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Found 1 structure.
Displayed structure 1
| -6)-a-D-Glcp-(1-4)-a-D-Glcp-(1-4)-a-D-Glcp-(1- | Show graphically |
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Structure type: polymer chemical repeating unit
Trivial name: pullulan
Compound class: EPS, O-polysaccharide, cell wall polysaccharide, glucan, polysaccharide
Contained glycoepitopes: IEDB_140629,IEDB_142488,IEDB_144998,IEDB_146664,IEDB_420419,IEDB_420420,IEDB_420421,IEDB_857742,IEDB_983931,SB_192
The batch production of different exopolysaccharides (alginate, xanthan, pullulan, dextran) by free and immobilized microbial cultures was investigated. First, conventional free-cell cultures were performed to obtain control fermentation parameters and macromolecular characteristics of exopolysaccharides. Then microbial cultures were immobilized in composite agar layer/microporous membrane structures and tested for polysaccharide production. The immobilized-cell system proved unsuitable for xanthan and pullulan production. Owing to the fouling of the microporous membrane by the polysaccharide, dextran production by immobilized Leuconostoc mesenteroides also was inefficient. More promising results have been obtained with immobilized Azotobacter vinelandii cultures. The amount of alginate produced by immobilized A. vinelandii represented about 60% of that recovered from a free-cell culture, whereas the polysaccharide yield reached 35% instead of 9% for the free counterpart. These results are compared to the macromolecular characteristics of exopolysaccharides.
Publication DOI: 10.1016/0141-0229(94)90141-4Three water-soluble, non-ionic extracellular polysaccharides (EPS) obtained from lactic acid bacteria (S. thermophilus THS, L. helveticus K16 and S. thermophilus ST1) were subjected to a comparative study by means of multidetector size-exclusion chromatography, providing distributions and averages of molar masses, radii of gyration and intrinsic viscosities. All polysaccharides displayed random coil character. Further analysis of the data reveals differences in chain stiffness and extension that could be well correlated to structural features. The calculated persistence lengths ranged from 5 to 10nm and fall within the range typical for many single-stranded bacterial or plant polysaccharides. The ST1 polysaccharide had the highest molar mass but the lowest persistence length, which is attributed to the presence of the flexible (1→6)-linkage in the main chain.
Lactic acid bacteria, EPS, intrinsic viscosity, Persistence length, SEC-MALLS
NCBI PubMed ID: 26917378This study examined the response of various forms and sources of glucans toward two different Limulus amebocyte lysate (LAL) methods, the modified LAL, and Glucatell. The glucans studied were curdlan, laminarin, yeast glucan, barley glucan, paramylon, pullulan, pustulan, mannan, and pachyman (as part of the Glucatell kit). Both methods provided largely similar results for each of the glucans; however, the Glucatell method yielded slightly higher responses to certain structures that may not necessarily be of fungal origin, leading to falsely greater positive results. The performance of each method to measure fungal glucan concentration specifically was then assessed.
glucan, Curdlan, dectin receptor, glucatell, LAL, Limulus amebocyte lysate
NCBI PubMed ID: 21830869ZG16p is a soluble mammalian lectin that interacts with mannose and heparan sulfate. Here we describe detailed analysis of the interaction of human ZG16p with mycobacterial phosphatidylinositol mannosides (PIMs) by glycan microarray and NMR. Pathogen-related glycan microarray analysis identified phosphatidylinositol mono- and di-mannosides (PIM1 and PIM2) as novel ligand candidates of ZG16p. Saturation transfer difference (STD) NMR and transferred NOE experiments with chemically synthesized PIM glycans indicate that PIMs preferentially interact with ZG16p by using the mannose residues. The binding site of PIM was identified by chemical-shift perturbation experiments with uniformly 15N-labeled ZG16p. NMR results with docking simulations suggest a binding mode of ZG16p and PIM glycan; this will help to elucidate the physiological role of ZG16p
NMR spectroscopy, chemical synthesis, lectins, carbohydrate microarrays, microarrays, phosphatidyl inositol mannoside
NCBI PubMed ID: 25919894Currently, despite the thoroughgoing scientific research carried out in the area of wound healing management, the treatment of skin injuries, regardless of etiology remains a big provocation for health care professionals. An optimal wound dressing should be nontoxic, non-adherent, non-allergenic, should also maintain a humid medium at the wound interfacing, and be easily removed without trauma. For the development of functional and bioactive dressings, they must meet different conditions such as: The ability to remove excess exudates, to allow gaseous interchange, to behave as a barrier to microbes and to external physical or chemical aggressions, and at the same time to have the capacity of promoting the process of healing by stimulating other intricate processes such as differentiation, cell adhesion, and proliferation. Over the past several years, various types of wound dressings including hydrogels, hydrocolloids, films, foams, sponges, and micro/nanofibers have been formulated, and among them, the electrospun nanofibrous mats received an increased interest from researchers due to the numerous advantages and their intrinsic properties. The drug-embedded nanofibers are the potential candidates for wound dressing application by virtue of: Superior surface area-to volume ratio, enormous porosity (can allow oxy-permeability) or reticular nano-porosity (can inhibit the microorganisms'adhesion), structural similitude to the skin extracellular matrix, and progressive electrospinning methodology, which promotes a prolonged drug release. The reason that we chose to review the formulation of electrospun nanofibers based on polysaccharides as dressings useful in wound healing was based on the ever-growing research in this field, research that highlighted many advantages of the nanofibrillary network, but also a marked versatility in terms of numerous active substances that can be incorporated for rapid and infection-free tissue regeneration. In this review, we have extensively discussed the recent advancements performed on electrospun nanofibers (eNFs) formulation methodology as wound dressings, and we focused as well on the entrapment of different active biomolecules that have been incorporated on polysaccharides-based nanofibers, highlighting those bioagents capable of improving the healing process. In addition, in vivo tests performed to support their increased efficacy were also listed, and the advantages of the polysaccharide nanofiber-based wound dressings compared to the traditional ones were emphasized.
polysaccharides, cellulose, Alginates, Dextran, pullulan, chitosan, electrospinning, electrospun nanofibers, gums, hyaluronic acid, pectins, starch, wound dressings, wound healing
NCBI PubMed ID: 33080849A mutant strain of the deuteromycete Aureobasidium pullulans deficient in melanin synthesis was used to investigate the production of the exopolysaccharide pullulan and biomass, respectively. Shake-flask experiments with different carbon sources showed significant differences in pullulan elaboration. Sucrose was most suitable for pullulan synthesis among the carbon sources examined. Fermentations were carried out both batch-wise and continuously in a stirred vessel fermentator. In batch fermentations about 45% of the glucose offered was converted into pullulan at maximum formation rates of 0.16 g/l per hour using standard medium. The yield of polysaccharide could be maintained at 45% in continuous fermentations. At a dilution rate of 0.05 l/h, the formation rate of polysaccharide increased up to 0.35 g/l per hour. Alterations in the nitrogen content of the feed significantly affected the consumption rate of glucose and the production rate of polysaccharide, but final concentrations of biomass were hardly affected.
polysaccharide, fermentation, pullulan, Aureobasidium pullulans, Biomass
Publication DOI: 10.1007/BF00228599Isopullulanase (IPU) from Aspergillus niger A.T.C.C. (American Type Culture Collection) 9642 hydrolyses pullulan to isopanose. IPU is important for the production of isopanose and is used in the structural analysis of oligosaccharides with α-1,4 and α-1,6 glucosidic linkages. We have isolated the ipuA gene encoding IPU from the filamentous fungi A. niger A.T.C.C. 9642. The ipuA gene encodes an open reading frame of 1695 bp (564 amino acids). IPU contained a signal sequence of 19 amino acids, and the molecular mass of the mature form was calculated to be 59 kDa. IPU has no amino-acid-sequence similarity with the other pullulan-hydrolysing enzymes, which are pullulanase, neopullulanase and glucoamylase. However, IPU showed a high amino-acid-sequence similarity with dextranases from Penicillium minioluteum (61%) and Arthrobacter sp. (56%). When the ipuA gene was expressed in Aspergillus oryzae, the expressed protein (recombinant IPU) had IPU activity and was immunologically reactive with antibodies raised against native IPU. The substrate specificity, thermostability and pH profile of recombinant IPU were identical with those of the native enzyme, but recombinant IPU (90 kDa) was larger than the native enzyme (69-71 kDa). After deglycosylation with peptide-N-glycosidase F, the deglycosylated recombinant IPU had the same molecular mass as deglycosylated native enzyme (59 kDa). This result suggests that the carbohydrate chain of recombinant IPU differed from that of the native enzyme.
