Natural Product Mixtures Research Articles

Evaluation of curcuma and ginger mixture ability to prevent ROS production induced by bisphenol S: an in vitro study

The use of bisphenol S (BPS) as a substitute of Bisphenol A is increasing in several products and it can be found in different environmental and biological matrices. Its toxicity has been studied at different levels and one of BPS toxic mechanisms at high concentrations seems to be the induction of oxidative stress through the generation of reactive oxygen species (ROS). This study evaluates the ability of a curcuma and ginger (CG) mixture to exert an antioxidant effect on rat hepatocytes treated with BPS. The effects of the mixture were compared to those of a well-known antioxidant (Trolox). Three different BPS concentrations were used in order to verify ROS production. 70 µg/mL and 150 µg/mL of BPS generated a significant ROS increase (p < 0.01) as compared to control, while CG mixture was able to decrease this ROS production in hepatic cells, as compared to cells treated with 70 µg/ml of BPS (p < 0.01) restoring control levels. BPS 70 µg/mL was tested for total antioxidant capacity (TEAC), superoxide dismutase (SOD) and total thiols. TEAC and SOD significant decreased (p < 0.05 and p < 0.01, respectively) as compared to controls and CG mixture was able to restore control values. Given the widespread BPS use, results obtained in this study can be of high impact for the community, demonstrating the ability of a mixture of natural products to prevent BPS-induced oxidative stress.

Methanol linear gradient counter-current chromatography for the separation of natural products: Sinopodophyllum hexandrum as samples

Counter-current chromatography (CCC) is a unique, liquid-liquid partition chromatography process. Both the mobile and stationary phases are liquids, so no solid support matrix is used. CCC has gained wide acceptance as a preparative technique in a variety of fields. Because the mobile and stationary phases are both liquids, gradient elution is difficult to perform with CCC. Phase equilibrium must be maintained, so any change in the composition of one phase may induce a compositional change in the other. In this work, a new linear gradient elution method was developed for CCC. Biphasic solvent systems containing heptane, ethyl acetate, methanol, and water (HepEMWat) in various ratios were prepared and used to optimize both isocratic and linear gradient CCC separation with methanol. We first separated a test mixture of four standard compounds with partition coefficients ranging from 0.8 to 7.8. The separation resembled a reversed-phase process, and elution was performed while progressively decreasing the polarity of the mobile phase. Target molecules with small partition coefficients eluted first in the lower phase of the optimized HepEMWat solvent system. Elution of constituents with large partition coefficients was quite slow under isocratic conditions. Separation time was significantly reduced when elution was performed with a linear gradient using methanol and the optimal HepEMWat system. Elution with a 3:7:4:6 (v/v/v/v) HepEMWat system took approximately 200 min. This included an 80-min isocratic step, followed by gradient elution with methanol from 0% to 30%. The optimized methanol linear gradient CCC method was then used to separate a complex mixture of natural products isolated from Sinopodophyllum hexandrum (Royle) Ying roots. Twelve compounds with a wide range of polarities were well-resolved in a single separation. We have developed a convenient and cost-effective strategy for the separation of complex mixtures. No tedious mobile phase preparation step is required. The volume of unused mobile phase is minimal, so little solvent is wasted. The method is an important advance for the separation of mixtures that contain many compounds with a large range of polarities and partition coefficients, which are common features of natural products.

Design and Application of a High-Throughput, High-Content Screening System for Natural Product Inhibitors of the Human Parasite Trichomonas vaginalis.

It is estimated that Trichomonas vaginalis affects an astonishing 3.9% of the world's population, and while many of those infected are asymptomatic, progression of the disease can lead to serious health problems. Currently, the nitroimidazoles constitute the only drug class approved to treat trichomoniasis in the United States, which makes the spread of drug resistance a realistic concern. We developed a new image-based, high-throughput, and high-content assay for testing natural products (purified compounds and extracts) for antitrichomonal activity. Applying this assay system to a library of fungal natural product extracts led to the identification of three general classes of natural product inhibitors that exhibited moderate to strong activities against T.vaginalis: anthraquinones, xanthone-anthraquinone heterodimers, and decalin-linked tetramic-acid-containing metabolites. The tetramate natural products emerged as the most promising candidate molecules with pyrrolocin A (51) exhibiting potent activity against the parasite (EC50 = 60 nM), yet this metabolite showed limited toxicity to mammalian cell lines (selectivity index values of 100 and 167 versus 3T3 fibroblast and Ect1 normal cervical cells, respectively). The imaging-based assay system is a powerful tool for the bioassay-guided purification of single-component antitrichomonal biomolecules from complex natural product mixtures.

