Special Issue-"Isolation, Structure Elucidation and Biological Activity of Natural Products".

This Special Issue of Molecules gathers fourteen research studies and three review papers covering developments in the scope of the isolation, structure elucidation and biological activity of natural products [...].

Synthesis and Antimicrobial Activity of Ursolic Acid Ester Derivatives.

Infections caused by microorganisms are a major cause of morbidity and mortality worldwide, and natural products continue to be important sources for the discovery of new antimicrobial agents. Ursolic acid is a triterpene with known antibacterial action, being naturally found in plants, such as Jaracanda oxyphylla and Jacaranda caroba (Bignoniaceae). Ursolic acid derivative esters have revealed potential biological activities, such as antitumor, antiviral, and antibacterial activity. In this study, sixteen esters (1-16) were synthesized from ursolic acid using DIC/DMAP and characterized by infrared (IR), nuclear magnetic resonance (1 H- and 13 C-NMR) and mass spectrometry. All ursolic acid esters were evaluated against Bacillus cereus, Staphylococcus aureus, Escherichia coli, Salmonella typhimurium, and the yeast Candida albicans. Six compounds are herein described for the first time (3, 9, 11, 13, 14 and 16) with yields up to 91.6 %. Compounds 11 (3ß-(3,4-dimethoxybenzoyl)ursolic acid) and 15 (3ß-nicotinoylursolic acid) displayed promising antifungal activity, with inhibition of C. albicans growth of 93.1 and 95.9 %, respectively.

Plant Spices as a Source of Antimicrobial Synergic Molecules to Treat Bacterial and Viral Co-Infections.

The COVID-19 pandemic exposed the lack of antiviral agents available for human use, while the complexity of the physiological changes caused by coronavirus (SARS-CoV-2) imposed the prescription of multidrug pharmacotherapy to treat infected patients. In a significant number of cases, it was necessary to add antibiotics to the prescription to decrease the risk of co-infections, preventing the worsening of the patient's condition. However, the precautionary use of antibiotics corroborated to increase bacterial resistance. Since the development of vaccines for COVID-19, the pandemic scenario has changed, but the development of new antiviral drugs is still a major challenge. Research for new drugs with synergistic activity against virus and resistant bacteria can produce drug leads to be used in the treatment of mild cases of COVID-19 and to fight other viruses and new viral diseases. Following the repurposing approach, plant spices have been searched for antiviral lead compounds, since the toxic effects of plants that are traditionally consumed are already known, speeding up the drug discovery process. The need for effective drugs in the context of viral diseases is discussed in this review, with special focus on plant-based spices with antiviral and antibiotic activity. The activity of plants against resistant bacteria, the diversity of the components present in plant extracts and the synergistic interaction of these metabolites and industrialized antibiotics are discussed, with the aim of contributing to the development of antiviral and antibiotic drugs. A literature search was performed in electronic databases such as Science Direct; SciELO (Scientific Electronic Library Online); LILACS (Latin American and Caribbean Literature on Health Sciences); Elsevier, SpringerLink; and Google Scholar, using the descriptors: antiviral plants, antibacterial plants, coronavirus treatment, morbidities and COVID-19, bacterial resistance, resistant antibiotics, hospital-acquired infections, spices of plant origin, coronaviruses and foods, spices with antiviral effect, drug prescriptions and COVID-19, and plant synergism. Articles published in English in the period from 2020 to 2022 and relevant to the topic were used as the main inclusion criteria.

Phytochemicals of Avocado Residues as Potential Acetylcholinesterase Inhibitors, Antioxidants, and Neuroprotective Agents.