NCBI PubMed ID: 9169610Pullulan is a water-soluble glucan gum produced aerobically by growing a yeast like fungus Aureobasidium pullulans. It is a regularly repeating copolymer with the chemical structure {→6)-α-d-glucopyranosyl-(1→4)-α-d-glucopyranosyl-(1→4)-α-d-glucopyranosyl-(1→}n. Thus the polysaccharide is viewed as a succession of α-(1→6)-linked (1→4)-α-d-triglucosides i.e. maltotriose (G3). Pullulan have a wide range of commercial and industrial applications in many fields like food science, health care, pharmacy and even in lithography. Due to its strictly linear structure, pullulan is also very valuable in basic research as well as a well-defined model substance. This review attempts to critically appraise the current literature on fungal exopolysaccharide (EPS) 'pullulan' considering its microbial sources, structural geometry, upstream processing, downstream processing, peculiar characteristics and applications.
pullulan, Aureobasidium pullulans, Maltotriose, Upstream processing, Downstream processing
NCBI PubMed ID: 26048217The structure of pullullan-like polysaccharides produced as exocellular material by different strains of Cryphonectria parasitica, the fungus responsible for chestnut tree cankers, was investigated with nuclear magnetic resonance (NMR) techniques. 13C, mono- and bidimensional 1H, and 1H–13C heteronuclear correlated NMR spectra (HSQC and HMBC) were recorded. Advanced analysis of the NMR spectra allowed the main resonance of the atoms in the maltotriose and in the maltotetraose repeat units of pullulan-like polysaccharides from C. parasitica to be recognised with confidence. In all cases investigated, the presence of large amounts of α-(1→6) maltotetraose subunits was evidenced, in addition to the α-(1→6) maltotriose subunits, corresponding to the repeating unit of pullulan produced by Aureobasidium pullulans and other fungi. The results were in agreement with other data from this laboratory, obtained with independent techniques. The belief that in ‘pullulans’ the maximum amount of α-(1→6) maltotetraose subunits is about 7% can thus be considered as definitely outdated.
structure, NMR spectroscopy, Cryphonectria parasitica, Exopolysaccharide(s), Pullulan(s)
Publication DOI: 10.1016/j.carbpol.2005.11.012O-linked heterogalactomannans with similar structural features have been purified from the fungal walls of the entomopathogenic fungi Lecanicillium muscarium, Beauveria bassiana, Beauveria brongniartii, and Cordyceps sphingum. Their composition and structure have been determined using acid hydrolysis, methylation analysis, gas-liquid chromatography-mass spectrometry (GC-MS) and Nuclear magnetic resonance spectroscopy (NMR). All structures have an α-(1→6)-mannose backbone, but one of the two strains of L. muscarium included in this study contained an acidic heterogalactomannan instead of the neutral polysaccharide isolated in the rest of the species analyzed. Sequence analysis of the internal transcribed spacer (ITS) region of this strain indicated that it belongs to the related genus Simplicillium, displaying low identity (83 %) with the closest Lecanicillium species. This is a new demonstration of the structural diversity of fungal wall heteromannans and validates their interest as chemotaxonomic markers. The production of a pullulan-like extracellular polysaccharide in strain CBS 413.70C of L. muscarium is also reported.
cell wall, nuclear magnetic resonance spectroscopy, fungi, galactomannans, entomopathogenic, O-linked polysaccharides
NCBI PubMed ID: 21872183Phenotypic and molecular characterization of five yeast-like fungal isolates from flowers of wild plants showed that they are related to Aureobasidium pullulans. Compared to other isolates, an osmotolerant and non-pigmented isolate A. pullulans RBF-4A3 produced 26.35 g l-1 of melanin-free exopolysaccharide (EPS) in 96 h at 30 °C in 5% glucose containing medium. At higher concentrations of glucose (7.5-25% (w/v)), the EPS produced by this organism increased from 34.68 to 66.79 g l-1 up to 15% (w/v) glucose, with a productivity of 16.69 g l-1 per day. Beyond 15% (w/v) glucose concentration, the EPS production decreased gradually to 43.29 g l-1 at 25% (w/v) glucose. Fourier-transform infrared (FTIR) spectroscopy confirmed that chemical structures of the exopolysaccharide produced by A. pullulans RBF-4A3 and standard pullulan were identical. This is the first report of pullulan production at 15% (w/v) concentration of glucose by an osmotolerant strain of A. pullulans.
exopolysaccharide, fermentation, pullulan, Aureobasidium pullulans, osmotolerant
Publication DOI: 10.1016/j.carbpol.2010.10.003Pullulan is a linear glucosic polysaccharide produced by the polymorphic fungus Aureobasidium pullulans, which has long been applied for various applications from food additives to environmental remediation agents. This review article presents an overview of pullulan's chemistry, biosynthesis, applications, state-ofthe- art advances in the enhancement of pullulan production through the investigations of enzyme regulations, molecular properties, cultivation parameters, and bioreactor design. The enzyme regulations are intended to illustrate the influences of metabolic pathway on pullulan production and its structural composition. Molecular properties, such as molecular weight distribution and pure pullulan content, of pullulan are crucial for pullulan applications and vary with different fermentation parameters. Studies on the effects of environmental parameters and new bioreactor design for enhancing pullulan production are getting attention. Finally, the potential applications of pullulan through chemical modification as a novel biologically active derivative are also discussed.
biosynthesis, production, pullulan, Aureobasidium pullulans, Applications
NCBI PubMed ID: 21800030Aureobasidium pullulans is the source of commercially produced pullulan, a high molecular weight polysaccharide that is used in the manufacture of edible films. It has been proposed that α-amylase decreases the molecular weight of pullulan in late cultures. Based on a recent phylogenetic analysis, five representative strains were chosen to study the relationship between α-amylase and pullulan production. In sucrose-grown cultures, pullulan yields increased over time while the molecular weight of pullulan generally decreased. However, no α-amylase activity was detected in these cultures. Low levels of α-amylase were present in starch-grown culture, but pullulan analysis was complicated by residual starch. To facilitate further studies on the role of α-amylase in the reduction of pullulan molecular weight, the α-amylase gene from A. pullulans NRRL Y-12974 was cloned and characterized. The coding region of the complete α-amylase gene contains 2,247 bp, including 7 introns and 8 exons. The putative mRNA was 1,878 bp long, encoding an α-amylase of 625 amino acid residues. Southern blot analysis indicated that there was only one copy of this gene in the genome. Reverse transcription-polymerase chain reaction (RT-PCR) analysis indicated that the gene was transcribed in both sucrose- and starch-grown cultures. It is possible that very low levels of α-amylase attack the minor maltotetraose subunits of pullulan and cause the reduction of molecular weight.
molecular weight, pullulan, Aureobasidium pullulans, α-amylase
NCBI PubMed ID: 21113644Pullulan is a water-soluble extracelluler polysaccharide produced aerobically by fungus Aureobasidium pullulans and some other species. Current work focused on developing an A. pullulans strain with enhanced pullulan productivity and raw material utilization ratio using a new method for genome shuffling of A. pullulans N3.387. After the third cycle of the protoplast fusion, a mutant, designated as F3-2, producing 179.7% (20.7 g/L) more pullulan than the wild-type strain was obtained. Furthermore, the raw material utilization ratio of F3-2 was determined as 97.9%, which was 29.0% higher than the wild-type strain. Genetic stability analysis of F3-2 showed that ranges of the pullulan production and the raw material utilization ratio of the generation were 0.13 g/L and 1.93%, respectively, suggesting that F3-2 was genetically stable. Elucidation of this new genome shuffling process is also interesting for enhancing production of extracellular polysaccharide in both A. pullulans and other fungus.
pullulan, Aureobasidium pullulans, genetic stability, genome shuffling, inactive protoplast fusion, protoplast formation
Publication DOI: 10.1016/j.procbio.2010.11.004The carbon-13 nmr spectra of two cyclodextrins and several linear glucans have been completely assigned. These assignments were made by comparison with the spectra of glucose, some of its specifically O-methylated derivatives, and a number of differently linked glucobioses and glucotrioses. This technique enables the composition, structure, and major sequence of a number of glucans to be determined. Conformational effects are also apparent in some of the spectra. The 1→4, 1→4, and 1→6 linkage sequence of the glucans from T. mesenterica and P. pullulans give, in addition to an anomeric signal for the 1→6 linkage, two separate anomeric signals for each of the 1→4 linkages. This multiplicity, apparent also in other carbon signals of the spectra, is due to the sensitivity of these carbons to the type of linkage on the neighboring glucose units. Some evidence that conformational effects are involved in this multiplicity is provided by a comparison of the spectra of the 1→4-linked cyclodextrins and linear glucans, where appreciable chemical shift differences are apparent for C1 and C4. These effects are influenced by pH modification and are attributed to differences in rotational isomers at the linked carbons.
