Vasorelaxant Activity of (2S)-Sakuranetin and Other Flavonoids Isolated from the Green Propolis of the Caatinga Mimosa tenuiflora.

Some in vitro and in vivo evidence is consistent with the cardiovascular beneficial activity of propolis. As the single actors responsible for this effect have never been identified, an in-depth investigation of flavonoids isolated from the green propolis of the Caatinga Mimosa tenuiflora was performed and their mechanism of action was described. A comprehensive electrophysiology, functional, and molecular docking approach was applied. Most flavanones and flavones were effective CaV1.2 channel blockers with a potency order of (2S)-sakuranetin > eriodictyol-7,3'-methyl ether > quercetin 3-methyl ether > 5,4'-dihydroxy-6,7-dimethoxyflavanone > santin > axillarin > penduletin > kumatakenin, ermanin and viscosine being weak or modest stimulators. Except for eriodictyol 5-O-methyl ether, all the flavonoids were also effective spasmolytic agents of vascular rings, kumatakenin and viscosine also showing an endothelium-dependent activity. (2S)-Sakuranetin also stimulated KCa1.1 channels both in single myocytes and vascular rings. In silico analysis provided interesting insights into the mode of action of (2S)-sakuranetin within both CaV1.2 and KCa1.1 channels. The green propolis of the Caatinga Mimosa tenuiflora is a valuable source of multi-target vasoactive flavonoids: this evidence reinforces its nutraceutical value in the cardiovascular disease prevention arena.

In vitro Evaluation of the Antileishmanial and Antischistosomal Activities of p-Coumaric Acid Prenylated Derivatives.

We have evaluated eight p-coumaric acid prenylated derivatives in vitro for their antileishmanial activity against Leishmania amazonensis promastigotes and their antischistosomal activity against Schistosoma mansoni adult worms. Compound 7 ((E)-3,4-diprenyl-4-isoprenyloxycinnamic alcohol) was the most active against L. amazonensis (IC50=45.92 µM) and S. mansoni (IC50=64.25 µM). Data indicated that the number of prenyl groups, the presence of hydroxyl at C9, and a single bond between C7 and C8 are important structural features for the antileishmanial activity of p-coumaric acid prenylated derivatives.

3,3'-O-dimethylquercetin: A bi-functional vasodilator isolated from green propolis of the Caatinga Mimosa tenuiflora.

In the search for novel, bi-functional compounds acting as CaV1.2 channel blockers and K+ channel stimulators, which represent an effective therapy for hypertension, 3,3'-O-dimethylquercetin was isolated for the first time from Brazilian Caatinga green propolis. Its effects were investigated through electrophysiological, functional, and computational approaches. In rat tail artery myocytes, 3,3'-O-dimethylquercetin blocked Ba2+ currents through CaV1.2 channels (IBa1.2) in a concentration-dependent manner, with the inhibition being reversed upon washout. The compound also shifted the voltage dependence of the steady-state inactivation curve to more negative potentials without affecting the slope of the inactivation and activation curves. Furthermore, the flavonoid stimulated KCa1.1 channel currents (IKCa1.1). In silico simulations provided additional evidence for the binding of 3,3'-O-dimethylquercetin to KCa1.1 and CaV1.2 channels and elucidated its mechanism of action. In depolarized rat tail artery rings, the flavonoid induced a concentration-dependent relaxation. Moreover, in rat aorta rings its antispasmodic effect was inversely related to the transmembrane K+ gradient. In conclusion, 3,3'-O-dimethylquercetin demonstrates effective in vitro vasodilatory properties, encouraging the exploration of its scaffold to develop novel derivatives for potential use in the treatment of hypertension.

Efficacy and safety of guttiferone E in melanoma-bearing mice.

