The use of peracetic acid for estrogen removal from urban wastewaters: E2 as a case study.

17ß-Estradiol (E2) is a natural estrogen produced by the feminine endocrine system. It is excreted mainly through urine and feces. Exposure to E2 may affect the reproductive system of both animals and humans, especially since the removal of E2 in conventional processes and technologies present in the wastewater treatment plants is not sufficient. Chlorine is one of the most studied and used oxidant worldwide. Although there are studies that demonstrate the endocrine disrupting compounds removal like E2, its reaction with organic matter can originate by-products, namely, trihalomethanes, which are known to have high toxic potential. The main aim of the present study was to evaluate the removal of E2 (50 µg E2 L-1-maximum concentration) using peracetic acid (PAA), a seeming cleaner and innocuous alternative to chlorine. To this end, a series of jar tests were performed, using different peracetic acid concentrations (1, 5, 10, and 15 mg L-1) and contact times (10, 15, and 20 min). The results obtained showed that a peracetic acid concentration of 15 mg L-1 with a contact time of 20 min had a removal efficacy of approximately 100%. The second main goal of this study was to evaluate the ecotoxicological potential of the tested treatments on the zebrafish Danio rerio. Several oxidative stress biomarkers were evaluated, namely glutathione S-transferase, lipid peroxidation, and catalase, besides vitellogenin. Both peracetic acid and E2 caused significant increases in the oxidative stress biomarkers, although this did not lead to increased lipid peroxidation levels. In addition, peracetic acid significantly decreased the estrogenic activity of E2, as indicated by decreased vitellogenin levels. Peracetic acid demonstrated to have great potential as an alternative disinfectant for chlorine treatments, and indications for future research are discussed.

Efficacy assessment of peracetic acid in the removal of synthetic 17α-ethinyl estradiol contraceptive hormone in wastewater.

Increasing concerns have been raised on endocrine disrupting chemicals like the sex hormone 17α-ethinylestradiol (EE2), the more since traditional wastewater (WW) treatments appear to be ineffective for their removal. The efficacy of the relatively novel disinfectant peracetic acid (PAA) in EE2 removal was evaluated, as well as its potential effects on WW quality parameters. The treatments tested for EE2 removal were also evaluated in terms of toxicity, through the determination of biochemical responses (antioxidant enzymes, lipid peroxidation and vitellogenin induction) using zebrafish (Danio rerio) as a biological model. PAA contact times less than 20 min appeared insufficient regardless of the PAA dose tested, but a 100% EE2 removal was attained at a PAA concentration of 15 mg/L with a contact time of 20 min. Total suspended solids, chemical oxygen demand and pH in PAA treatments remained well within levels set in European legislation for WW discharge. EE2 induced significant increased vitellogenin (VTG) levels in both female and male fish, indicating increased estrogenic activity, especially in males suggesting an endocrine disruption effect. With the addition of PAA (15 mg/L), however, VTG levels in both sexes returned to control values. Although this PAA treatment showed increased levels of the antioxidant enzyme catalase, the lipid peroxidation levels were similar or even lower than in controls. Overall the results suggest that the use of PAA appears a promising way forward as a less toxic alternative to chlorine disinfection with high efficiency in the removal of EDC like EE2.

Recovery of phenolic compounds from multi-component solution by a synthesized activated carbon using resorcinol and formaldehyde.

The adsorption of four phenolic compounds (gallic acid, protocatechuic acid, vanillic acid and syringic acid) is investigated using a synthesized mesoporous carbon on both single and multi-component synthetic solutions. Some correlation of the adsorption capacity of the carbon and the nature of adsorbate could be made, except for gallic acid whose concentration decrease seems to be not exclusively due to adsorption but also to polymerization reaction. In the multi-component mixture, negative effects in the adsorption capacity are observed probably due to competition for the active centers of the adsorbent surface. In desorption studies, ethanol presents better performance than water and acetonitrile. Vanillic acid is the compound with the higher adsorption and interestingly it is then possible to desorb a relatively high amount of it from the adsorbent, which may represent a possibility for a selective recovery of vanillic acid. These results present a potential way to treat the wastewater from the cork industry.

