Ukrainian Antarctic Journal

Vol 23 No 1(30) (2025): Ukrainian Antarctic Journal
Articles

Collagenolytic enzymes from Antarctic hydrobionts: a promising biotechnological approach for accelerating purulent-necrotic wound healing

Nataliia Raksha
Educational and Scientific Center “Institute of Biology and Medicine” of Taras Shevchenko National University of Kyiv, Kyiv, 01601, Ukraine
Tetiana Halenova
Educational and Scientific Center “Institute of Biology and Medicine” of Taras Shevchenko National University of Kyiv, Kyiv, 01601, Ukraine
Tetiana Vovk
Educational and Scientific Center “Institute of Biology and Medicine” of Taras Shevchenko National University of Kyiv, Kyiv, 01601, Ukraine
Tetiana Koval
Educational and Scientific Center “Institute of Biology and Medicine” of Taras Shevchenko National University of Kyiv, Kyiv, 01601, Ukraine
Nataliia Nikitina
Educational and Scientific Center “Institute of Biology and Medicine” of Taras Shevchenko National University of Kyiv, Kyiv, 01601, Ukraine
Liudmyla Stepanova
Educational and Scientific Center “Institute of Biology and Medicine” of Taras Shevchenko National University of Kyiv, Kyiv, 01601, Ukraine
Tetyana Beregova
Educational and Scientific Center “Institute of Biology and Medicine” of Taras Shevchenko National University of Kyiv, Kyiv, 01601, Ukraine
Olexiy Savchuk
Educational and Scientific Center “Institute of Biology and Medicine” of Taras Shevchenko National University of Kyiv, Kyiv, 01601, Ukraine
Published July 29, 2025
Keywords
  • complicated wounds,
  • marine organisms,
  • proteinases
How to Cite
Raksha, N., Halenova, T., Vovk, T., Koval, T., Nikitina, N., Stepanova, L., Beregova, T., & Savchuk, O. (2025). Collagenolytic enzymes from Antarctic hydrobionts: a promising biotechnological approach for accelerating purulent-necrotic wound healing. Ukrainian Antarctic Journal, 23(1(30), 51-63. https://doi.org/10.33275/1727-7485.1.2025.743

Abstract

The objective of this study was to isolate the fractions of proteolytic enzymes from Antarctic hydrobionts, specifically Sterechinus neumayeri and Odontaster validus, and assess their wound-healing potential on purulent-necrotic wounds in rats. The purification procedure included ammonium sulfate precipitation and ion-exchange chromatography on Q-Sepharose. The enzymes derived from both hydrobionts exhibited significant collagenolytic activity, averaging around 14 rel. units per mg of total proteins, and were different in molecular weight and substrate specificity. Enzyme- electrophoresis with gelatin as a substrate revealed that enzymes from S. neumayeri were mostly above 40 kDa, while those from O. validus ranged between 15–35 kDa. The inhibitory analysis demonstrated that O. validus enzymes were predominantly metal-dependent proteases (67%), whereas S. neumayeri contained both serine proteinases (44%) and metal-dependent enzymes (34%). This enzymatic heterogeneity suggests broad substrate specificity, which is advantageous for cleansing various wound types. The enzymes exhibited optimal activity at an alkaline pH (8.0–9.0), which
aligns with the pH conditions of chronic and infected wounds, making them particularly effective in the early stages of wound healing. To evaluate the therapeutic potential, a composition based on the mixture of collagenolytic enzymes from both hydrobionts was applied to purulent-necrotic wounds in rats. The group of rats treated with the composition showed faster wound reduction than the control group. By the 12th day, wound size in the treated group had reduced to 24.4 ± 6.7 mm2, significantly smaller than in the control group – 49.8 ± 6.9 mm2. Complete epithelialization occurred by day 27 in the treated group, while the control group achieved full healing by day 30. The results confirm that collagenolytic enzymes from Antarctic hydrobionts are effective in enzymatic debridement, promoting faster wound healing. Combining serine and metal-dependent proteases enhances the range of protein breakdown, improving the efficiency of wound cleansing. The enzymes' high activity at alkaline pH is beneficial for treating both acute and chronic wounds, with enzyme activity decreasing as healing progresses, minimizing potential damage to healthy tissues.

