Bisbenzylisoquinoline (BBIQ) alkaloids are a diverse group of natural products that demonstrate a range of biological activities. In this study, the in vitro antiplasmodial activity of three BBIQ alkaloids (cycleanine [compound 1], isochondodendrine [compound 2], and 2′-norcocsuline [compound 3]) isolated from the Triclisia subcordata Oliv. medicinal plant traditionally used for the treatment of malaria in Nigeria are studied alongside two semisynthetic analogues (compounds 4 and 5) of cycleanine.

Background Cylicodiscus gabunensis Harms (Family Leguminosae ) (CG) is an African medicinal plant used as a treatment of various ailments including malaria, liver diseases, and gastrointestinal disturbances. Its extracts showed potent in vitro antibacterial activity. However, the antibacterial components are unknown. Methods In this study, the stem bark of the CG plant was extracted and its antibacterial property against a panel of Gram-negative and Gram-positive bacterial strains assessed using the disk diffusion assay method. Bioassay-guided fractionation of the bioactive extracts was employed to identify bioactive constituents using both gas and liquid chromatography mass spectrometry. Chemical synthesis was used to make the analogues of gallic acid. Microplate dilution assays and scanning electron microscopy (SEM) were used to evaluate the antibacterial properties and mechanism of action of the active fractions and pure compounds. Results The most bioactive sub-fractions derived from CG comprised of ethyl gallate, gallic acid and polyphenols. Five alkyl/alkenyl gallates were synthesized. A preliminary structure-activity relationship of gallic acid derivatives was obtained using the synthetic analogues and a series of commercially available phenolic compounds. Increasing the length of alkyl chains generally increases the potency of the alkyl gallates. Introducing a double bond with restricted conformations of the C-5 side chain has little effect on the antibacterial property. SEM analysis of the effect of alkyl gallates on Staphylococcus aureus indicates that they appear to interrupt S. aureus bacterial cell wall integrity. Conclusions The results of this research rationalise the ethnobotanical use of C. gabunensis and suggest that gallate derivatives may serve as promising antibacterial agents for the treatment of infectious diseases.

Modification of metal surfaces with antimicrobial peptides is a promising approach to reduce bacterial adhesion. Here, cyclic peptides or cycloids, possessing remarkable stability and antimicrobial activities, were extracted and purified from Viola philippica Cav., and identified using mass spectrometry. Cyclotides were subsequently utilized to modify stainless steel surfaces via polydopamine-mediated coupling. The resulting cyclotide-modified surfaces were characterized by Fourier transform infrared (FTIR) spectroscopy and contact angle analysis. The antibacterial capacity of these cyclotides against Staphylococcus aureus was assessed by Alamar blue assay. The antibiofilm capacity of the modified surfaces was assessed by crystal violet assay, and scanning electron microscopy (SEM). A composite of Kalata b1, Varv A, Viba 15 and Viba 17 (P1); Varv E (P2); and Viphi G (P3) were isolated and identified. FTIR analysis of the modified surfaces demonstrated that cyclotides bound to the surfaces and induced reduction of contact angles. Antimicrobial effects showed an order P3 > P1 and P2, with P3-treated surfaces demonstrating the strongest antibiofilm capacity. SEM confirmed reduced biofilm formation for P3-treated surfaces. This study provides novel evidence for cyclotides as a new class for development of antibacterial and antibiofilm agents.

Our previous studies demonstrated that cycleanine, a macrocyclic bisbenzylisoquinoline (BBIQ) alkaloid, showed potent anti-ovarian cancer activity via apoptosis induction. Here, we synthesized two novel (aminoalkyl)cycleanine analogues (2 and 3) through a simple and efficient two-step reaction starting from cycleanine isolated from Triclisia subcordata Oliv. These analogues showed greater potency than the unmodified cycleanine in three human ovarian cancer cell lines. Both 2 and 3 induced apoptosis in ovarian cancer cells by activations of caspases 3/7, cleavage of PARP, increase in subG1 cell cycle phase and in the percentage of apoptotic cells. Further confocal fluorescence microscopy analysis confirmed the cellular uptake of alkaloids in ovarian cancer cells by using the unique (alkynyl)cycleanine (3) via click chemistry reaction. Our results suggest that cycleanine could be a hit compound for the future development in attacking ovarian cancer.

