Selective Liquid Chromatography Research Articles

Percutaneous Microdialysis and Pharmacokinetic Study of Tongluo-Qutong Rubber Plaster in Rats by UPLC-MS/MS

Tongluo-Qutong rubber plaster (TQRP) is a well-known traditional Chinese medicine prescription for the treatment of osteoarthritis and cervical spondylosis. Due to a lack of in vivo permeation data, the active ingredients of TQRP have not been fully elucidated, presenting a huge obstacle to quality evaluation, pharmacokinetic studies, and safety assessment of TQRP for clinical application. In this study, a selective and reproducible ultraperformance liquid chromatography tandem mass spectrometry method was developed and validated for percutaneous microdialysis and pharmacokinetic experiments. In the percutaneous microdialysis study, the mean area under the concentration–time curve (AUC0-24h) of emodin (EMO) and piperine (PIP) were 127.1 and 2603.6 h·ng/mL, respectively. In the pharmacokinetic study, ferulic acid (FA), EMO, and PIP were determined in plasma samples. The mean AUC0-32h values of FA, EMO, and PIP in plasma were 15441.5, 202.0, and 1704.5 h·ng/mL, respectively. The in vivo exposure levels of active ingredients such as FA, EMO, and PIP after dermal administration of TQRP provide insights and data to support identification of its bioactive components and further study of its mechanism of action.

Driving Under the Influence of Drugs: A Single Parallel Monitoring-Based Quantification Approach on Whole Blood

Driving under the influence of psychoactive substances is a major cause of motor vehicle crashes. The identification and quantification of substances most frequently involved in impaired-driving cases in a single analytic procedure could be an important asset in forensic toxicology. In this study, a highly sensitive and selective liquid chromatography (LC) approach hyphenated with Orbitrap high-resolution mass spectrometry (HRMS) was developed for the quantification of the main drugs present in the context of driving under the influence of drugs (DUID) using 100 μL of whole blood. This procedure involves a simple sample preparation and benefit from the selectivity brought by parallel reaction monitoring (PRM) allowing to solve most DUID cases using a single multi-analyte injection. The method was fully validated for the quantification of the major classes of psychoactive substances associated with impaired-driving (cannabinoids, cocaine and its metabolites, amphetamines, opiates and opioids, and the major benzodiazepines and z-drugs). The validation guidelines set by the “Société Française des Sciences et des Techniques Pharmaceutiques” (SFSTP) were respected for 22 psychoactive substances using 15 internal standards. Trueness was measured to be between 95.3 and 107.6% for all the tested concentrations. Precision represented by repeatability and intermediate precision was lower than 12% while recovery (RE) and matrix effect (ME) ranged from 49 to 105% and from −51 to 3%, respectively. The validated procedure provides an efficient approach for the simultaneous and simple quantification of the major drugs associated with impaired driving benefiting from the selectivity of PRM.

Preclinical study of simultaneous pharmacokinetic and pharmacodynamic herb-drug interactions between Yin-Chen-Hao-Tang and spironolactone

BackgroundThe prevalence and therapeutic effects of the use of herbal remedies for chronic liver diseases make the combined administration of herbal products with conventional treatment unable to be ignored. This study investigated the pharmacokinetic and pharmacodynamic herb-drug interactions between the herbal formula Yin-Chen-Hao-Tang (YCHT) and spironolactone.MethodsA selective high-performance liquid chromatography (HPLC) method was developed and validated for the detection of spironolactone and its metabolite canrenone in rat urine. The interaction study was conducted by collecting urine samples after oral administration of spironolactone alone or in combination with YCHT for 5 days. Urine pharmacokinetic parameters and urinary sodium, potassium, volume, and weight were analyzed.ResultsThe results revealed significant increases in the cumulative amount and the area under the rate curve (AURC) of the metabolite canrenone after pretreatment with the high dose of YCHT. The urine weight and volume were significantly reduced dose-dependently as a result of pretreatment with YCHT. The urinary sodium-to-potassium ratio, which indicates diuretic effects, was also reduced in the high-dose YCHT condition.ConclusionsHerb-drug pharmacokinetic and pharmacodynamic interactions between YCHT and spironolactone were observed in the study. The herb-drug interaction that appeared with a single dose of spironolactone should be considered when patients are being treated with a continuous administration of this drug.

