Raman spectroscopy is a widely used analytical technique; however, the choice of container material can significantly affect repeatability due to noise and unwanted Raman bands. This paper describes the interaction of excitation radiation with the sample’s environment and the materials surrounding or containing it. Polytetrafluoroethylene (PTFE) and Barium Sulphate are analyzed as contact surfaces with the containers. We aim to investigate how different container materials affect the Raman spectrum and identify the optimal material to minimize measurement bias. We compared the Raman spectra of containers made of dissimilar materials, including plastic, glass, quartz, and aluminum, to assess their effects on the spectra, and we propose a container design that enhances Raman spectroscopy signals. We consider Raman bands from biological samples of glucose (in water) and urine (urea and creatinine), which serve as references for comparison in the in vitro analysis of these substances across different containers and backgrounds.