A greenhouse field experiment involving tomato (Solanum lycopersicum) was performed using different nitrogen (N) management regimes: sole application of differing rates of chemical N fertilizer (SC) (SC treatments: N0, N1, N2, and N3) and combined application of manure and chemical N fertilizer (MC) (MC treatments: MN0, MN1, MN2, and MN3). These were used to understand the relationship between comprehensive fruit composition, yield, and N fractions (soil mineral N; soil soluble organic N; soil microbial biomass N, and soil fixed ammonium) under greenhouse conditions. The results showed that the MC treatments significantly increased vitamin C and soluble sugar content compared with SC treatments. In addition, the MN2 treatment produced a high yield and had a positive effect on fruit composition. The N3 (563 kg N/ha) and MN3 (796 kg N/ha) treatments resulted in a high loss of N below the root zone (0–30 cm), consequently reducing N use efficiency. Soil mineral N, soil soluble organic N, and soil fixed ammonium tended to be higher during the first fruiting period, whereas soil microbial biomass N tended to be higher during the second fruiting period. MC treatments significantly increased the N fraction in the 0- to 30-cm soil layer; N fractions tended to be higher with the MN2 treatment. According to an optimum regression equation, soil fixed ammonium during the first fruiting period and soil microbial biomass N during the second fruiting period had a more significant influence on tomato yield and fruit composition. Overall, application MC at an appropriate rate (MN2: 608 kg N/ha) is a promising approach to achieving high yields and optimum taste, and it offers a more sustainable fertilizer management strategy compared with chemical N fertilization.