Improving Biodegradation of Clofibric Acid by Trametes pubescens through the Design of Experimental Tools

International audienceClofibric acid (CLF) is the main pharmacologically active metabolite in composition of the pharmaceutical products used for controlling blood lipid content. This xenobiotic compound is highly persistent in the aquatic environment and passes unchanged or poorly transformed in wastewater treatment plants. A white-rot fungal strain of Trametes pubescens was previously selected, for its ability for clofibric acid biodegradation (up to 30%) during cultivation in submerged system under aerobic conditions at an initial CLF concentration of 15 mg L−1. Plackett-Burman design (PBD) and response surface methodology (RSM) were used for experimental planning, mathematical modelling and statistical analysis of data of the biotechnological process of CLF biotransformation by Trametes pubescens fungal strain. After optimization, the capacity of the selected Trametes pubescens strain to degrade CLF was increased by cultivation in a liquid medium containing 3 g·L−1 yeast extract, 15 g·L−1 peptone, 5 g·L−1 glucose and mineral salts, inoculated at 2% (v/v) vegetative inoculum and cultivated at pH 5.5, during 14 days at 25 °C and 135 rpm. In these optimized biotechnological conditions, the CLF biotransformation yield was 60%

Screening of Biotechnological Parameters for Fructofuranosidases Production by a Newly Isolated Fungal Strain Using Plackett-Burman Design

Previously some fungal strains with good inulinase potential biosynthesis were screening and the most performing strain is the newly fungal strain, Rhizoctonia spp. coded I-PN4. In this study, the Plackett Burman design was used to identify the effect of process parameters upon fructofuranosidases (inulinase and invertase) production with new selected fungal strain. The agitation rate, yeast extract, buckwheat flour and K2HPO4 concentration from fermentative medium composition, were determined as the most influential variables that have positive effects on fructofuranosidases biosynthesis. Temperature has a negative effect on both invertase and inulinase biosynthesis yield. In optimal established fermentative conditions, the maximum yields for extracellular inulinase was 6.2198 UI and 4.7664 UI for invertase respectively. The linear model fits well for variable screening and is globally significant, as R2 is 88.36% for inulinase production and 84.24% for invertase production respectively

Spectroscopic and Molecular Modeling Investigation on the Interaction between Folic Acid and Bovine Lactoferrin from Encapsulation Perspectives

The impact of thermal treatment on the ability of lactoferrin (FL) to bind folic acid (FA) was investigated by employing fluorescence spectroscopy, molecular dynamics and docking tests. The structural and conformational particularities of LF upon heating at 80 °C and 100 °C were first estimated based on the intrinsic fluorescence changes in respect to the native protein. The emission spectra indicated gradual unfolding events accompanied by Trp exposure with increasing temperature. In agreement with the experimental results, molecular modeling investigations showed that the secondary and tertiary structure of LF are slightly affected by the thermal treatment. Some minor unfolding events related particularly to the α-helical regions of LF were observed when the temperature increased to 100 °C. The LF fluorescence quenching upon FA addition indicated that a static mechanism stands behind LF-FA complex formation. Regardless of the simulated temperature, the hydrogen bonds played an important role in regulating the interaction between the protein and ligand. FA binding to LF equilibrated at different temperatures occurred spontaneously, and all complexes displayed good thermodynamic stability. The obtained results support the suitability of LF as biocompatible material, for obtaining micro- and nanoparticles for delivery of dietary supplements or for enhancing the functionality of target delivery systems

Screening of Biotechnological Parameters for Fructofuranosidases Production by a Newly Isolated Fungal Strain Using Plackett-Burman Design

Previously some fungal strains with good inulinase potential biosynthesis were screening and the most performing strain is the newly fungal strain, Rhizoctonia spp. coded I-PN4. In this study, the Plackett Burman design was used to identify the effect of process parameters upon fructofuranosidases (inulinase and invertase) production with new selected fungal strain. The agitation rate, yeast extract, buckwheat flour and K2HPO4 concentration from fermentative medium composition, were determined as the most influential variables that have positive effects on fructofuranosidases biosynthesis. Temperature has a negative effect on both invertase and inulinase biosynthesis yield. In optimal established fermentative conditions, the maximum yields for extracellular inulinase was 6.2198 UI and 4.7664 UI for invertase respectively. The linear model fits well for variable screening and is globally significant, as R2 is 88.36% for inulinase production and 84.24% for invertase production respectively