Modified Atmosphere Packaging (MAP) for Seaweed Conservation: Impact on Physicochemical Characteristics and Microbiological Activity.

Conventional conservation techniques such as drying, salting or freezing do not allow for preserving the original characteristics of seaweeds. The present work aims to study the impact of minimal processing, in particular "Modified Atmosphere Packaging" (MAP), on the physicochemical characteristics and food safety of two seaweed species, "laver" (Porphyra umbilicalis) and "sea-lettuce" (Ulva lactuca), stored at 6 °C for 15 days. Different parameters were evaluated using analytical methods, namely the composition of headspace gases, color, texture, microorganisms, and volatile organic compounds (VOCs). The main findings of this study were that the MAP treatment was able to inhibit the respiration rate of minimally processed seaweeds, also preserving their color and texture. There was a remarkable reduction in the microbial load for P. umbilicalis treated under modified and vacuum atmospheres, and U. lactuca exhibited relatively steady values with no notable differences between the treatments and the control. Therefore, during the 15-day study period, both seaweeds met the requirements for food safety. GC-TOF-MS allowed to conclude that both MAP and vacuum treatments were more efficient in maintaining the odor characteristics of U. lactuca compared to P. umbilicalis with no significant differences throughout the storage days. Metabolic responses to diverse sources of abiotic stress seemed to account for most of the changes observed.

Determination of microcystin-LR in waters in the subnanomolar range by sol-gel imprinted polymers on solid contact electrodes.

The present work reports new sensors for the direct determination of Microcystin-LR (MC-LR) in environmental waters. Both selective membrane and solid contact were optimized to ensure suitable analytical features in potentiometric transduction. The sensing layer consisted of Imprinted Sol-Gel (ISG) materials capable of establishing surface interactions with MC-LR. Non-Imprinted Sol-Gel (NISG) membranes were used as negative control. The effects of an ionic lipophilic additive, time of sol-gel polymerization, time of extraction of MC-LR from the sensitive layer, and pH were also studied. The solid contact was made of carbon, aluminium, titanium, copper or nickel/chromium alloys (80 : 20 or 90 : 10). The best ISG sensor had a carbon solid contact and displayed average slopes of 211.3 mV per decade, with detection limits of 7.3 × 10(-10) M, corresponding to 0.75 µg L(-1). It showed linear responses in the range of 7.7 × 10(-10) to 1.9 × 10(-9) M of MC-LR (corresponding to 0.77-2.00 µg L(-1)), thus including the limiting value for MC-LR in waters (1.0 µg L(-1)). The potentiometric-selectivity coefficients were assessed by the matched potential method for ionic species regularly found in waters up to their limiting levels. Chloride (Cl(-)) showed limited interference while aluminium (Al(3+)), ammonium (NH(4)(+)), magnesium (Mg(2+)), manganese (Mn(2+)), sodium (Na(+)), and sulfate (SO(4)(2-)) were unable to cause the required potential change. Spiked solutions were tested with the proposed sensor. The relative errors and standard deviation obtained confirmed the accuracy and precision of the method. It also offered the advantages of low cost, portability, easy operation and suitability for adaptation to flow methods.

Metabolite identification of ibuprofen biodegradation by Patulibacter medicamentivorans under aerobic conditions.

