Lactiplantibacillus plantarum MIUG BL21 paraprobiotics: Evidences on inactivation kinetics and their potential as cytocompatible and antitumor alternatives.

Two new -biotics concepts, such as paraprobiotics and postbiotics were introduced, with beneficial effects beyond the viability of probiotic. In this study, the effect of individual (thermal, ohmic heating, high pressure, and ultrasound) and combined (ohmic, high pressure and ultrasound in combination with heating) treatments on the inactivation kinetics of Lactiplantibacillus plantarum was investigated. Different inactivation rates were obtained, up to 8.18 after 10 min at 90 °C, 2.07 after 15 min at a voltage gradient of 20 V/cm, 6.62 after 10 min at 600 MPa and 3.6 after ultrasound treatment for 10 min at 100 % amplitude. The experimental data were fitted to Weibullian model proposed by Peleg, allowing to estimate the inactivation rate coefficient (b) and the shape of the inactivation curves (n). At lower concentration, the samples showed both cytocompatibility and antiproliferative effect, stimulating the cell proliferation on both murine fibroblast and human colorectal adenocarcinoma cell lines.

Kombucha and Water Kefir Grains Microbiomes' Symbiotic Contribution to Postbiotics Enhancement.

Wild artisanal cultures, such as a symbiotic culture of bacteria and yeasts (SCOBY) and water kefir grains (WKG), represent a complex microorganism consortia that is composed of yeasts and lactic and acetic acid bacteria, with large strains of diversity and abundance. The fermented products (FPs) obtained by the microbiome's contribution can be included in functional products due to their meta-biotics (pre-, pro-, post-, and paraprobiotics) as a result of complex and synergistic associations as well as due to the metabolic functionality. In this study, consortia of both SCOBY and WKG were involved in the co-fermentation of a newly formulated substrate that was further analysed, aiming at increasing the postbiotic composition of the FPs. Plackett-Burman (PBD) and Response Surface Methodology (RSM) techniques were employed for the experimental designs to select and optimise several parameters that have an influence on the lyophilised starter cultures of SCOBY and WKG activity as a multiple inoculum. Tea concentration (1-3%), sugar concentration (5-10%), raisins concentration (3-6%), SCOBY lyophilised culture concentration (0.2-0.5%), WKG lyophilised culture concentration (0.2-0.5%), and fermentation time (5-7 days) were considered the independent variables for mathematical analysis and fermentation conditions' optimisation. Antimicrobial activity against Bacillus subtilis MIUG B1, Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 25922, and Aspergillus niger MIUG M5, antioxidant capacity (DPPH), pH and the total acidity (TA) were evaluated as responses. The rich postbiotic bioactive composition of the FP obtained in optimised biotechnological conditions highlighted the usefulness of the artisanal co-cultures, through their symbiotic metabolic interactions for the improvement of bioactive potential.

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.

Attempts for Developing Novel Sugar-Based and Sugar-Free Sea Buckthorn Marmalades.

Sea buckthorn (Hippophae rhamnoides L.) is recognized as a valuable source of vitamin C and antioxidants, frequently used as nutraceuticals and cosmeceuticals. In the present study, attempts are made to produce and characterize a novel type of marmalade using sea buckthorn berries processed at 102 °C into marmalade in two combinations, with whole cane or stevia sugar. Changes in the phytochemical profile, antioxidant activity, color, shelf-life, texture, microbiological, and sensorial characteristics were determined. The total carotenoids content in the marmalades were significantly different, with values of 0.91 ± 0.03 mg/g dry weight (DW) in the sample with whole sugar cane (Cz) and 2.69 ± 0.14 mg/g DW in the sample with Stevia sugar (Cs). Significant values of polyphenols were found, of 59.41 ± 1.13 mg GAE/g DW in Cz and 72.44 ± 2.31 mg GAE/g DW in Cs, leading to an antioxidant activity of 45.12 ± 0.001 µMol Trolox/g DW and 118.07 ± 0.01 µMol Trolox/g DW, respectively. Accelerated storage study showed a decrease in all the phytochemicals, however no significant changes were found in antioxidant activity. Values of <100 CFU/g for yeasts and molds and <5 CFU/g for Enterobacteriaceae after 21 days of storage at the room temperature of the marmalades were determined. The sensorial and color results were more than acceptable. Overall, the results highlighted the potential of using sea buckthorn as a potential rich source of bioactive compounds to be used in the sugar-based products manufacturing.

