Microwave-assisted extraction of antioxidant compounds from by-products of Turkish hazelnut (Corylus avellana L.) using natural deep eutectic solvents: Modeling, optimization and phenolic characterization.

An environmentally friendly method using natural deep eutectic solvents (NADES) and microwave-assisted extraction (MAE) for the recovery of bioactive compounds from hazelnut pomace (a hazelnut oil process by-product) was developed to contribute to their sustainable valorization. Eight different NADES were prepared for the extraction of antioxidant constituents from hazelnut pomace, and choline chloride:1,2-propylene glycol (CC-PG) was determined as the most suitable NADES, considering their extraction efficiency and physicochemical properties. After selecting suitable NADES, operational parameters for the MAE process of antioxidants from hazelnut pomace were optimized and modeled using response surface methodology. For the highest recovery of antioxidants, the operational parameters of the MAE process were found to be 24% water, 38 min, 92 °C and 18 mL/0.1 g-DS. Under optimized conditions, extracts of both pomace as a by-product and unprocessed hazelnut flours of three different hazelnut samples (Tombul, Çakildak, and Palaz) were prepared, and their antioxidant capacities were evaluated by spectrophotometric methods. Antioxidant capacities of CC-PG extracts of all hazelnut samples were 2-3 times higher than those of ethanolic extracts. In addition, phenolic characterization of the prepared extracts was carried out using the UPLC-PDA-ESI-MS/MS system. The results of this study suggest that hazelnut by-products can potentially be considered an important and readily available source of natural antioxidants. Furthermore, the modeled MAE procedure has the potential to create an effective and sustainable alternative for pharmaceutical and food industries.

Colorimetric sensors and nanoprobes for characterizing antioxidant and energetic substances.

The development of analytical techniques for antioxidant compounds is important, because antioxidants that can inactivate reactive species and radicals are health-beneficial compounds, also used in the preservation of food and protection of almost every kind of organic substance from oxidation. Energetic substances include explosives, pyrotechnics, propellants and fuels, and their determination at bulk/trace levels is important for the safety and well-being of modern societies exposed to various security threats. Most of the time, in field/on site detection of these important analytes necessitates the use of colorimetric sensors and probes enabling naked-eye detection, or low-cost and easy-to-use fluorometric sensors. The use of nanosensors brings important advantages to this field of analytical chemistry due to their various physico-chemical advantages of increased surface area, surface plasmon resonance absorption of noble metal nanoparticles, and superior enzyme-mimic catalytic properties. Thus, this critical review focuses on the design strategies for colorimetric sensors and nanoprobes in characterizing antioxidant and energetic substances. In this regard, the main themes and properties in optical sensor design are defined and classified. Nanomaterial-based optical sensors/probes are discussed with respect to their mechanisms of operation, namely formation and growth of noble metal nanoparticles, their aggregation and disaggregation, displacement of active constituents by complexation or electrostatic interaction, miscellaneous mechanisms, and the choice of metallic oxide nanoparticles taking part in such formulations.

Comparison of endometrial prostanoid profiles in three infertile subgroups: the missing part of receptivity?

