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Betulinic acid (BA), a betulin derivative, is an important plant-based natural product. BA has many biological activities and attracts attention especially as a promising antitumour drug. Extraction from plants or semi-synthesis from betulin are the common methods to obtain BA, but these methods have some drawbacks. In recent years, microbial biotransformation of betulin to BA has been anticipated as an alternative method. Betulin is abundant in the outer bark of birch (Betula spp.) and obtained from these plants, however endophytes of Betula spp. have not been investigated for biotransformation of betulin to BA before. Therefore, this study aimed to investigate the culturable endophytes of Betula pubescens var. litwinowii (Doluch.) Ashburner & McAll and to determine their biotransformation capacity of betulin to BA. To this end, bacterial and fungal endophytes were isolated from the surface-sterilized root, stem, and branch samples. A total of 37 endophytes (11 bacteria and 26 fungi) were identified by polyphasic approach. All endophytes were screened for the biotransformation of betulin to BA. Betulin and BA content of the extracts were determined by RP-HPLC analysis. Two different bacterial and fungal biotransformation processes were designed. It was found that the fungus BRF59 produced BA via betulin biotransformation under the Fungal Biotransformation Process-2 when the fungus was acclimatized to betulin before biotransformation, and glycerol was used as a carbon source, instead of glucose, during bioprocess. The highest BA yield and BA concentration was obtained as 12.22±0.94 % and 80.85±2.64 µg/ml, respectively, under non-optimized conditions. The fungal isolate BRF59, belonging to the genus Fusarium according to ITS analysis, was further identified based on RPB2 sequence analysis and found that Fusarium lacertarum. In conclusion, this study demonstrated that betulin was biotransformed to BA by an endophytic fungus isolated from the genus Betula for the first time. F. lacertarum BRF59 might be a good candidate for production of BA through betulin biotransformation and, using glycerol as a carbon source during biotransformation might enable this by-product to convert value-added chemical. [Display omitted] • Culturable endophytes of Betula pubescens var. litwinowii (Doluch) was investigated. • Biotransformation of betulin to betulinic acid by endophytes was investigated. • Fusarium lacertarum BRF59 produced betulinic acid via betulin biotransformation. • The highest betulinic acid production was 80.85±2.64 µg/ml. • Acclimatization to betulin and using glycerol induced betulinic acid production. [ABSTRACT FROM AUTHOR]

Phenolic and triterpenoid compounds are essential components in stone fruit skin and flesh tissues. They are thought to possess general antimicrobial activity. However, regarding brown rot disease, investigations were only confined to a limited number of phenolics, especially chlorogenic acid. The activity of triterpenoids against Monilinia spp. , as an essential part of the peach cuticular wax, has not been studied before. In this work, the anti-fungal effect of some phenolics, triterpenoids, and fruit surface compound (FSC) extracts of peach fruit at two developmental stages were investigated on Monilinia fructicola and Monilinia laxa characteristics during in vitro growth. A new procedure for assaying anti-fungal activity of triterpenoids, which are notoriously difficult to assess in vitro because of their hydrophobicity, has been developed. Measurements of colony diameter, sporulation, and germination of second-generation conidia were recorded. Furthermore, the expression of twelve genes of M. fructicola associated with germination and/or appressorium formation and virulence-related genes was studied relative to the presence of the compounds. The study revealed that certain phenolics and triterpenoids showed modest anti-fungal activity while dramatically modulating gene expression in mycelium of M. fructicola on culture medium. MfRGAE1 gene was overexpressed by chlorogenic and ferulic acids and MfCUT1 by betulinic acid, at 4- and 7- days of mycelium incubation. The stage II FSC extract, corresponding to the period when the fruit is resistant to Monilinia spp., considerably up-regulated the MfLAE1 gene. These findings effectively contribute to the knowledge of biochemical compounds effects on fungi on in vitro conditions. • Peach surface compounds may have antifungal activity. • A protocol to assay hydrophobic compounds has been developed. • Few compounds have demonstrated limited antifungal capacity against Monilinia spp. • Many compounds dramatically modulated M. fructicola gene expression on culture medium. [ABSTRACT FROM AUTHOR]

• 38 strains of endophytic fungi were isolated from silver birch. The presence of Betulin and Betulinic acid was confirmed in 32 isolates. • The highest amount of betulin in Fusarium cicatricum (311.604 mg/ml) and Penicillium citrinum (409/381 mg/ml) and the highest amount of Betulinic acid in Alternaria alternate (431.275 mg/ml) was found. • Cellulose nanofibers increased the amount of terpenes by 27.5 and 8 times respectively in P. citrinum and F. fuckelii compared to the control. This study specifically aims to produce Betulin and Betulinic acid derivatives, which are anticancer terpenes by endophytic fungi isolated from the bark of silver Birch (Betula pendula Roth) for the first time. Betulin can be converted or decomposed into its derivatives over time, but all these derivatives are known, as terpenes how, are exciting prospects for application in medicine and cosmetics. In order to carry out this research, bark samples were collected from silver birch trees in Siahmarzkooh – Golsetan – Iran. Samples were cultured in potato dextrose agar (PDA) after surface sterilization. Thirty-eight endophytic fungi were isolated. The ability of Betulin and Betulinic acid induction by endophytic fungi was measured by HPLC and three isolates with the highest amount were selected and molecularly recognized (PCR-sequencing). In the next step, elicitors were induced into 2-year-old mycelium reserves of birch endophytic fungi. The cultures were extracted with three concentrations and three different times of cellulose nanofibers and sodium nitrate, and the changes of terpenes were measured by HPLC. The results showed that 32 of 38 isolates have terpene. Fusarium fuckelii (311.604 µg/mL), Penicillium citrinum (381/409 µg/mL) had the highest amount of Betulin, and Alternaria alternate (431.275 µg/mL) had the highest amount of Betulinc acid. The results reconfirmed the conversion of Betulin into its derivatives over time. Cellulosic nanofibers had a more significant effect on the increase of terpene as an elicitor, so 100 mg/L of it induced 27.5 and 8 times in P. citrinum and F. fuckelii , respectively, compared to the control. This study showed that the endophytic fungi isolated from the silver birch trees could produce, increase terpenes, and probably can be more easily used in the cosmetic or pharmaceutical industries. [ABSTRACT FROM AUTHOR]

The liana Urera gravenreuthii Engl. is an endemic plant to Cameroon which was not yet been chemically and pharmacologically investigated so far despite its uses for medical purposes in local folk medicine to treat several illnesses including dysentery, neuralgia and malaria. This study mainly aimed to contribute to providing further information on the chemical composition of the plant and to identify its antiplasmodial compounds which might support its effectiveness against malaria. The purification of the methanolic extract of U. gravenreuthii led to the isolation and characterization of fifteen distinct compounds (1 ‒ 15) which were structurally confirmed using their NMR data. Thirteen compounds (1 ‒ 11, 14 ‒ 15) were evaluated for their antiplasmodial activity against P. falciparum 3D7 and the results showed that the most active were betulinic acid (2) (2.2 ± 0.2 μ M), rutin (9) (2.8 ± 1.4 μ M), quercetin (8) (3.7 ± 1.3 μ M) and lupeol (1) (4.8 ± 0.3 μ M) which displayed a good activity with IC 50 < 5 μ M. Furthermore, the hypotheses on the structure-activity relationship of the tested compounds and the chemotaxonomic significance of the study have been discussed. The results support the identification of the plant as a member of the family Urticaceae, strengthen its use for the management of malaria and qualify it as an important source of good candidates in new antiplasmodial drug development. [Display omitted] • This is the first chemical and biological investigation of Urera gravenreuthii. • Fifteen compounds have been isolated from the methanolic extract. • Betulinic acid (2) and rutin (9) were the most active with IC 50 values of (2.2 ± 0.2 μM) and (2.8 ± 1.4 μM), respectively, against P. falciparum 3D7. • The structure-activity relationship of the isolated compounds has been discussed. • The obtained results classifies the plant as an important source of antiplasmodial leads for new drugs discovery. [ABSTRACT FROM AUTHOR]

The application of numerous chemotherapeutic drugs has been limited due to poor solubility, adverse side effects, and even multidrug resistance in patients. Polymeric micelles with reversibly cross-linked structures provide a promising solution to these issues. Herein, we optimized and synthesized programable-released disulfide cross-linked micelle (PDCM) based on our previous well-defined dendrimers to deliver the antitumor drug betulinic acid (BA) and paclitaxel (PDCM@PTX) and evaluated the therapeutic efficacy of multidrug-resistant (MDR) simulative orthotopic intraperitoneal ovarian cancer mice models. Comprehensive results demonstrated that PDCM@PTX formed stable nanoparticles able to improve the pharmacokinetic profile and circulation time of PTX, allowing for increased tumor penetration. Furthermore, in the tumor microenvironment, the programable-switches (ester bonds and disulfide cross-linking) of PDCM@PTX were cleaved by the high concentration of glutathione (tumor microenvironment) and esterase (intracellular) present in the tumor, allowing for in situ release of PTX and BA, resulting in intensive therapeutic efficacy in MDR ovarian cancer. [Display omitted] [ABSTRACT FROM AUTHOR]

Immunotherapy combined with effective therapeutics such as chemotherapy and photodynamic therapy have been shown to be a successful strategy to activate anti-tumor immune responses for improved anticancer treatment. However, developing multifunctional biodegradable, biocompatible, low-toxic but highly efficient, and clinically available transformed nano-immunostimulants remains a challenge and is in great demand. Herein, we report and design of a novel carrier-free photo-chemotherapeutic nano-prodrug COS-BA/Ce6 NPs by combining three multifunctional components—a self-assembled natural small molecule betulinic acid (BA), a water-soluble chitosan oligosaccharide (COS), and a low toxic photosensitizer chlorin e6 (Ce6)—to augment the antitumor efficacy of the immune adjuvant anti-PD-L1-mediated cancer immunotherapy. We show that the designed nanodrugs harbored a smart and distinctive "dormancy" characteristic in chemotherapeutic effect with desired lower cytotoxicity, and multiple favorable therapeutic features including improved 1O 2 generation induced by the reduced energy gap of Ce6, pH-responsiveness, good biodegradability, and biocompatibility, ensuring a highly efficient, synergistic photochemotherapy. Moreover, when combined with anti-PD-L1 therapy, both nano-coassembly based chemotherapy and chemotherapy/photodynamic therapy (PDT) could effectively activate antitumor immunity when treating primary or distant tumors, opening up potentially attractive possibilities for clinical immunotherapy. Natural small-molecule-mediated photosensitive pro-drug nano-assemblies that harbored a distinctive chemotherapeutic dormancy function, which can effectively generate robust immunological responses with the assistance of anti-PD-L1 to cause highly efficient, safe, and synergistic anti-tumor chemotherapy/photodynamic therapy/immunotherapy. [Display omitted] [ABSTRACT FROM AUTHOR]