Co-Microencapsulated Black Rice Anthocyanins and Lactic Acid Bacteria: Evidence on Powders Profile and In Vitro Digestion.

Two multi-functional powders, in terms of anthocyanins from black rice (Oryza sativa L.) and lactic acid bacteria (Lactobacillus paracasei, L. casei 431®) were obtained through co-microencapsulation into a biopolymer matrix composed of milk proteins and inulin. Two extracts were obtained using black rice flour as a raw material and hot water and ethanol as solvents. Both powders (called P1 for aqueous extract and P2 for ethanolic extract) proved to be rich sources of valuable bioactives, with microencapsulation efficiency up to 80%, both for anthocyanins and lactic acid bacteria. A higher content of anthocyanins was found in P1, of 102.91 ± 1.83 mg cyanindin-3-O-glucoside (C3G)/g dry weight (DW) when compared with only 27.60 ± 17.36 mg C3G/g DW in P2. The morphological analysis revealed the presence of large, thin, and fragile structures, with different sizes. A different pattern of gastric digestion was observed, with a highly protective effect of the matrix in P1 and a maximum decrease in anthocyanins of approximatively 44% in P2. In intestinal juice, the anthocyanins decreased significantly in P2, reaching a maximum of 97% at the end of digestion; whereas in P1, more than 45% from the initial anthocyanins content remained in the microparticles. Overall, the short-term storage stability test revealed a release of bioactive from P2 and a decrease in P1. The viable cells of lactic acid bacteria after 21 days of storage reached 7 log colony forming units (CFU)/g DW.

Fostering Lavender as a Source for Valuable Bioactives for Food and Pharmaceutical Applications through Extraction and Microencapsulation.

Lavender flowers were used in this study as a source of phytochemicals as naturally occurring antioxidants. Two different extraction techniques were applied, such as ultrasound-assisted (UAE) and supercritical fluids (SCE) methods. The comparative evaluation of the phytochemicals profile evidenced a higher content of chlorophyll a and b of 5.22 ± 0.12 mg/g dry weight (D.W.) and 2.95 ± 0.16 mg/g D.W, whereas the carotenoids content was 18.24 ± 0.04 mg/g D.W. in the SCE extract. Seven main compounds were found in both extracts: ß-linalool, eucalyptol, linalool acetate, ß-trans-ocimene, and limonene in SCE and linalool acetate, ß-linalool, 6-methyl-2-(2-oxiranyl)-5-hepten-2-ol, linalool oxide, lavandulyl acetate and camphor in UAE. The (n-3) acids had a higher contribution in SCE. The extracts were microencapsulated in different combinations of wall materials based on polysaccharides and milk proteins. The four variants showed different phytochemical and morphological profiles, with a better encapsulating efficiency for proteins (up to 98%), but with a higher content of encapsulated carotenoids for polysaccharides, the latter showing remarkable antimicrobial activity against selected microorganisms. Carboxymethyl cellulose and whey proteins led to a double encapsulation of lipophilic compounds. The powders were tested in two food matrices as ingredients, with multiple targeted functions, such as flavoring, antimicrobial, antioxidant activity that can successfully replace synthetic additives.

Yarrowia lipolytica and Lactobacillus paracasei Solid State Fermentation as a Valuable Biotechnological Tool for the Pork Lard and Okara's Biotransformation.

