Search Results (17)

Lactic acid bacteria (LAB) are central to food, feed, and health biotechnology, yet their genomes have long resisted rapid, precise manipulation. This review charts the evolution of LAB genome-editing strategies from labor-intensive RecA-dependent double-crossovers to state-of-the-art CRISPR and CRISPR-associated transposase systems. Native homologous recombination, transposon mutagenesis, and phage-derived recombineering opened the door to targeted gene disruption, but low efficiencies and marker footprints limited throughput. Recent phage RecT/RecE-mediated recombineering and CRISPR/Cas counter-selection now enable scar-less point edits, seamless deletions, and multi-kilobase insertions at efficiencies approaching model organisms. Endogenous Cas9 systems, dCas-based CRISPR interference, and CRISPR-guided transposases further extend the toolbox, allowing multiplex knockouts, precise single-base mutations, conditional knockdowns, and payloads up to 10 kb. The remaining hurdles include strain-specific barriers, reliance on selection markers for large edits, and the limited host-range of recombinases. Nevertheless, convergence of phage enzymes, CRISPR counter-selection and high-throughput oligo recombineering is rapidly transforming LAB into versatile chassis for cell-factory and therapeutic applications. Full article

Sourdough fermentation has emerged as a promising biotechnological approach to reducing gluten content and modifying gluten proteins in wheat-based products. This review assesses the current scientific literature on the enzymatic degradation and hydrolysis of gluten during lactic acid bacteria (LAB) sourdough fermentation. It explores implications for individuals with gluten-related disorders, including celiac disease, non-celiac gluten sensitivity and intolerance, as well as irritable bowel syndrome (IBS). In addition, LAB sourdough effect on fermentable oligo-, di-, monosaccharides and polyols (FODMAPs), amylase-trypsin inhibitors (ATIs), and phytate are revised. Selected homo- and heterofermentative LAB are capable of degrading gluten proteins, especially the polypeptides derived from the action of native cereal proteases. Mixed cultures of LAB degrade gluten peptides more effectively than monocultures. However, LAB sourdough is not sufficient to remove the toxic peptides to the minimal level (<20 ppm). This goal is achieved only if sourdough is combined with fungal proteases during sourdough fermentation. LAB sourdough directly contributes to lower FODMAPs but not ATIs and phytate. Phytate is reduced by the endogenous cereal phytases activated at acidic pHs (pH < 5.0), conditions generated during sourdough fermentation. ATIs are also lowered by endogenous cereal proteases instead of LAB proteases/peptidases. Despite LAB sourdough not fully degrading the gluten or directly reducing the ATIs and phytate, it participates through peptidases activity and acidic pH that trigger the action of endogenous cereal proteases and phytases. Full article

Electrochemical enzyme biosensors are extensively utilized in clinical analysis and environmental monitoring, yet achieving effective enzyme immobilization while maintaining high activity remains a challenge. In this work, we developed a flow-through enzyme biosensor system using a 3D-printed flow-through electrochemical cell fabricated from commercially available poly (lactic acid). After modification with thiacalixarene-functionalized oligo (lactic acids) (OLAs), the material enabled efficient immobilization of uricase on the inner surface of a replaceable reactor of the cell. Swelling and hydrolytic stability of OLAs in cone, partial cone, and 1,3-alternate conformations were studied, with 1,3-alernate conformation demonstrating superior stability and enzyme immobilization performance. The use of OLAs enhanced immobilization efficiency by over 30% and protected the reactor from swelling, hydrolytic degradation, and enzyme loss. The biosensor was validated for amperometric uric acid determination, with a screen-printed carbon electrode modified with carbon black and Prussian Blue. This modification reduced the cathodic potential for uric acid detection to –0.05 V. The biosensor exhibited a linear detection range of 10 nM to 30 μM with a detection limit of 7 nM, and it performed effectively in artificial urine and synthetic blood plasma. The novel cell design, featuring easy assembly and low-cost replaceable parts, makes this biosensor a promising candidate for routine clinical analysis and other practical applications. Full article

