Oxygen Radical Absorbance Capacity Assay Research Articles

Use of Botanical Ingredients: Nice Opportunities to Avoid Premature Oxidation of NABLABs by Increasing Their ORAC Values Strongly Impacted by Dealcoholization or Pasteurization

Even when fresh, non-alcoholic, and low-alcoholic beers (NABLABs) exhibit significant staling defects due to premature oxidation. In this study, the antioxidant power of eleven fresh commercial NABLABs was assessed by means of three different assays: the oxygen radical absorbance capacity (ORAC), the linoleic acid-induced oxidation (TINH), and the indicator time test (ITT). Only the first two assays, both involving radicalar degradations initiated by AAPH, were found to correlate with each other. NABLABs displayed lower ORAC values than conventional beers (on average, 6127 μmol eq. Trolox/L), except for three samples made with special-colored malts or dry-hopped. Dealcoholization was the step with the greatest impact on the ORAC value (up to a 95% loss) and on flavan-3-ols, sotolon, and polyfunctional thiols, while pasteurization strongly affected color, TBA, and Strecker aldehydes. ORAC assays applied to hop, alternative cereals, and various botanical ingredients indicated that mashing with red sorghum, dry hopping/spicing, and wood maturation could bring the antioxidant power of a NABLAB close to those of conventional beers. With an ORAC value not reached by any other tested botanical ingredient (5234 µmol eq. Trolox/g), African Vernonia amygdalina leaves (traditionally used for Rwandan Ikigage beers) emerged here as the best candidate.

Bay Laurel of Northern Morocco: A Comprehensive Analysis of Its Phytochemical Profile, Mineralogical Composition, and Antioxidant Potential

Laurus nobilis, sometimes referred to as laurel, has been used for medicinal and culinary purposes for a very long time. The main subjects of this study are the phytochemical composition, mineralogical profile, and potential antioxidant properties of Laurus nobilis in Tangier, Northern Morocco. For phytochemical analysis of methanolic extracts, high-performance liquid chromatography (HPLC-UV-MS) was used, and Fourier transformation infrared spectroscopy (FT-IR) was used to identify each individual component. Minerals were studied by inductively coupled plasma atomic emission spectroscopy (ICP-AES) and wavelength dispersive X-ray fluorescence (WD-XRF). Total tannin, flavonoid, and phenolic amounts were quantified using aqueous and methanolic extracts. The antioxidant properties were assessed using 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2′-azino-bis (3ethylbenzothiazoline-6-sulfonic acid) (ABTS), ferric reducing antioxidant power (FRAP), and oxygen radical absorbance capacity (ORAC) assays. Research has revealed a complex array of phytochemicals, including tannins, flavonoids, and phenolic acids. Mineral analysis has revealed the existence of vital components that are beneficial to health. Comparing the methanolic extract to the water extract, it demonstrated higher levels of phenols, flavonoids, and tannins as well as stronger antioxidant activity, indicating greater health benefits. This comprehensive study highlights the importance of Laurus nobilis from Northern Morocco as a reliable botanic resource with potential pharmaceutical, nutritional, and cosmetic uses.

Flow-based bioconjugation of coumarin phosphatidylethanolamine probes: Optimised synthesis and membrane molecular dynamics studies

A series of phosphatidylethanolamine fluorescent probes head-labelled with 3-carboxycoumarin was prepared by an improved bioconjugation approach through continuous flow synthesis. The established procedure, supported by a design of experiment (DoE) set-up, resulted in a significant reduction in the reaction time compared to the conventional batch method, in addition to a minor yield increase. The characterization of these probes was enhanced by an in-depth molecular dynamics (MD) study of the behaviour of a representative probe of this family, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine labelled with 3-carboxycoumarin (POPE-COUM), in bilayers of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC)/1-stearoyl-2-linoleoyl-sn-glycero-3-phosphocholine (SLPC) 2:1, mimicking the composition of the egg yolk lecithin membranes recently used experimentally by our group to study POPE-COUM as a biomarker of the oxidation state and integrity of large unilamellar vesicles (LUVs). The MD simulations revealed that the coumarin group is oriented towards the bilayer interior, leading to a relatively internal location, in agreement with what is observed in the nitrobenzoxadiazole fluorophore of commercial head-labelled NBD-PE probes. This behaviour is consistent with the previously stated hypothesis that POPE-COUM is entirely located within the LUVs structure. Hence, the delay on the oxidation of the probe in the oxygen radical absorbance capacity (ORAC) assays performed is related with the inaccessibility of the probe until alteration of the LUV structure occurs. Furthermore, our simulations show that POPE-COUM exerts very little global and local perturbation on the host bilayer, as evaluated by key properties of the unlabelled lipids. Together, our findings establish PE-COUM as suitable fluorescent lipid analogue probes.