Avocado (Persea americana) is a widely consumed fruit and a rich source of nutrients and phytochemicals. Its industrial processing generates peels and seeds which represent 30% of the fruit. Environmental issues related to these wastes are rapidly increasing and likely to double, according to expected avocado production. Therefore, this work aimed to evaluate the potential of hexane and ethanolic peel (PEL-H, PEL-ET) and seed (SED-H, SED-ET) extracts from avocado as sources of neuroprotective compounds. Minerals, total phenol (TPC), total flavonoid (TF), and lipid contents were determined by absorption spectroscopy and gas chromatography. In addition, phytochemicals were putatively identified by paper spray mass spectrometry (PSMS). The extracts were good sources of Ca, Mg, Fe, Zn, ω-6 linoleic acid, and flavonoids. Moreover, fifty-five metabolites were detected in the extracts, consisting mainly of phenolic acids, flavonoids, and alkaloids. The in vitro antioxidant capacity (FRAP and DPPH), acetylcholinesterase inhibition, and in vivo neuroprotective capacity were evaluated. PEL-ET was the best acetylcholinesterase inhibitor, with no significant difference (p > 0.05) compared to the control eserine, and it showed neither preventive nor regenerative effect in the neuroprotection assay. SED-ET demonstrated a significant protective effect compared to the control, suggesting neuroprotection against rotenone-induced neurological damage.

Antiviral fungal metabolites and some insights into their contribution to the current COVID-19 pandemic.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) outbreak, which started in late 2019, drove the scientific community to conduct innovative research to contain the spread of the pandemic and to care for those already affected. Since then, the search for new drugs that are effective against the virus has been strengthened. Featuring a relatively low cost of production under well-defined methods of cultivation, fungi have been providing a diversity of antiviral metabolites with unprecedented chemical structures. In this review, we present viral RNA infections highlighting SARS-CoV-2 morphogenesis and the infectious cycle, the targets of known antiviral drugs, and current developments in this area such as drug repurposing. We also explored the metabolic adaptability of fungi during fermentation to produce metabolites active against RNA viruses, along with their chemical structures, and mechanisms of action. Finally, the state of the art of research on SARS-CoV-2 inhibitors of fungal origin is reported, highlighting the metabolites selected by docking studies.

New AChE inhibitors from microbial transformation of trachyloban-19-oic acid by Syncephalastrum racemosum.

Trachyloban-19-oic acid (1) is a diterpene very abundant in nature and its structural modification can furnish new bioactive compounds. Biotransformation of 1 by fungus Syncephalastrum racemosum provided three derivatives, two hydroxylated products (2-3) and one product of rearrangement (4). Products 3 and 4 have never been reported so far, to the best of our knowledge. Structure of 3 was formed after oxidation and rearrangement of compound 2. Compounds 1-4 were evaluated for inhibition of acetylcholinesterase, enzyme linked to the symptomatic control of Alzheimer's disease. All the compounds presented inhibitory activity higher than starting material 1, and product 3 presented IC50 = 0.06 µM, which is about six times higher than activity found for galanthamine (IC50 = 0.38 µM), the positive control used in this assay.

Phytotoxic activity and metabolism of Botrytis cinerea and structure-activity relationships of isocaryolane derivatives.

Research has been conducted on the biotransformation of (8S,9R)-isocaryolan-9-ol (4a) and (1S,2S,5R,8S)-8-methylene-1,4,4-trimethyltricyclo[6.2.1.0(2,5)]undecan-12-ol (5a) by the fungal phytopathogen Botrytis cinerea. The biotransformation of compound 4a yielded compounds 6-9, while the biotransformation of compound 5a yielded compounds 10-13. The activity of compounds 4a and 5a against B. cinerea has been evaluated. (8R,9R)-Isocaryolane-8,9-diol (6), a major metabolite of compound 4a, shows activity compared to its parent compound 4a, which is inactive. The effect of isocaryolanes 3, 4a, and 5a, together with their biotransformation products 6-8, 10, and 14-17, on the germination and radicle and shoot growth of Lactuca sativa (lettuce) has also been determined. Compounds 7-13 are described for the first time.

Atomization of Cocoa Honey Using Whey Protein Isolate to Produce a Dry Formulation with Improved Shelf Life for Industrial Application.

