In vitro antiplasmodial activities and synergistic combinations of differential solvent extracts of the polyherbal product, Nefang

Protus Arrey Tarkang, Kathrin Diehl Franzoi, Sukjun Lee, Eunyoung Lee, Diego Vivarelli, Lucio Freitas-Junior, Michel Liuzzi, Tsabang Nolé, Lawrence S Ayong, Gabriel A Agbor, Faith A Okalebo, Anastasia N Guantai, Protus Arrey Tarkang, Kathrin Diehl Franzoi, Sukjun Lee, Eunyoung Lee, Diego Vivarelli, Lucio Freitas-Junior, Michel Liuzzi, Tsabang Nolé, Lawrence S Ayong, Gabriel A Agbor, Faith A Okalebo, Anastasia N Guantai

Abstract

Nefang, a polyherbal product composed of Mangifera indica (bark and leaf), Psidium guajava, Carica papaya, Cymbopogon citratus, Citrus sinensis, and Ocimum gratissimum (leaves), is a potential therapy against P. falciparum malaria. In vitro antiplasmodial activities of its constituent solvent extracts were analyzed on CQ-sensitive (3D7) and multidrug resistant (Dd2) P. falciparum strains. The interactions involving the differential solvent extracts were further analyzed using a variable potency ratio drug combination approach. Effective concentration 50 (EC50) values were determined by nonlinear regression curve-fitting of the dose-response data and used in calculating the fractional inhibitory concentration 50 (FIC50) and combination indices (CI) for each pair. The derived EC50 values (3D7/Dd2, μ g/mL) are Nefang-96.96/55.08, MiB-65.33/34.58, MiL-82.56/40.04, Pg-47.02/25.79, Cp-1188/317.5, Cc-723.3/141, Cs-184.4/105.1, and Og-778.5/118.9. Synergism was obtained with MiB/Pg (CI = 0.351), MiL/Pg (0.358), MiB/Cs (0.366), MiL/Cs (0.482), Pg/Cs (0.483), and Cs/Og (0.414) when analyzed at equipotency ratios. Cytotoxicity testing of Nefang and the solvent extracts on two human cell lines (Hep G2 and U2OS) revealed no significant toxicity relative to their antiplasmodial activities (SI > 20). Taken together, our data confirm the antimalarial activities of Nefang and its constituent plant extracts and identified extract pairs with promising synergistic interactions for exploitation towards a rational phytotherapeutic and evidence-based antimalarial drug discovery.

Figures

Figure 1
Figure 1
Isobolograms of the in vitro interactions between differential solvent extracts of Nefang at variable potency ratios. (a): Control CQ/CQ; (b): Control ART/CQ; (c): Cp/Og-EtOH; (d): MiB/Pg-Aq; (e): MiB/Cs- Aq; (f): Cp/Cs-Aq. CQ: Chloroquine; ART: Artemisinin; FIC50 = Fractional Inhibitory Concentration 50. A concave isobologram is consistent with synergy, a convex one is consistent with antagonism, and a straight line is consistent with additivity. Axes are EC50s normalized to 1.