Opportunities and Limitations for Untargeted Mass Spectrometry Metabolomics to Identify Biologically Active Constituents in Complex Natural Product Mixtures.

Compounds derived from natural sources represent the majority of small-molecule drugs utilized today. Plants, owing to their complex biosynthetic pathways, are poised to synthesize diverse secondary metabolites that selectively target biological macromolecules. Despite the vast chemical landscape of botanicals, drug discovery programs from these sources have diminished due to the costly and time-consuming nature of standard practices and high rates of compound rediscovery. Untargeted metabolomics approaches that integrate biological and chemical data sets potentially enable the prediction of active constituents early in the fractionation process. However, data acquisition and data processing parameters may have major impacts on the success of models produced. Using an inactive botanical mixture spiked with known antimicrobial compounds, untargeted mass spectrometry-based metabolomics data were combined with bioactivity data to produce selectivity ratio models subjected to a variety of data acquisition and data processing parameters. Selectivity ratio models were used to identify active constituents that were intentionally added to the mixture, along with an additional antimicrobial compound, randainal (5), which was masked by the presence of antagonists in the mixture. These studies found that data-processing approaches, particularly data transformation and model simplification tools using a variance cutoff, had significant impacts on the models produced, either masking or enhancing the ability to detect active constituents in samples. The current study highlights the importance of the data processing step for obtaining reliable information from metabolomics models and demonstrates the strengths and limitations of selectivity ratio analysis to comprehensively assess complex botanical mixtures.

A role of mast cells in ovalbumin-induced mouse allergic asthma responses suppressed by natural product mixture (Hwangchango)

A role of mast cells in ovalbumin-induced mouse allergic asthma responses suppressed by natural product mixture (Hwangchango)

Inhibitory Effects of Roseoside and Icariside E4 Isolated from a Natural Product Mixture (No-ap) on the Expression of Angiotensin II Receptor 1 and Oxidative Stress in Angiotensin II-Stimulated H9C2 Cells.

Hypertension is a major risk factor for the development of cardiovascular diseases. This study aimed to elucidate whether the natural product mixture No-ap (NA) containing Pine densiflora, Annona muricate, and Monordica charantia, or its single components have inhibitory effects on hypertension-related molecules in Angiotensin II (Ang II)-stimulated H9C2 cells. Individual functional components were isolated and purified from NA using various columns and solvents, and then their structures were analyzed using ESI–MS, 1H-NMR, and 13H-NMR spectra. H9C2 cells were stimulated with 300 nM Ang II for 7 h. NA, telmisartan, ginsenoside, roseoside (Roseo), icariside E4 (IE4), or a combination of two components (Roseo and IE4) were administered to the cells 1 h before Ang II stimulation. The expression and activity of hypertension-related molecules or oxidative molecules were determined using RT-PCR, western blot, and ELISA. Ang II stimulation increased the expression of Ang II receptor 1 (AT1), tumor necrosis factor-α (TNF-α), monocyte chemoattractant protein-1 (MCP-1), tumor growth factor-β (TGF-β) mRNA, and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity and the levels of hydrogen peroxide (H2O2) and superoxide anion (•O2−) and reduced anti-oxidant enzyme activity. NA significantly improved the expression or activities of all hypertension-related molecules altered in Ang II-stimulated cells. Roseo or IE4 pretreatment either decreased or increased the expression or activities of all hypertension-related molecules similar to NA, but to a lesser extent. The pretreatment with a combination of Roseo and IE4 (1:1) either decreased or increased the expression of all hypertension-related molecules, compared to each single component, revealing a synergistic action of the two compounds. Thus, the combination of single components could exert promising anti-hypertensive effects similar to NA, which should be examined in future animal and clinical studies.

