Advanced nanofibrous sorbents for the extraction of pollutants from river water and protein-containing matrices.

The extraction efficiencies of thirty types of fibers produced by meltblown, alternating current electrospinning, and meltblown-co-electrospinning technologies were tested as advanced sorbents for on-line solid-phase extraction in a high-performance liquid chromatography system have been tested and compared with a commercial C18 sorbent. The properties of each fiber, which were often depended on the production process, and their applicability were demonstrated with the extraction of the model analytes nitrophenols and chlorophenols from various matrices including river water and to purify complex matrix human serum and bovine serum albumin from macromolecular ballast. Polycaprolactone fibers outperformed other polymers and were selected for subsequent modifications including (i) incorporation of hybrid carbon nanoparticles, i.e., graphene, activated carbon, and carbon black into the polymer prior to fiber fabrication, and (ii) surface modification by dip coating with polyhydroxy modifiers including graphene oxide, tannin, dopamine, hesperidin, and heparin. These novel fibrous sorbents were comparable to commercial C18 sorbent and provided excellent analyte recoveries of 70-112% even from the protein-containing matrices.

Effect of P-glycoprotein and Cotreatment with Sofosbuvir on the Intestinal Permeation of Tenofovir Disoproxil Fumarate and Tenofovir Alafenamide Fumarate.

PURPOSE: We aimed to compare the effects of P-glycoprotein (ABCB1) on the intestinal uptake of tenofovir disoproxil fumarate (TDF), tenofovir alafenamide fumarate (TAF), and metabolites, tenofovir isoproxil monoester (TEM) and tenofovir (TFV), and to study the molecular mechanism of drug-drug interaction (DDI) between sofosbuvir (SOF) and TDF/TAF. METHODS: Bidirectional transport experiments in Caco-2 cells and accumulation studies in precision-cut intestinal slices prepared from the ileal segment of rodent (rPCIS) and human (hPCIS) intestines were performed. RESULTS: TDF and TAF were extensively metabolised but TAF exhibited greater stability. ABCB1 significantly reduced the intestinal transepithelial transfer and uptake of the TFV(TDF) and TFV(TAF)-equivalents. However, TDF and TAF were absorbed more efficiently than TFV and TEM. SOF did not inhibit intestinal efflux of TDF and TAF or affect intestinal accumulation of TFV(TDF) and TFV(TAF)-equivalents but did significantly increase the proportion of absorbed TDF. CONCLUSIONS: TDF and TAF likely produce comparable concentrations of TFV-equivalents in the portal vein and the extent of permeation is reduced by the activity of ABCB1. DDI on ABCB1 can thus potentially affect TDF and TAF absorption. SOF does not inhibit ABCB1-mediated transport of TDF and TAF but does stabilise TDF, albeit without affecting the quantity of TFV(TDF)-equivalents crossing the intestinal barrier. Our data thus suggest that reported increases in the TFV plasma concentrations in patients treated with SOF and TDF result either from a DDI between SOF and TDF that does not involve ABCB1 or from a DDI involving another drug used in combination therapy.

Small nanofibrous disks for preconcentration of environmental contaminants followed by direct in-vial elution and chromatographic determination.

A novel method for the extraction of river water contaminants as model analytes of ranging polarities, including bisphenols A, C, S, Z, fenoxycarb, kadethrin, and deltamethrin, using small compact fibrous disks has been developed and validated. Polymer nanofibers and microfibers prepared from poly(3-hydroxybutyrate), polypropylene, polyurethane, polyacrylonitrile, poly(lactic acid), and polycaprolactone doped with graphene were evaluated in terms of extraction efficiency, selectivity, and stability in organic solutions. Our novel extraction procedure comprised preconcentration of analytes from 150 mL river water to 1 mL of eluent using a compact nanofibrous disk freely vortexed in the sample. Small nanofibrous disks with a diameter of 10 mm were cut from a compact and mechanically stable 1-2 mm thick micro/nanofibrous sheet. After 60 min extraction in a magnetically stirred sample located in a beaker, the disk was removed from the liquid and washed with water. Then, the disk was inserted into a 1.5 mL HPLC vial and extracted with 1.0 ml methanol upon short intensive shaking. Our approach avoided the undesired problems related to the manual handling typical of "classical" SPE procedure since the extraction was carried out directly in the HPLC vial. No sample evaporation, reconstitution, or pipetting was required. The nanofibrous disk is affordable, needs no support or holder, and its use avoids creation of plastic waste originating from disposable materials. Recovery of compounds from the disks was 47.2-141.4% depending on the type of polymer used and the relative standard deviations calculated from 5 extractions ranged from 6.1 to 11.8% for poly(3-hydroxybutyrate), 6.3-14.8% for polyurethane, and 1.7-16.2% for polycaprolactone doped with graphene. A small enrichment factor was obtained for polar bisphenol S using all sorbents. A higher preconcentration reaching up to 40-fold was achieved for lipophilic compounds such as deltamethrin when using poly(3-hydroxybutyrate) and graphene-doped polycaprolactone.

