Ultrasound-assisted extraction and derivatization of sterols and fatty alcohols from olive leaves and drupes prior to determination by gas chromatography–tandem mass spectrometry

A method for simultaneous determination of sterols and fatty alcohols in olive leaves and drupes based on ultrasound-assisted extraction and derivatization prior to individual identification–quantitation by chromatographic separation and mass spectrometry detection (single ion monitoring mode) is reported here. The sample preparation procedure involves the following steps: (i) leaching of the raw material accelerated by ultrasound; (ii) saponification of the leachate, also accelerated by ultrasound, and separation of the unsaponifiable matter; (iii) cleaning of the extract by solid-phase extraction; (iv) silylation of the target analytes—also assisted by ultrasound; (v) injection into the gas chromatograph for identification–simultaneous quantitation of the two families of compounds. Individual separation–determination of the fatty alcohols and sterols provide limits of detection (LOD) in the range 9.8 × 10⁻² to 2 μg/l and 5.0–6.0 μg/l, respectively. The LOQs range from 0.3 to 0.9 μg/l and 17.0 to 21.0 μg/l, and the linear dynamic ranges are between LOQ and 25.0 μg/ml. The between-day precision, expressed as relative standard deviation (RSD), ranges between 3.6 and 6.1% and the within-laboratory reproducibility, also expressed as RSD, between 6.4 and 9.2%. Within the study of the metabolomic profile of the unsaponifiable fraction in olive tree, the method has been applied to the determination of the target analytes in different varieties of olive trees cultivated in the same zone, so that differences in this unsaponifiable fraction can be attributed to characteristics of the target varieties. As compared with its European Union counterpart, the method is endowed with similar analytical characteristics and drastic shortening of the operational time.     

Targeted analysis of multiple pharmaceuticals,plant toxins and other secondary metabolites in herbal dietary supplements by ultra-high performance liquid chromatography–quadrupole-orbital ion trap mass spectrometry

In this study, an ultra-high performance liquid chromatography–quadrupole-orbital ion trap mass spectrometry (UHPLC–Q-orbitrap MS) method was developed and validated for simultaneous determination of 96 pharmaceuticals, plant toxins, and other plant secondary metabolites in herbal dietary supplements. Target analytes were extracted from samples using the QuEChERS (quick easy cheap effective rugged safe) procedure. The instrument was operated in full MS–data dependent tandem mass spectrometry (full MS–dd-MS/MS) acquisition mode which enabled collection of quantitative high resolution (HR) full mass spectral data and confirmatory HR MS/MS data in a single run. The method provided excellent selectivity in both full MS and dd-MS/MS mode. Under optimized collision energy settings, product ion spectra containing both precursor and two or more product ions were obtained for most of the analytes. Limits of detection (LODs) and limits of quantification (LOQs) for the method differed significantly for the examined matrices. LODs ≤ 10 μg kg⁻¹ and LOQs ≤ 50 μg kg⁻¹ were obtained for 48 to 81% of target compounds across five different matrices. With the exception of highly polar analytes, the optimized QuEChERS extraction procedure provided acceptable recoveries in the range 70%–120%. The precision of the method, characterized as the relative standard deviation (RSD, n = 5), was ≤25% and ≤18% at spiking concentrations of 50 μg kg⁻¹ and 500 μg kg⁻¹, respectively. Because of variations in matrix effects in extracts of herbal dietary supplements that differed in composition, the method of standard additions and an approach based on dilution of matrix components followed by quantification using solvent standards were applied for quantification. The procedure was used to examine commercial dietary supplements for the 96 analytes of interest. To the best of our knowledge, this is the first report of an integrated analysis and quantification of this wide range of compounds.     

