Snail1 controls TGF-ß responsiveness and differentiation of mesenchymal stem cells.

The Snail1 transcriptional repressor plays a key role in triggering epithelial-to-mesenchymal transition. Although Snail1 is widely expressed in early development, in adult animals it is limited to a subset of mesenchymal cells where it has a largely unknown function. Using a mouse model with inducible depletion of Snail1, here we demonstrate that Snail1 is required to maintain mesenchymal stem cells (MSCs). This effect is associated to the responsiveness to transforming growth factor (TGF)-ß1 that shows a strong Snail1 dependence. Snail1 depletion in conditional knockout adult animals causes a significant decrease in the number of bone marrow-derived MSCs. In culture, Snail1-deficient MSCs prematurely differentiate to osteoblasts or adipocytes and, in contrast to controls, are resistant to the TGF-ß1-induced differentiation block. These results demonstrate a new role for Snail1 in TGF-ß response and MSC maintenance.

Critical review on recent developments in solventless techniques for extraction of analytes.

The most recent contributions on solventless extraction techniques have been reviewed. This paper deals with those techniques that use solid phases, such as solid-phase microextraction, liquid phases, such as single-drop microextraction and hollow-fibre liquid-phase microextraction, and subcritical fluids, such as subcritical water extraction. In all cases, the most recent publications have been critically studied. Direct extraction and derivatization processes to facilitate the extraction of analytes in different areas have been included. Hyphenated approaches, if available, are also included in this review. Comparison of techniques organized by analytes and matrices also enhances this critical overview of solventless techniques.

Efficiency of whole and skimmed powdered milk for trapping volatile compounds released from plastic containers in high-temperature applications.

Plastic food containers used for high-temperature applications are not completely inert, and potentially harmful chemicals may be transferred to foodstuffs when such containers are heated. The aim of this work was to investigate the role of food fat content on the efficiency of trapping volatile organic compounds from heated plastic packaging. Relatively simple food matrices such as powdered skimmed and whole milk were evaluated with respect to their retention of several selected migrants: toluene, 1-octene, ethylbenzene, o-, m-, and p-xylene, styrene, and 1,4-dichlorobenzene released from containers made of polypropylene (random and copolymer), polycarbonate, and styrene-acrylonitrile copolymer, which are all commonly used in high-temperature applications. The analytical method (purge and trap gas chromatography and mass spectrometry) was optimized for each matrix. The developed procedure had detection limits of 0.01 to 1.2 ng, depending on the analyte and sample matrix, and both reproducibility and repeatability (expressed as relative standard deviation) were below 15%. This method was applied to the different plastic materials. The concentrations of the volatile compounds in both matrices were well below the established specific migration limits. Temperature and fat content of powdered milk were the most influential variables in mass transfer processes. These values were compared with those obtained with either Tenax TA (alternative test medium for fatty food simulants) or Porapak Q (another widely used sorbent). Similar results were found in skimmed powdered milk and Tenax TA, but significant differences were observed for whole powdered milk.

New cinnamon-based active paper packaging against Rhizopusstolonifer food spoilage.

A new active paper package based on the incorporation of cinnamon essential oil to solid wax paraffin as an active coating is proposed, developed, and evaluated. The antifungal activity of the active paper is tested against Rhizopusstolonifer, and the results demonstrate that 6% (w/w) of the essential oil in the active coating formulation completely inhibits the growth of R. stolonifer, whereas 4% still has strong antimicrobial activity in in vitro conditions. Then, active paper is evaluated with actual food, sliced bread, using different storage times. After 3 days of storage, almost complete inhibition is obtained with 6% cinnamon essential oil. Qualitative analysis by solid-phase microextraction and determination of cinnamaldehyde in the sliced bread were also performed and confirmed the strong correspondence between the inhibition of the mold and the amount of cinnamaldehyde in the bread.

Determination of bile acids in human serum by on-line restricted access material-ultra high-performance liquid chromatography-mass spectrometry.

