Preparative and scaled‐up separation of high‐purity α‐linolenic acid from perilla seed oil by conventional and pH‐zone refining counter current chromatography

α‐Linolenic acid is an essential omega‐3 fatty acid needed for human health. However, the isolation of high‐purity α‐linolenic acid from plant resources is challenging. The preparative separation methods of α‐linolenic acid by both conventional and pH‐zone refining counter current chromatography were firstly established in this work. The successful separation of α‐linolenic acid by conventional counter current chromatography was achieved by the optimized solvent system n‐heptane/methanol/ water/acetic acid (10:9:1:0.04, v/v), producing 466 mg of 98.98% α‐linolenic acid from 900 mg free fatty acid sample prepared from perilla seed oil with linoleic acid and oleic acid as by‐products. The scaled‐up separation in 45× is efficient without loss of resolution and extension of separation time. The separation of α‐linolenic acid by pH‐zone refining counter current chromatography was also satisfactory by the solvent system n‐hexane/methanol/water (10:5:5, v/v) and the optimized concentration of trifluoroacetic acid 30 mM and NH₄OH 10 mM. The separation can be scaled up in 180× producing 9676.7 mg of 92.79% α‐linolenic acid from 18 000 mg free fatty acid sample. pH‐zone refining counter current chromatography exhibits a great advantage over conventional counter current chromatography with 20× sample loading capacity on the same column.     

A rapid and efficient extraction method based on industrial MCM‐41‐miniaturized matrix solid‐phase dispersion extraction with response surface methodology for simultaneous quantification of six flavonoids in Pollen typhae by ultra‐high‐performance liquid chromatography

An industrial MCM‐41‐miniaturized matrix solid‐phase dispersion extraction coupled with response surface methodology was explored to determine L‐epicatechin, typhaneoside, isorhamnetin‐3‐O‐neohespeidoside, naringenin, kaempferol, and isorhamnetin in Pollen typhae by ultra‐high performance liquid chromatography connected to a photodiode array detection. Several variables were optimized in detail, including mesh number of sieve, type of adsorbent, mass ratio of sample to adsorbent, grinding time, methanol concentration, and elution volume. Central composite design was applied to optimize the best conditions for the maximum yields of the total flavonoids. The results displayed a good linear relationship (R > 0.9992) and the recoveries ranged from 92.9 to 103% (RSD < 4.53%) of the six flavonoids. The optimal method with high efficiency and low consumption was obviously better than heating reflux and ultrasonic extraction. It was proven that the developed industrial MCM‐41‐miniaturized matrix solid‐phase dispersion extraction coupled with simple ultra‐high performance liquid chromatography method could be a rapid and efficient tool for extraction and determination of flavonoids in natural products.     

Ionic‐liquid‐based ultrasound‐assisted extraction of isoflavones from Belamcanda chinensis and subsequent screening and isolation of potential α‐glucosidase inhibitors by ultrafiltration and semipreparative high‐performance liquid chromatography

The separation of a compound of interest from its structurally similar homologues to produce high‐purity natural products is a challenging problem. This work proposes a novel method for the separation of iristectorigenin A from its structurally similar homologues by ionic‐liquid‐based ultrasound‐assisted extraction and the subsequent screening and isolation of potential α‐glucosidase inhibitors via ultrafiltration and semipreparative high‐performance liquid chromatography. Ionic‐liquid‐based ultrasound‐assisted extraction was successfully applied to the extraction of tectorigenin, iristectorigenin A, irigenin, and irisflorentin from Belamcanda chinensis . The optimum conditions for the efficient extraction of isoflavones were determined as 1.0 M 1‐ethyl‐3‐methylimidazolium tetrafluoroborate with extraction time of 30 min and a solvent to solid ratio of 30 mL/g. Ultrafiltration with liquid chromatography and mass spectrometry was applied to screen and identify α‐glucosidase inhibitors from B. chinensis , followed by the application of semipreparative high‐performance liquid chromatography to separate and isolate the active constituents. Four major compounds including tectorigenin, iristectorigenin A, irigenin, and irisflorentin were screened and identified as α‐glucosidase inhibitors, and then the four active compounds abovementioned were subsequently isolated by semipreparative high‐performance liquid chromatography (99.89, 88.97, 99.79, and 99.97% purity, respectively). The results demonstrate that ionic liquid extraction can be successfully applied to the extraction of isoflavones from B. chinensis .     