References

  1. Anjum, K., Abbas, S. Q., Akhter, N., Shagufta, B. I., Shah, S. A. A., & Hassan, S. S. U. (2017). Emerging biopharmaceuticals from bioactive peptides derived from marine organisms. Chemical Biology & Drug Design, 90(1), 12–30. https://doi.org/10.1111/cbdd.12925
  2. Bennison, L. R., Miller, C. N., Summers, R. J., Minnis, A. M. B., Sussmann, G., & McGuiness, W. (2017). The pH of wounds during healing and infection: a descriptive literature review. Wound Practice and Research, 25(2), 63–69.
  3. Bradford, M. M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72(1–2), 248–254. https://doi.org/10.1016/0003-2697(76)90527-3
  4. Carroll, A. R., Copp, B. R., Davis, R. A., Keyzers, R. A., & Prinsep, M. R. (2022). Marine natural products. Natural Product Reports, 39(1), 1122–1171. https://doi.org/10.1039/D1NP00076D
  5. Cavicchioli, R., Charlton, T., Ertan, H., Mohd Omar, S., Siddiqui, K.S., & Williams, T. J. (2011). Biotechnological uses of enzymes from psychrophiles. Microbial Biotechnology, 4(4), 449–460. https://doi.org/10.1111/j.1751-7915.2011.00258.x
  6. Gethin, G. (2007). The significance of surface pH in chronic wounds. Wounds UK, 3(3), 52–56.
  7. Han, G., & Ceilley, R. (2017). Chronic wound healing: A review of current management and treatments. Advances in Therapy, 34, 599–610. https://doi.org/10.1007/s12325-017-0478-y
  8. Hoyoux, A., Jennes, I., Dubois, P., Genicot, S., Dubail, F., François, J. M., Baise, E., Feller, G., & Gerday, C. (2001). Cold-adapted β-galactosidase from the Antarctic psychrophile Pseudoalteromonas haloplanktis. Applied and Environmental Microbiology, 67(4), 1529–1535. https://doi.org/10.1128/AEM.67.4.1529-1535.2001
  9. Huang, Y.-Z., Jin, Z., Wang, Z.-M., Qi, L.-B., Song, S., Zhu, B.-W., & Dong, X.-P. (2021). Marine bioactive compounds as nutraceutical and functional food ingredients for potential oral health. Frontiers in Nutrition, 8, 686663. https://doi.org/10.3389/fnut.2021.686663
  10. Kusampudi, S., Meganathan, V., Keshava, S., & Boggaram, V. (2023). Purification and characterization of a serine protease from organic dust and elucidation of its inductive effects on lung inflammatory mediators. American Journal of Physiology – Lung Cellular and Molecular Physiology, 325(1), L74–L90. https://doi.org/10.1152/ajplung.00309.2022
  11. Martins, A., Vieira, H., Gaspar, H., & Santos, S. (2014). Marketed marine natural products in the pharmaceutical and cosmeceutical industries: Tips for success. Marine Drugs, 12(2), 1066–1101. https://doi.org/10.3390/md12021066
  12. Mayer, A. M. S., Rodríguez, A. D., Taglialatela-Scafati, O., & Fusetani, N. (2013). Marine pharmacology in 2009–2011: Marine compounds with antibacterial, antidiabetic, antifungal, anti-inflammatory, antiprotozoal, antituberculosis, and antiviral activities; affecting the immune and nervous systems, and other miscellaneous mechanisms of action. Marine Drugs, 11(7), 2510–2573. https://doi.org/10.3390/md11072510
  13. Moore, S., & Stein, W. H. (1954). A modified ninhydrin reagent for the photometric determination of amino acids and related compounds. Journal of Biological Chemistry, 211(2), 907–913. https://doi.org/10.1016/S0021-9258(18)71178-2
