Exploring the bioactive potential of algae residue extract via subcritical water extraction: Insights into chemical composition and biological activity.

Gelidium sesquipedale is valued in the Spanish agar industry, but its production generates substantial waste, often discarded despite its nutritional and bioactive content. Subcritical water extraction (SWE) at 175 °C and 50 bar for 130 min was performed on this waste after agar extraction, comparing it to conventional ethanol extraction. The SWE extract exhibited superior nutritional profile, including proteins (170.6 ± 1.0 mg/gfreeze-dried-extract), essential amino acids (18.1%), carbohydrates (148.1 ± 0.3 mg/gfreeze-dried-extract), total phenolic content (57 ± 7 mg-EqGA/gfreeze-dried-extract), and also containing Maillard reaction compounds, such as 5-hydroxymethylfurfural, furfural, 2-furanmethanol, 1-(2-furanyl)-ethanone, and 5-methyl-2-furfural, influencing color, aroma and flavor. This extract showed better antioxidant and anti-inflammatory properties than the conventional extract, and higher xanthine oxidase, tyrosinase, and acetylcholinesterase inhibition activities. Toxicological assessment on human cells indicated the safety of the SWE extract. Therefore, SWE technology offers a promising method to valorize G. sesquipedale residue, yielding a bioactive and nutrient-rich extract suitable for food and nutraceutical applications.

Onion (Allium cepa L.) Skin Waste Valorization: Unveiling the Phenolic Profile and Biological Potential for the Creation of Bioactive Agents through Subcritical Water Extraction.

Onion skin waste (OSW), the primary non-edible byproduct from onion processing, offers a renewable source of bioactive compounds. This study aims to valorize OSW through subcritical water extraction (SWE), aligning with a circular economy and biorefinery principles. SWE was carried out at 145 °C and 50 bar for 50 min in a discontinuous reactor, producing a phenolic-rich extract (32.3 ± 2.6 mg/g) dominated by protocatechuic acid (20.3 ± 2.5 mg/g), quercetin-4'-O-glucoside (7.5 ± 0.2 mg/g), and quercetin (3.2 ± 0.6 mg/g). Additionally, the extract contains sugars (207.1 ± 20.3 mg sucrose-Eq/g), proteins (22.8 ± 1.6 mg BSA-Eq/g), and free amino acids (20.4 ± 1.2 mg arginine-Eq/g). Its phenolic richness determines its scavenging activity against ●NO and O2●- radicals and its α-glucosidase and aldose-reductase inhibition without affecting α-amylase. Notably, the extract demonstrates significant α-glucosidase inhibition (IC50 = 75.6 ± 43.5 µg/mL), surpassing acarbose (IC50 = 129.5 ± 1.0 µg/mL) in both pure enzyme and cell culture tests without showing cytotoxicity to AGS, HepG2, and Caco-2 human cell lines. The extract's bioactivity and nutritional content make it suitable for developing antioxidant and antidiabetic nutraceutical/food components, highlighting SWE's potential for OSW valorization without using organic solvents.

Mitochondrial Dysfunction in Skeletal Muscle of Rotenone-Induced Rat Model of Parkinson's Disease: SC-Nanophytosomes as Therapeutic Approach.

The development of new therapeutic options for Parkinson's disease (PD) requires formulations able to mitigate both brain degeneration and motor dysfunctions. SC-Nanophytosomes, an oral mitochondria-targeted formulation developed with Codium tomentosum membrane polar lipids and elderberry anthocyanin-enriched extract, promote significant brain benefits on a rotenone-induced rat model of PD. In the present work, the effects of SC-Nanophytosome treatment on the skeletal muscle tissues are disclosed. It is unveiled that the rotenone-induced PD rat model exhibits motor disabilities and skeletal muscle tissues with deficient activity of mitochondrial complexes I and II along with small changes in antioxidant enzyme activity and skeletal muscle lipidome. SC-Nanophytosome treatment mitigates the impairment of complexes I and II activity, improving the mitochondrial respiratory chain performance at levels that surpass the control. Therefore, SC-Nanophytosome competence to overcome the PD-related motor disabilities should be also associated with its positive outcomes on skeletal muscle mitochondria. Providing a cellular environment with more reduced redox potential, SC-Nanophytosome treatment improves the skeletal muscle tissue's ability to deal with oxidative stress stimuli. The PD-related small changes on skeletal muscle lipidome were also counteracted by SC-Nanophytosome treatment. Thus, the present results reinforces the concept of SC-Nanophytosomes as a mitochondria-targeted therapy to address the neurodegeneration challenge.