Rapid determination of lipophilic vitamins in human serum by ultra‐high performance liquid chromatography using a fluorinated column and high‐throughput miniaturized liquid–liquid extraction

A high‐throughput miniaturized liquid–liquid extraction procedure followed by a simple ultra‐high performance liquid chromatography method coupled with fluorescence detection for bioanalytical analysis of all tocopherol isomers and retinol in human serum has been developed and validated. In the extraction procedure, a synthetic internal standard tocol was used, which does not occur in the human body. The separation of structurally related vitamins was achieved using a new generation of pentafluorophenyl propyl core–shell stationary phase with elution using methanol and an aqueous solution of ammonium acetate. The fluorescence of retinol and tocopherol isomers was detected at λ_ex = 325, 295 nm and λ_em = 480, 325 nm, respectively. The rapid baseline separation of all analytes was accomplished within 4.0 min. The sensitivity of method was demonstrated with lower limits of quantification: retinol 0.01 μM, α‐tocopherol 0.38 μM, β‐tocopherol 0.18 μM, γ‐tocopherol 0.14 μM, and δ‐tocopherol 0.01 μM. Possible application of this method in clinical practice was confirmed by the analysis of human serum samples from healthy volunteers. Finally, the simultaneous determination of retinol and all tocopherol isomers in human serum can enable the clarification of their role in metabolism and in diseases such as cancer.     

Three‐phase hollow‐fiber microextraction combined with ion‐pair high‐performance liquid chromatography for the simultaneous determination of five components of compound α‐ketoacid tablets in human urine

The determination of α‐ketoacid concentration is demanded to evaluate the absorption and metabolic behavior of compound α‐ketoacid tablets taken by chronic kidney disease patients. To eliminate the interference of endogenous substance of urine and enrich the analytes, a three‐phase hollow‐fiber liquid‐phase microextraction combined with ion‐pair high‐performance liquid chromatography method was established for the determination of d ,l ‐α‐hydroxymethionine calcium, d ,l ‐α‐ketoisoleucine calcium, α‐ketovaline calcium, α‐ketoleucine calcium, and α‐ketophenylalanine calcium of compound α‐ketoacid tablets in human urine samples. The extraction parameters, such as organic solvent, pH of donor phase and acceptor phase, stirring rate, and extraction time were optimized. Under the optimal conditions, the obtained enrichment factors were up to 11‐, 110‐, 198‐, 202‐, and 50‐fold, respectively. The calibration curves for these analytes were linear over the range of 0.1–10 mg/L for α‐ketovaline calcium, d ,l ‐α‐ketoisoleucine calcium, and α‐ketoleucine calcium, 0.5–10 mg/L for d ,l ‐α‐hydroxymethionine calcium, and α‐ketophenylalanine calcium with r > 0.99. The relative standard deviations (n = 5) were less than 6.27% and the LODs were 100.7, 10.0, 5.8, 7.8, and 8.6 μg/L (based on S/N = 3), respectively. Good recoveries from spiked urine samples (92–118%) were obtained. The proposed method demonstrated excellent sample clean‐up and analytes enrichment to determine the five components in human urine.     