Development and validation of a sensitive HPLC-HESI-MS/MS method for quantitative determination of bitopertin in rat and marmoset plasma

Bitopertin is a potent glycine transporter 1 (GlyT1) inhibitor that has undergone clinical trials for diverse disorders and has a well-documented pharmacokinetic (PK) profile in humans. Even though pre-clinical studies have demonstrated potential therapeutic effects on cognition and neuropathic pain, the PK profile of bitopertin in the rat has been partly disclosed and no study reporting its PK profile in the common marmoset has been published. The aim of this study was to develop and validate a sensitive and selective high-performance liquid chromatography coupled with heat assisted electrospray ionisation tandem mass spectrometry (HPLC-HESI-MS/MS) assay to quantify bitopertin in the rat (Sprague-Dawley) and the common marmoset (Callithrix jacchus) plasma after administration of 1.0 mg/kg subcutaneously. The analytical method consisted of protein precipitation followed by HPLC-HESI–MS/MS. Chromatographic separation was carried out on a Thermo Scientific Aquasil C18 analytical column (100 x 2.1 mm I.D., 5.0 μm) kept at 50°C using acetonitrile and water both fortified at 0.1% (v/v) with formic acid at a ratio 55:45 as mobile phase with a constant flow rate of 250 μL/min. The calibration function was linear in the range of 0.3-200.0 ng/mL in rat plasma. The intra-day and inter-day precision and accuracy were within ± 15% at all concentrations. The limit of detection (LOD) and quantitation (LOQ) in rat plasma were 0.08 and 0.3 ng/mL, respectively. This method has demonstrated high sensitivity and specificity and was successfully applied to measure bitopertin in rat and marmoset plasma, allowing the investigation of its PK properties in both species.

Determination of levetiracetam in human plasma by online heart-cutting liquid chromatography: Application to therapeutic drug monitoring.

Levetiracetam is an antiepileptic drug for the treatment of psychiatric patients. In this study, a selective, straightforward, and rapid online heart-cutting liquid chromatography method was developed for the therapeutic drug monitoring of levetiracetam. This method allows for the determination of levetiracetam in human plasma without complex sample preparation. The mobile phases consisted of 30mM aq. orthophosphoric acid solution/methanol (70:30) at a flow rate of 1mL/min for the first system and 10mM aq. orthophosphoric acid solution/methanol (55:45) at a flow rate of 1mL/min for the second system. The first separation was carried out on a GL Sciences Intersil ODS-3 column (4.6mm × 150mm, 3µm) and the second separation was carried out on a Restek Ultra PFPP column (4.6mm × 150mm, 5µm). The detection was carried out at 205nm for both systems. The method was validated for selectivity and linearity, which were in the 6-60µg/mL range. Intra- and interassay accuracies were<112.6%, and the intra- and interassay precisions were<6.4% for all quality control samples. The lower limit of quantitation was 6µg/mL. The developed method was successfully applied for therapeutic drug monitoring of plasma samples from patients.

Phenylethanoids, phenylpropanoids, and phenolic acids quantification vis-à-vis gene expression profiling in leaf and root derived callus lines of Rhodiola imbricata (Edgew.)