Antifungal Activity of Guiera senegalensis: From the Chemical Composition to the Mitochondrial Toxic Effects and Tyrosinase Inhibition.

Pest resistance against fungicides is a widespread and increasing problem, with impact on crop production and public health, making the development of new fungicides an urgent need. Chemical analyses of a crude methanol extract (CME) of Guiera senegalensis leaves revealed the presence of sugars, phospholipids, phytosterols, guieranone A, porphyrin-containing compounds, and phenolics. To connect chemical composition with biological effects, solid-phase extraction was used to discard water-soluble compounds with low affinity for the C18 matrix and obtain an ethyl acetate fraction (EAF) that concentrates guieranone A and chlorophylls, and a methanol fraction (MF) dominated by phenolics. While the CME and MF exhibited poor antifungal activity against Aspergillus fumigatus, Fusarium oxysporum and Colletotrichum gloeosporioides, the EAF demonstrated antifungal activity against these filamentous fungi, particularly against C. gloeosporioides. Studies with yeasts revealed that the EAF has strong effectiveness against Saccharomyces cerevisiae, Cryptococcus neoformans and Candida krusei with MICs of 8, 8 and 16 µg/mL, respectively. A combination of in vivo and in vitro studies shows that the EAF can function as a mitochondrial toxin, compromising complexes I and II activities, and as a strong inhibitor of fungal tyrosinase (Ki = 14.40 ± 4.49 µg/mL). Thus, EAF appears to be a promising candidate for the development of new multi-target fungicides.

An Experimental Approach to Address the Functional Relationship between Antioxidant Enzymes and Mitochondrial Respiratory Complexes.

Mitochondrial dysfunction and cytosolic oxidative stress are pathological biomarkers interlinked in several chronic diseases and cellular toxicity promoted by high-energy radiation or xenobiotics. Thus, assessing the activities of the mitochondrial redox chain complexes and the cytosolic antioxidant enzymes in the same cell culture system is a valuable approach to addressing the challenge of chronic diseases or unveiling the molecular mechanisms underlying the toxicity of physical and chemical stress agents. The present article gathers the experimental procedures to obtain, from isolated cells, a mitochondria-free cytosolic fraction and a mitochondria-rich fraction. Furthermore, we describe the methodologies to evaluate the activity of the main antioxidant enzymes in the mitochondria-free cytosolic fraction (superoxide dismutase, catalase, glutathione reductase and glutathione peroxidase), and the activity of the individual mitochondrial complexes I, II and IV, as well as the conjugated activity of complexes I-III and complexes II-III in the mitochondria-rich fraction. The protocol to test the citrate synthase activity was also considered and used to normalize complexes. The procedures were optimized within an experimental setup to allow that each condition to be tested only requires sampling of one T-25 flask of cells 2D cultured, as the typical results presented and discussed here.

The administration of methyl and butyl parabens interferes with the enzymatic antioxidant system and induces genotoxicity in rat testis: possible relation to male infertility.

Parabens are esters of p-hydroxybenzoic acid, used for decades as a preservative in many products, including agrochemicals, pharmaceuticals, foods and cosmetics. Concerns regarding parabens toxicity include adverse effects on endocrine activity, carcinogenesis, infertility, spermatogenesis, and adipogenesis. The present study aimed to investigate the in vivo administration of methyl and butylparaben at concentrations of 100 and 200 mg/kg body weight, by subcutaneous injection, in variable murinometric measurements, antioxidant systems and genotoxicity. The administration of parabens did not affect the consumption of water and food. However, there was a decrease in the weight of the testes and the seminal vesicle (p < 0.05). The administration of parabens caused an increase in superoxide dismutase for methylparaben (200 mg/kg) and both concentrations of butylparaben (p < 0.05). Catalase showed increased activity in all groups treated with parabens. In contrast, glutathione reductase and glutathione S-transferase suffered a decrease in the groups treated with both parabens. These results show that parabens, especially butyl, can affect the rat testis enzymatic antioxidant system, decreasing the cellular antioxidant capacity, which was confirmed by the decrease in the glutathione reducing power, expressed by the reduced glutathione/oxidized glutathione ratio. Therefore, an increase in lipid peroxidation was observed, which was significant in the case of butyl. Genetic Damage Indicator values show that butylparaben treatments displayed significantly higher values than the control. This study shows for the first time that parabens can induce genotoxicity in the rat male reproductive organ.

