Isolation of anticancer and anti-trypanosome secondary metabolites from the endophytic fungus Aspergillus flocculus via bioactivity guided isolation and MS based metabolomics

Abstract

This study aims to identify bioactive anticancer and anti-trypanosome secondary metabolites from the fermentation culture of Aspergillus flocculus endophyte assisted by modern metabolomics technologies. The endophyte was isolated from the stem of the medicinal plant Markhamia platycalyx and identified using phylogenetics. Principle component analysis was employed to screen for the optimum growth endophyte culturing conditions and revealing that the 30-days rice culture (RC-30d) provided the highest levels of the bioactive agents. To pinpoint for active chemicals in endophyte crude extracts and successive fractions, a new application of molecular interaction network is implemented to correlate the chemical and biological profiles of the anti-trypanosome active fractions to highlight the metabolites mediating for bioactivity prior to purification trials. Multivariate data analysis (MVDA), with the aid of dereplication studies, efficiently annotated the putatively active anticancer molecules. The small-scale RC-30d fungal culture was purified using high-throughput chromatographic techniques to yield compound 1, a novel polyketide molecule though inactive. Whereas, active fractions revealed from the bioactivity guided fractionation of medium scale RC-30d culture were further purified to yield 7 metabolites, 5 of which namely cis-4-hydroxymellein, 5-hydroxymellein, diorcinol, botryoisocoumarin A and mellein, inhibited the growth of chronic myelogenous leukaemia cell line K562 at 30 μM. 3hydroxymellein and diorcinol exhibited a respective inhibition of 56% and 97% to the sleeping sickness causing parasite Trypanosoma brucei brucei. More interestingly, the antitrypanosomal activity of A. flocculus extract appeared to be mediated by the synergistic effect of the active steroidal compounds i.e. ergosterol peroxide, ergosterol and campesterol. The isolated structures were elucidated by using 1D, 2D NMR and HR-ESIMS.

Melleins—Intriguing Natural Compounds

“…Compounds 18, 34, 49 and 1 inhibited the growth of chronic myelogenous leukemia cell line K562 at 30 µM. Compound 31 exhibited an inhibition of 56% to the sleeping-sickness-causing parasite Trypanosoma brucei brucei [80]. Compound 49, characterized as (3R)-3-methoxymellein, was also previously obtained from the fermentation culture of Botryosphaeria sp.…”

“…Apiospora montagnei [40] larvicidal [59] Aspergillusflocculus [55] citotoxicity [80] Aspergillus melleus [3] Aspergillusochraceus [21,22,58] HCV protease inhibitor [38] Aspergillus oniki 1784 [19] Botryosphaeria mamane PSU-M76 [42] Botryosphearia obtusa phytotoxicity [29,44] Bothryosphaeria rhodina antibacterical [45] Colletotrichum gloeosporioides [82] Diplodia africana [48] Diplodia fraxini [72] Diplodia mutila [78] Diplodia seriata [78] Epicoccum nigrum [51] Eudistoma vannamei [55] Hypoxylon deustum [26] Hypoxylon fragiforme [26] Hypoxylon howeianum [26] Hypoxylon haematostroma [26] Hypoxylon venusfuissimum [26] Lachnum palmae [75,76] Lasiodiplodia euphorbicola [70] Lasiodiplodia theobromae [20,71] Lasiodiplodia sp. [63] Microsphaeropsis sp.…”

“…Apiospora montagnei larvicidal [59] Aspergillus flocculus [55] citotoxicity [80] Botryosphaeria mamane PSU-M76 larvicidal [42] Botryosphaeria obtusa phytotoxic [30,44] Botryosphaeria rhodina antibacterical [45] Diplodia africana [48] Diplodia seriata [78] Eudistoma vannamei [55] Lasiodiplodia euphorbicola [70] Lachnum palmae [75,76] Microsphaeropsis sp. [37] Neofusicoccum parvum phytotocicity [47,69] Neofusicoccum luteum [78] Sardiniella urbana [79] Seimatosporium sp.…”

“…Aspergillus flocculus [80] Botryosphaeria obtusa phytotoxic [30,31] Botryosphaeria rhodina antibacterial [45] Biscogniauxia cylindrospora BCRC 33717 [49] Lachnum palmae [75,76] Paraconiothyrium sporulosum [73] Xylaria cubensis [54] (3R,4S)-4-hydroxymellein (20, Figure 2)…”

Agricultural and Other Biotechnological Applications Resulting from Trophic Plant-Endophyte Interactions

“…Table 4 presents a summary of the most popular bioactive compounds synthesized by endophytic microbiota. Usually, cytotoxic activity [129][130][131] followed by anticancer and antibacterial activities [132][133][134], antiviral effects [135], and protective impact against pathogens [134] were detected. Many such cases have been documented in literature older than from the last two years are included in Table 4.…”