Nanoarchitectonics of an acetogenin-enriched nanosystem mediated by an aqueous extract ofAnnona cherimolaMill with anti-inflammatory and proapoptotic activity against HepG2 cell line.

For the first time, this study shows the nanoarchitectonic process to obtain an acetogenin-enriched nanosystem (AuNPs-Ac) using an aqueous extract fromAnnona cherimolaMill (ACM) composed of gold nanoparticles embedded in an organic matrix that acts as stabilizing agent and presents anti-inflammatory activity and cytotoxical effect against HepG2 cell line, promoting apoptosis. The synthesis of AuNPs-Ac was confirmed by x-ray diffraction analysis, showing metallic gold as the only phase, and the scanning transmission microscope showed an organic cap covering the AuNPs-Ac. Fourier-transformed infrared suggests that the organic cap comprises a combination of different annonaceous acetogenins, alkaloids, and phenols by the presence of bands corresponding to aromatic rings and hydroxyl groups. High-Performance Liquid Chromatography has demonstrated the presence of annonacin, a potent acetogenin, in the extract of ACM. Anin vitroanti-inflammatory activity of the extract of ACM and the AuNPs-Ac was performed using the albumin denaturation method, showing a nonlinear response, which is better than sodium diclofenac salt in a wide range of concentrations that goes from 200 to 400µg ml-1with both samples. The viability assay was studied using trypan blue, treating IMR90 and HepG2 at different concentrations of AuNPs-Ac. The results defined a median lethal dose of 800µg ml-1against HepG2 through apoptosis according to the ratio of caspase-cleaved 9/alpha-tubulin evaluated. It was also demonstrated that the nanosystem presents a higher cytotoxic effect on the HepG2 cell line than in IMR90, suggesting a targeted mechanism. In addition, the nanosystem performs better than using only the extract of ACM in the anti-inflammatory or antiproliferative test, attributed to their higher surface area.

Acetogenins from the Stem of Uvaria rufa and Their Cytotoxic Activity.

Four new adjacent bis-tetrahydrofuran acetogenins, bullacin C (7), uvarirufin (9), and uvariasolins III (12) and IV (13), along with 11 known acetogenins, were isolated from the stem of Uvaria rufa. Their structures were elucidated based on spectroscopic data analysis, including 1D and 2D NMR, HRESIMS, and MALDI-MS/MS of the lithium adducts. Absolute configurations were assigned using Mosher ester analysis and ECD measurements. Uvarirufin (9) possesses a unique C-39 skeleton among acetogenins. Most tested acetogenins exhibited cytotoxicity against human cancer cell lines (HCT 116, 22Rv1, MDA-MB-435, OVCAR3). Squamocin (8) and uvarirufin (9) were found to be the most potent, with an IC50 value of 1.2 µM for both in HCT 116 colon cancer cells. Additionally, a new application of Dragendorff's reagent is proposed herein for the TLC detection of acetogenins.

Annonaceous Acetogenins Synergistically Inhibit Hepatocellular Carcinoma with Sorafenib.

Sorafenib was first approved as the standard treatment for advanced hepatocellular carcinoma (HCC). Despite providing an advantage in terms of patient survival, sorafenib has shown poor clinical efficacy and severe side effects after long-term treatment. Thus, combination treatment is a potential way to increase the effectiveness and reduce the dose-limiting toxicity of sorafenib. Extracts of the seeds of Annona montana have shown synergistic antitumor activity with sorafenib, and seven annonaceous acetogenins, including three new acetogenins, muricin P (2), muricin Q (3), and muricin R (4), were isolated from the extracts by bioguided fractionation and showed synergy with sorafenib. The structures of these compounds were determined using spectroscopic and chemical methods. Annonacin (1) and muricin P (2), which reduced intracellular ATP levels and promoted apoptosis, exhibited synergistic cytotoxicity with sorafenib in vitro. In vivo, annonacin (1) displayed synergistic antitumor activity by promoting tumor cell apoptosis. Moreover, the potential mechanism of annonacin (1) was predicted by transcriptomic analysis, which suggested that SLC33A1 is a potential target in HCC. Annonacin (1) might be a novel candidate for combination therapy with sorafenib against advanced HCC.