Antimicrobial Activities of AC-ALL from Natural Sources against Acne

Background/Objectives: AC-ALL is the extract mixture of natural products. It is known that AC-ALL has anti-microbial activity on Propionibacterium acnes ( P. acnes ), the causative organism of acne which is gram positive bacterium. The anti-microbial activity against P. acnes is very important to develop cosmetics or drug preventing pimple on face. Here we identified components of AC-ALL which have anti-microbial activity on P. acnes . Methods/Statistical analysis: P. acnes was provided from Korean Collection for Type Cultures (KCTC). AC-ALL is provided from GL Biotech. Co., Ltd. P. acnes was culture in autoclaved GMA broth (Gifu Anaerobic Broth, Nissui 5422, Japan) with anaeropack (MGC, A=0.4, Japan) at 37 o C and the speed of 150 rpm in rectangular jar (MGC, 2.5L A-110, Japan). It also was cultured on autoclaved Plate Count Agar (PCA) with anaeropack at 37 o C for 3 days. The anti-microbial activity of AC-ALL was measured by growth inhibition of P. acnes . To test the anti-microbial activity, the growth of P. acnes was measured by OD 600 in the presence or in the absence of AC-ALL. AC-ALL was analyzed with Gas Chromatography-Mass Spectrometer (Clarus 600 series+TurboMatrix HSS Trap, Perkin Elmer, USA). Findings: In this study, the anti-microbial effect of AC-ALL on P. acnes was tested. We designed experiments to identify the components of AC-ALL showing efficacy to inhibit the growth of P. acnes . First, we tested the anti-microbial activity of AC-ALL by following the growth of P. acnes in the presence or in the absence of AC-ALL. It showed AC-ALL has good inhibitory activity on growth of P. acnes . Even only 2% of the mixes extract can inhibit 90% of the growth of P. acnes . We then analyzed the component of AC-ALL through Gas Chromatography-Mass Spectrometer (GC-MS) and identified six major component peaks such as eucalyptol, Linalool, D-camphor, borneol, caryophyllen and linalyl acetate. We then purchased component compounds from companies one by one and tested the anti-microbial activity of those compounds on P. acnes . When six major compound at the component compound ratio of AC-ALL was added on the growth medium during the culture of P. acnes , the growth of P. acnes was inhibited as much as AC-ALL. Even though any single compound except linalyl acetate was added on growth medium, the growth of P. acnes was also inhibited more or less. These results from this study suggest that six major components have real inhibitory ingredients of AC-ALL on the growth of P acnes. Improvements/Applications: AC-ALL can be applied to develop new cosmetics or drugs to treat the acne. The source of AC-ALL is natural products. So AC-ALL can have advantage over currently used drugs to treat acne in terms of side effect.

Synergy and antagonism in natural product extracts: when 1 + 1 does not equal 2.

Covering: 2000 to 2019 According to a 2012 survey from the Centers for Disease Control and Prevention, approximately 18% of the U.S. population uses natural products (including plant-based or botanical preparations) for treatment or prevention of disease. The use of plant-based medicines is even more prevalent in developing countries, where for many they constitute the primary health care modality. Proponents of the medicinal use of natural product mixtures often claim that they are more effective than purified compounds due to beneficial "synergistic" interactions. A less-discussed phenomenon, antagonism, in which effects of active constituents are masked by other compounds in a complex mixture, also occurs in natural product mixtures. Synergy and antagonism are notoriously difficult to study in a rigorous fashion, particularly given that natural products chemistry research methodology is typically devoted to reducing complexity and identifying single active constituents for drug development. This report represents a critical review with commentary about the current state of the scientific literature as it relates to studying combination effects (including both synergy and antagonism) in natural product extracts. We provide particular emphasis on analytical and Big Data approaches for identifying synergistic or antagonistic combinations and elucidating the mechanisms that underlie their interactions. Specific case studies of botanicals in which synergistic interactions have been documented are also discussed. The topic of synergy is important given that consumer use of botanical natural products and associated safety concerns continue to garner attention by the public and the media. Guidance by the natural products community is needed to provide strategies for effective evaluation of safety and toxicity of botanical mixtures and to drive discovery in botanical natural product research.

The effect of fermentation time on the volatile aromatic profile of tarhana dough.