Characterisation of Newly Polar Psychrotrophic Streptomycetes Isolates from Polar Soils with Cold Adapted Bioremediation Potential

Novel polar streptomycetes coded 4 Alga, P2C4 and MIUG 12P, isolated from soil and vegetation samples prelevated from East Antarctica, were genetically and biochemically characterized in order to set up the phylogeny and to identify the capability to grow at lower temperature and to generate amylase and protease cold-active. The 16S rRNA gene sequence of isolate coded 4 Alga was 100% identical to sequences of Streptomyces sp. isolates from Norway and from Solomon Islands, which also were the closest relatives of P2C4 (99.53%) and MIUG 12P (99.53%) isolates. The data emphasised the skills of the strains Streptomyces 4 Alga, Streptomyces P2C4 and Streptomyces MIUG 12P to biosynthesis cold-adapted amylases and proteases suitable in bioconversion processes at low temperature. These results are quit valuable, because only few streptomycetes, particularly psychrotolerant ones, have so far been explored for their enzymatic potential in cold conditions

Tailoring the Optimized Fermentation Conditions of SCOBY-Based Membranes and Milk Kefir Grains to Promote Various Functional Properties

Kombucha culture (named SCOBY-Symbiotic Culture of Bacteria and Yeasts) and milk kefir grains represent multiple consortia of wild microorganisms that include lactic acid bacteria, acetic acid bacteria and yeasts with valuable functional properties. Their fermentative potential provides a wide range of derivate metabiotics (prebiotics, probiotics, postbiotics and paraprobiotics) with valuable in vitro and in vivo benefits. This study targeted the evaluation of the functionality of a co-culture of SCOBY-based membranes and milk kefir grains, used as freeze-dried starter cultures, for the fermentation of a newly formulated medium based on black tea infusion, supplemented with bovine colostrum and sugar, in order to produce bioactive compounds with functional properties. The design and optimization of the biotechnological process were achieved by using the Plackett–Burman experimental design (six factorial points, three center points) and the response surface methodology and central composite design (three factorial points, six axial points and two center points in axial) tools. The statistical analysis and the mathematical modelling of the responses such as the pH, titratable acidity, antioxidant activity and antimicrobial activity (against Bacillus subtilis, Escherichia coli, Staphylococcus aureus and Aspergillus niger) were investigated. Further, the composition of organic acids, polyphenols and flavonoids of the fermented product obtained under the optimized fermentation conditions was also analyzed. The fermentation of the medium containing 6.27% (w/v) bovine colostrum powder, 1.64% (w/v) black tea, 7.5% (w/w) sugar, pH 6.7, with an inoculum based of 0.36% (w/v) milk kefir grains powder and 0.5% (w/v) SCOBY-based membrane (both as freeze-dried culture), at 30 °C, for 5 days, in an aerobic stationary system, revealed an antifungal activity between 80 and 100% against Aspergillus niger, an antibacterial activity of 8–22 mm against Escherichia coli and Bacillus spp. And a titratable acidity of 445 °Th. The chemical composition of the obtained product had a positive impact on the functional properties of the fermented products in terms of the antimicrobial and antioxidant properties

Multifunctional Ingredient from Aqueous Flavonoidic Extract of Yellow Onion Skins with Cytocompatibility and Cell Proliferation Properties

Significant quantities of onion are cultivated annually, such that industrial processing leads to an appreciable amount of by-products, estimated at around 500,000 tons. Onion skins are considered an important source of naturally occurring antioxidant compounds, particularly flavonoid compounds. Our study follows the development of a sustainable solution in order to manage the by-products of yellow onion skins by designing ingredients with multifunctional activities. A green solvent aqueous extraction of flavonoids was applied to obtain a safe, flavonoid-enriched extract, yielding a total flavonoid content of 50.21 ± 0.09 mg quercetin equivalent (QE)/g dry weight (DW), and an antioxidant activity of 250.81 ± 6.76 mM Trolox/g DW. Complex biopolymeric matrices consisting of whey protein isolates, whey protein hydrolysates, maltodextrin, and pectin were further dissolved in the flavonoid-enriched aqueous extract, followed by freeze-drying. Two powders were obtained, both showing satisfactory phytochemical content and good stability during storage. The application of confocal microscopy revealed that the microscopic structure of the powders have a distribution of the bioactive compounds within the biopolymeric matrices. The in vitro digestion suggested remarkable stability in the gastric tract and a flavonoid-controlled release in the intestinal phase. A significant compatibility range of up to 1 mg/mL for both powders was found, whereas concentrations between 10 and 250 µg/mL stimulated cell proliferation after 24 h of cultivation. The powders showed satisfactory thermal and pH stability, which favors their addition to different food matrices