Melanoma, an aggressive and potentially fatal skin cancer, is constrained by immunosuppression, resistance, and high toxicity in its treatment. Consequently, there is an urgent need for innovative antineoplastic agents. Therefore, this study investigated the antimelanoma potential of guttiferone E (GE). In an allogeneic murine B16 melanoma model, GE was administered subcutaneously and intraperitoneally. Antitumor evaluation included tumor volume/weight measurements and histopathological and immunohistochemical analysis. Furthermore, the toxicity of the treatments was evaluated through body/organ weights, biochemical parameters, and genotoxicity. Subcutaneous administration of 20 mg/kg of GE resulted in a significant reduction in both tumor volume and weight, effectively suppressing melanoma cell proliferation as evidenced by a decrease in mitotic figures. The tumor growth inhibition rate was equivalent to 54%. This treatment upregulated cleaved caspase-3, indicating apoptosis induction. On the other hand, intraperitoneal administration of GE showed no antimelanoma effect. Remarkably, GE treatments exhibited no toxicity, evidenced by non-significant differences in body weight gain, as well as organ weight, biochemical parameters of nephrotoxicity and hepatotoxicity, and genotoxic damage. This study revealed, for the first time, the efficacy of subcutaneous administration of GE in reducing melanoma, in the absence of toxicity. Furthermore, it was observed that the apoptotic signaling pathway is involved in the antimelanoma property of GE. These findings offer valuable insights for further exploring GE's therapeutic applications in melanoma treatment.

Leishmanial activity of Brazilian brown propolis and its diterpenes.

Propolis is a natural product widely used in folk medicine. Among its various applications, its antiparasitic properties stand out. Due to its great biodiversity, Brazil is a major producer of several types of propolis. This study proposes to evaluate the leishmanicidal properties of the hydroalcoholic extract of propolis collected in the southern region of Brazil (Brown propolis - HEBP) and its main isolated compounds: abietic acid (1), 13-epi-cupressic acid (2), 13-epi-torulosol (3), dehydroabietic acid (4), cis-communic acid (5) and ent-agatic acid (6). In general, the diterpenes did not show activity against the promastigotes of Leishmania (Leishmania) amazonensis at the evaluated concentrations. However, the HEBP was very active with an inhibition concentration of 50% at 8.32 µg/mL. Moreover, transmission electron microscopy (TEM) and scanning electron microscopy (SEM) assays showed morphological and structural alterations in promastigote forms of L. (L.) amazonensis when incubated with HEBP.

Copaifera langsdorffii Oleoresin-Loaded Nanostructured Lipid Carrier Emulgel Improves Cutaneous Healing by Anti-Inflammatory and Re-Epithelialization Mechanisms.

The skin is essential to the integrity of the organism. The disruption of this organ promotes a wound, and the organism starts the healing to reconstruct the skin. Copaifera langsdorffii is a tree used in folk medicine to treat skin affections, with antioxidant and anti-inflammatory properties. In our study, the oleoresin of the plant was associated with nanostructured lipid carriers, aiming to evaluate the healing potential of this formulation and compare the treatment with reference drugs used in wound healing. Male Wistar rats were used to perform the excision wound model, with the macroscopic analysis of wound retraction. Skin samples were used in histological, immunohistochemical, and biochemical analyses. The results showed the wound retraction in the oleoresin-treated group, mediated by α-smooth muscle actin (α-SMA). Biochemical assays revealed the anti-inflammatory mechanism of the oleoresin-treated group, increasing interleukin-10 (IL-10) concentration and decreasing pro-inflammatory cytokines. Histopathological and immunohistochemical results showed the improvement of re-epithelialization and tissue remodeling in the Copaifera langsdorffii group, with an increase in laminin-γ2, a decrease in desmoglein-3 and an increase in collagen remodeling. These findings indicate the wound healing potential of nanostructured lipid carriers associated with Copaifera langsdorffii oleoresin in skin wounds, which can be helpful as a future alternative treatment for skin wounds.

Brazilian Green Propolis' Artepillin C and Its Acetylated Derivative Activate the NGF-Signaling Pathways and Induce Neurite Outgrowth in NGF-Deprived PC12 Cells.

