Natural Products & Metabolomics

Alzheimer´s disease (AD) is the most common form of dementia affecting predominantly elderly people from developed countries. One aspect of the illness is that patients suffer from an impaired memory due to deposition of aggregated A-peptides forming amyloid plaques. According to the glutaminyl cyclase (QC) hypothesis this enzyme plays a key role in generating neurotoxic amyloid peptides (amyloid-β or Aβ) by modifying the N-terminus of peptides to N-pyroglutamated derivatives. These modified proteins are resistant to degradation and are at the same time “seeds” for the formation of toxic A-oligomers in the brain. In order to screen for natural inhibitors of QC, strains of different species of algae belonging to Chlorophyceae and Eustigmatophyceae were cultivated in 100 L tubular photobioreactors. The resulting crude extracts of algae from exponential and stationary growth phases were tested for their inhibition properties of glutaminyl cyclase (QC). Here 27 of the 72 tested extracts inhibited the QC. Fractions separated by Sephadex G-15 column chromatography also showed QC inhibition activity.

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Trigonelline (3-carboxy-1-methyl pyridinium) was identified as a relevant bioactivity and taste imparting component in Balanites aegyptiaca fruit, using 1H NMR of crude extracts without any fractionation or isolation step. The structural integrity of trigonelline was established within the extract matrix via1H NMR, 1H–1H COSY, HMQC and HMBC and by comparison with authentic standard. A quantitative 1H NMR method (qHNMR) was used to determine trigonelline concentrations in the peel and pulp of B. aegyptiaca fruit of 8 and 13 mg g−1, respectively. Trigonelline so far has not been reported from B. aegyptiaca or its genus as it easily escapes LC–MS based detection. Its discovery provides novel insight into the balanite fruits antidiabetic properties as the compound is known for a pronounced hypoglycemic effect. In addition, it is likely to impart the perceptible bitter taste portion to balanites sweet bitter taste. UPLC–MS of the crude extract additionally revealed the fruit flavonoid pattern showing quercetin/isorhamnetin flavonol conjugates in addition to epicatechin, the latter being present at much lower levels.

Glycyrrhiza glabra, commonly known as licorice, is a popular herbal supplement used for the treatment of chronic inflammatory conditions and as sweetener in the food industry. This species contains a myriad of phytochemicals including the major saponin glycoside glycyrrhizin (G) of Glycyrrhetinic acid (GA) aglycone. In this study, 2D-ROESY NMR technique was successfully applied for distinguishing 18α and 18β glycyrrhetinic acid (GA). ROESY spectra acquired from G. glabra, Glycyrrhiza uralensis and Glycyrrhiza inflata crude extracts revealed the presence of G in its β-form. Anti-inflammatory activity of four Glycyrrhiza species, G, glabra, G. uralensis, G. inflata, and G. echinata roots was assessed against COX-1 inhibition revealing that phenolics rather than glycyrrhizin are biologically active in this assay. G. inflata exhibits a strong cytotoxic effect against PC3 and HT29 cells lines, whereas other species are inactive. This study presents an effective NMR method for G isomer assignment in licorice extracts that does not require any preliminary chromatography or any other purification step.

Bacterial wilts of potato, tomato, pepper, and or eggplant caused by Ralstonia solanacearum are among the most serious plant diseases worldwide. In this study, the issue of developing bactericidal agents from natural sources against R. solanacearum derived from plant extracts was addressed. Extracts prepared from 25 plant species with antiseptic relevance in Egyptian folk medicine were screened for their antimicrobial properties against the potato pathogen R. solancearum by using the disc‐zone inhibition assay and microtitre plate dilution method. Plants exhibiting notable antimicrobial activities against the tested pathogen include extracts from Acacia arabica and Punica granatum. Bioactivity‐guided fractionation of A. arabica and P. granatum resulted in the isolation of bioactive compounds 3,5‐dihydroxy‐4‐methoxybenzoic acid and gallic acid, in addition to epicatechin. All isolates displayed significant antimicrobial activities against R. solanacearum (MIC values 0.5–9 mg/ml), with 3,5‐dihydroxy‐4‐methoxybenzoic acid being the most effective one with a MIC value of 0.47 mg/ml. We further performed a structure–activity relationship (SAR) study for the inhibition of R. solanacearum growth by ten natural, structurally related benzoic acids.

Bauhinia L. (Fabaceae) comprises ca. 300–350 plant species, many of which are traditionally used in folk medicine for their antidiabetic, antioxidant and anti-inflammatory effects. Bauhinia s.l. recently has been subdivided into 9 genera based on phylogenetic data: Bauhinia s.str., Barklya, Brenierea, Gigasiphon, Lysiphyllum, Phanera, Piliostigma, Schnella (American Phanera) and Tylosema. The aerial parts of 8 species corresponding to 5 genera were analyzed: Bauhinia forficata, Bauhinia variegata, B. variegata var. candida, Bauhinia galpinii, Schnella glabra, Piliostigma racemosa, Phanera vahlii and Lysiphyllum hookeri. Leaves and shoots were subjected to metabolite profiling via UHPLC–PDA–qTOF-MS coupled to multivariate data analyzes to identify compound compositional differences. A total of 90 metabolites were identified including polyphenols and fatty acids; flavonoid conjugates accounted for most of the metabolite variation observed. This study provides a comprehensive map of polyphenol composition in Bauhinia and phytochemical species aggregations are consistent with recent Bauhinia genus taxonomic relationship derived from phylogenetic studies. DPPH radical scavenging and α-glucosidase inhibitory assays were also performed to assess selected aspects of the antioxidant and antidiabetic potential for the examined species with respect to metabolite profiles.

