Chromatography-electrospray Ionization-mass Spectrometry Method Research Articles

Distribution of Vicine, Convicine and Levodopa in Faba Bean Plant Tissues Determined by UltraHigh Performance Liquid Chromatography-Electrospray Ionization Mass Spectrometry

Faba bean is an emerging pulse crop for the Canada Prairies and is becoming an increasingly important source of plant sourced protein. It also has a high content of levodopa (up to 0.1 % of the dry seed weight), a nutraceutical compound for treating Parkinson's Disease (PD). The presence of pyrimidine glycosides, vicine and convicine, however, restricts the use of faba due to their potential to cause favism in susceptible individuals. For the rapid separation and quantification of levodopa, vicine and convicine, a 5-min ultrahigh performance liquid chromatography-electrospray ionization mass spectrometry method (UHPLC-ESI MS) was developed. This method provided a wide linear range of quantification (4.8 nM to 60 µM for vicine, and 6.0 nM to 24 µM for convicine and levodopa), and low limit of detection (1.2 to 3.0 nM). The method was applied in the analysis of diverse faba bean samples, from dehulled mature faba bean and seed coat, to fresh plant tissues such as flowers, leaves, stems, roots and pods. The results indicated that fresh faba bean plant tissues, such as leaves and flowers, had the highest concentration of levodopa and low level of VC, which makes them an alternative source of plant-based l-DOPA with potential as a food-based treatment for Parkinson's Disease.

Effectiveness and Biochemical Impact of Flubendiamide and Flonicamid Insecticides against Bemisia tabaci (Hemiptera: Aleyrodidae) and Residue Dissipation in Cherry Tomato Plants and Soil under Greenhouse Conditions

Abstract We investigated the effectiveness and biochemical impact of the insecticides flubendiamide and flonicamid compared with azadirachtin and acetamiprid against the whitefly Bemisia tabaci (Grennadius) (Hemiptera: Aleyrodidae) infesting cherry tomato (Solanum lycopersicum var. cerasiforme L.) grown under greenhouse conditions. The dissipation of both insecticides in the plants and in the soil also was determined using the QuEChERS (quick, easy, cheap, effective, rugged, and safe) and liquid chromatography–electrospray ionization–mass spectrometry methods. Both insecticides were more effective for reducing B. tabaci populations than were either acetamiprid or azadirachtin. Biochemical analysis revealed that esterase may play an important role in whitefly adaptation to flubendiamide and flonicamid. The QuEChERS method was determined suitable for quickly detecting residues of flubendiamide and flonicamid in complex matrices. The recovery rates on tomato fruit samples were 92.8–106.0%, with a relative standard deviation (RSD) range of 0.46–2.65%. For soil samples, the recovery rates were 81.3–95.7% with RSDs of 1.20–3.86%. We further determined that flubendiamide had dissipation half-lives of 3.13, 3.63, and 3.68 d in tomato fruit, tomato leaves, and soil, respectively. Flonicamid had half-lives of 4.25, 3.54, and 2.60 d in fruit, leaves, and soil, respectively. These results suggest that preharvest intervals of 3 and 7 d are appropriate for flubendiamide and flonicamid, respectively, in cherry tomato production. The risk quotient was >1 by the day 5 after application; however, that value declined to <1 on day 7 after application, indicating little long-term risk to human health.

Jiaotai Pill () Alleviates Insomnia through Regulating Monoamine and Organic Cation Transporters in Rats.