Artepillin C is the most studied compound in Brazilian Green Propolis and, along with its acetylated derivative, displays neurotrophic activity on PC12 cells. Specific inhibitors of the trkA receptor (K252a), PI3K/Akt (LY294002), and MAPK/ERK (U0126) signaling pathways were used to investigate the neurotrophic mechanism. The expression of proteins involved in axonal and synaptic plasticity (GAP-43 and Synapsin I) was assessed by western blotting. Additionally, physicochemical properties, pharmacokinetics, and drug-likeness were evaluated by the SwissADME web tool. Both compounds induced neurite outgrowth by activating the NGF-signaling pathways but through different neuronal proteins. Furthermore, in silico analyses showed interesting physicochemical and pharmacokinetic properties of these compounds. Therefore, these compounds could play an important role in axonal and synaptic plasticity and should be further investigated.

Development and Characterization of High-Absorption Microencapsulated Organic Propolis EPP-AF® Extract (i-CAPs).

The demand for organic and functional food continues to increase yearly. Among the available functional foods, propolis is a bee product that has various beneficial properties, including antimicrobial, antioxidant, and anti-inflammatory activities. However, it generally is only available in ethanol solution, which has poor bioavailability, as it is relatively insoluble in water. The use of such ethanol extracts is often objectionable because of the alcohol content and because they have a strong and striking taste. Development of alternatives that can efficiently and safely increase solubility in water, and that meet organic production specifications, has been a challenge. To address these concerns, microcapsules were developed using spray-dryer technology from an emulsion based on EPP-AF® propolis and gum arabic (i-CAPS). These propolis-loaded microcapsules were characterized using FT-IR, SEM, TGA, HPLC, and spectrophotometric techniques, along with determination of antimicrobial, antioxidant, antitumor, anti-inflammatory, and antihypercholesterolemic activities, as well as permeability in in vitro models. The production system resulted in microcapsules with a spherical shape and an encapsulation efficiency of 93.7 ± 0.7%. They had IC50s of 2.654 ± 0.062 and 7.342 ± 0.058 µg/mL by FRAP and DPPH antioxidant methods, respectively. The EPP-AF® i-CAPS also had superior antimicrobial activity against Gram-positive bacteria. Antitumor activity was calculated based on the concentration that inhibited 50% of growth of AGS, Caco-2, and MCF-7 cell strains, giving results of 154.0 ± 1.0, 117 ± 1.0, and 271.0 ± 25 µg/mL, respectively. The microcapsule presentation reduced the permeation of cholesterol by 53.7%, demonstrating antihypercholesterolemic activity, and it improved the permeability of p-coumaric acid and artepillin C. The IC50 for NO production in RAW 264.7 cells was 59.0 ± 0.1 µg/mL. These findings demonstrate the potential of this new propolis product as a food and pharmaceutical ingredient, though additional studies are recommended to validate the safety of proposed dosages.

Polyalthic Acid from Copaifera lucens Demonstrates Anticariogenic and Antiparasitic Properties for Safe Use.

This study aimed at evaluating the potential of Copaifera lucens, specifically its oleoresin (CLO), extract (CECL), and the compound ent-polyalthic acid (PA), in combating caries and toxoplasmosis, while also assessing its toxicity. The study involved multiple assessments, including determining the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) against cariogenic bacteria. CLO and PA exhibited MIC and MBC values ranging from 25 to 50 µg/mL, whereas CECL showed values equal to or exceeding 400 µg/mL. PA also displayed antibiofilm activity with minimum inhibitory concentration of biofilm (MICB50) values spanning from 62.5 to 1000 µg/mL. Moreover, PA effectively hindered the intracellular proliferation of Toxoplasma gondii at 64 µg/mL, even after 24 h without treatment. Toxicological evaluations included in vitro tests on V79 cells, where concentrations ranged from 78.1 to 1250 µg/mL of PA reduced colony formation. Additionally, using the Caenorhabditis elegans model, the lethal concentration (LC50) of PA was determined as 1000 µg/mL after 48 h of incubation. Notably, no significant differences in micronucleus induction and the NDI were observed in cultures treated with 10, 20, or 40 µg/mL of CLO. These findings underscore the safety profile of CLO and PA, highlighting their potential as alternative treatments for caries and toxoplasmosis.

Evaluation of anti-inflammatory activity of kaurenol: Experimental evaluation and mechanistic insights.