The ethyl acetate fraction of Psorospermum aurantiacum stems has furnished cadensin H (1) a new compound belonging to the class of xanthonolignoid alongside with six compounds: 1,5,6-trihydroxy-7-methoxyxanthone (2), betulinic acid (3), 1,7-dihydroxyxanthone (4), betulin (5), lupeol (6) and stigmasterol glucoside (7). The crude methanolic extract and ethyl acetate fraction, as well as some major isolated compounds were evaluated for antibacterial and antioxidant activities. Shigella flexneri, Staphylococcus aureus, SSalmonella. typhi, EEscherichia coli and Klebsiella pneumonia , were used as test organisms. The crude extract has moderate activity against Shigella flexneri , methicillin-resistant Staphylococcus aureus, SSalmonella. typhi and EEscherichia. coli strains with MICs values ranging from 125 to 500 µ g/mL, and MBC of 500 µ g/mL for Staphylococcus aureus, SSalmonella. typhi and EEscherichia. coli. While the ethyl acetate fraction has moderate activity against S. flexneri , methicillin resistant S. aureus, SSalmonella. typhi and EEscherichia. coli with MIC ranging from 250 to 1000 µg/mL and MBC from 500 to 1000 µg/mL for S. flexneri, S. aureus, E. coli strains. Among all the compounds tested, only 1,5,6-trihydroxy-7-methoxyxanthone showed moderate activity on all bacterial strains with MICs values from 12.5 to 50 µ g/mL and MBC of 25 µg/mL and 100 µg/mL on methicillin-resistant S. aureus , 100 µg/mL on EEscherichia. coli. DPPH for radical scavenging and iron reducing antioxidant (FRAP) were used to evaluate antioxidant activity. Compound (2) showed important activity with the first reagent while compound (1) showed moderate activity with the second reagent. The structures of the isolated compounds were established using modern spectroscopic methods including 1D and 2D NMR and High-resolution mass spectrometry and by comparing with data reported in the literature. [Display omitted] • Isolation of 7 compounds from the stems of psorospermum aurantiacum (Hypericaceae). • Four of the compounds isolated are found for the first time in the genus psorospermum. • First report of antioxidant activities of compounds isolated from the stems of psorospermu m. [ABSTRACT FROM AUTHOR]

Betulinic acid (BA) is a natural pentacyclic triterpenoid with broad-spectrum anticancer activity, which has great development potential as an anti-cancer drug. In this study, a novel hyaluronic acid (HA)-modified BA liposome (BA-L) was developed for use in targeted liver cancer therapy. The size, polymer dispersity index (PDI), zeta potential, and entrapment efficiency were measured. Cell viability, cell migration and clonogenicity, cellular uptake, immunohistochemistry of CD44, and protein expression of ROCK1/IP3/RAS were also investigated. BA, BA-L, and HA-BA-L had no inhibitory effect on the activity of LO2 normal hepatocytes, but they inhibited the proliferation of HepG2 and SMMC-7721 cells in a dose- and time-dependent manner, with HA-BA-L exhibiting the most prominent inhibitory effect. Compared with the BA-L group, the expression of CD44 in HepG2 cells in the HA-BA-L group was decreased. The results of WB showed that BA, BA-L, and HA-BA-L downregulated the expression of ROCK1, IP3, and RAS in HepG2 cells, and the expression level in the HA-BA-L group was significantly decreased. The easily prepared HA-BA-L was demonstrated to be an excellent CD44-mediated intracellular delivery system capable of targeting effects. Further mechanistic research revealed that the inhibition of HA-BA-L on HepG2 cells may be mediated by blocking the ROCK1/IP3/RAS signaling pathways. [Display omitted] [ABSTRACT FROM AUTHOR]

[Display omitted] • A simple and universal strategy was developed for the fabrication of self-stacked hydrogels of bioactive divalent metal ions coordinated with pharmacological small molecules. • We elucidated and summarised the relationship between the molecular structure and gelling ability of these natural self-stacked small molecule gels. • The synergistic effects of betulinic acid and Mg2+ were harnessed within the self-stacked hydrogel, enhancing its pharmacological impact. • In vivo studies displayed that the BA-Mg/BC composite hydrogel significantly accelerated scar-free wound healing. Despite significant advancements in self-stacked hydrogels of herbal small molecules in recent years, the emergence of new pharmacological small molecule self-stacked hydrogel systems remains largely reliant on serendipity and painstaking efforts. To address this challenge, we proposed a 'bioactive bivalent metal ions-coordinated pharmacological small molecules' strategy. This approach enabled the preparation of triterpenoid betulinic acid-based self-stacked hydrogels with excellent injectable and self-healing properties via a simple two-step method. We extended this strategy to self-stacking hydrogels based on other pentacyclic triterpene small molecules, anthraquinone small molecules, flavonoid small molecules and synthetic steroid small molecules. This universal strategy is capable of extracting hydrophobic pharmacological small molecules to construct novel self-stacked biomaterials in a cost-effective, efficient, large-scale, and environmentally friendly manner. Importantly, we successfully synthesized Mg2+-coordinated betulinic acid (BA-Mg) self-stacked hydrogel at low concentrations in the bacterial cellulose (termed BC) hydrogel network, overcoming the mechanical limitations of self-stacked hydrogels for biomedical applications. The BA-Mg/BC hybrid hydrogel demonstrated exceptional biocompatibility, anti-inflammatory, and angiogenic properties, significantly accelerating the skin wound healing process through the synergistic pharmacological effects of Mg2+ and BA. [ABSTRACT FROM AUTHOR]

• Betulinic acid improves the cognitive functions of zebrafish subsequent TiO 2 NP treatment. • Betulinic acid significantly decreases proinflammatory markers such as TNF-α, IL-1β, and IL-6 in the brain of the zebrafish brain. • Betulinic acid mitigates oxidative stress via activating the Nrf-2/HO-1 signaling in the TiO 2 NP model of zebrafish confirmed from histopathology and immunohistochemistry analysis. Betulinic acid (BA) is a natural triterpenoid extracted from Bacopa monnieri. BA has been reported to be used as a neuroprotective agent, but their molecular mechanisms are still unknown. Therefore, in this study, we attempted to investigate the precise mechanism of BA for its protective effect against Titanium dioxide nanoparticles (TiO2NP) induced neurotoxicity in zebrafish. Hence, our study observation showed that 10 µg/ml dose of TiO2NP caused a rigorous behavioral deficit in zebrafish. Further, biochemical analysis revealed TiO 2 NP significantly decreased GSH, and SOD, and increased MDA, AChE, TNF-α, IL-1β, and IL-6 levels, suggesting it triggers oxidative stress and neuroinflammation. However, BA at doses of 2.5,5,10 mg/kg improved behavioral as well as biochemical changes in zebrafish brain. Moreover, BA also significantly raised the levels of DA, NE, 5-HT, and GABA and decreased glutamate levels in TiO 2 NP-treated zebrafish brain. Our histopathological analysis proved that TiO 2 NP causes morphological changes in the brain. These changes were expressed by increasing pyknotic neurons, which were dose-dependently reduced by Betulinic acid. Likewise, BA upregulated the levels of NRF-2 and HO-1, which can reduce oxidative stress and neuroinflammation. Thus, our study provides evidence for the molecular mechanism behind the neuroprotective effect of Betulinic acid. Rendering to the findings, we can consider BA as a suitable applicant for the treatment of AD-like symptoms. [ABSTRACT FROM AUTHOR]

Betulinic acid (BA), which is a pentacyclic triterpenoid found in the bark of plane, birch, and eucalyptus trees, has emerged as a compound of significant interest in scientific research due to its potential therapeutic applications. BA has a range of well-documented pharmacological and biological effects, including antibacterial, immunomodulatory, diuretic, antiviral, antiparasitic, antidiabetic, and anticancer activities. Although numerous research studies have explored the potential anticancer effects of BA, there is a noticeable gap in the literature, highlighting the need for a more up-to-date and comprehensive evaluation of BA's anticancer potential. The aim of this work is to critically assess the reported cellular and molecular mechanisms underlying the cancer preventive and therapeutic effects of BA. Relevant research on the inhibitory effects of BA against cancerous cells was searched using Science Direct, Scopus, Web of Science, and PubMed following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. The anticancer properties of BA are mediated by the activation of cell death and cell cycle arrest, production of reactive oxygen species, increased mitochondrial permeability, modulation of nuclear factor-κB and Bcl-2 family signaling. Emerging evidence also underscores the combined anticancer effects of BA with other natural bioactive compounds or approved drugs. Notably, several novel BA nanoformulations have been found to exhibit encouraging antineoplastic activities. BA, whether used alone or in combination, or as a form of nanoformulation, shows significant potential for cancer prevention and treatment. Nevertheless, further detailed studies are necessary to confirm the therapeutic effectiveness of this natural compound. [Display omitted] [ABSTRACT FROM AUTHOR]

