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Antibacterial activity and stability of mouthwash formulated with ethanolic extract of sappan wood (Caesalpinia sappan L.) against Streptococcus mutans ATCC 25173 and Pseudomonas aeruginosa ATCC 10145

DOI
https://doi.org/10.14719/pst.10427
Submitted
3 July 2025
Published
29-01-2026
Versions

Abstract

This study aimed to develop and evaluate a herbal mouthwash formulation containing an ethanolic extract of sappan wood (Caesalpinia sappan L.) as a potential natural alternative to chlorhexidine-based products. The extract and derived mouthwash were assessed for antibacterial activity against Streptococcus mutans Clarke and Pseudomonas aeruginosa (Schroeter) Migula using the agar well diffusion and minimum inhibitory concentration (MIC) methods at concentrations of 1 %, 3 % and 5 %. Positive controls included 0.2 % chlorhexidine and a commercial mouthwash, while negative controls consisted of mouthwash without extract and 96 % ethanol. The stability of the mouthwash was evaluated through a six-cycle cycling test by monitoring organoleptic properties, homogeneity, pH, sedimentation and redispersion. The mouthwash exhibited a maximum inhibition zone of 21.5 mm and 15.2 mm against S. mutans and P. aeruginosa, respectively, with MIC values of 1 % and 3 %. It remained physically stable during six storage cycles with consistent pH (4.6–6.5) and homogeneous dispersion. These results demonstrate that sappan wood extract possesses strong antibacterial activity and good formulation stability, supporting its potential as a safe and effective herbal mouthwash. The study highlights the novelty of exploring a plant-based mouthwash as a promising natural alternative to chlorhexidine, showing comparable in vitro antibacterial activity with potentially fewer adverse effects.

