Determination of alkenes in cracking products by normal-phase high-performance liquid chromatography–diode array detection

Alkene content determinations in fluid catalytic cracking (FCC) liquid products were performed by means of normal-phase high-performance liquid chromatography (NP-HPLC) with diode array detection (UV/DAD). Separation of alkenes from aromatic hydrocarbons was performed on amino-modified silica gel column with n-heptane as mobile phase. The column has a little affinity to alkenes and saturated hydrocarbons and a pronounced affinity to aromatic compounds. The problem of alkenes and saturates co-elution on this column type was overcome with the detection system, UV/DAD, sensitive and selective to alkenes, while saturates are inactive in UV field. Total alkene content was determined as a sum of mono- and dialkene groups quantified by external standard method. Validation and verification of the developed method proved their applicability. The following criteria were used to validate the HPLC–DAD method: selectivity, linearity, precision, limits of detection and quantification. Alkene contents were quantified with the external standard method of wide calibration range, so both low and high alkene contents can be determined by the single calibration. Correlation coefficients were higher than 0.99. Precision was evaluated as repeatability and intermediary precision with relative standard deviations less than 5%. Some structural investigation of alkene groups was performed to confirm the assumption. Proposed method was compared with certified NMR method. Six commercial motor gasoline samples were analyzed by these two methods. Obtained results indicate good agreement between alkene content determined by both methods. The developed method was applied to the determination of alkene content in liquid FCC products in the boiling range from 70 °C to 190 °C.     

Determination of migrated phthalic acid residues into edible oils using a green mode of air-assisted liquid–liquid microextraction followed by high-performance liquid chromatography–diode array detector

In this study, for the first time, an organic solvent-free air-assisted liquid–liquid microextraction method has been reported for the extraction and preconcentration of phthalic acids (o-phthalic acid, m-phthalic acid, and p-phthalic acid) from edible oil samples. The method is based on the repeated aspirating/injection of an alkaline aqueous solution and the oil sample mixture in a conical bottom centrifuge tube to form a cloudy solution. After phase separation by centrifuging, the sedimented phase is directly analyzed by high-performance liquid chromatography–diode array detection. Under the optimum extraction conditions, the method showed low limits of detection and quantification between 0.11–0.29 and 0.28–0.91 ng mL^?1, respectively. Extraction recoveries and enrichment factors were from 81 to 97% and 406 to 489, respectively. The relative standard deviations for the analysis of 5 ng mL^?1 of each analyte were less than 5.9% for intraday (n = 6) and interday (n = 5) precisions. Finally, different oil samples were successfully analyzed using the proposed method and m-phthalic acid, and p-phthalic acid were determined in some of them at ng mL^?1 level.     

Determination of triazines in honey by dispersive liquid–liquid microextraction high-performance liquid chromatography

Dispersive liquid–liquid microextraction (DLLME) high-performance liquid chromatography (HPLC) was developed for extraction and determination of triazines from honey. A room temperature ionic liquid, 1-hexyl-3-methylimidazolium hexafluorophosphate [C₆MIM][PF_6.], was used as extraction solvent and Triton X 114 was used as dispersant. A mixture of 175 μL [C₆MIM][PF₆] and 50 μL 10% Triton X 114 was rapidly injected into the 20 mL honey sample by syringe. After extraction, phase separation was performed by centrifugation and the sedimented phase was analyzed by HPLC. Some experimental parameters, such as type and volume of extraction solvent, concentration of dispersant, pH value of sample solution, salt concentration and extraction time were investigated and optimized. The detection limits for chlortoluron, prometon, propazine, linuron and prebane are 6.92, 5.84, 8.55, 8.59 and 5.31 μg kg⁻¹, respectively. The main advantages of the proposed method are simplicity of operation, low cost, high enrichment factor and extraction solvent volume at microliter level. Honey samples were analyzed by the proposed method and obtained results indicated that the proposed method provides acceptable recoveries and precisions.     

Semi-automated alignment and quantification of peaks using parallel factor analysis for comprehensive two-dimensional liquid chromatography–diode array detector data sets

Parallel factor analysis was used to quantify the relative concentrations of peaks within four-way comprehensive two dimensional liquid chromatography–diode array detector data sets. Since parallel factor analysis requires that the retention times of peaks between each injection are reproducible, a semi-automated alignment method was developed that utilizes the spectra of the compounds to independently align the peaks without the need for a reference injection. Peak alignment is achieved by shifting the optimized chromatographic component profiles from a three-way parallel factor analysis model applied to each injection. To ensure accurate shifting, components are matched up based on their spectral signature and the position of the peak in both chromatographic dimensions. The degree of shift, for each peak, is determined by calculating the distance between the median data point of the respective dimension (in either the second or first chromatographic dimension) and the maximum data point of the peak furthest from the median. All peaks that were matched to this peak are then aligned to this common retention data point. Target analyte recoveries for four simulated data sets were within 2% of 100% recovery in all cases. Two different experimental data sets were also evaluated. Precision of quantification of two spectrally similar and partially coeluting peaks present in urine was as good as or better than 4%. Good results were also obtained for a challenging analysis of phenytoin in waste water effluent, where the results of the semi-automated alignment method agreed with the reference LC–LC MS/MS method within the precision of the methods.     

