Application of metabolomics and molecular networking in investigating the chemical profile and antitrypanosomal activity of British bluebells (Hyacinthoides non-scripta)

Bulb, leaf, scape and flower samples of British bluebells (Hyacinthoides non-scripta) were collected regularly for one growth period. Methanolic extracts of freeze-dried and ground samples showed antitrypanosomal activity, giving more than 50% inhibition, for 20 out of 41 samples. High-resolution mass spectrometry was used in the dereplication of the methanolic extracts of the different plant parts. The results revealed differences in the chemical profile with bulb samples being distinctly different from all aerial parts. High molecular weight metabolites were more abundant in the flowers, shoots and leaves compared to smaller molecular weight ones in the bulbs. The anti-trypanosomal activity of the extracts was linked to the accumulation of high molecular weight compounds, which were matched with saponin glycosides, while triterpenoids and steroids occurred in the inactive extracts. Dereplication studies were employed to identify the significant metabolites via chemotaxonomic filtration and considering their previously reported bioactivities. Molecular networking was implemented to look for similarities in fragmentation patterns between the isolated saponin glycoside at m/z 1445.64 [M + formic-H]− equivalent to C64H104O33 and the putatively found active metabolite at m/z 1283.58 [M + formic-H]− corresponding to scillanoside L-1. A combination of metabolomics and bioactivity-guided approaches resulted in the isolation of a norlanostane-type saponin glycoside with antitrypanosomal activity of 98.9% inhibition at 20 µM

Bioactive secondary metabolites from the endophytic fungus Chaetomium sp. isolated from Salvia officinalis growing in Morocco

This study reports the chemical investigation and cytotoxic activity of the secondary metabolites produced by the endophytic fungus Chaetomium sp. isolated from Salvia officinalis growing in Morocco. This plant was collected from the Beni-Mellal Mountain in Morocco and belongs to the Lamiaceae family and is named in Morocco 'Salmia'. The endophytic fungus Chaetomium sp. was isolated from the tissues of the stem of this plant. The fungal strain was identified by PCR. The crude organic extract of the fungal strain was proven to be active when tested for cytotoxicity against L5178Y mouse lymphoma cells. Chemical investigation of the secondary metabolites showed that cochliodinol is the main component beside isocochliodinol. The structures of the isolated compounds were determined on the basis of NMR analysis (1H, 13C, COSY and HMBC) as well as by mass spectrometry using ESI (Electron Spray Ionisation) as source

Metabolomics and dereplication strategies in the discovery of natural product derived drugs

Metabolomics is the technology designed to provide general qualitative and quantitative profile of metabolites in organisms exposed to different conditions. Metabolomics is applied in many aspects of natural drug discoveries, particularly in bioactivity screening to improve dereplication and identification procedures. Fast dereplication of known compounds and identification of lead bioactive metabolites is important in the primary stages of metabolomics profiling prior to an intensive isolation work. Two levels of metabolomics were used in this study. First was metabolites fingerprinting which aimed for rapid classification of samples by comparing the metabolites patterns or fingerprints. Second was metabolites profiling and dereplication study for class of compounds related to a specific pathway in order to individually identify and quantify these metabolites. This study involved isolation of endophytic fungus Aspergillus aculeatus from Egyptian medicinal plants, Terminalia laxiflora. Identification of the strains has been achieved through molecular biological methods. Metabolomic profiling, using NMR and HR-MS were done at different stages of the growth phase for both solid and liquid culture media. Dereplication studies were accomplished by utilizing the MZmine 2.10 software with aid of the AntiBase and DNP databases. By end of the dereplication process metabolites were sorted out into three levels; level 1: identified compounds, level 2: putatively annotated compound class and level 3: completely unidentified and unclassified compounds. Multivariate data analysis was employed by using PCA in order to classify samples into groups, trends and outliers, which maximize the information, can be obtained from spectral data. OPLS-DA was used to correlate the chemical profile with tested biological activity. Metabolomics has been shown to be a powerful facilitator in the discovery of natural products, which are considered an excellent source for novel leads

Bioactive secondary metabolites from the endophytic fungus Chaetomium sp. isolated from Salvia officinalis growing in Morocco

