Lower back pain (LBP) is one of the most common musculoskeletal complaints worldwide. Intervertebral disc degeneration (IDD) is considered to be a significant contributor to LBP; however, the mechanisms underlying IDD remain to be fully elucidated. One of the major features of IDD is the decreased content of type II collagen and proteoglycans in the nucleus pulposus (NP). The present study aimed to investigate the biochemical mechanisms of IDD at the microscopic level using Raman spectroscopy. Raman spectroscopy, based on inelastic scattering of light, is an emerging optical technique that may measure the chemical composition of complex biological samples, including biofluids, cells and tissues. In the present study, 30 NP tissue samples from 30 patients who were diagnosed with lumbar disc herniation and received spinal fusion surgery to relieve LBP were obtained and analyzed. Routine pre-operative 3.0T, T2-weighed MRI was used to classify the cases according to Pfirrmann grades and the T2 signal intensity value of the NP was measured. Subsequently, all NP samples were scanned and analyzed using a Laser MicroRaman Spectrometer at room temperature. The Raman spectral results demonstrated that the relative content of proteoglycans, expressed as the relative intensity ratio of two peaks (I₁₀₆₄/I₁₀₀₄), was significantly inversely correlated with the Pfirrmann grade (ρ=-0.6462; P<0.0001), whereas the content of collagen (amide I) was significantly positively correlated with the Pfirrmann grade (ρ=0.5141; P<0.01). In conclusion, the higher relative intensity of the ratio of two peaks (I₁₆₇₀/I₁₆₄₀; Amide I) represented a higher fractional content of disordered collagen, which suggested that the defective collagen structure may lead to NP abnormalities.