Rapid separation and identification of myoglobin and hemoglobin in urine by centrifugation through a...

Urinary myoglobin (Mr = 16 500) is readily separated from hemoglobin (Mr = 64 000) by centrifugation through a microconcentrator membrane with a 30 000-Da cutoff. Myoglobin, but not hemoglobin, will pass through the membrane, after which each fraction may be separately analyzed by spectrophotometry. This technique is advantageous over direct spectrophotometry of urine because it is not restricted to analyzing the oxy forms of these hemoproteins and obviates the difficulty of discriminating small (3 nm) differences in their spectra. In addition, this method of separation is more complete than that attained by differential "salting out" with ammonium sulfate, and it is simpler than gel filtration or ultrafiltration under reduced pressure.