Reliability of dissociation constants and resolution capability of SQUAD(84) and SPECFIT/32 in the regression of multiwavelength spectrophotometric pH-titration data

The resolving power of multicomponent spectral analysis and the computation reliability of the stability constants and molar absorptivities determined for five variously protonated anions of physostigmine salicylate by the SQUAD(84) and SPECFIT/32 programs has been examined with the use of simulated and experimental spectra containing overlapping spectral bands. The reliability of the dissociation constants of drug was proven with goodness-of-fit tests and by examining the influence of pre-selected noise level sinst(A) in synthetic spectra regarding the precision s(pK) and also accuracy of the estimated dissociation constants. Precision was examined as the linear regression model s(pK) = β0 + β1 sinst(A). In all cases the intercept β0 was statistically insignificant. When an instrumental error sinst(A) is small and less than 0.5 mAU, the parameters’ estimates are nearly the same as the bias ΔpK = pKa,calc − pKa,true is quite negligible. In all four dissociation constants the bias seems to be quite small even though for pKa4 it is a little bit higher, i.e., +0.05 for sinst(A) about 1.0 mAU. In the interval of sinst(A) from 0.1 to 1.0 mAU all four dissociation constants pKi are accurate enough. Of the various regression diagnostics considered, the goodness-of-fit is the most efficient criterion of whether the parameters found adequately represent the data. The magnitude of instrumental error sinst(A) only slightly affects the shape of a Cattel's scree graph sk(A) = f(k) to determine the true number of light-absorbing species in the equilibrium mixture.

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► Reliability of pK and ɛ for anions of physostigmine salicylate was examined.
► When instrumental error <0.5 mAU, bias of parameters is negligible.
► In error interval 0.1–1.0 mAU all four pKi are accurate enough.
► Error slightly affects the shape of Cattel's plot to determine number of species.