Cocoa honey, a by-product obtained during the processing of cocoa, is a juice rich in pectin, organic acids, minerals and phenolic compounds with antioxidant properties. Fresh cocoa honey is quickly fermented due to its high content of reducing sugars, such as fructose and glucose, which limits its shelf life. Currently, cocoa honey is only commercialized in frozen form, as logistical challenges prevent the wide distribution or export of this by-product for applications in the market of sweets, jellies, beverages, confectionery, and nutraceutical foods among others. Spray-drying technology is a viable prospect for the large-scale stabilization of products such as cocoa honey, with less heat exposure compared to other conventional drying methods. This work aimed to evaluate the efficacy of drying adjuvants for a rapid removal of the water present in cocoa honey via atomization, since this process minimizes the effects of glass transition temperature (Tg) related to materials with high sugar contents. Physical parameters such as the moisture content, hygroscopicity, particle size, and yield of the products obtained were determined. Cocoa honey presented 85.3 ± 0.20 g/100 g of moisture. The formulations successfully decreased moisture content, which was lower than 11.72 ± 0.08 g/100 g in the formulations. Water activity ranged between 0.1464 ± 0.0043 and 0.1562 ± 0.029, with no significant difference between the formulations. The hygroscopicity of cocoa honey powders ranged from 29.29 to 29.87 g of water/100 g of cocoa honey. The combination of 20% maltodextrin and 1% whey protein isolate (WPI) led to the best yield, resulting in a free-flowing powder as the final product. On the other hand, the formulation composed of maltodextrin and whey protein isolate in the ratio of 29:1, respectively, led to the most stable product, with less loss of phenolic compounds during the drying process (6.04%). Regarding particle diameter, 90% of the accumulated distribution did not exceed 57 µm. The greatest dispersion of particles occurs in the Ma20W10 formulation with a span of 2.72, inferring greater variation in size between small (7.01 ± 0.06 µm), medium (18.25 ± 0.37 µm), and large (56.65 ± 1.17 µm) particles. The use of whey protein isolate as an adjuvant proved to be an efficient drying process in the production of cocoa honey powder, and was also advantageous for enriching the nutritional content of the final product due to its protein origin. Furthermore, the combination of spray-drying technology and the use of whey protein isolate as adjuvant led to a free-flowing cocoa honey powder with an adequate particle size and benefits in terms of shelf-life extension, providing new opportunities for the commercialization of cocoa honey as an ingredient for the food industry, with benefits for the circular economy.

Biotransformation of clovane derivatives. Whole cell fungi mediated domino synthesis of rumphellclovane A.

Here we describe the biotransformation of clovane derivatives by filamentary fungi Pestalotiopsis palustris and Penicillium minioluteum, and the application of the latter to the synthesis and determination of the absolute configuration of rumphellclovane A (2). Methoxyclovanol (1), a growth inhibitor of the phytopathogen Botrytis cinerea, is metabolised by P. palustris to yield rumphellclovane A (2), a natural compound recently isolated from the gorgonian coral Rumphella antipathies, two new compounds, (1R,2S,5S,8R,9S,10R)-2-methoxyclovane-9,10-diol (5) and (1S,2S,5S,7R,8R,9R)-2-methoxyclovane-7,9-diol (6), hydroxylated in positions not easily accessed by classic synthetic chemistry, and clovanodiols 3 and 4. P. minioluteum is able to selectively transform methoxyclovanol (1) into clovanodiols 3 and 4 and, in turn, lactone 8, the putative intermediate in the above mentioned synthesis of rumphellclovane A (2), into compound 2 via a domino process. The ability of P. minioluteum to carry out the cleavage of ethers on clovane derivatives is also evaluated.

Hibiscus, Rooibos, and Yerba Mate for Healthy Aging: A Review on the Attenuation of In Vitro and In Vivo Markers Related to Oxidative Stress, Glycoxidation, and Neurodegeneration.

The world is currently undergoing a demographic change towards an increasing number of elderly citizens. Aging is characterized by a temporal decline in physiological capacity, and oxidative stress is a hallmark of aging and age-related disorders. Such an oxidative state is linked to a decrease in the effective mechanisms of cellular repair, the incidence of post-translational protein glycation, mitochondrial dysfunction, and neurodegeneration, just to name some of the markers contributing to the establishment of age-related reduction-oxidation, or redox, imbalance. Currently, there are no prescribed therapies to control oxidative stress; however, there are strategies to elevate antioxidant defenses and overcome related health challenges based on the adoption of nutritional therapies. It is well known that herbal teas such, as hibiscus, rooibos, and yerba mate, are important sources of antioxidants, able to prevent some oxidation-related stresses. These plants produce several bioactive metabolites, have a pleasant taste, and a long-lasting history as safe foods. This paper reviews the literature on hibiscus, rooibos, and yerba mate teas in the context of nutritional strategies for the attenuation of oxidative stress-related glycoxidation and neurodegeneration, and, here, Alzheimer's Disease is approached as an example. The focus is given to mechanisms of glycation inhibition, as well as neuroprotective in vitro effects, and, in animal studies, to frame interest in these plants as nutraceutical agents related to current health concerns.

