Keywords
Staphylococcus aureus
anti-bacterial agents
pharmacologic actions
time-kill kinetics
How to Cite
Abstract
Staphylococcus aureus is one of the main pathogens in community and hospital infections that could cause mild skin infections to severe life-threatening bacteremia. Piper betle has shown antibacterial activities against S. aureus but its pharmacodynamics remains unclear despite its widespread availability in many over-the-counter preparations. This study aimed to describe the time-kill kinetics of P. betle extract against methicillin-sensitive Staphylococcus aureus ATCC 29213 (MSSA). Time-kill kinetics were conducted to describe the killing rate of P. betle extract by its effects on MSSA lag time duration, growth rate, and maximum optical density. The surviving colonies at different time-points after exposure to P. betle extract at minimum bactericidal concentration were also determined. Subinhibitory concentrations elucidated its concentration-dependent antibacterial activity by maintaining a trend of increasing lag time, decreasing growth rate, and decreasing maximum optical density as the concentration increases. In particular, concentration at 1250 μg/mL or 0.5 x minimum inhibitory concentration (MIC) showed consistent significant findings across these parameters. Concentrations at MIC and above did not show growth, indicating MSSA growth inhibition or death. P. betle extract’s bactericidal effect started immediately within two (2) hours and was sustained until no growth was observed from the eighth hour and beyond. P. betle extract maintained antibacterial activities against MSSA at subinhibitory concentrations and has also exhibited immediate and sustained bactericidal effect at minimum bactericidal concentration (MBC).
References
Balinado, L. O., & Chan, M. A. (2019). Assessment of Cytotoxic Activity of Five Common Philippine Medicinal Plants Using Brine Shrimp Lethality Assay. Mindanao Journal of Science and Technology, 17, 138–152.
Braga, P. C., Sala, M. T., & dal Sasso, M. (1999). Pharmacodynamic Effects of Subinhibitory Concentrations of Rufloxacin on Bacterial Virulence Factors. Antimicrobial Agents and Chemotherapy, 43(5), 1013–1019.
https://doi.org/10.1128/AAC.43.5.1013
Budiman, A., Rusnawan, D. W., & Yuliana, A. (2018). Antibacterial activity of Piper betle L. extract in cream dosage forms against Staphylococcus aureus and Propionibacterium acne. Journal of Pharmaceutical Sciences, 10, 4.
Chen, J., Zhou, H., Huang, J., Zhang, R., & Rao, X. (2021). Virulence alterations in Staphylococcus aureus upon treatment with the sub-inhibitory concentrations of antibiotics. Journal of Advanced Research, 31, 165–175.
https://doi.org/10.1016/j.jare.2021.01.008
CLSI. (2012). Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically; Approved Standard—Ninth Edition. Clinical and Laboratory Standards Institute.
Das, B., Mandal, D., Dash, S. K., Chattopadhyay, S., Tripathy, S., Dolai, D. P., Dey, S. K., & Roy, S. (2016). Eugenol Provokes ROS-Mediated Membrane Damage-Associated Antibacterial Activity against Clinically Isolated Multidrug-Resistant Staphylococcus aureus Strains. Infectious Diseases: Research and Treatment, 9, IDRT.S31741.
https://doi.org/10.4137/IDRT.S31741
Datta, A., Ghoshdastidar, S., & Singh, M. (2011). Antimicrobial Property of Piper betel Leaf against Clinical Isolates of Bacteria. International Journal of Pharma Sciences and Research, 2(3), 104–109.
Eagle, H., Fleischman, R., & Musselman, A. D. (1950). Effect of schedule of administration on the therapeutic efficacy of penicillin. The American Journal of Medicine, 9(3), 280–299.
https://doi.org/10.1016/0002-9343(50)90425-6
Eagle, H., & Musselman, A. D. (1949). The Slow Recovery of Bacteria from the Toxic Effects of Penicillin. Journal of Bacteriology, 58(4), 475–490.
https://doi.org/10.1128/jb.58.4.475-490.1949
Hall, B. G., Acar, H., Nandipati, A., & Barlow, M. (2014). Growth Rates Made Easy. Molecular Biology and Evolution, 31(1), 232–238.
https://doi.org/10.1093/molbev/mst187
Hyldgaard, M., Mygind, T., & Meyer, R. L. (2012). Essential Oils in Food Preservation: Mode of Action, Synergies, and Interactions with Food Matrix Components. Frontiers in Microbiology, 3.
https://doi.org/10.3389/fmicb.2012.00012
Jackson, K. A., Gokhale, R. H., Nadle, J., Ray, S. M., Dumyati, G., Schaffner, W., Ham, D. C., Magill, S. S., Lynfield, R., & See, I. (2019). Public Health Importance of Invasive Methicillin-sensitive Staphylococcus aureus Infections: Surveillance in 8 US Counties, 2016. Clinical Infectious Diseases, ciz323.
https://doi.org/10.1093/cid/ciz323
Jacobs, M. R. (2001). Optimisation of antimicrobial therapy using pharmacokinetic and pharmacodynamic parameters. Clinical Microbiology and Infection, 7(11), 589–596.