Ibuprofen (IBU) is a non-steroidal anti-inflammatory drug that is becoming increasingly recognized as an important micropollutant to be monitored in wastewater treatment plants (WWTP), since it has been detected in effluents at the µg L-1 level. The IBU metabolites from biological degradation are not completely understood and can represent a threat to natural aquatic systems. P. medicamentivorans was previously isolated from WWTP sludge and found to be capable of IBU degradation. The aerobic biodegradation of ibuprofen by this organism was investigated in a batch lab-scale reactor for the identification of the metabolites formed. The metabolites were analysed and putatively identified by HPLC-DAD-MS/MS and GC-MS and biodegradation pathways were proposed. The toxicity and the biodegradability potential of the metabolites were also investigated. The results showed that IBU biotransformation was achieved by hydroxylation followed by the formation of a carboxylic acid in the IBU molecule and by the formation of a catechol, allowing the aromatic ring cleavage. Two biodegradation pathways were proposed: in one, the metabolites generated from the enzymatic action correspond to a less biodegradable chemical structure of the intermediate products (isobutylbenzene and 3-isobutylphenol), with comparatively higher toxicity; in the other mechanism, more oxidable chemical structures were formed with less toxicity and higher biodegradability. This suggests that the biodegradation of IBU by P. medicamentivorans can take place by more than one mechanism regarding the enzymes formed by this Gram-positive bacterium, with subsequent oxidation of the parent compound to overall more soluble and less toxic compounds to fish, daphnia and green algae.

Analysis of Food by High Performance Liquid Chromatography Coupled with Coulometric Detection and Related Techniques: A Review.

The development of single coulometric cells in combination with high performance liquid chromatography to dual cells and to the coulometric electrode array detector is described. An overview is given about the application of these methods in food chemistry. Easily oxidizable compounds, such as phenolic substances, pesticides, or vitamins, can be determined, as well as substances with high oxidation potentials or electroinactive compounds. Substances exhibiting poor electrochemical activity can be transformed to electroactive compounds by precolumn derivatization, postcolumn photochemical reactions, postcolumn enzyme reactors, or by using the oxidative/reductive mode for coulometric electrode array detection. Furthermore, it is shown that the interesting combination of high performance liquid chromatography with electrochemistry and mass spectrometry has opened further possibilities with respect to interpretation of redox reactions, drug metabolism studies, metabolomics, and electrochemical derivatization.

Molecular motions in amorphous ibuprofen as studied by broadband dielectric spectroscopy.

The molecular mobility of amorphous ibuprofen has been investigated by broadband dielectric relaxation spectroscopy (DRS) covering a temperature range of more than 200 K. Four different relaxation processes, labeled as alpha, beta, gamma, and D, were detected and characterized, and a complete relaxation map was given for the first time. The gamma-process has activation energy E a = 31 kJ.mol (-1), typical for local mobility. The weak beta-relaxation, observed in the glassy state as well as in the supercooled state was identified as the genuine Johari-Goldstein process. The temperature dependence of the relaxation time of the alpha-process (dynamic glass transition) does not obey a single VFTH law. Instead two VFTH regimes are observed separated by a crossover temperature, T B = 265 K. From the low temperature VFTH regime, a T g (diel) (tau =100 s) = 226 K was estimated, and a fragility or steepness index m = 93, was calculated showing that ibuprofen is a fragile glass former. The D-process has a Debye-like relaxation function but the temperature dependence of relaxation time also follows the VFTH behavior, with a Vogel temperature and a pre-exponential factor which seem to indicate that its dynamics is governed by the alpha-process. It has similar features as the Debye-type process observed in a variety of associating liquids, related to hydrogen bonding dynamics. The strong tendency of ibuprofen to form hydrogen bonded aggregates such as dimers and trimers either cyclic or linear which seems to control in particular the molecular mobility of ibuprofen was confirmed by IR spectroscopy, electrospray ionization mass spectrometry, and MD simulations.

A new lupene triterpenetriol and anticholinesterase activity of Salvia sclareoides.

A new lupene triterpenetriol was isolated from the acetone extract of the aerial parts of Salvia sclareoides and characterised as (1beta,3beta)-lup-20(29)-ene-1,3,30-triol (1). In addition, nepetidin (2), nepeticin (3), lupendiol (4), (1beta,11alpha)-dihydroxy-lup-20(29)-en-3-one (5), ursolic acid (6), sumaresinolic acid (7) and hederagenin (8), were identified in this Salvia sp. To the best of our knowledge, the compounds 2 and 7 are new constituents in Salvia spp. The acetone, ethanol, butanol and water extracts of the plant were screened for the in vitro inhibitory activity of acetylcholinesterase (AChE) and butyrilcholinesterase (BChE), enzymes which play a role in the Alzheimer disease. All extracts inhibited acetylcholinesterase activity at 10 microg/ml, a remarkable activity since the standard drug rivastigmine does not inhibit acetylcholinesterase at the same concentration. Regarding the butyrilcholinesterase, the acetone extract at 1000 microg/ml was able to inhibit completely the enzyme activity and the butanol and ethanol extracts, at this concentration, produced a potent inhibition of BchE.

