Methanol extract and fraction of Anchomanes difformis root tuber modulate liver mitochondrial membrane permeability transition pore opening in rats

Document Type : Original Research Article


1 Laboratories for Biomembrane Research and Biotechnology, Department of Biochemistry, Faculty of Basic Medical Sciences, College of Medicine, University o Ibadan, Nigeria

2 Laboratories for Biomembrane Research and Biotechnology, Department of Biochemistry, Faculty of Basic Medical Sciences, College of Medicine, University of Ibadan, Nigeria.

3 Laboratories for Biomembrane Research and Biotechnology, Department of Biochemistry, Faculty of Basic Medical Sciences, College of Medicine, University of Ibadan, Nigeria


Objective: Extracts of Anchomanes difformis (AD) are used in folkloric medicine to treat several diseases and infections. However, their roles in mitochondrial permeability transition pore opening are not known.
Material and Methods: The viability of mitochondria isolated from Wistar rat liver used in this experiment, was assessed by monitoring their swelling amplitude in the absence of calcium and reversal of calcium-induced pore opening by spermine. The effects of methanol extract and fraction of A. difformis (MEAD and MFAD, respectively) on Mitochondrial Membrane Permeability Transition (MMPT) pore opening, ATPase activity, cytochrome c release and ferrous-induced lipid peroxidation were assessed spectrophotometrically. Phytochemical constituents of MEAD and MFAD were assessed using Gas Chromatography- Mass Spectrometry (GC-MS).
Results: The MEAD (10, 20, 40 and 80 μg/ ml) had no effect on MMPT pore opening in the absence of Ca2+, whereas MFAD at 80 μg/ml had a large amplitude pore opening effect. Both MEAD and MFAD reversed Ca2+‌‌-induced swelling with inhibition values of 18, 21, 24, 23% (for MEAD) and 41, 36, 35, and 26% (for MFAD) at 10, 20, 40 and 80 μg/ml, respectively. MFAD significantly enhanced F1F0 ATPase activity and caused cytochrome c release. Both MEAD and MFAD significantly inhibited ferrous-induced lipid peroxidation by 33.0, 64.0, 66, and 75% (for MEAD) and 24, 25, 30, and 45% (for MFAD), respectively. The GC-MS results revealed the presence of squalene as one of the major constituents of MEAD.
Conclusion: These findings suggest that MFAD can be used to induce cell death via mitochondrial permeability transition in isolated rat liver. Inhibition of lipid peroxidation by MEAD and MFAD showed that the pore opening effect of the extract and fraction was not mediated via peroxidation of mitochondrial membrane lipids.