BACKGROUND: Kaurenol, a diterpene alcohol found in Copaifera langsdorffii Desf. (known as "copaiba"), is historically used in traditional medicine for inflammatory conditions. OBJECTIVES: This study aims to comprehensively assess the potential anti-inflammatory and antinociceptive properties of kaurenol. METHODS: To this end, the following experiments were conducted to evaluated toxicity: locomotor performance and acute toxicity; nociception: acetic acid-induced writhing and formalin-induced antinociception; and anti-inflammatory activity: carrageenan and dextran-induced paw edema at 10, 20, and 40 mg/kg, and measurement of nitric oxide (NO), tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), and interleukin-10 (IL-10) in macrophages at 1, 3, and 9 µg/ml. RESULTS: Kaurenol did not show significant locomotor changes, acute toxicity, and central analgesic activity in the first phase of formalin test at dosages tested. Kaurenol showed 53%, 64%, 64%, and 58% of inhibition in the acetic acid-induced writhing, second phase of formalin test, carrageenan and dextran-induced paw edema, respectively. CONCLUSION: The anti-inflammatory activity was associated with the regulation of NO release and probably with the regulation of mediators, such as serotonin and prostaglandin in vascular permeability, as well as by being associated with the regulation of IL-6 and IL-10. Kaurenol display anti-inflammatory activity but has no analgesic activity.

Toxicological and chemoprevention studies of Brazilian brown propolis from Araucaria sp.

Brazilian brown propolis (BBP) is a natural product derived predominantly from the south region of Brazil, where Araucaria forests are dominant. Despite its potential as a source of bioactive compounds with leishmanicidal, anti-inflammatory, nociceptive, and antimicrobial properties, BBP has not been comprehensively studied compared to green propolis. Therefore, this study aimed to determine the safety and chemopreventive potential of BBP. The cytotoxicity attributed to BBP was assessed using two different assays, while the Salmonella/microsome assay was employed to evaluate mutagenicity. The acute toxicity attributed to BBP was determined using a zebrafish model, while the chemopreventive potential was investigated utilizing Chinese hamster lung (V79) cell lines. Data demonstrated that BBP exerted cytotoxic effects at concentrations greater than or equal to 10 µg/ml and did not exhibit mutagenicity in Salmonella typhimurium strains TA98 and TA100. However, at the highest concentration tested (4000 µg/plate), BBP induced a significant increase in revertant colonies in S. typhimurium TA102 strain. The LC50 equivalent to 8.83 mg/L was obtained in the acute toxicity evaluation in zebrafish. BBP also showed antigenotoxic effect by significantly reducing chromosomal damage induced by the mutagen doxorubicin in V79 cell cultures at a concentration of 2.5 µg/ml. Compared to Brazilian green and red propolis, BBP exhibited greater toxicity. On the other hand, at lower concentrations, BBP displayed chemopreventive potential, which may be associated with the antioxidant capacity of the extract. These findings contribute to a better understanding of the biological properties and potential applications of BBP in treating various diseases.

Suppression of VEGF and inflammatory cytokines, modulation of Annexin A1 and organ functions by galloylquinic acids in breast cancer model.

The ongoing development of novel drugs for breast cancer aims to improve therapeutic outcomes, reduce toxicities, and mitigate resistance to chemotherapeutic agents. Doxorubicin (Dox) is known for its significant side effects caused by non-specific cytotoxicity. In this study, we investigated the antitumor activity of galloylquinic acids (BF) and the beneficial role of their combination with Dox in an Ehrlich ascites carcinoma (EAC)-bearing mouse model, as well as their cytotoxic effect on MCF-7 cells. The EAC-mice were randomized into five experimental groups: normal saline, Dox (2 mg/kg, i.p), BF (150 mg/kg, orally), Dox and BF combined mixture, and a control group. Mice were subjected to a 14-day treatment regimen. Results showed that BF compounds exerted chemopreventive effects in EAC mice group by increasing mean survival time, decreasing tumor volume, inhibiting ascites tumor cell count, modulating body weight changes, and preventing multi-organ histopathological alterations. BF suppressed the increased levels of inflammatory mediators (IL-6 and TNF-α) and the angiogenic marker VEGF in the ascitic fluid. In addition, BF and their combination with Dox exhibited significant cytotoxic activity on MCF-7 cells by inhibiting cell viability and modulating Annexin A1 level. Moreover, BF treatments could revert oxidative stress, restore liver and kidney functions, and normalize blood cell counts.