Development of Thai-taste maoberry (Antidesma bunius (L.) Spreng.) vegan sorbet from fruit of different maturities with added inulin: Physicochemical and antioxidant properties

Maoberry (Antidesma bunius (L.) Spreng.) is a tropical fruit popularly consumed with Thai-style chili sugar flakes containing sugar, salt, and chili to enhance its flavour, and reduce its sour and bitter taste. The present work aimed to develop a Thai-taste maoberry vegan sorbet using maoberries at different stages of maturity with added inulin, and to evaluate its physicochemical properties, total phenolic and total flavonoid contents, and antioxidant capacities. Results showed that the three different stages of maoberry maturity caused significant changes on pH, total soluble solids, mixture viscosity, overrun, and firmness of the sorbets. The antioxidant activities assessed using ferric-reducing antioxidant power (FRAP), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging, 3-ethylbenzthiazoline-6-sulphonic acid (ABTS), radical cation decolourisation, and oxygen radical absorbance capacity (ORAC) assays indicated similar trends. Black maoberry sorbet exhibited total flavonoid contents and FRAP values almost twice those of red maoberry sorbet. However, adding inulin significantly increased the melting rate of the product (p < 0.05), but improved the mixture viscosity and overrun, which are important attributes of sorbets.

Evidence of the Relationship between the Oxygen Radical Absorbance Capacity (ORAC) and the Potential Exposure of White Wines to Atypical Aging (ATA).

Atypical aging (ATA) is a well-known wine defect that leads to unacceptable off-odors of wet mop, soap, waxy, furniture varnish, and dish rag. The main compound that is responsible for these undesired scents, 2-aminoacetophenone (AAP), results from the oxidative degradation of 3-indole acetic acid. This study aimed to investigate different technological factors for a possible effect on the oxygen radical absorption capacity (ORAC) and ATA development in white wine. Oxygenation during fermentation did not result in a significant effect on the development of ORAC or ATA development. Instead, the addition of ascorbic acid after vinification led to increased ORAC and decreased AAP formation. The first evidence of a tendency toward a correlation between ORAC and potential ATA development was found, with r = 0.365. This study showed the potential of the ORAC assay for an early assessment of the possible formation of ATA during wine storage.

The combination of allantoin, bisabolol, D-panthenol and dipotassium glycyrrhizinate mitigates UVB-induced PGE2 synthesis by keratinocytes.

Erythema, characterized by the redness of the skin, is a common skin reaction triggered by various endogenous and exogenous factors. This response is often a result of the activation of underlying inflammatory mechanisms within the skin. The objective of this study is to investigate the potential benefits of applying a combination of skincare ingredients, namely allantoin, bisabolol, D-panthenol and dipotassium glycyrrhizinate (AB5D), in the modulation of inflammatory factors associated with erythema. Additionally, the study aims to elucidate the mechanisms by which these ingredients exert their combined actions to alleviate erythema-associated inflammation. Human epidermal keratinocytes were exposed to UVB and subsequently treated with AB5D. Transcriptomics profiling was performed to analyse the dose-response effect of AB5D treatment on keratinocytes. The quantitation of inflammatory mediators, including PGE2 , IL-1α, IL-6, IL-8, IL-1RA and TNFα, was performed on cultured media. Additionally, the oxygen radical absorbance capacity (ORAC) assay was carried out to evaluate the total antioxidant capacity of both individual ingredients and the AB5D combination. To assess the in-vitro antioxidant effects of AB5D against UVB-induced oxidative stress in hTERT keratinocytes, real-time quantitation of mitochondrial superoxide was measured through live-cell imaging. The application of AB5D to UVB-exposed keratinocytes downregulated gene sets associated with inflammatory responses, highlighting the anti-inflammatory properties of AB5D. Specifically, AB5D effectively reduced the production of PGE2 , leading to the downregulation of inflammatory cytokines. Moreover, our findings indicate that AB5D exhibits antioxidative capabilities, functioning as both an antioxidant agent and a regulator of antioxidant enzyme expression to counteract the detrimental effects of cellular oxidative stress. We demonstrated that AB5D can reduce UVB-induced PGE2 , IL-1α, IL-6, IL-8, IL-1RA and TNFα as well as mitochondrial superoxide. These findings suggest that AB5D may alleviate erythema by modulating inflammation via PGE2 and through antioxidation mechanisms.

