Quantitative analysis of biomarkers of liver and kidney injury in serum and urine using ultra‐fast liquid chromatography with tandem mass spectrometry coupled with a hydrophilic interaction chromatography column: Application to monitor injury induced by Euphorbiae pekinensis Radix

Fast and sensitive monitoring of drug‐induced liver and kidney injury in early stage is beneficial. An ultrafast liquid chromatography with tandem mass spectrometry assay was developed and validated to simultaneously determine ten endogenous biomarkers in serum and urine, including hippuric acid, phenylacetylglycine, 5‐oxoproline, cholic acid, taurine, indoleacetic acid, 3‐indoxyl sulfate, guanidinosuccinic acid, guanidinoacetic acid and uric acid. A CAPCELL CORE PC column (2.1 × 150 mm, 2.7 μm) was adopted for analytes separation. Gradient elution was performed with acetonitrile and water containing 5 mM ammonium acetate. Simple protein precipitation was applied in sample preparation. Good linearities were achieved with all the regression coefficients above 0.9911. Accuracy was 2.9–14.2% in serum and 4.1–14.6% in urine. The mean recovery was above 70% with acceptable matrix effects. The method was applied to monitor injury induced by Euphorbiae pekinensis Radix with a subacute rats model. All the biomarkers showed obvious concentration changes during the injury period. Furthermore, several biomarkers showed significant changes in earlier stage when compared with the current clinical serum bio‐parameters. The method might be helpful for early diagnosis of drug induced liver and kidney injury in clinical after tested on more drugs.     

Determination of sophoraflavanone G and kurarinone in rat plasma by UHPLC–MS/MS and its application to a pharmacokinetic study

This study aimed to develop and validate a simple and sensitive ultra high performance liquid chromatography tandem mass spectrometry method for the simultaneous determination of sophoraflavanone G and kurarinone in rat plasma by using rutin as the internal standard. Then, the developed method was applied to investigate the pharmacokinetics of sophoraflavanone G and kurarinone in rats after dosing the flavonoid extract from Sophora flavescens. Plasma samples were processed using a liquid–liquid extraction procedure with ethyl acetate. The analysis was performed on a triple quadrupole tandem mass spectrometer by multiple reaction monitoring with an electrospray ionization source in negative ionization mode. Quantitative ion transitions of m/z 423.2→161.2, 437.2→161.1, and 609.3→300.3 were monitored for sophoraflavanone G, kurarinone, and rutin, respectively. The calibration curves of the two analytes exhibited good linearity (r²>0.9923) over the range of 0.1–200 ng/mL for sophoraflavanone G and 0.1–1000 ng/mL for kurarinone. Relative standard deviations were less than 13.2% for the intra‐ and inter‐day precisions and no more than 12.6% for the recovery, showing good precision and satisfactory accuracy of the developed method. The validated method was successfully applied to the pharmacokinetic study of sophoraflavanone G and kurarinone after a single intravenous (25 mg/kg) and oral (500 mg/kg) administration of the flavonoid extract from S. flavescens, and the absolute bioavailability for sophoraflavanone G and kurarinone was about 36 and 17%, respectively.     

An LC–MS/MS method for rapid and sensitive high‐throughput simultaneous determination of various protein kinase inhibitors in human plasma

A reliable, high‐throughput and sensitive LC–MS/MS procedure was developed and validated for the determination of five tyrosine kinase inhibitors in human plasma. Following their extraction from human plasma, samples were eluted on a RP Luna®‐PFP 100 Å column using a mobile phase system composed of acetonitrile and 0.01 m ammonium formate in water (pH ~4.1) with a ratio of (50:50, v /v) flowing at 0.3 mL min⁻¹. The mass spectrometer was operating with electrospray ionization in the positive ion multiple reaction monitoring mode. The proposed methodology resulted in linear calibration plots with correlation coefficients values of r ² = 0.9995–0.9999 from concentration ranges of 2.5–100 ng mL⁻¹ for imatinib, 5.0–100 ng mL⁻¹ for sorafenib, tofacitinib and afatinib, and 1.0–100 ng mL⁻¹ for cabozantinib. The procedure was validated in terms of its specificity, limit of detection (0.32–1.71 ng mL⁻¹), lower limit of quantification (0.97–5.07 ng mL⁻¹), intra‐ and inter assay accuracy (−3.83 to +2.40%) and precision (<3.37%), matrix effect and recovery and stability. Our results demonstrated that the proposed method is highly reliable for routine quantification of the investigated tyrosine kinase inhibitors in human plasma and can be efficiently applied in the rapid and sensitive analysis of their clinical samples.     