Senna alexandrina Mill (Cassia acutifolia and Cassia angustifolia) are used for the laxative medicine Senna. Leaves and pods from two geographically different sources were distinguished from each other via proton nuclear magnetic resonance (1H-NMR) and ultra performance liquid chromatography-mass spectrometry (UPLC-MS) analysis. Under optimized conditions, we were able to simultaneously quantify and identify 107 metabolites including 21 anthraquinones, 24 bianthrones (including sennosides), 5 acetophenones, 25 C/O-flavonoid conjugates, 5 xanthones, 3 naphthalenes, 2 further phenolics, and 9 fatty acids. Principal component analysis (PCA) and hierarchical clustering analysis (HCA) were used to define both similarities and differences among samples. For sample classification based on tissue type (leaf and pod), both UPLC-MS and NMR were found to be more effective in separation than on geographical origin. Results reveal that the amounts of the major classes of bioactives in Senna, i.e., flavonoids and sennosides, varied significantly among organs. Leaves contained more flavonoids and ω-3 fatty acids but fewer benzophenone derivatives than pods. In contrast, pods were more enriched in bianthrones (sennosides). PCA analysis was found to be equally effective in predicting the origin of the commercial Senna preparation using NMR and/or UPLC-MS datasets. Furthermore, a selection of six so far uninvestigated Senna species were analyzed by UPLC-MS. Results revealed that the Senna alata leaf in terms of secondary metabolite composition is the most closely related species to S. alexandrina Mill, showing the highest levels of the anthraquinone “rhein” and flavonoid conjugates. To the best of our knowledge, this study provides the first approach utilizing both UPLC-MS and NMR to reveal secondary metabolite compositional differences among Senna species.

Despite recent advances in studies on trade‐offs between plant defence traits, little is known about whether trade‐offs reflect (i) evolutionary constraints at the species level or (ii) allocation constraints at the individual level. Here, we asked to which degree physical and chemical carbon‐based leaf defence traits covary within and across species.We assessed leaf toughness, leaf total phenolic and tannin concentrations for 51 subtropical tree species. Species trait means, sample‐specific values and phylogenetically independent contrasts were used in regression analyses. Phylogenetic signals and trait evolution were assessed along the phylogeny.Analyses of species‐level trait means revealed significant negative trait covariations between physical and chemical defence traits in analyses over all species. These covariations were inconsistent at the within‐species level. All three defence aspects showed strong phylogenetic signals, but differed in the degree of conservatism along the phylogeny. Inclusion of intraspecific trait variability significantly decreased the strength of these covariations. Strong negative covariations were detected between physical and chemical defence traits when phylogenetic non‐independence was accounted for.Synthesis. We addressed two sources of variation (allocation and evolution) independently from each other in the context of trait interrelationships. The observed negative covariations hint at the existence of a trade‐off between physical and chemical defence traits. The finding that intraspecific trait variation contributed less to this relationship suggests that the trade‐off is dominated by evolutionary constraints rather than by carbon allocation constraints.

Only certain flavonoids have been shown to enhance life span. This was pointed out for e.g. myricetin in the nematode Caenorhabditis elegans. However, the structural requirements responsible for this effect are not known. We used methylated derivatives of myricetin (laricitrin, syringetin, myricetintrimethylether) to investigate if free OH moieties in the B-ring are necessary for the life span extending effect. In analogy to myricetin, all derivatives increased the life span, decreased oxidative stress (DCF) and decreased the accumulation of lipofuscin. In contrast to myricetin, the methylated compounds strongly enhanced the resistance against thermal stress. Furthermore, treatment with the derivatives induced a much stronger nuclear localization of the DAF-16 transcription factor (FoxO homologue). Additionally, no antioxidant effects and only minor effects on life span prolongation and stress resistance were detectable for the methylated compounds in a DAF-16 deficient nematode strain. Comparable to the dietary flavonoid myricetin, the methylated myricetin derivatives laricitrin, syringetin and myricetintrimethylether strongly enhance the life span of C. elegans. Therefore, OH groups of ring B are not necessary for this effect. Only the methylated compounds increase the stress resistance of the nematode which was dependent on DAF-16. These findings suggest that methylation of myricetin increases the biofunctionality.

Zanthoxylum lemairei is widely used in African folk medicine for its pharmacological relevance. Chemical investigation of the ethanol extract from the leaves of this plant lead to the isolation of two new tetraflavonoids, lemairones A (1) and B (2), along with three known compounds, lupeol, sitosterol, and sitosterol 3-O-β-d-glucopyranoside. The antibacterial screening of the leaves of this plant, characterization of compounds 1 and 2, and their antibacterial activity are reported for the first time. The isolation of the compounds was performed using different chromatographic methods while their structures were elucidated by spectroscopic techniques including MS and NMR, and by comparison of data with those of similar flavonoids reported in the literature. The isolated compounds and the crude extract were tested against ten Gram negative multi-resistance bacterial strains including clinical isolates using a broth dilution method. The crude ethanol extract showed weak activity against the tested bacteria strains with a minimal inhibitory concentration (MIC) ranging from 512 to 1024 μg/mL. Among the isolated metabolites, only the new tetraflavonoids were tested. Lemairone A displayed weak activity while lemairone B had moderate activity against the resistant Escherichia coli AG100 with MIC values of 128 μg/mL and 64 μg/mL respectively. In addition, both molecules displayed weak activity against Klebsiella pneumoniae KP55 (MIC 128 μg/mL).