This invention relates to a novel method and a device for determining middle and protein bound uremic toxins in the biological fluids. More specifically the present invention relates to an optical method utilizing fluorescence preferable fluorescence of the spent dialysate and a specific model including a unique set of optical spectral components at certain wavelengths to determine preferable on line the concentration of the middle and protein bound uremic toxins such as beta2 microglobulin (B2M) and indoxyl sulfate (IS).