Biotic stress caused by in vitro co-inoculation enhances the expression of acetylcholinesterase inhibitors by fungi.

Fungi are an inexhaustible source of bioactive metabolites that can be driven to find medicines for chronic diseases, as Alzheimer's disease. In the present work, five species of soil-originated fungi (Aspergillus chevalieri, Clonostachys rogersoniana, Fusarium nygamai, Penicillium sp., and Talaromyces calidicanius) were submitted to mutual biotic stress aiming at activating the expression of metabolites capable of inhibiting the enzyme acetylcholinesterase. HPLC profiles showed that the in vitro biotic stress triggered the biosynthesis of metabolite-mediated defense responses. Five compounds present in the complex co-culture matrix were identified by Paper Spray Mass Spectrometry (PS-MS). The approach enhanced the biosynthesis of acetylcholinesterase inhibitors (up to 99.6% inhibition) in comparison with the individual cultures. The mutual biotic stress between T. calidicanius and A. Chevalieri led to the biosynthesis of a pool of metabolites statistically as efficient as serine (p < 0.05), the positive control used in the experiments (99.6 and 99.1% inhibition, respectively).

Biotransformation of bioactive isocaryolanes by Botrytis cinerea.

The metabolism of the fungistatic agent (8R,9R)-8-methoxyisocaryolan-9-ol (4) by the fungus Botrytis cinerea has been investigated. Biotransformation of compound 4 yielded compounds 5 and 6-9. No dihydrobotrydial is observed after 4 days of incubation of compound 4. Separate biotransformation of (8R,9R)-isocaryolane-8,9-diol (5) yielded compounds 7-11. The evaluation of the fungistatic activity against B. cinerea of compounds 4, 5, and 6 is reported. (4R,8R,9R)-8-Methoxyisocaryolane-9,15-diol (6), a major metabolite of (8R,9R)-8-methoxyisocaryolan-9-ol (4), shows a much reduced biological activity when compared with the parent compound. Isocaryolane derivatives 6-11 are described for the first time.

Antioxidant role of morusin and mulberrofuran B in ethanol extract of Morus alba roots.

The white mulberry, Morus alba, is distributed worldwide and known for its antioxidant properties. Ethanol extract of six varieties of M. alba roots were studied (IZ 13/6, IZ 40, IZ 56/4, IZ 64, Indonesia and Tigreada). From variety IZ 40, two secondary metabolites were isolated and unambiguously identified by 2 D NMR experiments as morusin and mulberrofuran B. These were quantified in the varieties by liquid chromatography, recording 5.27-16.74% for the first, and 0.54-3.55% for the latter. Mulberrofuran B presented higher activity than morusin by phosphomolybdenum (1531.33 ± 20.28 mmol ascorbic acid/g) and ferrocyanide (14.39%) methods, ABTS (95.74 ± 4.21 µM) and DPPH (843.87 ± 10.65 µM) radical sequestration capacity, while morusin showed significant statistical correlation with antioxidant activity for some M. alba varieties. These results suggested that morusin influences antioxidant activity of M. alba in synergism with other compounds and can be a good chemical marker in this species.

Synthesis, Fungitoxic Activity against Botrytis cinerea and Phytotoxicity of Alkoxyclovanols and Alkoxyisocaryolanols.