This paper describes a new, fully automated on-line method combining restricted access material (RAM) extraction and ultra high-performance liquid chromatography (UHPLC) with mass spectrometric (MS) detection for determining congeners of bile acids (BAs) in human serum. In this method, low-pressure RAM and high-pressure UHPLC-MS are hyphenated by using a 2.5-mL loop-type interface. The compatibility problem between the large volume (1.2mL) of strong solvent (methanol) used for RAM elution and the need for a weak solvent in UHPLC injection has been addressed by using an auxiliary pre-column cross-flow of 0.1% aqueous formic acid. In this way, the complete 2.5mL loop volume can be injected into the UHPLC system, thereby maximizing sensitivity while maintaining good chromatographic performance. The optimised method allows the simultaneous analysis of 13 bile acids in a single run, including glycine- and taurine-conjugated bile acids, cholic acid (CA), deoxycholic acid (DCA), chenodeoxycholic acid (CDCA), ursodeoxycholic acid (UDCA), and litocholic acid. The complete analysis of a 100-microL single serum sample is performed in 30 min, providing detection limits in the pg range (corresponding with clinically relevant concentration levels) for all of the analytes except lithocholic acid, intra-day precision values (%R.S.D.) below 4% (except ursodeoxycholic acid) and inter-day precision lower than 15% (except ursodeoxycholic, glycoursodeoxycholic acid (GUDCA) and lithocholic acid).

Direct determination of carnosic acid in a new active packaging based on natural extract of rosemary.

A new antioxidant film is being developed that incorporates a natural extract of rosemary and is intended for contact with food. The rosemary extract has been screened and carnosic acid and carnosol have been determined as the major antioxidant components (6.96% and 0.88%, respectively) that are responsible for the antioxidant properties of the whole extract. Thus, a fast method for the direct determination of carnosic acid in the packaging material, in order to evaluate the antioxidant capacity of the new active plastic, has been developed and optimized. The method consists of extraction from the plastic with methanol, followed by anion exchange solid-phase extraction and final analysis by UPLC-MS. Using this process, the recovery of carnosic acid is about 99%. The complete analytical performance of the method developed here is also assessed. The analytical features of the method, such as the relative standard deviation, reproducibility, repeatability, linear range, and detection and quantification limits, are shown. This method can be subsequently modified to monitor other active components in different packages, and it constitutes a crucial step forward in research into new and improved commercial antioxidant packages.

Development of flexible antimicrobial films using essential oils as active agents.

The antimicrobial activity in the vapor-phase of laboratory-made flexible films of polypropylene (PP) and polyethylene/ethylene vinyl alcohol copolymer (PE/EVOH) incorporating essential oil of cinnamon ( Cinnamomum zeylanicum), oregano ( Origanum vulgare), clove ( Syzygium aromaticum), or cinnamon fortified with cinnamaldehyde was evaluated against a wide range of microorganisms: the Gram-negative bacteria Escherichia coli, Yersinia enterocolitica, Pseudomonas aeruginosa, and Salmonella choleraesuis; the Gram-positive bacteria Listeria monocytogenes, Staphylococcus aureus, Bacillus cereus, and Enterococcus faecalis; the molds Penicillium islandicum, Penicillium roqueforti, Penicillium nalgiovense, Eurotium repens, and A spergillus flavus and the yeasts Candida albicans, Debaryomyces hansenii, and Zigosaccharomyces rouxii. Films with a nominal concentration of 4% (w/w) of fortified cinnamon or oregano essential oil completely inhibited the growth of the fungi; higher concentrations were required to inhibit the Gram-positive bacteria (8 and 10%, respectively), and higher concentrations still were necessary to inhibit the Gram-negative bacteria. PP films were more effective than PE/EVOH films. The atmospheres generated by the antimicrobial films inside Petri dishes were quantitatively analyzed using headspace-single drop microextraction (HS-SDME) in combination with gas chromatography-mass spectrometry (GC-MS). The analyses showed that the oregano-fortified PP films released higher levels of carvacrol and thymol, and the cinnamon-fortified PP films released higher levels of cinnamaldehyde, during the first 3-6 h of incubation, than the corresponding PE/EVOH films. Shelf-life tests were also performed, demonstrating that the antifungal activities of the films persisted for more than two months after their manufacture. In addition, migration tests (overall and specific) were performed, using both aqueous and fatty simulants, to ensure that the films meet EU regulations regarding food contact materials. Following contact with the tested films, the substances that had migrated into the aqueous simulants were recovered by direct immersion-single drop extraction (DI-SDME) and then analyzed by GC-MS. The fatty stimulant (isooctane) was directly injected into the chromatographic system.