https://doi.org/10.1046/j.1198-743x.2001.00295.x
Jamelarin, E. M., & Balinado, L. O. (2019). Evaluation of Antibacterial Activity of Crude Aqueous, Ethanolic and Methanolic Leaf Extracts of Piper retrofractum Vahl. And Piper betle L. Asian Journal of Biological and Life Sciences, 8(2), 63–67.
https://doi.org/10.5530/ajbls.2019.8.11
Lao, R. C. C., Yabes, A. M., Tobias-Altura, M., Panganiban, L. C. R., & Makalinao, I. R. (2022). In Vitro Antibacterial and Antibiofilm Activities of Piper betle L. Ethanolic Leaf Extract on Staphylococcus aureus ATCC 29213 [Pre-print]. Acta Medica Philippina.
Missiakas, D. M., & Schneewind, O. (2013). Growth and Laboratory Maintenance of Staphylococcus aureus. Current Protocols in Microbiology, Chapter 9, Unit-9C.1.
https://doi.org/10.1002/9780471729259.mc09c01s28
NCCLS. (1999). Methods for Determining Bactericidal Activity of Antimicrobial Agents; Approved Guideline (Vol. 19).
Odenholt, I. (2001). Pharmacodynamic effects of subinhibitory antibiotic concentrations. International Journal of Antimicrobial Agents, 17(1), 1–8.
https://doi.org/10.1016/S0924-8579(00)00243-0
Otto, M. P., Martin, E., Badiou, C., Lebrun, S., Bes, M., Vandenesch, F., Etienne, J., Lina, G., & Dumitrescu, O. (2013). Effects of subinhibitory concentrations of antibiotics on virulence factor expression by community-acquired methicillin-resistant Staphylococcus aureus. Journal of Antimicrobial Chemotherapy, 68(7), 1524–1532.
https://doi.org/10.1093/jac/dkt073
Singh, D., Narayanamoorthy, S., Gamre, S., Majumdar, A. G., Goswami, M., Gami, U., Cherian, S., & Subramanian, M. (2018). Hydroxychavicol, a key ingredient of Piper betle induces bacterial cell death by DNA damage and inhibition of cell division. Free Radical Biology and Medicine, 120, 62–71.
https://doi.org/10.1016/j.freeradbiomed.2018.03.021
Taylor, T. A., & Unakal, C. G. (2021). Staphylococcus aureus. In StatPearls. StatPearls Publishing.
https://www.ncbi.nlm.nih.gov/books/NBK441868/
Trakranrungsie, N., Chatchawanchonteera, A., & Khunkitti, W. (2006). Antidermatophytic Activity of Piper betle Cream. Thai Journal of Pharmacology, 28(3).
https://li01.tci-thaijo.org/index.php/JBAP/article/view/200219/139965
Valle, D. L. J., Andrade, J. I., Puzon, J. J. M., Cabrera, E. C., & Rivera, W. L. (2015). Antibacterial activities of ethanol extracts of Philippine medicinal plants against multidrug-resistant bacteria. Asian Pacific Journal of Tropical Biomedicine, 5(7), 532–540.
https://doi.org/10.1016/j.apjtb.2015.04.005
Valle, D. L. J., Cabrera, E. C., Puzon, J. J. M., & Rivera, W. L. (2016). Antimicrobial Activities of Methanol, Ethanol and Supercritical CO2 Extracts of Philippine Piper betle L. on Clinical Isolates of Gram Positive and Gram Negative Bacteria with Transferable Multiple Drug Resistance. PLOS ONE, 11(1), e0146349.
https://doi.org/10.1371/journal.pone.0146349
Valle, D. L. J., Puzon, J. J. M., Cabrera, E. C., & Rivera, W. L. (2016). Thin Layer Chromatography-Bioautography and Gas Chromatography-Mass Spectrometry of Antimicrobial Leaf Extracts from Philippine Piper betle L. against Multidrug-Resistant Bacteria. Evidence-Based Complementary and Alternative Medicine : ECAM, 2016, 4976791.
https://doi.org/10.1155/2016/4976791
Valle, D. L. Jr., Puzon, J. J. M., Cabrera, E. C., Cena-Navarro, R. B., & Rivera, W. L. (2021). Antimicrobial efficacy and activity of ethanolic extract of Piper betle L. on Staphylococcus aureus-infected wound in mice and clinical isolates of multiple drug-resistant bacterial pathogens. Tropical Biomedicine, 38(2), 134–142.
https://doi.org/10.47665/tb.38.2.049
Wiegand, I., Hilpert, K., & Hancock, R. E. W. (2008). Agar and broth dilution methods to determine the minimal inhibitory concentration (MIC) of antimicrobial substances. Nature Protocols, 3(2), 163–175.
https://doi.org/10.1038/nprot.2007.521
Zhanel, G. G., Hoban, D. J., & Harding, G. K. (1992). Subinhibitory antimicrobial concentrations: A review of in vitro and in vivo data. The Canadian Journal of Infectious Diseases, 3(4), 193–201.
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