OBJECTIVE: To study the prostanoid profile of the endometria of patients with recurrent implantation failure (RIF), unexplained infertility (UIF), and recurrent miscarriages (RM), and to compare them with the endometria of healthy fertile controls. DESIGN: Prospective cohort study. SETTING: University hospital. PATIENT(S): Fifteen patients with RIF, 18 patients with UIF, 16 patients with RM, and 23 fertile controls were recruited. INTERVENTION(S): Endometrial samples were taken during the window of implantation. After tissue homogenization and extraction, analysis with ultra-performance liquid chromatography diode array detector electrospray ionisation tandem mass spectrometry was performed. MAIN OUTCOME MEASURES: Concentrations of prostaglandin (PG) D1, PGE1, PGF1α, 6-ketoPGF1α, PGD2, PGE2, PGF2α, 15-deoxy-Δ12,14-PGJ2, PGD3, PGE3, PGF3α, thromboxane B2, 13,14-dihydro-PGE1, 13,14-dihydro-PGF1α, 13,14-dihydro-PGF2α, 13,14-dihydro-15-keto-PGE1, 13,14-dihydro-15-keto-PGE2, and 13,14-dihydro-15-keto-PGF2α were assessed. RESULT(S): Comparison of the endometria of patients with UIF and the controls showed no statistically significant differences. When the endometria of patients with RIF were compared with the controls, thromboxane B2 (TXB2) was found significantly higher (843.1 pg/mg vs. 133.5 pg/mg). When the endometria of patients with RM were compared with controls, 13,14-dihydro-15-keto PGF2α and TXB2 were found significantly higher (3907.30 pg/mg vs. 17.80 pg/mg and 858.7 pg/mg vs. 133.5 pg/mg respectively). CONCLUSION(S): We identified increased endometrial presence of TXB2 in patients with RM and RIF, and 13,14-dihydro-15-keto PGF2α in patients with RM. Although common ground is observed for RM and RIF, prostanoids, on the other hand, might make their own contribution to endometrial receptivity as important as genes and proteins. Attempts to normalize the prostaglandin profile of the endometrium via enzymatic activity can open new therapeutic options.

Screening Method for Argan Oil Adulteration with Vegetable Oils: An Online HPLC Assay with Postcolumn Detection Utilizing Chemometric Multidata Analysis.

This study is focused on examining the tocopherol isomers (α-, γ-, and δ-) fingerprinting by online RP-HPLC analysis with post column detection using CUPRAC (cupric reducing antioxidant capacity) methodology for argan oil authenticity. The proposed online assay was validated with good precision, reproducibility, and linearity. Sixteen argan oil samples (100% pure-certified and other commercial argan oils), possible adulterating vegetable oils (i.e., olive, sunflower, corn, and soya oils), and virgin argan oil blended with olive, sunflower, corn, and soya oils at levels of 5%, 10%, 15%, and 20% were analyzed. Spectrophotometric CUPRAC, DPPH, and ABTS assays were applied. Discrimination of fraudulent argan oils from virgin samples was performed by utilizing orthogonal partial least-squares discriminant analysis (OPLS-DA) regression modeling with good sensitivity and specificity. We suggested [γ-toc/α-toc] value as a new first screening adulteration factor (AF) that could be used to assess fraudulent argan oil samples. The distinct decrement in AF value was observed by the increase of adulteration rate. The AF values for virgin argan oils were ranged from 11.8 (lower limit) to 18.6 (upper limit). The presence of ß-sitosterol detected in commercial argan oils (with AF values out of limit values) was evaluated as fraudulent which was in accordance with the proposed assay. Our method enabled the detection of argan oil samples at adulteration levels of >5% in the case of sunflower, olive, and soya oils, >15% in the case of corn oil. This method may be an alternative and specific assay for the authentication and quality detection of commercial argan oils.

A novel colorimetric sensor for measuring hydroperoxide content and peroxyl radical scavenging activity using starch-stabilized gold nanoparticles.