To reveal the effect and mechanism of Jiaotai Pill (, JTP) on insomniac rats. The insomniac model was established by intraperitoneal injection of p-chlorophenylalanine (PCPA). In behavioral experiments, rats were divided into control, insomniac model, JTP [3.3 g/(kg•d)], and diazepam [4 mg/(kg•d)] groups. The treatment effect of JTP was evaluated by weight measurement (increasement of body weight), open field test (number of crossings) and forced swimming test (immobility time). A high performance liquid chromatography-electrochemical detection (HPLC-ECD) method was built to determine the concentration of monoamine transmitters in hypothalamus and peripheral organs from normal, model, JTP, citalopram [30 mg/(kg•d)], maprotiline [40 mg/(kg•d)] and bupropion [40 mg/(kg•d)] groups. Expressions of serotonin transporter (SERT), dopamine transporter (DAT), and norepinephrine transporter (NET) were analyzed by quantitative polymerase chain reaction (qPCR) and Western blot in normal, model and JTP groups. A high performance liquid chromatography-electrospray ionization mass spectrometry (HPLC-ESI-MS/MS) method was established to determine the pharmacokinetics, urine cumulative excretion of metformin in vivo, and tissue slice uptake in vitro, which were applied to assess the activity of organic cation transporters (OCTs) in hypothalamus and peripheral organs. Compared with the insomniac model group, the body weight and spontaneous locomotor were increased, and the immobility time was decreased after treatment with JTP (P<0.01). Both serotonin and dopamine contents in hypothalamus and peripheral organs were increased (P<0.01). The norepinephrine content was increased in peripheral organs and decreased in hypothalamus (P<0.05 or P<0.01). At the same time, SERT, DAT, OCT1, OCT2, and OCT3 were down-regulated in hypothalamus and peripheral organs (P<0.05). NET was down-regulated in peripheral organs and up-regulated in hypothalamus (P<0.05 or P<0.01). Moreover, the activity of OCTs in hypothalamus and peripheral organs was inhibited (P<0.05). JTP alleviates insomnia through regulation of monoaminergic system and OCTs in hypothalamus and peripheral organs.

Analysis of flavonoids with unified chromatography-electrospray ionization mass spectrometry-method development and application to compounds of pharmaceutical and cosmetic interest.

In this project, we aimed at analysing flavonoid-type compounds with unified chromatography (joining supercritical fluid chromatography and enhanced fluidity liquid chromatography with carbon dioxide-methanol mobile phases covering a wide range of compositions) and diode-array and electrospray ionization mass spectrometric detection (UC-DAD-ESI-MS). First, the chromatographic method was developed for 9 standard flavonoid molecules from three different families (flavanols, flavanones and flavonols, glycosylated or not), with a strong focus on mobile phase composition to achieve the elution of a wide range of flavonoids with good chromatographic quality (efficiency and resolution). For this purpose, two stationary phases were selected (ACQUITY UPC2 DEA and Diol), and five different additives (formic acid, citric acid, phosphoric acid, methanesulfonic acid and ammonium hydroxide) were successively introduced in the methanol co-solvent. The composition containing 0.1% methanesulfonic acid in methanol was retained as it provided the best chromatographic quality together with the possibility of hyphenating the chromatography to mass spectrometry. The DEA column appeared to provide the best efficiency and was retained for further method development. The gradient method was then optimized to achieve a fast analysis, which involved elution with a wide range of mobile phase composition (from 20 to 100% co-solvent in methanol) together with reversed flow rate and reversed pressure gradients at fixed temperature. The final gradient lasted 10min, followed by 2.5min of re-equilibration. Then, ESI-MS detection was optimized. Because the single-quadrupole mass spectrometer employed (ACQUITY UPC2 QDa) allowed the variation of only a few parameters, a design of experiments was used to define the best compromise for three parameters (probe temperature, cone voltage and capillary voltage). The make-up fluid introduced before entering the MS was also varied: different compositions of methanol-water containing either formic acid, ammonium hydroxide or sodium chloride were tested. The best results in terms of signal-to-noise ratio were obtained with methanol containing 20mM ammonium hydroxide and 2% water. The optimal UC-DAD-ESI-MS method was then applied to two different flavonoid formulation ingredients. The first one, hidrosmin (5-O-(β-hydroxyethyl)diosmin), is known for its vasoprotective properties and therefore employed in pharmaceutical formulations. The second one, α-glucosyl-hesperidin (sometimes referred to as vitamin P), is employed in cosmetic formulations. Identification of the major compounds in each sample was achieved with the help of MS detection. Graphical abstract.