A novel sorbent based on carbon nanotube/amino‐functionalized sol–gel for the headspace solid‐phase microextraction of α‐bisabolol from medicinal plant samples using experimental design

A novel sol–gel coating on a stainless‐steel fiber was developed for the first time for the headspace solid‐phase microextraction and determination of α‐bisabolol with gas chromatography and flame ionization detection. The parameters influencing the efficiency of solid‐phase microextraction process, such as extraction time and temperature, pH, and ionic strength, were optimized by the experimental design method. Under optimized conditions, the linear range was between 0.0027 and 100 μg/mL. The relative standard deviations determined at 0.01 and 1.0 μg/mL concentration levels (n = 3), respectively, were as follows: intraday relative standard deviations 3.4 and 3.3%; interday relative standard deviations 5.0 and 4.3%; and fiber‐to‐fiber relative standard deviations 6.0 and 3.5%. The relative recovery values were 90.3 and 101.4% at 0.01 and 1.0 μg/mL spiking levels, respectively. The proposed method was successfully applied to various real samples containing α‐bisabolol.     

Determination of four sulfonylurea herbicides in tea by matrix solid‐phase dispersion cleanup followed by dispersive liquid–liquid microextraction

Matrix solid‐phase dispersion combined with dispersive liquid–liquid microextraction has been developed as a new sample pretreatment method for the determination of four sulfonylurea herbicides (chlorsulfuron, bensulfuron‐methyl, chlorimuron‐ethyl, and pyrazosulfuron) in tea by high‐performance liquid chromatography with diode array detection. The extraction and cleanup by matrix solid‐phase dispersion was carried out by using CN‐silica as dispersant and carbon nanotubes as cleanup sorbent eluted with acidified dichloromethane. The eluent of matrix solid‐phase dispersion was evaporated and redissolved in 0.5 mL methanol, and used as the dispersive solvent of the following dispersive liquid–liquid microextraction procedure for further purification and enrichment of the target analytes before high‐performance liquid chromatography analysis. Under the optimum conditions, the method yielded a linear calibration curve in the concentration range from 5.0 to 10 000 ng/g for target analytes with a correlation coefficients (r²) ranging from 0.9959 to 0.9998. The limits of detection for the analytes were in the range of 1.31–2.81 ng/g. Recoveries of the four sulfonylurea herbicides at two fortification levels were between 72.8 and 110.6% with relative standard deviations lower than 6.95%. The method was successfully applied to the analysis of four sulfonylurea herbicides in several tea samples.     

Simultaneous determination and qualitative analysis of six types of components in Naoxintong capsule by miniaturized matrix solid‐phase dispersion extraction coupled with ultra high‐performance liquid chromatography with photodiode array detection and quadrupole time‐of‐flight mass spectrometry

A simple and effective sample preparation process based on miniaturized matrix solid‐phase dispersion was developed for simultaneous determination of phenolic acids (gallic acid, chlorogenic acid, ferulic acid, 3,5‐dicaffeoylqunic acid, 1,5‐dicaffeoylqunic acid, rosmarinic acid, lithospermic acid, and salvianolic acid B), flavonoids (kaempferol‐3‐O‐rutinoside, calycosin, and formononetin), lactones (ligustilide and butyllidephthalide), monoterpenoids (paeoniflorin), phenanthraquinones (cryptotanshinone), and furans (5‐hydroxymethylfurfural) in Naoxintong capsule by ultra high‐performance liquid chromatography. The optimized condition was that 25 mg Naoxintong powder was blended homogeneously with 100 mg Florisil PR for 4 min. One milliliter of methanol/water (75:25, v/v) acidified by 0.05% formic acid was selected to elute all components. It was found that the recoveries of the six types of components ranged from 61.36 to 96.94%. The proposed miniaturized matrix solid‐phase dispersion coupled with ultra high‐performance liquid chromatography was successfully applied to simultaneous determination of the six types of components in Naoxintong capsules. The results demonstrated that the proposed miniaturized matrix solid‐phase dispersion coupled with ultra high‐performance liquid chromatography could be used as an environmentally friendly tool for the extraction and determination of multiple bioactive components in natural products.     