References

  1. 1. Peres MA, Macpherson LMD, Weyant RJ, Daly B, Venturelli R, Mathur MR, et al. Oral diseases: a global public health challenge. Lancet. 2019;394(10194):249-60. https://doi.org/10.1016/S0140-6736(19)31146-8
  2. 2. Center for Data and Information, Ministry of Health of the Republic of Indonesia. InfoDATIN: National dental health. Jakarta: Ministry of Health of the Republic of Indonesia; 2019.
  3. 3. Saputri D. Radiographic features of periodontal disease. J Syiah Kuala Dent Soc. 2018;3(1):16-21.
  4. 4. Puttipan R, Chansakaow S, Khongkhunthian S, Okonogi S. Caesalpinia sappan: A promising natural source of antimicrobial agent for inhibition of cariogenic bacteria. Drug Discov Ther. 2018;12(4):197-205. https://doi.org/10.5582/ddt.2018.01035
  5. 5. Hajardhini P, Susilowati H, Yulianto HDK. Oral cavity as a potential reservoir for Pseudomonas aeruginosa infection. Odonto Dent J. 2020;7(2):125-33. https://doi.org/10.30659/odj.7.2.125-133
  6. 6. Rasyadi Y. Mouthwash formulations from leaf extract of Artocarpus altilis (Parkinson ex F.A. Zorn) Fosberg breadfruit. Chempublish J. 2018;3(2):76-84. https://doi.org/10.22437/chp.v3i2.5767
  7. 7. Asfar AMIA, Yasser M. Qualitative phytochemical analysis of flavonoid content of secang wood (Caesalpinia sappan L.) extract from ultrasonic assisted solvent extraction method. J Chem. 2018;19(2):15-25. https://doi.org/10.35580/chemica.v19i2.12772
  8. 8. Batubara I, Mitsunaga T, Ohashi H. Brazilin from Caesalpinia sappan wood as an antiacne agent. J Wood Sci. 2010;56(1):77-81. https://doi.org/10.1007/s10086-009-1046-0
  9. 9. Nirmal NP, Rajput MS, Prasad RGSV, Ahmad M. Brazilin from Caesalpinia sappan heartwood and its pharmacological activities: a review. Asian Pac J Trop Med. 2015;8(6):421-30. https://doi.org/10.1016/j.apjtm.2015.05.014
  10. 10. Reddy VLN, Ravikanth V, Lakshmi VVNSJ, Murty US, Venkateswarlu Y. Inhibitory activity of homoisoflavonoids from Caesalpinia sappan against Beauveria bassiana. Fitoterapia. 2003;74(6):600-2. https://doi.org/10.1016/S0367-326X(03)00153-9
  11. 11. Puttipan R, Wanachantarak P, Khongkhuthian S, Okonogi S. Effects of Caesalpinia sappan on pathogenic bacteria causing dental caries and gingivitis. Drug Discov Ther. 2017;11(6):316-22. https://doi.org/10.5582/ddt.2017.01055
  12. 12. Seephonkai P, Sedlak S, Wongpakam K, Sangdee K, Sangdee A. Time-kill kinetics and mechanism of action of Caesalpinia sappan L. and Ochna integerrima (Lour.) Merr. water extracts against pathogenic bacteria. J Pharm Pharmacogn Res. 2021;9(6):813-23. https://doi.org/10.56499/jppres21.1095_9.6.813
  13. 13. Kadchumsang S, Sirisard P, Sookkhee S, Chansakaow S. Antibacterial and antioxidant activities of Lanna medicinal plants used in mahoog formula. Int J Pharm Pharm Sci. 2015;7(9):366-70.
  14. 14. Pattananandecha T, Apichai S, Julsrigival J, Ogata F, Kawasaki N, Saenjum C. Antibacterial activity against foodborne pathogens and inhibitory effect on anti-inflammatory mediators’ production of brazilin-enriched extract from Caesalpinia sappan Linn. Plants. 2022;11:1698. https://doi.org/10.3390/plants11131698
  15. 15. Sugiaman VK, Jeffrey J, Widowati W, Dewi NSM, Ferdiansyah R, Muchtar DM, et al. Anti-inflammatory activity of sappan wood extract cream (Caesalpinia sappan) in Porphyromonas gingivalis-induced gingivitis rat models. Open Vet J. 2024;14(9):2224-36. https://doi.org/10.5455/OVJ.2024.v14.i9.10
  16. 16. Ministry of Health of the Republic of Indonesia. Indonesian herbal pharmacopoeia. 2nd ed. Jakarta: Ministry of Health of the Republic of Indonesia; 2017.
  17. 17. Arsiningtyas IS. Antioxidant profile of heartwood and sapanwood of Caesalpinia sappan L. trees’ part grown in Imogiri Nature Preserve, Yogyakarta. IOP Conf Ser Earth Environ Sci. 2021;810:012040. https://doi.org/10.1088/1755-1315/810/1/012040
  18. 18. Organisation for Economic Co-operation and Development. Test No. 407: repeated dose 28-day oral toxicity study in rodents. Paris: OECD Publishing; 2014.
  19. 19. Kumar T, Jain V. Appraisal of total phenol, flavonoid contents and antioxidant potential of folkloric Lannea coromandelica using in vitro and in vivo assays. Scientifica. 2015;203679:1-15. https://doi.org/10.1155/2015/203679
  20. 20. Barma MD, Kannan SD, Indiran MA, Rajeshkumar S, Kumar RP. Antibacterial activity of mouthwash incorporated with silica nanoparticles against Staphylococcus aureus, Streptococcus mutans, Enterococcus faecalis: an in vitro study. J Pharm Res Int. 2020;32(16):25-33. https://doi.org/10.9734/jpri/2020/v32i1630646
  21. 21. Wang X, Shen Y, Thakur K, Han J, Zhang JG, Hu F, et al. Antibacterial activity and mechanism of ginger essential oil against Escherichia coli and Staphylococcus aureus. Molecules. 2020;25(17):3955. https://doi.org/10.3390/molecules25173955
  22. 22. Gurning D, Nathaniel D, Meila O, Sagala Z. Antibacterial activity test of mouthwash preparations from 70 % ethanol extract of life-sustaining stems (Gynura procumbens (Lour.) Merr.) against Streptococcus mutans. Pharmacon. 2018;15(2):58-64. https://doi.org/10.23917/pharmacon.v15i2.5880