A rapid method with ultra‐high‐performance liquid chromatography–tandem mass spectrometry for simultaneous determination of five type B trichothecenes in traditional Chinese medicines

A speedy and selective ultra‐HPLC‐MS/MS method for simultaneous determination of deoxynivalenol (DON), 3‐acetyldeoxynivalenol (3‐ADON), 15‐ADON, nivalenol and fusarenon X in traditional Chinese medicines (TCMs) was developed. The method was based on one‐step sample cleanup using reliable homemade cleanup cartridges. A linear gradient mobile‐phase system, consisting of water containing 0.2% aqueous ammonia and acetonitrile/methanol (90:10, v/v) at a flow rate of 0.4 mL/min, and an Acquity UPLC HSS T3 column (100 mm×2.1 mm, 1.8 μm) were employed to obtain the best resolution of the target analytes. [¹³C₁₅]–DON was used as the internal standard to accomplish as accurate as possible quantitation. The established method was further validated by determining the linearity (R²≥0.9990), sensitivity (LOQ, 0.29–0.99 μg/kg), recovery (88.5–119.5%) and precision (RSD≤15.8%). It was shown to be a suitable method for simultaneous determination of DON, 3‐ADON, 15‐ADON, nivalenol and fusarenon X in various TCM matrices. The utility and practical impact of the method was demonstrated using different TCM samples.     

Rapid discovery of absorbed constituents and metabolites in rat plasma after the oral administration of Zi Shen Wan using high‐throughput UHPLC–MS with a multivariate analysis approach

Zi Shen Wan is a typical formula consisting of three herbs, Phellodendri Amurensis Cortex, Rhizoma Anemarrhenae, and Cortex Cinnamomi, and has been widely used for treating prostatitis and infection diseases. However, it lacks in‐depth research of the constituents of Zi Shen Wan in vivo and in vitro. In this work, ultra high performance liquid chromatography coupled with quadrupole‐time‐of‐flight mass spectrometry and MassLynx software was established to characterize the chemical compositions of Zi Shen Wan in vivo and in vitro. In total, 92 peaks were characterized in vitro and 33 peaks were characterized in vivo based on mass spectrometry and tandem mass spectrometry data. Among the 33 compounds characterized in rat plasma, 22 prototype components absorbed in rat serum and 11 metabolites were identified in vivo. This work was fully reports the chemical constituents of traditional Chinese formula of Zi Shen Wan, it demonstrated that ultra high performance liquid chromatography combined with quadrupole time‐of‐flight mass spectrometry coupled to MassLynx software and multivariate data processing approach could be successfully applied for rapid screening and comprehensive analysis of chemical constituents in vitro and prototype components or metabolites in vivo of traditional Chinese medicine.     

Enhancement by Glycyrrhizae Radix of hepatic metabolism of hypaconitine,a major bioactive and toxic component of Aconiti Laterlis Radix,evaluated by HPLC‐TQ‐MS/MS analysis

Glycyrrhizae Radix (GR) is often prescribed together with Aconiti Laterlis Radix (ALR) (a so‐called compatible drug pair) in traditional Chinese medicinal practice to reduce toxicity of ALR. However, the mechanisms involved remain to be addressed. In this study, the metabolic interactions between GR–ALR drug pair were investigated for the first time. First, an HPLC‐TQ‐MS/MS method was developed to analyze hypaconitine, a major bioactive and toxic component of ALR, in rat liver S9. Then the in vitro metabolic rates of hypaconitine by different rat liver S9 were compared using the established method. The experiments were designed in four groups: pure hypaconitine (group I) and ALR extract (group II) incubated with liver S9 of normal rats, and pure hypaconitine (group III) and ALR extract (group IV) incubated with liver S9 of GR‐pretreated rats. When incubated for more than 4 h, the metabolic rates of hypaconitine in group III were significantly higher than those in group I, and when incubated for more than 2 h, the metabolic rates of hypaconitine in group IV were significantly higher than those in group II, suggesting that GR can enhance metabolic rate of hypaconitine, the mechanism of which might be related to hepatic metabolizing enzyme induction by GR. Copyright © 2012 John Wiley & Sons, Ltd.     