Influence of Nutrient Deprivation on the Antioxidant Capacity and Chemical Profile of Two Diatoms from Genus Chaetoceros.

The limited availability of phosphate, nitrogen and silicon in the growth media affects the growth, cellular processes, and metabolism of diatoms. Silicon deficiency primarily affects diatom morphology, while phosphate deficiency reduces the production of nucleic acids and phospholipids. Differences in pigment and protein composition are mainly due to nitrogen deficiency. In this study, Chaetoceros socialis and Chaetoceros costatus were cultured under phosphate, nitrogen, and silicon deprivation conditions. The diatom biomass was collected during the stationary growth phase and extracted with 70% ethanol under ultrasonication. The chemical profiles of the extracts were analyzed by high-performance liquid chromatography with high-resolution mass spectrometry with electrospray ionisation (UHPLC-ESI-HRMS), while the antioxidant capacity was determined by 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging and oxygen radical absorbance capacity (ORAC) assays. Pigments, fatty acids, sterols, and derivatives were detected in both species. The total phenolic content in the extracts ranged from 46.25 ± 1.08 to 89.38 ± 6.21 mg of gallic acid equivalent (GAE)/L and from 29.58 ± 1.08 to 54.17 ± 1.18 mg GAE/L. for C. costatus and C. socialis, respectively. Antioxidant activity was higher in C. costatus extracts, especially those obtained from nitrogen-deprived media. The results of this study contribute to the existing knowledge and the ongoing efforts to overcome application and commercialization barriers of microalgae for wide-ranging potential in different industries.

A Kinetic Approach to Oxygen Radical Absorbance Capacity (ORAC): Restoring Order to the Antioxidant Activity of Hydroxycinnamic Acids and Fruit Juices.

This study introduces a kinetic model that significantly improves the interpretation of the oxygen radical absorbance capacity (ORAC) assay. Our model accurately simulates and fits the bleaching kinetics of fluorescein in the presence of various antioxidants, achieving high correlation values (R2 > 0.99) with the experimental data. The fit to the experimental data is achieved by optimizing two rate constants, k5 and k6. The k5 value reflects the reactivity of antioxidants toward scavenging peroxyl radicals, whereas k6 measures the ability of antioxidants to regenerate oxidized fluorescein. These parameters (1) allow the detailed classification of cinnamic acids based on their structure-activity relationships, (2) provide insights into the interaction of alkoxyl radicals with fluorescein, and (3) account for the regeneration of fluorescein radicals by antioxidants. The application of the model to different antioxidants and fruit extracts reveals significant deviations from the results of traditional ORAC tests based on the area under the curve (AUC) approach. For example, lemon juice, rich in 'fast' antioxidants such as ascorbic acid, shows a high k5 value, in contrast to its low AUC values. This finding underscores the limitations of the AUC approach and highlights the advantages of our kinetic model in understanding antioxidative dynamics in food systems. This study presents a comprehensive, quantitative, mechanism-oriented approach to assessing antioxidant reactivity, demonstrating a significant improvement in ORAC assay applications.