A multi-residue method for pesticide residue analysis in rice grains using matrix solid-phase dispersion extraction and high-performance liquid chromatography–diode array detection

Pesticides are widely used in rice cultivation, often resulting in detection of their residues in rice grains. So far, no analytical method has been available for the simultaneous determination of most rice pesticides in rice grains. This paper reports the development and validation of such a method for the determination of eight rice pesticides (penoxsulam tricyclazole, propanil, azoxystrobin, molinate, profoxydim, cyhalofop-butyl, deltamethrin) and 3,4-dichloroaniline, the main metabolite of propanil. Pesticide extraction and clean-up was performed by an optimized matrix solid-phase dispersion (MSPD) protocol on neutral alumina (5 g) using acetonitrile as the elution solvent. Samples were analyzed in a high-performance liquid chromatography–diode array detection (HPLC-DAD) system. Pesticide separation was achieved with a mobile phase of acetonitrile/water in a linear elution gradient from 30:70% (v/v) to 100:0% (v/v) in 14 min at a flow rate of 0.8 mL min^?1. Method validation was performed by means of linearity, intra-day accuracy, inter-day precision and sensitivity. Linear regression coefficients (R ²) were always above 0.9948. Limits of detection (LOD) and quantification (LOQ) varied from 0.002 to 0.200 mg kg^?1 and 0.006 to 0.600 mg kg^?1, respectively. Recoveries were investigated at three fortification levels and were found to be acceptable (74–127%) with relative standard deviations (RSD) below 12%. Application of the method for the analysis of five commercial rice grain samples showed that the pesticide levels were below the LOD. Overall, the method developed is suitable for the determination of residues of most rice pesticides in rice grains at levels below the established MRLs.     

Isolation of phenolic compounds from hop extracts using polyvinylpolypyrrolidone: Characterization by high-performance liquid chromatography–diode array detection–electrospray tandem mass spectrometry

The aim of the present work was the development of a suitable methodology for the separation and determination of phenolic compounds in the hop plant. The developed methodology was based on the sample purification by adsorption of phenolic compounds from the matrix to polyvinylpolypyrrolidone (PVPP) and subsequent desorption of the adsorbed polyphenols with acetone/water (70:30, v/v). At last, the extract was analyzed by HPLC–DAD and HPLC–ESI-MS/MS. The first phase of this work consisted of the study of the adsorption behavior of several classes of phenolic compounds (e.g. phenolic acids, flavonols, and flavanols) by PVPP in model solutions. It has been observed that the process of adsorption of the different phenolic compounds to PVPP (at low concentrations) is differentiated, depending on the structure of the compound (number of OH groups, aromatic rings, and stereochemistry hindrance). For example, within the phenolic acids class (benzoic, p-hydroxybenzoic, protocatechuic and gallic acids) the PVPP adsorption increases with the number of OH groups of the phenolic compound. On the other hand, the derivatization of OH groups (methylation and glycosylation) resulted in a greatly diminished binding. The use of PVPP revealed to be very efficient for adsorption of several phenolic compounds such as catechin, epicatechin, xanthohumol and quercetin, since high adsorption and recovery values were obtained. The methodology was further applied for the extraction and isolation of phenolic compounds from hops. With this methodology, it was possible to obtain high adsorption values (≥80%) and recovery yield values (≥70%) for the most important phenolic compounds from hops such as xanthohumol, catechin, epicatechin, quercetin and kaempferol glycosides, and in addition it allows the identification of about 30 phenolic compounds by HPLC–DAD and HPLC–ESI-MS/MS.     