• 1,2,3-Triazole-based betulinic acid derivatives were synthesized as α-glucosidase inhibitors. • All the derivatives exhibited excellent inhibition against α-glucosidase. • The most active compound d23 acted as a mixed-type inhibitor for α-glucosidase. • Compound d23 exerted its inhibitory activity by changing the conformation of α-glucosidase. • Compound d23 reduced fasting blood glucose level and improved glucose tolerance and dyslipidemia. A series of 1, 2, 3-triazole-based betulinic acid derivatives d1 – d31 were synthesized as α-glucosidase inhibitors with hypoglycemic activity. All the derivatives exhibited excellent inhibition against α-glucosidase, and compound d23 was the most active (IC 50 = 2.83 ± 0.19 μM). Inhibition kinetics showed that compound d23 was a mixed-type inhibitor for α-glucosidase. Spectroscopic studies based on 3D fluorescence and circular dichroism spectra suggested that compound d23 exerted its inhibitory activity by changing the conformation of α-glucosidase. Molecular docking revealed that compound d23 was well nested into the active pocket of α-glucosidase through hydrogen-bonding and hydrophobic interactions. In vivo experiments showed that compound d23 could not only reduce the level of fasting blood glucose, but also improve glucose tolerance and dyslipidemia. The present findings strongly suggest that compound d23 is exploitable as a leading compound for the development of α-glucosidase inhibitors in the treatment of type 2 diabetes. [Display omitted] [ABSTRACT FROM AUTHOR]

The NF-κB pathway plays a pivotal role in impeding the diabetic wound healing process, contributing to prolonged inflammation, diminished angiogenesis, and reduced proliferation. In contrast to modern synthetic therapies, naturally occurring phytoconstituents are well-studied inhibitors of the NF-κB pathway that are now attracting increased attention in the context of diabetic wound healing because of lower toxicity, better safety and efficacy, and cost-effectiveness. This study explores recent research on phytoconstituent-based therapies and delve into their action mechanisms targeting the NF-κB pathway and potential for assisting effective healing of diabetic wounds. For this purpose, we have carried out surveys of recent literature and analyzed studies from prominent databases such as Science Direct, Scopus, PubMed, Google Scholar, EMBASE, and Web of Science. The classification of phytoconstituents into various categorie such as: alkaloids, triterpenoids, phenolics, polyphenols, flavonoids, monoterpene glycosides, naphthoquinones and tocopherols. Noteworthy phytoconstituents, including Neferine, Plumbagin, Boswellic acid, Genistein, Luteolin, Kirenol, Rutin, Vicenin-2, Gamma-tocopherol, Icariin, Resveratrol, Mangiferin, Betulinic acid, Berberine, Syringic acid, Gallocatechin, Curcumin, Loureirin-A, Loureirin-B, Lupeol, Paeoniflorin, and Puerarin emerge from these studies as promising agents for diabetic wound healing through the inhibition of the NF-κB pathway. Extensive research on various phytoconstituents has revealed how they modulate signalling pathways, including NF-κB, studies that demonstrate the potential for development of therapeutic phytoconstituents to assist healing of chronic diabetic wounds. [Display omitted] • Phytoconstituents show promise in diabetic wound healing by inhibiting NF-κB, with lower toxicity and better safety. • NF-kB in diabetic wound healing coordinates cytokine secretion, growth factor release, and cell proliferation. • NF-κB is regulated by inhibitors like IκBα, IκBβ, and IκBγ, maintaining NF-κB pathway inactivation. [ABSTRACT FROM AUTHOR]

[Display omitted] • Modified in the A ring betulinic acid derivatives were synthesized. • Methodology to access 2α-(aminomethyl)betulinic acid derivatives has been achieved. • 3α-Alkynylbetulinic acid derivatives by the reaction of lithium acetylides with betulonic acid ester. • Catalytic system composed of AuCl 3 –AgOTf induced the elimination reaction. • The anticancer activities were investigated against various tumor cell lines. Betulonic acid benzyl ester 1 has been subjected to a series of structural modifications for the purpose of new triterpenoid synthesis and evaluating for anticancer activity. The one-pot two step synthesis of 2α-(aminomethyl)betulinic acid benzyl ester derivatives 3a–f (yield 46–69 %) was achieved by the Mannich reaction of compound 1 with methyleneiminium salts, generated in situ from N, N -disubstituted bis(amino)methanes 2a–f by the action of acetyl chloride in dichloromethane, and subsequent reduction of aminomethylation products with sodium borohydride. Minor 2β-(aminomethyl) triterpenoids 4c , d , f were also isolated (yield 6–15 %). We found, that the stereoselective reaction of triterpenoid 1 with acetylides, generated at –78 °C from alkynes in the presence of n -BuLi, has been useful and noteworthy as the key step in providing of new alkyne substituted triterpenoids – benzyl 3-alkynyl-3-deoxy-2(3),20(29)-lupadiene-28-oates or 3-deoxy-2(3)-dehydro-28-oxoallobetulin derivative. The new compounds were examined for anticancer activity against the human cell lines (MTT assay). All tested derivatives were non-toxic on human fibroblasts. The 3‐(phenylethynyl)lupa-2(3),20(29)-diene 9 showed selective cytotoxicity on cervical cancer cell lines. Tumor cells death trigged by the most active compound 4f resulted from apoptotic processes. These data make the series of synthesized 2 or 3 substituted lupane derivatives as promising compounds with anticancer potential. [ABSTRACT FROM AUTHOR]

Target identification of bioactive compounds is important for understanding their mechanisms of action and provides critical insights into their therapeutic utility. While it remains a challenge, unbiased chemoproteomics strategy using clickable photoaffinity probes is a useful and validated approach for target identification. One major limitation of this approach is the efficient synthesis of appropriately substituted clickable photoaffinity probes. Herein, we describe an efficient and consistent method to prepare such probes. We further employed this method to prepare a highly stereo-congested probe based on naturally occurring triterpenoid betulinic acid. With this photoaffinity probe, we identified tropomyosin as a novel target for betulinic acid that can account for the unique biological phenotype on cellular cytoskeleton induced by betulinic acid. A new method was developed to reliably prepare stereo-hindered clickable diazirines for target identification. Tropomyosin was identified as a new target for betulinic acid. [Display omitted] [ABSTRACT FROM AUTHOR]

• New 3-isopropyl-2,6-diphenyl-1-(2-(thiophen-2yl)acetyl)piperidin-4-one (IPTP) single crystals analysis. • The Hirshfeld surface analysis was employed to examine intermolecular contacts. • Compare the spectral studies of DFT experimental and theoretical and vibrational energy distribution analysis (VEDA) analysis. • Anti-HIV activities were observed for synthesized compounds. • FDA-approved anti-HIV drug of Nelfinavir molecular docking and ADME (Computational study) have been predicted. The 3-isopropyl-2,6-diphenyl-1-(2-(thiophen-2yl)acetyl)piperidin-4-one (IPTP) single crystals were grown by slow evaporation method and the structure is confirmed through FT-IR, 1H NMR, and 13C NMR spectral analyses. Single crystal X-ray diffraction revealed that the title compound having chair conformation of the piperidine ring, with an axial orientation of phenyl rings at C-2 and C-6 positions and an isopropyl group at C-3. Hirshfeld surface analysis was employed to examine intermolecular contacts and strong and weak attractive, repulsive, and van der Waals interactions in the molecule were determined using the reduced density gradient method (RDG). The geometrical parameters obtained through Density Functional Theory are compared with experimental values and also conducted to optimize structural parameters, frontier molecular orbitals (FMOs), and molecular electrostatic potential surface analysis (MEPS). NBO calculations were employed to investigate intermolecular and intramolecular charge movement, as well as the molecule's stability. Vibrational Energy Distribution Analysis (VEDA) software facilitated potential energy decomposition (PED) analysis using FT-IR wave numbers. The title compound molecular docking data were compared with the markedly available FDA-approved anti-HIV drug of Nelfinavir and their results were discussed, ADME analysis studies were discussed concerning FDA-approved anti-HIV drugs, including Nelfinavir, Betulinic Acid, Haloperidol, and Donepezil. [Display omitted] [ABSTRACT FROM AUTHOR]

• Novel betulinic acid derivatives containing 1,2,4-triazole- derived schiff base f1–f35 were synthesized and screened against α-glucosidase. • Compound f32 showed the highest α-glucosidase inhibitory with IC 50 of 1.52±0.16 μM. • Inhibition kinetics showed that compound f32 was a non-competitive inhibitor for α-glucosidase. • Spectroscopic studies based on three dimensional fluorescence and circular dichroism and molecular docking suggested that compound f32 interacted with α-glucosidase. • In vivo experiments showed that compound f32 could not only reduce the level of fasting blood glucose, but also improve glucose tolerance and dyslipidemia. A series of novel betulinic acid (BA) derivatives containing 1,2,4-triazole- derived schiff base have been designed and synthesized as α-glucosidase inhibitors. All the title compounds showed higher α-glucosidase inhibitory activity than acarbose and BA, with compound f32 showing the highest α-glucosidase inhibitory activity (IC 50 = 1.52±0.16 μM). Lineweaver-Burk plot analysis suggested that compound f32 was a non-competitive inhibitor for α-glucosidase. 3D fluorescence and circular dichroism spectroscopy disclosed the interaction of compound f32 with α-glucosidase by changing the secondary structure of α-glucosidase. Molecular docking showed that compound f32 could bind to the active site of α-glucosidase through hydrogen bonding and hydrophobic interaction. More importantly, compound f32 could not only reduce the level of fasting blood glucose and postprandial blood glucose in mice, but also ameliorate dyslipidemia. The current findings suggest that compound f32 may serve as a leading compound for the discovery of α-glucosidase inhibitors in the treatment of type 2 diabetes. [Display omitted] [ABSTRACT FROM AUTHOR]