Chemical and biological characterization of Melaleuca subulata (Cheel) Craven leaves’ volatile constituents supported by chemometric analysis and molecular docking

BackgroundThe genus Melaleuca (Myrtaceae) comprises dozens of essential oil (EO)-rich species that are appreciated worldwide for their various medicinal values. Additionally, they are renowned in traditional medicine for their antimicrobial, antifungal, and other skin-related activities. The current study investigated the chemical profile and skin-related activities of volatile constituents derived from M. subulata (Cheel) Craven (Synonym Callistemon subulatus) leaves cultivated in Egypt for the first time.MethodsThe volatile components were extracted using hydrodistillation (HD), headspace (HS), and supercritical fluid (SF). GC/MS and Kovat’s retention indices were implemented to identify the volatile compounds, while the variations among the components were assessed using Principal Component Analysis and Hierarchical Cluster Analysis. The radical scavenging activity was assessed using 2,2-diphenyl-1-picrylhydrazyl (DPPH), oxygen radical absorbance capacity (ORAC) and β-carotene assays. Moreover, the anti-aging effect was evaluated using anti-elastase, and anti-collagenase, while the antimicrobial potential was deduced from the agar diffusion and broth microdilution assays. Lastly, the molecular docking study was executed using C-docker protocol in Discovery Studio 4.5 to rationalize the binding affinity with targeted enzymes.ResultsThe SF extraction approach offered the highest EO yield, being 0.75%. According to the GC/MS analysis, monoterpene hydrocarbons were the most abundant volatile class in the HD oil sample (54.95%), with α-pinene being the most copious component (35.17%). On the contrary, the HS and SF volatile constituents were pioneered with oxygenated monoterpenes (72.01 and 36.41%) with eucalyptol and isopulegone being the most recognized components, representing 67.75 and 23.46%, respectively. The chemometric analysis showed segregate clustering of the three extraction methods with α-pinene, eucalyptol, and isopulegone serving as the main discriminating phytomarkers. Concerning the bioactivity context, both SF and HD-EOs exhibited antioxidant effects in terms of ORAC and β-carotene bleaching. The HD-EO displayed potent anti-tyrosinase activity, whereas the SF-EO exhibited significant anti-elastase properties. Moreover, SF-EO shows selective activity against gram-positive skin pathogens, especially S. aureus. Ultimately, molecular docking revealed binding scores for the volatile constituents; analogous to those of the docked reference drugs.ConclusionsM. subulata leaves constitute bioactive volatile components that may be indorsed as bioactive hits for managing skin aging and infection, though further in vivo studies are recommended.

Bioaccessibility and Digestibility of Proteins in Plant-Based Drinks and Cow's Milk: Antioxidant Potential of the Bioaccessible Fraction.

During the last years, a strong increase in the sales volume and consumption of plant-based drinks was observed, which were partly used as an alternative to cow's milk. As milk is a relevant protein source in many countries, we have investigated the protein bioaccessibility and digestibility of soy, almond, and oat drinks in comparison to milk using the tiny-TIMsg gastrointestinal model. The relative protein digestibility of all products was between 81% (soy drink) and 90% (milk). The digestible indispensable amino acid score (DIAAS) in vitro method was used to estimate the protein nutritional quality. The highest DIAAS values were obtained for milk in tryptophan (117%) and soy drink in sulfur containing amino acids (100%). Oat drink was limited in lysine (73%), almond drink in lysine (34%) and the sulfur containing amino acids (56%). Additionally, the antioxidant activity of the bioaccessible fractions was analyzed using Trolox equivalent antioxidative capacity and oxygen radical absorbance capacity assays, revealing a higher antioxidative potential of milk and soy drink compared to oat and almond drink.

Antioxidant Capacity of Free and Peptide Tryptophan Residues Determined by the ORAC (Oxygen Radical Absorbance Capacity) Assay Is Modulated by Radical-Radical Reactions and Oxidation Products.

The ORAC (Oxygen Radical Absorbance Capacity) assay is commonly employed for determining the antioxidant capacity of bioactive peptides. To gain insights into the meaning of this index for peptides containing a single Trp, we studied the consumption of this residue and fluorescein (FLH, the probe of ORAC method), induced by radicals generated by AAPH (2,2'-Azo-bis(2-amidinopropane) dihydrochloride) thermolysis. ORAC values were rationalized from kinetics and computational calculations of bond dissociation energies (BDE) of the N-H bond (indole ring of Trp). Free Trp, di- and tri- peptides, and three larger peptides were studied. Solutions containing 70 nM FLH, 1-5 μM free Trp or peptides, and 10 mM AAPH were incubated at 37 °C in phosphate buffer. Kinetic studies showed that FLH minimally affected Trp consumption. However, a clear protection of FLH, characterized by pseudo-lag times, was evidenced, reflecting radical-radical reactions and FLH repairing. Peptides showed similar ORAC values (~1.9-2.8 Trolox equivalents), while BDE varied between 91.9 and 103.5 kcal. These results, added to the protection of FLH observed after total consumption of Trp, indicate a lack of discrimination of the assay for the chemical structure of peptides and the contribution of oxidation products to the index.