Tarhana is a traditional food produced by fermentation of a dough prepared by mixing wheat flour, yoghurt, tomato, paprika, onion and mint. The fermentation of mixture of natural products gives the tarhana an aromatized desired food characteristic. Therefore, we aimed to determine the effects of fermentation time at home and commercial scale on the aromatic volatile profile of tarhana. In this respect, nine different tarhana dough samples were prepared at home and commercial scale and analysed for volatile and organic acid composition during fermentation which were then evaluated by principal component analysis. In all dough samples, the lactic, succinic and acetic acids were the most produced organic acids while the amounts of ketones decreased, and the alcohols, in particular the esters, increased within fermentation. Particularly, ethyl-lactate and ethyl-acetate increased significantly as well as ethyl-caprylate, ethyl-capronate and ethyl-hexanote responsible for fruity and flower flavour were also accumulated. Tarhana doughs were clustered in two main groups after five days of fermentation with principal component analysis and these clusters remained stable until the end of fermentation. As a conclusion, fermentation time determines the desirable tarhana by ensuring the formation of aromatic volatiles particularly esters. It is necessary to ferment tarhana for at least five days in order to obtain typical aromatic properties.

Curcumin and Novel Synthetic Analogs in Cell-Based Studies of Alzheimer's Disease.

Alzheimer’s disease (AD) is a chronic neurodegenerative disorder that is associated with the most common type of dementia and is characterized by the presence of deposits of the protein fragment amyloid beta (Aβ) in the brain. The natural product mixture of curcuminoids that improves certain defects in innate immune cells of AD patients may selectively enhance Aβ phagocytosis by alteration of gene transcription. In this work, we evaluated the protective effects of curcuminoids in cells from AD patients by investigating the effect on NF-κB and BACE1 signaling pathways. These results were compared to the gene expression profile of the clearance of Aβ. The minor curcumin constituent, bisdemethoxycurcumin (BDC) showed the most potent protective action to decrease levels of NF-κB and BACE1, decrease the inflammatory cascade and diminish Aβ aggregates in cells from AD patients. Moreover, mannosyl-glycoprotein 4-beta-N-acetylglucosaminyltransferase (MGAT3) and vitamin D receptor (VDR) gene mRNAs were up-regulated in peripheral blood mononuclear cells from AD patients treated with BDC. BDC treatment impacts both gene expression including Mannosyl (Beta-1,4-)-Glycoprotein Beta-1,4-N-Acetylglucosaminyltransferase, Vitamin D and Toll like receptor mRNA and Aβ phagocytosis. The observation of down-regulation of BACE1 and NF-κB following administration of BDC to cells from AD patients as a model system may have utility in the treatment of asymptomatic AD patients.

Shedding Light on Chemically Mediated Tri-Trophic Interactions: A 1H-NMR Network Approach to Identify Compound Structural Features and Associated Biological Activity.

Diverse mixtures of plant natural products play an important role in plant-herbivore-parasitoid interactions. In the pursuit of understanding these chemically-mediated interactions, we are often faced with the challenge of determining ecologically and biologically relevant compounds present in complex phytochemical mixtures. Using a network-based approach, we analyzed binned 1H-NMR data from 196 prepared mixtures of commonly studied secondary metabolites including alkaloids, amides, terpenes, iridoid glycosides, saponins, phenylpropanoids, flavonoids and phytosterols. The mixtures included multiple dimensions of chemical diversity, including molecular complexity, mixture complexity and differences in relative compound concentrations. This approach yielded modules of co-occurring chemical shifts that were correlated with specific compounds or common structural features shared across compounds. This approach was then applied to crude phytochemical extracts of 31 species in the phytochemically diverse tropical plant genus Piper (Piperaceae). Combining the activity of crude plant extracts in an array of bioassays with our 1H-NMR network approach, we identified a potential prenylated benzoic acid from these mixtures that exhibits antifungal properties and identified small structural differences that were potentially responsible for antifungal activity. In an intraspecific analysis of individual Piper kelleyi plants, we also found ontogenetic differences in chemistry that may affect natural plant enemies. In sum, this approach allowed us to characterize mixtures as useful network modules and to combine chemical and ecological datasets to identify biologically important compounds from crude extracts.