Anti-Inflammatory Activity of Olive Oil Polyphenols-The Role of Oleacein and Its Metabolites.

The anti-inflammatory potential of oleacein, the main polyphenolic compound found in olive oil, and its main metabolites were characterized by their effects on RAW 264.7 macrophages challenged with lipopolysaccharide (LPS), and by their ability to inhibit enzymes of the arachidonic acid metabolism with a key role in the synthesis of pro-inflammatory lipid mediators. Oleacein at 12.5 µM significantly decreased the amount of L-citrulline and ●NO generated by LPS-stimulated macrophages. Hydroxytyrosol, hydroxytyrosol acetate and hydroxytyrosol acetate sulfate were also able to reduce the cellular amount of ●NO, although to a lesser extent. In contrast, hydroxytyrosol glucuronide and sulfate did not show detectable effects. Oleacein was also able to inhibit the coupled PLA2 + 5-LOX enzyme system (IC50 = 16.11 µM), as well as the 5-LOX enzyme (IC50 = 45.02 µM). Although with lower activity, both hydroxytyrosol and hydroxytyrosol acetate were also capable of inhibiting these enzymes at a concentration of 100 µM. None of the other tested metabolites showed a capacity to inhibit these enzymes. In contrast, all compounds, including glucuronides and sulfate metabolites, showed a remarkable capacity to inhibit both cyclooxygenase isoforms, COX-1 and COX-2, with IC50 values lower than 3 µM. Therefore, oleacein and its metabolites have the ability to modulate ●NO- and arachidonic acid-dependent inflammatory cascades, contributing to the anti-inflammatory activity associated with olive oil polyphenols.

Brain Effects of SC-Nanophytosomes on a Rotenone-Induced Rat Model of Parkinson's Disease-A Proof of Concept for a Mitochondria-Targeted Therapy.

Mitochondria are an attractive target to fight neurodegenerative diseases due to their important functions for cells and the particularly close relationship between the functional connectivity among brain regions and mitochondrial performance. This work presents a mitochondria-targeted therapy designed to modulate the functionality of the mitochondrial respiratory chain and lipidome, parameters that are affected in neurodegeneration, including in Parkinson's disease (PD). This therapy is supported by SC-Nanophytosomes constructed with membrane polar lipids, from Codium tomentosum, and elderberry anthocyanin-enriched extract, from Sambucus nigra L. SC-Nanophytosomes are nanosized vesicles with a high negative surface charge that preserve their properties, including anthocyanins in the flavylium cation form, under conditions that mimic the gastrointestinal tract pH changes. SC-Nanophytosomes, 3 µM in phospholipid, and 2.5 mg/L of EAE-extract, delivered by drinking water to a rotenone-induced PD rat model, showed significant positive outcomes on disabling motor symptoms associated with the disease. Ex vivo assays were performed with two brain portions, one comprising the basal ganglia and cerebellum (BG-Cereb) and the other with the cerebral cortex (C-Cortex) regions. Results showed that rotenone-induced neurodegeneration increases the α-synuclein levels in the BG-Cereb portion and compromises mitochondrial respiratory chain functionality in both brain portions, well-evidenced by a 50% decrease in the respiratory control rate and up to 40% in complex I activity. Rotenone-induced PD phenotype is also associated with changes in superoxide dismutase and catalase activities that are dependent on the brain portion. Treatment with SC-Nanophytosomes reverted the α-synuclein levels and antioxidant enzymes activity to the values detected in control animals. Moreover, it mitigated mitochondrial dysfunction, with positive outcomes on the respiratory control rate, the activity of individual respiratory complexes, and the fatty acid profile of the membrane phospholipids. Therefore, SC-Nanophytosomes are a promising tool to support mitochondria-targeted therapy for neurodegenerative diseases.