NMR, oligosaccharide, Bacterial polysaccharide, amylose
NCBI PubMed ID: 4464319Many widely used polymers undergo main chain scission or crosslinking when exposed to radiation. Effects of electron beam irradiation at different doses up to 500 kGy on the biodegradable pullulan polymer films have been investigated by ultraviolet-visible (UV-VIS), Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), and thermogravimetric analysis (TGA) techniques. UV-Visible study shows increase of optical absorbance with increase of doses, attributing to the formation some groups or radicals. FTIR and TGA results reveal the processes of both crosslinking and degradation of polymer taking place depending upon the dose of e-beam radiation. The surface morphology of the film is found to be altered by the e-beam radiation as indicated by SEM micrographs.
degradation, pullulan, FTIR, electron beam
Publication DOI: 10.1007/s10967-012-1691-4Response surface methodology was applied to understand the interactions amongst the components of a complex medium for pullulan production by a novel Aureobasidium pullulans strain using a second order quadratic model. A total of 20 experimental runs were carried out and three dimensional response surfaces were generated to study the interaction. Results indicated that the concentration of yeast extract plays a critical role and has significant influence in pullulan production compared with peptone. Understanding the effect of complex nitrogen sources in the media using statistical methods has resulted in 70.43. g/L pullulan production which is higher as compared to earlier published reports.
exopolysaccharide, response surface methodology, pullulan, Aureobasidium pullulans, central composite design, effect of nitrogen sources
Publication DOI: 10.1016/j.bcab.2012.02.003Ever increasing demand for fossil fuels is a major factor for rapid depletion of these non-renewable energy resources, which has enhanced the interest of finding out alternative sources of energy. In recent years jatropha seed oil has been used extensively for production of bio-diesel and has shown significant potential to replace petroleum fuels at least partially. De-oiled jatropha seed cake (DOJSC) which comprises of approximately 55 to 65% of the biomass is a byproduct of bio-diesel industry. DOJSC contains toxic components like phorbol esters which restricts its utilization as animal feed. Thus along with the enhancement of biodiesel production from jatropha, there is an associated problem of handling this toxic byproduct. Utilization of DOJSC as a feed stock for production of biochemicals may be an attractive solution to the problem. Pullulan is an industrially important polysaccharide with several potential applications in food, pharmaceuticals and cosmetic industries. However, the major bottleneck for commercial utilization of pullulan is its high cost. A cost effective process for pullulan production may be developed using DOJSC as sole nutrient source which will in turn also help in utilization of the byproduct of bio-diesel industry. Results: In the present study, DOJSC has been used as a nutrient for production of pullulan, in place of conventional nutrients like yeast extract and peptone. Process optimization was done in shake flasks, and under optimized conditions (8% DOJSC, 15% dextrose, 28°C temperature, 200 rpm, 5% inoculum, 6.0 pH) 83.98 g/L pullulan was obtained. The process was further validated in a 5 L laboratory scale fermenter.Conclusion: This is the first report of using DOJSC as nutrient for production of an exopolysaccharide. Successful use of DOJSC as nutrient will help in finding significant application of this toxic byproduct of biodiesel industry. This in turn also have a significant impact on cost reduction and may lead to development of a cost effective green technology for pullulan production.
exopolysaccharide, fermentation, pullulan, Aureobasidium pullulans, Jatropha, value addition to waste
NCBI PubMed ID: 22462652To create a drug delivery system that allows the controlled release of proteins, such as growth factors, over a long-term period, cholesteryl group- and acryloyl group-bearing pullulan (CHPOA) nanogels were aggregated to form fast-degradable hydrogels (CHPOA/hydrogels) by cross-linking with thiol-bearing polyethylene glycol. The gold standard of clinical bone reconstruction therapy with a physiologically active material is treatment with recombinant human bone morphogenetic protein 2 (BMP2); however, this approach has limitations, such as inflammation, poor cost-efficiency, and varying interindividual susceptibility. In this study, two distinct growth factors, BMP2 and recombinant human fibroblast growth factor 18 (FGF18), were applied to a critical-size skull bone defect for bone repair by the CHPOA/hydrogel system. The CHPOA-FGF18/hydrogel displayed identical results to the control CHPOA-PBS/hydrogel, and the CHPOA-BMP2/hydrogel treatment imperfectly induced bone repair. By contrast, the CHPOA-FGF18 + BMP2/hydrogel treatment strongly enhanced and stabilized the BMP2-dependent bone repair, inducing osteoprogenitor cell infiltration inside and around the hydrogel. This report indicates that the CHPOA/hydrogel system can successfully deliver two different proteins to the bone defect to induce effective bone repair. The combination of the CHPOA/hydrogel system with the growth factors FGF18 and BMP2 might be a step towards efficient bone tissue engineering.
drug delivery, acryloyl group-modified cholesterol-bearing pullulan (CHPOA), bone regeneration, osteoblasts, recombinant human bone morphogenetic protein 2 (BMP2), recombinant human fibroblast growth factor 18 (FGF18)
NCBI PubMed ID: 22800537This study evaluated the potential of utilizing transfected pTGFβ-1 gene-engineered rat mesenchymal stem cells (MSCs) using nonviral vector to promote cartilage regeneration. Pullulan-spermine was used as the nonviral gene vector and gelatin sponge was used as the scaffold. MSCs were engineered with TGF-β1 gene with either the three-dimensional (3D) reverse transfection system or the two-dimensional (2D) conventional transfection system. For the 3D reverse transfection system, pullulan-spermine/pTGF-β1 gene complexes were immobilized to the gelatin sponge, followed by the seeding of MSCs. Pullulan-spermine/pTGF-β1 gene complexes were delivered to MSCs cultured in the plate to perform the 2D conventional transfection system, and then MSCs were seeded to the gelatin sponge. Then, TGF-β1 gene-transfected MSC seeded gelatin sponge was implanted to the full-thickness cartilage defect. Compared with the control group, both groups of TGF-β1 gene-engineered MSCs improved cartilage regeneration through optical observation and histology staining. So, with pullulan-spermine as the nonviral vector, TGF-β1-gene engineered MSCs can induce cartilage regeneration in vivo.
pullulan, nonviral gene vector, cartilage defect, mesenchymal stem cells, reverse transfection, three dimensional, spermin
NCBI PubMed ID: 23586825The aim of this study was to determine the effects of three novel fibers on satiety and serum parameters. In a randomized, double-blind, crossover design, fasted subjects (n = 20) consumed a low-fiber control breakfast or one of four breakfasts containing 25 g of fiber from soluble corn fiber (SCF) or resistant starch (RS), alone or in combination with pullulan (SCF+P and RS+P). Visual analog scales assessed appetite, and blood samples were collected to measure glucose, insulin, ghrelin, and glucagon-like peptide-1 (GLP-1). The fiber treatments did not influence satiety or energy intake compared to control. RS+P significantly reduced glucose, insulin, and GLP-1, but neither SCF treatment differed from control. To conclude, these fibers have little impact on satiety when provided as a mixed meal matched for calories and macronutrients. Additional research regarding the physiological effects of these novel fibers is needed to guide their use as functional ingredients in food products.