Enhancing the physicochemical stability and antioxidant activity of cape gooseberry calyx extract through nanoencapsulation in soy lecithin liposomes

The focus of this study was on the development, physicochemical characterisation and evaluation of the antioxidant activity of cape gooseberry calyx extract loaded into nanoliposomal systems. Various nanoliposomes were prepared and optimised using the ethanol injection method and characterised based on particle size, polydispersity and zeta potential measurements. Subsequently, the encapsulation efficiency and in vitro release profile of the natural antioxidant extract (NAE) were evaluated, and its antioxidant activity was assessed using the oxygen radical absorbance capacity assay. The results revealed that NAE-loaded nanoliposomes described desired quality features (e.g., particle size of < 200 nm, polydispersity index of < 0.3, zeta potential of > −40 mV and encapsulation efficiency of ∼70%). Furthermore, it was found that NAE release is controlled by various stages, and its antioxidant activity improves by around 30% when loaded into the nanoliposomes, suggesting that it could be a promising antioxidant functional raw material.

Physicochemical, Antioxidant, and Antimicrobial Properties of Three Medicinal Plants from the Western Part of the Rhodope Mountains, Bulgaria.

The present study examined the physicochemical, antioxidant, and antimicrobial properties of three medicinal plants: thyme (Thymus callieri Borbás ex Velen), cotton thistle (Onopordum acanthium L.), and hawthorn fruit (Crataegus monogyna Jacq.) from the Western Rhodope Mountains, Bulgaria. The first stage determined the physicochemical characteristics (moisture, ash, carbohydrates, proteins, and vitamin C) of the three herbs. The second stage investigated four types of extracts (aqueous, oil, methanolic, and ethanolic) of each herb and evaluated their total phenolic content, the presence of phenolic compounds (flavonoids and phenolic acids), their antioxidant activity, and antimicrobial properties. Thyme was characterised by the highest ash, protein, and vitamin C content (6.62%, 11.30%, and 571 mg/100 g, respectively). Hawthorn fruit showed the highest moisture and carbohydrate content (8.50% and 4.20%, respectively). The 70% ethanolic extracts of the three herbs exhibited the highest levels of phenolic compounds and, consequently, pronounced antioxidant activity, compared to the other three types of extracts. The aqueous, oil, methanolic, and ethanolic thyme extracts demonstrated the highest total phenolic content-TPC (27.20 mg GAE/g, 8.20 mg GAE/g, 31.70 mg GAE/g, and 310.00 mg GAE/g, respectively), compared to the extracts of the other two plants. These results were consistent with the highest antioxidant activity of the thyme extracts determined using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay, the oxygen radical absorbance capacity (ORAC) assay, and the hydroxyl radical averting capacity (HORAC) assay (except for the oil extract examined using the DPPH method). The results from the high-performance liquid chromatography (HPLC) analysis revealed that the flavonoid quercetin-3-ß-glucoside had the highest concentration in thyme (374.5 mg/100 g), while myricetin dominated in the cotton thistle (152.3 mg/100 g). The phenolic acid content analysis showed prevalence of rosmaric acid in the thyme (995 mg/100 g), whereas chlorogenic acid was detected in the highest concentration in the cotton thistle and hawthorn fruit (324 mg/100 g and 27.7 mg/100 g, respectively). The aqueous, methanolic, and ethanolic extracts showed moderate to high antibacterial potential but limited antifungal activity. None of the oil extracts inhibited the test microorganisms used in the study.

Evaluation of Crocin Content and In Vitro Antioxidant and Anti-Glycation Activity of Different Saffron Extracts.

