The Ethanolic Leaf Extracts of Dissotis multiflora (Sm) Triana and Paullinia pinnata Linn Exert Inhibitory Effect on Escherichia coli Through Membrane Permeabilization, Loss of Intracellular Material, and DNA Fragmentation
Abstract
Background: Dissotis multiflora (Sm) Triana and Paullinia pinnata Linn are widely used in Cameroonian traditional medicine to treat infectious diseases. These plants were found to be a reservoir of antioxidant and antimicrobial agents and have the potential to be used in clinic. Objective: To determine the mechanism of action of the ethanolic leaves extracts of Dissotis multiflora and Paullinia pinnata on Escherichia coli. Methodology: The microbroth dilution method was used to determine the minimum inhibitory concentrations (MICs) and the minimum bactericidal concentrations (MBCs) of D. multiflora and P. pinnata ethanolic leaves extracts. The above samples were tested for their rate of killing of E. coli cells at 1 MIC and 2 MICs. Sorbitol protection, outer membrane permeability, loss of 260-nm-absorbing material, fluorescence microscopy, and DNA degradation assay were used to examine the ultrastructural changes in bacteria induced by the extracts. Results: D. multiflora and P. pinnata extracts inhibited bacterial growth with MICs of 390.62 and 781.25 µg/mL respectively, while the MBCs values were found to be 781.25 and 1562.5 µg/mL respectively. Treatment with extracts had shorter kill-time in a time-dependent manner with effect most pronounced at 2 MICs than 1 MIC. The MIC of D. multiflora increased 4x from 390.62 µg/mL after 24 h of incubation to 1562.5 µg/mL after 7 days in the presence of an osmoprotectant indicating the inhibition of synthesis of cell wall constituents. P. pinnata had no effect on cell wall. Both extracts exhibited the greatest leakage and release of DNA materials at 30, 60, 90, and 120 min in concentration-dependent manner. Treated groups had higher values than control. At low concentrations (1/2 MIC and 1 MIC), these extracts effectively permeate the intact outer membrane of Gram-negative bacteria. Both extract were implicated in DNA fragmentation. Moreover, fluorescent cells observed further confirmed its inhibitory effect against the tested pathogen. The antibacterial action involved disruption of membrane potential, increase of membrane permeabilization, leakage of cellular material, and death suggesting them to be an alternative to antibiotics. Conclusion: These findings contribute to the understanding of the antibacterial inhibitory effect of D. multiflora and P. pinnata.
Keywords:
D. multiflora,, P. pinnata, Escherichia coli, Antibacterial activity, Membrane permeabilization, DNA Fragmentation, Ethanolic leaf extractsDOI
https://doi.org/10.22270/jddt.v11i2-S.4774References
Chitemerere TA, Mukanganyama S. Evaluation of cell membrane integrity as a potential antimicrobial target for plant products. BMC Complement Altern Med 2014; 14:278.
Mueller M, Tainter CR. Escherichia coli. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2021.
Bajpai VK, Shukla S, Paek WK, Lim J, Kumar P, Kumar P, et al. Efficacy of (C)-Lariciresinol to control bacterial growth of Staphylococcus aureus and Escherichia coli O157:H7. Front Microbiol 2017; 8:804.
Zhang J, Ye KP, Zhang X, Pan DD, Sun YY, Cao JX. Antibacterial Activity and mechanism of action of Black pepper essential oil on meat-borne Escherichia coli. Front Microbiol 2017; 7:2094.
Reduan FH, Shaari RM, Sayuti NS Acute and subacute dermal toxicity of ethanolic extract of Melastoma malabrathricum leaves in Sprague-Dawley rats. Toxicol Res 2020; 36:203-210.
Hana MA, Insha S, Vijay K, Irfan AR, Hashem AS, Arif TJ et al. Plant-based phytochemicals as possible alternative to antibiotics in combating bacterial drug resistance. Antibiotics 2020; 9:480.
Mohammed KO, Abdulrahman AA, Saud SA, Walid HS, Altaf A, Litaker MS et al. Antibacterial and antifungal activity of the extracts of different parts of Avicennia marina (Forssk.) Vierh. Plants 2021; 10:252.
Ting SU, Ye Q, Xuesi H, Bi Y, Haoming L, Da L et al. Novel opportunity to reverse antibiotic resistance: to explore traditional chineses medicine with potential activity against antibiotics-resistance bacteria. Front Microbiol 2020; 11:610070.
Bahman K, Milad I, Vahid S, Bibi SF. A review on antimicrobials: a mechanistic viewpoint. Antimicrob Resist Infect Control 2019; 8:118.