Reversed‐phase ion‐pair solid‐phase extraction and ion chromatography analysis of pyrrolidinium ionic liquid cations in environmental water samples

A method of reversed‐phase ion‐pair solid‐phase extraction combined with ion chromatography for determination of pyrrolidinium ionic liquid cations (N‐methyl‐N‐ethyl pyrrolidinium, N‐methyl‐N‐propyl pyrrolidinium, and N‐methyl‐N‐butyl pyrrolidinium) in water samples was developed in this study. First, ion‐pair reagent sodium heptanesulfonate was added to the water samples after static, centrifugation and filteration. Then, pyrrolidinium cations in the samples were enriched and purified by a reversed‐phase solid‐phase extraction column, and eluted from the column with methanol aqueous solution as eluent. Finally, the eluate collected was analyzed by ion chromatography. The separation and direct conductivity detection of these pyrrolidinium cations by ion‐exchange column using 1.0 mM methanesulfonic acid (in water)/acetonitrile (97:3, v:v) as mobile phase was achieved within 10 min. By using this method, pyrrolidinium cations in Songhua River and Hulan River were successfully extracted with the recoveries ranging from 74.2 to 97.1% and the enrichment factor assessed as 60. Pyrrolidinium cations with the concentration of 0.001?0.03 mg/L can be enriched and detected in the water samples. The developed method for the determination of pyrrolidinium ionic liquid cations in water samples is simple and reliable, which provides a reference for the study of the potential impact of ionic liquids on the environment.     

Preparation of five high‐purity iridoid glycosides from Gardenia jasminoides Eills by molecularly imprinted solid‐phase extraction integrated with preparative liquid chromatography

Five iridoid glycosides were prepared using molecularly imprinted solid‐phase extraction combined with preparative high‐performance liquid chromatography. Hydrophilic molecularly imprinted polymers were synthesized using α‐1‐allyl‐2‐N‐acetyl glucosamine, which introduced an abundance of hydrophilic groups into the polymers. Using molecularly imprinted solid‐phase extraction as the sample pretreatment procedure, five iridoid glycosides, gardenoside, geniposide, shanzhiside, geniposidic acid, and genipin‐1‐O‐gentiobioside, were selectively enriched from Gardenia fructus extracts. Preparative high‐performance liquid chromatography then provided iridoid glycosides with a purity >98%. The structures were elucidated by using nuclear magnetic resonance spectroscopy, optical rotation and melting point measurements, and mass spectrometry. The results demonstrate that molecularly imprinted solid‐phase extraction combined with preparative high‐performance liquid chromatography was an efficient, rapid, and economical method for the preparation of bioactive compounds from natural products.     

Sulfonated poly(styrene‐divinylbenzene) modified with amines and the application for pipette‐tip solid‐phase extraction of carbendazim in apples

Sulfonated poly(styrene‐divinylbenzene) modified with five kinds of amine functional groups was applied to the determination of carbendazim in apple samples with a pipette‐tip solid‐phase extraction method. The structures of the polymers were characterized by scanning electron microscopy, Fourier transform infrared spectroscopy, and thermogravimetric analysis. Five different modifications of the solid‐phase extraction sorbent based on sulfonated poly(styrene‐divinylbenzene) were tested under static and pipette‐tip solid‐phase extraction conditions. The polymer modified with p‐methoxyaniline showed the best recognition capacity and adsorption amount for carbendazim. Under the optimum conditions, 3.00 mg of the adsorbent, 1.00 mL of ethyl acetate as washing solvent, and 1.00 mL of ammonia/acetonitrile (5:95, v/v) as elution solvent were used in the pretreatment procedure of apple samples. The calibration graphs of carbendazim in methanol were linear over 5.00–200.00 μg/mL, and the limits of detection and quantification were 0.01 and 0.03 μg/mL, respectively. The method recoveries of carbendazim were in the range of 91.31–98.13% with associated intraday relative standard deviations of 0.76–2.13% and interday relative standard deviations of 1.10–1.85%. Sulfonated poly(styrene‐divinylbenzene) modified with p‐methoxyaniline showed satisfactory results (recovery: 97.96%) and potential for the rapid purification of carbendazim in apple samples combined with the pipette‐tip solid‐phase extraction.     