Development and validation of a sensitive and high‐throughput LC‐MS/MS method for the simultaneous determination of esomeprazole and naproxen in human plasma

A sensitive and high‐throughput LC‐MS/MS method has been developed and validated for the combined determination of esomeprazole and naproxen in human plasma with ibuprofen as internal standard. Solid‐phase extraction was used to extract both analytes and internal standard from human plasma. Chromatographic separation was achieved in 4.0 min on XBridge C₁₈ column using acetonitrile–25 mM ammonium formate (70:30, v/v) as mobile phase. Mass detection was achieved by ESI/MS/MS in negative ion mode, monitoring at m/z 344.19 → 194.12, 229.12 → 169.05 and 205.13 → 161.07 for esomeprazole, naproxen and IS, respectively. The calibration curves were linear from 3.00 to 700.02 ng/mL for esomeprazole and 0.50 to 150.08 ng/mL for naproxen. The intra‐ and inter‐batch precision and accuracy across four quality control levels met established criteria of US Food and Drug Administration guidelines. The assay is suitable for measuring accurate esomeprazole and naproxen plasma concentrations in human bioequivalence study following combined administration. Copyright © 2013 John Wiley & Sons, Ltd.     

Simultaneous identification of three pseudoallergic components in Danshen injection by using high‐expression Mas‐related G protein coupled receptor X2 cell membrane chromatography coupled online to HPLC–ESI‐MS/MS

Adverse drug reactions of Danshen injection mainly manifested as pseudoallergic reactions. In the present study, salvianolic acid A and a pair of geometric isomers (isosalvianolic acid C and salvianolic acid C) were identified as pseudoallergic components in Danshen injection by a high‐expression Mas‐related G protein coupled receptor X2 cell membrane chromatography coupled online with high‐performance liquid chromatography with electrospray ionization tandem mass spectrometry. Their pseudoallergic activities were evaluated by in vitro assay, which were consistent with the retention times on the cell membrane chromatography column. Salvianolic acid C, the most outstanding compound, was further found to induce pseudoallergic reaction through Mas‐related G protein coupled receptor X2. All the results above indicated that the system developed in this study is an effective method for simultaneously analyzing pseudoallergic components, even those with similar structures and the microcomponents in complex samples (salvianolic acid C in Danshen injection).     

A highly sensitive method for the quantification of fludrocortisone in human plasma using ultra‐high‐performance liquid chromatography tandem mass spectrometry and its pharmacokinetic application

A simple and high sensitive ultra‐high‐performance liquid chromatography tandem mass spectrometry method for the determination of fludrocortisone in human plasma was developed and validated as per guidelines. The analyte and internal standard (IS), fludrocortisone‐d₅, were extracted from human plasma via liquid–liquid extraction using tert‐butyl methyl ether. The chromatographic separation was achieved on a Chromolith RP_18e column using a mixture of acetonitrile and 2 mm ammonium formate (70:30, v/v) as the mobile phase at a flow rate of 0.7 mL/min. Quantitation was performed on a triple quadrupole mass spectrometer employing electrospray ionization technique, operating in multiple reaction monitoring and positive ion mode. The precursors to product ion transitions monitored for fludrocortisone and IS were m/z 381.2 → 343.2 and 386.2 → 348.4, respectively. The assay was validated with linear range of 40–3000 pg/mL. The intra‐ and inter‐day precisions (relative standard deviation) were within 0.49–7.13 and 0.83–5.87%, respectively. The proposed method was successfully applied to pharmacokinetic studies in humans. Copyright © 2015 John Wiley & Sons, Ltd.     

Ultra‐fast LC–ESI‐MS/MS method for the simultaneous determination of six highly toxic Aconitum alkaloids from Aconiti kusnezoffii radix in rat plasma and its application to a pharmacokinetic study

