EVERGREEN

Joint Journal of Novel Carbon Resource Sciences and Green Asia Strategy

ISSN:2189-0420 (Print until Mar 2020)
ISSN:2432-5953 (Online)

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Ozone Technology for Improving the Microbiological Quality of Milk from Medicated Dairy Cows

Putty Anggraeni1, Biatna Dulbert Tampubolon1, Endi Hari Purwanto1, Widia Citra Anggundari1, Ary Budi Mulyono1, Daryono Restu Wahono1, Yonan Prihhapso1, Bambang Prasetya1, Lilik Eka Radiati2,*, Abdul Manab2
1Research Center for Equipment Manufacturing Technology, National Research and Innovation Agency, Indonesia
2Faculty of Animal Science, Brawijaya University, Indonesia
*Author to whom correspondence should be addressed:
E-mail: lilik.eka@ub.ac.id (LER)
Evergreen, Vol. 13, Iss. 02, pp. 596–610, 2026
DOI: 10.5109/7420071
Creative Commons Attribution 4.0 International Creative Commons Attribution 4.0 International
Received: May 28, 2025 | Revised: December 29, 2025 | Accepted: February 17, 2026 | Published: June 2026
Abstract
This study examines the effects of vaccination and medication on bacterial contamination in milk and evaluates the potential of ozone technology for bacterial reduction. A total of 52 milk samples were collected, representing 26 paired observations analyzed statistically from dairy cows aged 2–9 years in Subang, West Java, were analyzed before and after ozonation. Bacterial counts were assessed for compliance with SNI 3141.1:2011. Non-parametric statistical methods were applied because the microbial data did not follow a normal distribution, as indicated by Shapiro-Wilk and Kolmogorov-Smirnov normality tests (p < 0.05). After ozonation, a statistically significant reduction in bacterial counts was observed. Paired analysis using the Wilcoxon Signed-Rank test (paired N = 26) showed a significant decrease in Total Plate Count (Z = 4.198, p < 0.001), with a large effect size (r = 0.82). Mean TPC decreased from 6.07 × 10⁸ CFU/mL (log₁₀ = 8.78) before treatment to 1.68 × 10⁷ CFU/mL (log₁₀ = 7.23) after 15 minutes of ozonation, corresponding to an average reduction of 1.55 log₁₀ CFU/mL. Following ozonation, compliance with SNI 3141.1:2011 increased from 3.8% to 26.9%. These findings demonstrate that ozone treatment effectively improves the microbiological quality milk from vaccinated and medicated dairy cows and can serve as a residue-free post- harvest intervention to enhance milk safety and quality in herds that are undergoing therapeutic treatments.
Keywords
antibiotic; bacterial count; dairy cow; milk; ozonation; vaccination
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References
  1. 1) A. Kuipers, W.J. Koops, and H. Wemmenhove, "Antibiotic use in dairy herds in the netherlands from 2005 to 2012," J. Dairy Sci., 99 (2) 1632-1648 (2016) doi:10.3168/jds.2014-8428
  2. 2) P.D. Constable, K.W. Hinchcliff, S.H. Done, and W. Grünberg, "Veterinary medicine: a textbook of the diseases of cattle, horses, sheep, pigs and goats," Elsevier Health Sciences, 2016
  3. 3) D. Tomanić, M. Samardžija, and Z. Kovačević, "Alternatives to antimicrobial treatment in bovine mastitis therapy: a review," Antibiotics, 12 (4) 683 (2023) doi:10.3390/antibiotics12040683
  4. 4) R. Floris, I. Recio, B. Berkhout, and S. Visser, "Antibacterial and antiviral effects of milk proteins and derivatives thereof," Curr. Pharm. Des., 9 (16) 1257-1275 (2003) doi:10.2174/1381612033454810
  5. 5) I. Hotea, M. Dragomirescu, A. Berbecea, and I. Radulov, "The role of nutrition in enhancing sustainability in sheep production," (2023) doi:10.5772/intechopen.113938
  6. 6) E.I. Stoltz, and D.J. Hankinson, "Antibiotics and lactic acid starter cultures," Appl. Microbiol., 1 (1) 24-29 (1953)
  7. 7) S. Nurmala, and D.O. Gunawan, "Pengetahuan penggunaan obat antibiotik pada masyarakat yang tinggal di kelurahan babakan madang," J Ilm Farm, 10 (1) 22-31 (2020) doi:10.33751/jf.v10i1.1728