Pain and inflammation are the most frequent reasons for which people seek medical care. Currently available analgesics against these conditions produce fatal adverse effects. NPK 500 capsules is an alternative herbal analgesic employed to treat dysmenorrhea, peptic ulcer and pain. NPK 500 is produced from Cassia sieberiana. A plant used in traditional medicine to treat pain and inflammation. This study reports the analysis, phytochemical characterization and mechanism of analgesic and anti-inflammatory activities of two NPK 500 capsules, called old and new NPK500 capsules (ONPK500 and NNPK500) respectively. Physicochemical, organoleptic, GC-MS and LC-MS methods were employed to analyze the NPK 500 capsules. Analgesic activity was evaluated using tail immersion, Randall-Selitto and acetic acid induced writing tests. Anti-inflammatory activity was evaluated using carrageenan-induced rat paw inflammation. Additionally, pro-inflammatory mediators such as prostaglandin E2 (PGE2), inducible nitric oxide synthase (iNOS), cyclooxygenase 1 and 2 (COX-2 and COX-1) were quantified in the sera of the rats using Enzyme Linked Immunosorbent Assay (ELISA) kits. Thirteen major compounds were characterized in the NNPK 500 capsules via the GC-MS and LC-MS spectroscopies. Kaempferol was the major compound characterized in addition to physcion, β-sitosterol 3-O-β-D-glucopyranoside, betulinic acid and nine others. Physicochemical and organoleptic indices of the capsules were also derived for its authentication and quality control. Furthermore, NNPK 500 0.5–1.5 mg/kg p.o. produce significant (P < 0.5) analgesic activity (160–197%) higher than that of ONPK500 (109.8%) and Morphine (101%) in the tail immersion test. The analgesic activity of NNPK 500 0.5–1.5 mg/kg p.o. (171.0–258.3%) and ONPK 500 (179.5%) were also significant (P < 0.01) and higher than that of Aspirin (103.00%) in the Randall-Selitto test. Both capsules also demonstrated significant (P < 0.5) analgesic and anti-inflammatory activities in the acetic acid-induced writhing and carrageenan-indued paw edema tests respectively. The two NPK500 capsules also, significantly (P < 0.5) inhibited PGE2 and iNOS but not COX-2 and COX-1 in the carrageenan-indued paw edema test. These results show that NNPK 500 and ONPK 500 capsules possessed potent analgesic and anti-inflammatory activities via inhibition of PGE2 and iNOS as a result of their chemical constituents. NPK500 capsules thus, relief acute pain and inflammation without causing gastrointestinal, renal or hepatic injuries, since they did not inhibit COX-1. [Display omitted] • The major active ingredient identified in NPK500 capsules was kaempferol. • Physcion, β-sitosterol 3-O-β-D-glucopyranoside and betulinic acid were also present. • Analgesic activity of NPK500 was 171.0–258.3% against 103.00% for Aspirin. • NPK500 also produced significant anti-inflammatory activity. • NPK500 capsules inhibited PGE2 and iNOS during inflammation. [ABSTRACT FROM AUTHOR]

Fifteen betulonic/betulinic acid conjugated with nucleoside derivatives were synthesized to enhance antitumor potency and water solubility. Among these, the methylated betulonic acid-azidothymidine compound (8c) exhibited a broad-spectrum of antitumor activity against three tested tumor cell lines, including SMMC-7721 (IC 50 = 5.02 μM), KYSE-150 (IC 50 = 5.68 μM), and SW620 (IC 50 = 4.61 μM) and along with lower toxicity (TC 50 > 100 μM) estimated by zebrafish embryos assay. Compared to betulinic acid (<0.05 μg/mL), compound 8c showed approximately 40-fold higher water solubility (1.98 μg/mL). In SMMC-7721 cells, compound 8c induced autophagy and apoptosis as its concentration increased. Transcriptomic sequencing analysis was used to understand the potential impacts of the underlying mechanism of 8c on SMMC-7721 cells. Transcriptomic studies indicated that compound 8c could activate autophagy by inhibiting the PI3K/AKT pathway in SMMC-7721 cells. Furthermore, in the xenograft mice study, compound 8c significantly slowed down the tumor growth, as potent as paclitaxel treated group. In conclusion, methylated betulonic acid-azidothymidine compound (8c) not only increases water solubility, but also enhances the potency against hepatocellular carcinoma cells by inducing autophagy and apoptosis, and suppressing the PI3K/Akt/mTOR signaling pathway. [Display omitted] • Conjugated triterpenes with nucleoside derivatives exhibited increased anti-tumor activity and solubility. • The methylated betulonic acid-azidothymidine derivative inducing autophagy and apoptosis in hepatocellular carcinoma cells. • Transcriptomic studies revealed its activation of autophagy by suppressing the PI3K/Akt/mTOR pathway in SMMC-7721 cells. [ABSTRACT FROM AUTHOR]

The present study deals with the phytochemical study of the leaf extract of C. klainei , a plant used in folk medicine to treat diabetes, venereal diseases, hypertension, malaria, cough and hemorrhoids. The liquid-liquid extraction and chromatographic separation of the dichloromethane/methanol (1:1) extract of this plant led to the isolation of eleven (11) known compounds identified from the analysis of their spectroscopic and physical data, and in comparison, with data from the literature. These compounds were identified to hentriancontane (1), β -sitosterol (2), stigmasterol (3), β -amyrin (4), betulinic acid (5), sitosterol-3- O - β - D -glucopyranoside (6), diosmetin (7), iristectorigenin A (8) , gallic acid (9), bergenin (10) and scyllitol (11) respectively. Furthermore, the chemotaxonomic significance of these compounds was discussed. [Display omitted] • Many classes of secondary metabolites are found in Coelocaryon klainei. • Nine of the eleven isolated compounds are newly reported from the genus Coelocaryon. • Polyphenols could be considered as chemotaxonomic markers of the genus Coelocaryon. [ABSTRACT FROM AUTHOR]

The phytochemical investigation on Mesua lepidota T. Anderson led to the identification of eight Mammea coumarins [lepidotols A-B (1–2), lepidotol E (3), lepidotins A-C (4–6), mammea A/BB cyclo F (7), and ochrocarpin E (8)], five triterpenes [friedelin (9), friedelinol (10), betulinic acid (11), glutinol (12), and α-amyrin (13)], one xanthone, namely pyranojacareubin (14), and two sterols [β-sitosterol (15) and stigmasterol (16)]. Identification was conducted by 13C-NMR dereplication and after a conventional isolation process. The structural elucidation of the isolated compounds was conducted through the analysis of spectroscopic data and comparison with literature data. The chemotaxonomic significance of all compounds was discussed, highlighting that most compounds were isolated only in the Mesua genus, or specifically from Mesua lepidota. In addition, glutinol (12) was isolated for the first time from the genus Mesua. • Sixteen compounds were identified from Mesua lepidota T. Anderson. • Glutinol was first reported from Mesua genus. • The hypothetical biogenesis pathway of isolated Mammea coumarins was discussed. • The chemotaxonomic significance of isolated metabolites was discussed. [ABSTRACT FROM AUTHOR]

• Betulinic acid is effective for damaging interkingdom biofilms Candida albicans and Staphylococcus aureus. • Betulinic acid inhibits adhesion and reduces preformed biofilms. • Betulinic acid is able to interact with C. albicans cell membrane (composition, hydrophobicity, fluidity). • Betulinic acid decreases the transcription of S. aureus biofilm-related gene sas F. The demand for antibiofilm molecules has increased over several years due to their potential to fight biofilm-associated infections, such as those including the interkingdom Staphylococcus aureus – Candida albicans occurring in clinical settings worldwide. Recently, we identified a pentacyclic triterpenoid compound, betulinic acid, from invasive macrophytes, with interesting antibiofilm properties. The aim of the present study was to provide insights into the mechanism of action of betulinic acid against the clinically relevant bi-species S. aureus – C. albicans biofilms. Microscopy examinations, flow cytometry and crystal violet assays confirmed that betulinic acid was effective at damaging mature S. aureus – C. albicans biofilms or inhibiting their formation, reducing biofilm biomass by 70% on average and without microbicidal activity. The results suggested an action of betulinic acid on cell membranes, inducing changes in properties such as composition, hydrophobicity and fluidity as observed in C. albicans , which may hinder the early adhesion step, biofilm growth and the physical interactions of both microbial species. Further results of real-time polymerase chain reaction argued in favour of a reduction in S. aureus – C. albicans physical interaction due to betulinic acid by the modulation of biofilm-related gene expression, as observed in early stages of biofilm formation. This study revealed the potential of betulinic acid as a candidate agent for the prevention and treatment of S. aureus – C. albicans biofilm-related infections. [Display omitted] [ABSTRACT FROM AUTHOR]

Patients with myocardial infarction have a much worse prognosis when they have myocardial ischemia-reperfusion (I/R) injury. Further research into the molecular basis of myocardial I/R injury is therefore urgently needed, as well as the identification of novel therapeutic targets and linkages to interventions. Three cysteine residues are present in DJ-1 at amino acids 46, 53, and 106 sites, with the cysteine at position 106 being the most oxidation-prone. This study sought to understand how oxidized DJ-1(C106) contributes to myocardial I/R damage. Rats' left anterior descending branches were tied off to establish a myocardial I/R model in vivo. A myocardial I/R model in vitro was established via anoxia/reoxygenation (A/R) of H9c2 cells. The results showed that autophagy increased after I/R, accompanied by the increased expression of oxidized DJ-1 (ox-DJ-1). In contrast, after pretreatment with NAC (N-acetylcysteine, a ROS scavenger) or Comp-23 (Compound-23, a specific antioxidant binding to the C106 site of DJ-1), the levels of ox-DJ-1, autophagy and LDH release decreased, and cell survival rate increased. Furthermore, the inhibition of interaction between ox-DJ-1 and PTEN could increase PTEN phosphatase activity, inhibit the p-IKK/NF-κB/Beclin1 pathway, reduce injurious autophagy, and alleviate A/R injury. However, BA (Betulinic acid, a NF-κB agonist) was able to reverse the protective effects produced by Comp-23 pretreatment. In conclusion, ox-DJ-1 could activate detrimental autophagy through the PTEN/p-IKK/NF-κB/Beclin1 pathway and exacerbate myocardial I/R injury. [ABSTRACT FROM AUTHOR]