Sensitive determination of nitrophenol isomers by reverse-phase high-performance liquid chromatography in conjunction with liquid–liquid extraction

A method for the highly sensitive determination of 2-, 3- and 4-nitrophenols was developed using reverse-phase high-performance liquid chromatography (RP-HPLC) with a UV photodiode array detector. Using a reverse-phase column and 40% aqueous acetonitrile as an eluent (i.e. isocratic elution), the integrated peak area of detector output was linear up to 300 mg/L and the detection limit was 150 µg/L. The sensitivity of this detection method was improved by pretreating the sample solutions with a solvent extraction procedure that makes use of the high partition coefficient of ethyl acetate (EA)/water system. To find an optimum condition for the extraction procedure, this process was simulated by plotting the concentration of nitrophenol extracted in organic solvent against the volume multiplication factor at various partition coefficient of solute. This simulation demonstrated that EA is a superior extractant to other organic solvents. With the newly developed method, the detection limit was extended to 0.3 µg/L. This method offers fast, reliable and more sensitive determination of nitrophenol isomers than any other HPLC method.     

Comparison of two different isolation methods of benzimidazoles and their metabolites in the bovine liver by solid-phase extraction and liquid chromatography–diode array detection

A new analytical method that uses high performance liquid chromatography–diode array detector (HPLC–DAD) was developed for the analysis of 14 benzimidazoles residues, including metabolites, in bovine liver. Samples were extracted using two different extraction procedures: with phosphate buffer after enzymatic hydrolysis (method A) or using organic solvent, i.e. acetonitrile (method B). Then, samples were purified on a strong cation exchange (SCX) cartridge and analyzed in HPLC/DAD. The recovery percentages, obtained spiking the matrix (liver) at concentrations of 500 and 100 μg kg^?1 with a standard mixture of benzimidazoles, were in the range 6–101% and 80–102% for methods A and B, respectively. The repeatability of the methods was assessed in all cases by the % of correlation value (CV) that was lower than 19%. The limits of quantification (LOQs) in the matrix for methods A and B were in the range 40–60 and 20–50 μg kg^?1, respectively. The best of the two methods, method B, was used for the analysis of 10 bovine liver samples.     

Identification of natural dyes used in works of art by pyrolysis–gas chromatography/mass spectrometry combined with in situ trimethylsilylation

Samples of four natural dyes from different organic families—natural madder (anthraquinonoid), curcuma (curcuminoid), saffron (carotenoid) and indigo (indigotic)—were analysed using a new procedure based on pyrolysis–gas chromatography/mass spectrometry (Py–GC/MS), which includes the on-line derivatisation of the natural dyes using hexamethyldisilazane (HMDS). In addition, a previous procedure involving the addition of a 10% H₂SO₄ aqueous solution to the dye and further separation with ethyl acetate has been tested. This procedure enhances the sensitivity of the method by extracting the colouring compounds from the rest of the compounds present in the natural dye. Two possible derivatising reagents—HMDS and tetramethylammonium hydroxide (TMAH)—were compared in order to assess their effectiveness in the proposed method. Characteristic peaks from trimethylsilyl derivatives of alizarin, quinizarin, xanthopurpurin and purpurin were obtained for madder; peaks from safranal, isophorone and trimethylsilyl derivative of crocetin for saffron; peaks from 4-(4-hydroxy-3-methoxy)phenyl-3-buten-2-one and 4-(4-hydroxy-3-methoxy)phenyl-2-butanone, which are primary pyrolysis products of curcuma, and peaks from indole, 2-methylindole and 2,3-dihydroindol-2-one, which are primary pyrolysis products of indigo, among others, were obtained. The reported procedure leads to the unambiguous identification of the four studied dyes from solid samples formed by individual dyes.Electronic Supplementary Material Supplementary material is available for this article at     

Determination of pesticides in water samples from the Wieprz-krzna Canal in the Leczyńsko-Włodawskie Lake District of southeastern Poland by thin-layer chromatography with diode array scanning and high-performance column liquid chromatography with diode array detection

High-performance thin-layer chromatography with diode array scanning (TLC-DAD) and high-performance column liquid chromatography with a diode array detector (HPLC-DAD) were used to screen water samples for pesticides. Atrazine, clofentezine, chlorfenvinphos, hexaflumuron, terbuthylazine, lenacyl, neburon, bitertanol, and metamitron were enriched from canal water samples by solid-phase extraction (SPE) on octadecyl silane (C18)/styrene-divinylbenzene-1, C18, C18 Polar Plus, and cyanopropyl (CN) cartridges. Recovery rates were high for all extraction materials except CN, for which values for all pesticides were lower. SPE was used for both preconcentration and fractionation of the analytes. Analytes were eluted by means of methanol and dichloromethane. Methanol eluates were analyzed by HPLC-DAD and dichloromethane eluates by TLC-DAD. The method was validated for precision, repeatability, and accuracy. Calibration graphs were linear between 0.1 and 50.0 microg/mL for all pesticides, and correlation coefficient (r) values were between 0.9994 and 1.000 as determined by HPLC-DAD. Calibration graphs were linear between 0.1 and 1.5 microg/spot for all pesticides, and r values were between 0.9899 and 0.9987 as determined by TLC-DAD. The limit of detection was between 0.04 and 0.23 microg/spot for TLC-DAD and 0.02 and 0.45 microg/mL for HPLC-DAD.     