Matijaševi´ D, Panti´ M, Raškovi´ B, Pavlovi´ V, Duvnjak D, Sknepnek A, et al. The antibacterial activity of coriolus versicolor methanol extract and its effect on ultrastructural changes of Staphylococcus aureus and Salmonella Enteritidis. Front Microbiol 2016; 7:1226.
Collet JF, Cho SH, Iorga BI, Goemans CV. How the assembly and protection of the cell envelope depend on cysteine residues. Antimicrob. J Biol Chem 2020; 21:11984-11994.
Mani-Lopez E, Cortes-Zavaleta O,Lopez-Malo A. A review of the methods used to determine the target site or the mechanism of action of essential oils and their components against fungi. SN Appl Sci 2021; 3:44.
Lv F, Liang H, Yuan Q, Li C. In vitro antimicrobial effects and mechanism of action of selected plant essential oil combinations against four food-related microorganisms. Food Res Int 2011; 44:3057-3064.
Bajpai VK, Sharma A, Baek KH. Antibacterial mode of action of Cudrania tricuspidata fruit essential oil, affecting membrane permeability and surface characteristics of food-borne pathogens. Food Control 2013; 32:582-590.
Militello M, Settanni L, Aleo A, Mammina C, Moschetti G, Giammanco GM, et al. Chemical composition and antibacterial potential of Artemisia arborescens L. essential oil. Curr Microbiol 2011; 62:1274-1281.
Akinpelu DA, Alayande KA, Aiyegoro OA, Akinpelu OF, Okoh AI. Probable mechanisms of biocidal action of Cocos nucifera Husk extract and fractions on bacteria isolates. BMC Complement Altern Med 2015; 15:116.
Robert EM, Ngono BD, Auguste AA, Mbabi NN, Moni NE, Betote DP et al. Rauvolfianine, a new antimycobacterial glyceroglycolipid and other constituents from Rauvolfia caffra. Sond (Apocynaceae). Nat Prod Res 2017; 1-6.
Afagnigni AD, Nkonpa KR, Teugwa MC. Phytochemical analysis, antimicrobial, and radical scavenging properties of methanol extracts of Dracaena deisteliana (Dracaenaceae) and Sporobulus indicus (Poaceae). Global J Med Res 2020; 20(6): 41-50.
Yamani HA, Pang EC, Martin N, Deighton MA. Antimicrobacterial activity of Tulsi (Ocimum tenuiflorum) essential oil and their major constituents against three species of bacteria. Front Microbiol 2016; 7:681.
Saritha K, Rajesh A, Manjulatha K, Setty OH, Yenugu S. Mechanism of antibacterial action of the alcoholic extracts of Hemidesmus indicus (L.) R. Br. ex Schult, Leucas aspera (Wild.), Plumbago zeylanica L., and Tridax procumbens (L.) R. Br. ex Schult. Front Microbiol 2015; 6:577.
Afagnigni AD, Nyegue MA, Ndoye FC, Voundi SH, Fonkoua MC, Etoa F-X. Antibacterial and antioxidant activities of ethanolic leaves extracts of Dissotis multiflora Triana (Melastomataceae). Int J Pharm Sci Drug Res 2016; 8:50-56.
Afagnigni AD, Nyegue, MA, Ndoye, FC, Njankouo, NY, Njayou, FN, Fonkoua, MC et al. Antidiarrheal activity of Dissotis multiflora (Sm) Triana (Melastomataceae) leaf extract in Wistar rats and sub-acute toxicity evaluation. Evid Based Complement Alternat Med 2017; 1-9.
Afagnigni AD, Nyegue MA, Djova SV, Etoa FX. LC-MS analysis, 15-lipoxygenase inhibition, cytotoxicity, and genotoxicity of Dissotis multiflora (Sm) Triana (Melastomataceae) and Paullinia pinnata Linn (Sapindaceae). J Trop Med 2020; 1-8.
Lasisi A.A., Ayinde B.W., Adeleye A.O., Onocha P.A., Oladosu I.A., Idowu P.A. New triterpene isovanniloyl and antibacterial activity of constituents from the roots of Paullinia pinnata Linn (Sapindaceae). J Saudi Chem Soc 2011;19:117-122.
Lunga PK, Tamoukou JD, Fodouop SP, Kuiate JR, Gatsing TD. Antityphoid and radical scavenging properties of the methanol extracts and compounds from the aerial part of Paullinia pinnata Linn (Sapindaceae). Springerplus 2014; 3: 302.