In Vitro and In Silico Evaluation of Red Algae Laurencia obtusa Anticancer Activity.

Studies estimate that nearly 2 million new cases of gastric cancer will occur worldwide during the next two decades, which will increase mortality associated with cancer and the demand for new treatments. Marine algae of the Laurencia genus have secondary metabolites known for their cytotoxic action, such as terpenes and acetogenins. The species Laurencia obtusa has demonstrated cytotoxicity against many types of tumors in previous analyses. In this study, we determined the structure of terpenes, acetogenins, and one fatty acid of Laurencia using mass spectrometry (ESI-FT-ICR/MS). In vitro cytotoxicity assays were performed with adenocarcinoma gastric cells (AGS) to select the most cytotoxic fraction of the crude extract of L. obtusa. The Hex:AcOEt fraction was the most cytotoxic, with IC50 9.23 µg/mL. The selectivity index of 15.56 shows that the Hex:AcOEt fraction is selective to cancer cells. Compounds obtained from L. obtusa were tested by the analysis of crystallographic complexes. Molecular docking calculations on the active site of the HIF-2α protein showed the highest affinity for sesquiterpene chermesiterpenoid B, identified from HEX:AcOEt fraction, reaching a score of 65.9. The results indicate that L. obtusa presents potential compounds to be used in the treatment of neoplasms, such as gastric adenocarcinoma.

A new isomaneonene derivative from the red alga Laurencia cf. mariannensis.

Determining the structures of new natural products from marine species not only enriches our understanding of the diverse chemistry of these species, but can also lead to the discovery of compounds with novel and/or important biological activities. Herein, we describe the isolation of isomaneonene C (1), a new halogenated C15 acetogenin, and three known compounds, α-snyderol (2), cis-maneonene D (3), and isomaneonene B (4), from the organic extract obtained from the red alga Laurencia cf. mariannensis collected from Iheya Island, Okinawa, Japan. The structures of these secondary metabolites were elucidated spectroscopically. All compounds were inactive at 30 µg/disc against methicillin-resistant Staphylococcus aureus (MRSA) in combination treatment with a ß-lactam drug, meropenem.

Identification of Annonaceous Acetogenins and Alkaloids from the Leaves, Pulp, and Seeds of Annona atemoya.

Annonaceae is a large family composed of more than 119 genera and more than 2500 species that are distributed in both tropical and subtropical areas. The Annona genus is a member of Annonaceae family, which encompasses about 175 species, most of which are native to Brazil and tropical America. This plant is commonly found on tropical and subtropical continents. Annona atemoya is a commercially important hybrid of A. squamosa and A. cherimola. Phytochemical investigations of A. atemoya leaves, fruit, and seeds have been conducted in limited studies. The purpose of this study was to investigate the constituents of the leaves, fruit pulp, and seeds of A. atemoya because few studies have reported their constituents. Annonaceous acetogenins were identified in the leaves and pulp of A. atemoya for the first time. Twenty compounds were identified: sixteen were acetogenins and four were alkaloids. Additionally, two compounds were isolated, and their structures were confirmed by spectroscopic analysis and compared with the results of previous studies. The concentration of acetogenins in the pulp was very low compared with that in the leaves, whereas the seeds were found to contain the highest concentrations and greatest diversity of compounds.

LCMS/MS Phytochemical Profiling, Molecular, Pathological, and Immune-Histochemical Studies on the Anticancer Properties of Annona muricata.

Annona muricate is a tropical plant that is well-known for its edible fruit of therapeutic interest. LCMS/MS analyses were applied to identify phytoconstituents of the ethanolic extract of the whole fruits and the aqueous extract of the edible fruit part, in addition to the investigation of their anticancer properties against Ehrlich ascites carcinoma (EAC) in male albino mice. LCMS/MS analyses resulted in the identification of 388 components, representing a wide array of classes of compounds, including acetogenins as the major constituents, alkaloids, flavonoids, and phenolics. Among them, four compounds were tentatively characterized as new compounds (1-4), including an acid derivative, protocatechuic-coumaroyl-quinic acid (1), and three flavonoid derivatives, dihydromyricetin galloyl hexoside (2), apigenin gallate (3), and dihydromyricetin hexouronic acid hexoside (4). Induction with EAC cells resulted in abnormalities in the gene expression of pro-apoptotic genes (Bax and caspase-3) and anti-apoptotic gene (Bcl-2) in the tumor mass. Moreover, microscopic, histopathological, and immune-histochemical examinations of the tumor mass and liver tissues exhibited extensive growth of malignant Ehrlich carcinoma cells and marked hydropic degeneration of hepatocytes and infiltration by tumor cells to liver tissue with marked inflammatory reaction. These abnormalities were markedly ameliorated aftertreatment of EAC mice with A. muricata extracts.