Synthesis and Antibacterial Activity of Polyalthic Acid Analogs.

Polyalthic acid (PA) is a diterpene found in copaiba oil. As a continuation of our work with PA, we synthesized PA analogs and investigated their antibacterial effects on preformed biofilms of Staphylococcus epidermidis and determined the minimal inhibitory concentration (MIC) of the best analogs against planktonic bacterial cells. There was no difference in activity between the amides 2a and 2b and their corresponding amines 3a and 3b regarding their ability to eradicate biofilm. PA analogs 2a and 3a were able to significantly eradicate the preformed biofilm of S. epidermidis and were active against all the Gram-positive bacteria tested (Enterococcus faecalis, Enterococcus faecium, S. epidermidis, Staphylococcus aureus), with different MIC depending on the microorganism. Therefore, PA analogs 2a and 3a are of interest for further in vitro and in vivo testing to develop formulations for antibiotic drugs against Gram-positive bacteria.

Advances in the phytochemical screening and biological potential of propolis.

Propolis is a natural resinous product collected from different parts of plants by bees and mixed with their salivary secretions. The occurrence of more than 180 different chemotypes has flavonoids, phenolic acids, esters, and phenolic aldehydes, as well as balsamic resins, beeswax, pollen, and essential and aromatic oils, among others. Its biological potential documented throughout the world justifies the need, from time to time, to organize reviews on the subject, with the intention of gathering and informing about the update on propolis. In this review (CRD42020212971), phytochemical advances, in vitro, in vivo, and clinical biological assays of pharmacological interest are showcased. The focus of this work is to present propolis clinical safety assays, antitumor, analgesic, antioxidant, anti-inflammatory, and antimicrobial activities. This literature review highlights propolis' promising biological activity, as it also suggests that studies associating propolis with nanotechnology should be further explored for enhanced bioprocessing applications.

Polyalthic acid and oleoresin from Copaifera trapezifolia Hayne reduce Toxoplasma gondii growth in human villous explants, even triggering an anti-inflammatory profile.

Due to the lack of efficient antiparasitic therapy and vaccines, as well as emerging resistance strains, congenital toxoplasmosis is still a public health issue worldwide. The present study aimed to assess the effects of an oleoresin obtained from the species Copaifera trapezifolia Hayne (CTO), and an isolated molecule found in the CTO, ent-polyalthic acid (ent-15,16-epoxy-8(17),13(16),14-labdatrien-19-oic acid) (named as PA), against T. gondii infection. We used human villous explants as an experimental model of human maternal-fetal interface. Uninfected and infected villous explants were exposed to the treatments; the parasite intracellular proliferation and the cytokine levels were measured. Also, T. gondii tachyzoites were pre-treated and the parasite proliferation was determined. Our findings showed that CTO and PA reduced efficiently the parasite growth with an irreversible action, but without causing toxicity to the villi. Also, treatments reduced the levels of IL-6, IL-8, MIF and TNF by villi, what configures a valuable treatment option for the maintenance of a pregnancy in an infectious context. In addition to a possible direct effect on parasites, our data suggest an alternative mechanism by which CTO and PA alter the villous explants environment and then impair parasite growth, since the pre-treatment of villi resulted in lower parasitic infection. Here, we highlighted PA as an interesting tool for the design of new anti-T. gondii compounds.

Brazilian red propolis extract free and encapsulated into polymeric nanoparticles against ovarian cancer: formulation, characterisation and biological assays in 2D and 3D models.

