Ultrasound‐assisted emulsification microextraction combined with injection‐port derivatization for the determination of some chlorophenoxyacetic acids in water samples

An efficient method based on ultrasound‐assisted emulsification microextraction followed by injection‐port derivatization GC analysis was developed to determine 2,4‐dichlorophenoxyacetic acid (2,4‐D) and 4‐chloro‐2‐methylphenoxyacetic acid (MCPA) in natural water samples. In this procedure, 12.5 μL of 1‐undecanol was injected slowly into a 12 mL home‐designed centrifuge glass vial containing an aqueous sample of the analytes located inside an ultrasonic water bath. The resulting emulsion was centrifuged, and 1 μL of the separated organic solvent together with 1 μL of the derivatization reagent were injected into a GC equipped with a flame ionization detector. Several factors that influence the derivatization and extraction were optimized. Under the optimal conditions, the LODs were 0.33 and 1.7 μg/L for MCPA and 2,4‐D, respectively. Preconcentration factors of 670 and 836 were obtained for MCPA and 2,4‐D, respectively. The precision of the proposed method was evaluated in terms of repeatability, which was <5.7% (n = 5). The applicability of the proposed method was evaluated by extraction and determination of chlorophenoxyacetic acids from some natural waters, which indicated that the matrices of natural waters have no significant effect on the extraction and derivatization efficiency of this method.     

Development of single‐drop microextraction and simultaneous derivatization followed by GC‐MS for the determination of aliphatic amines in tobacco

In this work, for the first time, headspace (HS) single‐drop microextraction and simultaneous derivatization followed by GC‐MS was developed to determine the aliphatic amines in tobacco samples. In the HS extraction procedure, the mixture of derivatization reagent and organic solvent was employed as the extraction solvent for HS single‐drop microextraction and in situ derivatization of aliphatic amine in the samples. Fast extraction and simultaneous derivatization of the analytes were performed in a single step, and the obtained derivatives in the microdrop extraction solvent were analyzed by GC‐MS. The optimized experiment conditions were: sample preparation temperature of 80°C and time of 30 min, HS extraction solvent (the mixture of benzyl alcohol and 2,3,4,5,6‐pentafluorobenzaldehyde) volume of 2.0 μL, extraction time of 90 s. With the optimal conditions, the method validations were also studied. The method has good linearity (R² more than 0.99), accepted precision (RSD less than 13%), good recovery (98–104%) and low limit of detection (0.11–0.97 μg/g). Finally, the proposed technique was successfully applied to the analyses of aliphatic amines in tobacco samples of seven different brands. It was further demonstrated that the proposed method offered a simple, low‐cost and reliable approach to determine aliphatic amines in tobacco samples.     

Trace Analysis of Methyl Tert‐butyl Ether in Water Samples Using New Ultrasound‐assisted Dispersive Liquid‐liquid Microextraction and Gas Chromatography‐flame Ionization Detection

In this study, simple and efficient ultrasound‐assisted dispersive liquid‐liquid microextraction combined with gas chromatography (GC) was developed for the preconcentration and determination of methyl‐tert‐butyl ether (MTBE) in water samples. One hundred microliters of benzyl alcohol was injected slowly into 10 mL home‐designed centrifuge glass vial containing an aqueous sample with 30% (w/v) of NaCl that was located inside the ultrasonic water bath. The formed emulsion was centrifuged and 2 μL of separated benzyl alcohol was injected into a gas chromatographic system equipped with a flame ionization detector (GC‐FID) for analysis. Several factors influencing the extraction efficiency such as the nature and volume of organic solvent, extraction temperature, ionic strength and centrifugation times were investigated and optimized. Using optimum extraction conditions a detection limit of 0.05 μg L^‐1 and a good linearity (r² = 0.998) in a calibration range of 0.1‐500 μg L^‐1 were achieved. This proposed method was successfully applied to the analysis of MTBE in tap, well and a ground water sam ple contaminated by leaking gasoline from an underground storage tank (LUST) in a gasoline service station.     

