Behavior of serum in the borate modification of the carbazole reaction.
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Borate/boric acid solutions have distinctive effects on the absorption and fluorescence emission spectra of dityrosine. In the presence of excess borate/boric acid, the fluorescence emission maximum of the singly ionized dityrosine chromophore shifts from 407 nm (quantum yield = 0.80) to 374 nm (quantum yield = 0.14). Fluorescence measurements performed as a function of pH and concentration are consistent with a 1:1 complex which may dissociate to either boric acid and singly ionized dityrosine (K1 = 17 mM) or to monoborate ion and unionized dityrosine (K2 = 0.10 mM). As a consequence of the pKa values characteristic of dityrosine and boric acid, complex formation is maximal near pH 8. 2,2'-Dihydroxy-biphenyl shows similar interactions. The fluorescence of dityrosyl calmodulin (0 Ca2+) also responds to the addition of boric acid, giving K1 = 42 mM and K2 = 2 mM. Singly ionized dityrosine produced through dissociation occurring in the excited state does not interact with boric acid.
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