Cancer incidence worldwide is alarming and among the cancers that affect women ovarian cancer is the most fatal. Many side effects are associated with conventional therapies and none of them are completely effective, so the development of new treatments is necessary. Brazilian red propolis extract is a natural product with complex composition and great potential for cancer treatment. However, its clinical application is harmed due to unfavourable physicochemical characteristics. To enable its application encapsulation in nanoparticles can be used. OBJECTIVES: The aims of this work were to develop polymeric nanoparticles with Brazilian red propolis extract and compare their action with the free extract against ovarian cancer cells. METHODS: Box Behnken design was used and nanoparticles were characterised using the techniques dynamic light scattering, nanoparticle tracking analysis, transmission electron microscopy, differential scanning calorimetry and encapsulation efficiency. Activity against OVCAR-3 was also tested on 2D and 3D models. KEY FINDINGS: Nanoparticles' sizes were ~200 nm with monomodal size distribution, negative zeta potential, spherical shape and with extract molecularly dispersed. Encapsulation efficiency was above 97% for the biomarkers chosen. Nanoparticles had greater efficacy in comparison with free propolis in OVCAR-3. CONCLUSIONS: So far, the nanoparticles here described have the potential to be a chemotherapy treatment in the future.

Development and Characterization of New Green Propolis Extract Formulations as Promising Candidates to Substitute for Green Propolis Hydroalcoholic Extract.

The technologies used to produce the different dosage forms of propolis can selectively affect the original propolis compounds and their biological activities. The most common type of propolis extract is hydroethanolic. However, there is considerable demand for ethanol-free propolis presentations, including stable powder forms. Three propolis extract formulations were developed and investigated for chemical composition and antioxidant and antimicrobial activity: polar propolis fraction (PPF), soluble propolis dry extract (PSDE), and microencapsulated propolis extract (MPE). The different technologies used to produce the extracts affected their physical appearance, chemical profile, and biological activity. PPF was found to contain mainly caffeic and p-Coumaric acid, while PSDE and MPE showed a chemical fingerprint closer to the original green propolis hydroalcoholic extract used. MPE, a fine powder (40% propolis in gum Arabic), was readily dispersible in water, and had less intense flavor, taste, and color than PSDE. PSDE, a fine powder (80% propolis) in maltodextrin as a carrier, was perfectly water-soluble and could be used in liquid formulations; it is transparent and has a strong bitter taste. PPF, a purified solid with large amounts of caffeic and p-Coumaric acids, had the highest antioxidant and antimicrobial activity, and therefore merits further study. PSDE and MPE had antioxidant and antimicrobial properties and could be used in products tailored to specific needs.

How Does Brazilian Green Propolis Act in Normal and Tumor Cells? Identification of Compounds in or out Monocytes or HEp-2 Cells.

The aim of this study was to identify propolis compounds after incubation of normal and tumor cells (monocytes and HEp-2 cells, respectively) with Brazilian green propolis, in the lysate and supernatant of cell cultures and within these cells by gas chromatography-mass spectrometry (GC/MS). Cinnamic acid derivatives were generally localized in the lysate of both cell lines after incubation, suggesting these compounds are actively transported across the membrane into the cytoplasm. Terpenes were also found in the lysate. Artepillin C, in contrast, was localised only in the supernatant. Some constituents were unobservable after incubation, especially in monocytes, suggesting the compounds had been degraded. Our findings shed light on the possible sites of action (intracellular or via a cell membrane protein) and the bioavailability of various constituents of propolis, as well as possible modes of delivery of bioactive constituents.

Brazilian Brown Propolis: an Overview About Its Chemical Composition, Botanical Sources, Quality Control, and Pharmacological Properties.

Brazil is one of the largest propolis producers in the world. Propolis is produced by bees from plant exudates and tissues, leading to many variations in the types of propolis. Generally, Brazilian propolis types are green, brown, and red. Despite not being the main research focus as the green and red propolis, brown propolis is the second most produced propolis type in Brazil and has tremendous economic and medicinal importance. Propolis has drawn attention with the rise in the search for healthier lifestyles, functional foods, biocosmetics, and natural products as therapeutic sources. This review covers the main chemical constituents identified in different types of Brazilian brown propolis, and their botanical sources, chemistry, and biological activities. The economic aspect of brown propolis is also presented. There are many gaps to be filled for brown propolis regarding the development of analytical methods, and quality control to allow its standardization, limiting its applicability in the food and pharmaceutical industries. Future perspectives regarding brown propolis research were discussed, especially biological activities, to support the medicinal uses of different types of brown propolis. Supplementary Information: The online version contains supplementary material available at 10.1007/s43450-023-00374-x.