Rapid determination of bisphenol A in drinking water using dispersive liquid‐phase microextraction with in situ derivatization prior to GC‐MS

This paper described a novel approach for the determination of bisphenol A by dispersive liquid‐phase microextraction with in situ acetylation prior to GC‐MS. In this derivatization/extraction method, 500 μL acetone (disperser solvent) containing 30.0 μL chlorobenzene (extraction solvent) and 30.0 μL acetic anhydride (derivatization reagent) was rapidly injected into 5.00 mL aqueous sample containing bisphenol A and K₂CO₃ (0.5% w/v). Within a few seconds the analyte was derivatized and extracted at the same time. After centrifugation, 1.0 μL of sedimented phase containing enriched analyte was determined by GC‐MS. Some important parameters, such as type and volume of extraction and disperser solvent, volume of acetic anhydride, derivatization and extraction time, amount of K₂CO₃, and salt addition were studied and optimized. Under the optimum conditions, the LOD and the LOQ were 0.01, 0.1 μg/L, respectively. The experimental results indicated that there was linearity over the range 0.1–50 μg/L with coefficient of correlation 0.9997, and good reproducibility with RSD 3.8% (n = 5). The proposed method has been applied for the analysis of drinking water samples, and satisfactory results were achieved.     

Solvent‐assisted dispersive micro‐SPE by using aminopropyl‐functionalized magnetite nanoparticle followed by GC‐PID for quantification of parabens in aqueous matrices

In this research, solvent‐assisted dispersive micro‐SPE was introduced as a simple modified technique for the determination of parabens in water and cosmetic samples. Aminopropyl‐functionalized magnetite nanoparticles (MNPs) were successfully synthesized and applied. GC with photoionization detector was used for the separation and detection of parabens. In this method, hexylacetate (15 μL) as a solvent and aminopropyl‐functionalized MNPs (5 μg) as a sorbent were added to an aqueous sample (10 mL) and then the sample was sonicated. Dispersed magnetite was collected in the bottom of the conical tube by using a strong magnet and then ACN was added as a desorption solvent. Forty microliters of this solvent was transferred into a microvial and then acetic anhydride and pyridine were added, thus derivatization was performed by acetic anhydride. After evaporation, 1 μL of derivatized sample was injected into a gas chromatograph for analysis. Several important parameters, such as kind of organic solvent, desorption solvent and volume, amount of aminopropyl‐functionalized MNPs and effect of salt addition were investigated. Under optimum conditions, the limits of detection achieved were between 50 and 300 ng/L, with RSDs (n = 5) lower than 8%. Under the optimum conditions, the enrichment factors ranged from 217 to 1253 and the extraction recoveries ranged from 10 to 62%. The recoveries were obtained for the analytes in river water and mouthwash solution and hand cream in the range of 87–103%. The advantages of proposed method are simplicity of operation, rapidity, high extraction yields, and environmental friendly character.     

Determination of haloperidol in biological samples with the aid of ultrasound‐assisted emulsification microextraction followed by HPLC‐DAD

A rapid and simple quantitative method for preconcentration and determination of haloperidol in biological samples was developed using ultrasound‐assisted emulsification microextraction, based on the solidification of floating organic droplet combined with HPLC‐DAD. The effects of several factors were investigated. A total of 30 μL of 1‐undecanol as an extraction solvent was injected slowly into a glass‐centrifuge tube containing 4 mL alkaline sample solution that was located inside the ultrasonic water bath. The formed emulsion was centrifuged and the fine droplets of solvent were floated at the top of the test tube, then it was cooled in an ice bath and the solidified solvent was transferred into a conical vial, after melt, the analysis of the extract was carried out by HPLC. Under the optimal conditions, the extraction efficiencies were more than 90% and the preconcentration factors were obtained between 119–122. The LOQs were obtained between 4–8 μg/L and the calibration curves were linear within the range of 4–1000 μg/L. Finally this method was applied to the determination of haloperidol in plasma and urine samples in the range of μg/L and satisfactory results were achieved (RSDs <7%).     

Determination of 3,5,6‐trichloro‐2‐pyridinol,phoxim and chlorpyrifos‐methyl in water samples using a new pretreatment method coupled with high‐performance liquid chromatography

A novel low‐density solvent‐based vortex‐assisted surfactant‐enhanced‐emulsification liquid–liquid microextraction with the solidification of floating organic droplet method coupled with high‐performance liquid chromatography was developed for the determination of 3,5,6‐trichloro‐2‐pyridinol, phoxim and chlorpyrifos‐methyl in water samples. In this method, the addition of a surfactant could enhance the speed of the mass transfer from the sample solution into the extraction solvent. The extraction solvent could be dispersed into the aqueous by the vortex process. The main parameters affecting the extraction efficiency were investigated and the optimum conditions were established as follows: 80 μL 1‐undecanol as extraction solvent, 0.2 mmol/L of Triton X‐114 selected as the surfactant, the vortex time was fixed at 60 s with the vortex agitator set at 3000 rpm, the concentration of acetic acid in sample solution was 0.4% v/v and 1.0 g addition of NaCl. Under the optimum conditions, the enrichment factors were from 172 to 186 for the three analytes. The linear ranges were from 0.5 to 500 μg/L with a coefficient of determination (r²) of between 0.9991 and 0.9995. Limits of detections were varied between 0.05 and 0.12 μg/L. The relative standard deviations (n = 6) ranged from 0.26 to 2.62%.     