Evaluation of Genotoxicity and Toxicity of Annona muricata L. Seeds and In Silico Studies.

Cancer is a multifactorial organic dysfunction for which great efforts are being devoted in searching for new treatments and therapeutic adjuvants. Annona muricata is a fruit that has promising activity against several types of cancer, as it contains acetogenins, the metabolite group associated with this action. Thus, the objective of this study was to evaluate, in experimental models, the toxic behavior of an extract and fraction rich in acetogenins from A. muricata seeds and study the acetogenin, Annonacin, in silico. Phytochemical characterization was made by thin layer chromatography, spectroscopy in the infrared region and nuclear magnetic resonance. Toxicity was evaluated by tests of Allium cepa and Artemia salina, and in silico studies using the SwissDock servers DockThor, PharmMapper, ADMETLab, PreADME, Osiris and ProTox. The extract and fraction showed genotoxic activity against meristematic cells of A. cepa, reducing the mitotic index; however, the extract produced great deleterious effects on the system, even causing cell necrosis. In A. Saline, the extract was more toxic than the fraction, but both samples were considered toxic. Annonacin was effectively linked to complex I, and presented different activities regarding toxicity. Thus, the results of this study are promising, highlighting the anticancer potential of acetogenins.

Pharmacological Activities of Soursop (Annona muricata Lin.).

Soursop (Annona muricata Lin.) is a plant belonging to the Annonaceae family that has been widely used globally as a traditional medicine for many diseases. In this review, we discuss the traditional use, chemical content, and pharmacological activities of A.muricata. From 49 research articles that were obtained from 1981 to 2021, A.muricata's activities were shown to include anticancer (25%), antiulcer (17%), antidiabetic (14%), antiprotozoal (10%), antidiarrhea (8%), antibacterial (8%), antiviral (8%), antihypertensive (6%), and wound healing (4%). Several biological activities and the general mechanisms underlying the effects of A.muricata have been tested both in vitro and in vivo. A.muricata contains chemicals such as acetogenins (annomuricins and annonacin), alkaloids (coreximine and reticuline), flavonoids (quercetin), and vitamins, which are predicted to be responsible for the biological activity of A.muricata.

New C15 Acetogenins from Two Species of Laurencia from the Aegean Sea.

The chemical diversity of the approximately 1,200 natural products isolated from red algae of the genus Laurencia, in combination with the wide range of their biological activities, have placed species of Laurencia in the spotlight of marine chemists' attention for over 60 years. The chemical investigation of the organic (CH2Cl2/MeOH) extracts of Laurencia microcladia and Laurencia obtusa, both collected off the coasts of Tinos island in the Aegean Sea, resulted in the isolation of 32 secondary metabolites, including 23 C15 acetogenins (1-23), 7 sesquiterpenes (24-30) and 2 diterpenes (31 and 32). Among them, six new C15 acetogenins, namely 10-acetyl-sagonenyne (2), cis-sagonenyne (3), trans-thuwalenyne C (4), tinosallene A (11), tinosallene B (12) and obtusallene XI (17), were identified and their structures were elucidated by extensive analysis of their spectroscopic data. Compounds 1-3, 5-11, 13 and 15-32 were evaluated for their antibacterial activity against Staphylococcus aureus and Escherichia coli.

Antileishmanial Effects of Acetylene Acetogenins from Seeds of Porcelia macrocarpa (Warm.) R.E. Fries (Annonaceae) and Semisynthetic Derivatives.