Determination of pyridine, 2-picoline, 4-picoline and quinoline from mainstream cigarette smoke by solid-phase extraction liquid chromatography/electrospray ionization tandem mass spectrometry

The present paper describes the development and validation of a new reversed-phase liquid chromatography–electrospray ionization tandem mass spectrometric method (RP-HPLC–ESI-MS/MS) for simultaneous determination of pyridine, 2-picoline, 4-picoline and quinoline from mainstream cigarette smoke. Liquid–liquid extraction followed by solid-phase extraction was applied to extract the target analytes from cigarette smoke. Baseline chromatographic separation was achieved by utilizing a Zorbax SB-Aq (4.6 × 150 mm, 5 μm) column in gradient chromatographic conditions with acetonitrile and ammonium acetate buffer as mobile phases. Popular commercially available Indian brand filtered and non-filtered cigarettes were analyzed using the same method. The identification of each chemical was established by chromatographic retention times, analyte specific fragmentation patterns and relative peak area ratios of two product/precursor ion pairs. The limit of detection of this method ranged from 1.74 to 14.32 ng/cig using an injection volume of 20 μl. The reproducibility of this method is excellent and better standard deviations were obtained compared to literature reported values for these chemicals. RSD value is less than 9% for all analytes.     

Analysis of volatiles and semivolatiles in drinking water by microextraction and thermal desorption

A new technique to analyze aqueous samples for nanograms per liter levels of volatile and semivolatile compounds using microextraction and thermal desorption into a gas chromatograph/ion trap mass spectrometer (GC/MS) is described. This method is inherently sensitive (50 mL of aqueous sample is extracted prior to each desorption), uses no solvents, and detects volatiles and semivolatiles in the same analysis. Aqueous standards and environmental samples are pumped through a length of porous-layer open-tubular capillary column, which is then thermally desorbed onto a 30 m × 0.25 mm i.d. analytical column interfaced to an ion trap mass spectrometer for subsequent separation and detection. Sharp chromatographic peaks and reproducible retention times (RT) were observed. Replicate injections of surrogates (n = 6) averaged 32.6% R.S.D. Analysis of domestic tap water detected 55 analytes, some at the low-nanograms per liter level, and detected 3 halogenated ethenes, not previously reported in drinking water. Analysis of an aqueous sample from a municipal ground water source detected the presence of numerous semivolatile compounds at trace-levels.     

Simultaneous quantification of multiple classes of phenolic compounds in blood plasma by liquid chromatography–electrospray tandem mass spectrometry

A method using high performance liquid chromatography–electrospray tandem mass spectrometry (LC–ESI-MS/MS) in positive ion mode was developed for the simultaneous analysis of 30 phenolic compounds, including four estrogens, bisphenol A (BPA), 10 hydroxylated polybrominated dephenyl ethers (OH-PBDEs) and 15 bromophenols (BRPs), in blood plasma. In the present method, derivatization with dansyl chloride was employed, and all the derivertized target compounds were well resolved on a 100 mm Xbridge C18 column with acetonitrile and 0.1% formic acid as the mobile phases. Purification procedures, such as liquid–liquid extraction and silica-gel chromatography, were applied to reduce matrix effects in the sample extract and remove excess derivatizing reagents, thus permitting selective and sensitive detection of the target phenolic compounds. The limit of quantification for all analytes, with a signal-to-noise ratio of ∼10, was 2–30 pg/g (plasma weight) except for 6-OH-BDE-137 (30 pg/g) and 3-BRP (60 pg/g). The method was validated for recoveries (68–100%), accuracy (84–110%) and precision (3.7–11%) using charcoal-stripped bovine blood plasma spiked with all target compounds (500 and 5000 pg/mL). Finally, the method was applied to analyze six blood plasma samples from frogs and cormorants, where two natural estrogens, one BPA, one OH-PBDE and four BRPs were detected. The greatest total concentrations of estrogens coincided with the least total concentrations of other phenolic compounds for both species. The proposed method based on derivatization followed by LC–MS/MS provides a novel method to simultaneously monitor multiple groups of phenolic compounds in blood plasma.     