Improved Dried Blood Spot-Based Metabolomics Analysis by a Postcolumn Infused-Internal Standard Assisted Liquid Chromatography-Electrospray Ionization Mass Spectrometry Method.

Dried blood spots (DBSs) have gained increasing attention recently with their growing importance in precision medicine. DBS-based metabolomics analysis provides a powerful tool for investigating new biomarkers. Until now, very few studies have discussed measures for improving analytical accuracy with the consideration of the special characteristics of DBSs. The present study proposed a postcolumn infused-internal standard (PCI-IS) assisted strategy to improve data quality for DBS-based metabolomics studies. An efficient sample preparation protocol with 80% acetonitrile as the extraction solvent was first established to improve the metabolite recovery. The PCI-IS assisted liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS) method was used to simultaneously estimate the blood volume and correct the signal change caused by ion source contamination and the matrix effect to evaluate the spot volume effect and hematocrit (Hct) variation effect on target metabolites. Phenylalanine-d8 was selected as the single PCI-IS to correct the matrix effect. For calibration of errors caused by the blood volume difference, 75% of the test metabolites showed good correlation (R2 ≥ 0.9) between the spot volume and the signal intensity after PCI-IS correction compared to less than 50% metabolites with good correlation before calibration. The spot volume was further calibrated by the same PCI-IS. Investigation of the Hct variation effect on target metabolites revealed that it affected the concentrations of metabolites in the DBS samples depending on their abundance in the red blood cell (RBC) or plasma; it is essential to preinvestigate the distribution of metabolites in blood to minimize the comparison bias in metabolomics studies. Finally, the PCI-IS assisted method was applied to study acetaminophen-induced liver toxicity. The results indicated that the proposed PCI-IS strategy could effectively remove analytical errors and improve the data quality, which would make the DBS-based metabolomics more feasible in real-world applications.

Liquid Chromatography/Mass Spectrometry Method for the Simultaneous Determination of Atorvastatin and Telmisartan in Human Plasma

A simple liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS) method has been developed and validated for the simultaneous determination of Atorvastatin (ATO) and telmisartan (TEL) in human plasma. After liquid-liquid extraction step, the analytes and internal standard, rosuvastatin (ROS) were separated using an isocratic mobile phase consisting of 10 mM ammonium acetate (pH=4)-methanol (20:80, v/v) on a reverse-phase C18 column. Detection was carried out on mass spectrometer by using electrospray ionization (ESI) source using their respective [M+H] + ions, m/z 482 for ROS, m/z 515 for TEL, and m/z 559 for ATO. For both drugs, the method was linear in the range of 1-35 ng/mL. Acceptable precision and accuracy were obtained for concentrations in the calibration range.

HILIC/ESI-MS determination of gangliosides and other polar lipid classes in renal cell carcinoma and surrounding normal tissues

Negative-ion hydrophilic liquid chromatography-electrospray ionization mass spectrometry (HILIC/ESI-MS) method has been optimized for the quantitative analysis of ganglioside (GM3) and other polar lipid classes, such as sulfohexosylceramides (SulfoHexCer), sulfodihexosylceramides (SulfoHex2Cer), phosphatidylglycerols (PG), phosphatidylinositols (PI), lysophosphatidylinositols (LPI), and phosphatidylserines (PS). The method is fully validated for the quantitation of thestudied lipids in kidney normal and tumor tissues of renal cell carcinoma (RCC) patients based on the lipid class separation and the coelution of lipid class internal standard with the species from the same lipid class. The raw data are semi-automatically processed using our software LipidQuant and statistically evaluated using multivariate data analysis (MDA) methods, which allows the complete differentiation of both groups with 100% specificity and sensitivity. In total, 21 GM3, 28 SulfoHexCer, 26 SulfoHex2Cer, 10 PG, 19 PI, 4 LPI, and 7 PS are determined in the aqueous phase of lipidomic extracts from kidney tumor tissue samples and surrounding normal tissue samples of 20 RCC patients. S-plots allow the identification of most upregulated (PI 40:5, PI 40:4, GM3 34:1, and GM3 42:2) and most downregulated (PI 32:0, PI 34:0, PS 36:4, and LPI 16:0) lipids, which are primarily responsible for the differentiation of tumor and normal groups. Another confirmation of most dysregulated lipids is performed by the calculation of fold changes together with T and p values to highlight their statistical significance. The comparison of HILIC/ESI-MS data and matrix-assisted laser desorption/ionization mass spectrometric imaging (MALDI-MSI) data confirms that lipid dysregulation patterns are similar for both methods. Graphical abstract ᅟ.