[Display omitted] • Triterpenoic acids benzylamides were synthesized and sulfamoylated. • These compounds were tested as inhibitors of carbonic anhydrases (CA) I, II, VA and IX. • An Asiatic acid derived trisulfamate held strong inhibitory effects for CA VA. • This compound showed a K i = 36.2 nM thus outperforming standard acetalzolamide. This investigation delves into the inhibitory capabilities of a specific set of triterpenoic acids on diverse isoforms of human carbonic anhydrase (h CA). Oleanolic acid (1), maslinic acid (2), betulinic acid (3), platanic acid (4), and asiatic acid (5) were chosen as representative triterpenoids for evaluation. The synthesis involved acetylation of parent triterpenoic acids 1 – 5 , followed by sequential reactions with oxalyl chloride and benzylamine, de-acetylation of the amides, and subsequent treatment with sodium hydride and sulfamoyl chloride, leading to the formation of final compounds 21 – 25. Inhibition assays against h CAs I, II, VA, and IX demonstrated noteworthy outcomes. A derivative of betulinic acid, compound 23 , exhibited a K i value of 88.1 nM for h CA VA, and a derivative of asiatic acid, compound 25 , displayed an even lower K i value of 36.2 nM for the same isoform. Notably, the latter compound displayed enhanced inhibitory activity against h CA VA when compared to the benchmark compound acetazolamide (AAZ), which had a K i value of 63.0 nM. Thus, this compound surpasses the inhibitory potency and isoform selectivity of the standard compound acetazolamide (AAZ). In conclusion, the research offers insights into the inhibitory potential of selected triterpenoic acids across diverse h CA isoforms, emphasizing the pivotal role of structural attributes in determining isoform-specific inhibitory activity. The identification of compound 25 as a robust and selective h CA VA inhibitor prompts further exploration of its therapeutic applications. [ABSTRACT FROM AUTHOR]

The phytochemical investigation of the leaves of Adinandra glischroloma Hand.–Mazz. resulted in the isolation of fourteen known compounds including nine triterpenoids: 28- nor -urs-12-ene-3 β ,17 β -diol (1), micromeric acid (2), 23-hydroxy ursolic acid (3), euscaphic acid (4), pomolic acid (5), 3,13-dihydroxy ursolic acid 28,13-olide (6), ursolic acid (7), betulinic acid (8), and oleanolic acid (9); one diterpene: ent -kaur-16-en-19-oic-acid (10); one megastigmane: (3 S, 5 R, 6 S, 9 R)-megastigmane-3,9-diol (11), one flavonoid: isoquercetin (12); one lignan: syringaresinol (13), and one sterol: β -sitosterol (14). The chemical structures were determined by NMR, MS spectroscopic analyses and comparison with literature values. All of these compounds were isolated from A. glischroloma for the first time. Furthermore, this is the first report on the isolation of compounds 1 – 3 , 10 , and 11 from the genus Adinandra and the Pentaphylacaceae family. Compounds 1 – 3 , 10 , and 11 may be used as chemotaxonomic markers to differentiate between A. glischromola from other species of the Adinandra genus. Our results showed that A. glischroloma plant is rich in pentacyclic triterpenes and possessed a strong relationship with other Adinandra species in Vietnam. [Display omitted] • Fourteen compounds 1 – 14 were identified from the leaves of Adinandra glischroloma. • Compounds 1 – 3 , 10 , and 11 are reported for the first time from the Pentaphylacaceae family. • The chemical composition of A. glischroloma plant is rich in pentacyclic triterpenes. [ABSTRACT FROM AUTHOR]

T-2 toxin, a type-A trichothecene mycotoxin, exists ubiquitously in mildewed foods and feeds. Betulinic acid (BA), a pentacyclic triterpenoid derived from plants, has the effect of relieving inflammation and oxidative stress. The purpose of this study was to investigate whether BA mitigates lung impairment caused by T-2 toxin and elucidate the underlying mechanism. The results indicated that T-2 toxin triggered the inflammatory cell infiltration, morphological alterations and cell apoptosis in the lungs. It is gratifying that BA ameliorated T-2 toxin-caused lung injury. The protein expression of nuclear factor erythrocyte 2-related factor 2 (Nrf2) pathway and the markers of antioxidative capability were improved in T-2 toxin induced lung injury by BA mediated protection. Simultaneously, BA supplementation could suppress T-2 toxin-induced mitogen-activated protein kinase (MAPK)/nuclear factor-kappa B (NF-κB)-dependent inflammatory response and mitochondrial apoptotic pathway. Therefore, T-2 toxin gave rise to pulmonary toxicity, but these changes were moderated by BA administration through regulation of the Nrf2/MAPK/NF-κB pathway, which maybe offer a viable alternative for mitigating the lung impairments caused by the mycotoxin. BA protects against T-2 toxin-induced lung damage by enhancing the antioxidant capacity, inhibiting the inflammatory response and cell apoptosis, all of which are associated with the activation of Nrf2-mediated antioxidant defense and the suppression of MAPK/NF-κB-mediated inflammation signaling pathways. [Display omitted] • BA ameliorated T-2 toxin-induced lung damage. • BA alleviated pulmonary oxidative stress via activating Nrf2 pathway. • BA inhibited pulmonary inflammation via regulating NF-κB/MAPK pathways. • BA reduced pulmonary apoptosis by down-regulating mitochondrial apoptotic pathway. [ABSTRACT FROM AUTHOR]

• Optimal culture media for callus induction was NT+1.5 mg/l of 2,4-D and 0.5 mg/l BAP. • Nano-elicitation can increase betulin and betulinic acid in cell cultures. • 1 day elicitation in 0.5% Cellulose nano-fiber induced the highest amount of betulin. • 3 days elicitation in 0.5% Chitin nano-fiber induced the highest amount of betulinic acid. • Maximum antioxidant activity was created in 2 days elicitation with 0.5% Chitosan nano-fiber. This study aimed to develop a protocol for callus induction and also to evaluate the effects of chitosan nano-fiber (CTS-N), chitin nano-fiber (CT-N), and cellulose nano-fiber (Cellul-N) as elicitors on betulin and betulinic acid production alterations in the produced stem cells of a hybrid of Betula pendula. Inner birch bark samples were cultured in three different WPM, NT, and B5 media applying three hormonal treatments. Callogenesis was repeated in the selected medium providing the maximum percentage of callus induction (i.e., NT medium enriched with 1.5 mg.L−1 of 2,4-D + 0.5 mg.L−1 of BAP) and was observed for 5 months. Calluses were re-cultured in the medium with CTS-N, CT-N, and Cellul-N as elicitors at different concentrations of 0.5, 0.75, and 1 mg.L−1 and were extracted after specific time intervals (i.e., one, two, and three days) followed by determining their fresh and dry weights. Also, betulin and betulinic acid levels were determined by High Performance Liquid Chromatography (HPLC). As the results revealed, the heaviest fresh and dry weights (1.5-fold heavier than the control) were obtained for calli treated with 1% CTS-N for three days. Furthermore, one day elicitation using 0.5% Cellul-N induced the highest amount of betulin (0.7 mg.g-1), and the highest amount of betulinic acid (0.98 mg.g-1) was obtained in the callies treated by 0.5% CT-N for 3 days, while the corresponding weights for the control treatments were measured 0.08 mg.g-1 and 0.002 mg.g-1, respectively. And the maximum antioxidant activity of 93.9% was achieved from the elicitation utilizing 0.5% CTS-N for two days. This is the first report on the noticeable impact of nanomaterials, i.e., CTS-N, CT-N, and Cellul-N, on increasing botulin and betulinic acid accumulation along with the antioxidant activity, which, of course, requires further scrutiny. [ABSTRACT FROM AUTHOR]

Natural products are one of the important sources for the discovery of new drugs. Betulinic acid (BA), a pentacyclic triterpenoid widely distributed in the plant kingdom, exhibits powerful biological effects, including antitumor activity against various types of cancer cells. A considerable number of BA derivatives have been designed and prepared to remove their disadvantages, such as poor water solubility and low bioavailability. This review summarizes the current studies of the structural diversity of antitumor BA derivatives within the last five years, which provides prospects for further research on the structural modification of betulinic acid. [Display omitted] [ABSTRACT FROM AUTHOR]

The need of novel antileishmanial compounds with a wide therapeutic index has been a challenge for the pharmaceutical industry in reducing the side effects resulted from treatment with chemotherapeutics. The present study aims to investigate the antileishmanial and cytotoxic activities of the ethanol extract (EtOH), the dichloromethane fraction (DCM) and its isolated constituent betulinic acid (BA) obtained from the stem bark of Mimosa caesalpiniifolia Benth. (Fabaceae). The antiproliferative effect on Leishmania amazonensis promastigotes was assessed by the Alamar Blue® colorimetric assay. The intramacrophagic amastigotes were exposed to different concentrations of EtOH, DCM and BA to determine the survival index. Selectivity indexes towards parasites were calculated for each treatment performed. The modulation of macrophage activation state was investigated by observing structural (phagocytic and lysosomal activities) and cellular (nitrite production) changes. The EtOH, DCM and BA were effective against Leishmania amazonensis promastigote forms (IC 50 : 4.63 ± 0.69, 0.38 ± 0.42 and 29.63 ± 4.63 µg•mL−1, respectively). The same was observed for axenic amastigotes forms (IC 50 : 4.8 ± 1.27, 0.56 ± 0.39 and 14.2 ± 2.01 µg•mL−1), with selectivity index higher than 20 for DCM and BA. The macrophages infection and the number of intracellular amastigotes were reduced to 54.6%, 52.7% and 27.5%, respectively, after treatments. On the other hand, both DCM and BA were not able to promote macrophages activation, but only EtOH. Since DCM have shown highest potency as antileishmanial agent, BA presented a highest selectivity index. These results demonstrate the importance of M. caesalpiniifolia as a source of promising compounds since both DCM and BA might be investigated as promising and safe candidates for treatment of leishmaniasis. This work opens for further studies with M. caesalpiniifolia and betulinic acid focused on elucidation of possible involved mechanisms of antileishmanial action, as well as the potential in vivo treatment in experimental models of leishmaniasis. [Display omitted] • Mimosa caeasalpiniifolia -derived products induces antileishmanial effect. • Betulinic acid (BA) and dichloromethane fraction (DCM) presents highest selectivity to parasites. • Macrophages activation probably underlies the ethanol extract (EtOH)-induced antileishmanial effect. [ABSTRACT FROM AUTHOR]