Salidroside and rosavins have been found as the most potent ingredients used in Rhodiola based herbal formulations. Rhodiola imbricata is a medicinal herb of the trans-Himalayan Ladakh region of India. Since, the natural supply of this herb is rapidly decreasing, due to its over-exploitation, high altitude region, and traditional usage in the Amchi system of medicine. In the present study, friable callus culture was developed for the first time from leaf and root explants of R. imbricata. Furthermore, callus cell lines were also evaluated for rapid growth rate and high metabolite accumulation. The results revealed that callus induction frequency observed in juvenile leaves (100 %) and roots (87.50 %) in MS medium enriched with 0.5 mg/L TDZ and 1 mg/L NAA. Scanning electron microscopy (SEM) analysis was done for histological observations of the callus surface. A selective and efficient ultra-performance liquid chromatography with PDA detector (UPLC-PDA) method was developed and validated for the quantification of phenylethanoids, phenylpropanoids, and phenolic acids. The maximum salidroside was detected in leaf derived friable green calli (3.59 mg/g DW) subsequently followed by leaf derived friable white calli (2.31 mg/g DW). While rosavin and rosarin were detected maximum in root derived compact green calli (0.15 mg/g DW) and root derived friable green calli (0.07 mg/g DW). Genes encoding enzymes involved in salidroside and rosavins biosynthesis were also investigated for transcript abundance in wild as well as in vitro cultures using the RT-qPCR approach. The present study explained the friable callus culture as a potential alternative approach to obtain higher metabolite yield in R. imbricata. Further, it can be used for sequential scale-up of metabolite at bioreactor level to meet the industrial demand.

Stationary-phase optimized selectivity in supercritical fluid chromatography using a customized Phase OPtimized Liquid Chromatography kit: comparison of different prediction approaches.

The use of stationary-phase optimized selectivity in liquid chromatography (SOS-LC) was shown to be successful for HPLC to analyze complex mixtures using a Phase OPtimized Liquid Chromatography (POPLC) kit. This commercial kit contains five stationary-phase types of varying lengths, which can be coupled to offer an improved separation of compounds. Recently, stationary-phase optimized selectivity supercritical fluid chromatography (SOS-SFC) has been introduced, transferring the methodology to SFC. In this study, the applicability of a customized POPLC expert kit for isocratic SFC runs was explored. Five stationary-phase chemistries were selected as potentially most suitable for achiral separations of polar compounds: aminopropyl (amino), cyanopropyl (CN), diol, ethylpyridine (EP), and silica. The retention factors (k) on the individual stationary phases were used for the prediction of the best stationary-phase combination, based on the POPLC algorithm (via the included software). As an alternative, the best column combination was predicted using multiple linear regression (MLR) models on the results obtained from a simplex centroid mixture design with only three stationary-phase types (amino, silica, and EP). A third approach applied the isocratic POPLC algorithm on the same three stationary-phase data. The proposed combinations were assembled and tested. The predicted and experimental retention factors were compared. The predictions based on the POPLC algorithm provided a stationary phase showing a complete separation of the mixture. The stationary phase suggested by the MLR models, on the other hand, showed co-elution of two compounds, due to an unexpected experimental retention shift. Overall, the customized POPLC kit showed good potential to be applied in SFC. Graphical abstract.

A rapid and sensitive bioanalytical LC-MS/MS method for the quantitation of a novel CDK5 inhibitor 20-223 (CP668863) in plasma: Application to in vitro metabolism and plasma protein-binding studies.

A rapid, selective, and sensitive liquid chromatography coupled with tandem mass spectrometry (MS/MS) method was developed and validated for the quantitation of the novel CDK5 inhibitor '20-223' in mouse plasma. Separation of analytes was achieved by a reverse-phase ACE Excel C18 column (1.7 μm, 100 × 2.1 mm) with gradient elution using 0.1% formic acid (FA) in methanol and 0.1% FA as the mobile phase. Analytes were monitored by MS/MS with an electrospray ionization source in the positive multiple reaction monitoring mode. The MS/MS response was linear over the concentration range 0.2-500 ng/mL for 20-223. The within- and between-batch precision were within the acceptable limits as per Food and Drug Administration guidelines. The validated method was successfully applied to plasma protein binding and in vitro metabolism studies. Compound 20-223 was highly bound to mouse plasma proteins (>98% bound). Utilizing mouse S9 fractions, in vitro intrinsic clearance (CLint ) was 24.68 ± 0.99 μL/min/mg protein. A total of 12 phase I and II metabolites were identified with hydroxylation found to be the major metabolic pathway. The validate method required a low sample volume, was linear from 0.2 to 500 ng/mL, and had acceptable accuracy and precision.

