Successive Fermentation of Aguamiel and Molasses by Aspergillus oryzae and Saccharomyces cerevisiae to Obtain High Purity Fructooligosaccharides.

Fructooligosaccharides (FOS) are usually synthesized with pure enzymes using highly concentrated sucrose solutions. In this work, low-cost aguamiel and molasses were explored as sucrose alternatives to produce FOS, via whole-cell fermentation, with an Aspergillus oryzae DIA-MF strain. FOS production process was optimized through a central composite experimental design, with two independent variables: initial sucrose concentration in a medium composed of aguamiel and molasses (AgMe), and inoculum concentration. The optimized process-165 g/L initial sucrose in AgMe (adjusted with concentrated molasses) and 1 × 107 spores/mL inoculum concentration-resulted in an FOS production of 119 ± 12 g/L and a yield of 0.64 ± 0.05 g FOS/g GFi. Among the FOSs produced were kestose, nystose, 1-fructofuranosyl-nystose, and potentially a novel trisaccharide produced by this strain. To reduce the content of mono- and disaccharides in the mixture, run a successive fermentation was run with two Saccharomyces cerevisiae strains. Fermentations run with S. cerevisiae S227 improved FOS purity in the mixture from 39 ± 3% to 61.0 ± 0.6% (w/w) after 16 h of fermentation. This study showed that agro-industrial wastes such as molasses with aguamiel are excellent alternatives as substrate sources for the production of prebiotic FOS, resulting in a lower-cost process.

The secondary metabolites from Beauveria bassiana PQ2 inhibit the growth and spore germination of Gibberella moniliformis LIA.

Fungal secondary metabolites with antimicrobial properties are used for biological pest control. Their production is influenced by several factors as environment, host, and culture conditions. In the present work, the secondary metabolites from fermented extracts of Beauveria bassiana PQ2 were tested as antifungal agents against Gibberella moniliformis LIA. The L18 (21 × 37) orthogonal array from Taguchi methodology was used to assess 8 parameters (pH, agitation, sucrose, yeast extract, KH2PO4, MgSO4, NH4NO3, and CaCl2) in B. bassiana PQ2 submerged fermentation. The ability of the fermented extracts to slow down the growth rate of G. moniliformis LIA was evaluated. The results from 18 trials were analyzed by Statistica 7 software by evaluating the signal-to-noise ratio (S/N) to find the lower-the-better condition. Optimal culture conditions were pH, 5; agitation, 250 rpm; sucrose, 37.5 g/L-1; yeast extract, 10 g/L-1; KH2PO4, 0.8 g/L-1; MgSO4, 1.2 g/L-1; NH4NO3, 0.1 g/L-1; and CaCl2, 0.4 g/L-1, being the agitation at the highest level the most significant factor. The optimal conditions were validated in a sparged bottle bioreactor resulting in a higher S/N value (12.48) compared to the estimate. The extract obtained has the capacity to inhibit the germination of G. moniliformis spores at 24 h. HPLC-ESI-MS2 allowed to identify the water-soluble red pigment as oosporein (m/z 304.9). The secondary metabolites from B. bassiana PQ2 are a suitable alternative to control the growth and sporulation of G. moniliformis.

A simple quantitative method using TLC-image analysis to determine fructooligosaccharides (FOS) in food samples.

The thin-layer chromatography technique (TLC) is a simple and inexpensive analysis commonly used to identify qualitatively the presence of carbohydrates in food samples such as mono- di and oligosaccharides particularly. TLC assay could be improved using image processing software for the semiquantitative determination of this type of compound. In the present work, TLC-image analysis with Silica Gel 60 TLC plates was used for the semiquantitative determination of 6 standards of carbohydrates (glucose, fructose, sucrose, 1-kestose, nystose, and fructofuranosylnystose). Subsequently, the areas of the spots of each compound were determined by digitizing in a conventional office scanner. Then, the segmentation of the images is carried out using software for image processing. The calibration curves were plotted in the Excel software using the average of the areas of the pigmentations obtained in pixels. In this study, the technique of thin-layer chromatography was also used to quantitatively determine the presence of carbohydrates in food samples such as honey, garlic, and onion. Values of determination coefficient (R2) greater than 0.97 in all the calibration curves were obtained. This technique could be useful for detecting carbohydrates (monosaccharides, disaccharides, and oligosaccharides) in analytical assays and food samples without needing specialized analytical equipment. In this work, it was possible to determine the concentration of carbohydrates in samples of garlic and onion that showed the presence of prebiotic carbohydrates in addition to sucrose, glucose, and fructose.

Characterization of a Biofilm Bioreactor Designed for the Single-Step Production of Aerial Conidia and Oosporein by Beauveria bassiana PQ2.