glucose, pullulan, starch, fiber, gut hormones, satiety, visual analog scale, insulin
NCBI PubMed ID: 23136915Two hypovirulent and one virulent strains of Cryphonectria parasitica, the causal pathogen of chestnut blight, produced a large amount of pullulan and a minor polysaccharide fraction containing galactose, mannose and, in one case, also rhamnose. The minor fractions of all strains elicited higher phytotoxicity on chestnut (Castanea sativa L.) leaves, twigs and seedlings or on tomato cuttings (Lycopersicon esculentum L.) than that given both by pullulan and native extracellular polysaccharide fractions. One of the components of the minor fraction was identified to be a galactan whose structure, on the basis of chemical and spectroscopic methods consisted of the repeat unit: [→6)-β-D-Galf-(1→5)-β-D-Galf-(1→]n. This is the first report on the production of polysaccharides by C. parasitica and their phytotoxic activity
polysaccharide, phytotoxicity, pullulan, Cryphonectria parasitica, gelatinous zone
Publication DOI: 10.1016/S0144-8617(98)00050-2In this study, in order to directly and efficiently convert inulin into pullulan, the INU1 gene from Kluyveromyces maximum KM was integrated into the genomic DNA and actively expressed in the high pullulan producer Aureobasidium melanogenum P16 isolated from the mangrove ecosystem. After the ability to produce pullulan from inulin by different transformants was examined, it was found that the recombinant strain EI36, one of the transformants, produced 40.92 U/ml of inulinase activity while its wild-type strain P16 only yielded 7.57 U/ml of inulinase activity. Most (99.27 %) of the inulinase produced by the recombinant strain EI36 was secreted into the culture. During the 10-l fermentation, 70.57 ± 1.3 g/l of pullulan in the fermented medium was attained from inulin (138.0 g/l) within 108 h, high inulinase activity (42.03 U/ml) was produced within 60 h, the added inulin was actively hydrolyzed by the secreted inulinase, and most of the reducing sugars were used by the recombinant strain EI36. This confirmed that the genetically engineered yeast of A. melanogenum strain P16 was suitable for direct pullulan production from inulin
inulin, Metabolic engineering, pullulan production, inulinase gene, A. pullulans
NCBI PubMed ID: 25985744Statistical experimental designs were applied to optimize the fermentation medium for exopolysaccharide (EPS) production. Plackett-Burman design was applied to identify the significance of seven medium variables, in which sweet potato and yeast extract were found to be the significant variables for EPS production. Central composite design was applied to evaluate the optimum condition of the selected variables. Maximum EPS production of 9.3 g/L was obtained with the predicted optimal level of sweet potato 10 %, yeast extract 0.75 %, 5.5 pH, and time 100 h. The determined (R 2) value was 0.97, indicating a good fitted model for EPS production. Results of this study showed that sweet potato can be utilized as a low-cost effective substrate for pullulan production in submerged fermentation
exopolysaccharide, response surface methodology, central composite design, sweet potato
NCBI PubMed ID: 28324414With the purpose of understanding the metabolic network of Aureobasidium pullulans, the central metabolic pathways were confirmed by the activities of the key enzymes involved in different pathways. The effect of different iodoacetic acid concentrations on pullulan fermentation was also investigated in this paper. The activities of phosphofructokinases and glucose-6-phosphate dehydrogenase existed in A. pullulans CGMCC1234, whereas 2-keto-3-deoxy-6-phosphogluconate aldolase activity was not detected. We proposed that the central metabolic pathways of A. pullulans CGMCC1234 included EMP and PPP, but no ED. Pullulan production declined fast as the iodoacetic acid increased, while cell growth offered upgrade firstly than descending latter tendency. Compared to the control group, the ratio of ATP/ADP of 0.60 mM iodoacetic acid group was lower at different stages of pullulan fermentation. The findings revealed that low concentration of iodoacetic acid might impel carbon flux flow toward the PPP, but reduce the flux of the EMP
pullulan, Aureobasidium pullulans, central metabolic
NCBI PubMed ID: 26428132The emergence of modern microbial technology and product biotechnology had significantly made over the way, scientists and researchers view differently the microbes and the product they biosynthesize. Polymers create a significant role inside the natural environment and also play a vital action in the fashionable industrial economics. Some of the natural biopolymers like proteins and nucleic acids pass the biological information both in the intracellular and extracellular environment. Other polymers such as sugar based polysaccharides provide metabolic energy for the cellular activity and play important role with their potential applications in the different sectors. For instance, those can be used as lubricants, absorbents, adhesives, cosmetics, drug delivery, structural materials, lithography and for the manufacturing of computational switching devices). The main constraints and major obstacles for the full commercialization are due to their production cost, which is related to cost of the substrates and downstream processing. In this research study purification and in vitro analysis of pullulan will be discussed
polysaccharide, microorganism, pullulan
Journal NLM ID: 101602478The effect of Tween 80 on the fermentative production of polymalic acid (PMA) and pullulan using Aureobasidium pullulans CCTCC M2012223 was investigated. Tween 80 is beneficial for the biosynthesis of PMA and pullulan, and can regulate the ratio of PMA to pullulan in a dose-dependent manner. After adding 0.05 % Tween 80 to the media, the maximal PMA and pullulan production was 46.45 and 28.8 g/L at 60 h in a 5 L fermenter, with an increase of 75.08 and 27.21 % when compared to the control. Tween 80 could regulate and enhance oxygen uptake rate and carbon dioxide evolution rate in the early phase of fermentation, and change the cell morphology. The transcription levels of mitochondrial dicarboxylate transporter and transmembrane transporter were also dramatically upregulated. The present work will be helpful in deeply understanding the mechanism of Tween 80 on the effect of PMA and pullulan production
pullulan, Aureobasidium pullulans, gene transcription, polymalic acid, Tween 80
NCBI PubMed ID: 25413862Batch culture of Aureobasidium pullulans CCTCC M 2012259 for pullulan production at different concentrations of ammonium sulfate and yeast extract was investigated. Increased pullulan production was obtained under nitrogen-limiting conditions, as compared to that without nitrogen limitation. The mechanism of nitrogen limitation favoring to pullulan overproduction was revealed by determining the activity as well as gene expression of key enzymes, and energy supply for pullulan biosynthesis. Results indicated that nitrogen limitation increased the activities of α-phosphoglucose mutase and glucosyltransferase, up-regulated the transcriptional levels of pgm1 and fks genes, and supplied more ATP intracellularly, which were propitious to further pullulan biosynthesis. The economic analysis of batch pullulan production indicated that nitrogen limitation could reduce more than one third of the cost of raw materials when glucose was supplemented to a total concentration of 70 g/L. This study also helps to understand the mechanism of other polysaccharide overproduction by nitrogen limitation
pullulan, Aureobasidium pullulans, nitrogen limitation, enzyme activity, 46 gene expression, ATP supply
NCBI PubMed ID: 25965490Mushrooms were considered as a special delicacy by early civilizations and valued as a credible source of nutrients including considerable amounts of dietary fiber, minerals, and vitamins (in particularly, vitamin D). Mushrooms are also recognized as functional foods for their bioactive compounds offer huge beneficial impacts on human health. One of those potent bioactives is b-glucan, comprising a backbone of glucose residues linked by β-(1-3)-glycosidic bonds with attached β-(1-6) branch points, which exhibits antitumor and immunostimulating properties. The commercial pharmaceutical products from this polysaccharide source, such as schizophyllan, lentinan, grifolan, PSP (polysaccharide-peptide complex) and PSK (polysaccharide-protein complex), have shown evident clinical results. The immunomodulating action of mushroom polysaccharides is to stimulate natural killer cells, T-cells, B-cells, neutrophils, and macrophage dependent immune system responses via differing receptors involving Dectin-1, the toll-like receptor-2 (a class of proteins that play a role in the immune system), scavengers and lactosylceramides. b-Glucans with various structures present distinct affinities toward these receptors to trigger different host responses. Basically, their antitumor abilities are influenced by the molecular mass, branching configuration, conformation, and chemical modification of the polysaccharides. This review aims to integrate the information regarding nutritional, chemical and biological aspects of polysaccharides in mushrooms, which will possibly be employed to elucidate the correlation between their structural features and biological functions
polysaccharides, mushroom, immune system, TLR, Dectin-1
Publication DOI: 10.1039/c2fo10279jBACKGROUND: Pullulan and glycogen have many applications and physiological functions. However, to date, it has been unknown where and how the pullulan is synthesized in the yeast cells and if cell wall structure of the producer can affect pullulan and glycogen biosynthesis. METHODS: The genes related to cell wall integrity were cloned, characterized, deleted and complemented. The cell wall integrity, pullulan biosynthesis, glycogen accumulation and gene expression were examined. RESULTS: In this study, the GT6 and GT7 genes encoding different α(1,2) mannosyltransferases in Aureobasidium melanogenum P16 were cloned and characterized. The proteins deduced from both the GT6 and GT7 genes contained the conserved sequences YNMCHFWSNFEI and YSTCHFWSNFEI of a Ktr mannosyltransferase family. The removal of each gene and both the two genes caused the changes in colony and cell morphology and enhanced glycogen accumulation, leading to a reduced pullulan biosynthesis and the declined expression of many genes related to pullulan biosynthesis. The swollen cells of the disruptants were due to increased accumulation of glycogen, suggesting that uridine diphosphate glucose (UDP-glucose) was channeled to glycogen biosynthesis in the disruptants, rather than pullulan biosynthesis. Complementation of the GT6 and GT7 genes in the corresponding disruptants and growth of the disruptants in the presence of 0.6 M KCl made pullulan biosynthesis, glycogen accumulation, colony and cell morphology be restored. GENERAL SIGNIFICANCE: This is the first report that the two α1,2 mannosyltransferases were required for colony and cell morphology, glycogen accumulation and pullulan biosynthesis in the pullulan producing yeast.
α1, A. melanogenum, glycogen accumulation, pullulan biosynthesis, 2 mannosyltransferases
NCBI PubMed ID: 29550432The β-glucans are the glucose polymers present in the cells walls of yeast, fungi and cereals. β-Glucans are the major compositions of various nutritional diets such as oats, barley, seaweeds and mushrooms. Various biological activities of β-glucans have been reported such as anticancer, antidiabetic, anti-inflammatory and immune-modulating effects. The importance of β-glucans in food processing industries such as bread preparation, yogurt and pasta have been well elucidated. In recent findings on food science research gut microbiota plays a significant role and vastly studied for its intermediate role in regulating health. Several reports have suggested that β-glucans should have a significant impact on the gut microbiota changes and in turn on human health. The review was aimed to accumulate the evidence on types of β-glucans, their functional properties and the mechanism by how the β-glucans regulate the gut microbiota and human health. The various in vitro, in vivo and clinical studies, have been summarized, in particular, the changes happening upon the β-glucans supplementation on the gut microbiota. Overall, this review updates the recent studies on β-glucans and gut microbiota and also inputs the demanding questions to be addressed in β-glucans-microbiota research in the future.