As part of our continuous studies involving the prospection of natural products from Brazilian flora aiming at the discovery of prototypes for the development of new antiparasitic drugs, the present study describes the isolation of two natural acetylene acetogenins, (2S,3R,4R)-3-hydroxy-4-methyl-2-(n-eicos-11'-yn-19'-enyl)butanolide (1) and (2S,3R,4R)-3-hydroxy-4-methyl-2-(n-eicos-11'-ynyl)butanolide (2), from the seeds of Porcelia macrocarpa (Warm.) R.E. Fries (Annonaceae). Using an ex-vivo assay, compound 1 showed an IC50 value of 29.9 µM against the intracellular amastigote forms of Leishmania (L.) infantum, whereas compound 2 was inactive. These results suggested that the terminal double bond plays an important role in the activity. This effect was also observed for the semisynthetic acetylated (1a and 2a) and eliminated (1b and 2b) derivatives, since only compounds containing a double bond at C-19 displayed activity, resulting in IC50 values of 43.3 µM (1a) and 23.1 µM (1b). In order to evaluate the effect of the triple bond in the antileishmanial potential, the mixture of compounds 1 + 2 was subjected to catalytic hydrogenation to afford a compound 3 containing a saturated side chain. The antiparasitic assays performed with compound 3, acetylated (3a), and eliminated (3b) derivatives confirmed the lack of activity. Furthermore, an in-silico study using the SwissADME online platform was performed to bioactive compounds 1, 1a, and 1b in order to investigate their physicochemical parameters, pharmacokinetics, and drug-likeness. Despite the reduced effect against amastigote forms of the parasite to the purified compounds, different mixtures of compounds 1 + 2, 1a + 2a, and 1b + 2b were prepared and exhibited IC50 values ranging from 7.9 to 38.4 µM, with no toxicity for NCTC mammalian cells (CC50 > 200 µM). Selectivity indexes to these mixtures ranged from >5.2 to >25.3. The obtained results indicate that seeds of Porcelia macrocarpa are a promising source of interesting prototypes for further modifications aiming at the discovery of new antileishmanial drugs.

Structure-Activity Relationships of Thiophene Carboxamide Annonaceous Acetogenin Analogs: Shortening the Alkyl Chain in the Tail Part Significantly Affects Their Growth Inhibitory Activity against Human Cancer Cell Lines.

In a previous study, we found that the thiophene carboxamide solamin analog, which is a mono-tetrahydrofuran annonaceous acetogenin, showed potent antitumor activity through the inhibition of mitochondrial complex I. In this study, we synthesized analogs with short alkyl chains instead of the n-dodecyl group in the tail part. We evaluated their growth inhibitory activities against human cancer cell lines. We found that the alkyl chain in the tail part plays an essential role in their activity.

An Overview of the Chemical Characteristics, Bioactivity and Achievements Regarding the Therapeutic Usage of Acetogenins from Annona cherimola Mill.

Annona cherimola Mill., or the custard apple, is one of the species belonging to the Annonaceae family, is widely used in traditional medicine, and has been reported to be a valuable source of bioactive compounds. A unique class of secondary metabolites derived from this family are Annonaceous acetogenins, lipophilic polyketides considered to be amongst the most potent antitumor compounds. This review provides an overview of the chemical diversity, isolation procedures, bioactivity, modes of application and synthetic derivatives of acetogenins from A. cherimola Mill.

Antiproliferative Activity of Mycalin A and Its Analogues on Human Skin Melanoma and Human Cervical Cancer Cells.

Mycalin A, a polybrominated C15 acetogenin isolated from the encrusting sponge Mycale rotalis, displays an antiproliferative activity on human melanoma (A375) and cervical adenocarcinoma (HeLa) cells and induces cell death by an apoptotic mechanism. Various analogues and degraded derivatives of the natural substance have been prepared. A modification of the left-hand part of the molecule generates the most active substances. A structurally simplified lactone derivative of mycalin A, lacking the C1-C3 side chain, is the most active among the synthesized compounds exhibiting a strong cytotoxicity on both A375 and HeLa cells but not but not on human dermal fibroblast (HDF) used as healthy cells. Further evidence on a recently discovered chlorochromateperiodate-catalyzed process, used to oxidise mycalin A, have been collected.