Simply enhancing throughput of free‐flow electrophoresis via organic‐aqueous environment for purification of weak polarity solute of phenazine‐1‐carboxylic acid in fermentation of Pseudomonas sp. M18

Herein, a simple novel free‐flow electrophoresis (FFE) method was developed via introduction of organic solvent into the electrolyte system, increasing the solute solubility and throughput of the sample. As a proof of concept, phenazine‐1‐carboxylic acid (PCA) from Pseudomonas sp. M18 was selected as a model solute for the demonstration on feasibility of novel FFE method on account of its faint solubility in aqueous circumstance. In the developed method, the organic solvent was added into not only the sample buffer to improve the solubility of the solute, but also the background buffer to construct a uniform aqueous‐organic circumstance. These factors of organic solvent percentage and types as well as pH value of background buffer were investigated for the purification of PCA in the FFE device via CE. The experiments revealed that the percentage and the types of organic solvent exerted major influence on the purification of PCA. Under the optimized conditions (30 mM phosphate buffer in 60:40 (v/v) water‐methanol at an apparent pH 7.0, 3.26 mL/min background flux, 10‐min residence time of injected sample, and 400 V), PCA could be continuously purified from its impurities. The flux of sample injection was 10.05 μL/min, and the recovery was up to 93.7%. An 11.9‐fold improvement of throughput was found with a carrier buffer containing 40% (v/v) methanol, compared with the pure aqueous phase. The developed procedure is of evident significance for the purification of weak polarity solute via FFE.     

Acid‐induced homogenous liquid‐phase microextraction: Application of medium‐chain carboxylic acid as extraction phase

In this research, a novel homogeneous liquid‐phase microextraction method was successfully developed based on applying octanoic acid as low‐density extraction solvent. The method was applied for extraction and determination of chlorophenols (CPs) as model compounds. Twelve milliliter of the sample solution was poured into a home‐designed glass vial. Sixty microliter of octanoic acid was solved in water sample by adjusting pH and ionic strength. By rapid addition of 75 μL of concentrated HCl (6 M), a cloudy solution was obtained. Phase separation occurred at 5000 rpm for 5 min. After that, 20 μL of the collected phase (approximately 26 μL) was injected into the HPLC‐UV instrument for analysis. The effect of some parameters such as the volume of concentrated HCl (phase separation reagent), ionic strength, extraction time, centrifugation time, and the volume of extracting phase on the extraction efficiency of the CPs were investigated and optimized. The preconcentration factors in a range of 159–218 were obtained under the optimal conditions. The linear range, detection limits (S/N = 3), and precision (n = 3) were 1– 200, 0.3–0.5 μg/L, and 4.6–5.1%, respectively. Tap water, seawater, and river water samples were successfully analyzed for the existence of CPs using the proposed method and satisfactory results were obtained.     

Comparison of solidification of floating drop and homogenous liquid–liquid microextractions for the extraction of two plasticizers from the water kept in PET‐bottles

Two approaches based on solidification of floating drop microextraction (SFDME) and homogenous liquid–liquid microextraction (HLLE) were compared for the extraction and preconcentration of di‐(2‐ethylhexyl) phthalate (DEHP) and di‐(2‐ethylhexyl) adipate (DEHA) from the mineral water samples. In SFDME, a floated drop of the mixture of acetophenone/1‐undecanol (1:8) was exposed on the surface of the aqueous solution and extraction was permitted to occur. In HLLE, a homogenous ternary solvent system was used by water/methanol/chloroform and the phase separation phenomenon occurred by salt addition. Under the optimal conditions, the LODs for the two target plasticizers (DEHA and DEHP), obtained by SFDME–GC‐FID and HLLE–GC‐FID, were ranged from 0.03 to 0.01 μg/L and 0.02 to 0.01 μg/L, respectively. HLLE provided higher preconcentration factors (472.5‐ and 551.2‐fold) within the shorter extraction time as well as better RSDs (4.5–6.9%). While, in SFDME, high preconcentration factors in the range of 162–198 and good RSDs in the range of 5.2–9.6% were obtained. Both methods were applied for the analysis of two plasticizers in different water samples and two target plasticizers were found in the bottled mineral water after the expiring time and the boiling water was exposed to a polyethylene vial.     