β-Glucans, immunomodulation, microbiota, Antidiabetic, Anticancer, SCFA
NCBI PubMed ID: 30196242Extractive fermentation technique or in situ product recovery process is a novel technique to segregate the desired product simultaneously in a fermentation process. For economic and high yield production of pullulan, Extractive fermentation process was applied fermentation process of A. pullulans. Aqueous Two Phase system (ATPS) systems were designed with various molecular mass of PEG (400, 600, 4000 and 6000) and dextran or mono/bi-sodium phosphate salts. Systems with short Tie Line length (TLL) 6.7 and 7.5% w/w for PEG-Salt and PEG-dextran respectively were chosen. Volume ratio for all the systems was kept constant at 1.0 and pH 7.0 for PEG-dextran and PEG-NaH2PO4 was maintained, whereas pH 9.0 was kept for PEG-Na2HPO4. A. pullulans, was found to be viable with PEG-NaH2PO4 and PEG-dextran systems. The biomass partitioned in the PEG rich top phase and the exopolysaccharide pullulan shown affinity towards the bottom phase. A maximum yield (36.47 g/L) was found with PEG 4000-Dextran 500 system of extractive fermentation process. The proposed process aptly integrates upstream and downstream process for continuous production and recovery of pullulan from the biomass, thus reducing the time quotient of the whole process.
production, fermentation, pullulan, Aureobasidium pullulans
NCBI PubMed ID: 30631089In this study, a new yeast-like fungal was obtained from leaf surfaces collected from Kheyroodkenar forest, Mazandaran, Iran. The properties of this strain, such as morphology, DNA molecular, and product showed that it is related to Aureobasidium pullulans family and named A. pullulans MG271838. The pullulan production conditions by this strain were optimized using a Box-Behnken design. The results showed that the optimum production yield (37.55 ± 0.45 g/l) was obtained in pH of 6.76, sucrose concentration of 6 %w/v and yeast extract concentration of 0.2%w/v. The pullulan had a concentration-dependent flow behavior, amorphous structure based on XRD pattern and high thermal stability (decomposition temperature of 300 °C). Also, the chemical structure of pullulan was confirmed by FTIR spectroscopy. In addition, there was a direct relationship between pullulan yield and the gene expression of fks, pgm and ugp as the most important genes in pullulan production.
optimization, pullulan, Aureobasidium pullulans, thermal stability
NCBI PubMed ID: 31340178In order to develop pullulan/gelatin-based gels as potential extracellular matrix-mimetic scaffolds, a "one-step" synthesis method using 1,1'-carbonyldiimidazole (CDI) as activator in DMSO under mild conditions was reported for the first time. Particularly, in contrast to conventional critical requirement of absolute dryness for CDI, it was interesting to find that the formation of gels could be accomplished in the presence of aqueous solvent within a much shorter time, while obtaining improved mechanical properties as demonstrated by compressive tests. UV-Visible spectroscopy, NMR spectrum, TEM and SEM analysis have been employed to evaluate the underlying reaction mechanisms. This method implied that absolute dryness might not be necessary for using CDI as activator in certain cases. This method also represents a new rapid and efficient approach for synthesizing a great variety of chemically crosslinked polysaccharide/protein-based gels as promising material platform potential for tissue engineering and drug delivery applications.
1, pullulan, gels, 1′-carbonyldiimidazole, gelatin, nano-gels
NCBI PubMed ID: 30223056Pullulan is a biopolymer used in food industry produced by Aureobasidium pullulans from starch. In the present work, sugarcane bagasse (SCB) hydrolysate was evaluated as an alternative substrate in fermentation process assisted by blue LED lights. The best fermentation conditions in Erlenmeyer flasks were 25.3 °C, stirring speed of 232 rpm and yeast extract concentration of 1.88 g/L, yielding 25.19 g/L of pullulan, that corresponded to yield of 0.48 g/g and 0.28 g/(L·h) of volumetric productivity. By using a column bubble photobioreactor, similar yield values were obtained. Thermal properties of the produced pullulan as glass transition (Tg) and melting (Tm) temperatures were 38 °C and 160 °C, respectively, which were similar to the corresponding values of commercial food grade pullulan. Therefore, SCB hydrolysate is a promising substrate to produce good quality pullulan (86% of purity) with application in food industry, besides to represent a new alternative for biorefineries.
pullulan, biorefinery, sugarcane bagasse
NCBI PubMed ID: 30639656A novel yeast strain Aureobasidium melanogenum 13-2 isolated from the Taklimakan desert was found to be able to produce a high level of extracellular polysaccharide (EPS). Under the optimal conditions, the yeast strain could yield 73.25 ± 2.3 g/L of EPS within 5 days at a flask level. During a 10-liter fermentation, the yeast strain could produce 78.05 ± 3.5 g/L of EPS within 120 h. The FT-IR spectra of the standard pullulan from Sigma and the purified EPS produced by A. melanogenum 13-2 were almost identical and the purified EPS could be actively hydrolyzed by a pullulanase, demonstrating that the purified EPS was pullulan. The molecular weight (Mw) of the purified pullulan was estimated to be 770300 g/moL. Disruption of a pullulan synthase gene (PUL1) made a mutant DAG27 lose the ability to synthesize any pullulan. The mutant DAG27 was more sensitive to radiation of UV light, high NaCl concentration, heat treatment, strong oxidation of H2O2 and desiccation than its wild type strain 13-2, suggesting that the produced pullulan could play an important role in resistance of the yeast cells to various stresses. This was the first time to show that the yeast strain obtained from the desert could produce such high level pullulan and the produced pullulan had an obviously protective effect on its producer.
exopolysaccharide, Aureobasidium melanogenum, harsh environment, pullulan function, The Taklimakan desert
NCBI PubMed ID: 31145957The materials used in food packaging based on non-biodegradable synthetic polymers pose a serious threat of pollution to the environment. Hence, research is now focused on developing eco-friendly and biodegradable packaging obtained from natural polymers. Pullulan is a microbial exopolysaccharide, obtained on a commercial scale by the yeast-like fungus Aureobasidium pullulans. It is a water-soluble, non-toxic and non-mutagenic edible biopolymer with excellent film-forming abilities and adhesive properties. Furthermore, pullulan presents great potential to fabricate thin, transparent, odorless and tasteless edible films and coating used as packaging material. This review article presents an overview on the basic mechanical and barrier properties of a pullulan-based film. It also describes the modification methods applied in order to obtain multifunctional materials in terms of satisfactory physico-mechanical performance and antimicrobial activity for food packaging.
biopolymer, pullulan, antimicrobial properties, food packaging, mechanical and barrier properties
Publication DOI: 10.1515/ijfe-2019-0030Pullulan is a polysaccharide as obtained from the black yeast fermentation. Soft and flexible pullulan hydrogels and multi-walled carbon nanotube/pullulan hydrogel composites were fabricated by a solvent casting method. The effects of crosslinking agent, MWCNT content, and electric field strength on the electromechanical behaviours were investigated. The storage modulus (G′) or the elasticity monotonically increased with increasing crosslinking agent and electric field strength. The MWCNT/pullulan hydrogel composite with 0.01%v/v MWCNT provided the highest storage modulus sensitivity of 71.2 at the applied electric field strength of 800 V/mm, relative to other hydrogel composites previously reported. The storage and loss moduli were further transformed to the creep compliance through the relaxation spectrum and the retardation spectrum, respectively. The creep compliance of the pristine hydrogels and composites decreased with increasing crosslinking agent amount and electric field strength. The 01MWCNT/Pullulan_5STMP hydrogel composite yielded the lowest creep compliance function, illustrating the ability to resist deformation through electric field. The dielectrophoresis force density of the MWCNT/pullulan hydrogel composite with 0.01%v/v MWCNT was the highest obtained at 0.2258 mN/mm3, suggesting as the most suitable for practical actuator applications.
pullulan, hydrogel, multi-walled carbon nanotube, electromechanical properties
Publication DOI: 10.1016/j.eurpolymj.2019.109231In this study, the production rate of both water-insoluble EPS, bacterial cellulose, and water-soluble EPS, P, was improved through сultivation of their producers on a nutrient media containing industrial wastes, and their material properties were analyzed. The growth rate and productivity of Gluconoacetobacter xylinus C3 strain on media with industrial wastes was investigated. An optimal nutrient medium based on molasses was selected for the bacterial cellulose producer. The nutrient medium contains 2% molasses, 1% yeast extract and peptone in a 1: 1 ratio, 0.3% sodium hydrogen phosphate, 0.1% citric acid and 1% ethanol. Cultivation of Gluconoacetobacter xylinus C3 strain on this medium for 7 days at 25–30 °С ensures its high productivity – 8.21 g/L. The composition of the optimized medium with molasses provides high mechanical properties (tensile strength – 37.12 MPa and relative elongation at break – 3.28%) of bacterial cellulose and does not affect the polymer microfibrillar structure. A modified Czapek-Dox medium with 10% molasses and 1% peptone is preferable for the exopolysaccharide accumulation by A. pullulans C8 strain. The optimized media has an advantage over the traditionally used media in terms of the efficiency of exopolysaccharide accumulation and cost reduction. The pullulan yield in media was 10.08 g/l, that is 1.5 times higher than in a standard Czapek-Dox medium. The surface morphology and microstructure of the pullulan samples obtained on different media showed minor changes. Therefore, the replacement of carbon source for molasses in a Czapek-Dox media for pullulan production did not alter the polymer content and viscosity.