Crystalline‐Sponge‐Based Structural Analysis of Crude Natural Product Extracts

AbstractCharacterization of complex natural product mixtures to the absolute structural level of their components often requires significant amounts of starting materials and lengthy purification process, followed by arduous structure elucidation efforts. The crystalline sponge (CS) method has demonstrated utility in the absolute structure elucidation of isolated organic compounds at miniscule quantities compared to conventional methods. In this work, we developed a new CS‐based workflow that greatly expedites the in‐depth structural analysis of crude natural product extracts. Using a crude extract of the red alga Laurencia pacifica, we showed that CS affinity screening prior to compound isolation enables prioritization of analytes present in the extract, and we subsequently resolved the molecular structures of six sesquiterpenes with stereochemical clarity from around 10 mg crude extract. This study demonstrates a new chemotyping workflow that can greatly accelerate natural product discovery from complex samples.

Crystalline-Sponge-Based Structural Analysis of Crude Natural Product Extracts.

Characterization of complex natural product mixtures to the absolute structural level of their components often requires significant amounts of starting materials and lengthy purification process, followed by arduous structure elucidation efforts. The crystalline sponge (CS) method has demonstrated utility in the absolute structure elucidation of isolated organic compounds at miniscule quantities compared to conventional methods. In this work, we developed a new CS-based workflow that greatly expedites the in-depth structural analysis of crude natural product extracts. Using a crude extract of the red alga Laurencia pacifica, we showed that CS affinity screening prior to compound isolation enables prioritization of analytes present in the extract, and we subsequently resolved the molecular structures of six sesquiterpenes with stereochemical clarity from around 10 mg crude extract. This study demonstrates a new chemotyping workflow that can greatly accelerate natural product discovery from complex samples.

The Mevalonate Pathway and Terpenes: a Diversity of Chemopreventatives

The goal of the present review is to give an overview of the most recent papers demonstrating the chemoprevention effects of terpenes. We were interested in showing the structural diversity of these compounds and elucidating overlapping mechanistic factors that may be responsible for chemoprevention by the terpenes in general in these effects. The studies reviewed point to chemoprevention effects across the broad array of structural classes of the terpenes, from the very small-molecule monoterpenes (10 carbon atoms) to the tetraterpenes (40 carbon atoms). These compounds have chemopreventative effects in cancers of the colon, bladder, skin, and liver. Some studies demonstrated that natural product mixtures have more robust effects and that synergy between compounds can be found in these natural products. Terpenes are the most diverse class of compounds on earth. The complex biology of animals including the development of cancers has evolved with these compounds enabling numerous structurally diverse groups of the terpenes to have similar effects on the development and progression of cancers. Mechanistically, these actions tend to cluster into effects of reactive oxygen species and mediators of inflammation which are key players in carcinogenesis.

Acute Modulatory Effects of Apple Cider Vinegar, Garlic, Ginger, Lemon and Honey Mixture, with and Without Exercise on Postprandial Glycemia in Non-Diabetic Females

Postprandial hyperglycemia is independently related to cardiovascular disease. Garlic, ginger, lemon, honey and apple cider vinegar are known to have anti-glycemic properties. However, the effectiveness of combination of these natural products on reducing postprandial glycemia is uncertain. The aim of the present study was to investigate the glucoselowering effect of a novel mixture consisting of apple cider vinegar, garlic, ginger, lemon, and honey; alone and in combination with exercise in response to a high-carbohydrate meal in non-diabetic individuals. Ten, female subjects (mean age: 25 ± 2.67 years, mean BMI: 22.6 ± 3.5 kg/m2 ) participated in this randomised, cross-over intervention consisting of four trials: control (CON), mixture only (MIX), exercise only (EX), and exercise + mixture (EX-MIX). All trials involved consumption of a high-carbohydrate breakfast, then followed by rest in CON, consumption of natural product mixture in MIX, brisk-walking exercise in EX, and combination of mixture and exercise in EX-MIX. Blood glucose was measured at fasting, and at 30, 60, 90, 120 minutes post meal. Postprandial glucose response was calculated as area under the glucose curve. Two-way repeated measures ANOVA showed a significant group and time interaction (p < 0.001). Compared to CON, postprandial glucose responses were 8%, 13% and 15% lower in MIX (p = 0.049), EX (p = 0.001) and EX-MIX (p = 0.005) respectively. Postprandial glucose was 8% lower in EX-MIX compared to MIX (p = 0.002). In conclusion, consuming natural product mixture containing garlic, ginger, lemon, honey and apple cider vinegar reduced postprandial glycemia to a certain extent, however, combining mixture with exercise produced a greater attenuation effect compared to consuming mixture alone. This finding is indicative of a potential benefit of the novel mixture as a complementary management of hyperglycemia in high-risk individuals. DOI : http://dx.doi.org./10.17576/JSKM-2018-15