Low‐density solvent‐based vortex‐assisted surfactant‐enhanced emulsification liquid–liquid microextraction and its application

For the first time, the low‐density solvent‐based vortex‐assisted surfactant‐enhanced emulsification liquid–liquid microextraction, followed by GC‐flame photometric detection has been developed for the determination of eight organophosphorus pesticides in aqueous samples. A small volume of organic extraction solvent (toluene) was dispersed into the aqueous samples by the assistance of surfactant and vortex agitator. The extraction was performed in a special disposable polyethylene pipette, allowing using the reagents with lower density than water as extraction solvents. The influence parameters were systemically investigated and optimized: toluene (30 μL) and Triton X‐100 (0.2 mmol/L) were used as the extraction solvent and the surfactant, respectively, and the extraction was performed for 1 min under room temperature without adding sodium chloride. Under the optimum conditions, the validation parameters such as the RSD (n = 6; 2.1–11.3%), LOD (0.005 and 0.05 μg/L), and linear range (0.1–50.0 μg/L with correlation coefficients (0.9958–0.9992) showed the method was satisfying. The proposed method has been successfully applied to the determination of the organophosphorus pesticides in real samples with recoveries between 82.8 and 100.2%.     

Application of continual injection liquid‐phase microextraction method coupled with liquid chromatography to the analysis of organophosphorus pesticides

A liquid‐phase microextraction coupled with LC method has been developed for the determination of organophosphorus pesticides (methidation, quinalphos and profenofos) in drinking water samples. In this method, a small amount (3 μL) of isooctane as the acceptor phase was introduced continually to fill‐up the channel of a 1.5 cm polypropylene hollow fiber using a microsyringe while the hollow fiber was immersed in an aqueous donor solution. A portion of the acceptor phase (ca. 0.4 μL) was first introduced into the hollow fiber and additional amounts (ca. 0.2 μL) of the acceptor phase were introduced to replenish at intervals of 3 min until set end of extraction (40 min). After extraction, the acceptor phase was withdrawn and transferred into a 2 mL vial for a drying step prior to injection into a LC system. Parameters that affect the extraction efficiency were studied including the organic solvent, length of fiber, volume of acceptor and donor phase, stirring rate, extraction time, and effect of salting out. The proposed method provided good enrichment factors of up to 189.50, with RSD ranging from 0.10 to 0.29%, analyte recoveries of over 79.80% and good linearity ranging from 10.0 to 1.25 mg/L. The LOD ranged from 2.86 to 82.66 μg/L. This method was applied successfully to the determination of organophosphorus pesticides in selected drinking water samples.     

Ionic‐liquid‐assisted microwave distillation coupled with headspace single‐drop microextraction followed by GC–MS for the rapid analysis of essential oil in Dryopteris fragrans

A rapid, green and effective miniaturized sample preparation technique, ionic‐liquid‐assisted microwave distillation coupled with headspace single‐drop microextraction was developed for the extraction of essential oil from dried Dryopteris fragrans. 1‐Ethyl‐3‐methylimidazolium acetate was the optimal ionic liquid as the destruction agent of plant cell walls and microwave absorption was medium. n‐Heptadecane (2.0 μL) was adopted as the suspended microdrop solvent in the headspace for the extraction and concentration of essential oil. The optimal parameters of the proposed method were an irradiation power of 300 W, sample mass of 0.9 g, mass ratio of ionic liquids to sample of 2.8, extraction temperature of 79°C, and extraction time of 3.6 min. In comparison to the previous reports, the proposed technique could equally monitor all the essential oil components with no significant differences in a simple way, which was more rapid and required a much lower amount of sample.     

Headspace solvent microextraction and gas chromatographic determination of some polycyclic aromatic hydrocarbons in water samples

Headspace solvent microextraction (HSME) was shown to be an efficient preconcentration method for extraction of some polycyclic aromatic hydrocarbons (PAHs) from aqueous sample solutions. A microdrop of 1-butanol (as extracting solvent) containing biphenyl (as internal standard) was used in this investigation. Extraction occurred by suspending a 3 μl drop of 1-butanol from the tip of a microsyringe fixed above the surface of solution in a sealed vial. After extraction for a preset time, the microdrop was retracted back into the syringe and injected directly into a GC injection port. The effects of nature of extracting solvent, microdrop and sample temperatures, stirring rate, microdrop and sample volumes, ionic strength and extraction time on HSME efficiency were investigated and optimized. Finally, the enrichment factor, dynamic linear range (DLR), limit of detection (LOD) and precision of the method were evaluated by water samples spiked with PAHs. The optimized procedure was successfully applied to the extraction and determination of PAHs in different water samples.     

