Bioactive properties of spearmint, orange peel, and baby sage oleoresins obtained by supercritical CO2 extraction and their integration into dark chocolate.

This study investigated the potential health benefits of spearmint, orange peel, and baby sage oleoresins extracted using supercritical CO2 and subsequently emulsified. The oleoresins were incorporated into dark chocolate, and their impact on physicochemical properties was evaluated. Characterization revealed rich sources of phenolic compounds, carotenoids, and volatile compounds in these oleoresins. In vitro studies demonstrated anti-obesogenic, antioxidant, anti-inflammatory, and neuroprotective properties of the emulsified oleoresins. However, only physicochemical properties were determined for the formulations of dark chocolate with these emulsified oleoresins. Chocolate formulations fortified with these emulsions displayed a softer texture, lower water activity, and solid-like behavior. The findings suggest that these oleoresins could serve as nutraceutical agents for mitigating metabolic syndrome and associated pathologies. Incorporating them into chocolate matrices offers a practical approach to formulating functional foods. Further research is warranted to explore the preventive and therapeutic efficacy in an in vivo model.

Development and Evaluation of Zinc and Iron Nanoparticles Functionalized with Plant Growth-Promoting Rhizobacteria (PGPR) and Microalgae for Their Application as Bio-Nanofertilizers.

Micronutrient deficiencies are widespread and growing global concerns. Nanoscale nutrients present higher absorption rates and improved nutrient availability and nutrient use efficiency. Co-application of nanofertilizers (NFs) with biological agents or organic compounds increases NF biocompatibility, stability, and efficacy. This study aimed to develop and evaluate zinc and iron bio-nanofertilizers formulated with plant growth-promoting rhizobacteria (PGPR) and microalgae. Nanoparticles (NPs) were synthesized with the co-precipitation method and functionalized with Pseudomonas species and Spirulina platensis preparation. NPs were characterized and evaluated on seed germination, soil microbial growth, and early plant response under seedbed conditions. NPs corresponded to zinc oxide (ZnO; 77 nm) and maghemite (γ-Fe2O3; 68 nm). Functionalized nanoparticles showed larger sizes, around 145-233 nm. The seedling vigor index of tomato and maize was significantly increased (32.9-46.1%) by bacteria-functionalized ZnO- and γ-Fe2O3-NPs at 75 ppm. NFs at 250 and 75 ppm significantly increased bacterial growth. NFs also improved early plant growth by increasing plant height (14-44%), leaf diameter (22-47%), and fresh weight (46-119%) in broccoli and radish, which were mainly influenced by bacteria capped ZnO- and γ-Fe2O3-NPs at 250 ppm. Beneficial effects on plant growth can be attributed to the synergistic interaction of the biological components and the zinc and iron NPs in the bio-nanofertilizers.

Fish oil and probiotics supplementation through milk chocolate improves spatial learning and memory in male Wistar rats.

Background: Cognition and brain function is critical through childhood and should be improved with balanced diets. Incorporating bioactive ingredients such as omega-3 polyunsaturated fatty acids (ω3 PUFAs) and probiotics into food formulations could be used as an approach to improve cognitive function. This study evaluated the effects on cognitive capacity of complementing rodent diets with chocolate, by itself and in combination with ω3 PUFAs from fish oil and probiotics. Methods: Spatial learning and memory in the rats were determined by the Barnes maze test in short- and long-term memory. Samples from the cecum were obtained to assess microbial counts (Lactobacillus, Bifidobacterium, Enterobacteriaceae, and total bacteria), and brains were recovered to analyze the neural morphology of the tissues. Also, glucose, brain weights, and epididymal tissue were analyzed. Results: The combination of chocolate with fish oil and probiotics improved the memory of rats compared to the result of each bioactive compound when evaluated separately. Treatments did not affect sugar level, epididymal adipose tissue, or brain weight. On the other hand, consuming probiotics alone or in combination with chocolate decreased Enterobacteria counts, while Lactobacillus and Bifidobacteria counts were not affected. Neural morphological analysis showed that combining chocolate with probiotics and ω3 PUFAs increased the number of neurons in the hippocampal CA1 and CA3 regions. Conclusion: Chocolate added with probiotics and ω3 PUFAs improved spatial memory and learning in the studied model.

Hydroxytyrosol inhibits cancer stem cells and the metastatic capacity of triple-negative breast cancer cell lines by the simultaneous targeting of epithelial-to-mesenchymal transition, Wnt/ß-catenin and TGFß signaling pathways.

