The invention provides a device and a method of multispectral imaging by diffraction based on a Nipkow disk, belonging to the technical field of imaging spectroscopy. The device comprises a diffraction lens, a chromatography and imaging system consisting of a light shield, the Nipkow disk and a micro-lens array disk, a detector, a relative position fixing mechanism, a first and a second rotation driving mechanisms and a linear actuator, wherein, when the relative position fixing mechanism is unlocked, the first and the second rotation driving mechanisms can drive the light shield, the micro-lens array disk and the Nipkow disk to rotate respectively; when the relative position fixing mechanism is locked up, a first driving mechanism can drive the chromatography and imaging system to scan the focal plane in the direction of an incident image so as to directly acquire the two-dimensional image and the spectral information of the incident ray at a single wavelength; and the linear actuatorcan drive the chromatography and imaging system to scan along the optical axis of the diffraction lens so as to acquire the image and the spectral information of incident rays at all wavelengths within the detected spectral range. The invention has the characteristics of low stray light, high spectral resolution and high spatial resolution.