exopolysaccharides, pullulan, bacterial cellulose, industrial wastes
Publication DOI: 10.18321/ectj866In the present investigation, polyethylene glycol (PEG 6000) was used for downstream processing of pullulan from Aureobasidium pullulans by aqueous phase separation (APS) technique. The cell-free broth was processed with PEG solution (10-35%, w/w) and pullulan formed a clear separate phase with all concentrations of PEG i.e. pullulan in lower phase and PEG solution along with impurities in upper phase. Maximum pullulan recovery from cell-free broth was obtained by PEG 25% (w/w). The sample handling in APS technique is quite easier due to lesser volume of PEG used in comparison to organic solvent precipitation i.e. minimum required ratio of cell-free broth to PEG for maximum pullulan yield was 0.5:1 (PEG:cell-free broth). Additionally, APS technique was found to be temperature independent. Further, structural attributes of pullulan recovered by APS was confirmed by FTIR, NMR and TLC. This is the first report on downstream processing of pullulan using PEG based aqueous solution.
pullulan, Aureobasidium pullulans, aqueous phase separation, polyethylene glycol
NCBI PubMed ID: 31427022A novel, yeast-like fungal strain, Aureobasidium melanogenum TN3-1, that was isolated from natural honey can actively transform 140.0 g/L of glucose into 110.29 ± 2.17 g/L of pullulan during fermentation, whereas A. melanogenum P16 and TN1-2 converted 140.0 g/L of glucose into only 45.81 ± 1.7 g/L and 48.7 ± 2.6 g/L of pullulan, respectively. It was noted that most of the cells in the culture of the strain TN3-1 were arthroconidia, while all of the yeast-like fungal cells of A. melanogenum P16 cultivated under the same conditions were blastoconidia. The cell sizes, cell walls and the number of small vacuoles of A. melanogenum TN3-1 were also much larger, thicker and higher, respectively, than those of A. melanogenum P16. The glycerol, trehalose and glycogen content in the A. melanogenum TN3-1 cells was higher than that of the A. melanogenum P16 and TN1-2 cells.
pullulan, arthroconidia, Aureobasidium spp., natural honey, osmophiles, small vacuoles
NCBI PubMed ID: 30827584Overexpression of the optimized Vitreoscilla hemoglobin (VHb) gene and the native flavohemoglobin (FHb) gene in Aureobasidium melanogenum P16 rendered a V6 strain and a F44 strain to overproduce pullulan compared to that produced by their wild type strain P16. The capacity to bind CO and oxygen in the V6 strain and the F44 strain was also obviously enhanced. At the same time, the transcriptional levels of the relevant genes were also increased in the V6 strain and the F44 strain and the fused vgbop + the gene encoding GFP and FHb gene + the gene encoding GFP were also actively expressed. During a 10-liter fermentation, the P16 strain produced only 72.0 ± 1.0 g/L pullulan, the yield was 0.77 g/g of sucrose, the productivity was 0.5 ± 0.01 g/L/h and only 79.4% of the total sugar was used. In contrast, the strain V6 yielded 102.3 ± 1.8 g/L of pullulan, the yield was 0.89 g/g of sucrose, the productivity was 0.7 ± 0.01 g/L/h and 96.0% of total sugar was used while 101.4 ± 2.9 g/L of pullulan was accumulated in the culture of the strain F44, the yield was 0.88 g/g of sucrose, the productivity was 0.7 ± 0.02 g/L/h and 96.4% of total sugar was utilized. These data strongly demonstrated that the concentration of pullulan, yield, productivity and sugar utilization were greatly enhanced by overexpression of the VHb and FHb. But their cell growth was almost the similar.
pullulan, Aureobasidium melanogenum, flavohemoglobin, Vitreoscilla hemoglobin
NCBI PubMed ID: 30953719Polysaccharide pullulan acts as a base matrix for the fabrication of pullulan/collagen blend film with dopant ZnO nanoparticles. From the FTIR data, it could be confirmed that there was significant interaction among the pullulan/collagen/ZnO-NPs. The weak interactions like H-bonding was evident by shift absorption band of hydroxyl groups into the lower frequency region. The powder X-ray diffraction study revealed the amorphous nature of pullulan/collagen blended film which was modified into a crystalline structure after the addition of ZnO nanoparticles. The transformation of the films into crystallinity and the presence of the Zn element was confirmed by SEM-EDAX. Further, the results of TGA revealed that the addition of nanoparticles influenced the thermal stability of the pullulan/collagen blended films. The addition of ZnO nanoparticles lead to an increase of the stiffness and mechanical resistance of the nanocomposite films; which showed an increase in their tensile strength evidenced by an increase in Young’s modulus. The fabricated films exhibited antifungal activity against Aspergillus niger with a maximum zone of inhibition of 18 mm for pullulan/collagen/ZnO-NPs (0.5%). However, the nanocomposite films easily dissolved in an aqueous medium, indicating their possible use as edible packaging films in the food industry.
polysaccharide, pullulan, acid soluble collagen, ZnO nanoparticles
Publication DOI: 10.1007/s10924-020-01953-yIn our previous study, it was found that Aureobasidium melanogenum TN3-1 was a high pullulan producing and osmotic tolerant yeast-like fungal strain. In this study, the HOG1 signaling pathway controlling glycerol synthesis, glycerol, trehalose and vacuoles were found to be closely related to its pullulan biosynthesis and high osmotic tolerance. Therefore, deletion of the key genes for the HOG1 signaling pathway, glycerol and trehalose biosynthesis and vacuole formation made all the mutants reduce pullulan biosynthesis and increase sensitivity of the growth of the mutants to high glucose concentration. Especially, abolishment of both the VSP11 and VSP12 genes which controlled the fission/fusion balance of vacuoles could cause big reduction in pullulan production (less than 7.4 ± 0.4 g/L) by the double mutant ΔDV-5 and increased sensitivity to high concentration glucose, while expression of the VSP11 gene in the double mutant ΔDV-5 made the transformants EV-2 restore pullulan production and tolerance to high concentration glucose. But cell growth of them were the similar. The double mutant ΔDV-5 had much bigger vacuoles and less numbers of vacuoles than the transformant EV-2 and its wild type strain TN3-1 while it grew weakly on the plate with 40% (w/v) glucose while the transformant EV-2 and its wild type strain TN3-1 could grow normally on the plate even with 60% (w/v) glucose. The double mutant ΔDV-5 also had high level of pigment and its cells were swollen. This was the first time to give the evidence that glycerol, trehalose and vacuoles were closely related to pullulan biosynthesis and high osmotic tolerance by A. melanogenum.
pullulan production, A. melanogenum, osmotic pressure, the whole genome duplication, vacuoles
NCBI PubMed ID: 32987074Although pullulan has many uses in industry, the detailed mechanisms of its biosynthesis still require clarification. In this study, it was found that a short α-1,4-glucosyl chain (pullulan primer) synthesized by the glycogenins Glg1 and Glg2 for initiation of glycogen biosynthesis was also needed for pullulan synthesis. The primers were also synthesized on sterol glycosides and glucosylceramides by catalysis of sterol glucosyltransferase (Sgt1) and ceramide β-glucosyltransferase (Gcs1). All the primers might be elongated to be long α-1,4-glucosyl chain (pullulan precursor) by catalysis of the glycogen synthetase domain of the AmAgs2 as previously reported. Then, the amylase domain of the same AmAgs2 was responsible for pullulan biosynthesis. Removal of all the genes encoding Glg1, Glg2, Gcs1 and Sgt1 made all the mutants produce much less pullulan than the strain P16. However, pullulan synthesis could not be stopped totally in these mutants, suggesting that any other unknown alternative pullulan primers may exist in the yeast cells. Complementation of all the genes in the mutants restored pullulan biosynthesis. This is the first time to report that like starch and glycogen biosynthesis, alternative primers are also required for pullulan biosynthesis in Aureobasidium melanogenum P16.
A. melanogenum, pullulan biosynthesis, pullulan primers, α-glucan synthetase
NCBI PubMed ID: 31923483Pullulan, a bio-polysaccharide, is being currently known as an edible coating shelf-life extension of fruits and vegetables. In this examination, edible coating method was optimized using the RSM technique and pullulan-based active edible coating formulations with Calcium chloride (1% w/v) and lemon juice (2% v/v) were coated on the Rastali and Chakkarakeli bananas and put away at 25 ± 1°C, 70% RH, for 20 days. Results have demonstrated that optimized coating emulsion at 60°C, dipping time for 10 min, and 10% w/v pullulan concentration was best for minimum (5.466%) weight loss. Further, physico-chemical properties for all coating solutions were compared with control (p < .05). This new edible coating formulation had shown low (64.92) color saturation, less (212.17) browning Index, 15% decreased peel‐pulp ratio, 19% reduced vitamin C content, 55% increased fruit firmness, and 12%–13% high total and residual sugar contents.
pullulan, bananas, food preservation, edible coating
Publication DOI: 10.1111/jfpp.14378In this study, a new strain suspected to be Aureobasidium pullulans was isolated from trees leaves. The molecular characterization and the resulting phylogenetic tree showed that the isolated strain was A. pullulans. Also, the results of methylation analysis, monosaccharide composition, FTIR, NMR and XRD confirmed that the obtained exo-polysaccharide from the mentioned strain was pullulan. The pullulan production optimization by central composite design (CCD) indicated that the maximum yield obtained under optimum conditions (pH of 6.5, sucrose concentration of 5.5% (w/v) and yeast extract concentration of 0.1% (w/v)) was 51.4 ± 0.50 g/L. The produced pullulan had an average molecular weight (Mw) of 207000 g/mol based on gel permeation chromatography results. The decomposition temperature (Td) of the produced pullulan was ~300 °C and also, the resulting pullulan had a Newtonian flow behavior in a wide range of concentrations.
