Abstract Spectral uncertainty is one of the most prominent spectral characteristics of hyperspectral images. Compared to the process of remote sensing hyperspectral imaging, hyperspectral imaging under land-based imaging conditions has the characteristics of variable detection directions, random imaging times, and complex environmental conditions, resulting in increased spectral uncertainty of targets in land-based hyperspectral images. The spectral uncertainty of the target mainly refers to the phenomenon of “Same spectral reflectance but different objects” and “Same object but different spectral reflectance” which poses significant difficulties for subsequent hyperspectral image target detection and recognition. In order to analyze the spectral uncertainty of hyperspectral images in land-based applications and address the issue of spectral uncertainty in similar targets, a spectral uncertainty evaluation index based on standard deviation vector was proposed. For the overall spectral separability between different types of targets, a quantitative index based on Jaccard Distance (JD-SSI) is proposed to measure the spectral separability of different kinds of targets. The experiment focused on grassland and its four typical camouflage materials, analyzing the spectral intra class differences and inter class separability of each target with grassland. It is a fundamental work for studying the spectral characteristics of land-based hyperspectral images, providing a new approach for subsequent spectral band extraction, hyperspectral image classification, and target detection tasks.