A novel colorimetric nanosensor was developed for evaluating peroxyl radical scavenging activity of phenolic antioxidants and for the detection of hydroperoxides formed during AAPH-induced oxidation of linoleic acid emulsions. Starch was used as a green reduction/stabilization agent for gold nanoparticles (AuNPs) synthesis in alkaline medium. When tert-butyl hydroperoxide (tert-BHP) was incubated with an excess of iodide ions in a 37 °C water bath for 90 min, triodide (I3-) was formed in an amount equivalent to tert-BHP concentration. Upon the addition of starch-stabilized gold nanoparticles (ss-AuNPs) solution to the incubation mixture, triiodide ions were rapidly adsorbed on the surface of AuNPs and caused their aggregation. A concomitant red shift (from 525 nm to 563 nm) of surface plasmon resonance (SPR) absorption of the nanoparticles was observed, absorbance linearly increasing with aqueous tert-BHP concentration. The method provided an LOD of 39 µM for tert-BHP, and was validated through linearity, precision and accuracy. The concentration of hydroperoxides estimated in linoleic acid peroxidation correlated well with those found by the reference ferric thiocyanate assay. Peroxyl radical scavenger antioxidants decreased the red-shifted SPR absorption of aggregated ss-AuNPs, thereby enabling an indirect estimation of antioxidant activity. This AuNPs-based colorimetric sensor is the first of its kind to directly determine peroxyl radical scavenging activity of polyphenols. The half-maximal inhibitive concentrations (IC50) of selected antioxidant compounds were calculated by utilizing the decrease in absorbance with increasing concentration of scavengers, and compared to those of classical oxygen radical absorbance capacity (ORAC) assay. The proposed nanosensor was superior over FL-based ORAC in determining the peroxyl radical scavenging activity of the lipophilic antioxidant α-tocopherol. The percentage scavenging of real samples such as green tea infusion and synthetic serum were determined. The proposed assay can be used for estimating the peroxyl scavenging of various food and biological samples in terms of its low cost, ease of use and compatibility.

Novel Spectroscopic and Electrochemical Sensors and Nanoprobes for the Characterization of Food and Biological Antioxidants.

Since an unbalanced excess of reactive oxygen/nitrogen species (ROS/RNS) causes various diseases, determination of antioxidants that can counter oxidative stress is important in food and biological analyses. Optical/electrochemical nanosensors have attracted attention in antioxidant activity (AOA) assessment because of their increased sensitivity and selectivity. Optical sensors offer advantages such as low cost, flexibility, remote control, speed, miniaturization and on-site/in situ analysis. Electrochemical sensors using noble metal nanoparticles on modified electrodes better catalyze bioelectrochemical reactions. We summarize the design principles of colorimetric sensors and nanoprobes for food antioxidants (including electron-transfer based and ROS/RNS scavenging assays) and important milestones contributed by our laboratory. We present novel sensors and nanoprobes together with their mechanisms and analytical performances. Our colorimetric sensors for AOA measurement made use of cupric-neocuproine and ferric-phenanthroline complexes immobilized on a Nafion membrane. We recently designed an optical oxidant/antioxidant sensor using N,N-dimethyl-p-phenylene diamine (DMPD) as probe, from which ROS produced colored DMPD-quinone cationic radicals electrostatically retained on a Nafion membrane. The attenuation of initial color by antioxidants enabled indirect AOA estimation. The surface plasmon resonance absorption of silver nanoparticles as a result of enlargement of citrate-reduced seed particles by antioxidant addition enabled a linear response of AOA. We determined biothiols with Ellman reagent-derivatized gold nanoparticles.

Modified Radical Scavenging and Antioxidant Activity Measurement of ß-Carotene with ß-Cyclodextrins Complexation in Aqueous Medium.

In order to evaluate the antioxidant capacity/activity of ß-carotene (BC) in aqueous media, we investigated the inclusion complexes of BC with methyl-ß-cyclodextrin (Me-ß-CD), 2-hydroxypropyl-ß-cyclodextrin (HP-ß-CD) and 2-hydroxyethyl-ß-cyclodextrin (HE-ß-CD) that enhance water solubility and chemical stability. The inclusion complexes (monitored by FTIR) exhibited higher solubility than free BC, and phase solubility studies showed a linear increase in the solubility with the Me-ß-CD concentration. Cupric ion-reducing antioxidant capacity (CUPRAC), ABTS-persulfate, peroxyl and hydroxyl radical scavenging assays were applied. CD-complexed ß-carotene exhibited less effective antioxidative and radical scavenging than free BC dissolved in acetone. ß-Carotene showed the highest antioxidant capacity in the presence of HE-ß-CD, and the lowest with Me-ß-CD, probably due to the deeper and more hydrophobic cavity of the latter. We believe that this is the first report on devising simple spectrophotometric methods for the wholistic assessment of antioxidant activity/capacity, hydroxyl and peroxyl radical scavenging activity of ß-carotene in aqueous solution with CDs.

