Co-administration of the prebiotic 1-kestose and the paraprobiotic Lactiplantibacillus plantarum FM8 in magellanic penguins promotes the activity of intestinal Lactobacillaceae and reduces the plc gene levels encoding Clostridium perfringens toxin.

Despite the well-known potential health benefits of prebiotics and non-viable probiotics (paraprobiotics) in various animal species, research regarding their use in penguins is scarce. Our study aimed to investigate the impact of a combined administration of prebiotics and paraprobiotics (referred to here as "parasynbiotics") on the gut microbiome and overall health of Magellanic penguins (Spheniscus magellanicus). The parasynbiotics consisted of 1-kestose, which is a fructooligosaccharide comprising sucrose and fructose, and heat-killed Lactiplantibacillus plantarum FM8, isolated from pickled vegetables. It was administered to eight penguins aged <3 years (Young-group) and nine penguins aged >17 years (Adult-group) for 8 weeks. Results from 16S rRNA sequencing revealed that compared to baseline, parasynbiotic administration significantly decreased the relative abundance of intestinal Clostridiaceae_222000 in both groups and significantly increased that of Lactobacillaceae in the Young-group. Quantitative real-time polymerase chain reaction revealed a significant decrease in the plc gene levels encoding alpha-toxin of Clostridium perfringens in the Young-group after parasynbiotic administration (P=0.0078). In the Young-group, parasynbiotic administration significantly increased the plasma levels of total alpha-globulin (P=0.0234), which is associated with inflammatory responses. Furthermore, exposure of dendritic cells to heat-killed L. plantarum FM8 promoted the secretion of interleukin 10, a major anti-inflammatory cytokine. Overall, parasynbiotic administration enhanced the activity of gut Lactobacillaceae, decreased the levels of C. perfringens and its toxin encoding plc gene, and reduced inflammatory response in penguins. These results provide novel insights into the potential benefits of parasynbiotics for improving penguin health.

Preparation and characterization of carboxymethylated Aureobasidium pullulans ß-(1 → 3, 1 → 6)-glucan and its in vitro antioxidant activity.

Aureobasidium pullulans ß-(1 â†’ 3, 1 â†’ 6)-glucan (APG) has a high degree of ß-(1 â†’ 6)-glucosyl branching and a regular triple helical structure similar to that of schizophyllan. In this study, APG was carboxymethylated to different degrees of substitution (DS = 0.51, 1.0, and 2.0, denoted CMAPG 1-3, respectively) using a heterogeneous reaction. With increasing DS, the triple-helix structure drastically decreased and converted to a random coil structure in CMAPG 3. Further, aqueous solutions of CMAPG changed from pseudoplastic fluids to perfect Newtonian liquids with increasing DS, indicating that the intra- and intermolecular hydrogen bonds had been cleaved by the substituents to form a random coil structure. In addition, APG and CMAPG solutions exhibited scavenging ability against hydroxyl, organic, and sulfate radicals. It was also found that the carboxymethylation of APG drastically enhanced the organic radical scavenging ability. On the basis of the relationship between the DS and radical scavenging ability of the CMAPG samples, we believe hydroxyl and organic radicals were preferably scavenged by the donation of hydrogen atoms from the glucose rings and the methylene moieties of the carboxymethyl groups, respectively. Considering the obtained results, CMAPG and APG are expected to have applications in pharmaceuticals, functional foods, and cosmetics as antioxidant polysaccharides.

Combined oral intake of short and long fructans alters the gut microbiota in food allergy model mice and contributes to food allergy prevention.