Rapid determination of α‐tocopherol in cereal grains using dispersive liquid–liquid microextraction followed by HPLC

The traditional for the determination of α‐tocopherol in cereal grains includes saponification of a sample followed by liquid–liquid extraction, and it is time‐ and solvent consuming. In this study, a dispersive liquid–liquid microextraction (DLLME) method was developed to extract α‐tocopherol in situ from the saponified grain sample solution. The DLLME experimental parameters including the type and volume of extractants, the volume of dispersers, the addition of salt and the extraction/centrifuging time were examined and optimized. The recommended analytical procedure showed excellent precision (relative SDs of the α‐tocopherol amount of 3.1% over intraday and 7.2% over interday), high sensitivity (the detection limit of 1.9 ng/mL), and strong recovery values (88.9–102.5%). In addition, statistical analyses showed no significant difference between the detected amounts of α‐tocopherol found by the standardized method and this new procedure. The method was successfully applied to determining the amounts and distribution of α‐tocopherol in 14 cereal grain samples.     

Dispersive solid‐phase extraction combined with dispersive liquid‐liquid microextraction for simultaneous determination of seven succinate dehydrogenase inhibitor fungicides in watermelon by ultra high performance liquid chromatography with tandem mass spectrometry

In this study, a simple and accurate sample preparation method based on dispersive solid‐phase extraction and dispersive liquid‐liquid microextraction has been developed for the determination of seven novel succinate dehydrogenase inhibitor fungicides (isopyrazam, fluopyram, pydiflumetofen, boscalid, penthiopyrad, fluxapyroxad, and thifluzamide) in watermelon. The watermelon samples were extracted with acetonitrile, cleaned up by dispersive solid‐phase extraction procedure using primary secondary amine, extracted and concentrated by the dispersive liquid‐liquid microextraction procedure with 1,1,2,2‐tetrachloroethane, and then analyzed by ultra high performance liquid chromatography with tandem mass spectrometry. The main experimental factors affecting the performance of dispersive solid‐phase extraction and dispersive liquid‐liquid microextraction procedure on extraction efficiency were investigated. The proposed method had a good linearity in the range of 0.1–100 µg/kg with correlation coefficients (r) of 0.9979–0.9999. The limit of quantification of seven fungicides was 0.1 µg/kg in the method. The fortified recoveries of seven succinate dehydrogenase inhibitor fungicides at three levels ranged from 72.0 to 111.6% with relative standard deviations of 3.4–14.1% (n = 5). The proposed method was successfully used for the rapid determination of seven succinate dehydrogenase inhibitor fungicides in watermelon.     

Determination of vitamin A in blood serum based on solid‐phase extraction using cetyltrimethyl ammonium bromide‐modified attapulgite

Cetyltrimethyl ammonium bromide‐modified attapulgite was prepared and utilized as a novel sorbent in a simple solid‐phase extraction method for the determination of vitamin A in blood serum. Several factors affecting extraction efficiency were systematically optimized, including the sampling solvent and its volume, as well as the elution solvent and its volume. Under the optimal solid‐phase extraction conditions, the adsorption capacity of vitamin A was as high as 28 mg/g according to the Langmuir isotherm model. Based on the developed solid‐phase extraction method, the level of vitamin A in 200 µL blood serum sample could be accurately determined by high‐performance liquid chromatography. The recoveries of vitamin A spiked in 10% v/v methanol aqueous solutions were in the range of 86.9–92.8%, with the relative standard deviations not more than 8.1%. The method was applied to the determination of vitamin A in serum samples from 20 pregnant women. Compared with the previously reported solid‐phase extraction methods for determination of vitamin A in serum, our developed cetyltrimethyl ammonium bromide‐modified attapulgite‐based solid‐phase extraction method used lower serum volume, omitted extra steps (i.e. evaporation and re‐dissolution), and eliminated internal standard. The results were promising for it to be used in routine monitoring during pregnancy.     

