Soil physical, chemical, and biological factors have been shown to affect root colonization, but the detailed mechanisms associated with the adequate colonization of root and microorganisms are not completely understood.Quantitative studies on the distribution of root colonies in time and space are needed to develop mathematical models that describe and predict the root colonization process; being a important factor, to know the effect of inorganic and organic fractions of soils on the ability of microorganisms to form these colonies.Here a methodology to the study of interaction between microorganisms and surface is described and its application for the study of the description of interaction and colonization mechanism between nitrogen-fixing bacteria and particle of soils is shown, special attention is directed toward the effect of humic acids on formation of biofilms.For this, humic acids was extracted from two types of soil samples and characterized by Infrared spectroscopy with Fourier Transform coupled to Attenuated Total Reflectance (FTIR-ATR) and contact angle (AC).Humic substances were deposited on cellulose previously activated with spectral marker for the mid-infrared and anew characterized.Later, the bacterium Azotobacter chroococcum (nitrogen-fixing bacterium) was deposited on the surface by direct contact.End surfaces were analyzed by FTIR-ATR, and AC.The results suggest that it is necessary to analyze different surfaces in order to describe several mechanisms associated with the nature of this through the use of instrumental techniques and physicochemical models with special emphasis on surface interactions, which makes this a promising tool for description of the process of microbial colonization of soils. Infrared spectroscopy Nitrogen-fixing bacteriaHumic substances Biofilm