Validation and use of a fast sample preparation method and liquid chromatography–tandem mass spectrometry in analysis of ultra-trace levels of 98 organophosphorus pesticide and carbamate residues in a total diet study involving diversified food types

This paper reports a comprehensive sensitive multi-residue liquid chromatography–tandem mass spectrometry (LC–MS/MS) method for detection, identification and quantitation of 73 pesticides and their related products, a total of 98 analytes, belonging to organophosphorus pesticides (OPPs) and carbamates, in foods. The proposed method makes use of a modified QuEChERS (quick, easy, cheap, effective, rigged, and safe) procedure that combines isolation of the pesticides and sample clean-up in a single step. Analysis is performed by liquid chromatography-electrospray ionization–tandem mass spectrometry operated in the multiple reaction monitoring (MRM) mode, acquiring two specific precursor-product ion transitions per target compound. Two main fragment ions for each pesticide were obtained to achieve the identification according to the SANCO guidelines 10684/2009. The method was validated with various food samples, including edible oil, meat, egg, cheese, chocolate, coffee, rice, tree nuts, citric fruits, vegetables, etc. No significant matrix effect was observed for tested pesticides, therefore, matrix-matched calibration was not necessary. Calibration curves were linear and covered from 1 to 20 μg L⁻¹ for all compounds studied. The average recoveries, measured at 10 μg kg⁻¹, were in the range 70–120% for all of the compounds tested with relative standard deviations below 20%, while a value of 10 μg kg⁻¹ has been established as the method limit of quantitation (MLOQ) for all target analytes. Similar trueness and precision results were also obtained for spiking at 200 μg kg⁻¹. Expanded uncertainty values were in the range 21–27% while the HorRat ratios were below 1. The method has been successfully applied to the analysis of 700 food samples in the course of a baseline monitoring study of OPPs and carbamates.     

Ultra-high-pressure liquid chromatography–tandem mass spectrometry method for the determination of alkylphenols in soil

A novel method has been developed for the determination of alkylphenols in soil by ultra-high-pressure liquid chromatography employing small particle sizes, combined with tandem mass spectrometry. Soil samples were extracted with pressurized liquid extraction (PLE) and then cleaned with solid-phase extraction (SPE). The extracts were separated on C18 column (1.7 μm, 50 mm × 2.1 mm) with a gradient elution and a mobile phase consisting of water and acetonitrile, and then detected by an electrospray ionization tandem mass spectrometry in negative ion mode with multiple reaction monitoring (MRM). Compared with traditional liquid chromatography, it took ultra-high-pressure liquid chromatography much less time to analyze alkylphenols. Additionally, the ultra-high-pressure liquid chromatography/tandem mass spectrometry method produces satisfactory reliability, sensitivity, and accuracy. The average recoveries of the three target analytes were 74.0–103.4%, with the RSD < 15%. The calibration curves for alkylphenols were linear within the range of 0.01–0.4 μg/ml, with the correlation coefficients greater than 0.99. When 10 g soil sample was used for analysis, the limits of quantification (LOQs) of the three alkylphenols were all 1.0 μg/kg.     

Selective separation and purification of highly polar basic compounds using a silica-based strong cation exchange stationary phase

Compared to moderately and weakly hydrophilic bases, highly polar basic compounds are even more difficult to separate due to their poor retention in reversed phase (RP) mode. This study described the successful applications of a strong cation exchange (SCX) stationary phase to achieve symmetric peak shape, adequate retention and selectivity in the separation of very polar basic compounds. Salt and acetonitrile concentrations were adjusted to optimize the separation. Good correlations (R² = 0.998–1.000) between the logarithm of the retention factor and the logarithm of salt or acetonitrile concentration were obtained. Gradients generated by changing salt or acetonitrile concentration were compared for the analysis of different highly polar bases. Although all of the analytes were eluted more quickly with an acetonitrile gradient, the effect of the gradients tested on peak width and peak shape varied with respect to analyte. In addition, the effects of different types of cation and anion additives were also investigated. After separation parameters were acquired, the SCX-based method was utilized to analyze highly hydrophilic alkaloids from Scopolia tangutica Maxim with high separation efficiency (plate numbers > 32,000 m⁻¹). Concurrently, one very polar alkaloid fraction was purified with symmetric peak shape using the current method. Our results suggest that SCX stationary phase can be used as an alternative to RP stationary phase in the analysis and purification of highly hydrophilic basic compounds.     

