Field experiments were conducted to examine the influence factors of cultivar, nitrogen application and irrigation on grain protein content, gluten content and grain hardness in three winter wheat cultivars under four levels of nitrogen and irrigation treatments. Firstly, the influence of cultivars and environment factors on grain quality were studied, the effective factors were cultivars, irrigation, fertilization, et al. Secondly, total nitrogen content around winter wheat anthesis stage was proved to be significant correlative with grain protein content, and spectral vegetation index significantly correlated to total nitrogen content around anthesis stage were the potential indicators for grain protein content. Accumulation of total nitrogen content and its transfer to grain is the physical link to produce the final grain protein, and total nitrogen content at anthesis stage was proved to be an indicator of final grain protein content. The selected normalized photochemical reflectance index (NPRI) was proved to be able to predict of grain protein content on the close correlation between the ratio of total carotenoid to chlorophyll a and total nitrogen content. The method contributes towards developing optimal procedures for predicting wheat grain quality through analysis of their canopy reflected spectrum at anthesis stage. Regression equations were established for forecasting grain protein and dry gluten content by total nitrogen content at anthesis stage, so it is feasible for forecasting grain quality by establishing correlation equations between biochemical constitutes and canopy reflected spectrum.