UPLC-MS as a powerful technique for screening the nonvolatile contaminants in recycled PET.

The possibility of using recycled polyethylene terephthalate as a food contact material is being seriously considered, but the potential migration of nonvolatile compounds from it must be assessed to ensure that it is safe to do so. In the study presented here, four samples of recycled PET were each exposed to three food simulants under the harsh extraction conditions stipulated by European legislation regarding migration tests. The nonvolatile compounds that migrated from them were determined by ultra performance liquid chromatography-mass spectrometry using three different cone voltages, and both positive and negative ionization modes. A total of 36 chemical compounds were detected, some of which were identified, including common additives such as N,N'-di-beta-naphthyl-p-phenylenediamine (antioxidant) and 2,4-di-tert-butyl-6-(5-chloro-2H-benzotriazol-2-yl)phenol (light stabilizer) as well as degradation compounds such as ethylene terephthalate dimers and trimers. In addition, specific migration values of three common components of polyethylene terephthalate (diethylene glycol, terephthalic acid, and isophthalic acid) were determined and found to occur at levels of <1 mg/kg-much lower than the specific migration limits stipulated by European legislation.

On-line strategies for determining trace levels of nitroaromatic explosives and related compounds in water.

We report the development and tests of several systems for the simultaneous determination of 18 energetic compounds and related congeners in untreated water samples. In these systems a Restricted Access Material trap or liquid-chromatography precolumn (with a C(18) or porous graphitic carbon, PGC, stationary phase) followed by a PGC analytical column are used for sample clean-up, enrichment and separation of the trace level analytes, which are then analyzed by mass spectrometry (MS). The relative merits of two MS ionization interfaces (atmospheric pressure chemical ionization, APCI, and atmospheric pressure photoionization, APPI) were also compared for the MS identification and quantification of these analytes. APCI was found to be superior in cases where both alternatives are applicable. A major drawback when applying APPI is that no signal is obtained for the cyclic nitramines and nitrate esters. Using APCI, a wide spectrum of unstable compounds can be determined in a single analysis, and the feasibility of using large volume samples (up to 100 mL) in combination with the sensitivity of the MS detection system provide method detection limits ranging from 2.5 pg/mL (for 2,4-dinitrotoluene and 2,6-diamino-6-nitrotoluene) to 563 pg/mL (for pentaerythritol tetranitrate, PETN), with repeatability ranging from 2 to 7%. Other chemometric parameters such as robustness, selectivity, repeatability, and intermediate precision were also evaluated in the validation of the extraction methods for use in water analysis. Tests with untreated groundwater and drinking water samples, spiked with 20 ng of the analytes, yielded results similar to those obtained with high purity water samples.

Solid- and vapor-phase antimicrobial activities of six essential oils: susceptibility of selected foodborne bacterial and fungal strains.