  8. 8) N.F. Amalia, D. Mahdiyah, and N. Noval, "Antibacterial Activity of Bungur Bark Extract (Lagerstroemia speciosa (L.) Pers) Againts Staphylococcus aureus ATCC 29213 By Diffusion and Dillution Methods.," in: Int. Conf. Heal. Sci., 2021: pp. 470-479
  9. 9) M.F.R. Shuvo, F.F. Haque, and K.M.A. Kabir, "Modeling the impact of combined effect of vaccination, testing, and treatment on epidemic dynamics," Evergr. Jt. J. Nov. Carbon Resour. Sci. Green Asia Strateg., 11 (03) 1498-1507 (2024) doi:10.5109/7236807
  10. 10) M.A. Cavalcante, B.R. de C. Leite Júnior, A.A.L. Tribst, and M. Cristianini, "Improvement of the raw milk microbiological quality by ozone treatment.," Int. Food Res. J., 20 (4) (2013)
  11. 11) G. Sydykova, S. Umbetova, Z. Baimakhanova, G. Abieva, and G. Kurmanbayev, "Modern applications of ozone technology," 10 (4) 2308-2316 (2023). doi: ttps://doi.org/10.5109/7160908
  12. 12) R.B. Afonso, R.H.R. Moreira, and P.L.R. de Almeida, "Can ozone be used as antimicrobial in the dairy industry? a systematic review," J. Dairy Sci., 105 (2) 1051-1057 (2022) doi:10.3168/jds.2021-20900
  13. 13) T.E. Susilorini, S. Maylinda, P. Surjowardojo, and S. Suyadi, "Importance of body condition score for milk production traits in peranakan etawah goats.," (2014)
  14. 14) N. Montayeva, S. Montayev, and A. Montayeva, "Bioactive mineral feed additive for poultry: therapeutic and preventive effects on haematological parameters in chickens," 11 (02) 984-992 (2024) doi:10.3390/ani12131623
  15. 15) P.C.N. Mantrawan, I.N.K. Besung, I.G.K. Suarjana, and N.K. Suwiti, "Total bakteri pada berbagai umur dan lokasi peternakan sapi bali di nusa penida," Bul. Vet. Udayana, 10 (2) 122-126 (2018) doi:10.24843/bulvet.2018.v10.i02.p03
  16. 16) Kementerian Pertanian, "Annual Report 2022," 2022
  17. 17) A.D.W.I. Setyawan, "Review: the diversity of local cattle in indonesia and the efforts to develop superior indigenous cattle breeds," 17 (1) 275-295 (2016) doi:10.13057/biodiv/d170139
  18. 18) F.F. Rahmah, F. Rahly, and Y. Yusriani, "Development planning ranch area based on the potential of feeding ruminants in aceh , indonesia," 05002 1-13 (2021) doi:10.1051/e3sconf/202130605002
  19. 19) P. Biswal, "ASSESSMENT of immunity, stress and production performance of bovines after foot and mouth disease vaccination", Deemed University. ICAR-Indian Veterinary Research Institute. Izatnagar - 243 122 (U.P.), India (2023)
  20. 20) E.A. Fuentes, J.A. Achy, D.F. da Silva, A.C.G. Graboschii, J. de O. Bernardo, J.G. Joaquim, A.B. Fraga, and P.B. Escodro, "Ozone use in the treatment of subclinical mastitis in dairy cows," J. Dairy Res., 90 (4) 382-386 (2023) doi:10.1017/S0022029923000808
  21. 21) I. Jarto, J.A. Lucey, and K.E. Smith, "Impact of processing temperature on production of milk protein permeate during microfiltration of skim or whole milk," Int. J. Dairy Technol., 71 (4) 844-848 (2018) doi:10.1111/1471-0307.12542
  22. 22) F. Trimboli, M. Ragusa, C. Piras, V. Lopreiato, and D. Britti, "Outcomes from experimental testing of nonsteroidal anti-inflammatory drug (nsaid) administration during the transition period of dairy cows," Animals, 10 (10) 1832 (2020) doi:10.3390/ani10101832
  23. 23) S.K. Njoroge, "Effect of dairy farming practices on intake of antibiotic residues in milk consumed in kiambu county, kenya," (2020)
  24. 24) E. De Jong, "Enhancing antimicrobial stewardship on dairy farms with a focus on selective treatment of clinical mastitis," (2024) doi:10.11575/PRISM/42812
  25. 25) N. Verma, N. Agarwal, and L. Misra, "Review of some diseases of dairy animals and treatment by ethno-veterinary medicines," Adv. Med. Plants Res. Https//Doi. Org/11, 9-32 (2023)
  26. 26) R. Paramasivam, D.R. Gopal, R. Dhandapani, R. Subbarayalu, M.P. Elangovan, B. Prabhu, V. Veerappan, A. Nandheeswaran, S. Paramasivam, and S. Muthupandian, "Is amr in dairy products a threat to human health? an updated review on the origin, prevention, treatment, and economic impacts of subclinical mastitis," Infect. Drug Resist., 155-178 (2023) doi:10.2147/IDR.S384776