A Rapid Determination and Quantification of Three Biologically Active Polyisoprenylated Benzophenones using Liquid Chromatography-Tandem Mass Spectrometry (MRM) Method in Five Garcinia species from Cameroon

Following investigation of Garcinia genus, a sensitive, rapid and simple reversed-phase high performance liquid chromatography-electrospray ionization mass spectrometry method has been developed for the identification and quantification of three polyisoprenylated benzophenones, garcinol (1), isogarcinol (2) and 7- epi-clusianone (3), in the extracts of five Garcinia species from Cameroon. The separation of those compounds was achieved on a RP-18 column using a solvent system consisting of a mixture of acetonitrile-water-formic acid as a mobile phase in a gradient elution mode. The identification of the three compounds was determined on a triple quadripole mass spectrometer with ESI interface operating in the negative mode. A multiple reaction monitoring (MRM) method was developed for the quantification of these polyisoprenylated benzophenones in the extracts of the Garcinia species. The method was validated through intra- and inter-day precision, with the relative standard deviation (RSD) less than 6%, limits of detection (LOD) and limits of quantification (LOQ) &lt;1 ng. Overall recoveries ranged from 94% to 104%, with RSDs ranging from 0.8% to 4.5%. The results indicated that the fruits of G. preussii and the roots of G. brevipedicellata are good source of garcinol (1) and isogarcinol (2) respectively.

Determination of hexabromocyclododecanes in ambient air by high performance liquid chromatography- electrospray ionization-mass spectrometry

A high performance liquid chromatography-electrospray ionization-mass spectrometry (HPLC-ESI-MS) method was developed for the determination of hexabromocyclododecanes (HBCDs) in ambient air samples. The samples were extracted by Soxhlet extractor with hexane, then purified on the composite gel column. At first, the interfering substances were rinsed with 50 mL hexane and 100 mL hexane-dichloromethane (9:1, v/v), then 180 mL hexane -dichloromethane (4:1, v/v) was used to elute the targets. The compounds were separated by gradient elution with acetonitrile-methanol-water as mobile phases on a UF-ODS column (150 mm×2.1 mm, 3.0 μm). Electrospray ionization negative ion source and selective ion monitoring (SIM) mode were adopted in MS detection. The results showed that α -HBCD, β -HBCD and γ -HBCD could be well separated, and the chromatographic peak area ratio of α -HBCD, β -HBCD and γ -HBCD to internal standard D18- γ -HBCD with their concentrations had a good linear relationship, with the correlation coefficients (R) ≥ 0.9988. The limits of detection (LODs, S/N=3) of α -HBCD, β -HBCD and γ -HBCD were 0.4, 0.5 and 0.4 μg/L, respectively. The limits of quantification (LOQs, S/N=10) were 1.4, 1.6 and 1.3 μg/L, respectively. The method detection limits (MDL) were 0.13, 0.17 and 0.13 pg/m3 (n=5), respectively. The recoveries of HBCDs spiked in the actual air samples were in the range of 74.8%-95.8%. It is demonstrated that the method has high sensitivity and good selectivity, and can meet the requirement of monitoring and analyzing HBCDs in air samples.