Crocin, a glycoside carotenoid that exhibits several health benefits, is mainly obtained from saffron (Crocus sativus L.), whose quality and content of phytochemicals can be strongly affected by environmental conditions. Therefore, in this work, the crocin content and in vitro antioxidant activity of saffron extracts obtained from three different varieties (Greek, Sicilian, and Iranian saffron) were assessed. Crocin content in saffron extracts was quantified via ultra-performance liquid chromatography coupled with mass spectrometry. The antioxidant activity of saffron extracts was evaluated using the oxygen radical absorbance capacity (ORAC) assay and nitric oxide (NO) radical scavenging test. The Maillard reaction was used to assess anti-glycation activity. Although the Sicilian and Iranian saffron extracts contained higher amounts of crocin (128 ± 6 ng/mL and 126 ± 4 ng/mL, respectively) compared to the Greek extracts (111 ± 2 ng/mL), ORAC values (50.9 ± 0.5) and % NO inhibition (35.2 ± 0.2) were higher for the Greek variety, which displayed a total phenolic content about two-fold greater than that of the other two extracts. Sicilian and Greek saffron had similar anti-glycation activities, while Iranian saffron was less effective. These results suggest that the antioxidant activity of saffron extracts could be ascribed to their naturally occurring complex mixture of phytochemicals, deserving further investigation as supplements to prevent pathological conditions induced by radical species.

Optimization of pressurized ethanol extraction for efficient recovery of hyperoside and other valuable polar antioxidant-rich extracts from Betula pendula Roth leaves

Silver birch (Betula pendula Roth) leaves, known for their long-standing uses in traditional medicine, are bioactive compound-rich by-products of the birch logging industry, abundantly available throughout the northern hemisphere. Considering the current demands for the sustainable valorization of various plant materials and their processing by-products, this study aimed to optimize pressurized ethanol extraction (PLE-EtOH) parameters in order to produce hyperoside and other valuable polar antioxidant-rich extracts from silver birch leaves. A central composite design along with response surface methodology (CCD-RSM) was used to optimize the temperature and time of PLE-EtOH for yield, total phenolic (TPC) and flavonoid (TFC) content, Trolox equivalent antioxidant capacity (TEAC) measured in oxygen radical absorbance capacity (ORAC), cupric ion reducing antioxidant capacity (CUPRAC), and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) assays, and hyperoside content measured by ultraperformance liquid chromatography-electrospray tandem mass spectrometry (UPLC-ESI/MS2) analysis. The optimal PLE-EtOH conditions were 86 °C and 39 min, which produced an extract with 32.2% yield, high TPC (97.3 mg gallic acid equivalents/g), TFC (37.3 mg quercetin equivalents/g) and proanthocyanidin (301.2 mg catechin hydrate equivalents/g) content, also strong in vitro antioxidant capacity in ORAC, CUPRAC and ABTS assays (999.3, 297.0 and 392.3 mg Trolox equivalents/g, respectively). Hyperoside (54.1 mg/g), betuloside (18.3 mg/g) and chlorogenic acid (11.5 mg/g) were the prime phenolic compounds in the PLE-EtOH extract. Additionally, a high sun protection factor (37.4 at 1 mg/mL) was determined by in vitro method for this extract. Such products from B. pendula leaves have potential cosmetical, nutraceutical, and pharmaceutical applications.

Liver protective effect of chloroform extract of Bauhinia purpurea leaves is attributed partly to its antioxidant action and the presence of flavonoids

Context Bauhinia purpurea L. (Fabaceae) is used in the Ayurvedic system to treat various oxidative-related ailments (e.g., wounds, ulcers etc.). Therefore, it is believed that the plant also has the potential to alleviate oxidative-related liver damage. Objective This study elucidates the hepatoprotective activity of chloroform extract of B. purpurea leaves (CEBP) in paracetamol (PCM)-induced liver injury (PILI) rats. Materials and methods Male Sprague-Dawley rats (n = 6) were pre-treated once daily (p.o.) with CEBP (50–500 mg/kg) for seven consecutive days before being administered (p.o.) a hepatotoxic agent, 3 g/kg PCM. Liver enzyme levels were determined from the collected blood, while the collected liver was used to determine the activity of endogenous antioxidant enzymes and for histopathological examination. CEBP was also subjected to radical scavenging assays and phytochemical analysis. Results CEBP significantly (p < 0.05) reversed the toxic effect of PCM by increasing the serum levels of AST and ALT, and the activity of endogenous catalase (CAT) and superoxide dismutase (SOD) while reducing the liver weight/body weight (LW/BW) ratio. Other than low TPC value and radical scavenging activity, CEBP had a high antioxidant capacity when evaluated using the oxygen radical absorbance capacity (ORAC) assay. UHPLC-ESI-MS analysis of CEBP showed the presence of flavonoids. Discussion and conclusions CEBP exerts its hepatoprotective activity through a non-free radical scavenging pathway that involves activation of the endogenous enzymatic antioxidant defense system. Further study is needed to identify the responsible bioactive compounds before the plant can be developed as a future alternative hepatoprotective medicament for clinical use.