Beauveria bassiana is an entomopathogenic fungus that is used for the biological control of different agricultural pest insects. B. bassiana is traditionally cultivated in submerged fermentation and solid-state fermentation systems to obtain secondary metabolites with antifungal activity and infective spores. This work presents the design and characterization of a new laboratory-scale biofilm bioreactor for the simultaneous production of oosporein and aerial conidia by B. bassiana PQ2. The reactor was built with materials available in a conventional laboratory. KLa was determined at different air flows (1.5-2.5 L/min) by two different methods in the liquid phase and in the exhaust gases. The obtained values showed that an air flow of 2.5 L/min is sufficient to ensure adequate aeration to produce aerial conidia and secondary metabolites by B. bassiana. Under the conditions studied, a concentration of 183 mg oosporein per liter and 1.24 × 109 spores per gram of support was obtained at 168 h of culture. These results indicate that the biofilm bioreactor represents a viable alternative for the production of products for biological control from B. bassiana.

Structural characterization of native and oxidized procyanidins (condensed tannins) from coffee pulp (Coffea arabica) using phloroglucinolysis and thioglycolysis-HPLC-ESI-MS.

Procyanidins from coffee pulp are responsible from the limited valorization of this by-product. Information about procyanidin structure is still scarce and imprecise. The aim of this work was to study the native and oxidized procyanidins from coffee pulp with respect to composition and structure. An aqueous acetone extract from coffee pulp was purified using Sephadex LH-20. Butanolysis, phloroglucinolysis and thioglycolysis coupled to HLPC-ESI-MS were applied for the characterization of the native and oxidized procyanidins. The purification allowed to recovery three fractions (aqueous, ethanolic and acetonic) and only acetone fraction showed a high concentration of procyanidins (98%, w/w). HPLC-ESI-MS of procyanidins-rich fraction without any reaction resulted in a UV-Vis chromatogram unresolved typical of the presence of procyanidins. The extracted ion chromatogram and MS2 analysis revealed the presence from dimers to pentamers of native procyanidins. Interestingly, by first time an A-type trimeric procyanidin (m/z of 863) was observed in coffee pulp. In our study, (-)-epicatechin was the constitutive unit of procyanidins with an aDP of 6.8 (oligomeric native procyanidins) according to the phloroglucinolysis assay. Two oxidation markers useful to characterization of oxidized procyanidins were observed in the procyanidins-rich fraction after thioglycolysis, a dimer A2-ext and a molecule that corresponds to a linkage between an extension and a terminal unit. Coffee pulp procyanidins were presented with only a minor class of oxidized procyanidins. As far as we know, this is the first study about characterization of the oxidized procyanidins from coffee pulp.

Characterization by HPLC-ESI-MS2 of native and oxidized procyanidins from litchi (Litchi chinensis) pericarp.

Procyanidins (PCs) are polyphenols highly accumulated in litchi fruit (Litchi chinensis). Despite their bioactivity, the molecular composition of native and oxidized procyanidins is little understood. In this paper, polyphenols from litchi pericarp were extracted using two solvents (methanol and acetone). The mean degree of polymerization (mDP) of native and identification of oxidized PCs were carried out by phloroglucinolysis- and thioglycolysis-HPLC-ESI-MS/MS, respectively. About 60% of extracted polyphenols corresponded to procyanidins from litchi pericarp. Native PCs were mainly oligomeric procyanidins (mDP 4). Only (-)-epicatechin was detected as terminal and extension units in PCs. Thioglycolysis-HPLC-ESI-MS identified five oxidation markers of PCs with [M-H]-m/z 575, 593, 609, 679 and 863. Intra- and intermolecular modifications of A and B-type procyanidins were identified. The method used for the characterization of PCs from litchi pericarp allowed understanding of the structural composition of its native and oxidized tannins.

Effect of ultrasound treatment on the extraction of antioxidants from Ardisia compressa Kunth fruits and identification of phytochemicals by HPLC-ESI-MS.

The influence of ultrasound-assisted extraction of phytochemicals from Ardisia compressa Kunth on the antioxidant capacity was investigated. The factors evaluated were: ultrasound extraction time (10, 20 and 30 min), ethanol concentration (0, 35, 70 %) and solid/liquid ratio (1:4, 1:8 and 1:12 g mL-1). The L9 (3)3 array was applied, and the DPPH• scavenging capacity of treatments was evaluated to obtain optimal extraction conditions. Finally, the phytochemicals were characterized by high-performance liquid chromatography electrospray ionization mass spectrometry (HPLC-ESI-MS). Ten minutes of ultrasound extraction using 0 % of ethanol and solid/liquid ratio 1:12 g mL-1 were the optimal conditions of extraction. The HPLC-ESI-MS analysis revealed the presence of gluconic acid, quercetin-3-O-glucoside, isorhamnetin-3-O-rutinoside, demethylligstroside, ponicidin, 4-caffeoylquinic acid, rosmarinic acid, and galloyl-hexoside. The optimal ultrasound-assisted extraction conditions were defined by applying the Taguchi methodology. The phytochemicals identified in A. compressa fruits suggest its use as a potential source of bioactive compounds.