Testing the variability of PSA expression by different human prostate cancer cell lines by means of a new potentiometric device employing molecularly antibody assembled on graphene surface.

Prostate Specific Antigen (PSA) is widely used as a biomarker for prostate cancer. Recently, an electrochemical biosensor for PSA detection by means of molecularly imprinted polymers (MIPs) was developed. This work evaluated the performance and the effectiveness of that PSA biosensor in screening the biomarker PSA in biological media with complex composition, collected from different human prostate cell line cultures. For that, the prostate cancer LNCaP and PC3 cells, and the non-cancerous prostate cell line PNT2 were cultured for 2, 7 and 14days in either α-MEM or RPMI in the presence of 10% or 30% fetal bovine serum. Human gingival fibroblasts were used as a non-cancerous non-prostatic control. The different culture conditions modulated cellular proliferation and the expression of several prostate markers, including PSA. The electrochemical biosensor was able to specifically detect PSA in the culture media and values obtained were similar to those achieved by a commercial Enzyme-Linked Immunosorbent Assay (ELISA) kit, the most commonly used method for PSA quantification in prostate cancer diagnosis. Thus, the tested biosensor may represent a useful alternative as a diagnostic tool for PSA determination in biological samples.

A novel pentacyclic triterpene from Leontodon filii.

A novel oleanene triterpenetetrol was isolated from the chloroform extract of the aerial parts of Leontodon filii. Its structure was shown to be 2beta,3beta,15alpha,21beta-olean-12-ene-2,3,15,21-tetrol by chemical and spectroscopic methods. The fungicidal efficacy of the chloroform and methanol extracts of the plant was also evaluated, a protective effect being found against Plasmopara viticola, Botrytis cinerea, particularly powerful against Pyricularia oryzae.

A new dihydroxysterol from the marine phytoplankton Diacronema sp.

Diacronema sp. was cultured and its sterols were separated by column chromatography on silica gel. The new sterol 24-ethyl-4alpha-methyl-cholestane-3,20-diol (1) was characterised by NMR and MS spectrometry, as well as (22E)-24-ethyl-4alpha-methyl-5alpha-cholest-22-en-3beta-ol (2) and beta-sitosterol, the major components of the sterol fractions. Neither the biosynthetic origin of the new dihydroxysterol nor its role in the biochemistry of Diacronema is known.

Novel sensory surface for creatine kinase electrochemical detection.

This work describes a novel concept of biosensor for quantifying enzymes, where the substrate is immobilized directly over the working area of a screen printed electrode (Au-SPE). This concept is applied here to creatine kinase isoenzyme (CK-MB), a cardiac biomarker in ischemic conditions. It acts as a phospho-transferase on creatine (Crea), requiring the presence of phosphate. So, the phosphorylated form of creatine (Pcrea) was immobilized on the Au/SPE previously aminated with cysteamine (Cys) by self-assembling monolayer technique. Electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) studies were used to follow the chemical modifications in the Au-SPE. Since Pcrea is an electroactive species at low potential, its consumption over the platform by the enzyme changed the electrical response of the biosensor. So, CK-MB determination has been achieved in mediator free-conditions due the redox proprieties of the Pcrea. The analytical features of the resulting biosensor were studied by square wave voltammetry (SWV). The limit of detection was 0.11 µg/mL and the slope was -0.029(± 0.0035) µA × mL/µg. The interference effect of troponin T (TnT), bovine serum albumin (BSA) and myoglobin (Myo) in the performance of the sensor was tested and good selectivity was observed. The biosensor was successfully applied to biological fluids, showing good stability at room temperature and excellent sensitivity and selectivity. This new concept of biosensor is especially useful for point of care (POC) applications, due to the low cost and small size of the final device.