Novel Reverse-Phase High-Performance Liquid Chromatography (RP-HPLC) Method Development and Validation of Atorvastatin and Fenofibrate in Tablets

A novel, simple, selective, precise, and accurate reverse-phase high-performance liquid chromatography (RP-HPLC) gradient method was developed for the simultaneous estimation of atorvastatin and fenofibrate in the combined formulation. The drugs atorvastatin calcium and fenofibrate were separated in the presence of their impurities atorvastatin related compound H, fenofibrate related compound A, and fenofibrate related compound B. The drugs and related compounds were separated on Kromasil C18 (250 x 4.6, 5μ) with reverse-phase gradient elution. Water adjusted pH 4.0 with phosphoric acid used as a buffer in pump A and acetonitrile used as a solvent in pump B as a mobile phase with gradient elution. The flow rate was 2.0 mL/min. 254 nm was the detection wavelength. The retention times were about 4.6 minutes for fenofibrate related compound A, 5.2 minutes for atorvastatin calcium, 5.7 minutes for fenofibrate related compound B, 8.7 minutes for atorvastatin related compound H, and 17.6 minutes for fenofibrate. The linearity ranges for atorvastatin calcium and fenofibrate were 5.00 to 15.00 and 80 to 240 mcg/mL, respectively, with correlation coefficient 0.999 for both. The proposed method validated statistically with respect to system suitability, specificity, linearity, precision, accuracy, range, robustness, and ruggedness. The method was accurate, linear, precise, specific, selective, and rapid suitable for the quantitative estimation of atorvastatin and fenofibrate in tablets.

Novel Isocratic RP-HPLC Method Development and Validation of Rosuvastatin and Fenofibrate in Tablets

A novel, simple, selective, precise, and accurate reverse-phase high-performance liquid chromatography (RP-HPLC) gradient method was developed for the simultaneous estimation of atorvastatin and fenofibrate in the combined formulation. The drugs atorvastatin calcium and fenofibrate were separated in the presence of their impurities atorvastatin related compound H, fenofibrate related compound A, and fenofibrate related compound B. The drugs and related compounds were separated on Kromasil C18 (250 x 4.6, 5μ) with reverse-phase gradient elution. Water adjusted pH 4.0 with phosphoric acid used as a buffer in pump A and acetonitrile used as a solvent in pump B as a mobile phase with gradient elution. The flow rate was 2.0 mL/min. 254 nm was the detection wavelength. The retention times were about 4.6 minutes for fenofibrate related compound A, 5.2 minutes for atorvastatin calcium, 5.7 minutes for fenofibrate related compound B, 8.7 minutes for atorvastatin related compound H, and 17.6 minutes for fenofibrate. The linearity ranges for atorvastatin calcium and fenofibrate were 5.00 to 15.00 and 80 to 240 mcg/mL, respectively, with correlation coefficient 0.999 for both. The proposed method validated statistically with respect to system suitability, specificity, linearity, precision, accuracy, range, robustness, and ruggedness. The method was accurate, linear, precise, specific, selective, and rapid suitable for the quantitative estimation of atorvastatin and fenofibrate in tablets.

Simultaneous Determination of Active Ingredients in Multicomponent Common Over the Counter Tablets in the Present of Parabens and 4-Aminophenol by HPLC

A new selective reverse-phase high-performance liquid chromatography method with an attitude of practical use in pharmaceutical laboratories for simultaneous determination of Ascorbic acid, Acetaminophen, phenylephrine, Caffeine, Pamabrom, Guaifenesin, Dextromethorphan, Pyrilamine, Chlorpheniramine maleate, Ibuprofen in the presence of Methyl, Ethyl, Butyl Parabens as preservatives has been developed and validated. These active pharmaceutical ingredients widely used in a variety of cough and cold medicines. The separation was carried out on an Atlantis dC18 column including 4.6 mm inner diameter, 150 mm length and particle size of 3 μm, using a mobile phase which consisted of an ion-pairing agent (sodium salt of heptane sulphonic acid) as a buffer solution mixed with acetonitrile as an organic modifier at a flow rate of 1 mL min−1 on a gradient system. The investigation was carried out on wavelength program detection to quantify the very low concentration of Dextromethorphan and Chlorpheniramine and high concentration of Acetaminophen, Guaifenesin, Ibuprofen, Ascorbic Acid at the wavelengths of 290 and 215 nm. The temperature was maintained at 40 °C. The method was shown to be linear (R2 > 0.996), accurate (recoveries 97.77–103.94%), precise (RSD ≤ 1.0%), sensitive, specific, simple, fast and robust. The proposed reverse phase-liquid chromatography method was successfully applied to the routine analysis of the Over-The-Counter tablets containing these active pharmaceutical ingredients without any interference with excipients and parabens and 4-aminophenol.

