FTIR spectroscopy monitoring of cell wall modifications during the habituation of bean (Phaseolus vulgaris L.) callus cultures to dichlobenil

The habituation of bean calluses to dichlobenil results from the acquisition of a modified cell wall, with an enhancement in pectins and a decrease in cellulose and hemicelluloses. In this work, Fourier transform infrared (FTIR) spectroscopy in conjunction with a set of multivariate analyses and other statistical tools, such as principal component analyses and Student's t-test applied to clusters from a dendrogram, were used to monitor the modifications occurring in the cell walls of bean callus cultures due to a habituation program to dichlobenil. Forty samples of calluses, differing in the dichlobenil concentration at which they were growing and in the number of subcultures in a given concentration of dichlobenil, corresponding to two habituation experiments, were analyzed. Multivariate analyses of the spectra showed that the type and the extent of cell wall modifications depended on the concentration of the inhibitor in the culture medium and the time that the callus had been present at a given concentration of the inhibitor, and the analyses distinguished among three groups of calluses with different levels of habituation to dichlobenil: (i) non-habituated and habituated to low dichlobenil concentrations, (ii) habituated to intermediate concentrations (up to 4 muM), and (iii) habituated to high concentrations (4-12 muM). A slight modification of cell walls was only detected after 13 subcultures in 0.5 muM dichlobenil. In the presence of a higher concentration of dichlobenil, the content of cellulose was clearly reduced while that of pectins was increased. We conclude that FTIR spectroscopy associated with a set of statistical tools is a powerful method for analyzing in muro-and more rapidly-the changes in polysaccharides related to dichlobenil habituation, and that it could be used in the future to identify cell wall changes related to habituation to other herbicides or stress factors. (C) 2004 Elsevier Ireland Ltd. All rights reserved.