This study reports the chemical investigation and cytotoxic activity of the secondary metabolites produced by the endophytic fungus Chaetomium sp. isolated from Salvia officinalis growing in Morocco. This plant was collected from the Beni-Mellal Mountain in Morocco and belongs to the Lamiaceae family and is named in Morocco “Salmia”. The endophytic fungus Chaetomium sp. was isolated from the tissues of the stem of this plant. The fungal strain was identified by PCR. The crude organic extract of the fungal strain was proven to be active when tested for cytotoxicity against L5178Y mouse lymphoma cells. Chemical investigation of the secondary metabolites showed that cochliodinol is the main component beside isocochliodinol. The structures of the isolated compounds were determined on the basis of NMR analysis (1H, 13C, COSY and HMBC) as well as by mass spectrometry using ESI (Electron Spray Ionisation) as source

In-vivo antimalarial activity of the endophytic actinobacterium, Streptomyces SUK 10

Endophytic bacteria, such as Streptomyces, have the potential to act as a source for novel bioactive molecules with medicinal properties. The present study was aimed at assessing the antimalarial activity of crude extract isolated from various strains of actinobacteria living endophytically in some Malaysian medicinal plants. Using the four day suppression test method on male ICR strain mice, compounds produced from three strains of Streptomyces (SUK8, SUK10, and SUK27) were tested in vivo against Plasmodium berghei PZZ1/100 in an antimalarial screen using crude extracts at four different concentrations. One of these extracts, isolated from Streptomyces SUK10 obtained from the bark of Shorea ovalis tree, showed inhibition of the test organism and was further tested against P. berghei-infected mice for antimalarial activity at different concentrations. There was a positive relationship between the survival of the infected mouse group treated with 50 μg/kg body weight (bw) of ethyl acetate-SUK10 crude extract and the ability to inhibit the parasites growth. The parasite inhibition percentage for this group showed that 50% of the mice survived for more than 90 days after infection with the parasite. The nucleotide sequence and phylogenetic tree suggested that Streptomyces SUK10 may constitute a new species within the Streptomyces genus. As part of the drug discovery process, these promising finding may contribute to the medicinal and pharmaceutical field for malarial treatment

Metabolomic tools to assess the chemistry and bioactivity of endophytic aspergillus strain

Endophytic fungi associated with medicinal plants are a potential source of novel chemistry and biology that may find applications as pharmaceutical and agrochemical drugs. In this study, a combination of metabolomics and bioactivity-guided approaches were employed to isolate anticancer secondary metabolites from an endophytic Aspergillus aculeatus. The endophyte was isolated from the Egyptian medicinal plant Terminalia laxiflora and identified using molecular biological methods. Metabolomics and dereplication studies were accomplished by utilizing the MZmine software coupled with the universal Dictionary of Natural Products database. Metabolic profiling, with aid of multivariate data analysis, was performed at different stages of the growth curve to choose the optimised method suitable for up-scaling. The optimised culture method yielded a crude extract abundant with biologically-active secondary metabolites. Crude extracts were fractionated using different high-throughput chromatographic techniques. Purified compounds were identified by HRESI-MS, 1D and 2D-NMR. This study introduced a new method of dereplication utilising both high-resolution mass spectrometry and NMR spectroscopy. The metabolites were putatively identified by applying a chemotaxonomic filter. We also present a short review on the diverse chemistry of terrestrial endophytic strains of Aspergillus, which has become a part of our dereplication work and this will be of wide interest to those working in this field. This article is protected by copyright. All rights reserved

Exploring the Chemical Space of Macro- and Micro-Algae Using Comparative Metabolomics

With more than 156,000 described species, eukaryotic algae (both macro- and micro-algae) are a rich source of biological diversity, however their chemical diversity remains largely unexplored. Specialised metabolites with promising biological activities have been widely reported for seaweeds, and more recently extracts from microalgae have exhibited activity in anticancer, antimicrobial, and antioxidant screens. However, we are still missing critical information on the distinction of chemical profiles between macro- and microalgae, as well as the chemical space these metabolites cover. This study has used an untargeted comparative metabolomics approach to explore the chemical diversity of seven seaweeds and 36 microalgal strains. A total of 1390 liquid chromatography-mass spectrometry (LC-MS) features were detected, representing small organic algal metabolites, with no overlap between the seaweeds and microalgae. An in-depth analysis of four Dunaliella tertiolecta strains shows that environmental factors may play a larger role than phylogeny when classifying their metabolomic profile

