Near-infrared spectroscopy (NIR) is a non-destructive, fast, and low-cost method to measure the grain quality of different cereals. However, the feasibility for determining the critical biochemicals, related to the classifications for food, feed, and fuel products are not adequately investigated. Fourier-transform (FT) NIR was applied in this study to determine the eight biochemicals in four types of sorghum samples: hulled grain flours, hull-less grain flours, whole grains, and grain flours. A total of 20 hybrids of sorghum grains were selected from the two locations in China. Followed by FT-NIR spectral and wet-chemically measured biochemical data, partial least squares regression (PLSR) was used to construct the prediction models. The results showed that sorghum grain morphology and sample format affected the prediction of biochemicals. Using NIR data of grain flours generally improved the prediction compared with the use of NIR data of whole grains. In addition, using the spectra of whole grains enabled comparable predictions, which are recommended when a non-destructive and rapid analysis is required. Compared with the hulled grain flours, hull-less grain flours allowed for improved predictions for tannin, cellulose, and hemicellulose using NIR data. This study aimed to provide a reference for the evaluation of sorghum grain biochemicals for food, feed, and fuel without destruction and complex chemical analysis.