Mitochondria research and neurodegenerative diseases: On the track to understanding the biological world of high complexity.

From the simple unicellular eukaryote to the highly complex multicellular organism like Human, mitochondrion emerges as a ubiquitous player to ensure the organism's functionality. It is popularly known as "the powerhouse of the cell" by its key role in ATP generation. However, our understanding of the physiological relevance of mitochondria is being challenged by data obtained in different fields. In this review, a short history of the mitochondria research field is presented, stressing the findings and questions that allowed the knowledge advances, and put mitochondrion as the main player of safeguarding organism life as well as a key to solve the puzzle of the neurodegenerative diseases.

Trichilia catigua and Turnera diffusa phyto-phospholipid nanostructures: Physicochemical characterization and bioactivity in cellular models of induced neuroinflammation and neurotoxicity.

Flavonoid-based therapies supported by nanotechnology are considered valuable strategies to prevent or delay age-related and chronic neurodegenerative disorders. Egg yolk phospholipids were combined with flavonoid-rich extracts obtained from Trichilia catigua A.Juss. (rich in flavan-3-ols and phenylpropanoid derivatives) or Turnera diffusa Willd. ex Schult (dominated by luteolin derivatives) to prepare nanophytosomes. The nanophytosomes showed that size and surface charge of the lipid-based vesicles are dependent of their phenolic composition. In vitro assays with SH-SY5Y cells showed that both formulations protect cells from glutamate-induced toxicity, but not from 6-hydroxydopamine/ascorbic acid. T. diffusa nanophytosomes promote a decrease of nitric oxide produced by BV-2 cells stimulated with interferon-γ. Nanophytosomes dialysed against a mannitol solution, and then lyophilised, allow to obtain freeze-dried products that after re-hydration preserve the essential physicochemical features of the original formulations, and exhibit improved colloidal stability. These results indicate that these flavonoid/phospholipid-based nanophytosomes have suitable features to be considered as tool in the development of therapeutic and food applications.

Topical fixed-dose combinations: Current in vitro methodologies for pre-clinical development.

The combination of two or more active pharmaceutical ingredients in the same dosage form - fixed-dose combination products - for topical administration represents a promising therapeutic approach for treating several pathologies, including pain. The pre-clinical development of fixed-dose combination products aims to characterize the interactions between the different APIs and ensure that the final medicinal product has the required safety characteristics. To this end, there are several regulatory accepted in vitro tests to assess the safety of medicinal products intended for cutaneous administration. In turn, the evaluation of anti-inflammatory activity should be based on models described in the scientific literature, as there are no models fully validated by competent entities. Therefore, this work presents the information regarding accepted in vitro tests to assess the safety of topical products and the most used methods to assess anti-inflammatory activity. Additionally, a new approach to select a fixed-dose combination product with the potential to enhance the therapeutic effects of the individual active pharmaceutical ingredients is rationalized by integrating the overall effects on several targets relevant for inflammation and pain management in one numeric index.

In vivo methodologies to assist preclinical development of topical fixed-dose combinations for pain management.

The combination in a fixed dose of two or more active pharmaceutical ingredients in the same pharmaceutical dosage form is an approach that has been used successfully in the treatment of several pathologies, including pain. In the preclinical development of a topical fixed-dose combination product with analgesic and anti-inflammatory activities for pain management, the main objective is to establish the nature of the interaction between the different active pharmaceutical ingredients while obtaining data on the medicinal product safety and efficacy. Despite the improvement of in vitro assays, animal models remain a fundamental strategy to characterise the interaction, efficacy and safety of active pharmaceutical ingredients at the physiological level, which cannot be reached by in vitro assays. Thus, the main goal of this review is to systematise the available animal models to evaluate the efficacy and safety of a new fixed-dose combination product for topical administration indicated for pain management. Particular emphasis is given to animal models that are accepted for regulatory purposes.

Inherited Metabolic Memory of High-Fat Diet Impairs Testicular Fatty Acid Content and Sperm Parameters.

