Comparisons of sensitivity and specificity of iron chromogens: measurement of serum iron with a new aqueous color reagent.

Methods presently used for the determi­ nation of serum iron and unsaturated ironbinding capacity have somewhat limited spectrophotometric sensitivity. Since molar absorptivity of a metal-ligand complex is a relative measure of sensitivity, an increase in the former improves accuracy in the de­ termination of trace constituents, especially when they are abnormally low. Obviously, low amounts of iron are best determined using the most sensitive, selective and spe­ cific reagents since in abnormal circum­ stances in which iron is the ion of interest, low concentrations are the most commonly encountered condition. Earlier procedures, if compared to modern ones, used the least sensitive reactants, and one might list thiocyanate, bipyridyl and 1 ,10-phenanthroline as almost defunct re­ agents. Several of the more sensitive colori­ metric techniques employ 4 ,7-diphenyl-l, 10-phenanthroline, its sulfonate (BPS) and 2,4,6,-tripyridyl-triazine (TPTZ) with mo­ lar absorptivities of 22,400, 22,140 and 22,600 respectively.1-9'27'30 A more sensitive but less known ligand than BPS and TPTZ is 2 ,6-di-[pyridyl-(2)]-4-[p-methoxyphenyl]pyridine and its sulfonate with a molar ab­ sorptivity of 26,900.2’23'24 Another new and very promising ligand, 3-(2-pyridyl)-5,6-diphenyl-1,2 ,4-triazine (PDT) with a molar absorptivity of 23,500,22 and its sulfonate called ferrozine with a molar absorptivity of 27,90025 may be capable of determining copper on the same specimen by sequential determination.22 Still another reagent for iron, 7-bromo-l ,3-dihydro-l-(3-dimethylaminopropyl)-5-(2-pyridyl)-2H-1 ,4-benzodiazepin-2-one dihydrochloride with a molar absorptivity of 17,000 has been described.8’18 However, its disadvantage at this time is its comparative insensitivity. The preparation of a sample for subse­ quent color formation is most commonly carried out by protein precipitation. How­ ever, other systems are available which in­ clude extraction,15 total destruction17 or di­ rect reaction without isolation of iron from its protein environment.10 The direct tech­ niques have appeal because of the apparent ease with which the determinations are carried out, and this approach will be con­ sidered further under Discussion. Measurements must often be made on serum samples of small volumes and of low concentrations of iron. In automated meth­ ods where only a portion of the iron is dialyzed, an increase in sensitivity would be beneficial. The highly insoluble but promis­ ing ligand, 2 ,6-bis(4-phenyl-2-pyridyl)-4phenyl pyridine (terosite) which has a visible range molar absorptivity of 30,200 would be useful for the determination of serum iron provided it could be converted to a watersoluble derivative. Since its molar absorp­ tivity is higher than TPTZ and BPS (30,200/ 22,600/22,140), the use of the compound could be advantageous for determining iron in the serum of anemic patients or where the handling of smaller samples is manda­ tory. The problem of converting this com­ pound to a water-soluble form was under­