Reverse ethnopharmacological-guided study of Aristolochia trilabiata leaves - unveiling its antioxidant potential and chemical profile

Oxidative stress, a contributing factor to numerous diseases such as inflammation, has sparked increased interest in the search for natural antioxidants known for their biodegradability and low toxicity. In this study, we investigated Aristolochia trilabiata, a plant native to the Brazilian Amazon, as a potential source of bioactive compounds. Using the "reverse ethnopharmacology" method, we collected valuable information on the medicinal uses of A. trilabiata. A total of 15 families were interviewed, and remarkably 40% reported medicinal uses of decoction and infusion of its leaves mainly to treat inflammation, pain, and urinary issues. Otherwise, 27% were aware without details, and 33% had no knowledge about the species. Since inflammation was described as medicinal use, our study focused on the antioxidant capacities of the aqueous extract (ExtH2O) and its methanolic (FrMeOH) and aqueous (FrH2O) fractions from the leaves of A. trilabiata, associated with their molecular profiles. Thus, the antioxidant capacities of aqueous extract and its fractions were evaluated by oxygen radical absorbance capacity (ORAC) assays. The molecular profile was assessed via Liquid chromatography-mass spectrometry (LC-MS/MS) and nuclear magnetic resonance (NMR) analysis. As a result, FrMeOH exhibited a potent antioxidant capacity (4.94 mmol TE/g), confirmed promising ability against oxygen-derived free radicals, and it was associated mainly with eriodictyol and blumenol C-derived compounds, which beyond potent antioxidant effects, could be associated with anti-inflammatory properties, making A. trilabiata a promising natural source of bioactive that opens exciting avenues for further research and utilization in medicine and natural product discovery.

Antioxidant peptides generated from chicken feet protein hydrolysates.

As major industrial poultry by-products, chicken feet are considered as notable sources of several bioactive molecules. The current work covers the processing of chicken feet proteins as substrates to be hydrolysed by combinations of three commercial enzymes (Alcalase®, Flavourzyme® and Protana® Prime) during different hydrolysis periods and the evaluation of the identified peptides having antioxidant activity after simulated gastrointestinal digestion. Enzymatic hydrolysis with Alcalase® and Protana® Prime combination for 4 h resulted in the highest activities. Reversed-phase high-performance liquid chromatographic separation of the purified hydrolysate yielded three active fractions that were further identified by nano-liquid chromatography-tandem mass spectrometry. The bioactivities of over 230 identified peptide sequences were estimated after simulated gastrointestinal digestion, and those peptides with the highest chance of exerting antioxidant activity were selected to be further synthesised and tested. In this sense, the synthesised dipeptides CF and GY showed the highest antioxidant capacity. CF presented IC50 values of 69.63 and 145.41 μmol L-1 in 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and oxygen radical absorbance capacity (ORAC) assays, respectively. In contrast, GY IC50 values were 15.27 and 10.06 μmol L-1 in ABTS and ORAC assays, respectively. Significant differences (P < 0.05) were registered between peptides in the same antioxidant assays. Overall, the findings emphasised the favourable impact of enzymatic hydrolysis with the obtaining of antioxidant peptides from poultry by-products that could be evaluated as a safe and economical source to retard oxidation in food systems. © 2023 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Fluorescent head-labelled phospholipid coumarin bioconjugate as a chemical reporter for antioxidant protection in liposomes

