The Near Infrared Spectroscopy (NIRS) of 55 natural water samples were collected by a PbS detector with 8 nm resolution at wavelengths from 400 nm to 2 500 nm in air background and three quartz sample cells in thickness of 1 mm, 2 mm and 4 mm respectively were used to investigate the effect of different optical path difference. The nitrogen concentrations were measured by the Chinese National Standard Method (NSM), and the value of TN is used as the standard. The calibration models of the NIRS were established with Stepwise Multiple Linear Regression (SMLR) and Partial Least-squares (PLS) to analyze the TN concentration. Experimental results show that the correlation coefficient of the calibration model is 0.97 when the sample cell is 1 mm, and the Root Mean Square Error of Cross Validation (RMSECV) is 0.490 mg/L. In order to validate the feasibility of the NIRS method for TN measurement of water, the results of the NIRS method and NSM method have been compared to each other by t-test. The experimental results demonstrate that there is no significant difference between the NIRS method and the NSM by t-test on the condition of the significance level greater than 0.05;The NIRS method is promising for detecting TN in waters and wastewaters. Moreover,because of the strong influence of the water absorbance, 1 mm thickness sample cell is more suitable for near infrared spectral analysis of TN.