methylation analysis, pullulan, molecular characterization
NCBI PubMed ID: 32088229Pullulan is a microbial exopolysaccharide with wide uses in various industries. The aim of this study was to investigate pullulan production from agro-industrial wastes and study of pH, molasses concentration and corn steep liquor concentration as independent variables and yield of pullulan as response. Briefly, 5% v/v of the inoculation media (yeast extract 3 g, malt extract 3 g, peptone 5 g and sucrose 10 g per liter of distilled water), including Aureobasidium pullulans were added into media, containing 100 ml of molasses (100, 150 and 200 g/l) and various corn steep liquor concentrations (20, 40 and 60 ml/l) at adjusted pH (4.5, 5.5 and 6.5). After extraction and separation of the biomass using centrifuge, two folds of the supernatant volume of cold ethanol were added to the samples and stored at 4 °C for 24 h. After centrifuging, pullulan was dried and analyzed using Fourier transform infrared spectroscopy, X-ray diffraction, thermogravimetric analysis and rheological tests. Findings revealed that the maximum production yield (18.29 g/l ±0.10) was achieved under optimum fermentation conditions (pH of 5.3, molasses concentration of 165 g/l and corn steep liquor concentration of 43 ml/l). Then, physiochemical and thermal properties of the pullulan under the highlighted conditions were investigated. Pullulan included 78.8% solubility with no hygroscopicity. Furthermore, structural analysis using Fou-rier transform infrared and X-ray diffraction verified presence of pullulan with an amorphous structure in the supernatant. The exopolysaccharide included acceptable thermal stability and gel-like behavior; in which, the elastic component was predominant based on the results of thermogravimetric analysis and rheological properties, respectively.
pullulan, molasses, corn steep liquor, rheological attributes, thermal analysis
Publication DOI: 10.22037/afb.v7i4.29747Yoghurt acts as a perfect delivery system for probiotic bacteria and some functional food ingredients, thereby enhancing their viability. Pullulan (PUL), a natural exopolysaccharide produced by yeast Aureobasidium pullulans is a low-calorie food ingredient and dietary fibre with potential prebiotic properties. The main aim of the present study was to investigate the effect of 0.5%–2.0% PUL addition on pH, sensory quality and viability of the yoghurt starter and probiotic bacteria in low-fat yoghurt during 28 days of storage. PUL addition enhanced the viability of Bifidobacterium and Lactobacillus, while it did not affect S. thermophilus count. The protective effect of PUL increased with concentration and was the highest for 1.5 and 2.0% PUL supplementation. Incorporation of PUL at 1.5% and 2.0% prolonged survival of B. animalis subsp. Lactis at a concentration above the therapeutic level (<10 6 cfu/mL) for the entire storage period. All the products showed a decrease in pH during the storage period. Further, the sensory analysis revealed that PUL incorporation improved thickness, smoothness, and viscosity of low-fat yoghurts, but decreased the intensity of milk taste and aroma, contributing to a decrease in overall acceptability.
yoghurt, probiotic, pullulan, prebiotic, storage
Publication DOI: 10.1016/j.lwt.2020.109414The fungal α-glucan synthases (Agss) are multi-domain proteins catalyzing biosynthesis of cell wall α-1,3-glucan which determines cell wall integrity or fungal pathogenicity and pullulan which is a maltotriosyl polymer made of α-1,4 and α-1,6 bound glucose units. The Agss family can be divided into 11 groups, some of which lost the original functions due to accumulation of harmful mutations or gene loss. Schizosaccharomyces pombe kept five kinds of Agss in the genome while Aspergillus spp. and Penicillium spp. lost one or two or three kinds of Agss. All the human, animal and plant pathogens kept only one single kind of Ags or only one active Ags for synthesis of cell wall α-1,3-glucan, a virulence factor. While the genus Aureobasidium spp. contained three kinds of Agss, of which only some of the Ags2 was involved in pullulan biosynthesis. Although many Agss contained Big_5 domain, only the Big_5 domain with conserved amino acids LQS from some strains of A. melanogenum could catalyze pullulan biosynthesis. This whole amino acid sequence and phylogenetic differences may cause non-α-1,3-glucan synthesizing activity of some fungal Agss.
α-1, 3-glucan, pullulan, fungal cell wall, Aureobasidium spp., α-glucan synthase
NCBI PubMed ID: 32569690Pullulan, a water-soluble polysaccharide consisting of maltotriose units used in the preparation of edible films and drug delivery, is generally produced from starch by Aureobasidium pullulans (de Bary & Löwenthal) G.Arnaud fungus. In this article, the characterisation of an alternative pullulan source − the stromata of Cyttaria hariotii E.Fisch. fungus − by elemental analysis, infrared spectroscopy and thermal analysis techniques is reported. With a view to a possible valorisation of this pullulan and its derivatives as bioactive formulations in agriculture, low-molecular-weight pullulan (<7 kDa) complexes with Mn(II), suitable for the remediation of Mn-deficiencies in winter cereal by foliar application, were synthesised and characterised by mass spectrometry.
mass spectrometry, FTIR spectroscopy, thermal analysis, E1204, llao-llao
Publication DOI: 10.1080/14786419.2020.1831493Pullulan has many useful characteristics but, its high cost limits its potential applications. In the present work, kitchen waste (KW), which otherwise has zero commercial value, was evaluated for the economical production of pullulan. Before fermentation, the KW was hydrolyzed into free sugars using an in-house produced cocktail of enzymes. During hydrolysis, 46 ± 3.5 g/l and 31 ± 2.2 g/l of total reducing sugars and glucose were released, respectively. Hydrolyzed kitchen waste was then used as substrate for fermentation by Aureobasidium pullulans MTCC 2013 yielding 20.46 ± 2.01 g/l pullulan. Further, effect of different nitrogen sources was evaluated and yeast extract (3%) was found to be the best, yielding (24.77 ± 1.06 g/l) exopolysaccharide (EPS). The pullulan produced from KW was characterized in terms of organoleptic properties, physical strength, Fourier-transform infrared spectroscopy (FTIR), and H nuclear magnetic resonance (H NMR) analysis. The results corroborated well with commercial pullulan. The biodegradable nature and water solubility of the film developed from pullulan was also confirmed. To the best of our knowledge, this is the first report on the validation of the biodegradability of in-house produced pullulan. Thus, kitchen waste appears to be a promising option for economical pullulan production. Additionally, the method may also prove to be helpful for managing the increasing load of municipal solid waste in an eco-friendly and scientific way.
enzymatic hydrolysis, pullulan, exopolysaccharide (EPS), Aureobasidium pullulans, kitchen waste
NCBI PubMed ID: 31838544Glucans are biopolymers made up of repeating units of D-glucose and are of biological origin with varying molecular weights. The differences in their linkage, chemical structure, branching and substitutions afford them unique properties. This review deals with the production and applications of α and β glucans from different sources such as cereal, yeast, fungi, plants, bacteria, and current global manufactures and their market value. The market value of glucans is more than 200 M US$ and expected to grow rapidly in medical, food and cosmetic industries. Applications of some of the industrially important α glucans, namely, dextran, amylopectin, glycogen, pullulan, amylose and β glucans; namely cereal glucan, yeast glucan, curdlan, laminarin, microbial cellulose, lentinan are described. Chemical modification of glucans can help in improving their properties for wider applications. A thorough understanding of these biopolymers could expand their applications in newer fields and may help in replacing some of the synthetic polymers which have several associated waste disposal and toxicity problems.
fermentation, Agrobacterium, Curdlan, Dextran, cyclodextrin
Publication DOI: 10.1080/00194506.2020.1798820The effects of several surfactants on the biosynthesis of β-1,3-D-glucan (β-glucan) and pullulan by Aureobasidium pullulans CCTCC M 2012259 were investigated, and Triton X-100 was found to decrease biomass formation but increase β-glucan and pullulan production. The addition of 5 g/L Triton X-100 to the fermentation medium and bioconversion broth significantly increased β-glucan production by 76.6% and 69.9%, respectively, when compared to the control without surfactant addition. To reveal the physiological mechanism underlying the effect of Triton X-100 on polysaccharides production, the cell morphology and viability, membrane permeability, key enzyme activities, and intracellular levels of UDPG, NADH, and ATP were determined. The results indicated that Triton X-100 increased the activities of key enzymes involved in β-glucan and pullulan biosynthesis, improved intracellular UDPG and energy supply, and accelerated the transportation rate of precursors across the cell membrane, all of which contributed to the enhanced production of β-glucan and pullulan. Moreover, a two-stage culture strategy with combined processes of batch fermentation and bioconversion was applied, and co-production of β-glucan and pullulan in the presence of 5 g/L Triton X-100 additions was further improved. The present study not only provides insights into the effect of surfactant on β-glucan and pullulan production but also presents a feasible approach for efficient production of analogue exopolysaccharides. Triton X-100 increased β-glucan and pullulan production under either batch fermentation or bioconversion. Triton X-100 increased the permeability of cell membrane and accelerated the transportation rate of precursors across cell membrane. Activities of key enzymes involved in β-glucan and pullulan biosynthesis were increased in the presence of Triton X-100. Intracellular UDPG levels and energy supply were improved by Triton X-100 addition.