Clovane and isocaryolane derivatives have been proven to show several levels of activity against the phytopathogenic fungus Botrytis cinerea. Both classes of sesquiterpenes are reminiscent of biosynthetic intermediates of botrydial, a virulence factor of B. cinerea. Further development of both classes of antifungal agent requires exploration of the structure-activity relationships for the antifungal effects on B. cinerea and phytotoxic effects on a model crop. In this paper, we report on the preparation of a series of alkoxy-clovane and -isocaryolane derivatives, some of them described here for the first time (2b, 2d, 2f-2h, and 4c-4e); the evaluation of their antifungal properties against B. cinerea, and their phytotoxic activites on the germination of seeds and the growth of radicles and shoots of Lactuca sativa (lettuce). Both classes of compound show a correlation of antifungal activity with the nature of side chains, with the best activity against B. cinerea for 2d, 2h, 4c and 4d. In general terms, while 2-alkoxyclovan-9-ols (2a-2e) exert a general phytotoxic effect, this is not the case for 2-arylalkoxyclovan-9-ols (2f-2i) and 8-alkoxyisocaryolan-9-ols (4a-4d), where stimulating effects would make them suitable candidates for application to plants.

Hydroxylation at carbon-2 of ent-16-oxo-17-norkauran-19-oic acid by Fusarium proliferatum.

A new product of biotransformation of ent-16-oxo-17-norkauran-19-oic acid (1) by Fusarium proliferatum was isolated and identified as a 2beta-hydroxy derivative (2). The structure of 2 was elucidated on the basis of spectroscopic data interpretation and single-crystal X-ray diffraction analysis. The allelopathic activity of compound 2 was evaluated on the growth of radicals and shoots of Lactuca sativa (lettuce). This is the first time that fungal hydroxylation at position C-2 has been reported on an ent-kaurane diterpene skeleton.

Edible flowers: Bioactive profile and its potential to be used in food development.

The consumption of flowers as food is reported in various cultures around the world as part of traditional cuisine or alternative medicine, in addition to their wide use as ornaments. However, many species of edible flowers can be considered more than a delicacy or a garnish due to their nutritional value as source of protein and essential aminoacids. In this scenario, flowers represent an important segment to expand food market, due to their suitable sensory and nutritional characteristics, as well as presence of bioactive compounds beneficial to human health. Research on consumer behavior and purchase intent have been unraveling ways to explore the different attributes of edible flowers, in order to conquer this promising market in the following decades. Socio-cultural factors involved in the consumption of edible flowers have been subject of several studies aiming at popularizing and expanding this growing industry and encouraging local use of traditional flowers is important to preserve endangered traditions. On the other hand, nutritional properties, pharmacological benefits, chemical composition and the forms of preparation of edible species have been increasingly studied with the growing search for natural and health foods. Modern and effective methods for extraction of bioactive compounds from flowers are also contributing to explore their components, allowing the development of functional ingredients for food industry. Some key information as proper taxonomy and toxicological profile are still necessary to stimulate the consumption of edible flowers, as well as the creation of a good practice manual for proper management (cultivation, handling and preparation) of flowers for commercialization. Scientific and technical information on nutritional, therapeutic and chemical features of edible flowers are reviewed and discussed, aiming at strengthening the knowledge, and, consequently, consuming habits and research on their benefits in human diet.

Selective activity of Mucor plumbeus reductase towards (-)-camphorquinone.

The biotransformation of 1R-(-)-camphorquinone, achieved by growing cells of four fungi species isolated from soil (Mucor plumbeus, Lecanicillium muscarium, Thamnostylum sp. and Syncephalastrum racemosum), was investigated in optimized culture media for each species. Fungi were grown aerobically under shaking and their activities with respect to camphorquinone were monitored for 20 days by gas chromatography coupled to mass spectrometry (GCMS). Camphorquinone was found to be stable in control flasks throughout the experiment. The most interesting results were found for M. plumbeus, which was only able to perform monoreduction of camphorquinone when cultivated on a glucose-peptone-yeast extract medium. Large-scale experiments were set up and the camphorquinone biotransformation products formed by M. plumbeus were purified by column chromatography and identified by (1)H and (13)C nuclear magnetic resonance (NMR). Theoretical calculations were employed as a complementary technique to unambiguously identify the biotransformation products. These findings suggest that M. plumbeus could be of great use for the selective reduction of camphorquinone and related compounds.

Edible mushrooms as a ubiquitous source of essential fatty acids.