SCOPE: Exposure to a high-fat diet (HFD) from early-life is associated with a testicular metabolic signature link to abnormal sperm parameters up to two generations after exposure in mice. Hereby, this study describes a testicular lipid signature associate with "inherited metabolic memory" of exposure to HFD, persisting up to two generations in mice. METHODS AND RESULTS: Diet-challenged mice (n = 36) are randomly fed after weaning with standard chow (CTRL); HFD for 200 days or transient HFD (HFDt ) (60 days of HFD + 140 days of standard chow). Subsequent generations (36 mice per generation) are fed with chow diet. Mice are euthanized 200 days post-weaning. Glucose homeostasis, serum hormones, testicular bioenergetics, and antioxidant enzyme activity are evaluated. Testicular lipid-related metabolites and fatty acids are characterized by 1 H-NMR and GC-MS. Sons of HFD display impaired choline metabolism, mitochondrial activity, and antioxidant defenses, while grandsons show a shift in testicular ω3/ω6 ratio towards a pro-inflammatory environment. Grandsons of HFDt raise 3-hydroxybutyrate levels with possible implications to testicular insulin resistance. Sperm counts decrease in grandsons of HFD-exposed mice, regardless of the duration of exposure. CONCLUSION: HFD-induced "inherited metabolic memory" alters testicular fatty acid metabolism with consequences to sperm parameters up to two generations.

A nanophytosomes formulation based on elderberry anthocyanins and Codium lipids to mitigate mitochondrial dysfunctions.

The development of nanomedicines to modulate the mitochondrial function is a great scientific challenge since mitochondrial dysfunction is a pathological hallmark of many chronic diseases, including degenerative brain pathologies like Parkinson's and Alzheimer's diseases. To address this challenge, the mitochondriotropic features of the elderberry anthocyanin-enriched extract (Sambucus nigra) were combined with the self-assembling properties of the membrane polar lipids from Codium tomentosum in an innovative SC-Nanophytosomes formulation. Membrane polar lipids, obtained by a new procedure as chlorophyll-free extract, are characterized by 26% of non-phosphorus polar lipids and 74% of phospholipids (dominated by anionic lipids) containing a high degree of polyunsaturated fatty acids. The anthocyanin-enriched extract is dominated by a mixture of four cyanidin-glycosides, representing about 86% of their phenolic content. SC-Nanophytosomes engineered with 600 µM algae membrane polar lipids and 0.5 mg/L of the anthocyanin-enriched extract are nanosized vesicles (diameter =108.74 ± 24.74 nm) with a negative surface charge (Zeta potential = -46.93 ± 6.63 mV) that exhibit stability during storage at 4 ºC. In vitro assays with SH-SY5Y cells showed that SC-Nanophytosomes have the competence to target mitochondria, improving the mitochondrial respiratory chain complexes I and II and preserving the mitochondrial membrane potential in the presence of rotenone. Additionally, SC-Nanophytosomes protect SH-SY5Y cells against the toxicity induced by rotenone or glutamate. Green-fluorescent labeled SC-Nanophytosomes were used to reveal that they are mainly internalized by cells via caveola-mediated endocytosis, escape from endosome and reach the cytoplasm organelles, including mitochondria. Overall, data indicate that SC-Nanophytosomes have the potential to support a mitochondria-targeted therapy for neurodegenerative diseases.

Homarine Alkyl Ester Derivatives as Promising Acetylcholinesterase Inhibitors.

Reversible acetylcholinesterase (AChE) inhibitors are key therapeutic tools to modulate the cholinergic connectivity compromised in several degenerative pathologies. In this work, four alkyl esters of homarine were synthesized and screened by using Electrophorus electricus AChE and rat brain AChE-rich fraction. Results showed that all homarine alkyl esters are able to inhibit AChE by a competitive inhibition mode. The effectiveness of AChE inhibition increases with the alkyl side chain length of the homarine esters, being HO-C16 (IC50 =7.57±3.32 µM and Ki =18.96±2.28 µM) the most potent inhibitor. The fluorescence quenching studies confirmed that HO-C16 is the compound with higher selectivity and affinity for the tryptophan residues in the catalytic active site of AChE. Preliminary cell viability studies showed that homarine esters display no toxicity for human neuronal SH-SY5Y cells. Thus, the long-chain homarine esters emerge as new anti-cholinesterase agents, with potential to be considered for therapeutic applications development.

Improving pollutants environmental risk assessment using a multi model toxicity determination with in vitro, bacterial, animal and plant model systems: The case of the herbicide alachlor.