Endophytic Streptomyces SUK10: its molecular characteristic and bioactivity against malaria parasite

It has been known that Streptomyces is the mains source for antibiotics. The present study found that the crude extract obtained from Streptomyces SUK10 living endophytically in Shorea ovalis tree has antimalarial activity against Plasmodium berghei strain PZZ1/100. Using validated surface sterilization and isolation method, Streptomyces SUK10 was isolated from the bark of Shorea ovalis by and cultured on the isolation agar. With spiral spore chain, microscopically, this strain has substrate mycelium in brownish yellow and aerial mycelium in whitish grey, other than yellow dissolved pigment. As showed in early molecular tests, the 23S rRNA sequence of Streptomyces SUK10 was 96% identical to Streptomyces hygroscopicus, the outstanding producer of Hygromycin B antibiotic. In early phase of antimalarial screening, the secondary metabolites from three different strains of endophytic Streptomyces isolated from three different trees were in-vivo tested on the mice ICR strain against P. berghei strain PZZ1/100. These were done by implementing the four days (4D) suppression test using their own ethyl acetate-crude extract at four different concentrations: 50, 100, 200 and 400 µg/kg body weight (bw). Upon obtaining significant results (p ≤ 0.05), ethyl acetate-SUK10 crude extract were screened for fully anti-malarial activity at five different narrower concentration range 5, 10, 50, 100 and 200 µg/kg bw. There was a positive relationship between the survival of the infected mice group treated with 50 µg/kg body weight of ethyl acetate-SUK10 crude extract and its ability to inhibit the parasites growth. Considering quinine hydrochloride at 10 mg/kg bw and 0.9% normal saline as positive and negative control respectively, the parasite inhibition percentage for this group at certain concentration showed more than 70% and 50% of this mice members (n=3) were also able to survive more than 90 days after the infection with the parasite. Nevertheless, Indole-3-lactic acid (C11H11NO3) was isolated from the anti-trypanosomal active fraction while Gancidin W or Maculosin 6 (L-leucyl-L-prolyl lactam, C11H18N2O2) was the major compound found from SUK 10 crude extract. Gancidin W has been previously reported to be anti-parasitic which we are still yet to be tested for anti-malarial activity and its cytotoxicity. These promising findings contribute to the medicinal and pharmaceutical field for future antimalarial treatment

Gancidin W, a potential low toxicity antimalarial agent isolated from an Endophytic Streptomyces SUK10

As it resisted against nearly all current drugs, malaria was reported remained as the most threatening human parasitic disease. In line with this, endophytic Streptomyces are potential sources for novel bioactive molecules. In this study, prolyl-leucyl-diketopiperazine (C11H18N2O2) or Gancidin W (GW) was successfully isolated from Streptomyces SUK10 that inhabited in the bark of Shorea ovalis. Using four days suppressive test (4DST), this compound was in-vivo tested against Plasmodium berghei NK65. At 6.25 and 3.125 µg kg-1 body weight (bw), there was a very significant relationship of the ability to inhibit the growth of P. berghei NK65 when GW exhibited an inhibition rate of nearly 80% on male ICR strain mice. Comparing GW with dH2O diluted quinine hydrochloride and 0.9% normal saline as positive and negative controls respectively, 50% of the mice group treated with 3.125 µg kg-1 bw was managed to survive more than eight months post-infection exposure where this survival ranged was exceeded half of the normal mice’s life span period. As in-vivo toxicity assessment towards selected vital organs and blood enzymes were also investigated, the ALT and AST enzymes level was slightly higher but there was no abnormalities and injuries were found on the tested organs of the mice group treated with GW at this concentration. These findings indicated that GW isolated from Streptomyces SUK10 exhibited very low toxicity and is a good candidate as a potential antimalarial agent