Pharmacokinetics of Picroside I, II, III, IV in Rat Plasma by UPLCMS/ MS

Background: Modern pharmacological studies show that rhizoma coptidis has protective effects on the liver, gallbladder, kidney, cerebral ischemia-reperfusion, local hypoxia injury, antiinflammatory, bone injury, nerve cells and myocardial cells. The effective components have been isolated from picroside I, II, III and IV. Introduction: A selective and sensitive ultra-performance liquid chromatography electrospray ionization tandem mass spectrometry (UPLC-ESI-MS/MS) method was developed for the simultaneous quantitative determination of picroside I, II, III and IV in rat plasma to aid the pharmacokinetics studies. Method: Sprague-Dawley (SD) rats were orally administered with 10 mg/kg, intravenously injected with 1 mg/kg for the mixture of picroside I, II, III and IV. The biological samples were collected at 0.083 3 h, 0.25 h, 1 h, 2 h, 4 h, 6 h, 8 h, 12 h, 24 h. A UPLC BEH C18 column (2.1 mm×50 mm, 1.7 μm) was used for chromatographic separation with the mobile phase consisting of acetonitrile and 0.1% formic acid by gradient elution. The flow rate was 0.4 mL/min. Multiple reaction monitoring (MRM) transitions were m/z 491.1→147.1 for picroside I, m/z 511.1→234.9 for picroside II, m/z 537.3→174.8 for picroside III and m/z 507.3→163.1 for picroside IV in negative ion mode. Result: The inter-day precision was less than 13%, the intra-day precision was less than 15%. The accuracy ranged from 89.4% to 111.1%. Recovery was higher than 79.1%, and the matrix effect ranged from 96.2% to 109.0%. Conclusion: The sensitive, rapid and selective UPLC-MS/MS method can be applied to the pharmacokinetic study of picroside I, II, III and IV in rats.

Liquid Chromatography Tandem Mass Spectrometry Based Label-Free Quantification Method for Assessment of Allergen-Induced Anaphylactoid Reactions.

Mast cells are essential in mediating inflammatory processes. When activated, mast cells can rapidly release characteristic granules and various mediators into the interstitium. Tryptase (TPS) and β-hexosaminidase (HEXB) are typical protease mediators stored in granules and released upon activation. They have been recognized as important biomarkers of anaphylaxis, and the released level is associated with the severity of allergic reactions. In this study, a sensitive, accurate, and selective liquid chromatography tandem mass spectrometry (LC-MS/MS) method for simultaneously quantifying the two biomarkers was developed and validated in LAD2 cell culture supernatant, and P14R was used as internal standard. Good linearity was observed in the range of 50-2500 ng/mL for TPS and 10-2000 ng/mL for HEXB both with R2 > 0.99. The matrix effect and recovery were both within acceptable limits. We quantified TPS and HEXB released from Laboratory of Allergic Disease 2 (LAD2) mast cells treated with several potential allergens, and the results demonstrate that the method can be used to investigate TPS and HEXB levels in LAD2 mast cell model during allergy research. We anticipate our approach to be a robust and sensitive assessment method for more biomarkers with similar kinetics characteristics and to be a major tool of allergic drug assessment or antiallergic drug development in research.