A fast, sensitive, and efficient ultra‐fast LC–ESI‐MS/MS method was developed for the simultaneous quantitation of six highly toxic Aconitum alkaloids, that is, aconitine, mesaconitine, hypaconitine, benzoylaconine, benzoylmesaconine, and benzoylhypaconine, in rat plasma after oral administration of crude ethanol extracts from Aconiti kusnezoffii radix by ultrasonic extraction, reflux extraction for 1 h, and reflux extraction for 3 h, respectively. The separation of six Aconitum alkaloids and aminopyrine (internal standard) was performed on an InertSustain® C₁₈ column, and the quantification of the analytes was performed on a 4000Q ultra‐fast LC–MS/MS system with turbo ion spray source in the positive ion and multiple‐reaction monitoring mode. Absolute recoveries ranged within 65.06–85.1% for plasma samples. The intra‐ and interday precision and accuracy of analytes were satisfactory. The methods were validated with sensitivity reaching the lower LOQ for aconitine, mesaconitine, hypaconitine, benzoylaconine, benzoylmesaconine, and benzoylhypaconine, which were 0.025, 0.025, 0.050, 0.025, 0.025, and 0.100 ng/mL, respectively. The method was successfully applied to a pharmacokinetic study of six Aconitum alkaloids in rat plasma after oral administration of crude ethanol extracts from the raw root of Aconitum kusnezoffii Reichb. by three different extraction processes.     

Development of rapid and high‐throughput LC–MS/MS method for quantification of olprinone in rabbit plasma

A simple, highly sensitive and rapid method for quantification of olprinone (phosphodiesterase 3 inhibitor) in rabbit plasma using liquid chromatography–tandem mass spectrometry with electrospray was developed. An aliquot of 50 μL of plasma sample was cleaned up and extracted using Ostro? 96‐well plate followed by dilution. Chromatographic separation of olprinone and olprinone‐d3 was carried out on a CORTECS^® T3 column within 3 min. Detection was achieved using a triple quadrupole mass spectrometer employing electrospray ionization operated in positive ion multiple reaction monitoring mode using the transitions m/z 251.07 → m/z 155.06 and m/z 254.21 → m/z 158.10 for olprinone and olprinone‐d3, respectively. The method was validated according to US Food and Drug Administration guideline for bioanalytical methods, and showed excellent linearity in the range 10.0–2000.0 ng/mL with coefficient of determination >0.99. The intra‐ and inter‐day precisions (CV) were <5.1% and the accuracies were within the range 99.7–103.2% at all quality control concentrations. Furthermore, olprinone was stable under various stability conditions. The developed method was used for quantification of olprinone in rabbit plasma after its intravenous administration at the dose of 1 mg/kg in order to better understand the metabolism of olprinone in a rabbit model of lung injury.     

Pharmacokinetics study of multiple components absorbed in rat plasma after oral administration of Stemonae radix using ultra‐performance liquid‐chromatography/mass spectrometry with automated MetaboLynx software analysis

Stemonae radix (Stemona tuberosa Lour, Bai Bu) is a traditional Chinese medicinal (TCM) plant known for its antitussive and anti‐ectoparasitic activity; however, the in vivo pharmacokinetic of its multiple bioactive components remains unknown. In this article, UPLC‐Q‐TOF‐high‐definition mass spectrometry (HDMS) coupled with automated data analysis MetaboLynx? software together were first developed to screen the potentially bioactive components in the rat plasma after oral administration of Stemonae radix. Time course of the absorbed components of Stemonae radix was built to evaluate pharmacokinetic behaviors. This rapid automated analysis method was successfully applied for identification, screening, and monitoring of the 28 constituents absorbed and metabolized studies of Stemonae radix after oral administration to rats. The results showed that the ongoing changes of 28 constituents including eight parent compounds and 20 metabolites in vivo were observed to find biomarkers. From the angle of behavior in vivo, it suggested that croomine and tuberostemonine would be potential efficacy markers. This work also demonstrated that the pharmacokinetics‐based UPLC‐Q‐TOF‐HDMS can provide a reliable means of identifying and screening potentially bioactive components contributing to pharmacological effects of medicinal herbs, and to better clarify its action mechanism, further prospecting natural products in the search for new leads in drug discovery.     

A high‐throughput LC–MS/MS method for determination of flunarizine in human plasma: Pharmacokinetic study with different doses

A high‐throughput and sensitive liquid chromatography–tandem mass spectrometry (LC–MS/MS) method has been developed and validated for the determination of flunarizine in human plasma. Liquid–liquid extraction under acidic conditions was used to extract flunarizine and flunarizine‐d8 from 100 μL human plasma. The mean extraction recovery obtained for flunarizine was 98.85% without compromising the sensitivity of the method. The chromatographic separation was performed on Hypersil Gold C₁₈ (50 × 2.1 mm, 3 μm) column using methanol–10 mm ammonium formate, pH 3.0 (90:10, v/v) as the mobile phase. A tandem mass spectrometer (API‐5500) equipped with an electrospray ionization source in the positive ion mode was used for detection of flunarizine. Multiple reaction monitoring was selected for quantitation using the transitions, m/z 405.2 → 203.2 for flunarizine and m/z 413.1 → 203.2 for flunarizine‐d8. The validated concentration range was established from 0.10 to 100 ng/mL. The accuracy (96.1–103.1%), intra‐batch and inter‐batch precision (CV ≤ 5.2%) were satisfactory and the drug was stable in human plasma under all tested conditions. The method was used to evaluate the pharmacokinetics of 5 and 10 mg flunarizine tablet formulation in 24 healthy subjects. The pharmacokinetic parameters C_max and AUC were dose‐proportional.     