  27. 27) M. Babur, "Usage areas of ozone in animal health as an alternative treatment method: usage ozone in animal health," Rats, 2 (1) 33-39 (2024) doi:10.5281/zenodo.12601232
  28. 28) C. Sharma, N. Rokana, M. Chandra, B.P. Singh, R.D. Gulhane, J.P.S. Gill, P. Ray, A.K. Puniya, and H. Panwar, "Antimicrobial resistance: its surveillance, impact, and alternative management strategies in dairy animals," Front. Vet. Sci., 4 237 (2018) doi:10.3389/fvets.2017.00237
  29. 29) M.C. Beltrán, R.L. Althaus, A. Molina, M.I. Berruga, and M.P. Molina, "Analytical strategy for the detection of antibiotic residues in sheep and goat’s milk," Spanish J. Agric. Res., 13 (1) e0501–e0501 (2015) doi:10.5424/sjar/2015131-6522
  30. 30) S. Pyörälä, "Treatment of mastitis during lactation," Ir. Vet. J., 62 1-5 (2009)
  31. 31) B.D. Tampubolon, B. Basuki, P. Anggraeni, W.C. Anggundari, A.B. Mulyono, E.H. Purwanto, A. Nurhakim, D.R. Wahono, B. Prasetya, and A. Manab, "Trend of High Microbial Contamination in Livestock Milk in Indonesia and Distribution Mapping," in: E3S Web Conf., EDP Sciences, 2024: p. 2003 doi:10.1051/e3sconf/202452702003
  32. 32) M. Virto, G. Santamarina-García, G. Amores, and I. Hernández, "Antibiotics in dairy production: where is the problem?" Dairy, 3 (3) 541-564 (2022) doi:10.3390/dairy3030039
  33. 33) R. Kupczyński, M. Bednarski, M. Sokołowski, W. Kowalkowski, and K. Pacyga, "Comparison of antibiotic use and the frequency of diseases depending on the size of herd and the type of cattle breeding," Animals, 14 (13) 1889 (2024) doi:10.3390/ani14131889
  34. 34) R. Botondi, M. Lembo, C. Carboni, and V. Eramo, "The use of ozone technology: an eco–friendly method for the sanitization of the dairy supply chain," Foods, 12 (5) 987 (2023) doi:10.3390/foods12050987
  35. 35) O. Sonia, and L. Fernandez, "Ozone therapy as a novel complementary therapeutic approach in refractory idiopathic granulomatous mastitis," (July) 1-8 (2023) doi:10.3389/fmed.2023.1174372
  36. 36) L.V. de Faria, T.P. Lisboa, N. da S. Campos, G.F. Alves, M.A.C. Matos, R.C. Matos, and R.A.A. Munoz, "Electrochemical methods for the determination of antibiotic residues in milk: a critical review," Anal. Chim. Acta, 1173 (xxxx) (2021) doi:10.1016/j.aca.2021.338569
  37. 37) H. De Beer, A. Jacoby, and K. Jansen, "Microbial quality of milk , produced by small scale farmers in a peri-urban area in south africa," (February 2014) (2010)
  38. 38) D. Sert, and E. Mercan, "Assessment of powder flow, functional and microbiological characteristics of ozone-treated skim milk powder," Int. Dairy J., 121 (2021) doi:10.1016/j.idairyj.2021.105121
  39. 39) L.V. Costa, C. Gebara, O. De F, N.E. Freitas, A. Nascimento, C.S. Prado, I.A. Nunes, F. Oliveira, S. Duarte, M.E. Lage, F.R. De Alencar, B. Aparecida, S. Machado, K. Val, and S. Hodel, "Antibiotic residues in raw cow ’ s milk: a systematic review of the last decade," 1-14 (2024) doi:10.17605/OSF.IO/3ZMJC
  40. 40) A. Fadli, "Total plate count, most probable number index, and coliform bacteria of fresh cow milk in jember traditional market," 1 (1) 7-13 (2024)
  41. 41) A.C. Khanashyam, M.A. Shanker, A. Kothakota, N.K. Mahanti, and R. Pandiselvam, "Ozone applications in milk and meat industry," Ozone Sci. Eng., 44 (1) 50-65 (2022) doi:10.1080/01919512.2021.1947776
  42. 42) E. Prayitno, R. Hartanto, and D.W. Harjanti, "Physicochemical and microbiological appearance of sapera goat’s milk on frozen storage," J. Sain Peternak. Indones., 16 (4) 308-314 (2021) doi:10.31186/jspi.id.16.4.308-314
  43. 43) M.A. Khadre, A.E. Yousef, and J.‐. Kim, "Microbiological aspects of ozone applications in food: a review," J. Food Sci., 66 1242-1252 (2001) doi:10.1111/j.1365-2621.2001.tb15196.x