Many different species of mushrooms are consumed around the world, after harvesting from nature or cultivated under controlled conditions. Fruiting bodies of mushrooms from the Agaricus, Lentinula, and Pleurotus genera are reported as the most cultivated and commercialized ones in many countries, while wild edible mushroom species vary greatly according to the growth location. Mushrooms are nutritionally well-balanced sources of carbohydrates and proteins, with low fat concentrations, usually ranging from 0.1 to 16.3% making them very health foods. Although mushrooms are not a choice source of lipids, they contain essential fatty acids such as linoleic, oleic, and linolenic in their lipid profiles, usually as the major constituents. Therefore, compared to other food of vegetal and animal origin, mushrooms have the advantage of possessing high levels of polyunsaturated fatty acids (PUFA). Percentage of these fatty acids (in 100 g of total fatty acids) in mushrooms varies greatly: linoleic acid ranges from 0.0-81.1%, oleic acid between 1.0 and 60.3%, and linolenic acid from 0.0-28.8%. A global overview of the lipid composition of mushrooms is presented in this review, emphasizing the presence and levels of monounsaturated fatty acids (MUFA) and PUFA. Literature points that, in mushrooms, unsaturated fatty acid levels are generaly greater than those of saturated ones, regardless of the continent where the mushroom is cultivated or harvested. Comparing individually different species of the same mushroom (Agaricus bisporus, Pleurotus ostreatus, and Boletus edulis) from different continents, similar fatty acid profile is also frequently observed. The great variety of edible mushrooms found worldwide and their consistent production of fatty acids, regardless the geographic source, make mushrooms an important source of essential fat acids for a human health diet.

Production of omega 3, 6, and 9 fatty acids from hydrolysis of vegetable oils and animal fat with Colletotrichum gloeosporioides lipase.

Hydrolysis of vegetable oils (Olive, corn, peanut, sesame, flaxseed, soy, canola, garlic, sunflower, almond, castor bean oils) and beef marrow bone oil by Colletotrichum gloeosporioides lipase was studied. The enzyme was capable of generating free fatty acids from all oils tested. The higher hydrolytic activity of the enzyme was towards olive (18.0 IU) and soybean (17.8 IU) oils. The average percentage of essential fatty acids generated from hydrolysis of the oils was 32.92% of omega 9 (as oleic acid C18:1), 26.24% of omega 6 (linoleic C18:2), and 5.86% of omega 3 (such as α-linolenic acid C18:3). Comparison between chromatographic profile of the oils and its enzymatic hydrolysate showed a good equivalence, stressing the applicability of these vegetable substrates under the action of lipase from C. gloeosporioides produce essential fatty acids, being more efficient production of α-linolenic acid from flaxseed oil, linoleic acid from sunflower oil, and oleic acid from olive.

Antimicrobial activity and acetylcholinesterase inhibition by extracts from chromatin modulated fungi.

Major health challenges as the increasing number of cases of infections by antibiotic multiresistant microorganisms and cases of Alzheimer's disease have led to searching new control drugs. The present study aims to verify a new way of obtaining bioactive extracts from filamentous fungi with potential antimicrobial and acetylcholinesterase inhibitory activities, using epigenetic modulation to promote the expression of genes commonly silenced. For such finality, five filamentous fungal species (Talaromyces funiculosus, Talaromyces islandicus, Talaromyces minioluteus, Talaromyces pinophilus, Penicillium janthinellum) were grown or not with DNA methyltransferases inhibitors (procainamide or hydralazine) and/or a histone deacetylase inhibitor (suberohydroxamic acid). Extracts from T. islandicus cultured or not with hydralazine inhibited Listeria monocytogenes growth in 57.66±5.98% and 15.38±1.99%, respectively. Increment in inhibition of acetylcholinesterase activity was observed for the extract from P. janthinellum grown with procainamide (100%), when compared to the control extract (39.62±3.76%). Similarly, inhibition of acetylcholinesterase activity increased from 20.91±3.90% (control) to 92.20±3.72% when the tested extract was obtained from T. pinophilus under a combination of suberohydroxamic acid and procainamide. Concluding, increases in antimicrobial activity and acetylcholinesterase inhibition were observed when fungal extracts in the presence of DNA methyltransferases and/or histone deacetylase modulators were tested.