Identification and Determination of Phenolics in Lamiaceae Species by UPLC-DAD-ESI-MS/MS.

This study reports the phenolic profile screening of aromatic Lamiaceae species such as marjoram (Origanum majorana L.), lavender (Lavandula officinalis) and pennyroyal (Mentha pulegium L.) using a novel and validated ultra performance liquid chromatography method coupled with DAD diode array detector and tandem mass spectrometry (MS/MS) in negative mode of electrospray ionization. Identification and quantification of phenolics in these plant extracts has been realized within 12 min. This method showed good precision (percentage relative standard deviation; RSD% 0.54-2.72 for intra-day, 1.71-4.64 for inter-day), reproducibility (percentage recovery, REC% 92.0-109.0) and linearity (r = 0.9988-0.9999). Limits of detection ranged from 0.02 to 18.2 ng/mL. The extraction of plants was performed using microwave-assisted extraction technique and 60% (v/v) aqueous methanol solvent medium was selected as suitable solvent because of maximum extraction efficiency. Total antioxidant capacity, total phenolic content and free radical scavenging activity of these plant extracts were tested and the results correlated well among each other. According to the Folin assay, phenolic contents of Origanum majorana L., Mentha pulegium L. and Lavandula officinalis were calculated as 119 ± 3.4, 85.1 ± 2.8 and 57.8 ± 2.1 mg GAE/g dry matter, respectively.

Spectroscopic study and antioxidant properties of the inclusion complexes of rosmarinic acid with natural and derivative cyclodextrins.

Measurement of total antioxidant activity/capacity of polyphenols in various solvent media necessitates the use of cyclodextrins to solubilize lipophilic antioxidants of poor aqueous solubility. The inclusion complexes of the slightly water soluble antioxidant, rosmarinic acid (RA), with α-cyclodextrin (α-CD), ß-cyclodextrin (ß-CD), 2-hydroxypropyl-ß-cyclodextrin (HP-ß-CD), 2-hydroxyethyl-ß-cyclodextrin (HE-ß-CD), and methyl-ß-cyclodextrin (M-ß-CD) were investigated for the first time. The effect of cyclodextrins (CDs) on the spectral features of RA was measured in aqueous medium using UV-vis and steady-state fluorescence techniques by varying the concentrations of CDs. The molar stoichiometry of RA-CD inclusion complexes was verified as 1:1, and the formation constants of the complexes were determined from Benesi-Hildebrand equation using fluorescence spectroscopic data. Among the CDs, maximum inclusion ability was measured in the case of M-ß-CD followed by HP-ß-CD, HE-ß-CD, ß-CD and α-CD. Solid inclusion complexes were prepared by freeze drying, and their functional groups were analyzed by IR spectroscopy. Antioxidant capacity of CD-complexed rosmarinic acid was measured to be higher than that of the lone hydroxycinnamic acid by the CUPric Reducing Antioxidant Capacity (CUPRAC) method. The mechanism of the TAC increase was interpreted as the stabilization of the 1-e oxidized o-catechol moiety of RA by enhanced intramolecular H-bonding in a hydrophobic environment provided by CDs, mostly by M-ß-CD.

Determination of antioxidants by a novel on-line HPLC-cupric reducing antioxidant capacity (CUPRAC) assay with post-column detection.

A novel on-line HPLC-cupric reducing antioxidant capacity (CUPRAC) method was developed for the selective determination of polyphenols (flavonoids, simple phenolic and hydroxycinnamic acids) in complex plant matrices. The method combines chromatographic separation, constituent analysis, and post-column identification of antioxidants in plant extracts. The separation of polyphenols was performed on a C18 column using gradient elution with two different mobile phase solutions, i.e., MeOH and 0.2% o-phosphoric acid. The HPLC-separated antioxidant polyphenols in the extracts react with copper(II)-neocuproine (Cu(II)-Nc) reagent in a post-column reaction coil to form a derivative. The reagent is reduced by antioxidants to the copper(I)-neocuproine (Cu(I)-Nc) chelate having maximum absorption at 450 nm. The negative peaks of antioxidant constituents were monitored by measuring the increase in absorbance due to Cu(I)-Nc. The detection limits of polyphenols at 450 nm (in the range of 0.17-3.46 microM) after post-column derivatization were comparable to those at 280 nm UV detection without derivatization. The developed method was successfully applied to the identification of antioxidant compounds in crude extracts of Camellia sinensis, Origanum marjorana and Mentha. The method is rapid, inexpensive, versatile, non-laborious, uses stable reagents, and enables the on-line qualitative and quantitative estimation of antioxidant constituents of complex plant samples.