β-1, 3-glucan, pullulan, Aureobasidium pullulans, triton X-100, UDPG
NCBI PubMed ID: 33170326The effects of mineral salts on the production of exopolysaccharides, including β-glucan and pullulan, by Aureobasidium pullulans CCTCC M 2012259 were investigated. Zinc sulfate at certain concentrations decreased dry biomass but favored to the biosynthesis of both exopolysaccharides. When 100 mg/L zinc sulfate was added to the fermentation medium, production of β-glucan and pullulan increased by 141.7 and 10.2%, respectively, when compared with that noted in the control without zinc sulfate addition. To reveal the physiological mechanism underlying improved β-glucan and pullulan production, key enzymes activities, energy metabolism substances, intracellular uridine diphosphate glucose (UDPG) levels, and gene expression were determined. The results indicated that zinc sulfate up-regulated the transcriptional levels of pgm1, ugp, fks, and kre6 genes, increased activities of key enzymes involved in the biosynthesis of UDPG, β-glucan and pullulan, enhanced intracellular UDPG content, and improved energy supply, all of which contributed to the increment in β-glucan and pullulan production. The present study not only provides a feasible approach to improve the production of exopolysaccharides but also contributes to better understanding of the physiological characteristics of A. pullulans.
gene expression, β-glucan, pullulan, Aureobasidium pullulans, UDPG, zinc sulfate
NCBI PubMed ID: 31867695Pullulan is an important polysaccharide. Although its synthetic pathway in Aureobasidium melanogenum has been elucidated, the mechanism underlying its biosynthesis as regulated by signaling pathway and transcriptional regulator is still unknown. In this study, it was found that the expression of the UGP1 gene encoding UDPG-pyrophosphorylase (Ugp1) and other genes which were involved in pullulan biosynthesis was controlled by the transcriptional activator Msn2 in the nuclei of yeast-like fungal cells. The Ugp1 was a rate-limiting enzyme for pullulan biosynthesis. In addition, the activity and subcellular localization of the Msn2 were regulated only by the cAMP-PKA signaling pathway. When the cAMP-PKA activity was low, the Msn2 was localized in the nuclei, the UGP1 gene was highly expressed, and pullulan was actively synthesized. By contrast, when the cAMP-PKA activity was high, the Msn2 was localized in the cytoplasm and the UGP1 gene expression was disabled so that pullulan was stopped, but lipid biosynthesis was actively enhanced. This study was the first to report that pullulan and lipid biosynthesis in yeast-like fungal cells were regulated by the Msn2 and cAMP-PKA signaling pathway. Elucidating the regulation mechanisms was important to understand their functions and enhance pullulan and lipid biosynthesis.
pullulan biosynthesis, Msn2, UDPG-pyrophosphorylase, yeast-like fungi, cAMP-PKA
NCBI PubMed ID: 32339573Mounting evidence indicated the capability of various microorganisms in biosynthesis of exopolysaccharides (EPSs). A wide range of evidence extensively investigated the ability of bacterial species for EPS synthesis and their favorable effects, so little is known regarding yeast species. Many factors like composition of growth media and fermentation conditions are related to the structural and physical properties of EPSs. The EPS protects the producer yeast strain against extreme environment. Researchers proposed that yeast EPSs have priority over bacterial EPSs because of high yields of EPS biosynthesis and easy separation methods from growth media. Besides, they have drawn increasing attention due to their interesting biological activities, food, pharmaceutical, and cosmetics applications. Although a limited number of studies exist, this review aims to highlight the EPS structure and various applications of known yeast species in detail.
yeast, exopolysccharide, health benefits, physiological activities
NCBI PubMed ID: 33604744Pullulan is an imperative microbial exo-polymer commercially produced by yeast like fungus Aureobasidium pullulans. Its structure contains maltosyl repeating units which comprises two α-(1→4) linked glucopyranose rings attached to one glucopyranose ring through α-(1→6) glycosidic bond. The co-existence of α-(1→6) and α-(1→4) glycosidic linkages endows distinctive physico-chemical properties to pullulan. It is highly biocompatible, non-toxic and non-carcinogenic in nature. It is extremely resistant to any mutagenicity or immunogenicity. The unique properties of pullulan make it a potent candidate for biomedical applications viz. drug delivery, gene delivery, tissue engineering, molecular chaperon, plasma expander, vaccination, etc. This review highlights the potential of pullulan in biomedical research and development.
pullulan, biomedical industry, pharmaceutical industry
NCBI PubMed ID: 33137388Drug distribution relies heavily on polymers, which also offer a variety of benefits like controlled release, targeted release, prolonged release, etc. Due to their low toxicity and great safety, biodegradable polymers are highly preferred. The exopolysaccharide known as pullulan is generated from a fungus known as Aureobasidium pullulan. It has many different qualities, including biodegradability, appropriate adhesion, antioxidant, film-forming capacity, blood compatibility, mucosal adhesion, etc. However, its application in the pharmaceutical industry is restricted by its insolubility in organic solvents, mechanical characteristics, and lack of macromolecule-carrying ability groups. This review provides an overview of the modifications made to pullulan, including periodate oxidation, etherification, esterification, sulfation, urethane derivatization, PEG incorporation, and cationization, to enhance its solubility in organic solvents, mechanical properties, pH sensitivity, drug delivery, anticoagulant, and antimicrobial properties. Pullulan has nine active hydroxyl groups in its structure that react chemically that can be used for physicochemical modification to produce pullulan derivatives. A key area of pullulan research has been pullulan modification, which has demonstrated enhanced solubility, pH-sensitive targeting, broadened horizons for delivery systems, anticoagulation, and antibacterial properties.
properties of pullulan derivatives, pullulan complex in drug delivery, pullulan derivatives in drug development, pullulan in drug delivery optimization
NCBI PubMed ID: 36369833Pullulan is the one of the most potent biocompatible polymer, which is basically synthesized by the Aureobasidium pullulans. This polymer appears to be a linear α-glucan of maltotriose units with occasional branching of glucosyl or maltosyl substitution. The employment and application of pullulan in biomedical and tissue engineering field is emerging owing to its biocompatible, nontoxic, non-immunogenic, and inert nature. It can be derivatized via various chemical reactions to increase its utility in the field of pharmaceuticals. In addition, pullulan and its derivatives have photographic, lithographic, and electronic applications in the biomedical instrumentation. This chapter provides comprehensive information about 'pullulan' considering its microbial sources, biosynthesis aspects, characterization, and functionalization. It also highlights the various applications of pullulan and its derivatives in the pharmaceutical and biomedical fields.
polysaccharide, application, pullulan, Aureobasidium pullulans, surface modification
Publication DOI: 10.1007/978-3-030-42215-8_15Microbial polysaccharides (MPs) are biopolymers secreted by microorganisms such as bacteria and fungi during their metabolic processes. Compared to polysaccharides derived from plants and animals, MPs have advantages such as wide sources, high production efficiency, and less susceptibility to natural environmental influences. The most attractive feature of MPs lies in their diverse biological activities, such as antioxidative, anti-tumor, antibacterial, and immunomodulatory activities, which have demonstrated immense potential for applications in functional foods, cosmetics, and biomedicine. These bioactivities are precisely regulated by their sophisticated molecular structure. However, the mechanisms underlying this precise regulation are not yet fully understood and continue to evolve. This article presents a comprehensive review of the most representative species of MPs, including their fermentation and purification processes and their biomedical applications in recent years. In particular, this work presents an in-depth analysis into the structure-activity relationships of MPs across multiple molecular levels. Additionally, this review discusses the challenges and prospects of investigating the structure-activity relationships, providing valuable insights into the broad and high-value utilization of MPs
structure-activity relationship, microbial polysaccharides, bioactivity, biomedicine
NCBI PubMed ID: 38616098The branched dextrins produced from waxy rice starch β-limit dextrin by Bacillus subtilis' saccharifying α-amylse [EC 3. 2. 1. 1] were isolated by the multiple development paper chromatography and their chemical structures were investigated. The analyses using β-amylase [EC 3. 2. 1. 2], glucoamylase [EC 3. 2. 1. 3], pullulanase, and pullulan α-1, 4-glucoside hydrolase revealed that they were doubly branched dextrins with the following structures: 63-α-(62-α-glucosylmaltosyl)-maltotriose, 63-α-, 65-α-diglucosylmaltopentaose, and 63-α-(63-α-glucosylmaltotriosyl)-maltotriose. The results were discussed in connection with the interior structure of β-limit dextrin.
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