Single‐drop microextraction followed by in‐syringe derivatization and GC‐MS detection for the determination of parabens in water and cosmetic products

A single‐drop microextraction (SDME) method followed by in‐syringe derivatization and GC‐MS determination has been developed for analysis of five parabens, including methyl, ethyl, isopropyl, n‐propyl and n‐butyl paraben in water samples and cosmetic products. N,O‐Bis(trimethylsilyl)acetamide (BSA) was used as derivatization reagent. Derivatization reaction was performed inside the syringe barrel using 0.4 μL of BSA. Parameters that affect the derivatization yield such as temperature and time of the reaction were studied. In addition, experimental SDME parameters such as selection of organic solvent, addition of salt, extraction time and extraction temperature were investigated and optimized. The RSD of the method for aqueous samples varied from 8.1 to 13%. The LODs ranged from 0.001 (n‐butyl paraben) to 0.015 (methyl paraben) μg/L, and the enrichment factors were between 23 and 150.     

Determination of phthalate esters in liquor samples by vortex‐assisted surfactant‐enhanced‐emulsification liquid–liquid microextraction followed by GC–MS

A novel method using vortex‐assisted surfactant‐enhanced‐emulsification liquid–liquid microextraction has been developed for the extraction of phthalate esters (PAEs) in Chinese liquor samples prior to analysis by GC–MS. In the proposed method, a high‐density extraction solvent (carbon tetrachloride) was dispersed into samples with the aid of a surfactant (Triton X‐100) and vortex agitation, resulting in a short extraction equilibrium (30 s). After centrifugation, a single microdrop of solvent was easily collected for GC–MS analysis. Key factors that affected the extraction efficiency were optimized. Under the optimum conditions, linearity was found in the range from 0.05 to 50 μg/L. Coefficients of determination varied from 0.9938 to 0.9971. LODs, based on an S/N of 3, ranged from 4.9 to 13 ng/L. Enrichment factors varied from 140 to 184. Reproducibility and recoveries were assessed by testing a series of three liquor samples that were spiked with different concentration levels. Finally, the proposed method was successfully applied to the determination of PAEs in 16 Chinese liquor samples. In this work, high‐density‐solvent vortex‐assisted surfactant‐enhanced‐emulsification liquid–liquid microextraction was applied for the first time for the extraction of PAEs in Chinese liquor samples and was proved to be simple, rapid, and sensitive.     

Preconcentration Ultra Trace of Cd(II) in Water Samples Using Dispersive Liquid‐Liquid Microextraction with Salen(N,N′‐Bis(Salicylidene)‐Ethylenediamine) and Determination Graphite Furnace Atomic Absorption Spectrometry

Dispersive liquid–liquid microextraction (DLLME) technique was successfully used as a sample preparation method for graphite furnace atomic absorption spectrometry (GF AAS). In this extraction method, 500 μL methanol (disperser solvent) containing 34 μL carbon tetrachloride (extraction solvent) and 0.00010 g Salen(N,N′‐bis(salicylidene)ethylenediamine) (chelating agent) was rapidly injected by syringe into the water sample containing cadmium ions (interest analyte). Thereby, a cloudy solution formed. The cloudy state resulted from the formation of fine droplets of carbon tetrachloride, which have been dispersed, in bulk aqueous sample. At this stage, cadmium reacts with Salen(N,N′‐bis(salicylidene)‐ethylenediamine), and therefore, hydrophobic complex forms which is extracted into the fine droplets of carbon tetrachloride. After centrifugation (2 min at 5000 rpm), these droplets were sedimented at the bottom of the conical test tube (25 ± 1 μL). Then a 20 μL of sedimented phase containing enriched analyte was determined by GF AAS. Some effective parameters on extraction and complex formation, such as extraction and disperser solvent type and their volume, extraction time, salt effect, pH and concentration of the chelating agent have been optimized. Under the optimum conditions, the enrichment factor 122 was obtained from only 5.00 mL of water sample. The calibration graph was linear in the range of 2‐21 ng L^?1 with a detection limit of 0.5 ng L^?1. The relative standard deviation (R.S.D._s) for ten replicate measurements of 20 ng L^?1 of cadmium was 2.9%. The relative recoveries of cadmium in tap, sea and rain water samples at a spiking level of 5 and 10 ng L^?1 are 99, 94, 97 and 96%, respectively. The characteristics of the proposed method have been compared with cloud point extraction (CPE), on‐line liquid‐liquid extraction, single drop microextraction (SDME), on‐line solid phase extraction (SPE) and co‐precipitation based on bibliographic data. Therefore, DLLME combined with GF AAS is a very simple, rapid and sensitive method, which requires low volume of sample (5.00 mL).     