Comparing adsorption performance of microfibers and nanofibers with commercial molecularly imprinted polymers and restricted access media for extraction of bisphenols from milk coupled with liquid chromatography.

Advanced solid phase extraction (SPE) fibrous sorbents including polyethylene, polypropylene poly (hydroxybutyrate), and polyamide 6 nanofibers, polycaprolactone microfibers/nanofibers, polycaprolactone microfibers/polyvinylidene difluoride nanofibers, and poly (hydroxybutyrate) microfibers/polypropylene microfibers composites, as well as commercial molecularly imprinted polymers and restricted access media sorbent were compared in terms of bisphenols extraction from milk and their clean-up efficiency. Three on-line SPE-HPLC methods were completely validated for the extraction and detection of bisphenols A, AF, C, A diglycidyl ether, and F diglycidyl ether in bovine milk. Polycaprolactone composite nanofibers compared favorably to restricted access media, enabled excellent clean-up of bisphenols from the proteinaceous matrix, and yielded recoveries 98.0-124.5% and 93.0-115.0%, respectively, with RSD less than 10%. Total analysis time including on-line SPE step lasted only 12 min, which represents a significant reduction in time compared with previously reported as well as official European Union and AOAC methods defined for the determination of bisphenols in various matrices.

Evaluation of Anthocyanin Profiles in Various Blackcurrant Cultivars over a Three-Year Period Using a Fast HPLC-DAD Method.

Anthocyanins are the most important polyphenolic substances contained in blackcurrant fruits. They are responsible for the various health benefits caused, in particular, by their high antioxidant activity. Anthocyanins derived from anthocyanidins cyanidin and delphinidin are typical for blackcurrant fruits, especially their rutinoside and glucoside forms. These four anthocyanins usually represent about 97-98% of total anthocyanins in blackcurrant fruits. In this study, we developed and validated a new HPLC-DAD method for rapid anthocyanin separation and determination in fifteen perspective blackcurrant cultivars ('Ruben', 'Ben Lomond', 'Ben Conan', 'Ceres', 'Moravia', 'Ometa', 'Lota', 'Fokus', 'Tenah', 'Sejanec', 'Consort', 'Triton', 'Ben Hope', 'Ben Gairn', and one gooseberry hybrid 'Josta'). Eight of them were monitored throughout the three-year experiment. The most represented anthocyanins in all monitored blackcurrant cultivars were delphinidin-3-rutinoside (36.7-63.6%), cyanidin-3-rutinoside (26.4-40.6%), delphinidin-3-glucoside (6.1-17.9%), and cyanidin-3-glucoside (1.3-9.9%). The individual anthocyanin proportion (%) in each cultivar was specific, and a similar profile was verified in a three-year period for eight available cultivars. Total anthocyanin content expressed as a sum of four major anthocyanins present in blackcurrants was compared with values expressed as the equivalent of cyanidin-3-glucoside, as many authors do. We revealed an underestimation of about 20% with the latter method. Cultivars with the highest average total anthocyanin content were 'Ben Gairn' (294.38 mg/100 g), 'Ceres' (281.31 mg/100 g), and 'Ometa' (269.09 mg/100 g).

Determination of Antiviral Drugs and Their Metabolites Using Micro-Solid Phase Extraction and UHPLC-MS/MS in Reversed-Phase and Hydrophilic Interaction Chromatography Modes.

