Banca de DEFESA: FRANCIS ARTHUR SECO PRANDO

Protocol for the preparation of chicken eggshell membrane (Gallus gallus domesticus) for use as a bioactive healing material in the treatment of injuries to the ocular surface.

Biological material, chicken egg, veterinary ophthalmology

Chicken eggshell membrane is a natural biomaterial that has gained increasing attention in the biomedical field due to its unique properties, versatility and sustainability. The objective of the present study was to develop a methodology for preparing chicken eggshell membrane (Gallus gallus domesticus) to be used as a bioactive healing material in the treatment of injuries to the ocular surface. 12 red chicken eggs were used, purchased at the local market. The method used was direct manual extraction of the shell membrane. After washing with physiological solution, the membrane was divided into 3 fragments (Control group; Treatment group 1 – 0.1% peracetic acid; Treatment group 2 – 0.2% peracetic acid) and each fragment divided into 4 parts. In all groups, the first part was placed in a vial containing 10% formaldehyde and sent for histological analysis. The second part was subdivided into 2 fractions, the first being inoculated in brain-heart infusion (BHI) broth and the second inoculated in Tetrationate broth.  The third part was inoculated on Sabouraud Agar with chloramphenicol. The fourth part was placed in a bottle containing sterile 99% glycerol and stored for 30 days at room temperature. In both treatments, the fragments were placed in an Erlenmeyer flask containing 20ml of peracetic acid (Perax Rio 0.2®) and kept under slight agitation for 20 minutes at room temperature. Then, the fragments were subjected to 3 washes of 15 minutes each , in 20ml of sterile PBS (pH 7.0), which was changed after each wash. This process was carried out at room temperature with slight agitation. After washing with PBS, each fragment was divided into 4 parts. The following steps were the same as those applied to the control group methodology. Histological analysis proved that both glycerolization and sterilization with peracetic acid do not promote cellular toxicity and destruction of tissue cells. In the microbiological analysis it was found that treatment with peracetic acid effectively reduces the microbial population in both concentrations. With the present study it was possible to conclude that manual extraction is a method that can be used to extract the eggshell membrane without compromising its structure or physical-chemical characteristics; Sterilization of the eggshell membrane is viable, especially when used at a concentration of 0.2% and glycerol is a good preservation medium.