RP-HPLC method for quantitative estimation of Efinaconazole in topical microemulsion and microemulsion-based-gel formulations and in presence of its degradation products

The aim of current research work was to establish robust, sensitive, precise and selective reversed-phase high-performance liquid chromatography (RP-HPLC) method for the quantification of efinaconazole from microemulsion and microemulsion-gel based formulations for onychomycosis treatment. Chromatographic separation of efinaconazole was performed on Waters Inertsil® HPLC column, 5 µm (ODS-3V C18, 250 × 4.6 mm). The mobile phase was optimized as mixture of X and Y components in 20:80 v/v proportion (X component contains 0.01M phosphate buffer having pH 5.5 and acetonitrile in 75:25 v/v proportion and Y component contains acetonitrile) and the flow rate optimized at 1 mL min−1. Efinaconazole detection and quantification was done at a wavelength of 210 nm using a ultra-violet (UV) detector. Linear relationship was observed in method calibration curve for the concentration range of 50–150 µg/mL with r2 value >0.99. Different stress conditions experiments were carried out for the efinaconazole method development such as in acidic, basic, oxidation, photo, thermal, and humidity condition and no unknown degradation products interference were found during the estimation of efinaconazole. Further, the matrix effect of a microemulsion based excipients like diethylene glycol monoethyl ether, caprylcaproyl macrogol glycerides, and glyceryl caprylate has been overcame by present method of analysis with no excipient's interference. The method for analysis of efinaconazole was accurate, precise with average recovery rate of 98.6–101.6% and percentage RSD for all parameters of chromatographic system were found to be not more than 2.0%. Further, the method can be rapidly and effectively applied for the quantification of efinaconazole drug content from a microemulsion and microemulsion based gel formulations.Abbreviations: AUC, area under a curve; RP-HPLC, Reversed-phase high-performance liquid chromatography; ICH, International Council for Harmonisation; LOD, limit of detection; LOQ, limit of quantification; UV, ultra-violet; RSD, relative standard deviation; N, number of theoretical plates; Smix, surfactant mixture.

Parallel Reaction Monitoring-Based Quantification of Cannabinoids in Whole Blood.

Cannabis is the most consumed drug of abuse, making it the primary target for identification and quantification in human whole blood regarding forensic and clinical toxicology analyses. Among biological matrices, blood is the reference for toxicological interpretation. A highly sensitive and selective liquid chromatography (LC) hyphenated with high-resolution mass spectrometry (HRMS) was developed for the quantification of Δ9-tetrahydrocannabinol (THC), 11-hydroxytetrahydrocannabinol (THC-OH), 11-nor-9-carboxy-tetrahydrocannabinol (THC-COOH) and cannabidiol (CBD). Those cannabinoids were extracted from 1mL of whole blood by a simple liquid-liquid extraction (LLE) in acidic conditions. HRMS was performed on an Orbitrap-based instrument using its trapping capabilities and increased selectivity for parallel reaction monitoring (PRM) quantification in positive polarity with a negative polarity switching for THC-OH and THC-COOH. Although selected reaction monitoring (SRM) and PRM-targeted methods have similar performance in terms of linearity, dynamic range, precision and repeatability, Orbitrap-based PRM provides a higher specificity due to the use of high-resolution mode separating background ions from the targeted molecules. The method was fully validated according to guidelines set forth by the "Société Française des Sciences et des Techniques Pharmaceutiques" (SFSTP). Trueness was measured below 107% for all tested concentrations. Repeatability and intermediate precision were found to be lower than 12% while the assay was found to be linear in the concentration range of 0.4-20ng/mL for THC, THC-OH and CBD and of 2-100ng/mL for THC-COOH. Recovery (RE) and matrix effect (ME) ranged from 70.6 to 102.5% and from -40 to 6.6%, respectively. The validated method provides an efficient procedure for the simultaneous and rapid quantification of cannabinoids in PRM mode providing an alternative over classical SRM.

Mass Spectrometry-Based Rapid Quantitative Bioanalysis of Flibanserin: Pharmacokinetic and Brain Tissue Distribution Study in Female Rats.