The antimicrobial activity of essential oils (EOs) of cinnamon (Cinnamon zeylanicum), clove (Syzygium aromaticum), basil (Ocimum basillicum), rosemary (Rosmarinus officinalis), dill (Anethum graveolens), and ginger (Zingiber officinalis) was evaluated over a range of concentrations in two types of contact tests (solid and vapor diffusion). The EOs were tested against an array of four Gram-positive bacteria (Staphylococcus aureus, Bacillus cereus, Enterococcus faecalis, and Listeria monocytogenes), four Gram-negative bacteria (Escherichia coli, Yersinia enterocolitica, Salmonella choleraesuis, and Pseudomonas aeruginosa), and three fungi (a yeast, Candida albicans, and two molds, Penicillium islandicum and Aspergillus flavus). The rationale for this work was to test the possibility of creating a protective atmosphere by using natural compounds that could extend the shelf life of packaged foodstuffs while minimizing organoleptic alterations. In the solid diffusion tests, cinnamon and clove gave the strongest (and very similar) inhibition, followed by basil and rosemary, with dill and ginger giving the weakest inhibition. The fungi were the most sensitive microorganisms, followed by the Gram-positive bacterial strains. The Gram-negative strain P. aeruginosa was the least inhibited. The composition of the atmosphere generated by the EOs, and their minimum inhibitory concentrations (MICs), were determined using a disk volatilization method, in which no inhibition from rosemary or basil was observed. Cinnamon and clove, once again, gave similar results for every microorganism. As a general rule, MIC (fungi) << MIC (bacteria) with no clear differences between Gram-positive or -negative strains except for P. aeruginosa, which was not inhibited by any of the EOs in the vapor phase. The atmosphere generated from the EOs was analyzed by means of solid-phase microextraction combined with gas chromatography-ion trap mass spectrometry. Differences among the volatiles in the EOs, which may be responsible for the differences in their antimicrobial performances, were found.

Supercritical fluid extraction of energetic nitroaromatic compounds and their degradation products in soil samples.

This paper explores the use of supercritical fluid extraction (SFE), in combination with various analyte collection strategies, for extracting energetic nitroaromatic compounds and their degradation products from soil samples. The required selectivity has been achieved by a combination of an SFE program and active trapping. Several different collection strategies were tested, using a selection of liquids (methanol, toluene, methyl tert-butyl ether, acetonitrile), inert and solid-phase extraction materials (Nexus, Oasis, LiChrolut), and 1-cm liquid chromatography precolumns (porous graphitic carbon, PGC). The best results were obtained using SFE in combination with a PGC precolumn. This setup allows on-line cleanup of the extract, and comparable results were obtained using either GC-ECD or GC-chemical ionization-MS for confirmatory analysis. The time required for a complete analysis was less than 60 min, and only 1 mL of toluene was needed for a 0.5-g representative sample. In contrast, the EPA standard method 8330 required 18-h sonication and 20 mL of acetonitrile for a 4.0-g sample and further time for sample cleanup and HPLC analysis. The method presented here provides method detection limits in the low-nanogram range, with relative standard deviations lower than 7%. The optimized method has been compared and validated with EPA method 8330 in terms of efficiency parameters such as robustness, accuracy (trueness and precision), and capability of detection. The validation demonstrated that the two analytical methodologies give comparable performance for the determination of nitroaromatic compounds, but SFE is superior for analyzing amine degradation products.

Determination of nitroaromatic compounds in air samples at femtogram level using C18 membrane sampling and on-line extraction with LC-MS.

This paper explores the use of C18 solid-phase extraction membranes for sampling very low concentrations of nitroaromatic compounds in the atmosphere. After sampling, analytes trapped in the membrane are desorbed on-line directly by a chromatographic mobile phase. The analytes are then separated onto a porous graphitic carbon (PGC) HPLC column. Finally, they are analyzed by an LC-MS/MS detector equipped with an atmospheric pressure chemical ionization (APCI) interface. The method was validated by controlled exposure of the membranes to standard gaseous mixtures of 2,4,6-trinitrotoluene (TNT) and 2,4-dinitrotoluene (2,4-DNT). The developed method was fully characterized, and no breakthrough was observed when sampling volumes up to 9.2 m3. Analyses of membranes following medium- and long-term storage demonstrated that samples could be stored on the C18 membranes without degradation or losses. In addition, the results obtained with this technique were compared with those obtained by a gas chromatographic method in which analytes were collected on Tenax TA and thermally desorbed. The developed method allows sampling at flow rates of 15 L/min and has method detection limits in the femtogram/liter range, with a relative standard deviation lower than 10%. An additional advantage of this method is that it separates most of the TNT and DNT isomers, as demonstrated by applying the method to the analysis of headspace over military-grade TNT explosives.