Plant protein ingredients are gaining attention for human nutrition, yet they differ significantly from animal proteins in functionality and nutrition. Fungal solid-state fermentation (SSF) can modulate the composition and functionality, increasing their applicability in foods. Quinoa flour (QF) served as a substrate for Aspergillus oryzae and Rhizopus oligosporus, resulting in two fermented ingredients (QFA and QFR) with different nutritional, functional, and aroma characteristics. A higher increase in protein (+35%) and nitrogen (+24%) was observed in the QFA, while fat was predominantly increased in the QFR (+78%). Fermentable oligo-, di-, monosaccharides and polyols (FODMAPs) decreased in the QFR but increased in the QFA due to polyol production. Metabolomic analysis revealed higher lactic acid concentrations in the QFA, and higher citric, malic, and fumaric acid contents in the QFR. The SSF reduced most antinutrients, while R. oligosporus produced saponins. Olfactometry showed the development of fruity ester compounds and a decrease in metallic and cardboard aromas. Both ingredients showed an enhanced water-holding capacity, with the QFA also demonstrating an increased oil-holding capacity. Complex formation increased the particle size, reduced the solubility, and decreased the foaming properties. Mycelium production darkened the ingredients, with the QFR having a higher differential colour index. This study highlights the potential of SSF to produce ingredients with improved nutritional, sensory, and functional properties. Full article

Polymer nanogels—considered as nanoscale hydrogel particles—are attractive for biological and biomedical applications due to their unique physicochemical flexibility. However, the aggregation or accumulation of nanoparticles in the body or the occurrence of the body’s defense reactions still pose a research challenge. Here, we demonstrate the fabrication of degradable nanogels using thermoresponsive, cytocompatible poly[oligo(ethylene glycol) methacrylate]s-based copolymers (POEGMA). The combination of POEGMA’s beneficial properties (switchable affinity to water, nontoxicity, non-immunogenicity) along with the possibility of nanogel degradation constitute an important approach from a biological point of view. The copolymers of oligo(ethylene glycol) methacrylates were partially modified with short segments of degradable oligo(lactic acid) (OLA) terminated with the acrylate group. Under the influence of temperature, copolymers formed self-assembled nanoparticles, so-called mesoglobules, with sizes of 140–1000 nm. The thermoresponsive behavior of the obtained copolymers and the nanostructure sizes depended on the heating rate and the presence of salts in the aqueous media. The obtained mesoglobules were stabilized by chemical crosslinking via thiol-acrylate Michael addition, leading to nanogels that degraded over time in water, as indicated by the DLS, cryo-TEM, and AFM measurements. Combining these findings with the lack of toxicity of the obtained systems towards human fibroblasts indicates their application potential. Full article

Delivery systems for biologically active substances such as proanthocyanidins (PCANs), produced in the form of electrospun nonwoven through the electrospinning method, were designed using a polymeric blend of poly(L-lactide-co-glycolide) (PLGA)and poly[(R,S)-3-hydroxybutyrate] ((R,S)-PHB). The studies involved the structural and thermal characteristics of the developed electrospun three-dimensional fibre matrices unloaded and loaded with PCANs. In the next step, the hydrolytic degradation tests of these systems were performed. The release profile of PCANs from the electrospun nonwoven was determined with the aid of UV–VIS spectroscopy. Approximately 30% of the PCANs were released from the tested electrospun nonwoven during the initial 15–20 days of incubation. The chemical structure of water-soluble oligomers that were formed after the hydrolytic degradation of the developed delivery system was identified through electrospray ionization mass spectrometry. Oligomers of lactic acid and OLAGA oligocopolyester, as well as oligo-3-hydroxybutyrate terminated with hydroxyl and carboxyl end groups, were recognized as degradation products released into the water during the incubation time. It was also demonstrated that variations in the degradation rate of individual mat components influenced the degradation pattern and the number of formed oligomers. The obtained results suggest that the incorporation of proanthocyanidins into the system slowed down the hydrolytic degradation process of the poly(L-lactide-co-glycolide)/poly[(R,S)-3-hydroxybutyrate] three-dimensional fibre matrix. In addition, in vitro cytotoxicity and antimicrobial studies advocate the use of PCANs for biomedical applications with promising antimicrobial activity. Full article

