<b>Two-dimensional (2D) correlation analysis was applied to characterize the attenuated total reflection (ATR) spectral intensity fluctuations of immature and mature cotton fibers. Prior to 2D analysis, the spectra were leveled to zero at the peak intensity of 1800 cm<sup>−1</sup> and then were normalized at the peak intensity of 660 cm<sup>−1</sup> to subjectively correct the variations resulting from ATR sampling. Next, normalized spectra were subjected to principal component analysis (PCA), and two clusters of immature and mature fibers were confirmed on the basis of the first principal component (PC1) negative and positive scores, respectively. The normalized spectra clearly demonstrated the intensity increase or decrease of the bands ascribed to different C-O confirmations of primary alcohols in the 1050-950 cm<sup>−1</sup> region, which was not apparent from raw ATR spectra. The PC1 increasing-induced 2D correlation analysis revealed remarkable differences between the immature and mature fibers. Of interest were that: (1) Both intensity increase of two bands at 968 and 956 cm<sup>−1</sup> and the shifting of 968 cm<sup>−1</sup> in immature fibers to 956 cm<sup>−1</sup> in mature fibers, together with the intensity decreasing and shifting of the 1048 and 1042 cm<sup>−1</sup> bands, are the characteristics of cotton fiber development and maturation. (2) Intensities of most bands in the 1800-1200 cm<sup>−1</sup> region decreased with the fiber growth, suggesting they are from either noncellulosic components or CH and OH fractions in amorphous celluloses. (3) The reverse sequence of intensity variations of the bands in the 1100-1000 cm<sup>−1</sup> and 1000-900 cm<sup>−1</sup> region of asynchronous spectra indicated a different mechanism of compositional and structural changes in developing cotton fibers at different growth stages.</b>

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