Application of β-cyclodextrin for the analysis of estrogenic steroids in human urine by high-performance liquid chromatography

Summary A procedure is described for simultaneous determination of estriol, estrone and 17-estradiol in human urine. After acid hydrolysis of the sulphate conjugates, the estrogens were extracted into diethyl ether. The ether extracts were concentrated and applied directly to an HPLC column. Using a 25 cm C-18 column and acetonitrile-water modified by the addition of -cyclodextrin as mobile phase, the separation of estriol, estradiol and estrone was achieved within 20 minutes. The extraction of estrogens from the biological matrix is excellent. Estrogens were detected using a UV-detector (280 nm) or a spectrofluorimetric detector (_exc=280nm, _em=312 nm). The latter detection system has been used for the determination of estrogens in the urine of non-pregnant women. The procedure is simple and can be used in clinical practice.     

Determination of ochratoxin A in pig tissues by liquid–liquid extraction and clean-up and high-performance liquid chromatography

A fast, simple, and sensitive HPLC–FD method is described for determination of ochratoxin A (OTA) in pig kidney and muscle; a small mass (<2.5 g) of sample and a relatively small volume (<15 mL) of a non-halogenated extraction solvent are required. Ochratoxin B, systematically absent from all the samples investigated, was used as internal standard. Liquid–liquid partition was used for sample clean-up. Recoveries at the 1 ng g^–1 level were 86±15% and 74±8% for kidney and muscle, respectively, and detection limits were 0.14 and 0.15 ng g^–1. Clean-up by solid-phase extraction (SPE) is required for pig liver. A survey of the OTA content of tissues of pigs slaughtered in southern Italy revealed that 52 out of 54 analysed samples were contaminated; the OTA concentration in kidney ranged between 0.26 and 3.05 ng g^–1. The effect of measurement precision on compliance with legal limits is also discussed.     

Dispersive liquid–liquid microextraction followed by high-performance liquid chromatography for the determination of three carbamate pesticides in water samples

A simple, rapid and efficient method, dispersive liquid–liquid microextraction (DLLME) in conjunction with high-performance liquid chromatography (HPLC), has been developed for the determination of three carbamate pesticides (methomyl, carbofuran and carbaryl) in water samples. In this extraction process, a mixture of 35 µL chlorobenzene (extraction solvent) and 1.0 mL acetonitrile (disperser solvent) was rapidly injected into the 5.0 mL aqueous sample containing the analytes. After centrifuging (5 min at 4000 rpm), the fine droplets of chlorobenzene were sedimented in the bottom of the conical test tube. Sedimented phase (20 µL) was injected into the HPLC for analysis. Some important parameters, such as kind and volume of extraction and disperser solvent, extraction time and salt addition were investigated and optimised. Under the optimum extraction condition, the enrichment factors and extraction recoveries ranged from 148% to 189% and 74.2% to 94.4%, respectively. The methods yielded a linear range in the concentration from 1 to 1000 µg L^?1 for carbofuran and carbaryl, 5 to 1000 µg L^?1 for methomyl, and the limits of detection were 0.5, 0.9 and 0.1 µg L^?1, respectively. The relative standard deviations (RSD) for the extraction of 500 µg L^?1 carbamate pesticides were in the range of 1.8–4.6% (n = 6). This method could be successfully applied for the determination of carbamate pesticides in tap water, river water and rain water.     

Enantiomeric analysis of drugs in water samples by using liquid–liquid microextraction and nano-liquid chromatography

The nano-LC technique is increasingly used for both fast studies on enantiomeric analysis and test beds of novel stationary phases due to the small volumes involved and the short conditioning and analysis times. In this study, the enantioseparation of 10 drugs from different families was carried out by nano-LC, utilizing silica with immobilized amylose tris(3-chloro-5-methylphenylcarbamate) column. The effect on chiral separation caused by the addition of different salts to the mobile phase was evaluated. To simultaneously separate as many enantiomers as possible, the effect of buffer concentration in the mobile phase was studied, and, to increase the sensitivity, a liquid–liquid microextraction based on the use of isoamyl acetate as sustainable extraction solvent was applied to pre-concentrate four chiral drugs from tap and environmental waters, achieving satisfactory recoveries (>70%).     

