Temanggung tobacco is a local tobacco, with high nicotine content. Nitrogen available in soil affects dry sliced yield and nicotine content. The experiment was conducted in glasshouse of IToFCRI Malang from March–August 2009. The treatment consist of six levels of N fertilizer (0, 1.62, 3.64, 4.86, 6.48, and 8.10 g N/plant), were arranged in randomized block design with three replications. Every threatment in one repli-cation received 2.70 g P2O5 + 1.35 kg manure. The results showed that physiological plant parameters (N-leaf, chlorophyll content, specific leaf weight, photosynthesis rate, respiration rate, light efficiency to CO2 re-duction (EF0), and coefficient of leaf maintenance respiration (CLMR)), dry slice yield, and nicotine content were affected by N rates. The response of these parameters on N fertilizer were expressed by closed qua-dratic curves, which maximum rate of N fertilizer were 7.89, 7.89, 54.75, 9.50, 26.25, 21.50, 10.00, 6.08, and 6.86 g N/plant respectively. Dry sliced yield were affected by chlorophyll content, respiration rate, and specific leaf weight. Nicotine content were affected by photosynthesis rate, CLMR, respiration rate, and chlo-rophyll content.