In this paper, in vivo spectra from 23 patients' blood samples with various Creatinine (Cr) concentration levels ranging from 0.96 to 12.5 mg/dL were measured using Fourier transform near-infrared spectrometer (FT-NIRS) and spectrum quantitative analysis method. Since Cr undergoes passive filtration, it serves as a key biomarker of kidneys function via the estimation of glomerular filtration rate. Thus, increased blood Cr concentration reflects impaired renal function. After spectra pre-processing and outlier exclusion, a spectral model was developed based on partial least squares regression (PLSR) method, wherein Cr concentrations correlated with filtered NIR spectra across several peaks, where Cr is known to absorb NIR light. Several statistical metrics were applied to estimate the model efficiency during data analysis. Comparison of spectra-derived concentrations to reference Cr measurements by the current gold-standard Jaffe's method held in hospital lab revealed a Cr prediction accuracy of 1.64 mg/dL with good correlation of R = 0.9. Bland-Altman plots were used to compare between our calculations and reference lab values and reveal minimal bias between the two. The finding presented the potential of FT-NIRS coupled with PLSR technique for Cr determination.