Collagen can be identified as the most abundant fibrous protein in the kingdom of Animalia. Approximately one-third of the total protein content in the human body is collagen. Being an important biopolymer, collagen consists of a unique repeating unit of tripeptide domain. This tripeptide domain is made from three specific amino acids called glycine, proline and hydroxyproline. The repeating units polymerized together to make the polypeptide chain of collagen. Collagen has certain structural integrity which is necessary to maintain proper functioning of body tissues. Characterization of these structural features and specific functionalities is important to evaluate performances of collagen in tissues like skin, cartilage, and tendons. Several conventional characterization techniques like X-ray diffraction (XRD), thermal analysis, electron microscopy and spectroscopic methods are utilized to analyze the structural and physiochemical properties of collagen. Many of these characterization techniques pose certain challenges due to the complex structure and sensitivity of collagen towards many external factors. However, vibrational spectroscopic characterization techniques like Fourier Transform Infrared (FTIR) spectroscopy and Raman spectroscopy can overcome most of these challenges and characterize collagen effectively. Both FTIR and Raman spectroscopic techniques aid to obtain comprehensive understanding about native and modified collagen states. These techniques collectively provide detailed insights into the collagen’s chemical structure, bonding types, molecular interactions and conformational changes. Most importantly, these techniques facilitate rapid, highly sensitive and nondestructive approaches to characterize collagen. These specific advantages make FTIR and Raman spectroscopic techniques preferable over many other conventional characterization methods. In this chapter, characterization of collagen using Fourier Transform Infrared (FTIR) spectroscopy and Raman spectroscopy is discussed. The major characteristic peaks observed in both techniques are mentioned while explaining their molecular origins. The sections are further clarifying the different spectral shifts and changes in intensity ratios due to structural changes in collagen-based systems. This chapter emphasizes the importance of employing FTIR and Raman spectroscopic methods to characterize collagen’s structure, denaturation, and chemical modifications.