Acute Modulatory Effects of Apple Cider Vinegar, Garlic, Ginger, Lemon and Honey Mixture, with and Without Exercise on Postprandial Glycemia in Non-Diabetic Females

Postprandial hyperglycemia is independently related to cardiovascular disease. Garlic, ginger, lemon, honey and apple cider vinegar are known to have anti-glycemic properties. However, the effectiveness of combination of these natural products on reducing postprandial glycemia is uncertain. The aim of the present study was to investigate the glucoselowering effect of a novel mixture consisting of apple cider vinegar, garlic, ginger, lemon, and honey; alone and in combination with exercise in response to a high-carbohydrate meal in non-diabetic individuals. Ten, female subjects (mean age: 25 ± 2.67 years, mean BMI: 22.6 ± 3.5 kg/m2 ) participated in this randomised, cross-over intervention consisting of four trials: control (CON), mixture only (MIX), exercise only (EX), and exercise + mixture (EX-MIX). All trials involved consumption of a high-carbohydrate breakfast, then followed by rest in CON, consumption of natural product mixture in MIX, brisk-walking exercise in EX, and combination of mixture and exercise in EX-MIX. Blood glucose was measured at fasting, and at 30, 60, 90, 120 minutes post meal. Postprandial glucose response was calculated as area under the glucose curve. Two-way repeated measures ANOVA showed a significant group and time interaction (p &lt; 0.001). Compared to CON, postprandial glucose responses were 8%, 13% and 15% lower in MIX (p = 0.049), EX (p = 0.001) and EX-MIX (p = 0.005) respectively. Postprandial glucose was 8% lower in EX-MIX compared to MIX (p = 0.002). In conclusion, consuming natural product mixture containing garlic, ginger, lemon, honey and apple cider vinegar reduced postprandial glycemia to a certain extent, however, combining mixture with exercise produced a greater attenuation effect compared to consuming mixture alone. This finding is indicative of a potential benefit of the novel mixture as a complementary management of hyperglycemia in high-risk individuals. DOI : http://dx.doi.org./10.17576/JSKM-2018-15

An Introduction to Plant Tissue Culture: Advances and Perspectives.

Plant tissue culture techniques are the most frequently used biotechnological tools for basic and applied purposes ranging from investigation on plant developmental processes, functional gene studies, commercial plant micropropagation, generation of transgenic plants with specific industrial and agronomical traits, plant breeding and crop improvement, virus elimination from infected materials to render high-quality healthy plant material, preservation and conservation of germplasm of vegetative propagated plant crops, and rescue of threatened or endangered plant species. Additionally, plant cell and organ cultures are of interest for the production of secondary metabolites of industrial and pharmaceutical interest. New technologies, such as the genome editing ones combined with tissue culture and Agrobacterium tumefaciens infection, are currently promising alternatives for the highly specific genetic manipulation of interesting agronomical or industrial traits in crop plants. Application of omics (genomics, transcriptomics, and proteomics) to plant tissue culture will certainly help to unravel complex developmental processes such as organogenesis and somatic embryogenesis, which will probably enable to improve the efficiency of regeneration protocols for recalcitrant species. Additionally, metabolomics applied to tissue culture will facilitate the extraction and characterization of complex mixtures of natural plant products of industrial interest. General and specific aspects and applications of plant tissue culture and the advances and perspectives are described in this edition.

A Combined LC-MS Metabolomics- and 16S rRNA Sequencing Platform to Assess Interactions between Herbal Medicinal Products and Human Gut Bacteria in Vitro: a Pilot Study on Willow Bark Extract.