References

    1. Stojanoski N. Development of health culture in Veles and its region from the past to the end of the 20th century. Veles: Society of Science and Art. 1999:13–34.
    1. WHO/TDR News no. 79, pp. 8–13, 2007.
    1. Dahanukar SA, Kulkarni RA, Rege NN. Pharmacology of medicinal plants and natural products. Indian Journal of Pharmacology. 2000;32(4):S81–S118.
    1. Petrovska BB. Historical review of medicinal plants usage. Pharmacognosy Review. 2012;6(11):1–5.
    1. McChesney JD, Venkataraman SK, Henri JT. Plant natural products: back to the future or into extinction? Phytochemistry. 2007;68(14):2015–2022.
    1. Newman DJ, Cragg GM, Snader KM. Natural products as sources of new drugs over the period 1981–2002. Journal of Natural Products. 2003;66(7):1022–1037.
    1. Vaghasiya YK, Shukla VJ, Chanda SV. Acute oral toxicity study of Pluchea arguta boiss extract in mice. Journal of Pharmacology and Toxicology. 2010;6(2):113–123.
    1. Kelly K. History of Medicine. New York, NY, USA: Facts on file; 2009.
    1. Hopkins AL. Network pharmacology. Nature Biotechnology. 2007;25(10):1110–1111.
    1. Hopkins AL. Network pharmacology: the next paradigm in drug discovery. Nature Chemical Biology. 2008;4(11):682–690.
    1. World Health Organisation (WHO) WPR/RC52/7: A Draft Regional Strategy for Traditional Medicine in Western Pacific. WHO Regional Committee, 52nd session Darussalam, 10–14 September 2001.
    1. Adjanohoun JE, Aboubakar N, Dramane K, et al. Contribution to Ethnobotanical and Floristic Studies in Cameroon. Addis Ababa, Ethiopia: Scientific, Technical and Research Commission, Organization of African Unity; 1996.
    1. Tarkang PA, Okalebo FA, Agbor GA, Tsabang N, Guantai AN, Rukunga GM. Indigenous knowledge and folk use of a polyherbal antimalarial by the Bayang Community, South West Region of Cameroon. Journal of Natural Product and Plant Resources. 2012;2(3):372–380.
    1. Tarkang PA, Atchan APN, Kuiate J, Okalebo FA, Guantai AN, Agbor GA. Antioxidant potential of a polyherbal antimalarial as an indicator of its therapeutic value. Advances in Pharmacological Sciences. 2013;2013:9 pages.678458
    1. Organisation Mondiale de la Santé (OMS) Paludisme. Aide mémoire No 94. WHO Media Centre, Geneva, Switzerland, 2007.
    1. Fidock DA, Rosenthal PJ, Croft SL, Brun R, Nwaka S. Antimalarial drug discovery: efficacy models for compound screening. Nature Reviews Drug Discovery. 2004;3(6):509–520.
    1. White NJ, Olliaro PL. Strategies for the prevention of antimalarial drug resistance: rationale for combination chemotherapy for malaria. Parasitology Today. 1996;12(10):399–401.
    1. Wagner H, Ulrich-Merzenich G. Synergy research: approaching a new generation of phytopharmaceuticals. Phytomedicine. 2009;16(2-3):97–110.
    1. Emanuel EJ, Wendler D, Killen J, Grady C. What makes clinical research in developing countries ethical? The benchmarks of ethical research. Journal of Infectious Diseases. 2004;189(5):930–937.
    1. Butterweck V, Nahrstedt A. What is the best strategy for pre-clinical testing of botanicals: a Critical perspective. Planta Medica. 2012;78(8):747–754.
    1. Patwardhan B. Drug discovery and development: traditional medicine and ethnopharmacology perspectives. SciTopics. 2009
    1. Handa SS, Khanuja SPS, Longo G, Rakesh DD. Extraction Technologies for Medicinal and Aromatic Plants. Trieste, Italy: ICS-UNIDO International Centre for Science and High Technology; 2008.
    1. Odebiyi OO, Sofowora EA. Phytochemical screening of Nigerian medicinal plants II. Lloydia. 1978;41(3):234–246.
    1. Harborne JB. Phytochemical Methods. 2nd edition. London, UK: Chapman and Hall; 1973.
    1. Ahmed SA, Gogal RM, Jr., Walsh JE. A new rapid and simple non-radioactive assay to monitor and determine the proliferation of lymphocytes: an alternative to [3H]thymidine incorporation assay. Journal of Immunological Methods. 1994;170(2):211–224.
    1. Nociari MM, Shalev A, Benias P, Russo C. A novel one-step, highly sensitive fluorometric assay to evaluate cell- mediated cytotoxicity. Journal of Immunological Methods. 1998;213(2):157–167.
    1. Brandão MGL, Krettli AU, Soares LSR, Nery CGC, Marinuzzi HC. Antimalarial activity of extracts and fractions from Bidens pilosa and other Bidens species (Asteraceae) correlated with the presence of acetylene and flavonoid compounds. Journal of Ethnopharmacology. 1997;57(2):131–138.
    1. Trager W, Jensen JB. Human malaria parasites in continuous culture. Science. 1976;193(4254):673–675.
    1. Lambros C, Vanderberg JP. Synchronization of Plasmodium falciparum erythrocytic stages in culture. Journal of Parasitology. 1979;65(3):418–420.
    1. Izumiyama S, Omura M, Takasaki T, Ohmae H, Asahi H. Plasmodium falciparum: development and validation of a measure of intraerythrocytic growth using SYBR Green I in a flow cytometer. Experimental Parasitology. 2009;121(2):144–150.
    1. Ohrt C, Willingmyre GD, Lee P, Knirsch C, Milhous W. Assessment of azithromycin in combination with other antimalarial drugs against Plasmodium falciparum in vitro . Antimicrobial Agents and Chemotherapy. 2002;46(8):2518–2524.
    1. Benoit-Vical F, Valentin A, Mallié M, Bastide JM, Bessière JM. In vitro antimalarial activity and cytotoxicity of Cochlospermum tinctorium and C. planchonii leaf extracts and essential oils. Planta Medica. 1999;65(4):378–381.