Comparative pharmacokinetic study on three formulations of Astragali Radix by an LC–MS/MS method for determination of formononetin in human plasma

Astragali Radix (AR) is a widely used traditional Chinese medicine for healing the cardiovascular, liver and immune systems. Recently, superfine pulverizing technology has been applied to developing novel formulations to improve bioavailability of the active constituents in herbs, such as ultrafine granular powder of AR. In this study, a universal and sensitive quantitative method based on LC–MS/MS was employed for determining formononetin, the main flavonoid in AR, in human plasma for comparative pharmacokinetics of three oral formulations of AR. Formononetin and IS (quercetin) were extracted by ethyl acetate from human plasma and were separated on a C₁₈ column with a mobile phase consisting of acetonitrile and 0.1% formic acid. Positive‐ion electrospray‐ionization mode was applied in mass spectrometric detection. The quantitative method was validated with regards to selectivity, linearity, accuracy and precision, matrix effect, extraction recovery and stability, and was applied to comparing the pharmacokinetics of ultrafine granular powder (UGP), ultrafine powder (UP) and traditional decoction pieces (TDP) of AR after oral administration. The peak concentration and areas under the concentration–time curve of formononetin in UGP and UP were significantly higher than those of TDP. UGP and UP could significantly improve the bioavailability of AR in human compared with TDP after oral administration.     

A validated HPLC‐MS/MS method for the quantification of caderofloxacin in human plasma and its application to a clinical pharmacokinetic study

A simple, selective and rapid HPLC‐MS/MS method was developed and validated for the determination of caderofloxacin in human plasma. Sparfloxacin was used as the internal standard (IS). After precipitation with methanol and dilution with the mobile phase, the samples were injected into the HPLC‐MS/MS system. The chromatographic separation was performed on a Zorbax XDB Eclipse C₁₈ column (150 × 4.6 mm, 5 µm) with a mobile phase of ammonium acetate buffer (20 mm, pH 3.0)–methanol, 45:55 (v/v). The MS/MS analysis was done in positive mode. The multiple reaction monitoring transitions monitored were m/z 412.3 → 297.1 for caderofloxacin and m/z 393.2 → 292.2 for the IS. The calibration curve was linear over the range of 50.0–8000 ng/mL with an aliquot of 100 μL plasma. The precision of the assay was 2.0–9.4 and 6.6–11.5% for the intra‐ and inter‐run variability, respectively. The intra‐ and inter‐run accuracy (relative error) was 4.4–10.0 and ?1.2–4.0%. The total run time was 3.5 min. The assay was fully validated in accordance with the US Food and Drug Administration guidance. It was successfully applied to a pharmacokinetic study of caderofloxacin in healthy Chinese volunteers. Copyright © 2015 John Wiley & Sons, Ltd.     

Characterization and identification of the metabolites of Menthae Haplocalycis Herba water extracts in rat plasma,urine, and feces by ultra‐high performance liquid chromatography with linear ion trap‐Orbitrap mass spectrometry

Menthae Haplocalycis Herba has been utilized for food and medicinal purposes in China for thousands of years. It has various efficacies, including dispelling wind and heat and relieving sore throat. M. Haplocalycis Herba has been also widely used in food, cosmetics, spices, and other fields. Exploring the constituents and detecting the metabolites of M. Haplocalycis are of great significance to clarify the effective substances. However, the in vivo metabolites of M. Haplocalycis Herba water extract are still unclear. Herein, a sensitive and specific method, ultra‐high performance liquid chromatography with linear ion trap‐Orbitrap mass spectrometry, established in this assay was used to study the metabolism of M. Haplocalycis Herba water extract in rat plasma, urine, and feces. We characterized and identified 9, 50, and 34 metabolites in plasma, urine, and feces, respectively. Seven metabolic pathways, including phase Ⅰ (isomerization, demethylation, hydroxylation, and dehydration) and phase Ⅱ (sulfation and glucuronidation) were mainly involved in the metabolism. It is the first systematic study on the metabolism of M. Haplocalycis Herba water extract in vivo, which enrich current understanding of the metabolic behavior of M. Haplocalycis Herba water extract and provide a metabolic rationale for further in‐depth in vivo biotransformation and pharmacokinetic analysis.     