Dispersive solid-phase extraction followed by dispersive liquid–liquid microextraction for the determination of some sulfonylurea herbicides in soil by high-performance liquid chromatography

Dispersive solid-phase extraction (DSPE) combined with dispersive liquid–liquid microextraction (DLLME) has been developed as a new approach for the extraction of four sulfonylurea herbicides (metsulfuron-methyl, chlorsulfuron, bensulfuron-methyl and chlorimuron-ethyl) in soil prior to high-performance liquid chromatography with diode array detection (HPLC-DAD). In the DSPE-DLLME, sulfonylurea herbicides were first extracted from soil sample into acetone–0.15 mol L⁻¹ NaHCO₃ (2:8, v/v). The clean-up of the extract by DSPE was carried out by directly adding C₁₈ sorbent into the extract solution, followed by shaking and filtration. After the pH of the filtrate was adjusted to 2.0 with 2 mol L⁻¹ HCl, 60.0 μL chlorobenzene (as extraction solvent) was added into 5.0 mL of it for DLLME procedure (the acetone contained in the solution also acted as dispersive solvent). Under the optimum conditions, the enrichment factors for the compounds were in the range between 102 and 216. The linearity of the method was in the range from 5.0 to 200 ng g⁻¹ with the correlation coefficients (r) ranging from 0.9967 to 0.9987. The method detection limits were 0.5–1.2 ng g⁻¹. The relative standard deviations varied from 5.2% to 7.2% (n = 5). The relative recoveries of the four sulfonylurea herbicides from soil samples at spiking levels of 6.0, 20.0 and 60.0 ng g⁻¹ were in the range between 76.3% and 92.5%. The proposed method has been successfully applied to the analysis of the four target sulfonylurea herbicides in soil samples, and a satisfactory result was obtained.     

Residue analysis of glucocorticoids in bovine milk by liquid chromatography–tandem mass spectrometry

A sensitive liquid chromatography–tandem mass spectrometry (LC-MS/MS) method for the simultaneous determination of 13 steroidal anti-inflammatory drugs in bovine milk is presented. Due to their weakly acid nature, analytes were separated by ion suppression reversed phase chromatography and detected in positive-ion mode by a high flow electrospray source. Dexamethasone-d4 was used as internal standard. The sample preparation was simple and reliable; it included acidic deproteinization of milk followed by sample enrichment and clean-up, utilizing a C18 solid phase extraction cartridge. Recoveries exceeded 70% with an intra-day precision not larger than 12%. The efficiency of the sample clean-up and internal standardization rendered negligible the matrix effect, estimated by comparing standard and matrix-matched calibration curves. A small-scale reconnaissance was carried out on several raw and whole fresh milk samples. A large number of analyzed samples showed a chromatographic peak, in the retention time window of cortisol, at levels included between its decision limit (CCα) and detection capability (CCβ). As a result of a heat-induced transformation, an isomeric product of triamcinolone was observed during the extract evaporation. Since this rearrangement might occur during the milk pasteurization process, LC-MS/MS and ¹H-NMR investigations were performed out to conclusively differentiate the two isomers. One- and two-dimensional proton NMR spectra were able to identify the transformation product as 9a-fluoro-11b,16a-trihydroxy-17b-hydroxymethyl-D-homoandrosta-1,4-diene-3,17a-dione.     

Determination of organophosphorus pesticides in environmental water samples by dispersive liquid–liquid microextraction with solidification of floating organic droplet followed by high-performance liquid chromatography

A simple dispersive liquid–liquid microextraction based on solidification of floating organic droplet coupled with high-performance liquid chromatography–diode array detection was developed for the determination of five organophosphorus pesticides (OPs) in water samples. In this method, the extraction solvent used is of low density, low toxicity, and proper melting point near room temperature. The extractant droplet could be collected easily by solidifying it in the lower temperature. Some important experimental parameters that affect the extraction efficiencies were optimized. Under the optimum conditions, the calibration curve was linear in the concentration range from 1 to 200 ng mL⁻¹ for the five OPs (triazophos, parathion, diazinon, phoxim, and parathion-methyl), with the correlation coefficients (r) varying from 0.9991 to 0.9998. High enrichment factors were achieved ranging from 215 to 557. The limits of detection were in the range between 0.1 and 0.3 ng mL⁻¹. The recoveries of the target analytes from water samples at spiking levels of 5.0 and 50.0 ng mL⁻¹ were 82.2–98.8% and 83.6–104.0%, respectively. The relative standard deviations fell in the range of 4.4% to 6.3%. The method was suitable for the determination of the OPs in real water samples.