Snakebites are considered a major neglected tropical disease, resulting in around 100,000 deaths per year. The recommended treatment by the WHO is serotherapy, which has limited effectiveness against the toxins involved in local tissue damage. In some countries, patients use plants from folk medicines as antivenoms. Aegiphila species are common plants from the Brazilian Amazon and are used to treat snakebites. In this study, leaves from Aegiphila integrifolia (Jacq) Moldenke were collected from Roraima state, Brazil and its ethanolic extract was evaluated through in vitro and in vivo experiments to verify their antiophidic activity against Bothrops atrox crude venom. The isolated compounds from A. integrifolia were analyzed and the chemical structures were elucidated on the basis of infrared, ultraviolet, mass, 1H and 1³C NMR spectrometry data. Among the described compounds, lupeol (7), betulinic acid (1), β -sitosterol (6), stigmasterol (5), mannitol (4), and the flavonoids, pectolinarigenin (2) and hispidulin (3), were identified. The ethanolic extract and flavonoids (2 and 3) partially inhibited the proteolytic, phospholipase A2 and hyaluronidase activities of B. atrox venom, and the skin hemorrhage induced by this venom in mice. Antimicrobial activity against different bacteria was evaluated and the extract partially inhibited bacterial growth. Thus, taken together, A. integrifolia ethanolic extract has promising use as an antiophidic and antimicrobial. [Display omitted] • Plant extracts can be used as snake antivenoms, neutralizing some of the toxins. • Leaves from the Amazon plant Aegiphila integrifolia was studied. • The flavonoids pectolinarigenin and hispidulin were isolated in huge amounts. • Ethanolic extract and flavonoids were active in B. atrox venom in vitro assays. • The extract showed prevention of muscle damage by the poison and toxins of B. atrox. [ABSTRACT FROM AUTHOR]

Huntington's disease (HD) is a neurodegenerative disorder characterized by unwanted choreatic movements, behavioral and psychiatric disturbances, and dementia. The activation of the hypoxic response pathway through the pharmacological inhibition of hypoxia-inducing factor (HIF) prolyl-hydroxylases (PHDs) is a promising approach for neurodegenerative diseases, including HD. Herein, we have studied the mechanism of action of the compound Betulinic acid hydroxamate (BAH), a hypoximimetic derivative of betulinic acid, and its efficacy against striatal neurodegeneration using complementary approaches. Firstly, we showed the molecular mechanisms through which BAH modifies the activity of the PHD2 prolyl hydroxylase, thus directly affecting HIF-1α stability. BAH treatment reduces PHD2 phosphorylation on Ser-125 residue, responsible for the control of its hydrolase activity. HIF activation by BAH is inhibited by okadaic acid and LB-100 indicating that a protein phosphatase 2A (PP2A) is implicated in the mechanism of action of BAH. Furthermore, in striatal cells bearing a mutated form of the huntingtin protein, BAH stabilized HIF-1α protein, induced Vegf and Bnip3 gene expression and protected against mitochondrial toxin-induced cytotoxicity. Pharmacokinetic analyses showed that BAH has a good brain penetrability and experiments performed in a mouse model of striatal neurodegeneration induced by 3-nitropropionic acid showed that BAH improved the clinical symptoms. In addition, BAH also prevented neuronal loss, decreased reactive astrogliosis and microglial activation, inhibited the upregulation of proinflammatory markers, and improved antioxidant defenses in the brain. Taken together, our results show BAH's ability to activate the PP2A/PHD2/HIF pathway, which may have important implications in the treatment of HD and perhaps other neurodegenerative diseases. [ABSTRACT FROM AUTHOR]

The paper examines the molecular mechanisms of the cytotoxicity of conjugates of betulinic acid with the penetrating cation F16. The in vitro experiments on rat thymocytes revealed that all the obtained F16-betulinic acid derivatives showed more than 10-fold higher cytotoxicity as compared to betulinic acid and F16. In this case, 0.5–1 μM of all conjugates showed mitochondria-targeted action, inducing superoxide overproduction and reducing the mitochondrial potential of cells. Experiments on isolated rat liver mitochondria revealed the ability of conjugates to dose-dependently reduce the membrane potential of organelles, as well as the intensity of respiration and oxidative phosphorylation, which is also accompanied by an increase in the production of hydrogen peroxide by mitochondria. It was shown that these actions of derivatives may be due to several effects: the reversion of ATP synthase, changes in the activity of complexes of the respiratory chain and permeabilization of the inner mitochondrial membrane. All compounds also demonstrated the ability to induce aggregation of isolated rat liver mitochondria. Using the model of lecithin liposomes, we found that the F6 conjugate (2 μM) induces the permeability of vesicle membranes for the fluorescent probe sulforhodamine B. High concentrations (25 μM) of the F6 derivative have been found to induce dynamic processes in the liposome membrane leading to aggregation and/or fusion of vesicle membranes. The paper discusses the relationship between the mitochondria-targeted effects of F16-betulinic acid conjugates and their cytotoxicity. [Display omitted] • Derivatives decrease viability, mitopotential and induce ROS burst in thymocytes. • Derivatives stimulate H 2 O 2 production in isolated rat liver mitochondria. • Derivatives inhibit respiratory chain complexes and show protonophore effect. • Derivatives cause aggregation of mitochondria and lecithin liposomes. • F6 derivative is the most active and induce membrane permeabilization. [ABSTRACT FROM AUTHOR]

Betulinic acid (BA), a natural pentacyclic triterpene, was extracted from the white birch tree, Triphyophyllum peltatum and the jujube tree. In a variety of human cancer cell lines, this substance displays anticancer properties. In this study, we examined how BA works to inhibit human laryngeal cancer growth. We discovered that BA minimally exhibited cytotoxicity in normal cells (human normal cell line GES-1), while remarkably inhibiting viability of AMC-HN-8, TU212, HEp-2 and M4e cells in a concentration-dependent manner. In AMC-HN-8 cancer cells, BA induced apoptosis, activated caspase-3/9/PARP, significantly reduced mitochondrial membrane potential (MMP), increased the expression of cytochrome C in the cytoplasm, transported Bax to the mitochondria, increased the production of reactive oxygen species (ROS), and the ROS scavenger N -acetylcysteine can reduce apoptosis. All data showed that BA triggered apoptosis via the mitochondrial pathway, in which ROS production was likely involved. The findings support the development of BA as a viable drug for the treatment of human laryngeal carcinoma. • Betulinic acid showed strong anti-laryngeal carcinoma activity. • Betulinic acid induced apoptosis via mitochondrial pathway. • Betulinic acid-induction apoptosis can be attenuated by the ROS scavenger N -acetylcysteine. [ABSTRACT FROM AUTHOR]

[Display omitted] • Betulinic acid derivatives show significant inhibitory activity against SARS-CoV-2 3CLpro. • The oxidation of the hydroxyl group at C-3 and esterification of the carboxylate group at C-28 can improve the inhibitory activity. • The inhibition rates of BA02 and BA05 are 78.1% and 82.5%, respectively. During the coronavirus reproduction process, 3-chymotrypsin-like protease (3CLpro) and papain-like protease (PLpro) are accountable for the fragmentation of two polyprotein precursors (pp1a/pp1ab) into substructural proteins. These two proteins are vital for the replication and transcription of the viral genome. Therefore, 3CLpro is a key protein and target for the design of coronavirus inhibitors. In previous studies, we found that betulinic acid has an inhibitory effect on 3CLpro, with 51.5 % inhibition of 3CLpro at 20 µM. Then, series of betulinic acid derivatives were designed, synthesized, and evaluated for their inhibition activities. The results showed that BA02 and BA05 showed significant inhibitory activity on 3CLpro with inhibitory rates of 78.1 % and 82.5 % at 20 µM, respectively. Further evaluation of these two compounds shows that their IC 50 values are 7.22 ± 0.14 μM and 6.40 ± 0.14 μM, respectively. [ABSTRACT FROM AUTHOR]

A green and efficient microwave and gravity-assisted solvent-free distillation followed by extraction (MGSFDE) approach was presented to extract essential oil, oleanoic acid, 3- O -acetyloleanolic acid and betulinic acid from Melaleuca bracteata F. Muell. 'Revolution Gold' leaves. In this process, essential oils were obtained by solvent-free distillation with the synergistic action of microwave and gravity using in situ water in fresh plant cells. Meanwhile, most of the water contained in fresh plants is removed, which simplifies the operation for the subsequent extraction of strongly alcohol-soluble triterpenic acids such as oleanoic acid, 3- O -acetyloleanolic acid and betulinic acid (avoiding the drying process of plant materials), increasing the yield of triterpenic acid. For the extraction process of essential oil, the second-order kinetic model showed a better fit than the first-order kinetic model. Through single-factor, face-centered central composite design and Box Behnken design, the operational parameters affecting MGSFDE were systematically optimized. Under optimized parameters, the yield of M. bracteata leaf essential oil was 6.05 ± 0.17 mL/kg DW, and the yields of oleanoic acid, 3- O -acetyloleanolic acid and betulinic acid were 28.99 ± 1.45, 9.21 ± 0.46 and 0.91 ± 0.05 mg/g DW, respectively. The types of compounds in the essential oil extracted at 540 W were the ones that were most similar to those in the traditional hydrodistillation (HD), and the relative content was higher, according to the results of GC MS, principal component analysis, and heatmap analysis. Because of its greater yields, lower power consumption, lower CO 2 emissions, and lower water wastage, MGSFDE is assumed to be an effective and ecologically friendly extraction process with the potential to replace traditional methods and provides a great reference for extraction studies of other aromatic plants containing volatile and nonvolatile active ingredients. [Display omitted] • Use an efficient microwave and gravity solvent-free distillation followed by extraction. • One-pot obtain essential oil, oleanoic acid, 3- O -acetyloleanolic acid & betulinic acid. • Test factors by single factor, face-centered central composite & Box Behnken design. • Use principal component & heatmap analysis to compare extraction methods. [ABSTRACT FROM AUTHOR]

