The estimation of leaf nitrogen concentration (LNC) in crop plants is an effective way to optimize nitrogen fertilizer management and to improve crop yield. The objectives of this study were to (1) analyse the spectral features, (2) explore the spectral indices, and (3) investigate a suitable modelling strategy for estimating the LNC of five species of crop plants (rice (Oryza sativa L.), corn (Zea mays L.), tea (Camellia sinensis), gingili (Sesamum indicum), and soybean (Glycine max)) with laboratory-based visible and near-infrared reflectance spectra (300–2500 nm). A total of 61 leaf samples were collected from five species of crop plant, and their LNC and reflectance spectra were measured in laboratories. The reflectance spectra of plants were reduced to 400–2400 and smoothed using the Savitzky–Golay smoothing (SG) method. The normalized band depth (NBD) values of all bands were calculated from SG-smoothed reflectance spectra, and a successive projections algorithm-based multiple linear regression (SPA-MLR) method was then employed to select the spectral features for five species. The SG-smoothed reflectance spectra were resampled using a spacing interval of 10 nm, and normalized difference spectral index (NDSI) and three-band spectral index (TBSI) were calculated for all wavelength combinations between 400 and 2400 nm. The NDSI and TBSI values were employed to calibrate univariate regression models for each crop species. The leave-one-out cross-validation procedure was used to validate the calibrated regression models. Study results showed that the spectral features for LNC estimation varied among different crop species. TBSI performed better than NDSI in estimating LNC in crop plants. The study results indicated that there was no common optimal TBSI and NDSI for different crop species. Therefore, we suggest that, when monitoring LNC in heterogeneous crop plants with hyperspectral reflectance, it might be appropriate to first classify the data set considering different crop species and then calibrate the model for each species. The method proposed in this study requires further testing with the canopy reflectance and hyperspectral images of heterogeneous crop plants.