Selecting two-dimensional cross-correlation functions to enhance interpretation of near-infrared spectra of proteins

Two-dimensional infrared (2D-1R) cross-correlation can facilitate the inter pretation of infrared spectra, However, the fluctuation of noise and the ov erlapped component bands often complicate the analysis of 2D-IR spectra, Th e ratio of asynchronous to synchronous correlation functions (F) can serve as a control of coherence for the cross peaks in 2D correlation maps. A syn chronous cross peak with an F value close to 0 corresponds to the good cohe rence of cross-correlated pair of signals, more likely reflecting a true sy nchronous cross peak, while a large F value associated with a synchronous c ross peak is more likely a false cross peak. Similarly, the noncoherence of asynchronous cross peaks can be checked by computing their F values. In th is case, large F values are associated with true asynchronous correlation p eaks, while small F values are more likely false asynchronous cross peaks. This approach has been tested on smoothed and unsmoothed near-infrared (NIR ) spectra of human serum albumin (HSA) in aqueous solutions. A set of NIR s pectra of HSA with increasing concentration were measured to generate 2D co rrelation maps. Unsmoothed spectra permitted the acquisition of synchronous and asynchronous 2D correlation maps containing all putative cross peaks i n addition to artifacts generating by the fluctuation of noise or band over lapping. The control of coherence of pairs of signals on each cross peak, b ased on their F values, facilitated the interpretation of 2D-NIR maps, by i mproving the selection of true cross peaks from false cross peaks. The smoo thed NIR spectra provided cross peaks without any artifacts in the synchron ous and asynchronous correlation maps. The selection of cross peaks based o n their F values confirmed the absence of false cross peaks in the case of smoothed spectra. Despite some limitations as discussed in this report, thi s procedure is straightforward to implement and will make the analysis of 2 D vibrational spectra much easier.