Two new ultra-high performance liquid chromatography (UHPLC) methods for analyzing 21 selected antivirals and their metabolites were optimized, including sample preparation step, LC separation conditions, and tandem mass spectrometry detection. Micro-solid phase extraction in pipette tips was used to extract antivirals from the biological material of Hanks balanced salt medium of pH 7.4 and 6.5. These media were used in experiments to evaluate the membrane transport of antiviral drugs. Challenging diversity of physicochemical properties was overcome using combined sorbent composed of C18 and ion exchange moiety, which finally allowed to cover the whole range of tested antivirals. For separation, reversed-phase (RP) chromatography and hydrophilic interaction liquid chromatography (HILIC), were optimized using extensive screening of stationary and mobile phase combinations. Optimized RP-UHPLC separation was carried out using BEH Shield RP18 stationary phase and gradient elution with 25 mmol/L formic acid in acetonitrile and in water. HILIC separation was accomplished with a Cortecs HILIC column and gradient elution with 25 mmol/L ammonium formate pH 3 and acetonitrile. Tandem mass spectrometry (MS/MS) conditions were optimized in both chromatographic modes, but obtained results revealed only a little difference in parameters of capillary voltage and cone voltage. While RP-UHPLC-MS/MS exhibited superior separation selectivity, HILIC-UHPLC-MS/MS has shown substantially higher sensitivity of two orders of magnitude for many compounds. Method validation results indicated that HILIC mode was more suitable for multianalyte methods. Despite better separation selectivity achieved in RP-UHPLC-MS/MS, the matrix effects were noticed while using both chromatographic modes leading to signal enhancement in RP and signal suppression in HILIC.

Determination of Ochratoxin A and Ochratoxin B in Archived Tokaj Wines (Vintage 1959-2017) Using On-Line Solid Phase Extraction Coupled to Liquid Chromatography.

According to the EU legislation, ochratoxin A contamination is controlled in wines. Tokaj wine is a special type of sweet wine produced from botrytized grapes infected by "noble rot" Botrytis cinerea. Although a high contamination was reported in sweet wines and noble rot grapes could be susceptible to coinfection with other fungi, including ochratoxigenic species, no screening of Tokaj wines for mycotoxin contamination has been carried out so far. Therefore, we developed an analytical method for the determination of ochratoxin A (OTA) and ochratoxin B (OTB) involving online SPE coupled to HPLC-FD using column switching to achieve the fast and sensitive control of mycotoxin contamination. The method was validated with recoveries ranging from 91.6% to 99.1% with an RSD less than 2%. The limits of quantification were 0.1 and 0.2 µg L-1 for OTA and OTB, respectively. The total analysis time of the online SPE-HPLC-FD method was a mere 6 min. This high throughput enables routine analysis. Finally, we carried out an extensive investigation of the ochratoxin contamination in 59 Slovak Tokaj wines of 1959-2017 vintage. Only a few positives were detected. The OTA content in most of the checked wines did not exceed the EU maximum tolerable limit of 2 µg L-1, indicating a good quality of winegrowing and storing.

A first approach using micellar electrokinetic capillary chromatography for the determination of fipronil and fipronil-sulfone in eggs.

Fipronil is an insecticide that is not approved in the European Union in food. In 2017, fipronil was involved in a European health alert due to its presence in fresh hen eggs because of an illicit use in poultry farms, so reliable methods are needed to determine fipronil and its main metabolites in these matrixes. In this work, we report the first approach to the study of fipronil and two metabolites, fipronil-sulfone and fipronil-sulfide by CE. MEKC mode was employed using a solution of 50 mM ammonium perfluorooctanoate pH 9.0 with 10% (v/v) methanol as background electrolyte. The proposed method was combined with a simple sample treatment based on salting-out assisted LLE (SALLE) using acetonitrile as extraction solvent and ammonium sulfate as salt. The SALLE-MEKC-UV method allowed the simultaneous quantification of fipronil and fipronil-sulfone. Validation parameters yielded satisfactory results, with precision, expressed as relative SD, below 14% and recoveries higher than 83%. Limits of detection were 90 µg/kg for fipronil and 150 µg/kg for fipronil-sulfone, so in terms of sensitivity further studies of sample treatments allowing extra preconcentration or the use of more sensitive detection, such as MS, would be needed.

Preparation of citrinin-selective molecularly imprinted polymer and its use for on-line solid-phase extraction coupled to liquid chromatography.

