Define the Principle behind Cyanmethemoglobin Method?
The principle behind the cyanmethemoglobin method is as follow:
Principle The haemoglobin is treated with a reagent containing potassium cyanide and sodium carbonate. With haemoglobin ferricyanide forms methaemoglobin which is converted to cyanmethaemoglobin or haemiglobincyanide (HiCN) with cyanide. The absorbance is measured at 540 nm. The reaction is presented herewith:
Haemoglobin + potassium ferricyanide ------→ Methaemoglobin
Methaemoglobin + potassium cyanide ------→ Cyanmethaemoglobin (HiCN)
in solution the ferrous ions of the haemoglobin are oxidized to the ferric state by potassium fericyanide to form methaemoglobin. In turn methaemoglobin reacts with cyanide ions (CN-) provided by potassium cyanide to form HiCN. The time required for full colour development is shortened by if dihydrogen potassium phosphate is substituted for sodium bicarbonate in the classic Drabkin's reagent. The colour is stable for more than 24 hours in well stoppered tubes kept in dark. An HiCN standard can be used to calibrate the spectrophotometer, The concentration of HiCN in the sample is calculated from the absorbance in a calibrated spectrophotometer or alternatively with the help of a standard curve.