Several environmental pollutants, including pesticides, herbicides and persistent organic pollutants play an important role in the development of chronic diseases. However, most studies have examined environmental pollutants toxicity in target organisms or using a specific toxicological test, losing the real effect throughout the ecosystem. In this sense an integrative environmental risk of pollutants assessment, using different model organisms is necessary to predict the real impact in the ecosystem and implications for target and non-target organisms. The objective of this study was to use alachlor, a chloroacetanilide herbicide responsible for chronic toxicity, to understand its impact in target and non-target organisms and at different levels of biological organization by using several model organisms, including membranes of dipalmitoylphosphatidylcholine (DPPC), rat liver mitochondria, bacterial (Bacillus stearothermophilus), plant (Lemna gibba) and mammalian cell lines (HeLa and neuro2a). Our results demonstrated that alachlor strongly interacted with membranes of DPPC and interfered with mitochondrial bioenergetics by reducing the respiratory control ratio and the transmembrane potential. Moreover, alachlor also decreased the growth of B. stearothermophilus and its respiratory activity, as well as decreased the viability of both mammalian cell lines. The values of TC50 increased in the following order: Lemna gibba < neuro2a < HeLa cells < Bacillus stearothermophilus. Together, the results suggest that biological membranes constitute a putative target for the toxic action of this lipophilic herbicide and point out the risks of its dissemination on environment, compromising ecosystem equilibrium and human health.

Trichilia catigua and Turnera diffusa extracts: In vitro inhibition of tyrosinase, antiglycation activity and effects on enzymes and pathways engaged in the neuroinflammatory process.

ETHNOPHARMACOLOGICAL RELEVANCE: Flavonoids interact with multiple targets in Central Nervous System resulting in a broad neuroprotection mediated by complementary processes and synergic interactions. Therefore, flavonoid-based therapies may input positive outcomes in the prevention and early management of neurodegenerative diseases. In Brazilian folk medicine Trichilia catigua is used for its neuroactive properties, such as neurostimulant, antioxidant and anti-neuroinflammatory, while Turnera diffusa is traditionally used as a tonic in neurasthenia. Both species are known to be rich in flavonoids. AIM OF THE STUDY: To study aqueous extracts of T. catigua and T. diffusa in terms of their antioxidant and antiglycation effects, inhibition of tyrosinase activity, and interaction with enzymes and pathways engaged in neuroinflammation. Moreover, whenever possible, to establish a relationship between the studied activities and the traditional usage of the species. MATERIALS AND METHODS: The phenolic profiles of the aqueous extracts were validated by HPLC-DAD. The effect of the extracts over mushroom tyrosinase and 5-lipoxygenase activities, as well as their capacity to impair bovine serum albumin glycation, were assessed by in vitro assays. The anti-neuroinflammatory potential of the same extracts was evaluated by their capacity to mitigate the pro-inflammatory stimulus induced in BV-2 microglia cells by interferon-gamma. RESULTS: T. catigua extract, a rich mixture of phenolic acids, catechins and flavonolignans, excels by its ability to decrease lipid peroxidation (EC50 = 227.18 ±â€¯9.04 µg/mL), and to work as anti-glycation agent, and inhibitor of both tyrosinase and 5-lipoxigenase (IC50 = 358.84 ±â€¯19.05 and 56.25 ±â€¯14.53 µg/mL, respectively). However, only T. diffusa extract, mainly composed by luteolin derivatives, is able to lower NO production by BV-2 microglia cells stimulated with interferon-gamma, despite its lower activities in the other assays. CONCLUSIONS: Overall, this work highlights the value of medicinal plant extracts as sources of bioactive flavonoid-rich extracts with neuroactive effects. Furthermore, these results support their application as alternative strategies to develop functional foods and therapeutics to fight chronic neurodegenerative disorders.

Valorisation of kitul, an overlooked food plant: Phenolic profiling of fruits and inflorescences and assessment of their effects on diabetes-related targets.