Determination of anthocyanins in cherry and cranberry by high-performance liquid chromatography–electrospray ionization–mass spectrometry

Anthocyanins are a group of widespread natural phenolic compounds in vegetables and fruits. The anthocyanins have a wide range of applications due to the antioxidant, anticancer and anti-inflammatory properties. In this study, anthocyanins (delphinidin-3-o-glucoside, cyanidin-3-o-glucoside, pelargonidin-3-o-glucoside and malvidin-3-o-glucoside) in cherry and cranberry were determined using high-performance liquid chromatography–electrospray ionization–mass spectrometry (HPLC–ESI–MS). The anthocyanins were separated using gradient elution and a reserved-phase analytical column before identification by high-performance liquid chromatography–electrospray ionization–mass spectrometry. A high-performance liquid chromatography–electrospray ionization–mass spectrometry method was optimized for the determination of anthocyanins in cherry and cranberry. Furthermore, in this study, we investigated extraction conditions of fruit samples as well as determination of optimum HPLC–ESI–MS conditions. This study is novel in terms of simultaneously examining both optimization of HPLC parameters and extraction conditions. Obtained optimum conditions were used for the determination as the quantitative and qualitative analysis of anthocyanins in cherry and cranberry. The content of anthocyanins on the basis of wet weight in cherry and cranberry samples was determined for delphinidin-3-o-glucoside <d.l. (detection limit) and <d.l., for cyanidin-3-o-glucoside varied from 3.5 ± 0.4 to 8.3 ± 1.1 mg kg−1 (average 5.8 ± 0.8 mg kg−1) and 9.8 ± 1.4 to 18 ± 3.0 mg kg−1 (average 13.2 ± 1.8 mg kg−1), for pelargonidin-3-o-glucoside <d.l. and varied from 136 ± 19 to 233 ± 35 mg kg−1 (average 185.3 ± 28 mg kg−1), for malvidin-3-o-glucoside <d.l. and <d.l., respectively.

Quantitation of acotiamide in rat plasma by UHPLC-Q-TOF-MS: method development, validation and application to pharmacokinetics.

A novel, sensitive and selective ultra-high-performance liquid chromatography-electrospray ionization mass spectrometry method was developed and validated for the quantification of acotiamide (ACT), a first-in-class drug used in functional dyspepsia, in rat plasma. A simple protein precipitation method with acetonitrile as precipitating solvent was used to extract ACT from rat plasma. ACT and an internal standard (mirabegron, IS) were separated on an Agilent poroshell EC C18 column (50 × 3.0 mm, 2.7 µm) using methanol-10 mM ammonium acetate binary gradient mobile phase at a flow rate of 0.4 mL/min over 4 min run time. Detection was performed using target ions of [M + H](+) at m/z 451.2010 for ACT and m/z 397.1693 for IS in selective ion mode. The method was validated in the calibration range of 1.31-1000 ng/mL. All the validation parameters were well within the limits. The method demonstrated good performances in terms of intra- and inter-day precision (3.27-12.60% CV) and accuracy (87.96-104.94%). Thus the present ultra-high-pressure liquid chromatograhy-high-resolution mass spectrometry method for determination of ACT in rat plasma, is highly sensitive and rapid with a short run-time of 4 min, can be suitable for high sample throughput and for large batches of biological samples in pharmacokinetic studies. This method can be extended to measure plasma concentrations of ACT in humans to understand drug metabolism, drug interaction and adverse effects.

Continuous comprehensive two-dimensional liquid chromatography-electrospray ionization mass spectrometry of complex lipidomic samples.

A new continuous comprehensive two-dimensional liquid chromatography-electrospray ionization mass spectrometry method has been developed for the lipidomic characterization of complex biological samples. The reversed-phase ultra-high-performance liquid chromatography with a C18 column (150 mm × 1 mm, 1.7 μm) used in the first dimension makes the separation of numerous lipid species differing in their hydrophobic part of the molecule, mainly fatty acyl chain lengths and the number and positions of double bonds, possible. Coeluted lipid species in the first dimension are resolved by the fast hydrophilic interaction liquid chromatography separation (50 mm × 3 mm, 2.7 μm, core-shell particles) of lipid classes according to their different polarities in the second dimension. Retention times in both dimensions, accurate m/z values, and tandem mass spectra provide high confidence in the identification of lipid species. The retention behavior of individual lipids in reversed-phase mode follows the equivalent carbon number pattern, which provides an additional tool for unambiguous identification. This analytical method is applied for the lipidomic characterization of total lipid extracts of human plasma and porcine brain samples, which resulted in the identification of 143 lipid species from four lipid categories and ten lipid classes.