Sarcosine oxidase composite screen-printed electrode for sarcosine determination in biological samples.

As the prostate cancer (PCa) progresses, sarcosine levels increase both in tumor cells and urine samples, suggesting that this metabolite measurements can help in the creation of non-invasive diagnostic methods for this disease. In this work, a biosensor device was developed for the quantification of sarcosine via electrochemical detection of H2O2 (at 0.6V) generated from the catalyzed oxidation of sarcosine. The detection was carried out after the modification of carbon screen printed electrodes (SPEs) by immobilization of sarcosine oxidase (SOX) on the electrode surface. The strategies used herein included the activation of the carbon films by an electrochemical step and the formation of an NHS/EDAC layer to bond the enzyme to the electrode, the use of metallic or semiconductor nanoparticles layer previously or during the enzyme immobilization. In order to improve the sensor stability and selectivity a polymeric layer with extra enzyme content was further added. The proposed methodology for the detection of sarcosine allowed obtaining a limit of detection (LOD) of 16nM, using a linear concentration range between 10 and 100nM. The biosensor was successfully applied to the analysis of sarcosine in urine samples.

Smart plastic antibody material (SPAM) tailored on disposable screen printed electrodes for protein recognition: application to myoglobin detection.

This work introduces two major changes to the conventional protocol for designing plastic antibodies: (i) the imprinted sites were created with charged monomers while the surrounding environment was tailored using neutral material; and (ii) the protein was removed from its imprinted site by means of a protease, aiming at preserving the polymeric network of the plastic antibody. To our knowledge, these approaches were never presented before and the resulting material was named here as smart plastic antibody material (SPAM). As proof of concept, SPAM was tailored on top of disposable gold-screen printed electrodes (Au-SPE), following a bottom-up approach, for targeting myoglobin (Myo) in a point-of-care context. The existence of imprinted sites was checked by comparing a SPAM modified surface to a negative control, consisting of similar material where the template was omitted from the procedure and called non-imprinted materials (NIMs). All stages of the creation of the SPAM and NIM on the Au layer were followed by both electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). AFM imaging was also performed to characterize the topography of the surface. There are two major reasons supporting the fact that plastic antibodies were effectively designed by the above approach: (i) they were visualized for the first time by AFM, being present only in the SPAM network; and (ii) only the SPAM material was able to rebind to the target protein and produce a linear electrical response against EIS and square wave voltammetry (SWV) assays, with NIMs showing a similar-to-random behavior. The SPAM/Au-SPE devices displayed linear responses to Myo in EIS and SWV assays down to 3.5 µg/mL and 0.58 µg/mL, respectively, with detection limits of 1.5 and 0.28 µg/mL. SPAM materials also showed negligible interference from troponin T (TnT), bovine serum albumin (BSA) and urea under SWV assays, showing promising results for point-of-care applications when applied to spiked biological fluids.

Artificial antibodies for troponin T by its imprinting on the surface of multiwalled carbon nanotubes: its use as sensory surfaces.

A novel artificial antibody for troponin T (TnT) was synthesized by molecular imprint (MI) on the surface of multiwalled carbon nanotubes (MWCNT). This was done by attaching TnT to the MWCNT surface, and filling the vacant spaces by polymerizing under mild conditions acrylamide (monomer) in N,N'-methylenebisacrylamide (cross-linker) and ammonium persulphate (initiator). After removing the template, the obtained biomaterial was able to rebind TnT and discriminate it among other interfering species. Stereochemical recognition of TnT was confirmed by the non-rebinding ability displayed by non-imprinted (NI) materials, obtained by imprinting without a template. SEM and FTIR analysis confirmed the surface modification of the MWCNT. The ability of this biomaterial to rebind TnT was confirmed by including it as electroactive compound in a PVC/plasticizer mixture coating a wire of silver, gold or titanium. Anionic slopes of 50 mV decade(-1) were obtained for the gold wire coated with MI-based membranes dipped in HEPES buffer of pH 7. The limit of detection was 0.16 µg mL(-1). Neither the NI-MWCNT nor the MWCNT showed the ability to recognize the template. Good selectivity was observed against creatinine, sucrose, fructose, myoglobin, sodium glutamate, thiamine and urea. The sensor was tested successfully on serum samples. It is expected that this work opens new horizons on the design of new artificial antibodies for complex protein structures.

