Mise en évidence et évaluation des “moyennes molécules” de la taille de la vitamine B 12 présentes dans les liquides biologiques de sujets normaux et de patients urémiques

Abstract

Determination of “middle molecules” presenting vitamin B 12 molecular size in normal and uremic body fluids Uremic solutes with the molecular size of vitamin B 12 are assumed to be toxic. An analytical method is proposed to detect and separate these solutes in body fluids using two combined techniques: gel filtration on Sephadex G-15 and ion-exchange chromatography on DEAE-Sephadex A-25. The vitamin B 12 molecular size has been localized by ultrafiltration through membranes with a defined cut-off. Normal and uremic body fluids (urine, plasma, hemodialysis fluid) have been separated into 9 ultraviolet-absorbing peaks (a to i) by high-speed gel filtration. Peaks b and e present the molecular size of vitamin B 12, 10–15 Å molecular diameter in pH 7 aqueous solution. Peak b, which correlates with uremic neuropathy, is separated into 6 sub-peaks (b 1 to b 6) by ion-exchange chromatography, sub-peak b 4.2 is the only one to correlate with uremic neuropathy. The coefficient of variation in the integrated area of a single peak is 16%. This method gives the chromatographic profile of the vitamin B 12 molecular size content from 500 μl of uremic plasma or 100 μl of normal urine within one hour.