Eucommia ulmoides Oliv. is a deciduous tree which contains various chemical ingredients. The main objective was to document the active chemical ingredients of Eucommia ulmoides Oliv. and their metabolic profiles in vivo, with a view to providing an experimental and theoretical basis for clarifying the mechanisms underlying the pharmacological activity of Eucommia ulmoides Oliv. against rheumatoid arthritis. Eight main active constituents of Eucommia ulmoides Oliv. bark (pinoresinol glucopyranoside, aucubin, geniposidic acid, geniposide, genipin, chlorogenic acid, quercetin and betulinic acid) were quantified using high-performance liquid chromatography (HPLC). This paper additionally identified and characterized prototype metabolites via ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) combined with the Human Metabolome Database (HMDB) and literature comparisons. Ultra pressure liquid chromatography-mass spectrometry/ mass spectrometry (UPLC-MS/MS) was subsequently employed to quantify these components in blood over time and evaluate their pharmacokinetic characteristics. The anti-rheumatoid arthritis effects of genipin, pinoresinol glucopyranoside and their combinations were assessed using in vitro cellular assays. We identified and characterized a total of 53 ingredients from Eucommia ulmoides Oliv. bark and plasma samples, among which 20 were confirmed as prototype metabolites. Meanwhile, this paper derived and analyzed the metabolic cleavage pathway of 8 index ingredients. Six of these compounds displayed rapid entry into blood, with high plasma exposure and fast elimination rates. Data from the in vitro cellular assay showed that aucubin, pinoresinol glucopyranoside, genipin, and combinations of these compounds effectively inhibit MH7A cell proliferation, reduce NO release, and decrease inflammatory factor levels. [Display omitted] [ABSTRACT FROM AUTHOR]

Investigation for a naturally occurring anti-obesity drug has become the need of society all over the world. Betulinic acid (BA) is a lupane-type pentacyclic triterpene and is sourced from various organisms. This high potential biologically active molecule is reported to have anti-obesity effect. In this study, we report the molecular mechanism of action of BA that underlies anti-obesity activity and also an improved method of its isolation common teak tree. Mouse pre-adipocyte cells were used to develop hyperlipidemic conditions in vitro. Change in expression of genes associated to adipogenesis was checked using quantitative real-time PCR (qPCR). Co-factor specificity of PPAR gamma was analyzed through immune precipitation and immunoblot. Betulinic acid was found to be effective in reducing the lipid content in 3T3L1 cells. Level of PPAR gamma and LXR alpha was reduced in connection to reduced adipogenesis. Change in steroid responsive co-activators (SRCs) during BA treatment proved that the compound can impart profound change in co-factor selectivity, which is crucial in determining the activity profile of PPAR gamma. BA treatment enhanced the SRC-1 interaction with PPAR gamma while reducing the levels of SRC-3. Present study has proved that betulinic acid, a promising candidate in anti-obesity drug development, has potential in regulating the activity of PPAR gamma through co-factor modulation. [ABSTRACT FROM AUTHOR]

Psoriasis is a common inflammatory skin disease. Current treatment for psoriasis relies on conventional immunosuppressive agents. However, long-term treatment with global immunosuppression may cause various side effects. Thus, it is compelling to seek alternative drugs for treating psoriasis with potentially less side effects. Betulinic acid (BA) is a naturally occurring pentacyclic triterpene, an ingredient that originally exists in natural plants and lacks systemic toxicity. BA can regulate immunity with anti-fibrotic, anti-inflammatory and antioxidant properties. However, it's unknown whether BA has a therapeutic effect on psoriasis. The objectives of this study were to investigate whether BA attenuates psoriatic skin inflammation and to identify its mechanisms of action. A murine model of imiquimod-induced psoriasis was utilized to evaluate skin lesion while flow cytometry, immunohistochemistry, quantitative RT-PCR and Western blotting analyses were performed for immunoassays. We found that BA treatment alleviated psoriatic symptoms and inflammatory skin lesion. BA lowered the PASI scores, decreased epidermal thickness and reduced T cell infiltration in the skin lesion. Moreover, BA reduced the frequency of IL-17A-expressing CD4+ and γδ T cells in psoriatic mice, but did not alter CD4+FoxP3+ Treg frequency. BA also reduced IL-17A production but increased anti-inflammatory cytokine IL-10 level in serum of the psoriatic mice. Furthermore, BA inhibited gene expression of pro-inflammatory mediators in skin lesions, including RORγt, IL-17A, IL-6 and TNFα. Importantly, it suppressed NFκB signaling in the skin lesion. Finally, BA inhibited T cell proliferation and IL-17A production by CD4+ T-Cells in vitro. Thus, BA attenuates psoriasis and inhibits Th17 development. • Betulinic acid alleviates psoriatic symptoms and the skin lesion in mice. • It reduces IL-17+CD4+ and IL-17+γδTCR+ T cell frequency in the psoriatic mice. • It also lowers proinflammatory cytokine levels in the skin lesion such as IL-17A. • Treatment with betulinic acid inhibits NFκB signaling in psoriatic mouse skin. [ABSTRACT FROM AUTHOR]

• Amides of cystamine with betulinic acid synthesized for biological investigation. • Antimicrobial activity, cytotoxicity and cell apoptosis were investigated. • Comparison of substitution at the C(3)-OH and C(17)-COOH on cytotoxicity studied. • Cytotoxicity of the target compounds on CEM found with TI = 20, and cell apoptosis documented. • Antimicrobial activity found high on Streptococcus mutans. Amides of betulinic acid with cystamine were synthesized to investigate their antimicrobial and antitumor activity, and their influence on the cell cycle and cell apoptosis. The former target amide (6) displayed cytotoxicity in CEM cell line after 72 h of treatment (IC 50 = 3.0 ± 0.7 μM; TI = 20), and induced apoptosis by caspase-3/7 activation in CEM cells. The latter target amide (9) displayed antimicrobial activity against Streptococcus mutans (MIC 3.125 μM; MBC 3.125 μM) and Bacillus cereus (MIC 25 μM; MBC 25 μM). The achieved results demonstrate enhancing of their biological activity over that of the parent compounds. However, two intermediate compounds (2 and 7) displayed either considerable cytotoxicity (2 ; 7.5 ± 0.8 μM; TI = 10, against G361) or antimicrobial activity (7 ; both against Actinomyces odontolycus and Clostridium perfrigens with MIC 12.5 µM and MBC 12.5 µM). The experimental data were compared with the in silico calculated physico-chemical and ADME parameters of the target compounds, including successful intermediates. [ABSTRACT FROM AUTHOR]

A series of novel fluorine-containing lupane triterpenoid acid derivatives with fluoroaromatic amide moieties at the C-28 position (1 – 8) or with 2-(fluoroacyl)cyclopentane-1,3-dione fragments at the C-3 position (9 – 18) of lupane skeleton was synthesized. A simple synthesis of novel lupane triterpenoid hybrids with 2-(fluoroacyl)-2-cyclopenten-1-one moieties was developed. An interaction of 2-acyl-3-chlorocyclopent-2-en-1-ones, obtained from corresponding cyclic β-triketones, with methyl 3-amino-3-deoxybetulinate gave 3β-isomers (9 – 13) and 3α-isomers (14 – 18) of target hybrids, which were isolated as individual compounds. Anti-inflammatory properties of selected synthesized compounds were studied in vivo using the histamine-, concanavalin A- and sheep erythrocytes immunization-induced mouse paw edema models. The antioxidant activity was investigated in vivo on the model of tetracycline-induced hepatitis. Majority of synthesized fluorine-containing lupane triterpenoid acid derivatives exhibited significant anti-inflammatory and antioxidant effects. Among studied compounds, 3β-hybrid 11 with 2-perfluorobutanoyl-2-cyclopenten-1-one moiety was the most potent bioactive compound. [ABSTRACT FROM AUTHOR]

Betulinic acid (BA), a pentacyclic triterpenoid, exhibits broad spectrum antiproliferative activity, but generally with only modest potency. To improve BA's pharmacological properties, fluorine was introduced as a single atom at C-2, creating two diastereomers, or in a trifluoromethyl group at C-3. We evaluated the impact of these groups on antiproliferative activity against five human tumor cell lines. A racemic 2-F-BA (compound 6) showed significantly improved antiproliferative activity, while each diastereomer exhibited similar effects. We also demonstrated that 2-F-BA is a topoisomerase (Topo) I and IIα dual inhibitor in cell-based and cell-free assays. A hypothetical mode of binding to the Topo I-DNA suggested a difference between the hydrogen bonding of BA and 2-F-BA to DNA, which may account for the difference in bioactivity against Topo I. [ABSTRACT FROM AUTHOR]

• BA induces apoptosis through mitochondria-mediated apoptotic pathway inhuman colorectal cells. • BA inhibited cancer cell metastasis via MMPs signaling pathway. • BA exhibited good anti-tumor effect in a tumor xenograft model of HCT116l cells. Betulinic acid (BA) is a pentacyclic triterpenoids extracted from birch with a wide range of biological properties. Recent studies have shown that BA has significant cytotoxicity to various types of human cancer cells, and shows potential in cancer treatment. However, the efficacy of BA on human colorectal cancer tumor cells is still unclear. The purpose of our study was to evaluate the anti-cancer activity of BA in human colorectal cancer cells in vitro and in vivo to investigate the possible mechanism. In this experiment, we found that BA inhibited colorectal cancer cell lines in vitro with a time-dependent and dose-dependent manner. Moreover, BA could induce cell apoptosis by upregulating expression of Bax and cleaved caspase-3 and downregulating protein of Bcl-2. BA could increase the production of reactive oxygen species and reduce mitochondrial membrane potential of cancer cell, suggesting that BA induced cancer cells apoptosis by mitochondrial mediated pathways. Furthermore, BA significantly inhibited the migration and invasion of colorectal cancer cells, reduced the expression of matrix metalloproteinase (MMPs) and increased the expression of MMPs inhibitor (TIMP-2). In addition, the growth of tumor was significantly suppressed by intraperitoneal administration of 20 mg/kg/day of BA in a xenograft tumor mouse model of HCT-116. Histopathological and immunohistochemical analysis showed that MMP-2+ cells and Ki-67+ cells were reduced and cleaved caspase-3+ cells were increased in tumor tissues of mice after BA administration. The results showed that BA not only promoted the apoptosis of colorectal cancer cells, but also inhibited the metastasis of cancer cells. Our results suggest that BA can be a potential natural drug to inhibit the growth and metastasis of colorectal cancer. [ABSTRACT FROM AUTHOR]