Solvent effects on the antioxidant capacity of lipophilic and hydrophilic antioxidants measured by CUPRAC, ABTS/persulphate and FRAP methods.

Antioxidants are health beneficial compounds that can protect cells and macromolecules (e.g., fats, lipids, proteins, and DNA) from the damage of reactive oxygen species (ROS). Solvent effect is a crucial parameter on the chemical behaviour of antioxidant compounds but there has been limited information regarding its role on antioxidant capacity and its assays. Therefore, the present study was undertaken to investigate the total antioxidant capacity (TAC) of some certain lipophilic and hydrophilic antioxidants, measured in different solvent media such as ethanol (EtOH) (100%), methanol (MeOH) (100%), methanol/water (4:1, v/v), methanol/water (1:1, v/v), dichloromethane (DCM)/EtOH (9:1, v/v). The cupric reducing antioxidant capacity (CUPRAC) values of selected antioxidants were experimentally reported in this work as trolox equivalent antioxidant capacity (TEAC), and compared to those found by reference TAC assays, i.e., 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid)/persulphate (ABTS/persulphate) and ferric reducing antioxidant power (FRAP) methods. The TAC values of synthetic mixtures of antioxidants were experimentally measured as trolox equivalents and compared to those theoretically found by making use of the principle of additivity of absorbances assuming no chemical interaction between the mixture constituents. Possible synergistic (e.g., BHT and BHA in DCM/EtOH) or antagonistic behaviours of these synthetic mixtures were investigated in relation to solvent selection.

Comparative evaluation of various total antioxidant capacity assays applied to phenolic compounds with the CUPRAC assay.

It would be desirable to establish and standardize methods that can measure the total antioxidant capacity level directly from vegetable extracts containing phenolics. Antioxidant capacity assays may be broadly classified as electron transfer (ET)- and hydrogen atom transfer (HAT)-based assays. The majority of HAT assays are kinetics-based, and involve a competitive reaction scheme in which antioxidant and substrate compete for peroxyl radicals thermally generated through the decomposition of azo compounds. ET-based assays measure the capacity of an antioxidant in the reduction of an oxidant, which changes colour when reduced. ET assays include the ABTS/TEAC, CUPRAC, DPPH, Folin-Ciocalteu and FRAP methods, each using different chromogenic redox reagents with different standard potentials. This review intends to offer a critical evaluation of existing antioxidant assays applied to phenolics, and reports the development by our research group of a simple and low-cost antioxidant capacity assay for dietary polyphenols, vitamins C and E, and human serum antioxidants, utilizing the copper(II)-neocuproine reagent as the chromogenic oxidizing agent, which we haved named the CUPRAC (cupric ion reducing antioxidant capacity) method. This method offers distinct advantages over other ET-based assays, namely the selection of working pH at physiological pH (as opposed to the Folin and FRAP methods, which work at alkaline and acidic pHs, respectively), applicability to both hydrophilic and lipophilic antioxidants (unlike Folin and DPPH), completion of the redox reactions for most common flavonoids (unlike FRAP), selective oxidation of antioxidant compounds without affecting sugars and citric acid commonly contained in foodstuffs and the capability to assay -SH bearing antioxidants (unlike FRAP). Other similar ET-based antioxidant assays that we have developed or modified for phenolics are the Fe(III)- and Ce(IV)-reducing capacity methods.