Mechanism of sequential order determination in bio-raman correlation analysis

Using the measured Raman spectra of triolein and cholesteryl linolenate, the contradiction caused in determining the sequential orders in the two-dimensional correlation spectroscopy was exemplified, in which time-profiles of four marker bands A, B, C, and D were modeled so that A→B→C→ D. Here 'A→B' is such notation that we read as 'A is occurred before B' or 'A earlier than B'. The two-dimensional correlation method gave the result B→C→D→A which was contradictive to the initial setting. We confirmed that the increments of distance between the peak positions of the Gaussian type time-profiles f and g, through a threshold, gave the unexpected switch in the sequential order. On the complex plane based on the synchronous and asynchronous axes, the vector g is identical to the synchronous axis in direction; the vector f is crossed through the asynchronous axis corresponding to the increments of distance between the peak positions of f and g. By the conventional rule of the correlation method, the vector f just crossing the asynchronous axis is not allowed to be in the second quadrant, but the vector f is transformed to the fourth quadrant with 180 degrees shifted. In the situation, the vector f is located in the later position for the standard vector g, that is, f is occurred 'after' g. Based on the revealed mechanism for the contradiction, the time series correlation analysis may be extended in a more versatile manner to allow bio-Raman correlation analysis on diverse dynamics of bio-molecules in living cells.