FODMAPs are fermentable oligo-, di-, monosaccharides, and polyols. The application of homofermentative lactic acid bacteria (LAB) has been investigated as a promising approach for producing low-FODMAP whole-wheat bread. The low-FODMAP diet is recommended to treat irritable bowel syndrome (IBS). Wheat flour is staple to many diets and is a significant source of fructans, which are considered FODMAPs. The reduction of fructans via sourdough fermentation, generally associated with heterofermentative lactic acid bacteria (LAB), often leads to the accumulation of other FODMAPs. A collection of 244 wild-type LAB strains was isolated from different environments and their specific FODMAP utilisation profiles established. Three homofermentative strains were selected for production of whole-wheat sourdough bread. These were Lactiplantibacillus plantarum FST1.7 (FST1.7), Lacticaseibacillus paracasei R3 (R3), and Pediococcus pentosaceus RYE106 (RYE106). Carbohydrate levels in flour, sourdoughs (before and after 48 h fermentation), and resulting breads were analysed via HPAEC-PAD and compared with whole-wheat bread leavened with baker’s yeast. While strain R3 was the most efficient in FODMAP reduction, breads produced with all three test strains had FODMAP content below cut-off levels that would trigger IBS symptoms. Results of this study highlighted the potential of homofermentative LAB in producing low-FODMAP whole-wheat bread. Full article

Mannanase with a molecular weight of 33.1 kDa was purified from Weissella cibaria F1. The F1 mannanase contained 289 amino acid residues and shared 70.0% similarity with mannanase from Bacillus subtilis (P55278 (MANB_BACIU)). The optimum reaction conditions of F1 mannanase were 50 °C and pH 6.5. After incubation at pH 4.5–8.0 and 30–60 °C for 2 h, the enzyme activity remained above 60%. The effects of metal ions on mannanase enzyme activity were measured, and Mn²⁺, Mg²⁺, and Cu²⁺ increased enzyme activity. The K_m (16.96 ± 0.01 μmol·mL⁻¹) and V_max (1119.05 ± 0.14 μmol·min⁻¹) values showed that the enzyme exhibited high affinity for locust bean gum. Mannanase was used to hydrolyze konjac glucomannan to produce konjac oligo-glucomannan (KOGM). KOGM increased the proliferation and phagocytosis of RAW264.7 macrophages and enhanced nitric oxide, and cytokine production in macrophages, which showed potent immunostimulatory activity. In this study, the advantages of mannanase derived from lactic acid bacteria were utilized to expand the application of KOGM in the medical field, which is helpful to explore the broad prospects of KOGM in functional food or medicine. Full article

Oligo(L-lactic acid) (OLLA) was synthesized by ring opening polymerization of L-lactides using stannous octoate (0.03 wt% of lactide). While this served as the initiator, L-lactic acids were the co-initiators at 140 °C for 10 h, wherein L-lactic acids were prepared by hydrolytic degradation of L-lactides at 100 °C for 1 h. The molecular weight or degree of polymerization was controlled with monomer/co-initiator ratio (mol/mol). α-cellulose and microcrystalline cellulose (MCC) were extracted from jute fiber by subsequent treatment with sodium chlorite (Na₂ClO₂), NaOH and H₂SO₄. Grafting of OLLA onto α-cellulose and MCC in toluene was carried out using para-toluene sulphonic acid as a catalyst and potassium persulphate (KPS) as an initiator at 130 °C under 380 mm (Hg) pressure for 3, 6, 9, 12, 15, and 18 h. New properties of α-cellulose and MCC were observed due to the successful grafting onto α-cellulose and MCC. Fourier-transform infrared spectroscopy (FTIR) and scanning electron microscope (SEM) were conducted in order to confirm grafting of OLLA onto cellulose and MCC. The FTIR analysis results showed there are some new characteristic absorption peaks appeared (1728 to 1732 cm⁻¹) in the spectrum, which confirmed the grafting of OLLA onto α-cellulose and MCC was successful. SEM images of α-cellulose and MCC before and after grafting revealed significant changes in surface morphology. Grafting of MCC could be more effective for further application in comparison to α-cellulose. Full article