A new selective molecularly imprinted polymer has been prepared and used for extraction in on-line SPE-HPLC to achieve the selective determination of citrinin. Four different imprinted polymers varying in combinations of components were prepared by bulk polymerization and evaluated in terms of binding capacity and selectivity. Imprinted polymer prepared from a mixture comprising 1-hydoxy-2-naphtoic acid as the template molecule, acrylamide as the structural monomer, ethylene dimethacrylate as the cross-linker (in a molar ratio of 1:4:16), and acetonitrile as the porogenic solvent exhibited the best properties. The selectivity of this sorbent was confirmed by comparison with the non-imprinted counterpart prepared using the same polymerization carried out in the absence of template. Imprinted polymer was packed in a 20 × 3 mm i.d. steel cartridge and coupled to the on-line SPE-HPLC system through a six-port switching valve. The method for determination of citrinin including the on-line extraction step was then developed and validated. The sample in the form of methanolic extract was loaded, cleaned, and preconcentrated in the imprinted SPE cartridge. Subsequent separation of citrinin from residual interferences was achieved using the analytical column Kinetex Biphenyl 100 × 4.6 mm i.d., 5 µm particle size, and fluorescence detection (Ex 335, Em 500 nm). The total analysis time was only 9.50 min. Our fully validated method was also applied to analysis of food supplements based on red yeast rice extracts, the control of which is implemented in European legislation. Only minor yet acceptable contamination was found in tested samples.

A Comparative Study of Advanced Stationary Phases for Fast Liquid Chromatography Separation of Synthetic Food Colorants.

Food analysis demands fast methods for routine control and high throughput of samples. Chromatographic separation enables simultaneous determination of numerous compounds in complex matrices, several approaches increasing separation efficiency and speed of analysis were involved. In this work, modern types of column with monolithic rod or superficially porous particles were employed and compared for determination of eight synthetic food dyes, their chromatographic performance was evaluated. During method optimization, cyano stationary phase Chromolith Performance CN 100 × 4.6 mm and Ascentis Express ES-CN 100 × 4.6 mm, 5 µm were selected for the separation of polar colorants. The separation was performed by gradient elution of acetonitrile/methanol and 2% water solution of ammonium acetate at flow rate 2.0 mL min-1. Mobile phase composition and the gradients were optimized in order to enable efficient separation on both columns. The method using fused-core particle column provided higher separation efficiency, narrow peaks of analytes resulted in increased peak capacity and shortening of analysis time. After the validation, the method was applied for analysis of coloured beers, soft drinks and candies.

Molecularly imprinted vs. reversed-phase extraction for the determination of zearalenone: a method development and critical comparison of sample clean-up efficiency achieved in an on-line coupled SPE chromatography system.

Sample preparation prior to chromatographic separation plays an important role in the analytical process. To avoid time-consuming and manual handling sample-prep, automated on-line techniques such as on-line SPE-HPLC are therefore preferred. In this study, two different on-line extraction approaches for mycotoxin/endocrine disruptor zearalenone (ZEA) determination using either molecularly imprinted polymer (MIP) with selective cavities and binding sites for extraction or a reversed-phase sorbent C18 providing non-selective interactions have been developed, validated, and compared. The validation characteristics were compared and the two methods were evaluated as being almost equal in terms of linearity, repeatability, precision, and recovery. Recoveries were in the range of 99.0-100.1% and limits of detection were found the same for both methods (1.5 µg L-1). Method precision calculated for spiked beer samples was better for C18 sorbent (2.5 vs. 5.4% RSD). No significant differences in the selectivity of either extraction method were observed. The possible reasons and further details associated with this finding are discussed. Finally, both validated methods were applied for the determination of ZEA contamination in beer samples. Due to ZEA's native fluorescence, chromatographic separation with fluorimetric detection (λex = 270 nm and λem, = 458 nm) was selected. Graphical abstract Determination of zearalenone in beer using an on-line extraction chromatography system.

Critical comparison of the on-line and off-line molecularly imprinted solid-phase extraction of patulin coupled with liquid chromatography.

Reaching trace amounts of mycotoxin contamination requires sensitive and selective analytical tools for their determination. Improving the selectivity of sample pretreatment steps covering new and modern extraction techniques is one way to achieve it. Molecularly imprinted polymers as selective sorbent for extraction undoubtedly meet these criteria. The presented work is focused on the hyphenation of on-line molecularly imprinted solid-phase extraction with a chromatography system using a column-switching approach. Making a critical comparison with a simultaneously developed off-line extraction procedure, evaluation of pros and cons of each method, and determining the reliability of both methods on a real sample analysis were carried out. Both high-performance liquid chromatography methods, using off-line extraction on molecularly imprinted polymer and an on-line column-switching approach, were validated, and the validation results were compared against each other. Although automation leads to significant time savings, fewer human errors, and required no handling of toxic solvents, it reached worse detection limits (15 versus 6 µg/L), worse recovery values (68.3-123.5 versus 81.2-109.9%), and worse efficiency throughout the entire clean-up process in comparison with the off-line extraction method. The difficulties encountered, the compromises made during the optimization of on-line coupling and their critical evaluation are presented in detail.