Effects of Acetogenins from Annona coriacea on the in Vitro Reactions of Photosynthesis.

Our search for candidates for photosynthesis inhibitors is allowing us to report the effect of two acetogenins identified in Annona coriacea Mart. leaves, ACG-A and ACG-B, a non-adjacent bis-THF and a mono-THF types, respectively. This is an important class of natural products which presents biological properties such as anticancer, neurotoxic, larvicidal and insecticidal. However, this is only the second report associated to its herbicidal activity. Their mechanisms of action on the light reactions of the photosynthesis were elucidated by polarographic techniques. Compounds inhibited the noncyclic electron transport on basal, phosphorylating, and uncoupled conditions from H2 O to methyl viologen (MV); therefore, they act as Hill reaction inhibitors. Studies on fluorescence of chlorophyll a (ChL a) indicated that they inhibited the acceptor side of PSII between P680 and PQ-pool, exactly as the commercial herbicide DCMU does.

Bio-Guided Isolation of Acetogenins from Annona cherimola Deciduous Leaves: Production of Nanocarriers to Boost the Bioavailability Properties.

Annonaceous acetogenins (ACGs) are lipophilic polyketides isolated exclusively from Annonaceae. They are considered to be amongst the most potent antitumor compounds. Nevertheless, their applications are limited by their poor solubility. The isolation of ACGs from Annona cherimola leaves, an agricultural waste, has not been reported to date. Molvizarin (1) cherimolin-1 (2), motrilin (3), annonacin (4) and annonisin (5) are isolated for the first time from A. cherimola deciduous leaves. Annonacin was found to be four- and two-times more potent in tumoral cells (HeLa, 23.6% live cells; IGROV-1, 40.8% live cells for 24 h) than in HEK-293 at 50 µM (24 h, 87.2% live cells). Supramolecular polymer micelles (SMPMs) were synthesized to encapsulate the major ACG isolated, annonacin, in order to improve its solubility in aqueous media. The bioavailability of this compound was increased by a factor of 13 in a simulated human digestive system when compared with free annonacin and an encapsulation efficiency of 35% was achieved. In addition, the cytotoxic activity of SMPMs that hosted annonacin (100 µM, 24 h, 5.8% live cells) was increased compared with free annonacin in water (100 µM, 24 h, 92% live cells). These results highlight the use of by-products of A. cherimola, and their pure compounds, as a promising source of anticancer agents. The use of SMPMs as nanocarriers of ACGs could be an alternative for their application in food field as nutraceutical to enhance the administration and efficacy.

Ultrasound-Assisted Extraction of Total Acetogenins from the Soursop Fruit by Response Surface Methodology.

The soursop fruit or Annona muricata (A. muricata) fruit is recognized by its bioactive compounds and acetogenins (ACG) are among the most important. The effect of ACGs, with greater importance in health, is that they present anti-tumor activity; however, the methods of extraction of ACGs are very slow and with a high expenditure of solvents. To our knowledge, there is no report of an optimal method for the extraction of acetogenins from the Annonaceae family by ultrasound-assisted extraction (UAE); therefore, the aim was to find the best UEA conditions of acetogenins from A. muricata fruit (peel, pulp, seed, and columella) by using response surface methodology. The effect of amplitude (40%, 70%, and 100%), time (5, 10, and 15 min), and pulse-cycle (0.4, 0.7, and 1 s) of ultrasound at 24 kHz was evaluated on the total acetogenin content (TAC). Optimal extraction conditions of acetogenins (ACGs) with UEA were compared with the extraction of ACGs by maceration. The optimal UEA conditions in the A. muricata pulp and by-products were dependent on each raw material. The highest TAC was found in the seed (13.01 mg/g dry weight (DW)), followed by the peel (1.69 mg/g DW), the pulp (1.67 mg/g DW), and columella (1.52 mg/g DW). The experimental TAC correlated well with the model (Adjusted R2 with values between 0.88 and 0.97). The highest effectiveness in ACG extraction was obtained in seeds and peels using UEA compared to extraction by maceration (993% and 650%, respectively). The results showed that A. muricata by-products are an important source of ACGs and that UAE could be a viable alternative, with high potential for large-scale extraction.