Docetaxel (DTX) is among the most frequently prescribed chemotherapy drugs and has recently been shown to extend survival in advanced prostate cancer patients. However, the poor water solubility of DTX prevents full exploitation of this potent anticancer drug. The current marketed formulation, Taxotere^®, contains a toxic co-solvent that induces adverse reactions following intravenous injection. Nano-sized polymeric micelles have been proposed to create safer, water-soluble carriers for DTX, but many have failed to reach the clinic due to poor carrier stability in vivo. In this study, we aimed to improve micelle stability by synthesizing an ester prodrug of DTX, oligo(lactic acid)₈-docetaxel (o(LA)₈-DTX), for augmented compatibility with the core of poly(ethylene glycol)-b-poly(lactic acid) (PEG-b-PLA) micelles. Due to the enhancement of drug-carrier compatibility, we were able to load 50% (w/w) prodrug within the micelle, solubilize 20 mg/mL o(LA)₈-DTX (~12 mg/mL DTX-equivalent) in aqueous media, and delay payload release. While the micelle core prohibited premature degradation, o(LA)₈-DTX was rapidly converted to parent drug DTX through intramolecular backbiting (t_1/2 = 6.3 h) or esterase-mediated degradation (t_1/2 = 2.5 h) following release. Most importantly, o(LA)₈-DTX micelles proved to be as efficacious but less toxic than Taxotere^® in a preclinical mouse model of prostate cancer. Full article

Poly(L-lactic acid) (PLLA) has attracted a great deal of attention for its use in biomedical materials such as biodegradable vascular scaffolds due to its high biocompatibility. However, its inherent brittleness and inflammatory responses by acidic by-products of PLLA limit its application in biomedical materials. Magnesium hydroxide (MH) has drawn attention as a potential additive since it has a neutralizing effect. Despite the advantages of MH, the MH can be easily agglomerated, resulting in poor dispersion in the polymer matrix. To overcome this problem, oligo-L-lactide-ε-caprolactone (OLCL) as a flexible character was grafted onto the surface of MH nanoparticles due to its acid-neutralizing effect and was added to the PLLA to obtain PLLA/MH composites. The pH neutralization effect of MH was maintained after surface modification. In an in vitro cell experiment, the PLLA/MH composites including OLCL-grafted MH exhibited lower platelet adhesion, cytotoxicity, and inflammatory responses better than those of the control group. Taken together, these results prove that PLLA/MH composites including OLCL-grafted MH show excellent augmented mechanical and biological properties. This technology can be applied to biomedical materials for vascular devices such as biodegradable vascular scaffolds. Full article

Fermentable oligo-, di-, and monosaccharides and polyols (FODMAPs) are associated with digestive disorders and with diseases such as irritable bowel syndrome. In this study, we determined the FODMAP contents of bread, bakery products, and flour and assessed the effectiveness of sourdough fermentation for FODMAP reduction. The fermentation products were analyzed to determine the DP 2–7 and DP >7 fructooligosaccharide (FOS) content of rye and wheat sourdoughs. FOSs were reduced by Acetobacter cerevisiae, Acetobacter okinawensis, Fructilactobacillus sanfranciscensis, and Leuconostoc citreum to levels below those in rye (−81%; −97%) and wheat (−90%; −76%) flours. The fermentation temperature influenced the sourdough acetic acid to lactic acid ratios (4:1 at 4 °C; 1:1 at 10 °C). The rye sourdough contained high levels of beneficial arabinose (28.92 g/kg) and mannitol (20.82 g/kg). Our study contributes in-depth knowledge of low-temperature sourdough fermentation in terms of effective FODMAP reduction and concurrent production of desirable fermentation byproducts. Full article

