from crime offenders, abusers or victims in the Republic of Ireland. To the best of our knowledge there are no hybrid LC–MS applications using MRM mode and product ion spectra in the linear ion trap mode for opioids, cocaines or amphetamines with validation data in urine.

Comparison of triple quadrupole, hybrid linear ion trap triple quadrupole, time‐of‐flight and LTQ‐Orbitrap mass spectrometers in drug discovery phase metabolite screening and identification in vitro – amitriptyline and verapamil as model compounds

Liquid chromatography in combination with mass spectrometry (LC/MS) is a superior analytical technique for metabolite profiling and identification studies performed in drug discovery and development laboratories. In the early phase of drug discovery the analytical approach should be both time- and cost-effective, thus providing as much data as possible with only one visit to the laboratory, without the need for further experiments. Recent developments in mass spectrometers have created a situation where many different mass spectrometers are available for the task, each with their specific strengths and drawbacks. We compared the metabolite screening properties of four main types of mass spectrometers used in analytical laboratories, considering both the ability to detect the metabolites and provide structural information, as well as the issues related to time consumption in laboratory and thereafter in data processing. Human liver microsomal incubations with amitriptyline and verapamil were used as test samples, and early-phase 'one lab visit only' approaches were used with all instruments. In total, 28 amitriptyline and 69 verapamil metabolites were found and tentatively identified. Time-of-flight mass spectrometry (TOFMS) was the only approach detecting all of them, shown to be the most suitable instrument for elucidating as comprehensive metabolite profile as possible leading also to lowest overall time consumption together with the LTQ-Orbitrap approach. The latter however suffered from lower detection sensitivity and false negatives, and due to slow data acquisition rate required slower chromatography. Approaches with triple quadrupole mass spectrometry (QqQ) and hybrid linear ion trap triple quadrupole mass spectrometry (Q-Trap) provided the highest amount of fragment ion data for structural elucidation, but, in addition to being unable to produce very high-important accurate mass data, they suffered from many false negatives, and especially with the QqQ, from very high overall time consumption.

Increased Fragmentation Efficiency of Ions in a Low Pressure Linear Ion Trap with an Added dc Octopole Field

In-trap fragmentation of ions in a hybrid linear ion trap triple quadrupole mass spectrometer occurs at pressures about 5e-5 torr. At these low pressures, efficient fragmentation of heavy ions (such as the singly charged homogenously substituted triazatriphosphorine of mass 2721.89 Da) can take a long time because of the relatively low collision frequency with the background gas and the high internal energy content required to produce fragmentation. Increasing the amplitude used for dipolar excitation leads to loss of the ion upon the quadrupole rods. In the work presented here, the addition of a dc octopolar field to a linear ion trap is described. The dc octopolar field was created by the addition of four auxiliary electrodes situated between the quadrupole rods at a distance of 10 mm from the axis. The inclusion of the dc octopolar field was shown to cause the ions’ frequency of motion to shift out of phase with the excitation signal at high radial amplitudes. This resulted in beat-like trajectories with periods of excitation and de-excitation as the ions’ frequency of motion shifted in and out of phase with the excitation signal. This led to a reduction in the loss of ions on the quadrupole rods during the excitation process. The result is an increased fragmentation efficiency relative to the fragmentation efficiency obtained when using an LIT constructed of round rods only. The inclusion of the dc octopolar field allowed the ion to be fragmented more efficiently in a relatively short excitation period.