Detection of total gluconic acids in matriz derived from microbial cultivation by High Performance Liquid Chromatography Efficiency (HPLC)

Abstract

The ability to produce organic acids and tolerates acidity is peculiar of the nitrogen-fixing bacterium Gluconacetobacter diazotrophicus. Besides the industrial application, various organic acids, including gluconic acids have been shown to be responsible for solubilization of various compounds found in non-available forms in the soil. For this reason, development of methods to quantify the production of these acids is essential to establish their role in plant growth promotion. However, due to their close structural form specific and quantitative determination of the substrate (D - glucose) and gluconic acids and its derivatives has been limited by High Performance Liquid Chromatography analysis (HPLC). The protocol presented here proposes the use of two detection methods to determine the amount of glucose and total gluconic acids in samples derived from in vitro cultures of G. diazotrophicus: enzyme-colorimetric (commercial) and by HPLC detection. Although it was not possible to separate the D - gluconic acid from their ketoderivatives, the method showed low level of D-glucose interference and very high linearity for the detection of total gluconic acids. The quantitative analysis of these compounds in samples of PAL5 culture supernatants indicated that 33,10 mg of total gluconic acids ml-1 were produced during growth of G. diazotrophicus. The chromatographic conditions analysis established in this study can be used as a tool for identification and characterization of microorganisms capable of promoting the solubilization of phosphates and other mineral by synthesizing total gluconic acids.