Myoglobin-biomimetic electroactive materials made by surface molecular imprinting on silica beads and their use as ionophores in polymeric membranes for potentiometric transduction.

Myoglobin (Mb) is among the cardiac biomarkers playing a major role in urgent diagnosis of cardiovascular diseases. Its monitoring in point-of-care is therefore fundamental. Pursuing this goal, a novel biomimetic ionophore for the potentiometric transduction of Mb is presented. It was synthesized by surface molecular imprinting (SMI) with the purpose of developing highly efficient sensor layers for near-stereochemical recognition of Mb. The template (Mb) was imprinted on a silane surface that was covalently attached to silica beads by means of self-assembled monolayers. First the silica was modified with an external layer of aldehyde groups. Then, Mb was attached by reaction with its amine groups (on the external surface) and subsequent formation of imine bonds. The vacant places surrounding Mb were filled by polymerization of the silane monomers 3-aminopropyltrimethoxysilane (APTMS) and propyltrimethoxysilane (PTMS). Finally, the template was removed by imine cleavage after treatment with oxalic acid. The results materials were finely dispersed in plasticized PVC selective membranes and used as ionophores in potentiometric transduction. The best analytical features were found in HEPES buffer of pH 4. Under this condition, the limits of detection were of 1.3 × 10(-6)mol/L for a linear response after 8.0 × 10(-7) mol/L with an anionic slope of -65.9 mV/decade. The imprinting effect was tested by preparing non-imprinted (NI) particles and employing these materials as ionophores. The resulting membranes showed no ability to detect Mb. Good selectivity was observed towards creatinine, sacarose, fructose, galactose, sodium glutamate, and alanine. The analytical application was conducted successfully and showed accurate and precise results.

Bioactivity studies and chemical profile of the antidiabetic plant Genista tenera.

AIM OF THE STUDY: Genista tenera is a plant endemic to the island of Madeira and is used in folk medicine to control diabetes. In the present work we evaluate the antihyperglycaemic activity of its n-butanol extract and determine its chromatographic profile. In addition, this extract, the ethyl acetate and diethyl ether plant extracts were studied in order to assess the plant antioxidant and acetylcholinesterase inhibitory activities, as well as its cyto- and genotoxicities. MATERIALS AND METHODS: HPLC-DAD-ESI-MS was used to analyze the flavonoid profile of the n-butanol extract. The antihyperglycaemic activity of this extract was performed over streptozotocin induced diabetic Wistar rats (200 mg/kg, bw/day), for 15 days. Antioxidant activity (DPPH assay) and acetylcholinesterase inhibitory effect (Ellman method) were also performed. Acute cytotoxicity and genotoxicity were assessed by proliferative index quantification and the short-term chromosomal aberration technique, after exposure of lymphocytes to the extracts. RESULTS AND CONCLUSIONS: The n-butanol extract, where 21 monoglycosyl and 12 diglycosyl flavonoids were detected, significantly lowered blood glucose levels, bringing them to normal values after 15 days of treatment. The best radical scavenging activity was observed for the ethyl acetate extract (48.7% at 139.1 microg/mL), which was also the most effective one at the minimal concentration tested. The highest acetylcholinesterase inhibitory activity (77.0% at 70.0 microg/mL) was also obtained with the ethyl acetate extract. In vitro toxicity studies showed no evidence for acute cytotoxicity or genotoxicity. This is the first report on antidiabetic activity of genus Genista.