PURPOSE: This study was aimed to determine the impact of hydroxytyrosol (HT), a minor compound found in olive oil, on breast cancer stem cells (BCSCs) and the migration capacity of triple-negative breast cancer (TNBC) cell lines through the alteration of epithelial-to-mesenchymal transition (EMT) and embryonic signaling pathways. METHODS: BCSCs self-renewal was determined by the mammosphere-forming efficiency in SUM159PT, BT549, MDA-MB-231 and Hs578T TNBC cell lines. Flow cytometric analysis of CD44+/CD24-/low and aldehyde dehydrogenase positive (ALDH+) subpopulations, migration by the "wound healing assay", invasion and Western blot of EMT markers and TGFß signaling were investigated in SUM159PT, BT549 and MDA-MB-231 cell lines. Wnt/ß-catenin signaling was assessed by Western blot in BT549 cells expressing WNT1 and MDA-MB-231 cells. Changes in TGFß activity was determined by SMAD Binding Element (SBE) reporter assay. RESULTS: HT reduced BCSCs self-renewal, ALDH+ (aldehyde dehydrogenase) and CD44+/CD24-/low subpopulations, tumor cell migration and invasion. Consistently, HT suppressed Wnt/ß-catenin signaling by decreasing p-LRP6, LRP6, ß-catenin and cyclin D1 protein expression and the EMT markers SLUG, ZEB1, SNAIL and VIMENTIN. Finally, HT inhibited p-SMAD2/3 and SMAD2/3 in SUM159PT, BT549 and MDA-MB-231 cells, what was correlated with a less TGFß activity. CONCLUSION: In conclusion, we report for the first time the inhibitory role of HT on BCSCs and tumor cell migration by targeting EMT, Wnt/ß-catenin and TGFß signaling pathways. Our findings highlight the importance of the chemopreventive compound HT as a novel candidate to be investigated as an alternative targeted therapy for TNBC.

Ex Vivo Cardiotoxicity of Antineoplastic Casiopeinas Is Mediated through Energetic Dysfunction and Triggered Mitochondrial-Dependent Apoptosis.

Casiopeinas are a group of copper-based antineoplastic molecules designed as a less toxic and more therapeutic alternative to cisplatin or Doxorubicin; however, there is scarce evidence about their toxic effects on the whole heart and cardiomyocytes. Given this, rat hearts were perfused with Casiopeinas or Doxorubicin and the effects on mechanical performance, energetics, and mitochondrial function were measured. As well, the effects of Casiopeinas-triggered cell death were explored in isolated cardiomyocytes. Casiopeinas III-Ea, II-gly, and III-ia induced a progressive and sustained inhibition of heart contractile function that was dose- and time-dependent with an IC50 of 1.3 ± 0.2, 5.5 ± 0.5, and 10 ± 0.7 µM, correspondingly. Myocardial oxygen consumption was not modified at their respective IC50, although ATP levels were significantly reduced, indicating energy impairment. Isolated mitochondria from Casiopeinas-treated hearts showed a significant loss of membrane potential and reduction of mitochondrial Ca2+ retention capacity. Interestingly, Cyclosporine A inhibited Casiopeinas-induced mitochondrial Ca2+ release, which suggests the involvement of the mitochondrial permeability transition pore opening. In addition, Casiopeinas reduced the viability of cardiomyocytes and stimulated the activation of caspases 3, 7, and 9, demonstrating a cell death mitochondrial-dependent mechanism. Finally, the early perfusion of Cyclosporine A in isolated hearts decreased Casiopeinas-induced dysfunction with reduction of their toxic effect. Our results suggest that heart cardiotoxicity of Casiopeinas is similar to that of Doxorubicin, involving heart mitochondrial dysfunction, loss of membrane potential, changes in energetic metabolites, and apoptosis triggered by mitochondrial permeability.

Use of Modified Phenolic Thyme Extracts (Thymus vulgaris) with Reduced Polyphenol Oxidase Substrates as Anthocyanin Color and Stability Enhancing Agents.

Residual enzymatic activity in certain foods, particularly of polyphenoloxidase (PPO), is responsible for the majority of anthocyanin degradation in food systems, causing also parallel losses of other relevant nutrients. The present work explored the feasibility of modifying phenolic profiles of thyme extracts, by use of chromatographic resins, to obtain phenolic extracts capable of enhancing anthocyanin colour and stability in the presence of PPO activity. Results indicated that pretreatment of thyme extracts with strong-anion exchange resins (SAE) enhanced their copigmentation abilities with strawberry juice anthocyanins. Phenolic chromatographic profiles, by HPLC-PDA, also demonstrated that thyme extracts subjected to SAE treatments had significantly lower concentrations of certain phenolic compounds, but extracts retained their colour enhancing and anthocyanin stabilization capacities though copigmentation. Additional testing also indicated that SAE modified extract had a lower ability (73% decrease) to serve as PPO substrate, when compared to the unmodified extract. Phenolic profile modification process, reported herein, could be potentially used to manufacture modified anthocyanin-copigmentation food and cosmetic additives for colour-stabilizing applications with lower secondary degradation reactions in matrixes that contain PPO activity.