Flibanserin (FLB) is the first United States Food and Drug Administration (USFDA) approved serotonin modulator recently marketed to treat acquired generalized women hypoactive sexual desire disorder. The scope of this study was to develop and validate a sensitive, selective and reliable ultra-performance liquid chromatography-mass spectroscopy/mass spectroscopy-based quantification method for FLB in rat plasma as well as brain tissue samples. The method includes a simple liquid-liquid sample extraction procedure. FLB was subjected to chromatographic separation using a poroshell C18 column with the mobile phase comprising a mixture of acetonitrile (ACN), 10mM ammonium acetate and acetic acid (90:10:0.1, v/v/v). Detection and quantification of FLB after positive electrospray ionization were carried out in selective ion monitoring mode. The fragment ions (m/z) of FLB (parent ion: 391.1741) and IS (parent ion: 448.1550) were monitored at 161.0704 and 285.0917, respectively. A linear response of FLB was observed over a concentration range of 2.5-600ng/mL in plasma and 5-500ng/mL in brain tissue homogenate. The intra- and inter-day precision and accuracy of the method met the acceptable limits specified in the USFDA bioanalytical method validation guideline. The analyte was found to be stable in benchtop, freeze-thaw, auto-injector and dry extract stability studies. The developed method was used to quantitate FLB in the plasma and brain tissue of a single-dose oral pharmacokinetic and brain tissue distribution study in female rats. Maximum FLB concentration in plasma and brain was achieved within an hour; however, the total amount of the drug that reached the brain was significantly less than in plasma. Rate of elimination of FLB from brain was also faster resulting in a lesser half-life in brain compared to the plasma.

The Effect of Grapefruit Juice on the Pharmacokinetics of Tadalafil in Rats.

We developed and validated a novel, sensitive, selective, and inexpensive high-performance liquid chromatography (HPLC) method for the determination of tadalafil in rats plasma and to investigate the effect of grapefruit juice on the pharmacokinetics of tadalafil in rats. The ZORBAX Eclipse XDB-C18 (4.6 × 150 mm, 5 μm) chromatography column can be used to separate tadalafil and carbamazepine (internal standard, IS). A mixture of acetonitrile-0.2% trifluoroacetic acid-water (48 : 10 : 42, V/V/V) was used as the mobile phase with a flow rate of 1.0 mL/min. The column temperature was set at 35.0°C. The detection wavelength was set at 286 nm. The tadalafil was extracted by ethyl acetate from plasma at the alkaline condition. 12 healthy male Sprague-Dawley (SD) rats were randomly divided into two groups, Group A (experimental group, received grapefruit juice 5 mL/kg for 7 days) and Group B (control group, received normal saline for 7 days). All the rats were given a single dose of tadalafil (5 mg/kg) after the last administration. The main pharmacokinetic parameters were calculated by DAS 2.0 software. Under the conditions of this experiment, the plasma concentrations of tadalafil in the range of 10–2000 ng/ml had a good linear relationship. The intra- and interday precision for tadalafil in plasma were less than 15%, and the relative recovery rate was good at low, medium, and high QC levels. The Cmax of tadalafil in the control group and the experimental group was (725.89 ± 161.59) ng/mL and (1271.60 ± 179.31) ng/mL, t1/2 was (9.28 ± 2.07) h and (11.70 ± 1.47) h, AUC (0-t) was (7399.61 ± 696.85) ng·h/mL and (9586.52 ± 2048.81) ng·h/mL, and AUC(0-∞) was (7995.50 ± 707.23) ng·h/mL and (10639.43 ± 2235.94) ng·h/mL, respectively. Results show that the Cmax of tadalafil in group A was 75.17% higher than that in group B, the Vz/F was also reduced, and the t1/2 was increased by 2.42 h. The developed HPLC–DAD method for the determination of tadalafil in rats plasma was accurate, reproducible, specific, and it was found to be suitable for the pharmacokinetics of tadalafil and food-drug interactions. Grapefruit juice can inhibit the metabolism of tadalafil and increase the exposure of tadalafil in rats.