Improvement of α‐tocopherols long‐term efficiency by modeling its heterogeneous natural environment in polyethylene

The natural antioxidant vitamin E (α‐tocopherol) is of interest to use in packaging applications to decrease the amount of toxic products migrating into food and drugs. We have earlier shown that the long‐term efficiency of α‐tocopherol in polyethylene (PE) films is poor. α‐Tocopherol is located in the lipid phase of the cell in vivo and it has been revealed that it is more efficient in a polar substrate. PE is more hydrophobic and homogenous than the heterogeneous and hydrophilic lipid phase. Three different additive systems were investigated to model α‐tocopherols heterogeneous natural environment in PE. Two of these had carboxylic acid groups, EAA and polyTRIM/PAA core‐shell particles (Core), and the third, oat starch, had no carboxylic acid groups. The materials were thermally aged and characterized by chemiluminescence (CL), FTIR, chromatography, and thermal analysis. The EAA system as well as the Core system improved the antioxidant properties of α‐tocopherol in PE, and the Core system had the best performance. We know that starch has stabilizing properties in PE, but it had no effect on the efficiency of α‐tocopherol. © 2006 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 44: 1660–1666, 2006     

Fe3O4 nanoparticles coated with polyhedral oligomeric silsesquioxanes and β‐cyclodextrin for magnetic solid‐phase extraction of carbaryl and carbofuran

In this study, porous sandwich structure Fe₃O₄ nanoparticles coated by polyhedral oligomeric silsesquioxanes and β‐cyclodextrin were prepared by surface polymerization and were used as the magnetic solid phase extraction adsorbent for the extraction and determination of carbaryl and carbofuran. The Fe₃O₄ nanoparticles coated with polyhedral oligomeric silsesquioxanes and β‐cyclodextrin were characterized by Fourier transform infrared spectroscopy, X‐ray diffraction, thermogravimetric analysis, vibrating sample magnetometry, and scanning electron microscopy. After optimizing the extraction conditions, a method that combined magnetic solid phase extraction with high‐performance liquid chromatography was developed for the determination of carbaryl and carbofuran in apple. The method exhibited a good linearity in the range of 2–400 μg/kg for carbaryl and carbofuran (R² = 0.9995), respectively. The limits of detection were 0.5 μg/kg of carbaryl and 0.7 μg/kg for carbofuran in apple, respectively. Extraction recoveries ranged from 94.2 to 103.1% with the preconcentration factor of 300 and the relative standard deviations were less than 5.9%. These results indicated that the method combined magnetic solid phase extraction with high‐performance liquid chromatography and was promising for the determination of carbaryl and carbofuran at trace amounts.     

Stereoisomer separation of flavanones and flavanone‐7‐O‐glycosides by means of nanoliquid chromatography employing derivatized β‐cyclodextrins as mobile‐phase additive

A nanoliquid chromatographic method for the stereoisomer separation of some flavanone aglycones and 7‐O‐glycosides has been proposed employing a C18 capillary column and a chiral mobile‐phase additive such as cyclodextrin. The chiral separation of eriodictyol, naringenin, and hesperitin was obtained by addition of carboxymethyl‐β‐cyclodextrin to the mobile phase, whereas eriocitrin, naringin, narirutin, and hesperidin diastereoisomers were resolved by using sulfobutyl ether‐β‐cyclodextrin. The influence of the composition of the mobile phase, the length of the capillary column, and the flow rate on the chiral recognition were investigated. At optimum conditions, baseline separation for the selected aglycones and glycosylated forms were achieved with a mobile phase consisting of 50 mM sodium acetate buffer pH 3 and 30% methanol containing 20 mM of carboxymethyl‐β‐cyclodextrin and 10 mM of sulfobutyl ether‐β‐cyclodextrin, respectively. Precision, linearity, and sensitivity of the method were tested. Limits of detection and quantification for the studied flavanone glycosides were in the range 1.3‐2.5 and 7.5‐12.5 µg/mL, respectively. The method was used for the determination of the diastereomeric composition of the flavanone‐7‐O‐glycosides in Citrus juices after solid‐phase extraction procedure.     