Caryota urens L. has long been valued as a traditional food, the edible fruits being eaten raw and the inflorescences commonly used on sweet sap and flour production. In the current work, the phenolic profile of methanol extracts obtained from the inflorescences and fruits was unveiled for the first time, nine caffeic acid derivatives being identified and quantified. Since kitul products have been reported for their antidiabetic properties, extracts radical scavenging activity and α-amylase, α-glucosidase and aldose reductase inhibitory activity were assessed. The inflorescences' extract was particularly active against yeast α-glucosidase (IC50 = 1.53 µg/mL), acting through a non-competitive inhibitory mechanism. This activity was also observed in enzyme-enriched homogenates obtained from human Caco-2 cells (IC50 = 64.75 µg/mL). Additionally, the extract obtained from the inflorescences showed no cytotoxicity on HepG2, AGS and Caco-2 cell lines. Our data suggest that C. urens inflorescences can support the development of new functional foods with α-glucosidase inhibitory activity.

Parabens enhance the calcium-dependent testicular mitochondrial permeability transition: Their relevance on the reproductive capacity in male animals.

Parabens, alkyl ester derivatives from p-hydroxybenzoic acid, are extensively used as antimicrobial preservatives. Nonetheless, due to its widespread and massive employment, several studies highlighted the association between parabens and alterations in the reproductive system. This study aimed to relate the adverse effect of the most commonly used parabens in testis mitochondria with male fertility. From all the parabens used, propyl and butyl were the ones that most negatively decreased the respiratory control ratio. In the case of butyl, inhibitions of 20% and 60% were observed, respectively, at the lowest and highest concentration, when compared to the control group. The membrane potential was only significantly affected by propyl (14%) and butyl (31%), and at a concentration of 250 µM. Succinate dehydrogenase, cytochrome c oxidase, and ATPase activities showed a nonsignificant decrease. Cytochrome c reductase, on the other hand, showed statistically significant inhibitions for both propyl (56%) and butylparaben (55%). The susceptibility to the mitochondrial permeability transition pore (MPTP) opening was increased by all parabens, although this increase was markedly significant for propyl and butyl. These results show that the susceptibility of mitochondria to parabens is dependent on the alkyl chain length and parabens hydrophobicity, and the main mitochondrial target is Complex II-III and MPTP. Hence, this study demonstrates the contribution of parabens exposition to the inhibition of testis mitochondrial function and their putative noxious effect on the male reproductive system.

Use of Parabens (Methyl and Butyl) during the Gestation Period: Mitochondrial Bioenergetics of the Testes and Antioxidant Capacity Alterations in Testes and Other Vital Organs of the F1 Generation.

Since the mid-1920s, parabens have been widely used as antimicrobial preservatives in processed foods and beverages, pharmaceuticals, and cosmetic products. Paraben use continues to generate considerable controversy, both in the general population and in the scientific community itself. The primary purpose of our study was to determine whether parabens (methyl and butyl at concentrations of 100 and 200 mg/kg body weight by subcutaneous injection) during pregnancy of adult female Wistar rats can have an impact on the F1 generation. As far as we know, we are the first to demonstrate that using parabens during pregnancy has negative repercussions on the mitochondrial bioenergetics and antioxidant activity of testicular germ cells in the F1 generation. Our study showed that there was a 48.7 and 59.8% decrease in the respiratory control index with 100 and 200 mg/kg of butylparaben, respectively. Cytochrome c oxidase activity was significantly inhibited (45 and 51%) in both groups. In addition, 200 mg/kg butylparaben promoted a marked decrease in citrate synthase activity, indicating that mitochondrial content decreased in the germ cells, especially spermatocytes and spermatids. Mitochondrial ROS production increased in groups exposed to parabens in a concentration-dependent manner, especially the butyl one (102 and 130%). The groups exposed to butylparaben showed an increase in superoxide dismutase (SOD) and catalase (CAT) activities, while glutathione reductase (GR) and glutathione S-transferase (GST) decreased. With methylparaben, only differences in SOD and GR were observed; for the latter, this only occurred with the highest concentration. The glutathione (GSH)/glutathione disulfide (GSSG) ratio did not undergo any significant change. However, there was a considerable increase in hydroperoxide content in animals exposed to butylparaben, with 100 and 200 mg/kg resulting in 98.6 and 188% increase, respectively. Furthermore, several other organs also showed alterations in antioxidant capacity due to paraben use. In summary, our study demonstrates that paraben use during pregnancy will cause severe changes in the mitochondrial bioenergetics and antioxidant capacity of testicular germ cells and the antioxidant capacity of several other F1 generation organs.