Ethnopharmacological relevance and aim: A decoction of the bark of Cylicodiscus gabunensis Harms is used as a traditional medicine in the treatment of malaria in Nigeria. This study aims to validate the antimalarial potency of this decoction in vitro against Plasmodium falciparum and define potential bioactive constituents within the C. gabunensis bark.Materials and methods: A bioassay-guided separation and fractionation protocol was applied to C. gabunensis extracts, exploiting the use of a Malaria Sybr Green I Fluorescence assay method to monitor antiproliferative effects on parasites as well as define 50% inhibition concentrations. Spectroscopic techniques, including GC-MS, TOF LC-MS and H-1 NMR were used to identify phytochemicals present in bioactive fractions. Analogues of gallic acid were synthesized de novo to support the demonstration of the antimalarial action of phenolic acids identified in C. gabunensis bark. In vitro cytotoxicity of plant extracts, fractions and gallate analogues was evaluated against the HepG2 cell line.Results: The antimalarial activity of ethanolic extracts of C. gabunensis bark was confirmed in vitro, with evidence for phenolic acids, primarily gallic acid and close analogues such as ethyl gallate, likely providing this effect. Further fractionation produced the most potent fraction with a 50% inhibitory concentration of 4.714/ml. Spectroscopic analysis, including H-1 NMR, LC-MS and GC-MS analysis of this fraction and its acid hydrolyzed products, indicated the presence of conjugates of gallic acid with oligosaccharides. The extracts/fractions and synthetic alkyl and alkenyl gallates showed moderate selectivity against P. falciparum.Conclusions: These results support the use of the bark of C. gabunensis as a traditional medicine in the treatment of human malaria, with phenolic acid oligosaccharide complexes evident in the most bioactive fractions.

Triclisia subcordata Oliv (Menispermeaceae) is an African medicinal plant traditionally used for the treatment of various diseases, including cancer. This study aimed to isolate and identify minor alkaloids present in this plant and assay their anticancer activities. Isochondodendrine (1) and 2'-norcocsuline (2) as two minor alkaloids together with the abundant cycleanine (3) [1] were isolated and identified by mass spectrometry and nuclear magnetic resonance spectroscopy. Both isochondodendrine and 2'-norcocsuline exhibited potent in vitro cytotoxicity in four ovarian cancer cell lines (A2780, Igrov-1, Ovcar-8, and Ovcar-4) with an IC50 range of 3.5–17 μM and 0.8–2.9 μM (assessed via sulforhodamine B dye assay), respectively. The IC50 in cell growth assays using normal human ovarian epithelial cells were 10.5 ± 1.2 μM and 8.0 ± 0.2 μM for isochondodendrine and 2'-norcocsuline, respectively. These alkaloids showed greater potencies for the cancer cells compared to normal cells. Apoptosis induction by these alkaloids was studied by caspase activity assay, western blot, and flow cytometry. They induced apoptosis in ovarian cancer cells by activation of caspases 3/7, cleavage of PARP, an increase in subG1 cell cycle phase and increases in both early and late apoptotic cells. Therefore isochondodendrine and 2'-norcocsuline are among the least abundant in T. subcordata, which also contribute to its cytotoxic activity and can be potential hit compounds for future development for the treatment of ovarian cancer.

Triclisia subcordata Oliv (Menispermeaceae) is used in herbal medicine for the treatment of cancer and other diseases in Africa.

Ovarian cancer remains one of the main causes of death in all gynecologic malignancies [1]. Natural products continue to be important sources of clinically approved anti-cancer drugs [2, 3]. Triclisia subcordata Oliv (Menispermeaceae) is a medicinal plant traditionally used for the treatment of various diseases [4], including cancer, in West Africa. This study aims to evaluate the in vitro anti-ovarian cancer activities of the crude extracts and the isolated components in T. subcordata. The ethanol extract of T. subcordata and its fractions (crude alkaloids) were screened for in vitro anti-ovarian cancer activities on Ovcar-8, Ovcar-4, A2780, and Igrov-1 ovarian cancer cell lines using the Sulforhodamine B assay method to measure cell growth. Bioassay-guided fractionation using silica gel column chromatography and HPLC were used to isolate the bioactive compound, whose identity and structure was identified by NMR and LC-MS techniques. Caspase and PARP cleavage assays were used to detect apoptotic activities. The effect of isolated pure compounds on cell cycle and apoptosis was analyzed by flow cytometry. Results: The crude alkaloids showed the strongest activity in cell growth assays on A2780 and Ovcar-8 cell lines (IC50 < 2.4 µg/mL). A bisbenzylisoquinoline alkaloid-cycleanine was isolated using HPLC and identified by MS and NMR analyses. The IC50values of cycleanine and tetrandrine ranged from 7 to 14µM on A2780, Ovcar-8, Ovcar-4 and Igrov-1 ovarian cancer cell lines. The IC50 of cycleanine on human normal ovarian surface epithelial cells was 35 ± 1µM hinting at modest selectivity towards cancer cells. Both cycleanine and tetrandrine caused apoptosis as shown by activation of caspases 3/7 and cleavage of poly (ADP) ribose polymerase (PARP) to form PARP-I. The percentage of Ovcar-8 cells in subG1 phase increased after exposure of cycleanine and tetrandrine to cells for 48h compared to control. In conclusion, cycleanine, like its isomer – tetrandrine isolated from T. subcordata, could be a potential new anti-ovarian cancer agent acting through the apoptosis pathway.

