The appropriateness of using cyanocobalamin as calibration standards in competitive binding assays of vitamin B12

Abstract The forms of vitamin B 12 that have been found in foods are cyanocobalamin, hydroxocobalamin, sulphitocobalamin, adenosylcobalamin and methylcobalamin. These cobalamins, with the exception of sulphitocobalamin, are also known to be present in serum and plasma. The addition of sodium cyanide, potassium cyanide, sodium metabisulphite or sodium nitrite in the extraction procedure employed for the determination of vitamin B 12 , however, can convert these cobalamins into derivatives which include dicyanocobalamin and nitritocobalamin. It was observed that cyanocobalamin has a binding affinity for hog intrinsic factor equal to that of methylcobalamin, dicyanocobalamin and nitritocobalamin, but not to that of hydroxocobalamin, sulphitocobalamin and adenosylcobalamin. Thus, these latter three cobalamins cannot be measured accurately using a competitive binding assay which employs hog intrinsic factor as its binding protein and cyanocobalamin as the calibration standards unless they are converted to cyanocobalamin, methylcobalamin, dicyanocobalamin or nitritocobalamin before the assay.