Adalberto Pessoa-Jr

Micellar solubilization of ibuprofen: influence of surfactant head groups on the extent of solubilization

An important property of micelles with particular significance in pharmacy is their ability to increase the solubility of poorly soluble drugs in water, thus increasing their bioavailability. In this work, the solubilization of ibuprofen (IBU) was studied in micellar solutions of three surfactants possessing the same hydrocarbon tail but different hydrophilic head groups, namely sodium dodecyl sulphate (SDS), dodecyltrimethylammonium bromide (DTAB), and n-dodecyl octa(ethylene oxide) (C12EO8). The results showed that, irrespective of the surfactant type, the solubility of IBU increased linearly with increasing surfactant concentration, as a consequence of the association between the drug and the micelles. The 80 mM DTAB and the 80 mM C12EO8 micellar solutions resulted in a 16-fold increase in solubility of IBU when compared to the buffer solution, whereas the 80 mM SDS micellar solution resulted in a 5.5-fold increase in IBU solubility. The highest value of molar solubilization capacity (c) was obtained with DTAB, c = 0.97, followed by C12EO8 ,c = 0.72, and finally SDS, c = 0.23. However, due to the stronger tendency of the nonionic surfactant in forming micelles in solution, at the same surfactant concentration, we obtained the same solubility of IBU in both DTAB and C12EO8.

Can affinity interactions influence the partitioning of glucose-6-phosphate dehydrogenase in two-phase aqueous micellar systems?

In this work, we provide an investigation of the role and strength of affinity interactions on the partitioning of the glucose-6-phosphate dehydrogenase in aqueous two-phase micellar systems. These systems are constituted of micellar surfactant solutions and offer both hydrophobic and hydrophilic environments, providing selectivity to biomolecules. We studied G6PD partitioning in systems composed of the nonionic surfactants, separately, in the presence and absence of affinity ligands. We observed that G6PD partitions to the micelle-poor phase, owing to the strength of excluded-volume interactions in these systems that drive the protein to the micelle-poor phase, where there is more free volume available.

Optimization of clavulanic acid production by Streptomyces daufpe 3060 by response surface methodology

Clavulanic acid is a β-lactam antibiotic which has a potent β-lactamase inhibiting activity. In order to optimize its production by the new isolate Streptomyces DAUFPE 3060, the influence of two independent variables, temperature and soybean flour concentration, on clavulanic acid and biomass concentrations was investigated in 250 mL-Erlenmeyers according to a 22 central composite design. To this purpose, temperature and soybean flour (SF) concentration were varied in the ranges 26-34°C and 10-50 g/L, respectively, and the results evaluated utilizing the Response Surface Methodology. The experimental maximum production of clavulanic acid (629 mg/L) was obtained at 32°C and 40 g/L SF after 48 h, while the maximum biomass concentration (3.9 g/L) at 30°C and 50 g/L soybean flour, respectively. These values are satisfactorily close to those (640 mg/L and 3.75 g/L, respectively) predicted by the model, thereby demonstrating the validity of the mathematical approach adopted in this study.

Caracterização da xilose redutase extraída por micelas reversas-CTAB a partir de homogenato de Candida guilliermondii

Xylosereductase (XR) (E.C.1.1.1.21), produced by Candida guilliermondii, grown in sugar cane bagasse hydrolysate, was separated directly from the cell homogenate by reversed micelles of cetyl trimethyl ammonium bromide (CTAB), attaining a recovery yield of 100% and enrichment factor of 5.6 fold. The extraction conditions were: pH=7.0, electrical conductivity= 14 mS/cm, T=5 oC, 5% (w/w) of hexanol, 22% (w/w) of butanol and 0.15 M CTAB. The XR after extraction was stable in pH interval of 6.0-6.5 and its heat inactivation constant was about 6.5 fold higher than that before extraction. The Vmax values against both xylose and NADPH for XR before and after extraction by reversed-micelles differed about 6%, whereas the difference on KM values were more pronounced. The (KM)xylose for XR after extraction was about 35% higher than before extraction, meanwhile (KM)NADPH was about 30% lower after than before extraction. As the KM variations indirectly signaled, the XR affinity simultaneously diminishes for xylose and increases for NADPH. This could probably explain why the Vmax values for XR before and after extraction were quite similar.

Microbial cell disruption methods for efficient release of enzyme L-asparaginase

Abstract The efficacy of a simple laboratory method for cell disruption based on the glass bead stirring, sonication, osmotic shock, freezing and grinding, or use of solvents and detergents was assessed in this study, via measurements of the release of total protein and L-asparaginase activity. Three different microbial sources of L-asparaginase were used: Escherichia coli BL21 (DE3), Leucosporidium muscorum, and Aspergillus terreus (CCT 7693). This study adjusted and identified the best procedure for each kind of microorganism. Sonication and glass bead stirring led to obtaining filamentous fungus cell-free extracts containing high concentrations of soluble proteins and specific activity; however, sonication was the best since it obtained 4.61 ± 0.12 IU mg−1 after 3 min of operation time. Mechanical methods were also the most effective for yeast cell disruption, but sonication was the technique which yielded a higher efficiency releasing 7.3 IUtotal compared to glass bead stirring releasing 2.7 IUtotal at the same operation time. For bacterium, sonication proved to be the best procedure due to getting the highest specific activity (9.01 IU mg−1) and total enzyme activity (61.7 IU). The data presented lead to conclude that the mechanical methods appeared to be the most effective for the disintegration of the all microbial cells studies. This is the first report related to the experimental comparison of L-ASNase extraction procedures from different microorganisms, which can also be used for extracting periplasm located enzymes from other organisms.

ADSORPTION OF ENDOTOXINS ON IDA-Ca+2 BY ION METAL AFFINITY CHROMATOGRAPHY

A method was established for the determination of deoxynivalenol (vomitoxin) in grain and its products based on solid-phase extraction coupled with high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The sample was firstly extracted by acetonitrile-water (84:16, v/v). The extract was then cleaned-up by an HLB solid phase extraction cartridge. The separation was carried out on a Phenomenex Kinetex C18 column (100 mm × 4.6 mm, 2.6 μm) with a gradient elution using 0.3 per mille ammonia solution-acetonitrile as mobile phases. The analysis of deoxynivalenol was performed under electrospray negative ionization mode. The limit of detection (LOD, S/N =3) and the limit of quantification (LOQ, S / N =10) were 20 μg/kg and 50 μg / kg, respectively. A good linearity (r ＞ 0.99) was achieved for the target compound over the range of 20 - 1000 μg/L. The recoveries at the three spiked levels (50, 100, 500 μg/kg) in the blank matrices such as flour, barley, soybean, rice, cornmeal, cassava and wheat, were varied from 75.6% to 111.0% with the relative standard deviations no more than 13.0%. The method is accurate, efficient, sensitive and practical. The cost of pretreatment is obviously reduced by replacing immunoaffinity columns and Mycosep columns with HLB columns which have the same purification effect.