Main Subjects

Adnan SNA, Ibrahim N, Yaacob WA. 2017. Disruption of methicillin-resistant Staphylococcus aureus protein synthesis by tannins. GERMS, 7: 186-192.
Alabi TD, Brooks N, Oguntibeju O. Medicinal activities of Anchomanes difformis and its potential in the treatment of diabetes mellitus and other diseases conditions: A review. In: The potency of medicinal plants in the treatment and management of Diabetes mellitus. Apple Academy Press. First Edition Chapter 9, pp 219-235.
Ankur R, Arti M, Seema R, Amarjeet D, Ashok K. 2012. Mitochondrial permeability transition pore: another review. Int Res J Pharm, 3: 106-108.
Appaix F, Minatchy MN, Riva-Lavieille C, Olivares J, Antonsson B, Saks VA. 2000. Rapid spectrophotometric method for quantitation of cytochrome c release from isolated mitochondria or permeabilized cells revisited. Biochim Biophys Acta, 1457: 175-181.
Ataman JE, Idu M. 2015. Renal effects of Anchomanes difformis crude extract in Wistar rats. Avicenna J Phytomed, 5: 17-25.
Baev AY, Negoda A, Abramov AY. 2017. Modulation of mitochondrial ion transport by inorganic polyphosphate- essential role in mitochondrial permeability transition pore. J Bioenerg Biomembr. 49: 49-55
Baines CP, Kaiser RA, Sheiko T, Craigen WJ, Molkentin JD. 2007. Voltage-dependent anion channels are dispensable for mitochondrial-dependent cell death. Nat Cell Biol, 9: 550–555.
Bero J, Hannaert V, Chataigné G, Hérent MF, Quetin-Leclercq J. 2009. In vitro antiplasmodial activity of plants used in Benin in traditional medicine to treat malaria. J Ethnopharmacol, 122: 432-444.
Bonora M, Pinton P. 2014. Shedding light on molecular mechanisms and identity of MPTP. Mitochondrion, 21:11.
Carreras MC, Franco MC, Peralta JG, Poderoso JJ. 2004. Nitric oxide, complex I, and the modulation of mitochondrial reactive species in biology and disease. Mol Aspects Med, 25: 125–39.
Evans JL, Goldfine ID, Maddux BA, Grodsky GM. 2002. Oxidative stress and stress-activated signaling pathways: a unifying hypothesis of Type 2 diabetes. Endocr Rev, 23: 599–622.
Geryani MA, Mahdian D, Mousavi SH, Hosseini A. 2016. Cytotoxic and apoptogenic effects of Perovskia abrotanoides flower extract on MCF-7 and HeLa cell lines. Av J Phytomed, 6: 410-417.
Handal SS, Khanuja SPS, Longo G, Rakesh DD. 2008. An overview of extraction techniques for medicinal and aromatic plants. In: Extraction technologies for medicinal and aromatic plants. International centre for science and high technologies, Trieste, pp 21-54.
Harper JF, Breton G, Harmon A. 2004. Decoding Ca2+ signals through plant protein kinases. Ann Rev Pl Physiol Pl Mol Biol, 55: 263–288.
Johnson D, Lardy H. 1967. Methods Enzymol, 10: 94-96.
Kim SK, Faith K. 2012. Biological importance and application of squalene and squalane. Adv. in Food Nutr Res, 65: 223-233.
Kinnally KW, Antonsson B. 2007. A tale of two mitochondrial channels, MAC and PTP, in apoptosis. Apoptosis, 12: 857-868.
Kowaltowski AJ, Castilho RF, Grijalba MT, Bechara EJH, Vercesi AE. 1996. Effect of inorganic phosphate concentration on the nature of inner mitochondrial membrane alterations mediated by Ca21 ions. J Biol Chem, 271: 2929-2934.
Kroemer G, Reed JC. 2000. Mitochondrial control of cell death. Nat Med, 6: 513-519.
Lapidus RG, Sokolove PM. 1993. Spermine inhibition of the permeability transition of isolated rat liver mitochondria: An investigation of mechanism. J Biochem Biophys Met, 64: 246-253.
Lardy HA, Wellman H. 1953. The catalyst effects of 2, 4-dinitrophenol on adenosine triphosphatase hydrolysis by cell particles and soluble enzymes. J Biol Chem, 201: 357-370.
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. 1951. Protein measurement with the Folin phenol reagent. J Biol Chem,193: 262-275.
Martin KR. 2006. Targeting apoptosis with dietary bioactive agents. Soc Exp Biol Med, 23: 117-129.
Moradzadeh M, Hosseini A, Rakhshandeh H, Aghaei A, Sadeghnia HR. 2018. Cuscuta campestris induces apoptosis by increasing reactive oxygen species generation in human leukemic cells. Av J Phytomed, 8: 237-245.
Motaez M, Emami SA, Najjaran ZT. 2015. Growth inhibition and apoptosis induction of Scutellaria luteo-coeruleaBornm. & Sint.on leukemia cancer cell lines K562 and HL-60. Av J Phytomed, 5: 553-559.
National Institute of Health, NIH. 2002. Animals in Research", public health service policy on Human Care and Use of Laboratory Animals, Health Research Extension Act of 1985, Public Law 99-158, reprinted 2002.
Nelson DL, Cox MM, Freeman WH. 2005. Lehninger principles of biochemistry.4th Ed. New York: 2005.pp 485-519.
Nguyen TT, Quan X, Hwang KH, Xu S, Das R, Choi SK, Wiederkehr A, Wollheim CB, Cha SK, Park KS. 2015. Mitochondrial oxidative stress mediates high-phosphate-induced secretory defects and apoptosis in insulin-secreting cells. Am J Physiol Endocrinol Metab, 308: E933-941.
Noumi E. 2010. Ethno medicines used for treatment of prostatic disease in Foumban, Cameroon. Afr J Pharm Pharmacol, 4: 793-805.
Oghale OV, Idu M. 2016. Phytochemistry, anti-asthmatic and antioxidant activities of Anchomanes difformis (Blume) Engl. leaf extract. Asian Pacific J Trop Med, 6: 225-231.
Pal D, Mishra P, Sachan N, Ghosh AK. 2011. Biological activities and medicinal properties of Cajanus cajan (L) Millsp. J Adv Pharm Technol Res, 2: 207-214.
Ruberto G, Baratta MT, Deans SG, Dorman HJD. 2000. Antioxidant and antimicrobial activity of Foeniculum vulgare and Crithmum maritimumessential oils. Planta Med, 66: 687-693.
Sánchez E, García S, Heredia N. 2010. Extracts of edible and medicinal plants damage membranes of Vibrio cholera. Appl Environ Microbiol, 76: 6888-6894.
Seidlmayer LK, Juettner VV, Kettlewell S, Pavlov EV, Blatter LA, Dedkova EN. 2015. Distinct mPTP activation mechanisms in ischaemia-reperfusion: contributions of Ca2+, ROS, pH, and inorganic polyphosphate. Cardiovasc Res, 106: 237-248.
Shigenaga M, Hagen T, Ames B. 1994. Oxidative damage and mitochondrial decay in aging. PNAS, 91: 10771-10778.
Wiench B, Eichhorn T, Paulsen M, Efferth T. 2012. Shikonin directly targets mitochondria and causes mitochondrial dysfunction in cancer cells. Evidence-Based Complemen Alt Med, 2012: 726025 eCAM.