hexacynoferrate (III) is used as the oxidant, another separation step
is needed in order to extract the fluorescent thiochrome from the
aqueous phase to organic phase, or it will be quenched in the presence
of hexacynoferrate (III). Although Hg
is an efficient oxidant for the
thiochrome reaction without quenching the fluorescence, it is highly
toxic and not suitable for used in the biological systems. While Cu
is not as toxic as Hg
2+
2+
, it may give false response in the presence of
other biomolecules (such as cysteine) due to the formation of other
fluorescent species (e.g., Cu nanoclusters [15]) under similar reaction
conditions.When hydrogen peroxide is used as the oxidant, additional
reagents such as enzymes are required to make the thiochrome
formation more efficient, because meantime hydrogen peroxide as
well as other strong oxidants such as KMnO
also tends to convert
thiamine into non-fluorescent thiamine disulfide.