Large-scale isolation of middle and higher molecular weight uremic toxins

Reverse osmosis (40 bar) using membranes with a nominal cut-off of 500 Dalton is a useful method for the desalting and concentration of hemofiltrates (20-30 L) or even dialysates (120-240 L) from patients with end-stage renal failure. The removal of electrolytes and lower molecular weight solutes from the middle and higher molecular weight fractions can be carried out in one step. The freeze-dried residues show characteristic differences in their molecular weight distributions which are dependent upon their origin (Cuprophane dialysates or RP-6-hemofiltrates). Subfractionation is performed using Sephadex G-15 macrocolumns (2 m x 5 cm), the LKB-Ultrogel AcA 54 and Sephacryl S-200 as well as ion exchange chromatography. The fractions are characterized by their ability to suppress the incorporation of 3H-thymidine into rat bone-marrow as well as HeLa cell cultures. The greatest inhibitory activity originates from those fractions which are not dialysable using Visking tubes in vitro. The results indicate that the middle and higher molecular weight spectra do in fact overlap, suggesting that also higher molecular weight substances are involved in uremic intoxication.