Optimization of two-dimensional gas chromatography time-of-flight mass spectrometry for separation and estimation of the residues of 160 pesticides and 25 persistent organic pollutants in grape and wine

Two-dimensional gas chromatography (GC × GC) coupled with time-of-flight mass spectrometric (TOFMS) method was optimized for simultaneous analysis of 160 pesticides, 12 dioxin-like polychlorinated biphenyls (PCBs), 12 polyaromatic hydrocarbons (PAHs) and bisphenol A in grape and wine. GC × GC–TOFMS could separate all the 185 analytes within 38 min with >85% NIST library-based mass spectral confirmations. The matrix effect quantified as the ratio of the slope of matrix-matched to solvent calibrations was within 0.5–1.5 for most analytes. LOQ of most of the analytes was ≤10 μg/L with nine exceptions having LOQs of 12.5–25 μg/L. Recoveries ranged between 70 and 120% with <20% expanded uncertainties for 151 and 148 compounds in grape and wine, respectively, with intra-laboratory Horwitz ratio <0.2 for all analytes. The method was evaluated in the incurred grape samples where residues of cypermethrin, permethrin, chlorpyriphos, metalaxyl and etophenprox were detected at below MRL.     

Determination of phosmet and its metabolites in olives by matrix solid-phase dispersion and gas chromatography-mass spectrometry

An accurate method based on matrix solid-phase dispersion (MSPD), and gas chromatography-mass spectrometry (GC/MS) was developed for determination of phosmet residues and its metabolites (phosmet-oxon, phthalimide, N-hydroxymethylphthalimide, and phthalic acid) in olive fruits. After testing different sorbents and eluents for MSPD extraction, C₁₈ and acetonitrile were found to be the most appropriate for clean-up of the samples, in terms of yields and efficient removal of interfering compounds. All analytes were determined in selective ion monitoring (SIM) mode following a derivatization step with N,O-bis-trimethylsilyltrifluoroacetamide (BSTFA) containing 1% of trimethylchlorosilane (TMCS), except for phosmet and phosmet-oxon which were analyzed directly. The method showed suitable linearity (correlation coefficients higher than 0.8919 for all the compounds) and suitable sensitivity (limit of detection lower than 0.06 mg/kg). It was successfully applied in the analysis of olive fruits collected during the preharvest interval and olive oil. Phosmet residues found in all samples were lower than the maximum residue limits established by legislation (2 mg/kg).     

Simultaneous determination of aflatoxins B1, B2, G1, G2, M1 and M2 in peanuts and their derivative products by ultra-high-performance liquid chromatography-tandem mass spectrometry

A reliable ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method for simultaneous determination of aflatoxins B1, B2, G1, G2, M1 and M2 in peanuts and their derivative products was developed. The sample was extracted by 84% of acetonitrile aqueous solution and the extract was purified by a reliable solid phase extraction-based clean-up method. Then, the analytes were separated on Acquity UPLC HSS T3 column (100 mm × 2.1 mm, 1.8 μm particle size), and eluted with a mobile phase consisting of (A) water containing 0.1% formic acid and (B) acetonitrile/methanol (50/50, v/v). The separated compounds were detected with a Waters Micromass Quattro Ultima Pt tandem quadrupole mass spectrometer operating in positive electro-spray ionization using multiple reaction monitoring mode. The established method was extensively validated by determining the linearity (R² ≥ 0.9990), average recovery (74.7-86.8%) and precision (relative standard deviation ≤ 10.9%). It was shown to be a suitable method for simultaneous determination of the six aflatoxins in peanuts and their derivative products. Finally, a total of 73 samples randomly collected from different areas in Zhejiang province were screened for aflatoxins with the proposed method. The results showed that 31 samples of peanut butter, 14 samples of fresh peanut and 5 samples of musty peanut were contaminated with aflatoxins. Meanwhile, this was the first report on aflatoxins M1 and M2, which were found in unprocessed peanuts and their derivative products.     

