Nowadays, it is still a challenge for commercial nitrate sensors to meet the requirement of high accuracy in a complex water. Based on deep-ultraviolet spectral analysis and a regression algorithm, a different measuring method for obtaining the concentration of nitrate in seawater is proposed in this paper. The system consists of a deuterium lamp, an optical fiber splitter module, a reflection probe, temperature and salinity sensors, and a deep-ultraviolet spectrometer. The regression model based on weighted average kernel partial least squares (WA-KPLS) algorithm together with corrections for temperature and salinity (TSC) is established. After that, the seawater samples from Western Pacific and Aoshan Bay in Qingdao, China with the addition of various nitrate concentrations are studied to verify the reliability and accuracy of the method. The results show that the TSC-WA-KPLS algorithm shows the best results when compared against the multiple linear regression (MLR) and ISUS (in situ ultraviolet spectrophotometer) algorithms in the temperatures range of 4–25 °C, with RMSEP of 0.67 µmol/L for Aoshan Bay seawater and 1.08 µmol/L for Western Pacific seawater. The method proposed in this paper is suitable for measuring the nitrate concentration in seawater with higher accuracy, which could find application in the development of in-situ and real-time nitrate sensors.