Ovarian cancer is the leading cause of death in the gynaecologic cancers within the UK and US. Presently the standard treatment for ovarian cancer entails the use of chemotherapy drugs paclitaxel and carboplatin after aggressive surgical reduction in order to prolong the patient’s life for multiple years. However, prolonged use of platinum-based chemotherapy often leads to drug resistance, which causes the ovarian cancer patient to relapse and potential death. Therefore there is an urgent medical need for breakthrough drugs with an effective therapeutic impact on ovarian cancer. Phytochemicals (plant-derived natural products) have been used for thousands of years as treatment for various diseases, because of their huge chemical diversity and wide range of biological activities. In this review, the role of phytochemicals as chemo-preventive compounds, potential sources of new drugs for ovarian cancer and the benefits of their adoption as monotherapeutic agents or as chemosensitizers when used in-conjunction with the conventional anti-cancer drugs is highlighted. We will describe the phytochemicals: 1) clinically approved drugs such as paclitaxel and camptothecin including its semi-synthetic derivatives topotecan and irinotecan; 2) currently in clinical trials such as epipodophyllotoxin derivatives etoposide and teniposide, ventfolide, phenoxodiol, and combretastatins; 3) in preclinical trials such as quercetin, baicalein, baicalin, thymoquinone, betulinic acid and tetrandrine; and recently discovered compounds which have high potency (IC50 less than 10 μM) and have been discovered recently (last 15 years). In particular, several new compounds including bufatrienolides, ipomoeassin D, 2'-(R)-O-acetylglaucarubinone, and molvizarin have IC50s lower than 100 nM in ovarian cancer cells and might have different mechanisms of action from those of platinum derivatives/paclitaxel, therefore providing potential ways to attack multidrug resistance in ovarian cancer without jeopardising the patient’s treatment.  Corresponding author: Dr. Wen-Wu Li. E-mail: w.li@keele.ac.uk Wen-Wu Li, Okiemute Rosa Johnson-Ajinwo, and Fidelia Ijeoma Uche 82

Triclisia subcordata Oliv (Menispermeaceae) is a medicinal plant traditionally used for the treatment of various diseases in West Africa. The ethanol extract of T. subcordata and its fractions were screened for in vitro anti-ovarian cancer activities using the Sulforhodamine B assay. The crude alkaloids showed the strongest activity in cell growth assays on Ovcar-8 and A2780 cell lines (IC50 < 2.4 mu g/mL). A bisbenzylisoquinoline alkaloid-cycleanine was isolated using HPLC and identified by mass spectrometry and nuclear magnetic resonance analyses. The IC50 values of cycleanine and tetrandrine (an alkaloid previously reported from this plant) ranged from 7 to 14 mu M on Ovcar-8, A2780, Ovcar-4, and Igrov-1 ovarian cancer cell lines. The IC50 of cycleanine on human normal ovarian surface epithelial cells was 35 +/- 1 mu M, hinting at modest selectivity toward cancer cells. Both cycleanine and tetrandrine caused apoptosis as shown by activation of caspases 3/7 and cleavage of poly(ADP-ribose) polymerase to form poly(ADP-ribose) polymerase-1 by using western blot analysis. Flow cytometry analyses showed that the percentages of apoptotic cells and cells in subG(1) phase increased after exposure of cycleanine and tetrandrine to Ovcar-8 cells for 48 h compared with control. Cycleanine, like its isomer tetrandrine isolated from T. subcordata, could be a potential new anti-ovarian cancer agent acting through the apoptosis pathway. Copyright (C) 2016 John Wiley & Sons, Ltd.