Quantitative analysis of bromate in non-alcoholic beer using ultra performance liquid chromatography-electrospray ionization mass spectrometry

Bromate (BrO3−) is a possible human carcinogen which is formed as a bromide containing source water ozone disinfection by-product. Thus, BrO3− is subject to assess its threat to humans. In the present study, an ultra performance liquid chromatography-electrospray ionization mass spectrometry method has been developed for the quantitative analysis of BrO3− in non-alcoholic beer. The chromatographic separation of both transitions 81BrO3− and 79BrO3− (isotope contributions of 79Br and 81Br) was carried out and achieved in 0.9999), precision (repeatability and reproducibility in terms of relative standard deviation) <1% at 1.0 μg mL−1 standard and <3% in the analysis of sample 1 (classic malt) and sensitivity with a low limit of detection (LOD, 81BrO3−, 0.03 ng mL−1 and 79BrO3−, 0.03 ng mL−1) and limit of quantification (LOQ, 81BrO3−, 0.1 ng mL−1 and 79BrO3−, 0.1 ng mL−1) in the standard. Nevertheless, the LOD and LOQ in sample 1 (classic malt) were obtained only for transition 81BrO3−, 0.05 ng mL−1 and 0.16 ng mL−1, respectively. For transition 79BrO3− the obtained peak was below the limit of detection. A total of 39 non-alcoholic beer samples of different flavors were analyzed and the 81BrO3− level in classic samples was found between 1.20 μg mL−1 and 3.73 μg mL−1, however, in other flavored samples the concentration levels reached up to 14.04 μg mL−1 with excellent recovery rates (96–99%). In most of the analyzed real samples, only transition 81BrO3− was found nevertheless, transition 79BrO3− was either <LOD or completely diminished from the obtained sample chromatogram. This is one of the main findings from the present study and the mechanism of this cause is unknown. The obtained BrO3− levels were much higher than the limits in drinking water (10–25 ng mL−1) recommended by several regulatory agencies.

Simultaneous determination of residues in pollen and high-fructose corn syrup from eight neonicotinoid insecticides by liquid chromatography–tandem mass spectrometry

The neonicotinoids have recently been identified as a potential contributing factor to the sudden decline in adult honeybee population, commonly known as colony collapse disorder (CCD). To protect the health of honeybees and other pollinators, a new, simple, and sensitive liquid chromatography-electrospray ionization mass spectrometry method was developed and validated for simultaneous determination of eight neonicotinoids, including acetamiprid, clothianidin, dinotefuran, flonicamid, imidacloprid, nitenpyram, thiacloprid, and thiamethoxam, in pollen and high-fructose corn syrup (HFCS). In this method, eight neonicotinoids, along with their isotope-labeled internal standards, were extracted from 2 g of pollen or 5 g of HFCS using an optimized quick, easy, cheap, effective, rugged, and safe extraction procedure. The method limits of detection in pollen and HFCS matrices were 0.03 ng/g for acetamiprid, clothianidin, dinotefuran, imidacloprid, thiacloprid, and thiamethoxam and ranged between 0.03 and 0.1 ng/g for nitenpyram and flonicamid. The precision and accuracy were well within the acceptable 20% range. Selectivity, linearity, lower limit of quantitation, matrix effect, recovery, and stability in autosampler were also evaluated during validation. This validated method has been used successfully in analyzing a set of pollen and HFCS samples collected for evaluating potential honeybee exposure to neonicotinoids.