    1. Berenbaum MC. A method for testing for synergy with any number of agents. Journal of Infectious Diseases. 1978;137(2):122–130.
    1. Saiman L, Mehar F, Niu WW, et al. Antibiotic susceptibility of multiply resistant Pseudomonas aeruginosa isolated from patients with cystic fibrosis, including candidates for transplantation. Clinical Infectious Diseases. 1996;23(3):532–537.
    1. Ogbe RJ, Adenkola AY, Anefu E. Aqueous ethanolic extract of Mangifera indica stem bark effect on the biochemical and haematological parameters of albino rats. Archives of Applied Science Research. 2012;4(4):1618–1622.
    1. Morsi RMY, El-Tahan NR, El-Hadad AMA. Effect of aqueous extract Mangifera indica leaves, as functional foods. Journal of Applied Sciences Research. 2010;6(6):712–721.
    1. Uboh FE, Okon IE, Ekong MB. Effect of aqueous extract of Psidium Guajava leaves on liver enzymes, histological integrity and hematological indices in rats. Gastroenterology Research. 2010;3(1):32–38.
    1. Tarkang PA, Agbor GA, Tchamgoue DA, Tchokouaha LRY, Kemeta D, Mengue YSN. Acute and chronic toxicity studies of the aqueous and ethanol leaf extracts of carica papaya linn in wistar rats. Journal of Natural Product Plant Resources. 2012;2(5):617–627.
    1. Tarkang PA, Agbor GA, Tsabang N, et al. Effect of long-term oral administration of the aqueous and ethanol leaf extracts of Cymbopogon citratus (DC. ex Nees) stapf. Annals of Biological Research. 2012;3(12):5561–55570.
    1. Tarkang PA, Agbor GA, Tchamgoue DA, Tchokouaha LRY, Kemeta D, Mengue YSN. Acute and sub-chronic toxicity studies of the aqueous and ethanol leaf extracts of Citrus sinensis (Linnaeus) osbeck (pro sp.) in wistar rats. Der Pharmacia Lettre. 2012;4(5):1619–1629.
    1. Ajibade AJ, Fakunle PB, Ehigie LO, Akinrinmade AO. Sub-chronic hepatotoxicity in adult wistar rats following administration of Ocimum gratissimum aqueous extract. European Journal of Medicinal Plants. 2012;2(1):19–30.
    1. Philippe G, Angenot L, De Mol P, et al. In vitro screening of some Strychnos species for antiplasmodial activity. Journal of Ethnopharmacology. 2005;97(3):535–539.
    1. Pink R, Hudson A, Mouriès M-A, Bendig M. Opportunities and challenges in antiparasitic drug discovery. Nature Reviews Drug Discovery. 2005;4(9):727–740.
    1. Bidla G, Titanji VPK, Joko B, El-Ghazali G, Bolad A, Berzins K. Antiplasmodial activity of seven plants used in African folk medicine. Indian Journal of Pharmacology. 2004;36(4):245–246.
    1. Ngemenya MN, Tane P, Berzins K, Titanji VPK. Antiplasmodial activity of some medicinal plants used in cameroon: preliminary toxicity studies of highly active extracts. Proceedings of the 11th Annual Conference of The Cameroon Bioscience Society; December 2004.
    1. Nundkumar N, Ojewole JAO. Studies on the antiplasmodial properties of some South African medicinal plants used as antimalarial remedies in zulu folk medicine. Methods and Findings in Experimental and Clinical Pharmacology. 2002;24(7):397–401.
    1. Oliveira AB, Dolabela MF, Braga FC, Jácome RLRP, Varotti FP, Póvoa MM. Plant-derived antimalarial agents: new leads and efficient phythomedicines. Part I. alkaloids. Anais da Academia Brasileira de Ciencias. 2009;81(4):715–740.
    1. Percario S, Moreira DR, Gomes BAQ, et al. Oxidative stress in malaria. International Journal of Molecular Sciences. 13(12):16346–16372.
    1. Batista R, de Jesus Silva Júnior A, de Oliveira AB. Plant-derived antimalarial agents: new leads and efficient phytomedicines. part II. non-alkaloidal natural products. Molecules. 2009;14(8):3037–3072.
    1. Benoit-Vical F, Imbert C, Bonfils J-P, Sauvaire Y. Antiplasmodial and antifungal activities of iridal, a plant triterpenoid. Phytochemistry. 2003;62(5):747–751.
    1. Elford BC. L-Glutamine influx in malaria-infected erythrocytes: a target for antimalarials? Parasitology Today. 1986;2(11):309–312.
    1. Kirmizibekmez H, Çalis I, Perozzo R, et al. Inhibiting activities of the secondary metabolites of Phlomis brunneogaleata against parasitic protozoa and plasmodial enoyl-ACP reductase, a crucial enzyme in fatty acid biosynthesis. Planta Medica. 2004;70(8):711–717.
    1. Tasdemir D, Güner ND, Perozzo R, et al. Anti-protozoal and plasmodial FabI enzyme inhibiting metabolites of Scrophularia lepidota roots. Phytochemistry. 2005;66(3):355–362.
    1. Stevenson MM, Riley EM. Innate immunity to malaria. Nature Reviews Immunology. 2004;4(3):169–180.
    1. Mojab F. Antimalarial natural products: a review. Avicenna Journal of Phytomedicine. 2012;2(2):52–62.
    1. Azas N, Laurencin N, Delmas F, et al. Synergistic in vitro antimalarial activity of plant extracts used as traditional herbal remedies in Mali. Parasitology Research. 2002;88(2):165–171.
    1. Gathirwa JW, Rukunga GM, Njagi ENM, et al. The in vitro anti-plasmodial and in vivo anti-malarial efficacy of combinations of some medicinal plants used traditionally for treatment of malaria by the Meru community in Kenya. Journal of Ethnopharmacology. 2008;115(2):223–231.
    1. van Vuuren S, Viljoen A. Plant-based antimicrobial studies methods and approaches to study the interaction between natural products. Planta Medica. 2011;77(11):1168–1182.

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