Determination of gaseous toluene diisocyanate by use of solid-phase microextraction with on-fibre derivatisation.

An SPME method was developed for sampling gaseous 2,4-toluene diisocyanate (2,4-TDI) involving derivatisation of the isocyanate by reacting with dibutylamine (DBA). The TDI-DBA derivative thus formed was determined by LC-MS-MS utilising atmospheric pressure chemical ionisation (APCI). As a first step, DBA was loaded onto a poly(dimethylsiloxane)/divinylbenzene (PDMS-DVB) fibre coating by direct vapour-phase extraction of a highly concentrated diethyl ether solution of DBA. The DBA-loaded fibre was then exposed to an artificially generated atmosphere of gaseous 2,4-TDI. The linearity of the method ranged from 52.8 to 3100 microg m(-3) (6.8 to 400 ppbv) with a sampling time of 60 min. The proposed method has been applied to 2,4-TDI determination in an artificially generated dynamic standard atmosphere, yielding an approximate method detection limit (MDL) of 2 microg m(-3) (0.25 ppbv). This concentration is one twentieth of the Occupational Safety and Health Administration (OSHA) 8-hour time-weighted average (TWA) exposure limit. The sampler with the PDMS-DVB-DBA coating was found to be stable and retains the required amount of DBA for at least 10 days, an important feature for sampling systems with potential in-situ applications.

Determination of plastic monomers in water by solid-phase microextraction coupled with liquid chromatography.

A method is presented for the determination of 2 major plastic monomers, terephthalic acid and vinyl acetate, which are widely used to manufacture plastics that come in contact with foods. The analytes are extracted from aqueous solution by using solid-phase microextraction, followed by quantitation by liquid chromatography (LC) with UV detection. Multivariate optimization was applied and is described. The optimized method has linear ranges of 5-150 microg/g for terephthalic acid and 7.5-100 microg/g for vinyl acetate. Coefficients of variation at a spiking level of 20 microg/g were 13.6% for terephthalic acid and 3.1% for vinyl acetate; detection and quantitation limits were 0.59 and 1,99 microg/g, respectively, for terephthalic acid and 1.56 and 5.20 microg/g, respectively, for vinyl acetate. The characteristics of both the extraction technique and its coupling with LC are described and discussed.

Development of a personal isocyanate sampler based on DBA derivatization on solid-phase microextraction fibers.

The design and evaluation of a portable diffusive sampler for isocyanates is described. The sampler employs dibutylamine (DBA) loaded onto 60-microm polydimethylsiloxane-divinylbenzene (PDMS-DVB) solid-phase microextraction (SPME) fibers. The DBA-isocyanate derivative is then desorbed by sonication and analyzed by LC-MS using atmospheric pressure chemical ionization (APCI). The samplers are calibrated (i.e. the uptake rate is calculated) by exposing them to a known concentration of hexamethylene diisocyanate (HDI) in a standard gas-generation chamber. The uptake rate for the proposed method, at room temperature (25 degrees C), is 1.13 pg (min ppb(v))(-1) and the method detection limit is 3.2 microg m(-3), equivalent to less than 10% of the airborne time-weighted average (TWA) exposure limits recommended by both the National Institute for Occupational Safety and Health (NIOSH) and the American Conference of Governmental Industrial Hygienists (ACGIH). Practical points that should be considered when using the SPME device as a diffusive sampler are discussed.