Herbal preparations are complex mixtures of natural products, many of which are able to reach the distal gut due to low oral bioavailability. There, they can influence the microbial communities, and can be metabolized into potentially absorbable bioactive compounds by the intestinal bacteria. This aspect has often been disregarded when searching for the active principles of medicinal plants and herbal medicinal products. The aim of this study was to establish an interdisciplinary platform to unravel interactions of herbal medicine and intestinal microbiota, using a combined LC-MS metabolomics and 16S rRNA microbiome sequencing approach. Willow bark extract (WBE), a herbal medicinal product with a long history of traditional use and a well-established anti-inflammatory activity, was incubated with human fecal suspension under anoxic conditions. Samples were taken after 0.5, 4, and 24 h of incubation. Microbiome analyses revealed that incubation with WBE had a marked effect on microbial community composition and functions. For example, the proportion of Bacteroides sp. was clearly enhanced when the fecal sample used in this study was incubated with WBE. LC-MS analysis showed that WBE constituents were readily metabolized by fecal bacteria. Numerous microbial metabolites could be annotated, allowing the construction of putative microbial degradation pathways for the main groups of WBE constituents. We suggest that studies of this type help to increase the knowledge on bioactive principles of medicinal plants, since gut microbial metabolites might have been underestimated as a source of bioactive compounds in the past.

An efficient computer-aided structural elucidation strategy for mixtures using an iterative dynamic programming algorithm

The identification of chemical structures in natural product mixtures is an important task in drug discovery but is still a challenging problem, as structural elucidation is a time-consuming process and is limited by the available mass spectra of known natural products. Computer-aided structure elucidation (CASE) strategies seek to automatically propose a list of possible chemical structures in mixtures by utilizing chromatographic and spectroscopic methods. However, current CASE tools still cannot automatically solve structures for experienced natural product chemists. Here, we formulated the structural elucidation of natural products in a mixture as a computational problem by extending a list of scaffolds using a weighted side chain list after analyzing a collection of 243,130 natural products and designed an efficient algorithm to precisely identify the chemical structures. The complexity of such a problem is NP-complete. A dynamic programming (DP) algorithm can solve this NP-complete problem in pseudo-polynomial time after converting floating point molecular weights into integers. However, the running time of the DP algorithm degrades exponentially as the precision of the mass spectrometry experiment grows. To ideally solve in polynomial time, we proposed a novel iterative DP algorithm that can quickly recognize the chemical structures of natural products. By utilizing this algorithm to elucidate the structures of four natural products that were experimentally and structurally determined, the algorithm can search the exact solutions, and the time performance was shown to be in polynomial time for average cases. The proposed method improved the speed of the structural elucidation of natural products and helped broaden the spectrum of available compounds that could be applied as new drug candidates. A web service built for structural elucidation studies is freely accessible via the following link (http://csccp.cmdm.tw/).

Enriching the biological space of natural products and charting drug metabolites, through real time biotransformation monitoring: The NMR tube bioreactor

BackgroundNatural products offer a wide range of biological activities, but they are not easily integrated in the drug discovery pipeline, because of their inherent scaffold intricacy and the associated complexity in their synthetic chemistry. Enzymes may be used to perform regioselective and stereoselective incorporation of functional groups in the natural product core, avoiding harsh reaction conditions, several protection/deprotection and purification steps. MethodsHerein, we developed a three step protocol carried out inside an NMR-tube. 1st-step: STD-NMR was used to predict the: i) capacity of natural products as enzyme substrates and ii) possible regioselectivity of the biotransformations. 2nd-step: The real-time formation of multiple-biotransformation products in the NMR-tube bioreactor was monitored in-situ. 3rd-step: STD-NMR was applied in the mixture of the biotransformed products to screen ligands for protein targets. ResultsHerein, we developed a simple and time-effective process, the “NMR-tube bioreactor”, that is able to: (i) predict which component of a mixture of natural products can be enzymatically transformed, (ii) monitor in situ the transformation efficacy and regioselectivity in crude extracts and multiple substrate biotransformations without fractionation and (iii) simultaneously screen for interactions of the biotransformation products with pharmaceutical protein targets. ConclusionsWe have developed a green, time-, and cost-effective process that provide a simple route from natural products to lead compounds for drug discovery. General significanseThis process can speed up the most crucial steps in the early drug discovery process, and reduce the chemical manipulations usually involved in the pipeline, improving the environmental compatibility.