Abstract
The use of plants for pharmacological purposes is very old and evolves with the history of humanity. Medicinal plants have been used by different cultures for the management of different diseases for several centuries. According to the World Health Organization, about 80% of the world’s population relies on herbal products for their primary health care. Major classes of anticancer and antimicrobial drugs are based on natural products derived from plants and microorganisms. In addition, some of the drugs used in the treatment of various cancers, infections, inflammation, diabetes, and more are often natural products or their derivatives. For instance, between 1981 and 2014 over 50% of newly developed drugs or phytomedicines were from natural products. Facing the increase in mortality associated with drug-resistance of pathogens to the existing anticancer and antimicrobial drugs, there is an urgent need for the sourcing of new molecules that could lead to the formulation of new anticancer and antimicrobial drugs. It is in line with the above that this study aimed at investigating the phytochemical and pharmacological activities of some selected medicinal plants used in folk medicine for the treatment of cancer, diabetes, and microbial infections.
To achieve this, different parts (fruits, leaves, twigs, and stem bark) of Cola lateritia K. Schum., Tetrapleura tetraptera (Schumach. & Thonn.) Taub. and Anonidium mannii (Oliv.) Engl. & Diels were collected at mount Kala, a locality in the Central region of Cameroon. The plant materials were then prepared and extracted with different solvent systems affording different extracts. The isolation and purification of compounds were performed using different chromatographic techniques with different solvent systems and polarities. The structures of the isolated compounds were characterized and identified by means of spectroscopic and spectrometry techniques (1D and 2D NMR, HRESIMS, FT-IR). Furthermore, the cytotoxic potential of the pure isolates and the crude extracts was evaluated on prostate cancer cell lines (LNCaP, DU145 and PC3) using the MTT assay followed by the cell proliferation assay using the BrdU kit. Then, the mechanisms of action of compounds that have shown good cytotoxic potential were further tested for their impact on the cell cycle progression and the type of cell death (apoptosis, necrosis) induced using Flow cytometry. Cell adhesion, cell migration and chemotaxis studies were carried out through the wound-healing/scratch assay and Milli cell inserts respectively. Using Western blot assay, some proteins of the epithelial-mesenchymal transition (EMT) were assayed. The cytotoxic activities of the isolates were also tested on adenocarcinoma of the cervix (HeLa) cells and on breast cancer cell lines (MCF-7, MDA-MB-
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231 and Vero). In addition, all the extracts and pure compounds were evaluated for their antibacterial activity against a panel of bacteria strains including Gram-positive strains (Staphylococcus epidermidis, Staphylococcus aureus, Enterococcus feacalis, Mycobacterium smegmatis and Bacillus subtilis) and Gram-negative strains (Salmonella thyphimurium, Enterobacter cloacae, Escherichia coli, Klebsiella oxytoca, Proteus vulgaris, Klebsiella pneumoniae, Proteus mirabilis and Klebsiella aerogenes).
From the stem bark and twigs of Cola lateritia, eleven compounds belonging to the classes of triterpenoids, phytosteroids and fatty acids were isolated. Those compounds were characterized and identified as betulinic acid (1), ursolic acid (2), oleanolic acid (3), maslinic acid (4), arjunolic acid (5), lupenone (6), lupeol (7), ß-sitosterol-3-O-ß-D-glucoside (8), margaric acid (9), ß-sitosterol (10), and stigmasterol (11). All these compounds were isolated for the first time from C. lateritia.
A new glyceride, 2,3-dihydroxypropyl-31-hydroxyhentriacontanoate (1), a new glucoside stigmasterol derivative, stigmasterol 3-O-β-D-glucopyranoside 6'-hexadecanoate (2) as well as the known compounds 3-[(2-acetamido-2-deoxy-β-D-glucopyranosyl) oxy]olean-12-ene-28-oic acid (3), sucrose (4), pinitol (5), 4-O-α-D-Glucopyranosyl-D-glucopyranose (6), hexacosanoic acid (7), tetracosanoic acid (8), oleanolic acid (9), betulenic acid (10), stigmasterol (11), stigmasterol 3-O-β-D-glucoside (12) and daucosterol (13) were isolated from T. tetraptera fruits.
Studies of T. tetraptera stem bark led to the isolation of a new glyceride derivative, monoglyceryl-1-dotriacontanoate (1), triacont-1-ene (2), hexacosanoic acid (3), oleanane-3-O-β-D-glucoside-2'-acetamide (4), stigmasterol-3-O-β-D-glucoside (5), aridanin (6), 2,3-dihydroxypropyl 26-hydroxyhexacosanoate (7) and stigmasterol (8).
The study of A. mannii leaves extracts led to two new phytosteroids trivially called anonimadiol A (1), anonimadiol B (2) and stigmasterol (3). All the isolated compounds belong to the classes of triperpenoids, phytosteroids, fatty acids, sugars and glycerides.
Regarding the cytotoxic potential, compounds (3), and (4) from C. lateritia exhibited greater cytotoxic potential on prostate cancer cells with CC50 ranging between 17 and 25 μM. With T. tetraptera, compounds (4) and (6) showed cytotoxic potential with estimated CC50 of 15, 23, 16 and 17, 26, 16 μg/mL on DU145, PC3 and LNCaP cells, respectively and a potent cell growth arrest on the different cancer cell lines. After 72 h of treatment, (4) (20 μg/mL) showed a significant (p < 0.001) and concentration-dependent inhibition of DU145, PC3, LNCaP and
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HMEC cells growth. Regarding the formation of clones, a significant inhibition (p < 0.01) was observed with (3) and (4) at (2.5 μg/mL). Both compounds showed a significant inhibition (p < 0.05) of DU 145 cells proliferation. A 15% (DU145) and 25% (LNCaP) increase in apoptotic cells was observed with (4) and (6) at 10 μg/mL and a 20-25 % increase of apoptotic cells was observed after treatment with (3) and (4) isolated from C. lateritia at 2.5 and 10 μM. With regards to cell cycle progression, compounds (4) and (6) at 2.5 and 10 μg/mL induced an increase in the number of cells blocked in G2/M phase and a decrease in cells in the S-phase as does (4) (10 μM). With T. tetraptera, compound (4) significantly increased the cell adhesion to the fibronectin matrix and compound (6) increased the adherence of DU145 cells to collagen (2.5 μg/mL) and to fibronectin (2.5 and 10 μg/mL). This was accompanied by an increase in the expression of integrin β-1 at 10 μg/mL and integrin β-4 at 2.5 μg/mL. Furthermore, a down-regulation of pcdk1, cdk2, cyclin B, Bcl-2, N-cad, vimentin and cytokeratine 8-18 was noticed while, p19, p27, p53 pAKT, Bax, caspase-3 and E-cad were up-regulated. All these results suggest that both T. tetraptera and C. lateritia possess secondary metabolites with promising anticancer effects against prostate cancer. In addition, compounds 12-13 from T. tetraptera and the crude extract were cytotoxic against MDA-MB-231 cells with their CC50 values ranging between 14.5 and 20.0 μg/mL.
Regarding the antibacterial studies, all the tested samples exhibited significant antibacterial activity with MIC ranging from 18.5 to 588 μg/mL against all the strains used.