Synthesis, Characterization, and Reactivity of Complex Tricyclic Oxonium Ions, Proposed Intermediates in Natural Product Biosynthesis.

Reactive intermediates frequently play significant roles in the biosynthesis of numerous classes of natural products although the direct observation of these biosynthetically relevant species is rare. We present here direct evidence for the existence of complex, thermally unstable, tricyclic oxonium ions that have been postulated as key reactive intermediates in the biosynthesis of numerous halogenated natural products from Laurencia species. Evidence for their existence comes from full characterization of these oxonium ions by low-temperature NMR spectroscopy supported by density functional theory (DFT) calculations, coupled with the direct generation of 10 natural products on exposure of the oxonium ions to various nucleophiles.

Antitrypanosomal Activity of Acetogenins Isolated from the Seeds of Porcelia macrocarpa Is Associated with Alterations in Both Plasma Membrane Electric Potential and Mitochondrial Membrane Potential.

As part of a drug discovery program aimed at the identification of anti- Trypanosoma cruzi metabolites from Brazilian flora, four acetogenins (1-4) were isolated from the seeds of Porcelia macrocarpa and were identified by NMR spectroscopy and HRESIMS. The new compounds 1 and 2 displayed activity against the trypomastigote (IC50 = 0.4 and 3.6 µM) and amastigote (IC50 = 23.0 and 27.7 µM) forms. The structurally related known compound 3 showed less potency to the amastigotes, with an IC50 value of 58 µM, while the known compound 4 was inactive. To evaluate the potential mechanisms for parasite death, parameters were evaluated by fluorometric assays: (i) plasma membrane permeability, (ii) plasma membrane electric potential (ΔΨp), (iii) reactive oxygen species production, and (iv) mitochondrial membrane potential (ΔΨm). The results obtained indicated that compounds 1 and 2 depolarize plasma membranes, affecting ΔΨp and ΔΨm and contributing to the observed cellular damage and disturbing the bioenergetic system. In silico studies of pharmacokinetics and toxicity (ADMET) properties predicted that all compounds were nonmutagenic, noncarcinogenic, nongenotoxic, and weak hERG blockers. Additionally, none of the isolated acetogenins 1-4 were predicted as pan-assay interference compounds.

Annonaceous acetogenin mimic AA005 suppresses human colon cancer cell growth in vivo through downregulation of Mcl-1.

Annonaceous acetogenins are a well-established family of natural products with significant bioactivities, especially high cytotoxic and antitumor activities. AA005 is an annonaceous acetogenin mimic that has shown significant cytotoxicity against a variety of cancer cell lines, but its in vivo antitumor effects have not been demonstrated so far, and its anticancer mechanisms remain ambiguous. In this study, we investigated the effects of AA005 on human colon cancer cell lines in vivo. Human colon carcinoma cell line SW620 xenograft nude mice were treated with AA005 (5 mg/kg/day, i.p.) for 21 days. AA005 administration markedly inhibited the tumor growth via promoting nuclear translocation of apoptosis-inducing factor (AIF) and inducing AIF-dependent cell death. Subsequent studies in human colon carcinoma cell lines SW620 and RKO in vitro revealed that after the colon cancer cells exposed to AA005, downregulation of a B-cell lymphoma 2 family protein, myeloid cell leukemia-1 (Mcl-1), was an early event due to the inhibition of Mcl-1 mRNA level and protein synthesis in a time-dependent manner. Intriguingly, knockdown of Mcl-1 using small interfering RNA markedly accelerated the nuclear translocation of AIF and upregulation of receptor interacting protein-1, and enhanced AA005-mediated lethality, whereas ectopic expression of Mcl-1 substantially attenuated AA005-mediated lethality in the colon cancer cells. Finally, silencing Mcl-1 expression markedly enhanced AA005-induced lethality in SW620 xenograft nude mice, demonstrating a pivotal role of Mcl-1 downregulation in mediating the in vivo antitumor effects of AA005. Taken together, this study demonstrates for the first time the anticancer effects of AA005 against human colon cancer cell lines in vivo, which is mediated through the downregulation of Mcl-1.