Simultaneous quantification of atractyloside and carboxyatractyloside in rat plasma by LC‐MS/MS: Application to a pharmacokinetic study after oral administration of Xanthii Fructus extract

Xanthii Fructus is extensively used as an herbal medicine. Ingestion of this herb is associated with severe hepatotoxicity and nephrotoxicity. Atractyloside and carboxyatractyloside are two dominative toxic constituents in Xanthii Fructus. However, their pharmacokinetic study is lacking. In this study, a novel high‐performance liquid chromatography‐tandem mass spectrometry method was developed to simultaneously quantify the rat plasma concentrations of atractyloside and carboxyatractyloside. After protein precipitation, the analytes were chromatographic separated on a ZORBAX Eclipse Plus column (2.1 × 150 mm id, 5 µm) under gradient elute. In the negative electrospray ionization mode, the transitions at m/z 725.3→645.4 for atractyloside, m/z 769.3→689.4 for carboxyatractyloside, and m/z 479.2→121.1 for paeoniflorin (the internal standard) were acquired by multiple reaction monitoring. This analytical method showed good linearity over 1–500 ng/mL for atractyloside and 2–500 ng/mL for carboxyatractyloside with acceptable precision and accuracy. No matrix effect, instability and carryover occurred in the analysis procedure. The extraction recoveries were greater than 85.0%. This method was applied to a preliminary pharmacokinetic study by orally administering Xanthii Fructus extract (9 g/kg) to rats, which was useful to evaluate the role of these two compounds in Xanthii Fructus‐induced toxicity.     

Development and validation of a high‐throughput and robust LC–MS/MS with electrospray ionization method for simultaneous quantitation of oseltamivir phosphate and its oseltamivir carboxylate metabolite in human plasma for pharmacokinetic studies

A rapid, sensitive and rugged solid‐phase extraction ultraperformance liquid chromatography tandem mass spectrometry (LC‐MS/MS) method was developed for determination of oseltamivir phosphate (OP) and oseltamivir carboxylate (OC) in human plasma. The procedure for sample preparation includes a simple SPE extraction procedure coupled with a Chromatopack C₁₈ column (50 × 3.0 mm, i.d., 3.0 µm) with isocratic elution at a flow‐rate of 0.600 mL /min and acyclovir was used as the internal standard. The analysis was performed on a triple‐quadrupole tandem mass spectrometer by multiple reaction monitoring mode via electrospray ionization. Using 500 µL plasma, the methods were validated over the concentration ranges 0.92–745.98 and 5.22–497.49 ng/mL for OP and OC, with a lower limit of quantification of 0.92and 5.22 ng/mL. The intra‐ and inter‐day precision and accuracy of the quality control samples were within 10.1%. The recovery was 68.72, 70.66 and 71.59% for OP, OC and IS, respectively. Total run time was only 1.0 min. The method was highly reproducible with excellent chromatography properties. Copyright © 2010 John Wiley & Sons, Ltd.     

UHPLC–MS/MS method for the simultaneous quantitation of five anthraquinones and gallic acid in rat plasma after oral administration of prepared rhubarb decoction and its application to a pharmacokinetic study in normal and acute blood stasis rats

Prepared rhubarb, as one of the main processed products of rhubarb, has a good effect on promoting blood circulation. In this paper we describe a rapid, sensitive, and selective ultra‐fast liquid chromatography with tandem mass spectrometry method for simultaneous quantification of five anthraquinones (rhein, aloe‐emodin, chrysophanol, emodin, and physcion) and gallic acid in plasma. Chromatographic separation was performed on an Extend C₁₈ column at the temperature of 30°C using a mobile phase that consisted of 0.1% aqueous formic acid and acetonitrile. Satisfactory linearity, precision, accuracy, extraction recovery, and matrix effect have been achieved. Then, the validated method was successfully applied to a comparative pharmacokinetic study. The results might be helpful for guiding clinical application of prepared rhubarb in the future.     

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. Copyright © 2015 John Wiley & Sons, Ltd.