Abstract The present study demonstrated the development of gemcitabine and betulinic acid co-encapsulated PLGA−PEG polymer nanoparticles for enhancing the chemotherapeutic response. This combinatorial PLGA-PEG nanoparticle was formulated using double emulsion and had size <200 nm. The developed nanoparticles were characterized using dynamic light scattering and transmission electron microscopy for their size and shape, respectively. The in vitro release of the drugs from combinatorial nanoparticles was predominantly followed by Fickian diffusion phenomenon. Study on hemocompatibilty approved the administration of this combinatorial nanoparticle for animal study. In vitro cytotoxicity study on Panc1 cells using MTT assay, reactive oxygen species production and cellular apoptotic assay demonstrated that combinatorial nanoparticle was more cytotoxic compared to native drugs solution. Furthermore, the combinatorial nanoparticle suppressed tumor growth more efficiently in Ehrlich (solid) tumor model than the native gemcitabine and betulinic acid at the same concentrations. These findings indicated that PLGA-PEG nanoparticle might be used to co-deliver multiple chemotherapeutic drugs with different properties for enhancing antitumor efficacy. Highlights • Development of gemcitabine and betulinic acid loaded PLGA−PEG nanoparticles. • Combinatorial nanoparticle was more cytotoxic compared to native drugs. • Mechanistic investigation was performed using ROS and apoptotic assay in PANC1 cells. • Combinatorial nanoparticle exhibited improved anti-tumor efficacy in solid tumor model. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • Phytochemical study of D. costata. • Two new furano and pyrano-isoflavones were isolated from Deguelia costata (Fabaceae). • Cytotoxic effect of betulinic acid on rat glioma cells. Abstract Phytochemical investigation of the roots, leaves and stems of Deguelia costata resulted in the isolation of two new prenylated isoflavones, 5,5"-dihydroxy-4'-methoxy-6",6"-dimethyl-4",5"-dihydropyrano [2",3":7,8]isoflavone (1), and 5-hydroxy-4'-methoxy-5"-hydroxyisopropyl-4",5"-dihydrofurano[2",3":7,8]isoflavone (2), together with nine known compounds, 4'-O-methylderrone (3), 4'-O-methylalpinumisoflavone (4); 5-hydroxy-3',4'-dimethoxy-2",2"-dimethylpyrano[5",6":7,6]isoflavone (5), gancaonin M (6), robustin (7), isorobustin (8), scandenin (9), lupeol (10), and betulinic acid (11). The structures of 1 and 2 were determined by analyses of their spectroscopic data [1H, 13C NMR, HSQC and HMBC] and mass spectrometry. The cytotoxicity of 4'-methylderrone (3), scandenin (9), lupeol (10), and betulinic acid (11) were evaluated on C6 glioma cells. Among them, betulinic acid (11) presented a cytotoxic effect on rat C6 glioma cells with an IC 50 value of 38.0 μM. The total flavonoid content of dichloromethane extract from leaves, the most enriched in isoflavones, was obtained using the AlCl 3 method (80.07 ± 1.55 μg Quercetin g−1 dry extract). This new study reveals the potential of D. costata as a source of new isoflavones, as well as the promising cytotoxic effect of betulinic acid on C6 glioma cells. [ABSTRACT FROM AUTHOR]

Background and Aim: Metabolic syndromes are prevalent worldwide and result in various complications including obesity, cardiovascular disease and type II diabetes. Betulinic acid (BA) is a naturally occurring triterpenoid that has anti-inflammatory properties. We hypothesized that treatment with BA may result in decreased body weight gain, adiposity and hepatic steatosis in a diet-induced mouse model of obesity.Methods and Results: Mice fed a high-fat diet and treated with BA showed less weight gain and tissue adiposity without any change in calorie intake. Gene expression profiling of mouse tissues and cell lines revealed that BA treatment increased expression of lipid oxidative genes and decreased that of lipogenesis-related genes. This modulation was mediated by increased AMP-activated protein kinase (AMPK) phosphorylation, which facilitates energy expenditure, lipid oxidation and thermogenic capacity and exerts protective effects against obesity and nonalcoholic fatty liver disease. Overall, BA markedly inhibited the development of obesity and nonalcoholic fatty liver disease in mice fed a high-fat diet, and AMPK activation in various tissues and enhanced thermogenesis are two possible mechanisms underlying the antiobesity and antisteatogenic effects of BA.Conclusions: The current findings suggest that treatment with BA is a potential dietary strategy for preventing obesity and nonalcoholic fatty liver disease. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • Heterologous biosynthesis of betulinic acid in Y. lipolytica was achieved. • Production titer was increased by P450 fusion and MVA pathway gene overexpression. • Substitution of glycerol for glucose enhanced betulinic acid production. • Glycerol improved titer by increasing acetyl-CoA supply and MVA pathway expression. Abstract Glycerol is the main byproduct of the biodiesel production process and can be converted into valuable products by Yarrowia lipolytica. Betulinic acid is a pentacyclic triterpene with a variety of pharmacological properties. Herein we report heterologous biosynthesis of betulinic acid in Y. lipolytica by co-expressing lupeol synthase opAtLUP1, NADPH-cytochrome P450 monooxygenase opCYP716A12, and NADPH-cytochrome P450 reductase opAtCPR1. Initially, the engineered Y. lipolytica produced 0.32 mg/L betulinic acid. This titer was increased to 9.41 mg/L following P450 enzyme fusion and overexpression of key genes from the upstream mevalonate (MVA) pathway. Substitution of glycerol for glucose further enhanced betulinic acid production to 16.98 mg/L, 1.8-fold higher than the titer obtained with glucose. 26.53 mg/L betulinic acid was obtained with 40 g/L glycerol as the sole carbon source, which was comparable to the titer obtained from Saccharomyces cerevisiae with 50 g/L glucose in shake flask cultivation. Glycerol improved betulinic acid titer by increasing the expression of key genes in the MVA pathway, and by increasing the supply of acetyl-CoA. Our study highlights the possible applications of glycerol as carbon source in the production of valuable terpenoids by engineered Y. lipolytica. [ABSTRACT FROM AUTHOR]

Graphical abstract Abstract Emerging evidence has suggested that targeting glycolysis may be a promising strategy for cancer treatment. Betulinic acid (BA) is a natural pentacyclic terpene that has been reported to be active in inhibiting various malignancies. Here, we showed that BA could inhibit aerobic glycolysis activity in breast cancer cell lines MCF-7 and MDA-MB-231 by hampering lactate production, glucose uptake and extracellular acidification rate (ECAR), as well as suppressing aerobic glycolysis-related proteins including c-Myc, lactate dehydrogenase A (LDH-A) and p-PDK1/PDK1 (pyruvate dehydrogenase kinase 1). Mechanistic studies validated Caveolin-1 (Cav-1) as one of key targets of BA in suppressing aerobic glycolysis, as BA administration resulted in Cav-1 upregulation, whereas silencing Cav-1 abrogated the inhibitory effect of BA on aerobic glycolysis. Further investigations demonstrated that BA suppressed aerobic glycolysis in breast cancer cells by regulating the Cav-1/NF-κB/c-Myc pathway. More meaningfully, BA significantly inhibited breast cancer growth and glycolytic activity in both the transgenic MMTV-PyVT+/− breast cancer spontaneous model and the zebrafish breast cancer xenotransplantation model without any detectable side effects in vivo. Taken together, our study sheds novel insights into BA as a promising candidate drug for suppressing aerobic glycolysis, highlighting Cav-1 as a potential molecular target of BA and aerobic glycolysis regulation. [ABSTRACT FROM AUTHOR]

Abstract Agaricus bisporus (Lange) Sing. Chaidam, a special brown mushroom with thick body indigenous in Chaidam basin, was used for fungal chitosan extraction. FTIR, XRD and DSC spectra showed that fungal chitosan was similar to commercial chitosan from aquatic sources. Fungal chitosan and commercial chitosan were used to coat on betulinic acid-loaded liposomes modified with biosurfactants mannosylerythritol lipid A (MEL-A), respectively. After chitosan coating, the mean size, zeta potential and encapsulation efficiency of both liposomes increased. The liposomes coated with fungal chitosan were discovered to have smaller size and higher zeta potential. Furthermore, the wall material MEL-A and coating material chitosan endue liposomes with increased antioxidant capacity. Fungal chitosan coated liposomes also have stronger antioxidant effects than commercial chitosan. The findings implied that the fungal chitosan coated liposomes modified with MEL-A can be considered as a promising delivery system with enhanced antioxidant effects for bioactive compounds. Graphical abstract Unlabelled Image [ABSTRACT FROM AUTHOR]

Abstract The aggressive phenotype displayed by fibroblast-like synoviocytes (FLSs) contributes to cartilage and bone destruction in rheumatoid arthritis (RA). Betulinic acid has been demonstrated to have a positive therapeutic effect on tumor, inflammation and immune disorder, however, the effects of betulinic acid on RA FLSs have not been verified. Therefore, in the present study, we observed the effect of betulinic acid on the migration and invasion of RA FLSs and explored its underlying signal mechanisms. Our results showed that betulinic acid treatment suppressed the migration, invasion and reorganization of the actin cytoskeleton of RA FLSs. In addition, we found that the mRNA expression of IL-1β, IL-6, IL-8 and IL-17A were markedly down-regulated by treatment with betulinic acid in TNF-α-induced RA FLSs. To gain insight into the molecular mechanisms, we evaluated the effect of betulinic acid on NF-κB activation in RA FLSs. The results indicated that betulinic acid treatment reduced the TNF-α-induced activation of NF-κB signal pathway and the NF-κB nuclear accumulation. We also observed that treatment with betulinic acid attenuated synovial inflammation and joint destruction in mice with CIA. Taken together, these results suggest that betulinic acid inhibits the migration and invasion of RA FLSs by blocking NF-κB signal pathway activation. Highlights • Betulinic acid suppressed migration, invasion of rheumatoid arthritis fibroblast-like synoviocytes (RA FLSs). • Betulinic acid treatment reduced TNF-α-induced activation of NF-κB signal pathway and the NF-κB nuclear accumulation. • Betulinic acid ameliorated synovial inflammation and joint destruction in mice with collagen induced arthritis (CIA). • Our data suggested that betulinic acid may be an effective therapeutic agent for RA. [ABSTRACT FROM AUTHOR]