Enhancement of fructosyltransferase and fructooligosaccharides production by A. oryzae DIA-MF in Solid-State Fermentation using aguamiel as culture medium.

The aim of this work was to improve the production of fructosyltransferase (FTase) by Solid-State Fermentation (SSF) using aguamiel (agave sap) as culture medium and Aspergillus oryzae DIA-MF as producer strain. SSF was carried out evaluating the following parameters: inoculum rate, incubation temperature, initial pH and packing density to determine the most significant factors through Box-Hunter and Hunter design. The significant factors were then further optimized using a Box-Behnken design and response surface methodology. The maximum FTase activity (1347U/L) was obtained at 32°C, using packing density of 0.7g/cm(3). Inoculum rate and initial pH had no significant influence on the response. FOS synthesis applying the enzyme produced by A. oryzae DIA-MF was also studied using aguamiel as substrate.

Ultrasound-assisted extraction of polyphenols from native plants in the Mexican desert.

Several plants that are rich in polyphenolic compounds and exhibit biological properties are grown in the desert region of Mexico under extreme climate conditions. These compounds have been recovered by classic methodologies in these plants using organic solvents. However, little information is available regarding the use of alternative extraction technologies, such as ultrasound. In this paper, ultrasound-assisted extraction (UAE) parameters, such as the liquid:solid ratio, solvent concentration and extraction time, were studied using response surface methodology (RSM) for the extraction of polyphenols from desert plants including Jatrophadioica,Flourensiacernua, Turneradiffusa and Eucalyptuscamaldulensis. Key process variables (i.e., liquid:solid ratio and ethanol concentration) exert the greatest influence on the extraction of all of the phenolic compounds (TPC) in the studied plants. The best conditions for the extraction of TPC involved an extraction time of 40min, an ethanol concentration of 35% and a liquid:solid ratio ranging from 8 to 12mlg(-1) depending on the plant. The highest antioxidant activity was obtained in the E. camaldulensis extracts. The results indicated the ability of UAE to obtain polyphenolic antioxidant preparations from desert plants.

Microplate quantification of total phenolic content from plant extracts obtained by conventional and ultrasound methods.

INTRODUCTION: There is increasing interest in phenolic compounds around the world because of their potential positive impact on human health. Phenolic compounds are largely found in fruits and vegetables. Extraction of phenolic compounds is a very important step in their recovery. The newly developed technique of ultrasound-assisted extraction (UAE) appears to be an advantageous alternative compared with conventional techniques, because it is simple and environmental friendly. The potential of UAE needs to be evaluated in each plant in order to demonstrate its efficiency. OBJECTIVE: The objective of the present study was to compare a conventional method and UAE on the extraction efficiency of phenolic compounds from Jatropha dioica, Fluorensia cernua, Turnera diffusa and Eucalyptus camaldulensis plants and evaluate the in vitro anti-oxidant potential. METHODS: Validation of the new method was carried out using mixed-model methodology and regression analysis. Feasibility of this new method was shown and applied using several plants extracts obtained by different extraction methods from semi-arid Mexican plants, which were characterised by high levels of polyphenols. Additionally, the anti-oxidant potential of these extracts was determined by 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity. RESULTS: Results showed that the new microplate method can be used to determine total phenolic content in plant extracts. Additionally, an alternative extraction method by ultrasound was less efficient compared with the conventional method. CONCLUSION: The tested plants are good candidates to obtain nutraceuticals and functional food ingredients.

Ultrasound-assisted extraction of phenolic compounds from Laurus nobilis L. and their antioxidant activity.

Bay leaves (BL) (Laurus nobilis L., Family: Laureceae) are traditionally used to treat some symptoms of gastrointestinal problems, such as epigastric bloating, impaired digestion, eructing and flatulence. These biological properties are mainly attributed to its phenolic compounds. In this paper, ultrasound-assisted extraction of phenolic compounds from Laurus nobilis L. (Laureceae) was studied. Effects of several experimental factors, such as sonication time, solid/liquid ratio and concentration of solvent on extraction of phenolic compounds were evaluated through a randomized complete block design with factorial treatment arrangement (3(3)). The best extraction conditions were: 1g plant sample with 12 mL of 35% ethanol, for 40 min, obtaining a yield of phenolic compounds of 17.32±1.52 mg g(-1) of plant. In addition, free radical-scavenging potential of DPPH and lipid oxidation inhibition, by linoleic acid peroxidation of the selected extract was measured in order to evidence their antioxidant properties. Results indicated that high amounts of phenolic compounds can be extracted from L. nobilis by ultrasound-assisted extraction technology.