This study reports the biovalorization of the two agri-food by-products (pork lard and freeze-dried okara) through solid-state fermentation using a monoculture of Yarrowia lipolytica or a co-culture of Y. lipolytica and Lactobacillus paracasei, for developing a valuable fermented product with antioxidant and antimicrobial activity. First, some yeast strains were selected based on their properties to produce enzymes (protease and lipase) by cultivation on 5% (w/v) pork lard or 2% (w/v) freeze-dried okara. Two selected strains, Y. lipolytica MIUG D5 and Y. lipolytica ATCC 18942, were further used for the fermentation alone or in a co-culture with L. paracasei MIUG BL2. The Plackett-Burman experimental design was used to establish the effects of the fermentation parameters in order to obtain a fermented product with improved antioxidant and antimicrobial activities. As the Plackett-Burman experimental design are independent variables, the concentrations of the freeze-dried okara, pork lard, glycerol, inoculums type, inoculum concentration, and the fermentation time were analyzed. The 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging potential and the antimicrobial activity against aerobic spore-forming microorganisms were assessed as responses. For the fermented products, an antioxidant potential between 6.77-17.78 mM TE/g was obtained while the antimicrobial activity against Aspergillus niger ranged from 24 to 64%. Based on the statistical analysis, the time of the yeast fermentation and the concentration of pork lard were selected as variables with the influence on the SSF fermentation process and the functional properties of the fermented product. In the general context of a circular economy, the results demonstrate the possibility of bio-transforming the freeze-dried okara and the pork lard using Y. lipolytica as a valuable workhorse for the lactic acid bacteria (LAB) metabolism and postbiotics production into a fermented product, which is recommended for use as a food and feed ingredient with biotic properties.

Three Types of Beetroot Products Enriched with Lactic Acid Bacteria.

Beetroot (Beta vulgaris L.) represents a very rich source of bioactive compounds such as phenolic compounds and carotenoids, among which the most important being betalains, mainly betacyanins and betaxanthins. The beetroot matrix was used in a fresh or dried form or as lyophilized powder. A 1012 CFU/g inoculum of Lactobacillus plantarum MIUG BL3 culture was sprayed on the vegetal tissue. The lactic acid bacteria (LAB) viability for all the products was evaluated over 21 days, by microbiological culture methods. The antioxidant activity of the obtained food products was correlated to the betalains content and the viability of LAB. The content of polyphenolic compounds varied between 225.7 and 1314.7 mg L-1, hence revealing a high content of bioactive compounds. Through the confocal laser scanning microscopy analysis, a large number of viable probiotic cells were observed in all the variants but especially in the fresh red beet cubes. After 21 days of refrigeration, the high content of Lb. plantarum (CFU per gram) of the food products was attributed to the biocompounds and the nutrients of the vegetal matrix that somehow protected the bacterial cells, and thus maintained their viability. The obtained food products enriched with probiotic LAB can be regarded as new functional food products due to the beneficial properties they possessed throughout the undertaken experiments.

Involvement of lytic enzymes and secondary metabolites produced by Trichoderma spp. in the biological control of Pythium myriotylum.

This investigation was aimed to evaluate the antimicrobial activities and involvement of extracellular lytic enzymes produced by four strains of Trichoderma in the inhibition of Pythium myriotylum. Antagonistic effects were tested by dual culture. Activities of lytic enzymes were evaluated from the filtrate of each strain after cultivation in selected media. Organic extracts were obtained from liquid media subsequent to the cultivation of Trichoderma in potato dextrose broth (PDB). Non-volatile organic compounds such as polyphenols and flavonoids were evaluated spectrophotometrically while volatile organic compounds (VOCs) were analyzed by gas chromatography coupled with mass spectrometry (GC-MS). The antimicrobial activity of the organic extracts was determined using the poisoning method. Results have shown that all the strains were antagonists against P. myriotylum. T. erinaceum (IT-58), T. gamsii (IT-62), T. afroharzianum (P8), and T. harzianum (P11) that were found to produce cellulase, protease, and xylanase. Over 20 compounds were identified in each extract, including esters, lactones, and organic acids. The organic extracts also contained high amounts of polyphenolic compounds and flavonoids and significantly inhibited the mycelial growth of P. myriotylum. The minimal inhibition concentrations were 80 µg/µL, 40 µg/µL, 20 µg/µL, and 10 µg/µL, for extracts obtained from T. erinaceum (IT-58), T. gamsii (IT-62), T. afroharzianum (P8), and T. harzianum (P11), respectively. There was significant correlation between the production of total polyphenol and flavonoid content and the antagonistic effects of the tested strains.