BACKGROUND: It has become clear that the intestinal microbiota plays a role in food allergies. The objective of this study was to assess the food allergy-preventive effects of combined intake of a short fructan (1-kestose [Kes]) and a long fructan (inulin ([Inu]) in an ovalbumin (OVA)-induced food allergy mouse model. RESULTS: Oral administration of fructans lowered the allergenic symptom score and alleviated the decreases in rectal temperature and total IgA levels and increases in OVA-specific IgE and IgA levels induced by high-dose OVA challenge, and in particular, combined intake of Kes and Inu significantly suppressed the changes in all these parameters. The expression of the pro-inflammatory cytokine IL-4, which was increased in the allergy model group, was significantly suppressed by fructan administration, and the expression of the anti-inflammatory cytokine IL-10 was significantly increased upon Kes administration. 16 S rRNA amplicon sequencing of the gut microbiota and beta diversity analysis revealed that fructan administration may induce gut microbiota resistance to food allergy sensitization, rather than returning the gut microbiota to a non-sensitized state. The relative abundances of the genera Parabacteroides B 862,066 and Alloprevotella, which were significantly reduced by food allergy sensitization, were restored by fructan administration. In Parabacteroides, the relative abundances of Parabacteroides distasonis, Parabacteroides goldsteinii, and their fructan-degrading glycoside hydrolase family 32 gene copy numbers were increased upon Kes or Inu administration. The concentrations of short-chain fatty acids (acetate and propionate) and lactate were increased by fructan administration, especially significantly in the Kes + Inu, Kes, and Inu-fed (Inu, Kes + Inu) groups. CONCLUSION: Combined intake of Kes and Inu suppressed allergy scores more effectively than single intake, suggesting that Kes and Inu have different allergy-preventive mechanisms. This indicates that the combined intake of these short and long fructans may have an allergy-preventive benefit.

Biological Activity of High-Purity ß-1,3-1,6-Glucan Derived from the Black Yeast Aureobasidium pullulans: A Literature Review.

The black yeast Aureobasidium pullulans produces abundant soluble ß-1,3-1,6-glucan-a functional food ingredient with known health benefits. For use as a food material, soluble ß-1,3-1,6-glucan is produced via fermentation using sucrose as the carbon source. Various functionalities of ß-1,3-1,6-glucan have been reported, including its immunomodulatory effect, particularly in the intestine. It also exhibits antitumor and antimetastatic effects, alleviates influenza and food allergies, and relieves stress. Moreover, it reduces the risk of lifestyle-related diseases by protecting the intestinal mucosa, reducing fat, lowering postprandial blood glucose, promoting bone health, and healing gastric ulcers. Furthermore, it induces heat shock protein 70. Clinical studies have reported the antiallergic and triglyceride-reducing effects of ß-1,3-1,6-glucan, which are indicators of improvement in lifestyle-related diseases. The primary and higher-order structures of ß-1,3-1,6-glucan have been elucidated. Specifically, it comprises a single highly-branched glucose residue with the ß-1,6 bond (70% or more) on a backbone of glucose with 1,3-ß bonds. ß-Glucan shows a triple helical structure, and studies on its use as a drug delivery system have been actively conducted. ß-Glucan in combination with anti-inflammatory substances or fullerenes can be used to target macrophages. Based on its health functionality, ß-1,3-1,6-glucan is an interesting material as both food and medicine.

Solid-state relaxation NMR dataset for a water-soluble ß-(1→3, 1→6)-glucan from Aureobasidium pullulans and schizophyllan from Schizophyllum commune.

We report the solid-state nuclear magnetic resonance (NMR) relaxation dataset for a triple helix and a random structure of water-soluble Aureobasidium pullulans ß-(1→3, 1→6)-d-glucan (APG) and those of schizophyllan from Schizophyllum commune (SPG), obtained by the Bruker BioSpin 500 MHz NMR spectrometer. These data include solid-state proton spin-lattice relaxation in the rotating frame (T 1ρH) and 13C spin-lattice relaxation (T 1C) of these two ß-(1→3, 1→6)-glucans, which are related to the subject of article in International Journal of Biological Macromolecules, entitled "Characterization of the secondary structure and order-disorder transition of a ß-(1→3, 1→6)-glucan from Aureobasidium pullulans" [1]. Data can help to investigate the structural characterization of the structural polysaccharides.

Characterization of the secondary structure and order-disorder transition of a ß-(1 → 3, 1 → 6)-glucan from Aureobasidium pullulans.

