Correlation between eggshell membrane ultrastructure and respiration intensity

Abstract: Eggshell is a highly ordered and mineralized structure for egg respiration. The shell membrane with the organic layer has been one of the most important eggshell ultrastructures for the necessary channel of egg respiration. It is a high demand to determine the relationship between the ultrastructure of the eggshell membrane and the respiration intensity for the preservation and storage of eggs. In this research，the ultrastructure morphology of eggshell membranes was first characterized by a Scanning Electron Microscope (SEM). An image processing software was then utilized to extract the structural parameters from the ultrastructure image of the eggshell membrane. Therefore, a correlation model was established for the fiber diameter, porosity, the thickness of eggshell membranes, and the respiration intensity of the egg. The result showed that there was a significant difference between the strong (≥3 mg/(kg?h)) and weak (≤1 mg/(kg?h)) respiration intensity with the fiber diameter, porosity, and the thickness of the eggshell membranes. Specifically, there was a very significant positive correlation between the fiber diameter, the porosity of eggshell membranes, and the respiration intensity (P<0.01), whereas, a very significant negative correlation between the thickness of the eggshell membranes, and the respiration intensity of the egg (P<0.01). The membrane fibers were disorderly and tightly arranged in the eggshell ultrastructure under the strong respiration intensity. There were outstanding mucopolysaccharide tumor-like attachments on the outer membrane of eggs under the weak and medium respiration strength, whereas, the granular attachments on the eggs under the strong respiration strength. The fiber diameter of the outer eggshell membrane was larger than that of the inner, and the thickness of the outer eggshell membrane was 3-4 times that of the inner. The fibers of the inner eggshell membrane presented more slender and straighter than those of the outer. A partial least squares regression model was established, where the 10 characteristic parameters of porosity, fiber diameter, and thickness on both sides of the eggshell membrane were taken as the independent variables, and the egg breathing intensity as the dependent variable, as well as the correlation coefficient RP was 0.976 7. It was found that the ultrastructure of the eggshell membrane was used to better characterize the respiration intensity. A partial least square discriminant analysis was also conducted to take the respiration intensity as the classification, where the accuracy of the test set was 83.33%. The outer surface porosity on the outside of the outer membrane, the fiber diameter on the outside of the inner membrane, the thickness of the outer membrane, and the fiber diameter on the inside of the outer membrane greatly contributed to distinguish between different respiratory intensities. Among them, the porosity of the outer membrane was attributed to the great respiratory intensity. Consequently, the ultrastructure of the eggshell membrane posed better discrimination on the respiration intensity of fresh eggs in an air environment of 25℃. The finding can also offer a promising prospect for the storage and preservation of eggs, particularly for the protein extraction of eggshell membranes.