Nitrogen metabolism is one of the basic physiological processes of plants and is directly related to growth. Nitrogen assimilation (N) by plants requires the absorption of nitrate (NO3–) from the soil and the reduction to nitrite (NO2–), and then the conversion of NO2– to ammonium (NH4+). This is followed by the inclusion of NH4+ into organic compounds. The activity of nitrate reductase (NR), the essential enzyme for nitrogen assimilation, is regarded as an indication of nitrogen metabolism Plants can take up nitrogen as both nitrate (NO3-) and ammonium (NH4+), but the absorption characteristics of each nitrogen source differ between plant types. There is insufficient information regarding chestnut cultivars and variants’ nitrogen assimilation capability. In this article, the changes seen during the vegetation period in the capacity of nitrogen assimilation were evaluated in four chestnut cultivars/genotypes (‘Alimolla’, ‘Sarıkestane’, ‘N-23-1’ and ‘Erfelek’), which are prominent in different areas where they are cultivated under the same ecological conditions. In the leaf samples which are taken up monthly from cultivars/genotypes between May and October, nitrate reduction activity (NRA), total nitrogen (N), chlorophyll, and biomass parameters were detected. The NRA of the samples was determined according to the in vivo test. This spectrophotometric method is based on the determination of the absorbance of nitrite (NO2-) formed by the reduction of nitrate in the incubation medium. The results of the study showed that the nitrogen assimilation ability of chestnut cultivars/genotypes differed from each other. These results are also closely related to other parameters such as chlorophyll content and biomass. The findings are important in terms of revealing the physiological characteristics of chestnut cultivars and learning their functioning.