Isolation and chemical identification of lipid derivatives from avocado (Persea americana) pulp with antiplatelet and antithrombotic activities.

Platelets play a pivotal role in physiological hemostasis. However, in coronary arteries damaged by atherosclerosis, enhanced platelet aggregation, with subsequent thrombus formation, is a precipitating factor in acute ischemic events. Avocado pulp (Persea americana) is a good source of bioactive compounds, and its inclusion in the diet as a source of fatty acid has been related to reduced platelet aggregability. Nevertheless, constituents of avocado pulp with antiplatelet activity remain unknown. The present study aims to characterize the chemical nature of avocado constituents with inhibitory effects on platelet aggregation. Centrifugal partition chromatography (CPC) was used as a fractionation and purification tool, guided by an in vitro adenosine diphosphate (ADP), arachidonic acid or collagen-platelet aggregation assay. Antiplatelet activity was initially linked to seven acetogenins that were further purified, and their dose-dependent effects in the presence of various agonists were contrasted. This process led to the identification of Persenone-C (3) as the most potent antiplatelet acetogenin (IC50=3.4 mM) among the evaluated compounds. In vivo evaluations with Persenone A (4) demonstrated potential protective effects against arterial thrombosis (25 mg kg⁻¹ of body weight), as coagulation times increased (2-fold with respect to the vehicle) and thrombus formation was attenuated (71% versus vehicle). From these results, avocado may be referred to as a functional food containing acetogenin compounds that inhibit platelet aggregation with a potential preventive effect on thrombus formation, such as those that occur in ischaemic diseases.

Activity-guided identification of acetogenins as novel lipophilic antioxidants present in avocado pulp (Persea americana).

Avocado fruit is a rich source of health-related lipophilic phytochemicals such as monounsaturated fatty acids, tocopherols, carotenes, acetogenins and sterols. However, limited information is available on the contribution of specific phytochemicals to the overall antioxidant capacity (AOC) of the fruit. Centrifugal partition chromatography was used as fractionation tool, guided by an in vitro chemical assay of oxygen radical absorbance capacity (ORAC). Subsequent experiments focused on isolation and characterization of the chemical nature of the main contributors to lipophilic AOC of avocado pulp. ORAC values obtained for acetogenins were contrasted with results from an isolated kidney mitochondria membrane lipid peroxidation bioassay. The present study established that lipophilic AOC of the pulp was significantly higher than its hydrophilic AOC. Our results confirmed the presence of acetogenins in the fractions with highest lipophilic AOC, and for the first time linked them as contributors to lipophilic-ORAC values. Further HPLC-PDA/MS-TOF analysis led to structural elucidation of two novel acetogenins, not previously reported as present in avocado pulp, along with five already known related-compounds. Antioxidant properties observed for avocado pulp acetogenins by the ORAC assay suggested that, in the presence of an emulsifying agent, acetogenins could serve as novel lipophilic antioxidants in a food matrix. Results from isolated mitochondria lipid peroxidation bioassay, indicated that L-ORAC values which may have relevance for food matrix applications, should not be interpreted to have a direct relevance in health-related claims, compounds need to be evaluated considering the complexity of biological systems.

Cardiotoxicity of acetogenins from Persea americana occurs through the mitochondrial permeability transition pore and caspase-dependent apoptosis pathways.

Acetogenins are cell-membrane permeable, naturally occurring secondary metabolites of plants such as Annonaceae, Lauraceae and other related phylogenic families. They belong to the chemical derivatives of polyketides, which are synthesized from fatty acid precursors. Although acetogenins have displayed diverse biological activities, the anti-proliferative effect on human cancer cells has been widely reported. Acetogenins are inhibitors of complex I in the electron transport chain therefore they interrupt ATP synthesis in mitochondria. We tested a new acetogenins-enriched extract from the seed of Persea americana in order to investigate if any toxicity was induced on cardiac tissue and determine the involved mechanism. In isolated perfused heart we found that contractility was completely inhibited at an accumulative dose of 77 µg/ml. In isolated cardiomyocytes, the acetogenins-enriched extract induced apoptosis through the activation of the intrinsic pathway at 43 µg/ml. In isolated mitochondria, it inhibited complex I activity on NADH-linked respiration, as would be expected, but also induced permeability transition on succinate-linked respiration. Cyclosporine A, a known blocker of permeability transition, significantly prevented the permeability transition triggered by the acetogenins-enriched extract. In addition, our acetogenins-enriched extract inhibited ADP/ATP exchange, suggesting that an important element in phosphate or adenylate transport was affected. In this manner we suggest that acetogenins-enriched extract from Persea americana could directly modulate permeability transition, an entity not yet associated with the acetogenins' direct effects, resulting in cardiotoxicity.