Obesity is becoming a leading cause of health problems world-wide. Obesity and overweight are associated with the structural and chemical changes in tissues; however, few methods exist that allow for concurrent measurement of these changes. Using Brillouin and Raman microspectroscopy, both the mechanical and chemical differences can be assessed simultaneously. We hypothesized that Brillouin spectroscopy can measure the adipose tissues’ stiffness, which increases in obesity. Samples of brown and white adipose tissues obtained from control and diet-induced obese adult rats were analyzed. The results show that both adipose tissues of the obese group exhibit a greater high-frequency longitudinal elastic modulus than the control samples, and that the brown fat is generally stiffer than white adipose. The Raman spectra indicates that the lipids’ accumulation in adipose tissue outpaces the fibrosis, and that the high-fat diet has a greater effect on the brown adipose than the white fat. Overall, the powerful combination of Brillouin and Raman microspectroscopies successfully assessed both the mechanical properties and chemical composition of adipose tissue simultaneously for the first time. The results indicate that the adipose tissue experiences an obesity-induced increase in stiffness and lipid content, with the brown adipose tissue undergoing a more pronounced change compared to white adipose.