The invention relates to the field of optical colorimetric analysis, in particular to a monochromator for microplate readers. The monochromator comprises a plurality of monochromatic light assemblies and light guide fiber components. The light guide fiber components focus monochromatic light generated by all the monochromatic light assemblies to colorimetric cups corresponding to the microplate readers respectively through different optical paths, and optical signals are converted into electrical signals through a photocell. The light guide fiber components comprise multi-channel optical fiber bundles and optical focus mechanisms. The input ends of the multi-channel optical fiber bundles receive the monochromatic light output by the monochromatic light assemblies, and the optical focus mechanisms are located at the output ends of the multi-channel optical fiber bundles and are focused to the centers of the colorimetric cups of the corresponding channels. By means of the monochromator, two to twelve bundles of monochromatic light of any input wave length and even in energy distribution can be obtained at any time, the response time is short, and no mechanical movement mechanism is needed.