Ferrous thioglycollate

Theory The limit test for Iron is based on the reaction between iron and thioglycollic acid in a medium buffered with ammonium citrate to give a purple colour, which is subsequently compared with the standard colour obtained with a known amount of iron (0.04 mg of Fe). Ferrous thioglycollate is a co-ordination compound that attributes the purple colour besides thioglycollic acid converts the entire Fe3+ into Fe2+. The reactions involved may be expressed as follows ... 

Ferric ions and complexes in aqueous media react readily with cysteine (13, 21) or thioglycollic acid (14, 22) to form purple complexes which rapidly change to the ferrous form with accompanying formation of disulfides. It was not unreasonable, therefore, to assume that such a reaction provided rapid production of RS radicals in the initial stages of co-oxidation and that this rate should decrease as reduction to ferrous occurred. Eventually reoxidation of ferrous to ferric by peroxide would assert itself, and the rate could rise again. 

Thioglycolic acid can be identified by its ir spectrum or by gas chromatography. Most of the by-products and self-esterification products are also detected by liquid chromatography, eg, thiodiglycolic acid, dithiodiglycolic acid, linear dimers, and polymers. Iron content can be assayed by the red sensitive complex of 1,10-phenanthroline [66-71-7] and ferrous ion of a mineralized sample. Ferric ion turns an aqueous ammonia solution deep red-violet. 

The reaction is approximately first-order with respect to each reactant (the second-order rate coefficient increases with increase of substrate concentration), and catalysis by hydroxide ions is observed. Henderson and Winkler studied the ferrous ion-catalysed oxidation of thioglycolicacidto dithioglycolic acid. The rate is sensitive to traces of metal ions, and reproducible results could not be obtained in the absence of the catalyst. The oxidation is first-order with respect to both peroxodisulphate and ferrous ions, and zero-order with respect to the substrate. The second-order rate coefficient is approximately equal to that determined in the absence of the substrate, so Henderson and Winkler suggested that the ratedetermining step is the oxidation of ferrous to ferric ions, as in reaction (96), and that this is followed by reaction (97) and then rapid oxidation of thioglycolic acid by ferric ions. 

Several chromogenic iron chelators have been developed [17] that can be used in the assay. Ferrozine [3-(2-pyridyl)5,6-bis(4-phenylsulfonic acid) 1,2,4 triazine] Is also popular because its ferrous complex has a molar absorptivity of 28000 at 562 nm, which is about 35% more sensitive than the phenanthroline chromogen used in the reference method. However, ferrozine binds copper too, but this positive bias error is small at normal serum concentrations [42,45a], and is eliminated with thioglycolate [45b]. Ferrene and pyridyl-azo chelators have also been utilized as chromogens [45c,d]. 

Traces of iron may be estimated colorimetrically with considerable accuracy. The formation of ferric thiocyanate, which was the basis of the B.P. 1932 method, is no longer used because of the many interferences which are possible. Strafford preferred the use of thioglycollic acid which produces a purple colour with traces of ferrous iron in ammoniacal solution (ferric salts also respond because the reagent has strongly reducing properties). Quantitative use of this reagent was described by Swank and... 

To determine dissolved iron, a chromatic comparison was made of the violet complex produced by the reaction of ferrous ions with a buffer of thioglycolate. The maximum error of 30%. 

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