A Rapid and Sensitive LC–MS Method for Determination of Ezetimibe Concentration in Human Plasma: Application to a Bioequivalence Study

A selective and highly sensitive high performance liquid chromatography-electrospray ionization mass spectrometry method has been developed for determination of ezetimibe concentrations in human plasma. Ezetimibe was extracted from plasma with ethyl acetate followed by evaporation of the organic layer and, then, reconstitution of the residue in mobile phase before injection to chromatograph. The mobile phase consisted of acetonitrile-ammonium acetate (10 mM, pH 3.0), 75:25 (v/v). An aliquot of 10 μL was chromatographically analyzed on a prepacked Zorbax XDB-ODS C18 column (2.1 × 100 mm, 3.5 micron). Detection of analytes was achieved by mass spectrometry with atmospheric pressure chemical ionization (APCI) interface in the negative ion mode operated under the multiple-reaction monitoring mode (m/z transition: ezetimibe 408–271). Standard curves were linear (r = 0.998) over the wide ezetimibe concentration range of 0.05–30.0 ng mL−1 with acceptable accuracy and precision. The limit of detection was 0.02 ng mL−1. The validated LC–APCI–MS method has been used successfully throughout a bioequivalence study on an ezetimibe generic product in 24 healthy male volunteers.

Exploration of toxicity reducing mechanism of aconite alkaloids during decoction process using liquid chromatography-mass spectrometry

A high performance liquid chromatography-electrospray ionization mass spectrometry method was developed for the determination of aconite alkaloids. It was used to investigate the degradation of alkaloids of Radix Aconiti Lateralis Preparata during decoction. Six alkaline degradation products were identified, and the degradation regularity of diester-diterpenoid alkaloids was confirmed during the test using standards. The dynamic changes of the amount of aconite alkaloids in the decoction of Radix Aconiti Lateralis Preparata were supervised. Along with the increase of decoction time, the concentrations of diester-diterpenoid alkaloids and lipo-alkaloid decreased significantly. The results can provide a scientific basis for the safety use of aconite.

Liquid chromatography–mass spectrometry method for the analysis of 1,4‐dimethylpyridinium in rat plasma – application to pharmacokinetic studies

A sensitive and specific liquid chromatography electrospray ionization-mass spectrometry method for determination of 1,4-dimethylpyridinium (1,4-DMP) in rat plasma has been developed and validated. Chromatography was performed on an Aquasil C(18) analytical column (4.6 × 150 mm, 5 µm, Thermo Scientific, Rockford, IL, USA) with isocratic elution using a mobile phase containing acetonitrile and water with an addition of 0.1% of formic acid. Detection was achieved by an Applied Biosystems MDS Sciex (Concord, Ontario, Canada) API 2000 triple quadrupole mass spectrometer. Electrospray ionization was used for ion production. The limit of detection in the single ion monitoring mode was found to be 10 ng/mL. The limit of quantification was 50 ng/mL. The precision and accuracy for both within-day and between-day determination of 1,4-dimethylpyridinium was 2.4-7.56 and 90.93-111.48%. The results of this analytical method validation allow pharmacokinetic studies to be carried out in rats. The method was used for the pilot study of the pharmacokinetic behavior of 1,4-DMP in rats after intravenous administration.

Development and validation of liquid chromatography–mass spectrometric method for simultaneous determination of moxifloxacin and ketorolac in rat plasma: application to pharmacokinetic study

A highly sensitive, selective and rapid liquid chromatography-electrospray ionization mass spectrometry (LC-MS) method has been developed and validated for simultaneous determination of moxifloxacin (MFX) and ketorolac (KTC) in rat plasma. Gemifloxacin (GFX) was used as an internal standard (IS). A simple protein precipitation method was used for the extraction of analytes from rat plasma. Effective chromatographic separation of MFX, KTC and GFX was achieved on a Kromasil C(18) column (100 × 4.6 mm, 5 µm) using a mobile phase consisting of acetonitrile-10 mm ammonium acetate (pH 2.5)-0.1% formic acid (50:25:25) in an isocratic elution, followed by detection with positive ion electrospray ionization mass spectrometry using target ions of [M + H](+) at m/z 402 for MFX, m/z 256 for KTC and m/z 390 for GFX in selective ion recording mode. The method was validated over the calibration range of 5-100 ng/mL for MFX and 10-6000 ng/mL for KTC. The method demonstrated good performances in terms of intra- and inter-day precision (0.97-5.33%) and accuracy (93.91-101.58%) for both MFX and KTC, including lower and upper limits of quantification. The recoveries from spiked control samples were >75% for MFX and >79% for KTC. The matrix effect was found to be negligible and the stability data were within acceptable limits. Further, the method was also successfully applied to a single-dose pharmacokinetic study in rats. This method can be extended to measure plasma concentrations of both drugs in human to understand drug interaction and adverse effects.