Yusuf AZ, Zakir A, Shemau Z, Abdullahi M, Halima SA. Phytochemical analysis of the methanol leaves extracts of Paullinia pinnata Linn. J Pharmacognosy Phytother 2014; 6:10-16.
Jackson N, Annan K, Mensah AY, Ekuadzi E, Mensah ML, Habtemariam S. A novel triterpene from the roots of Paullina pinnata: 6α-(3’-Methoxy-4’-hydroxybenzoyl)-lup-20(29)-ene-3-one. Nat Prod Commun 2015; 10:563564.
Lunga PK, Nkodo JM, Tamokou JD, Kuiate JR, Gatsing G, Tchoumboue J. Post-treatment evaluation of the side effects of methanol leaf extract from Paullinia pinnata Linn, an antityphoid plant. Pharmacologia 2015; 6:264-272.
Afagnigni AD, Nyegue MA, Ndoye FC, Voundi OS, Fonkoua MC, Etoa F-X. In vitro assessment of antibacterial and antioxidant activities of ethanolic leaves extracts of Paullinia pinnata Linn (Sapindaceae). World J Pharm Sci 2016; 4:173-182.
Nyegue MA, Afagnigni AD, Ndam YN, Djova SV, Fonkoua MC, Etoa F-X. Toxicity and activity of ethanolic leaf extract of Paullinia pinnata (Sapindaceae) in Shigella flexneri-induced diarrhea in Wistar rats. J Evid-Based Integr Med 2020; 25:1-9.
Eloff JN. A sensitive and quick microplate method to determine the minimum inhibitory concentration of plant extracts for bacteria. Plant Med 1998; 64:711-713.
Babii C, Bahrin LG, Neagu AN, Gostin I, Mihasan M, Birsa LM et al. Antibacterial activity and proposed action mechanism of a new class of synthetic tricyclic flavonoids. J Appl Microbiol 2016; 120:630-637.
Frost DJ, Brandt KD, Cugier D, Goldman R A whole-cell Candida albicans assay for the detection of inhibitors towards fungal cell wall synthesis and assembly. J Antibiot 1995; 48:306-310.
Bennis S, Chami F, Chami N, Bouchikhi T, Remmal A. Surface alteration of Saccharomyces cerevisiae induced by thymol and eugenol. Lett Appl Microbiol 2004; 38:454-458.
Hao G, Shi YH, Tang YI, Le GW. The membrane action mechanism of analogs of the antimicrobial peptide Buforin 2. Peptides 2009; 30:1421-1427.
Da Silva IF, De Oliveira RG, Soares IM, Alvim TC, Ascêncio SD, Martins DT. Evaluation of acute toxicity, antibacterial activity and mode of action of the hydroethanolic extract of Piper umbellatum L. J Ethnopharmacol. 2014 ; 151:137-143.
Lambert RJ, Skandamis PN, Coote PJ, Nychas GJ. A study of the minimum inhibitory concentration and mode of action of oregano essential oil, thymol and carvacrol. J Appl Microbiol 2001; 91:453-462.
Tambe AB, Nzefa LD, Nchang AN. Childhood diarrhea determinants in Sub-Saharan Africa: A cross sectional study of Tiko-Cameroon. Challenges 2015; 6:229-243.
Mekonnen B, Asrie AB, Wubneh ZB. Antidiarrheal activity of 80% methanolic leaf extract of Justicia schimperiana. Evid.-Based Complement. Alternat Med 2018; 3037120:1-10.
Ali I, Sharma P, Suri KA, Satti NK, Dutt P, Afrin A et al. In vitro antifungal activities of amphotericin B in combination with acteoside, a phenylethanoid glycoside from Colebrookea oppositifolia. J Med Microbiol 2011; 60: 1326-1336.
Kuete V, Fokou W F, Karaosmanoğlu O, Beng VP, Sivas H. Cytotoxicity of the methanol extracts of Elephantopus mollis, Kalanchoe crenata and 4 other Cameroonian medicinal plants towards human carcinoma cells. BMC Complement Altern Med 2017; 17:280.
Adeyemo SO, Makinde J. M. Acute and sub-acute toxicity studies of the methanol extract of the leaves of Paullinia pinnata Linn in Wistar mice and rats. Afr J Med Med Sci 2013; 42:81-90
Lunga PK, Tamoukou JD, Fodouop SP, Kuiate JR, Gatsing TD. Antityphoid and radical scavenging properties of the methanol extracts and compounds from the aerial part of Paullinia pinnata Linn (Sapindaceae). SpringerPlus. 2014; 3:302.