Simultaneous determination of nine trace mono- and di-chlorophenols in water by ion chromatography atmospheric pressure chemical ionization mass spectrometry

A novel analytical method was proposed for the rapidly simultaneous determination of nine mono-chlorophenols (MCPs) and di-chlorophenols (DCPs) in water samples using eluent generator ion chromatography (IC) coupled with an atmospheric pressure chemical ionization mass spectrometry (APCI-MS) in the negative mode. The IC separation was carried out on an IonPac^® AS11 analytical column (250 mm × 4.0 mm) using gradient KOH containing 15% acetonitrile as organic modifier at a constant flow rate of 1.0 mL/min. The molecular ions m/z [M − H]⁻ 127 and 161 were selected for the quantification in selected ion monitoring (SIM) mode for MCPs and DCPs, respectively. The average recoveries were between 80.6% and 92.6%. Within-day and day-to-day relative standard deviations were less than 12.1% and 13.3%, respectively. The method allowed the nine objective compounds in water samples to be determined at μg/L levels. It was confirmed that this method could be used in routine analysis.     

Determination of ent-kaurene in subcutaneous fat of Iberian pigs by gas chromatography multi-stage mass spectrometry with the aim to differentiate between intensive and extensive fattening systems

This work presents a gas chromatography multi-stage mass spectrometry (GC-MS³) method for the determination of ent-kaurene in subcutaneous fat of Iberian pig, present in adipose tissue of animals due to pasture ingestion (extensive fattening system). The method comprises a saponification and a liquid-liquid extraction of the unsaponifiable fraction, followed by an isolation of the hydrocarbon fraction by high performance liquid chromatography (HPLC) and analysis by GC-MS³ (ion trap) with electronic ionization. The GC-MS³ analysis allows the isolation and characterization of specific fragments from the original (MS¹) molecular structure, and particularly, those fragments originated from the precursor ion (m/z = 229) characteristic of ent-kaurene. The MS/MS product fragment m/z = 213 is used as a further precursor fragment giving rise to a MS³ spectrum specific for ent-kaurene. The limit of detection of the MS³ technique is lower than 0.2 μg kg⁻¹ and a linear regression has been found between 0.2 and 112 μg kg⁻¹. This method is applicable for the determination of the fattening system of the Iberian pig.     

Ultra performance liquid chromatography-tandem mass spectrometry for the determination of epirubicin in human plasma

A novel method has been developed for the determination of epirubicin in human plasma by ultra performance liquid chromatography combined with tandem mass spectrometry (UPLC-MS/MS). Epirubicin and internal standard epidaunorubicin were achieved from plasma via solid-phase extraction (SPE) using Oasis HLB cartridge. The analysis was performed on an AcQuity UPLC™ BEH C₁₈ column (1.7 μm, 50 mm × 1 mm i.d.) utilizing a gradient elution profile and a mobile phase consisting of 0.1% formic acid in water and acetonitrile. The analytes were detected using an electrospray ionization tandem mass spectrometry in positive ion mode with multiple reaction monitoring (MRM). This method combines both advantages of UPLC and MS/MS, producing superior reliability, sensitivity and accuracy to previously published methods. The calibration curve was linear (r² = 0.998) over the concentration range of 0.50-100.0 ng/ml. The limits of detection (LOD) and quantification (LOQ) for epirubicin were 0.10 and 0.50 ng/ml using 0.2 ml plasma sample, respectively. Recoveries of greater than 89% with intra- and inter-day precision (R.S.D.) less than 12% were obtained at concentrations above the LOQ. The present method has been successfully applied to analyze human plasma samples taken from patients administered epirubicin intravenously. Also, the principal metabolite epirubicinol was detected in all the patient plasma samples under investigation. The proposed method is very rapid, reliable and sensitive, and can be applicable to therapeutical drug monitoring and pharmacokinetic studies of epirubicin.     