Graphene‐doped electrochemical copolymer coating of 2,2‐bithiophene and 3‐methylthiophene for the highly effective solid‐phase microextraction of volatile benzene homologues

We report the electrochemical fabrication of a poly(2,2‐bithiophene‐co‐3‐methylthiophene)‐graphene composite coating and its application in the headspace solid‐phase microextraction and gas chromatography determination of benzenes (i.e., bromobenzene, 4‐bromotoluene, 2‐nitrotoluene, 3‐nitrotoluene and 1,2,4‐trichlorobenzene). The coating was uniform and showed cauliflower‐like microstructure. It had high thermal stability (up to 375°C) and could be used for at least 180 times of solid‐phase microextraction without a decrease in extraction performance. Furthermore, it presented high extraction capacity for the benzenes due to the hydrophobic effect and π–π interaction between the analytes and the coating. Under optimized extraction conditions, good linearity (correlation coefficients higher than 0.9946), wide linear range (0.01–50 μg/L), and low limits of detection (5.25–12.5 ng/L) were achieved for these analytes. The relative standard deviation was lower than 5.7% for five successive measurements with one fiber, and the relative standard deviation for fiber‐to‐fiber was 4.9–6.8% (n = 5). The solid‐phase microextraction and gas chromatography method was successfully applied for the determination of three real samples, and the recoveries for standards added were 89.6–106% for nail polish, 85.8–110% for hair dye, and 90–106.2% for correction fluid, respectively.     

Automatic on‐line solid‐phase extraction with ultra‐high performance liquid chromatography and tandem mass spectrometry for the determination of ten antipsychotics in human plasma

An automatic on‐line solid‐phase extraction with ultra‐high performance liquid chromatography and tandem mass spectrometry method was developed for the simultaneous determination of ten antipsychotics in human plasma. The plasma sample after filtration was injected directly into the system without any pretreatment. A Shim‐pack MAYI‐C₈ (G) column was used as a solid‐phase extraction column, and all the analytes were separated on a Shim‐pack XR‐ODS III column with a mobile phase consisting of 0.1% v/v formic acid in water with 5 mM ammonium acetate and acetonitrile. The method features were systematically investigated, including extraction conditions, desorption conditions, the equilibration solution, the valve switching time, and the dilution for column‐head stacking. Under the optimized conditions, the whole analysis procedure took only 10 min. The limits of quantitation were in the range of 0.00321–2.75 μg/L and the recoveries ranged from 75.9 to 122%. Compared with the off‐line ultra‐high performance liquid chromatography and the reported methods, this validated on‐line method showed significant advantages such as minimal pretreatment, shortest analysis time, and highest sensitivity. The results indicated that this automatic on‐line method was rapid, sensitive, and reliable for the determination of antipsychotics in plasma and could be extended to other target analytes in biological samples.     

Rapid high‐performance liquid chromatography–tandem mass spectrometry method for simultaneous measurement of venlafaxine and O‐desmethylvenlafaxine in human plasma and its application in comparative bioavailability study

A rapid high‐performance liquid chromatography–tandem mass spectrometry method has been developed and validated for simultaneous measurement of venlafaxine and O‐desmethylvenlafaxine in human plasma using fluoxetine as an internal standard. In the liquid–liquid extraction method, compounds and internal standard were extracted from plasma using methyl tertiary butyl ether as an extraction solvent. The HPLC separation of the analytes was performed on a Zorbax SB‐C₁₈, 50 × 4.6 mm, 5 µm column, using a isocratic elution program using a mobile phase consisting of HPLC‐grade methanol: 5 mm ammonium acetate (80:20 v/v) at a flow‐rate of 1.0 mL/min with a total runtime of 3.0 min. The proposed method has been validated with a linear range of 4–400 ng/mL for venlafaxine and 5–500 ng/mL for O‐desmethyl venlafaxine. The method was applied for a bio‐equivalence study of 75 mg tablets formulation in 32 Indian male healthy subjects under fasting conditions. Copyright © 2009 John Wiley & Sons, Ltd.