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Sterilization of Human Amniotic Membrane Using an Ozone Hydrodynamic System

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Abstract

Human amniotic membrane (hAM) is an important biomaterial for Tissue Engineering, due to its great regenerative properties and potential use as a scaffold. The most used procedure to sterilize biomaterials is gamma-irradiation, but this method can affect several properties, causing damage to the structure and reducing the growth factors. The present work evaluated the efficiency of a new method based on ozonated dynamic water for hAM sterilization. HAM fragments were experimentally contaminated with Staphylococcus aureus, Escherichia coli, Candida albicans, Staphylococcus epidermidis, and Clostridium sporogenes (106 CFU/mL) and submitted to sterilization process for 5, 10 and 15 min. The analyses did not reveal microbial activity after 10 min for S. aureus and C. sporogenes and after 15 min for E. coli and S. epidermidis. The microbial activity of C. albicans was reduced with the exposure time increase, but the evaluated time was insufficient for complete sterilization. The depyrogenation process was investigated for different ozonation times (15, 20, 25, 30, and 35 min) to evaluate the ozone sterilization potential and presented promising results after 35 min. The ozone effect on hAM structure was evaluated by histological analysis. A decrease in epithelium average thickness was observed with the exposure time increase. Furthermore, some damage in the epithelium was observed when hAM was exposed for 10 and 15 min. It can indicate that ozone, besides being effective in sterilization, could promote the hAM sample’s de-epithelization, becoming a possible new method for removing the epithelial layer to use hAM as a scaffold.

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Acknowledgments

B. A. Kawata acknowledges CAPES (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - Brazil) - Financing Code 001 and the Santa Casa de Misericórdia in Pindamonhangaba-SP, Brazil. P. A. Laurindo Igreja Marrafa acknowledges CAPES (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - Brazil) for the Doctorate scholarship - Financing Code 001. S. Móbille Awoyama acknowledges FAPESP (Fundação de Amparo à Pesquisa do Estado de São Paulo) for the PIPE Program Phase 1 financing (Process No. 2019/22907-4). A. Barrinha Fernandes and C. J. de Lima acknowledge the Anima Institute (AI), Universidade Anhembi Morumbi, São Paulo-SP, Brazil. A. Barrinha Fernandes thanks CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico) for the productivity fellowship (Process No. 310708/2021-4).

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  1. Center for Innovation, Technology and Education – CITÉ, Parque de Inovação Tecnológica de São José dos Campos, São José dos Campos, SP, 12247-016, Brazil

    Túlia Botelho, Bianca Akemi Kawata, Silvia Móbille Awoyama, Pedro Augusto Laurindo Igreja Marrafa, Henrique Cunha Carvalho, Carlos José de Lima & Adriana Barrinha Fernandes

  2. Faculdade Santo Antônio – FSA, Caçapava, SP, Brazil

    Túlia Botelho

  3. Universidade Anhembi Morumbi – UAM, Biomedical Engineering Institute, São Paulo, SP, 04546-001, Brazil

    Bianca Akemi Kawata, Pedro Augusto Laurindo Igreja Marrafa, Carlos José de Lima & Adriana Barrinha Fernandes

  4. Centro Universitário FUNVIC – UNIFUNVIC, College of Pharmacy, Pindamonhangaba, SP, 12412-825, Brazil

    Silvia Móbille Awoyama

  5. Universidade Tecnológica Federal do Paraná – UTFPR, Campo Mourão, PR, 87301-899, Brazil

    Henrique Cunha Carvalho

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Botelho, T., Kawata, B.A., Móbille Awoyama, S. et al. Sterilization of Human Amniotic Membrane Using an Ozone Hydrodynamic System. Ann Biomed Eng 52, 1425–1434 (2024). https://doi.org/10.1007/s10439-024-03467-3

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