Simultaneous quantification of selective serotonin reuptake inhibitors and metabolites in human plasma by liquid chromatography–electrospray mass spectrometry for therapeutic drug monitoring

A simple and sensitive liquid chromatography–electrospray ionization mass spectrometry method was developed for the simultaneous quantification in human plasma of all selective serotonin reuptake inhibitors (citalopram, fluoxetine, fluvoxamine, paroxetine and sertraline) and their main active metabolites (desmethyl-citalopram and norfluoxetine). A stable isotope-labeled internal standard was used for each analyte to compensate for the global method variability, including extraction and ionization variations. After sample (250μl) pre-treatment with acetonitrile (500μl) to precipitate proteins, a fast solid-phase extraction procedure was performed using mixed mode Oasis MCX 96-well plate. Chromatographic separation was achieved in less than 9.0min on a XBridge C18 column (2.1×100mm; 3.5μm) using a gradient of ammonium acetate (pH 8.1; 50mM) and acetonitrile as mobile phase at a flow rate of 0.3ml/min. The method was fully validated according to Société Française des Sciences et Techniques Pharmaceutiques protocols and the latest Food and Drug Administration guidelines. Six point calibration curves were used to cover a large concentration range of 1–500ng/ml for citalopram, desmethyl-citalopram, paroxetine and sertraline, 1–1000ng/ml for fluoxetine and fluvoxamine, and 2–1000ng/ml for norfluoxetine. Good quantitative performances were achieved in terms of trueness (84.2–109.6%), repeatability (0.9–14.6%) and intermediate precision (1.8–18.0%) in the entire assay range including the lower limit of quantification. Internal standard-normalized matrix effects were lower than 13%. The accuracy profiles (total error) were mainly included in the acceptance limits of ±30% for biological samples. The method was successfully applied for routine therapeutic drug monitoring of more than 1600 patient plasma samples over 9 months. The β-expectation tolerance intervals determined during the validation phase were coherent with the results of quality control samples analyzed during routine use. This method is therefore precise and suitable both for therapeutic drug monitoring and pharmacokinetic studies in most clinical laboratories.

Development and Validation of a LC–MS/MS Method for the Determination of Ilaprazole Enantiomers and Its Application to a Bioequivalence Study in Healthy Chinese Volunteers

A sensitive and specific liquid chromatography–electrospray ionization–mass spectrometry method has been developed and validated for the identification and quantification of ilaprazole enantiomers in human plasma using granisetron as the internal standard. The method involves a simple liquid–liquid extraction. The analytes were separated by isocratic gradient elution on a Chiralcel OZ-RH (5 μm, 150 × 4.6 mm i.d.) column and analyzed in multiple-reaction-monitoring mode with positive electrospray ionization interface using the respective [M + H]+ ions, m/z 367.2 → m/z 183.8 for ilaprazole enantiomers, m/z 313.3 → m/z 138.1 for granisetron. The method was validated over the concentration range of 1.229–1,258 μg L−1 for d-ilaprazole and 1.880–1,926 μg L−1 for l-ilaprazole. The lower limit of quantification was 1.229 μg L−1 for d-ilaprazole and 1.880 μg L−1 for l-ilaprazole. The average extraction recovery of d-ilaprazole and l-ilaprazole was 78 and 71%, respectively. Finally, the validated method was applied to a bioequivalence study of ilaprazole in 24 healthy Chinese volunteers.