Assessing the suitability of recycled plastics used as agricultural soil covers: migration study and experimental harvest.

The present work is focused on evaluating the suitability of recycling postconsumer agricultural plastic films again for the same use. The criteria to assess the suitability was based on migration study. Both overall and specific migration tests were performed, and the results obtained (ranging from 0.14 to 1.27 mg/dm(2) for overall migration and from not detectable to 6.98 microg/dm(2) for specific migration) show how, from this point of view, the recycled material can be safely proposed to be used again as agricultural soil covers. A theoretical discussion about the migration process is also presented and a simple mathematical model was applied to the data obtained, showing how total migration which is experimentally detected is theoretically predictable. These conclusions found were used to design and develop a controlled crop of tomato by using this recycled film. The use of the recycled plastic in the whole process and the behavior and properties of the pesticides absorbed in the postconsumer film are discussed.

Use of solid-phase microextraction for the analysis of bisphenol A and bisphenol A diglycidyl ether in food simulants.

A new method has been developed to simultaneously analyse bisphenol A (BPA) and bisphenol A diglycidyl ether (BADGE) in aqueous based food simulants. The method consists on direct immersion solid-phase microextraction (SPME) of the analytes from the liquid matrix and subsequent chromatographic analysis by gas chromatography-mass spectrometry. Using the proposed method, a whole analysis (including chromatographic step) can be completed in less than 40 min, with minimum sample handling. The SPME method shows good analytical performance for simultaneous BPA and BADGE analysis, except for BADGE determination in the aqueous alcohol (simulant C) solution. Detection limits ranging from 0.1 to 2.0 ng/g for BPA and from 13 to 15 ng/g from BADGE were obtained, with a linear range from the low-ng/g to several-microg/g range for BPA and from 0.1 microg/g to 40 microg/g for BADGE. A possible optimisation method has been also developed and introduced.

A systematic approach to optimize solid-phase microextraction. Determination of pesticides in ethanol/water mixtures used as food simulants.

The optimization of solid-phase microextraction (SPME) of several organochlorine and organophosphorus pesticides is presented, and the influence of variables is discussed. The optimized method is applied to several selected ethanol/water mixtures used as food simulants, and the influence of the ethanol content on the SPME performance is also discussed. Detection limits ranging from 0.02 to 0.04 ng/g for water simulant and from 38.7 to 205.5 ng/g for 95% ethanol simulant were obtained. The relative standard deviation (% RSD) was <20% in all cases. The optimized method is compared with classical liquid-liquid extraction (LLE).

Design of a test for migration studies in the vapour phase.

A new test for migration studies in the vapour phase has been developed. The system consists of placing the packaging material containing the potential migrants in a glass vial together with the solid adsorbent, Tenax GC. The vial is introduced into an oven at a controlled temperature for different periods of time. Once the migration test has finished, Tenax is extracted by supercritical fluid chromatography (SFE) and analysed by GC-ECD. Migration conditions of 7 days at 80 degrees C are proposed. The results obtained are discussed.

Sulphate determination after its reduction to hydrogen sulphide and volatile separation by molecular absorption spectrometry.

A rapid spectrophotometric method for sulphate determination in a discontinuous mode is described. The method is based on sulphate reduction to hydrogen sulphide followed by its volatilization and absorption in an alkaline solution. The reduction is obtained when a sulphate sample is heated to 287 degrees C for 15 min, with a mixture of Fe(o)/KI and phosphoric acid. The resulting gas is swept by nitrogen flow into a 0.1 M sodium hydroxide solution and the absorbance of the sulphide ions is measured directly at 230 nm. The proposed method enabled us to determine 50-700 mug of total sulphate with a relative standard deviation of the order of 5%. The method has been applied for the determination of sulphates in liquid (mineral waters) and solid (gypsiferous soils) samples.