The oxygen radical absorbance capacity (ORAC) assay is considered a valuable analytical tool for measuring the protection afforded by an antioxidant compound to a target probe. Fluorescein is the classical probe used in the ORAC assays. In addition to not being a biological target for oxidation, fluorescein acts as a free molecule within the reaction media. Moreover, the probe specific location is the aqueous medium where its oxidation occurs. Hence, fluorescein is not ideal for inferring about the interactions of radicals with biomimetic membrane models within the liquid-lipid interface or deeper in the bilayer system.The aim of the present work was to explore the use of POPE-COUM probe, a 2-oleoyl-1-palmitoyl-sn-glycero-3-phosphoethanolamine labelled with 3-carboxycoumarin, to report the protection of antioxidants interacting with biomembranes using liposomes as a biomimetic model. The probe was obtained applying an improved bioconjugation by using continuous flow synthesis to afford the desired compound in just 5 min in 80% yield. The lipid peroxidation induced by 2,2′-azobis(2-methylpropionamidine) dihydrochloride (AAPH) derived radicals was monitored directly by the fluorescence intensity decay of the POPE-COUM, without resorting to any other reporter, in large unilamellar vesicles (LUVs). The results obtained showed a delay on the oxidation of the probe compared to aqueous phase fluorescein which might indicate that POPE-COUM is not accessible to the attack by the AAPH-derived radicals and is protected towards the inside of the vesicle. Hence, the access to the embedded probe only happens when alteration of the LUV structure occurs. Fluorescence lifetime imaging microscopy studies support the fact that the fluorescent POPE-COUM probe is restricted to the lipidic layers, and no background fluorescence was observed.Ultimately, these outcomes represent the proof-of-concept for the use of POPE-COUM in ORAC assays and its suitability as biomarker of the oxidation state and integrity of cellular membrane biomimetic systems such as liposomes.

Chaya (Cnidoscolus aconitifolius (Mill.) I.M. Johnst) leaf extracts regulate mitochondrial bioenergetics and fatty acid oxidation in C2C12 myotubes and primary hepatocytes

Ethnopharmacological relevanceChaya (Cnidoscolus aconitifolius (Mill.) I.M. Johnst) is an important component of the regular diet and traditional medicine of indigenous communities in Mexico. Customarily, Chaya is consumed as a beverage made of macerated leaf, cooked, or prepared in teas or infusions to empirically treat obesity, diabetes, gastrointestinal disorders, and kidney stones. The beneficial effects of Chaya can be attributed to the presence of protein, dietary fiber, vitamins, and especially polyphenols, which regulate mitochondrial function. Therefore, polyphenols present in Chaya extracts could be used to develop novel strategies to prevent and treat metabolic alterations related to mitochondrial dysfunction in the muscle and liver of subjects with obesity, type 2 diabetes, and cardiovascular diseases. However, limited information is available concerning the effect of Chaya extracts on mitochondrial activity in those tissues. Aim of the studyThe aim of this study was to evaluate the antioxidant capacity of an aqueous extract (AE) or mixed (methanol/acetone/water) extract (ME) of Chaya leaf and their effect on C2C12 myotubes and primary hepatocyte mitochondrial bioenergetics and fatty acid oxidation (FAO). Materials and methodsTotal polyphenol content and antioxidant activity were determined using the Folin–Ciocalteu method and the oxygen radical absorbance capacity assay, respectively. The effect of AE and ME from Chaya leaf on mitochondrial activity and FAO of C2C12 myotubes and primary hepatocytes was evaluated using an extracellular flux analyzer. ResultsThe AE and ME from Chaya leaf exhibited antioxidant activity and a polyphenol content similar to nopal, another plant used in Mexican traditional medicine. AE significantly (p < 0.05) decreased the maximal respiration and spare respiratory capacity (SRC) of C2C12 cells, whereas ME had little effect on C2C12 mitochondrial function. Conversely, ME significantly (p < 0.05) decreased SRC in primary hepatocytes, whereas AE increased maximal respiration and SRC at low doses (5 and 10 μM). Moreover, low doses of Chaya AE significantly (p < 0.05) increased AMPK phosphorylation, acyl-coenzyme A oxidase protein abundance, and palmitate oxidation in primary hepatocytes. ConclusionThe AE of Chaya leaf increases mitochondrial function and FAO of primary hepatocytes, indicating its potential to treat hepatic mitochondrial dysfunction underlying metabolic diseases.