The extraction and determination of volatile compounds in mice urine were performed using headspace solid-phase microextraction and gas chromatography-mass spectrometry. In order to optimize the extraction conditions, experimental design was applied. A sample volume of 108 µl, a temperature of 148.6°C, and a time of 94 minutes were found to be the optimal conditions. Samples of male and female mouse urine were analyzed to determine volatile compound profiles. A total of 36 organic compounds including ketones, aldehydes, and terpenes were detected. The results revealed that compounds such as 2-isopropyl-4,5-dihydrothiazole, which is considered a male sexual pheromone, were only detected in male urine samples, whereas others like benzaldehyde were especially abundant in female mouse urine. A comparison of female samples corresponding to different stages of the estrous cycle was also performed.