Mariame E. Aboudoulatif D, Povi LE, Kodjo A, Kossi M, Essohana P et al. Repeated toxicological study and cardiotoxicity of hydroalcoholic root extract of Paullinia pinnata Linn (Sapindaceae). J Appl Pharm Sci 2016; 6:24-28.
Trombetta D, Castelli F, Sarpietro MG, Venuti V, Cristani M, Daniele C et al. Mechanisms of antibacterial action of three monoterpenes. Antimicrob Agents Chemother 2005; 49:2474-2478.
Khan MS, Ahmad I, Cameotra SS. Phenyl aldehyde and propanoids exert multiple sites of action towards cell membrane and cell wall targeting ergosterol in Candida albicans. AMB Express 2013; 3:54.
Cowan MM. Plant’s products as antimicrobial agents. Clin Microbiol Rev. 1999; 14:564-582.
Carson CF, Mee BJ, Riley TV. Mechanism of action of Melaleuca alternifolia (tea tree) oil on Staphylococcus aureus determined by time-kill, lysis, leakage, and salt tolerance assays and electron microscopy. Antimicrob. Agents Chemother 2002; 46:1914-1920.
Zablotowicz RM, Hoagland RE, Wagner SC. Effect of Saponins on the Growth and Activity of Rhizosphere Bacteria," In: Waller G.R., Yamasaki K. (eds) Saponins used in Food and Agriculture. Adv Exp Med Biol Springer, Boston MA 1996.
Shimada T. Salivary proteins as a defence against dietary tannins. J Chem Ecol 2006; 32:1149-1163.
Denyer SP. Mechanisms of action of biocides. Int Biodeterior Biodegr 1990; 26:89-100.
Patra JK, Baek KH. Antibacterial activity and action mechanism of the essential oil from Enteromorpha linza L. against foodborne pathogenic bacteria. Molecules 2016; 21:388.
Ghestem A, Seguin E, Paris M, Orecchioni AM. Le préparateur en pharmacie. Dossier 2, Botanique-Pharmacognosie-Phytothérapie-Homéopathie. Tec et Doc (ed), 2001; 272.
Bezic N, Skocibusic M, Dinkic V, Radonic A. Composition and antimicrobial activity of Achillea clavennae L. essential oil. Phytother Res 2003; 17:1037-1040.
Sharma A, Bajpai VK, Baek KH. Determination of antibacterial mode of action of Allium sativum essential oil against foodborne Pathogens using membrane permeability and surface characteristic parameters. J Food Saf 2013; 33:197-208.
Jiangwei M, Luxi J, Yu C, Jian K. Activities and mechanisms of eugenol and cinnamaldehyde against Legionella pneumophila. Int J Clin Exp Med 2017; 10:16460-16467.
Tassou C, Koutsoumanis K, Nychas GJ. Inhibition of Salmonella enteritidis and Staphylococcus aureus in nutrient broth by mint essential oil. Food Res Int 2000; 33 :273-280.
Bajpai VK, Al-Reza SM, Choi UK, Lee JH, Kang SC. Chemical composition antibacterial and antioxidant activities of leaf essential oil and extracts of Metasequoia glyptostroboides Miki ex Hu. Food Chem. Toxicol 2009; 47:1876-1883.
Famuyinde IM, Fasina FO, Eloff JN, McGaw LJ. The ultrastructural damage caused by Eugenia zeyheri and Syzygium legatii acetone leaf extracts on pathogenic Escherichia coli. BMC Vet Res 2020; 16:326.
Wang LH, Zhang ZH, Zeng XA, Gong DM, Wang MS. Combination of microbiological, spectroscopic and molecular docking techniques to study the antibacterial mechanism of thymol against Staphylococcus aureus: membrane damage and genomic DNA binding. Anal Bioanal Chem 2017, 409:3055-3055
Published
Abstract Display: 738 
PDF Downloads: 736 PDF Downloads: 997 How to Cite
Issue
Section
Authors who publish with this journal agree to the following terms:
- Authors retain copyright and grant the journal right of first publication with the work simultaneously licensed under a Creative Commons Attribution-NonCommercial 4.0 International (CC BY-NC 4.0). that allows others to share the work with an acknowledgment of the work's authorship and initial publication in this journal.
- Authors are able to enter into separate, additional contractual arrangements for the non-exclusive distribution of the journal's published version of the work (e.g., post it to an institutional repository or publish it in a book), with an acknowledgment of its initial publication in this journal.
- Authors are permitted and encouraged to post their work online (e.g., in institutional repositories or on their website) prior to and during the submission process, as it can lead to productive exchanges, as well as earlier and greater citation of published work (See The Effect of Open Access).
How to Cite
.