Whey Protein Isolate-Xylose Maillard-Based Conjugates with Tailored Microencapsulation Capacity of Flavonoids from Yellow Onions Skins

The objective of this study is to encapsulate flavonoids from yellow onion skins in whey protein isolates (WPI) and xylose (X), by Maillard-based conjugates, as an approach to improve the ability to entrap flavonoids and to develop powders with enhanced antioxidant activity. WPI (0.6%, w/v) was conjugated to X (0.3%, w/v) through the Maillard reaction at 90 °C for 120 min, in the presence of a flavonoid-enriched extract. Two variants of powders were obtained by freeze-drying. The glycation of WPI allowed a better encapsulation efficiency, up to 90.53 ± 0.29%, corresponding to a grafting degree of 30.38 ± 1.55%. The molecular modelling approach was used to assess the impact of X interactions with α-lactalbumin and β-lactoglobulin on the ability of these proteins to bind the main flavonoids from the yellow onion skins. The results showed that X might compete with quercetin glucosides to bind with α-lactalbumin. No interference was found in the case of β-lactoglobulin. The microstructural appearance of the powders revealed finer spherosomes in powder with WPI–X conjugates via the Maillard reaction. The powders were added to nachos, followed by a phytochemical characterization, in order to test their potential added value. An increase in antioxidant activity was observed, with no significant changes during storage

Thermal Degradation Kinetics of Anthocyanins Extracted from Purple Maize Flour Extract and the Effect of Heating on Selected Biological Functionality

The thermal degradation of the anthocyanins and antioxidant activity in purple maize extracts was determined between 80 and 180 °C. The anthocyanins were found to be thermostable in the temperature range of 80 to 120 °C, whereas at higher temperatures the thermal degradation of both anthocyanins and antioxidant activity followed a first-order kinetic model. The z-values started from 61.72 ± 2.28 °C for anthocyanins and 75.75 ± 2.87 °C for antioxidant activity. The conformational space of pairs of model anthocyanin molecules at 25 and 180 °C was explored through a molecular dynamics test, and results indicated the occurrence of intermolecular self-association reactions and intramolecular co-pigmentation events, which might help explaining the findings of the degradation kinetics. The relationship between thermal degradation of anthocyanins and antioxidant activity and the in vitro release was further studied. The unheated extracts showed a high stability under gastric environment, whereas after heating at 180 °C, the digestion ended quickly after 60 min. After simulated intestinal digestion, the anthocyanins were slowly decreased to a maximum of 12% for the unheated extracts, whereas an 83% decrease was found after preliminary heating at 180 °C. The thermal degradation of anthocyanins was positively correlated with the in vitro decrease of antioxidant activity

Cross-Linked Microencapsulation of CO2 Supercritical Extracted Oleoresins from Sea Buckthorn: Evidence of Targeted Functionality and Stability

Oleoresin supercritical extracts from sea buckthorn were microencapsulated in whey proteins isolate and casein, in two states: native (N) and cross-linked mediated by transglutaminase (TG). The encapsulation efficiency showed values higher than 92% for total carotenoids and lycopene. Phytochemicals content was 352.90 ± 1.02 mg/g dry weight (DW) for total carotenoids in TG and 302.98 ± 2.30 mg/g DW in N, with antioxidant activity of 703.13 ± 23.60 mMol Trolox/g DW and 608.74 ± 7.12 mMol Trolox/g DW, respectively. Both powders had an inhibitory effect on α-glucosidase, of about 40% for N and 35% for TG. The presence of spherosomes was highlighted, with sizes ranging between 15.23–73.41 µm and an agglutination tendency in N, and lower sizes, up to 35 µm in TG. The in vitro digestibility revealed a prolonged release in an intestinal environment, up to 65% for TG. Moisture sorption isotherms were studied at 20 °C and the shape of curves corresponds to sigmoidal type II model. The presence of cross-linked mediated aggregates in TG powders improved stability and flowability. Our results can be used as evidence that cross-linked aggregates mediated by transglutaminase applied for microencapsulation of oleoresins have the potential to become new delivery systems, for carotenoids and lycopene, being valuable in terms of their attractive color and biological and bioaccessibility properties