Development and validation of highly selective method for the determination of imatinib mesylate and dexketoprofen trometamol combination in three different media

Imatinib mesylate is a small molecule used in cancer therapy as a thyrosine kinase inhibitor. Dexketoprofen trometamol is a non-steroidal anti-inflammatory drug that has seen use in cancer therapy in combination with an anticancer drug to minimize tumor size and to reduce pain in patients. In the present study, imatinib mesylate and dexketoprofen trometamol were selected as potential model drugs to be used in combination. A new, simple and selective Ultra Performance Liquid Chromatography method was developed and validated to determine the drug substances in distilled water, in a pH 7.4 phosphate buffer and in Dulbecco’s Modified Eagle Medium. The proposed method was developed using a BEH C-18 column with isocratic elution. A mixture of methanol:acetonitrile (80:20, v/v) and pH 9.5, 0.05 M ammonium acetate were (70:30, v/v) used as a mobile phase. Detection was carried out with a flow rate of 0.3 mL/min, a column temperature of 30°C and an injection volume of 20 µL. The method was validated considering linearity, accuracy, precision, specificity, robustness, detection limit and quantitation limit values, and was found to be linear in a range from 0.05 to 20.0 µg/mL for the three different media.

Determination of licochalcone A in rat plasma by UPLC–MS/MS and its pharmacokinetics

The aim of this study was to establish a rapid, sensitive, and selective ultra-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) method to quantify the concentrations of licochalcone A and applicate the technique to its pharmacokinetic study. Analytes were separated on an UPLC ethylene bridged hybrid (BEH) C18 column (2.1 mm × 50 mm, 1.7 μm). The mobile phase was consisted of acetontrile and 0.1% formic acid with a flow rate of 0.4 mL/min in a gradient elution mode. Multiple-reaction monitoring (MRM) was carried out in a negative mode for licochalcone A (m/z 337.2 → 119.7) and the internal standard (IS) (m/z 609.0 → 300.9). The linearity of licochalcone A was great from 0.53 to 530 ng/mL. The lower limit of quantification and the lower limit of detection were 0.53 ng/mL and 0.26 ng/mL, respectively. The intra-day precision was less than 14%, and the inter-day precision was no more than 11%. The accuracy was from 91.5% to 113.9%, the recovery was over 90.5%, and the matrix effect was between 84.5% and 89.7%. The results of stability were in an acceptable range. The bioavailability was only 3.3%, exhibiting poor absorption. The developed method was successfully applicable for determining the concentrations of licochalcone A and its pharmacokinetic study.

Pharmacokinetics of 13 active components in a rat model of middle cerebral artery occlusion after intravenous injection of Radix Salviae miltiorrhizae-Lignum dalbergiae odoriferae prescription.

As a representative formulation of Radix Salviae miltiorrhizae (Danshen)-Lignum Dalbergiae odoriferae (Jiangxiang), Xiangdan injection is widely prescribed for cardio- and cerebrovascular diseases in practice. This necessitates a pharmacokinetic investigation of this formulation to make it safer and more broadly applicable. We developed and validated a sensitive, selective, and reliable high-performance liquid chromatography with tandem mass spectrometry method for the simultaneous determination of 11 phenolic compounds including danshensu plus two diterpenoid quinones like cryptotanshinone and tanshinone IIA in rat. We applied this method for the pharmacokinetic studies of the 13 compounds in a rat model of middle cerebral artery occlusion after intravenous injection of Xiangdan injection or Danshen injection. In sham-operated rats, the animals taking Xiangdan injection exhibited significant growth of the area under the curve for danshensu, protocatechuic aldehyde, and tanshinone IIA compared with the changes seen in the data of those administrated with Danshen injection. Such a pattern was also observed in middle cerebral artery occlusion rats, whereas increased the area under the curve values were observed for danshensu, protocatechuic aldehyde, caffeic acid, rosmarinic acid, and tanshinone IIA. These results demonstrated that synergistic interactions occurred between the components of Danshen and the active compounds of Jiangxiang both in sham-operated and middle cerebral artery occlusion rats, increasing the bioavailability of Danshen. The results presented herein can be used to determine a reference dose for the clinical application of Xiangdan injection, and to elucidate the synergistic mechanism of Danshen and Jiangxiang.