Evaluation of two sample treatment methodologies for large-scale pesticide residue analysis in olive oil by fast liquid chromatography–electrospray mass spectrometry

In this study, a comprehensive evaluation of two simple sample treatment methodologies has been carried out for the development of large-scale multi-residue methods for pesticide testing in olive oil. The proposed methodologies are based on (a) liquid–liquid partitioning with acetonitrile followed by dispersive solid-phase extraction clean-up using graphitized carbon black, primary-secondary amine and C₁₈ sorbents; (b) liquid partitioning with acetonitrile saturated with petroleum ether followed by matrix solid-phase dispersion (MSPD) using aminopropyl as sorbent material and a Florisil cartridge for final clean-up in the elution step. To evaluate the proposed sample treatment methodologies, 105 representative multi-class pesticides were studied using fast liquid chromatography–electrospray time-of-flight mass spectrometry (LC–TOFMS). For validation purposes, recoveries studies were carried out at 10 and 100 μg kg⁻¹ levels, yielding recovery rates in the range 70–130% for 72% of analytes using liquid–liquid procedure and for 57% analytes using MSPD procedure. The LC–MS method provided good linearity, precision and accuracy. The limits of detection obtained were lower than 10 μg kg⁻¹ for more than 85% analytes using both sample treatment methodologies. In addition, minor matrix effects (i.e. signal suppression or enhancement ≤20%) were observed in ca. 70% of the studied compounds. Data obtained shows that both sample treatment methodologies proposed can be successfully applied for large-scale pesticide testing in olive oil samples, showing the ability to quickly detect trace amount of over one hundred target species with different physicochemical properties, without requiring expensive instrumentation for sample treatment step and involving relatively low amounts of solvent consumption and waste generation.     

Determination of anthelmintic drug residues in milk using ultra high performance liquid chromatography–tandem mass spectrometry with rapid polarity switching

A new UHPLC–MS/MS (ultra high performance liquid chromatography coupled to tandem mass spectrometry) method was developed and validated to detect 38 anthelmintic drug residues, consisting of benzimidazoles, avermectins and flukicides. A modified QuEChERS-type extraction method was developed with an added concentration step to detect most of the analytes at <1 μg kg⁻¹ levels in milk. Anthelmintic residues were extracted into acetonitrile using magnesium sulphate and sodium chloride to induce liquid–liquid partitioning followed by dispersive solid phase extraction for cleanup. The extract was concentrated into dimethyl sulphoxide, which was used as a keeper to ensure analytes remain in solution. Using rapid polarity switching in electrospray ionisation, a single injection was capable of detecting both positively and negatively charged ions in a 13 min run time. The method was validated at two levels: the unapproved use level and at the maximum residue level (MRL) according to Commission Decision (CD) 2002/657/EC criteria. The decision limit (CCα) of the method was in the range of 0.14–1.9 and 11–123 μg kg⁻¹ for drugs validated at unapproved and MRL levels, respectively. The performance of the method was successfully verified for benzimidazoles and levamisole by participating in a proficiency study.     

Multi-residue analysis of pharmaceutical compounds in wastewaters by dual solid-phase microextraction coupled to liquid chromatography electrospray ionization ion trap mass spectrometry

The aim of this article is to present a new procedure based on dual solid-phase microextraction (dSPME) for the simultaneous extraction of 16 pharmaceutical compounds with acidic and basic characteristics in urban wastewaters. Water samples are divided into two aliquots of 2 mL each extracted by two CW-TPR fibers at different pH values (pH 3 and 11) and with a NaCl concentration of 300 g L⁻¹ at 75 °C for 30 min. The analytes in both fibers are desorbed one after the other in the desorption chamber in static mode with mobile phase for 10 min. The extracts are injected into an LC system coupled to an ion trap mass spectrometer, leading to the accurate quantification of 16 pharmaceutical compounds in wastewaters, in MS² mode. All the target compounds found in wastewaters provide good signals corresponding to the protonated precursor ion [M+H]⁺. The parameters influencing adsorption and desorption of the analytes on fiber were optimized. The assessment of the analytical method was performed by studying the linearity (LOQ to 10 ng mL⁻¹) and the intra- and interday accuracy (89.2–109.7%) and precision (RSD <13.6%). The quantification limits obtained ranged between 0.005 and 0.05 μg L⁻¹. The application of the method to real samples proves its effectiveness in identifying and detecting naproxen, valsartan, bezafibrate, torasemide, diclofenac, carbamazepine, citalopram, lorazepan, fluoxetine, imipramine and amitriptyline in influent and effluent wastewater treatment plant samples.     

