Recuperação e concentração de biomoléculas por flotação em sistemas aquosos bifásicos

Abstract

Solvent sublation (SS) is a technique based on the adsorption of active superficial substances at a gas / liquid interface and has been highlighted, in relation to liquid-liquid extraction (LLE), by some benefits such as simple operation, low dosage high solvent separation rates and product concentration; although, it is not suitable for treating all biomolecules. Derived from solvent sublation, aqueous two-phase flotation (ATPF) operates in an environment with high water content and has gained prominence in recent years in the recovery of phenolic components. In this context, the present thesis explores ATPF as a separation technique to recover molecules from different sources such as plant extract: gallic acid (GA) - from infusion of guava leaves, and polyphenols like (+) - catechin and quercetin of the depulped residue of a typical Amazonian fruit, camu-camu. In addition, the bactericidal agent 2-hydroxybiphenyl (2-HBP) produced via biodesulfurization of 4-methyl-dibenzothiophene by Rhodococcus rhodochrous was recovered by ATPF. In these studies, aqueous biphasic systems were formed from the polymer Polyethyleneglycol (PEG) and ammonium sulfate salt, and flotation was favored by the injection of compressed air to generate the ascending bubbles. The column in the ATPF system was made of glass with a 2.64 cm internal diameter and a capacity of 180 mL. The column was fitted to the bottom with a sintered glass disk of G4 porosity. A scale magnification, with the height: diameter ratio used as a criterion, was also performed (for ATPF-camu camu) in a glass column of dimensions 60.0 x 8.00 cm and a useful volume of 2.5 L. results showed that the optimal operating conditions for recovery of gallic acid by ATPF were: pH equal to 4.0, ammonium sulfate concentration of 450 g/L, PEG concentration of 80% (w/w), gallic acid concentration of 500 μg/mL, flotation time of 120 min, air flow equal to 37.5 mL/min and volumetric ratio of 0.1. When fed with guava leaf extract, the ATPF showed higher enrichment indexes and gallic acid distribution ratios than extraction in a conventional biphasic aqueous system. In the study with the camu-camu residue, the optimal experimental conditions were: pH equal to 5.0, ammonium sulfate concentration of 450 g/L, air flow equal to 37.5 mL/min, flotation time of 60 minm and volumetric ratio of 0.1. In this case, in the optimum condition, a separation efficiency of 81.02% and a concentration coefficient two times higher than the traditional two-phase aqueous extraction were obtained. In addition, the increase in antioxidant power in the upper phase was observed, resulting from the migration of some phenolic components such as gallic acid, syringic acid, quercetin and (+) - catechin. It should be noted that the scale-up was successful. Alternatively, a multivariate methodology was used to define the best operating conditions of ATPF to recover 2-HBP. After a fractional experimental design, only the variables of air flow, float time and volume of the top phase were optimized by the Box-Behnken design, in which it was possible to reach separation efficiency values of 43.55-57.55% and coefficients concentration of 3.84-5.88. As in the previous chapters, ATPF surpassed the two-phase aqueous system extraction, liquid-liquid extraction and solvent sublation in terms of enrichment factors. The flotation process was also successful in the recovery and concentration of 2-HBP from the fermented broth, although analysis by HPLC indicates the high concentration of contaminants in the top phase. Thus, the results showed that ATPF is a promising technique to recover and, mainly, concentrate phenolic components from complex media such as natural extracts and fermentation broths.