This study revealed the secondary structures of the water-soluble Aureobasidium pullulans ß-(1 â†’ 3, 1 â†’ 6)-d-glucan (APG) whose primary structural unit is a ß-(1 â†’ 3)-d-glucan backbone with four ß-(1 â†’ 6)-d-glucosyl branching units every six residues. Solid-state NMR spectroscopy, X-ray diffractometry (XRD), and small-angle X-ray scattering (SAXS) experiments involving samples prepared from lyophilized APG showed that APG forms a triple helix in H2O and a random structure in DMSO. In addition, it was revealed that the transformation from the triple helix of APG to the random structure occurs reversibly, and that the triple helix is recovered from the random structure in DMSO/H2O mixtures containing more than 30% H2O. Solid-state NMR and diffraction studies revealed that the triple helix of APG is more stable than that of schizophyllan (SPG) whose structure comprises a ß-(1 â†’ 3)-d-glucan backbone with one ß-(1 â†’ 6)-d-branching unit every three residues. The APG helical pitch is 1.82 nm, which is about 10% longer than that of the triple helix of SPG. These findings show that the ß-(1 â†’ 6) side-chain frequency strongly affects the stability and helical pitch of a ß-(1 â†’ 3, 1 â†’ 6)-d-glucan.

Hydrothermal processing of ß-glucan from Aureobasidium pullulans produces a low molecular weight reagent that regulates inflammatory responses induced by TLR ligands.

The Kururu no ß-glu® (KBG) is a commercial hydrothermal-treated Aureobasidium pullulans ß-glucan produced by a unique hydrothermal process that results in high solubility of the ß-glucan. In this study, we examined the biological activities of this reagent. RAW264.7 cells do not express Dictin-1 on the cell surface, but cells still respond to various pathogen molecular patterns. Lipopolysaccharide (LPS) induced nitrogen oxide (NO) synthesis and TNF-α production in RAW264.7 cells, and those were suppressed by KBG in a dose-dependent manner. The major signaling cell surface receptor respond to LPS is the TLR4/MD-2 complex. The UT12 antibody against to the TLR4/MD-2 complex mimics LPS function and induces cell responses. NO generation and TNF-α production were similarly induced in cells by stimulation with the antibody, but those were not suppressed by KBG. Cell responses induced by other TLR ligands, such as CPG (TLR9 ligand) and Pam3CSK4 (TLR1/TLR2 ligand), were also suppressed by KBG. Therefore, the target molecule for KBG is different from TLR receptors and Dictin-1. Although we also examined the suppressive activities of several other ß-glucan products, comparable activities were not detected with other reagents. A unique hydrothermal process may produce the active reagent. Reprocessing KBG increased low molecular weight fractions, and suppressive activities were markedly enhanced. Therefore, low molecular weight fractions obtained by hydrothermal processing of KBG may result in potential reagents that control inflammation induced by various pathogens.

NMR spectroscopic structural characterization of a water-soluble ß-(1→3, 1→6)-glucan from Aureobasidium pullulans.

An unambiguous structural characterization of the water-soluble Aureobasidium pullulans ß-(1→3, 1→6)-glucan is yet to be achieved, although this ß-(1→3, 1→6)-glucan is expected to exhibit excellent biofunctional properties. Thus, we herein report the elucidation of the primary structure of the A. pullulans ß-(1→3, 1→6)-glucan using nuclear magnetic resonance spectroscopy, followed by comparison of the obtained structure with that of schizophyllan (SPG). Structural characterization of the A. pullulans ß-(1→3, 1→6)-glucan revealed that the structural units are a ß-(1→3)-d-glucan backbone with four ß-(1→6)-d-glucosyl side branching units every six residues. In addition, circular dichroism spectroscopic analysis revealed that the ß-(1→3, 1→6)-glucan interacted with polyadenylic acid (poly(A)) chains in DMSO solution to form a complex similar to that obtained in the complexation of SPG/poly(A). This finding indicates that ß-(1→3, 1→6)-glucan forms a triple-helical conformation in aqueous solution but exhibits a random coil structure in DMSO solution, which is similar to the behavior of SPG.

Two-dimensional NMR data of a water-soluble ß-(1→3, 1→6)-glucan from Aureobasidium pullulans and schizophyllan from Schizophyllum commune.