  44. 44) E. da P.E. Sitoe, L.R.D. Faroni, E.A.F. Fontes, E.R. de Alencar, P.R. Cecon, M.V. de Assis Silva, R. Pandiselvam, and F.C. Pacheco, "Effect of application of ozone at low-pressure on the technological properties, phenolic compounds, and antioxidant capacity of common beans during storage," LWT, 212 116964 (2024) doi:10.1016/j.lwt.2024.116964
  45. 45) E.S. Rohaeni, N. Ilham, R.A. Saptati, H. Sulistyawati, and P. Rahayu, "Developing a sustainable beef cattle business model for smallholder farms in south kalimantan ’ s drylands developing a sustainable beef cattle business model for smallholder farms in south kalimantan’s drylands," (February) (2024) doi:10.18280/ijsdp.190207
  46. 46) E. Sulistyowati, M.E. Malinda, W. Afrianti, D. Suherman, E. Soetrisno, S. Suharyanto, I. Badarina, and T. Akbarillah, "Sensory profile, total plate count and production cost of yogurt made from dairy goat milk added with ananas comosus and cucumis melo fruits," AGRITROPICA J. Agric. Sci., 7 (1) 34-38 (2024) doi:10.31186/j.agritropica.7.1.34-38
  47. 47) H. Pudjaatmaka, D. Fardiaz, and T. Agus, "KAMUS KIMIA," Ministry of Education and Culture, Jakarta, 1993
  48. 48) A. Maria, O. Id, and E. Romaszko, "Study protocol: the role of milk matrix lipids in programming the immunoreactivity of proteins derived from lactic acid bacteria," 1-17 (2024) doi:10.1371/journal.pone.0301477
  49. 49) S. Chun, Y. Chih, H. Lin, I.A. Aljuffali, and J. You, "Current pathogenic escherichia coli foodborne outbreak cases and therapy development," Arch. Microbiol., 199 (6) 811-825 (2017) doi:10.1007/s00203-017-1393-y
  50. 50) Y.F. Rosmi, A. Jati, A. Wasesa, D. Majid, and M.N. Jauhari, "Implementation of assistive hydraulic exoskeleton technology for children with cerebral palsy," 11 (03) 2305-2316 (2024) doi:10.5109/7236873
  51. 51) B.D. Tampubolon, P. Anggraeni , W.C. Anggundari , A.B. Mulyono, E.H. Purwanto, Novitasari, D.R. Wahono, B. Prasetya, A. Manab, and L.E. Radiati, "High distribution of antibiotic residues in dairy milk and used testing methods in Indonesia – review," J. Environ. Sci. Health B., 1-11 (2026) doi:10.1080/03601234.2026.2640802
  52. 52) A.N. Vlasova, and L.J. Saif, "Bovine immunology: implications for dairy cattle," 12 (June) 1-18 (2021) doi:10.3389/fimmu.2021.643206
  53. 53) J.A. Souza-Corrêa, M.A. Ridenti, C. Oliveira, S.R. Araújo, and J. Amorim, "Decomposition of lignin from sugar cane bagasse during ozonation process monitored by optical and mass spectrometries," J. Phys. Chem. B, 117 (11) 3110-3119 (2013) doi:10.1021/jp3121879
  54. 54) D.W. Hajanti, and D.G. Kusumaningrum, "The Effect of Ozone Exposure Duration on the Microbiological Quality and Nutritional Content of Ettawa Crossbred Goat Milk," J. Apl. Teknol. Pangan, 10 (1) 1-5 (2021). 1 doi:10.17728/jatp.7252
  55. 55) H. Hogeveen, K. Huijps, and T.J.G.M. Lam, "Economic aspects of mastitis: new developments.," N. Z. Vet. J., 59 (1) 16-23 (2011) doi:10.1080/00480169.2011.547165
  56. 56) W. Suriyasathaporn, Y.H. Schukken, M. Nielen, and A. Brand, "Low somatic cell count : a risk factor for subsequent clinical mastitis in a dairy herd," 1248-1255 (2000) doi:10.3168/jds.S0022-0302(00)74991-5
  57. 57) G. Lutviandhitarani, D.W. Harjanti, and F. Wahyono, "Green antibiotic daun sirih (piper betle l.) sebagai pengganti antibiotik komersial untuk penanganan mastitis," J. Agripet, 15 (1) 28-32 (2015)
  58. 58) B.D. Tampubolon, A.T. Setyoko, D.A. Bangun, R.B.F. Gultom, W.C. Anggundari, P. Anggraeni, N. Kusnandar, B. Prasetya, D.A. Susanto, D.R. Wahono, A. Manab, and L.E. Radiati, "Effect of Ozonation on Microbiological Quality, Nutritional Value, and Shelf Life of Fresh Cow’s Milk," International Journal of Design and Nature and Ecodynamics, 20 (5) 1093-1102 (2025) doi:10.18280/ijdne.200514
  59. 59) Lilliefors Significance Correction
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