Multivariate optimization of surfactant‐assisted directly suspended droplet microextraction combined with GC for the preconcentration and determination of tramadol in biological samples

In this work, a novel procedure based on surfactant‐assisted directly suspended droplet microextraction for the determination of tramadol prior to GC with flame ionization detection is proposed. In this technique, a free microdroplet of solvent is transferred to the surface of an immiscible aqueous sample containing Triton X‐100 and tramadol while being agitated by a stirring bar placed on the bottom of the sample vial. After the predetermined time, the microdroplet of solvent is withdrawn by a syringe and analyzed. The effective parameters such as the type of organic solvent, extraction time, microdroplet volume, salt content of the donor phase, stirring speed, the source phase pH, concentration of Triton X‐100, and extraction temperature were optimized. For this purpose, a multivariate strategy was applied based on an experimental design in order to screen and optimize the significant factors. This method requires minimal sample preparation, analysis time, solvent consumption, and represents significant advantages over customary analytical methods. The linearity ranged from 10 to 2000 μg/L with RSDs (n = 5) of 7.3–10. Preconcentration factors and the LODs were 391–466 and 2.5–6.5 μg/L, respectively. Finally, this method was applied to the analysis of biological samples and satisfactory results were obtained.     

Headspace Single Drop Microextraction for the Analysis of Fire Accelerants in Fire Debris Samples

Fire accelerants such as gasoline, kerosene, and diesel have commonly been used in arson cases. Improved analytical methods involving the extraction of fire accelerants are necessary to increase sample yield and to reduce the number of uncertain findings. In this study, an analytical method based on headspace single drop microextraction (HS-SDME) followed by gas chromatography–flame ionization detection (GC-FID) has been developed for the analysis of simulated fire debris samples. Curtain fabric was used as the sample matrix. The optimized conditions were 2.5 μL benzyl alcohol microdrop exposed for 20 min to the headspace of a 10 mL aqueous sample containing accelerants placed in 15-mL sample vial and stirred at 1500 rpm. The extraction method was compared with the solvent extraction method using n-hexane for the determination of fire accelerants. The HS-SDME process is driven by the concentration difference of analytes between the aqueous phases containing the analyte and the organic phase constituting the microdrop of a solvent. The limit of detection of HS-SDME for kerosene was 1.5 μL. Overall, the HS-SDME coupled with GC-FID proved to be rapid, simple and sensitive and a good alternative method for the analysis of accelerants in fire debris samples.     

Quantitation and pharmacokinetics of 1,4‐diamino‐2,3‐dicyano‐1,4‐bis (2‐aminophenylthio) butadiene (U0126) in rat plasma by liquid chromatography‐tandem mass spectrometry

A simple, robust, and rapid LC‐MS/MS method was developed for the quantitation of U0126 and validated in rat plasma. Plasma samples (20 μL) were deproteinized using 200 μL ACN containing 30 ng/mL of chlorpropamide, internal standard. Chromatographic separation performed on an Agilent Poroshell 120 EC‐C₁₈ column (4.6 × 50 mm, 2.7 μm particle size) with an isocratic mobile phase consisting of a 70:30 v/v mixture of ACN and 0.1% aqueous formic acid. Each sample was run at 0.6 mL/min for a total run time of 2 min per sample. Detection and quantification were performed using a mass spectrometer in selected reaction‐monitoring mode with positive ESI at m/z 381 → 123.9 for U0126 and m/z 277 → 175 for the internal standard. The standard curve was linear over a concentration range of 20–5000 ng/mL with correlation coefficients greater than 0.9965. Precision, both intra‐ and interday, was less than 10.1% with an accuracy of 90.7–99.4%. No matrix effects were observed. U0126 in rat plasma degraded approximately 41.3% after 3‐h storage at room temperature. To prevent degradation, sample handling should be on an ice bath and all solutions kept at 4°C. This method was successfully applied to a pharmacokinetic study of U0126 at various doses in rats.