On-line coupling of molecularly imprinted solid phase extraction with liquid chromatography for the fast determination of coumarins from complex samples.

In this work, an on-line SPE-HPLC method with spectrophotometric detection was developed for the determination of coumarins in complex samples. For the on-line cleanup of samples, a molecularly imprinted polymer was packed into the column cartridge and coupled directly with HPLC (MISPE-HPLC) using a column switching system. The separation of coumarins was performed on a C18 core-shell column (100×4.6mm, 5µm) with a mobile phase consisting of 0.3% acetic acid/acetonitrile with gradient elution at a flow-rate of 1mLmin-1. The total time of the whole analytical run, including the extraction step, was 13.25min. The on-line MISPE-HPLC method was optimized and validated. The results showed good linearity (0.10-100µgmL-1) with correlation coefficients higher than 0.99. The LOD values were from 0.03 to 0.15µgmL-1. The proposed method was successfully applied for analysis of real samples (Cassia cinnamon, chamomile tea, and Tokaj specialty wines) and obtained recoveries varied from 78.7% to 112.2% with an RSD less than 9%.

Determination of green, blue and yellow artificial food colorants and their abuse in herb-coloured green Easter beers on tap.

Beer is one of the most popular alcoholic beverages worldwide. For consumer acceptance, significant factors are its taste, flavour and colour. This study determines selected synthetic green, blue and yellow food colorants in popular Easter herb-coloured green beers on tap produced in breweries on Holy Thursday. The abuse of beer colouring with Tartrazine (E 102), Quinoline yellow (E 104), Sunset yellow (E 110), Patent blue (E 131), Indigo carmine (E 132), Brilliant blue FCF (E 133), Green S (E 142) and Fast green FCF (E 143) was assessed in 11 green beer samples purchased in local restaurants. HPLC was used for the separation and detection of artificial colorants with diode-array detection and a Chromolith Performance CN 100 × 4.6 mm column with guard pre-column Chromolith CN 5 × 4.6 mm. Separation was performed in gradient elution with mobile phase containing methanol-aqueous 2% ammonium acetate at pH 7.0. The study showed that eight beers (70%) marketed in the Czech Republic contained artificial colorants (Tartrazine and Brilliant blue FCF). The concentration of colorants found in analysed green herb-coloured beers ranged from 1.58 to 3.49 mg l(-)(1) for Tartrazine, 0.45-2.18 mg l(-)(1) for Brilliant blue, while Indigo carmine was detected only once at concentration 2.36 mg l(-)(1). Only three beers showed no addition of the synthetic colorants. However, the levels of artificial colorants found in beers marketed in the Czech region were very low and did not show a serious risk for consumers' health.

A fully automated and fast method using direct sample injection combined with fused-core column on-line SPE-HPLC for determination of ochratoxin A and citrinin in lager beers.

A new fast and sensitive method based on on-line solid-phase extraction on a fused-core precolumn coupled to liquid chromatography with fluorescence detection has been developed for ochratoxin A (OTA) and citrinin (CIT) determination in lager beer samples. Direct injection of 100 µL filtered beer samples into an on-line SPE-HPLC system enabled fast and effective sample extraction including separation in less than 6 min. Preconcentration of OTA and CIT from beer samples was performed on an Ascentis Express RP C18 guard column (5 × 4.6 mm), particle size 2.7 µm, with a mobile phase of methanol/0.5% aqueous acetic acid pH 2.8 (30:70, v/v) at a flow rate of 2.0 mL min(-1). The flow switch from extraction column to analytical column in back-flush mode was set at 2.0 min and the separation was performed on the fused-core column Ascentis Express Phenyl-Hexyl (100 × 4.6 mm), particle size 2.7 µm, with a mobile phase acetonitrile/0.5% aqueous acetic acid pH 2.8 in a gradient elution at a flow rate of 1.0 mL min(-1) and temperature of 50 °C. Fluorescence excitation/emission detection wavelengths were set at 335/497 nm. The accuracy of the method, defined as the mean recoveries of OTA and CIT from light and dark beer samples, was in the range 98.3-102.1%. The method showed high sensitivity owing to on-line preconcentration; LOQ values were found to be 10 and 20 ng L(-1) for OTA and CIT, respectively. The found values of OTA and CIT in all tested light, dark and wheat beer samples were significantly below the maximum tolerable limits (3.0 µg kg(-1) for OTA and 2000 µg kg(-1) for CIT) set by the European Union.