This article contains two-dimensional (2D) NMR experimental data, obtained by the Bruker BioSpin 500 MHz NMR spectrometer (Germany) which can used for the determination of primary structures of schizophyllan from Schizophyllum commune (SPG) and a water-soluble ß-(1→3, 1→6)-glucan from Aureobasidium pullulans. Data include analyzed the 2D NMR spectra of these ß-glucans, which are related to the subject of an article in Carbohydrate Polymers, entitled "NMR spectroscopic structural characterization of a water-soluble ß-(1→3, 1→6)-glucan from A. pullulans" (Kono et al., 2017) [1]. Data can help to assign the 1H and 13C chemical shifts of the structurally complex polysaccharides.

Production of Gentiobiose from Hydrothermally Treated Aureobasidium pullulans ß-1,3-1,6-Glucan.

We report production of the functional disaccharide gentiobiose ß-D-Glcp-(1→6)-D-Glc by a hydrolysis reaction of hydrothermally treated Aureobasidium pullulans ß-1,3-1,6-glucan as the substrate and Kitalase as the enzyme. Gentiobiose was produced over the pH range 4-6 and the concentration of gentiobiose produced decreased above pH 7. The maximum value of gentiobiose production was unaffected by the enzyme concentration. The maximum concentration of gentiobiose produced was dependent on the substrate concentration whereas the maximum ratio of gentiobiose to glucose was not. The production of gentiobiose from yeast ß-1,3-1,6-glucan was lower than that from A. pullulans ß-1,3-1,6-glucan.

Production of the Functional Trisaccharide 1-Kestose from Cane Sugar Molasses Using Aspergillus japonicus ß-Fructofuranosidase.

We report the production of the functional trisaccharide 1-kestose, O-ß-D-fructofuranosyl-(2→1)-ß-D-fructofuranosyl α-D-glucopyranoside, by ß-fructofuranosidase from Aspergillus japonicus using sugar cane molasses as substrate. Sucrose in cane sugar molasses acted as a fructosyl donor and acceptor for the enzyme. The tetrasaccharide nystose, O-ß-D-fructofuranosyl-(2→1)-ß-D-fructofuranosyl-(2→1)-ß-D-fructofuranosyl α-D-glucopyranoside, was produced from 1-kestose. Cane sugar molasses mixed with water provided a better substrate solution for ß-fructofuranosidase compared to undiluted molasses due to the high concentration of product inhibitors such as glucose and fructose in molasses. The maximum concentration of 1-kestose obtained was 84.9 mg/ml and the maximum production efficiency was 32.3% after 24 h reaction at 40 °C. The maximum efficiency of combined fructo-oligosaccharide (1-kestose and nystose) production was 40.6%. 1-Kestose was therefore produced via a fructosyl-transfer reaction catalyzed by ß-fructofuranosidase from A. japonicus.

Characterization and enzymatic hydrolysis of hydrothermally treated ß-1,3-1,6-glucan from Aureobasidium pullulans.

The chemical structure of hydrothermally treated ß-1,3-1,6-glucan from Aureobasidium pullulans was characterized using techniques such as gas chromatography/mass spectrometry (GC/MS) and nuclear magnetic resonance (NMR). The chemical shifts of anomeric carbons observed in the 13C-NMR spectra suggested the presence of single flexible chains of polysaccharide in the sample. ß-1,3-1,6-Glucan from A. pullulans became water-soluble, with an average molecular weight of 128,000 Da after hydrothermal treatment, and the solubility in water was approximately 10% (w/w). Sample (3% w/v) was completely hydrolyzed to glucose by enzymatic reaction with Lysing enzymes from Trichoderma harzianum. Gentiobiose (Glcß1 â†’ 6Glc) and glucose were released as products during the reaction, and the maximum yield of gentiobiose was approximately 70% (w/w). The molar ratio of gentiobiose to glucose after 1 h reaction suggested that the sample is likely highly branched. Sample (3% w/v) was also hydrolyzed to glucose by Uskizyme from Trichoderma sp., indicating that it is very sensitive to enzymatic hydrolysis.