A rapid determination of propiverine and its N-oxide metabolite in human plasma by high performance liquid chromatography-electrospray ionization tandem mass spectrometry

A simple, fast and sensitive high-performance liquid chromatography (HPLC)-electrospray ionization (ESI) tandem mass spectrometric method (LC-MS/MS) has been developed for determination of propiverine and propiverine N-oxide metabolite in human plasma using oxybutynin as internal standard. Instead of extracting propiverine from plasma using organic solvents, which should be separated from the aqueous phase and evaporated before injecting the sample into the chromatograph, plasma sample containing propiverine and N-oxide was directly injected after precipitating proteins with acetonitrile. Numerous compounds in the plasma did not interfere with the highly specific multiple reaction monitoring in tandem mass spectrometric detection following C₈ reversed-phase chromatographic separation under conditions that eluted propiverine, N-oxide and oxybutynin within 2 min (0.1% formic acid in water/acetonitrile, 25:75, v/v). The LC-MS/MS method and an alternative LC-MS method, using methyl-t-butyl ether extraction and selected ion monitoring, were validated over 1-250 ng ml⁻¹ of propiverine and 2 to 500 ng ml⁻¹ of N-oxide, and successfully applied in a pharmacokinetic study. The lower limit of quantitation was 1 ng ml⁻¹ for propiverine and 2 ng ml⁻¹ for N-oxide in both methods.     

Determination of sulfonamides in swine muscle after salting-out assisted liquid extraction with acetonitrile coupled with back-extraction by a water/acetonitrile/dichloromethane ternary component system prior to high-performance liquid chromatography

A salting-out assisted liquid extraction coupled with back-extraction by a water/acetonitrile/dichloromethane ternary component system combined with high-performance liquid chromatography with diode-array detection (HPLC–DAD) was developed for the extraction and determination of sulfonamides in solid tissue samples. After the homogenization of the swine muscle with acetonitrile and salt-promoted partitioning, an aliquot of 1 mL of the acetonitrile extract containing a small amount of dichloromethane (250–400 μL) was alkalinized with diethylamine. The clear organic extract obtained by centrifugation was used as a donor phase and then a small amount of water (40–55 μL) could be used as an acceptor phase to back-extract the analytes in the water/acetonitrile/dichloromethane ternary component system. In the back-extraction procedure, after mixing and centrifuging, the sedimented phase would be water and could be withdrawn easily into a microsyringe and directly injected into the HPLC system. Under the optimal conditions, recoveries were determined for swine muscle fortified at 10 ng/g and quantification was achieved by matrix-matched calibration. The calibration curves of five sulfonamides showed linearity with the coefficient of estimation above 0.998. Relative recoveries for the analytes were all from 96.5 to 109.2% with relative standard deviation of 2.7–4.0%. Preconcentration factors ranged from 16.8 to 30.6 for 1 mL of the acetonitrile extract. Limits of detection ranged from 0.2 to 1.0 ng/g.     

Fast and selective determination of triterpenic compounds in olive leaves by liquid chromatography-tandem mass spectrometry with multiple reaction monitoring after microwave-assisted extraction

A method for fast and selective determination of the main triterpenic compounds present in olive leaves — oleanolic, ursolic and maslinic acids as triterpenic acids and, uvaol and erythrodiol as triterpenic dialcohols — is reported here. Quantitative isolation of the analytes has been accomplished in 5 min by microwave assistance using ethanol as extractant. Due to the medium polarity of triterpenic acids and dialcohols, different ethanol-water ratios were tested in order to select the optimum extractant composition for their solubilisation. Microwave assistance provided a significant shortening of the leaching time as compared to conventional procedures by maceration, which usually requires at least 5 h. After extraction, determination was carried out by liquid chromatography-tandem mass spectrometry (LC-MS-MS) with a triple quadrupole (qQq) mass detector without any clean-up step prior to chromatographic analysis. Highly selective identification of triterpenes was confirmed by multiple reaction monitoring (MRM) using the most representative transitions from the precursor ion to the different product ions, while the most sensitive transitions were used for MS-MS quantitation. Total analysis performed in 25 min enables the characterization of a fraction with particular interest in the pharmacological area.