  14. Munilla-Morán, R., & Stark, J. R. (1989). Protein digestion in early turbot larvae, Scophthalmus maximus (L.). Aquaculture, 81(3–4), 315–327. https://doi.org/10.1016/0044-8486(89)90156-7
  15. Ni, X., Chi, Z., Ma, C., & Madzak, C. (2008). Cloning, characterization, and expression of the gene encoding alkaline protease in the marine yeast Aureobasidium pullulans 10. Marine Biotechnology, 10(3), 319–327. https://doi.org/10.1007/s10126-007-9067-4
  16. Ostapchenko, L., Savchuk, O., & Burlova-Vasilieva, N. (2011). Enzyme electrophoresis method in analysis of active components of haemostasis system. Advances in Bioscience and Biotechnology, 2(1), 20–26. https://doi.org/10.4236/abb.2011.21004
  17. Park, P.-J., Lee, S.-H., Byun, H.-G., Kim, S.-H., & Kim, S.-K. (2002). Purification and characterization of a collagenase from the mackerel, Scomber japonicus. Korean Society for Biochemistry and Molecular Biology – BMB Reports, 35(6), 576–582. https://doi.org/10.5483/BMBRep.2002.35.6.576
  18. Raksha, N., Halenova, T., Vovk, T., Beregova, T., Maievska, T., Tomchuk, V., Savchuk, O., & Ostapchenko, L. (2023). Isolation and partial characterization of serine proteases from jellyfish of the Antarctic region. Journal of Applied Biology and Biotechnology, 11(2), 144–150. https://doi.org/10.7324/JABB.2023.110214
  19. Raksha, N., Udovychenko, I., Halenova, T., Vovk, T., Savchuk, O., & Ostapchenko, L. (2020). Purification and biochemical characterization of fibrino(geno)lytic enzymes from tissues of Antarctic hydrobionts. Ukrainian Antarctic Journal, (1), 69–81. https://doi.org/10.33275/1727-7485.1.2020.380
  20. Robbins, K. C., & Summaria, L. (1970). Human plasminogen and plasmin. Methods in Enzymology, 19, 184–199. https://doi.org/10.1016/0076-6879(70)19013-6
  21. Saggu, S. K., Bala, R., Hora, R., & Mishra, P. C. (2023). Purification and characterization of a high molecular weight serine protease from Microbacterium paraoxydans sp. SKS10. Biotechnology and Applied Biochemistry, 70(5), 1741–1753. https://doi.org/10.1002/bab.2472
  22. Sarmiento, F., Peralta, R., & Blamey, J. M. (2015). Cold and hot extremozymes: Industrial relevance and current trends. Frontiers in Bioengineering and Biotechnology, 3, 148. https://doi.org/10.3389/fbioe.2015.00148
  23. Stricklin, G. P., & Hibbs, M. S. (1988). Biochemistry and Physiology of Mammalian Collagenases. In M. E. Nimni (Ed.), Collagen (1st ed., pp. 20). CRC Press. https://doi.org/10.1201/9781351070799-8
  24. Thomas, D. C., Tsu, C. L., Nain, R. A., Arsat, N., Fun, S. S., & Lah, N. A. S. N. (2021). The role of debridement in wound bed preparation in chronic wound: A narrative review. Annals of Medicine and Surgery, 71 (Suppl. 1), 102876. https://doi.org/10.1016/j.amsu.2021.102876
  25. Vijay Kumar, E., Srijana, M., Kiran Kumar, K., Harikrishna, N., & Reddy, G. (2011). A novel serine alkaline protease from Bacillus altitudinis GVC11 and its application as a dehairing agent. Bioprocess and Biosystems Engineering, 34(4), 403–409. https://doi.org/10.1007/s00449-010-0483-x
  26. Wanderley, M. C. A., Wanderley Duarte Neto, J. M., de Lima Filho, J. L., de Albuquerque Lima, C., Teixeira, J. A. C., & Porto, A. L. F. (2017). Collagenolytic enzymes produced by fungi: a systematic review. Brazilian Journal of Microbiology, 48(1), 13–24. https://doi.org/101016/j.bjm.2016.08.001