  23. 23. Suryani N, Adini S, Stiani SN, Indriatmoko DD. Herbal mouthwash containing ethyl acetate extract of bintaro bark (Cerbera odollam Gaertn.) as an antibacterial of Streptococcus mutans that causes dental plaque. J Farmaka. 2019;17(2):48-56.
  24. 24. National Standardization Agency of Indonesia (BSN). Indonesian national standard (SNI) 12-3524-1995: toothpaste. Jakarta: BSN; 1995.
  25. 25. Nurul N, Indrayati A, Murrukmihadi M. Optimization of liquid soap (Caesalpinia sappan L.) ethanol extract using simplex lattice design and its antibacterial effect on Staphylococcus aureus ATCC 25259. Infokes. 2019;9(2):7-12.
  26. 26. Amelinda E, Widarta IWR, Darmayanti LPT. Effect of maceration time on antioxidant activity of temulawak rhizome extract (Curcuma xanthorrhiza Roxb.). J Ilmu Teknol Pangan. 2018;7(4):165-74. https://doi.org/10.24843/itepa.2018.v07.i04.p03
  27. 27. Holidah D, Dewi IP, Christianty FM, Muhammadiya NS, Huda N. Antidiabetic and antidyslipidemic activity of secang (Caesalpinia sappan L.) wood extract on diabetic rat. Res J Pharm Technol. 2021;14(5):2801-6. https://doi.org/10.52711/0974-360X.2021.00494
  28. 28. Ayele DT, Akele ML, Melese AT. Analysis of total phenolic contents, flavonoids, antioxidant and antibacterial activities of Croton macrostachyus root extracts. BMC Chem. 2022;16(1):30. https://doi.org/10.1186/s13065-022-00822-0
  29. 29. Handayani F, Warnida H, Nur SJ. Formulation and test of antibacterial activity of Streptococcus mutans from the mouthwash preparation of bay leaf extract (Syzygium polyanthum (Wight) Walp.). Media Sains. 2016;9(1):74-84.
  30. 30. Kao TT, Wang MC, Chen YH, Chung YT, Hwang PA. Propylene glycol improves stability of the anti-inflammatory compounds in Scutellaria baicalensis extract. Processes. 2021;9:894. https://doi.org/10.3390/pr9050894
  31. 31. Sumiati T, Masaenah E, Mustofa KN. Formulation of herbal mouthwash ethanol extract of pineapple peel (Ananas comosus (L.) Merr.) as antibacterial Streptococcus sanguinis. J Farmamedika. 2021;6(1):15-23. https://doi.org/10.47219/ath.v6i1.112
  32. 32. Bontjura S, Waworuntu OA, Siagian KV. Test of antibacterial effect of leilem leaf extract (Clerodendrum minahassae L.) against Streptococcus mutans. Pharmacon. 2015;4(4):96-101.
  33. 33. Faradina AS, Mastra N, Karta IW. Antibacterial activity test of gout root (Plumbago zeylanica L.) ethanol extract on the growth of Pseudomonas aeruginosa in vitro. Meditory J Med Lab. 2019;7(2):110-18.
  34. 34. Davis WW, Stout TR. Disc plate method of microbiological antibiotic assay. Appl Microbiol. 1971;22(4):659-65. https://doi.org/10.1128/am.22.4.659-665.1971
  35. 35. Budi HS, Soesilowati P, Wirasti MJ. Antibacterial activity of sappan wood (Caesalpinia sappan L.) against Aggregatibacter actinomycetemcomitans and Porphyromonas gingivalis. Syst Rev Pharm. 2020;11(3):349-53. https://doi.org/10.37506/v11/i2/2020/ijphrd/194988
  36. 36. Breijyeh Z, Jubeh B, Karaman R. Resistance of gram-negative bacteria to current antibacterial agents and approaches to resolve it. Molecules. 2020;25(6):1-23. https://doi.org/10.3390/molecules25061340
  37. 37. Giovagnorio F, Sepe L, Tosco A, Esposito C. Resistance in Pseudomonas aeruginosa: a narrative review of antibiogram interpretation and emerging treatments. Antibiotics. 2023;12(11):1621. https://doi.org/10.3390/antibiotics12111621
  38. 38. Halboub E, Al-Maweri SA, Al-Wesabi M, Al-Kamel A, Shamala A, Al-Sharani A, et al. Efficacy of propolis-based mouthwashes on dental plaque and gingival inflammation: a systematic review. BMC Oral Health. 2020;20(1):198. https://doi.org/10.1186/s12903-020-01185-5
  39. 39. Pokharana M, Vaishnav R, Goyal A, Shrivastava A. Stability testing guidelines of pharmaceutical products. J Drug Deliv Ther. 2018;8(2):169-75. https://doi.org/10.22270/jddt.v8i2.1564
  40. 40. Pramiastuti O, Agusetianti N. The formulation of mouthwash of starfruit leaf extract (Averrhoa bilimbi L.) by maceration method. J Farm Sains Indones. 2019;2(1):21-31.
  41. 41. Wijaya HM, Lina RN. Formulation and physical evaluation of suspension preparations of papaya seed extract (Carica papaya L.) and teki grass tuber (Cyperus rotundus L.). Cendekia J Pharm. 2021;5(2):166-76. https://doi.org/10.31596/cjp.v5i2.160
  42. 42. Roland WSU, van Buren L, Gruppen H, Driesse M, Gouka RJ, Smith G, et al. Bitter taste receptor activation by flavonoids and isoflavonoids. J Agric Food Chem. 2013;61(44):10454-66. https://doi.org/10.1021/jf403387p
  43. 43. Djafar F, Yamlean PVY, Siampa JP. Mouthwash formulation of water hyacinth (Eichhornia crassipes (Mart.) Solms) extract as antibacterial of dental caries (Streptococcus mutans). Pharmacon. 2021;10(4):1169-77.
  44. 44. van Swaaij BWM, Slot DE, Van der Weijden GA, Timmerman MF, Ruben J. Fluoride, pH value and titratable acidity of commercially available mouthwashes. Int Dent J. 2024;74(2):260-67. https://doi.org/10.1016/j.identj.2023.09.002
  45. 45. Pratiwi RI, Amananti W. Utilization of carboxy methyl cellulose and cassava starch as suspending agents in pandan leaf suspension preparations. J Ilm Manuntung. 2021;7(1):111-19.
  46. 46. Ministry of Health of the Republic of Indonesia. The Indonesian pharmacopoeia. 6th ed. Jakarta: Ministry of Health of the Republic of Indonesia; 2020.

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