In the perspective of producing a rigid renewable and environmentally friendly rigid packaging material, two comb-like copolymers of cellulose acetate (AC) and oligo(lactic acid) OLA, feeding different percentages of oligo(lactic acid) segments, were prepared by chemical synthesis in solvent or reactive extrusion in the melt, using a diepoxide as the coupling agent and were used as compatibilizers for poly(lactic acid)/plasticized cellulose acetate PLA/pAC blends. The blends were extruded at 230 °C or 197 °C and a similar compatibilizing behavior was observed for the different compatibilizers. The compatibilizer C1 containing 80 wt% of AC and 14 wt% of OLA resulted effective in compatibilization and it was easily obtained by reactive extrusion. Considering these results, different PLAX/pAC(100-X) compounds containing C1 as the compatibilizer were prepared by extrusion at 197 °C and tested in terms of their tensile and impact properties. Reference materials were the uncompatibilized corresponding blend (PLAX/pAC(100-X)) and the blend of PLA, at the same wt%, with C1. Significant increase in Young’s modulus and tensile strength were observed in the compatibilized blends, in dependence of their morphologic features, suggesting the achievement of an improved interfacial adhesion thanks to the occurred compatibilization. Full article

Chronic kidney disease (CKD) is a global epidemic with an increasing prevalence worldwide. Effective preventive strategies are urgently needed. This study aimed to investigate the effect of nutraceutical components, a fermented soybean product (ImmuBalance, IMB) and an oligo-lactic acid product (LAP), on the prevention of adenine-induced CKD in mice. Female C57BL/6 mice were randomly assigned into following experimental groups: negative control; model control; and models treated with IMB at 250 or 1000 mg/kg body weight (BW), LAP at 1000 or 2000 mg/kg BW, and IMB/LAP combinations. The CKD model was established by intraperitoneal injection of adenine daily for 4 weeks, and treatments started 2 weeks before adenine injection and ended after 10 weeks. Compared with the model control, the treatments did not significantly alter the body weight or food intake. Both IMB and LAP, especially their combination, significantly inhibited tubular dilation, tubulointerstitial degeneration or atrophy, interstitial chronic inflammation and acute inflammation in the kidneys of CKD mice, and significantly decreased serum cystatin C levels. IMB or LAP significantly reversed CKD-associated increases of circulating and kidney levels of inflammatory cytokines, circulating levels of kidney injury biomarkers, and kidney levels of stem cell biomarkers, and significantly reversed CKD-associated reduction of cecum Clostridium leptum group. Our results suggest that dietary supplementation of IMB or LAP may significantly delay the development and/or progression of CKD. Full article

It has been suggested that sourdough fermented products have beneficial health effects. Fungal proteases and selected sourdough lactic acid bacteria were used to produce wheat bread and pasta with a reduced-gluten content (<50% of traditional products). Fermentable oligo-, di- and mono- saccharides and polyols and amylase/trypsin inhibitors were also evaluated. The sensorial features of new products were similar to traditional ones. The efficacy of these new products in reducing the severity of symptoms in Irritable Bowel Syndrome (IBS) patients were compared to traditional bread and pasta using a randomized, crossover-controlled trial. While on a strict gluten-free diet, patients were randomized to consume a reduced- or normal-gluten diet for 2weeks; then, patients from both arms started the wash-out period of one week, and subsequently started the final 2-week period on a normal or reduced-gluten diet. Compared to normal-gluten content, the administration of a reduced-gluten content diet resulted in a decrease of the Visual Analogue Scale score (p = 0.042), while no differences were found in the IBS-Severity Score, Hospital Anxiety and Depression Scale, and IBS-Quality of Life. Data herein reported are novel encouraging findings that should spur a new avenue of research aiming to develop products specifically designed for IBS patients. Full article