Abstract
Efforts are underway to develop technology for automatically determining the sex of chick embryos, aimed at establishing a stable and efficient poultry farming system while also addressing animal welfare concerns. This study investigated the possibility of chick sexing through blood analysis using Raman spectroscopy. Raman spectra were obtained from whole blood and its constituents, such as red blood cells (RBCs) and blood plasma, collected from chicks aged 1-2 days, using a 785-nm excitation wavelength. Principal component analysis (PCA) revealed statistically significant sex-dependent spectral variations in whole blood and RBCs, whereas blood plasma showed less clear dependency. These spectral differences between male and female chicks were attributed to differences in the proportion of spectral components from oxygenated (oxy-) and deoxygenated (deoxy-) RBCs, with males exhibiting a slightly stronger contribution of oxy-RBCs compared to females. This reflects the higher oxygen affinity of hemoglobin (Hb) in males compared to females. A model for discriminating chick sex was built using the ratios of certain Raman band characteristics of oxy-RBCs and deoxy-RBCs, achieving a sensitivity of 100%. This spectroscopic method holds promise for developing technology to discriminate the sex of early chicken embryos in ovo by detecting differences in oxygen saturation of RBCs based on sex.