Search

Melanin is a high value bioproduct generated through the fermentation of Aureobasidium pullulans, playing a crucial role in various fields, including food, medicine, environmental protection, and materials science. However, its high production costs and low synthetic yields significantly limit its applications. Therefore, it is essential to identify high-yield strains, reduce production costs, and optimize fermentation strategies. In this study, a high melanin-yielding Aureobasidium pullulans 53LC7 was screened and identified, and the fermentation process was optimized based on melanin yield, color value, and pullulan yield. The results indicated that the melanin yield peaked at an initial pH of 6.0, temperature of 27 °C, fermentation time of 6.5 d, and inoculation quantity of 2.5%, achieving a melanin yield of 16.33 g/L. Subsequently, huangjiu lees, a byproduct of huangjiu production, was incorporated into the fermentation medium, resulting in a melanin yield of 5.91 g/L. This suggests that the Aureobasidium pullulans was not effectively utilizing huangjiu lees. To address this, we employed an adaptive evolution strategy, which increased the melanin yield to 8.72 g/L. The enhanced production was correlated with the expression of key genes, including FKS, PKS, and Cmr1. Finally, cellulase was utilized to convert the crude fibers in huangjiu lees, which were difficult to utilize, into usable substrates, while pullulanase was employed to minimize byproduct formation in the fermentation system, resulting in a melanin yield of 19.07 g/L. This study not only provides promising strains for further research but also offers valuable insights for resource production technologies. [ABSTRACT FROM AUTHOR]

This review explores the production of the fungal polysaccharide pullulan by mutants and natural isolates of Aureobasidium species using strain improvement. Pullulan is a neutral polysaccharide gum whose structure is a maltotriose-containing glucan. This polysaccharide gum has applications in the fields of food, pharmaceuticals, biomedical and wastewater treatment. The strain improvement of Aureobasidium species has focused on the pullulan production process, including the isolation of strains exhibiting reduced pigmentation, polysaccharide overproduction, the production of pullulan with variable molecular weight, and increased osmotolerant strains promoting pullulan production at high carbon source concentrations and pullulan production on hemicellulosic substrates. The majority of studies have emphasized the isolation of reduced pigmentation and pullulan hyperproducer strains since the goal of large-scale commercial pullulan production is to synthesize non-pigmented polysaccharides. A promising area of strain improvement is the isolation of strains that synthesize authentic pullulan from hemicellulosic substrates. If strain improvement in this area is successful, the goal of commercially producing pullulan at a competitive cost will eventually be achieved. [ABSTRACT FROM AUTHOR]

Pullulan is a microbial exopolysaccharide produced by Aureobasidium spp. with excellent physical and chemical properties, resulting in great application value. In this study, a novel strain RM1603 of Aureobasidium pullulans with high pullulan production of 51.0 ± 1.0 g·L− 1 isolated from rhizosphere soil was subjected to atmospheric and room temperature plasma (ARTP) mutagenesis, followed by selection of mutants to obtain pullulan high-producing strains. Finally, two mutants Mu0816 and Mu1519 were obtained, with polysaccharide productions of 58.7 ± 0.8 and 60.0 ± 0.8 g∙L− 1 after 72-h fermentation, representing 15.1 and 17.6% increases compared with the original strain, respectively. Transcriptome analysis of the two mutants and the original strain revealed that the high expression of α/β-hydrolase (ABHD), α-amylase (AMY1), and sugar porter family MFS transporters (SPF-MFS) in the mutants may be related to the synthesis and secretion of pullulan. These results demonstrated the effectiveness of ARTP mutagenesis in A. pullulans, providing a basis for the investigation of genes related to pullulan synthesis and secretion. [ABSTRACT FROM AUTHOR]

Pullulan is a biopolymer produced by various strains of the fungi Aureobasidium pullulans. It is made up of maltotriose units consisting of α-(1,6) and α-(1,4) glycosidic bonds. Pullulan was discovered by Bauer in 1938, characterized by Bernier in 1958 and named as 'pullulan' by Bender in 1959. Submerged fermentation is widely employed for pullulan production followed by downstream processing for recovery of pullulan. Factors such as nutrients, minerals, pH, temperature, surfactants, light intensity, and melanin intermediate affect its production. Various optimization methods such as Plackett Burman design, response surface methodology, and artificial neural networks have been utilised for enhancing the yield of pullulan. Pullulan yield as high as 62.52 g/L can be achieved using a 5 L fermenter under optimised conditions. It can be characterized by techniques such as nuclear magnetic resonance, Fourier transform infrared spectroscopy, X-ray diffraction, scanning electron microscopy. The biopolymer has remarkable properties which makes it an ideal candidate to be used in various applications such as the biomedical and pharmaceutical sectors, cosmetics, food, and wastewater treatment. Pullulan's applications can be expanded by chemically modifying to obtain grafted pullulan, polymer blends, nanocomposites and hydrogels which have been gaining importance in recent years. Over the years, many patents using pullulan have been granted for different applications worldwide. This review sheds light on all these aspects of pullulan including its future scope. [ABSTRACT FROM AUTHOR]

Abstract Objectives The ascomycotic yeast-like fungus Aureobasidium exhibits the natural ability to synthesize several secondary metabolites, like polymalic acid, pullulan, or polyol lipids, with potential biotechnological applications. Combined with its polyextremotolerance, these properties make Aureobasidium a promising production host candidate. Hence, plenty of genomes of Aureobasidia have been sequenced recently. Here, we provide the annotated draft genome sequence of the polyol lipid-producing strain A. pullulans NRRL 62042. Data description The genome of A. pullulans NRRL 62042 was sequenced using Illumina NovaSeq 6000. Genome assembly revealed a genome size of 24.2 Mb divided into 39 scaffolds with a GC content of 50.1%. Genome annotation using Genemark v4.68 and GenDBE yielded 9,596 genes.

Objectives: The ascomycotic yeast-like fungus Aureobasidium exhibits the natural ability to synthesize several secondary metabolites, like polymalic acid, pullulan, or polyol lipids, with potential biotechnological applications. Combined with its polyextremotolerance, these properties make Aureobasidium a promising production host candidate. Hence, plenty of genomes of Aureobasidia have been sequenced recently. Here, we provide the annotated draft genome sequence of the polyol lipid-producing strain A. pullulans NRRL 62042. Data description: The genome of A. pullulans NRRL 62042 was sequenced using Illumina NovaSeq 6000. Genome assembly revealed a genome size of 24.2 Mb divided into 39 scaffolds with a GC content of 50.1%. Genome annotation using Genemark v4.68 and GenDBE yielded 9,596 genes. [ABSTRACT FROM AUTHOR]

The objective of this study was to optimize the fermentation parameters by using response surface methodology for increasing pullulan production of Aureobasidium pullulans AZ-6 in synthetic fermentation medium. The optimum conditions were determined as follows: the initial sucrose concentration (X1): 100 g/L; the initial peptone concentration (X2): 11.31 g/L; the initial pH (X3): 6.48; and the temperature (X4): 24.2 °C. In the optimum conditions, the maximum exopolysaccharide (EPS) concentration (Y1), the maximum pullulan concentration (Y2), the specific growth rate (Y3), the maximum specific pullulan formation rate (Y4), and the pullulan yield (Y5) were expected to be obtained as 37.078 and 35.372 g/L, 0.062 h−1, 0.021 [g pullulan/(g mo.h)], and 53.681%, respectively. The observed values of Y1, Y2, Y3, Y4, and Y5 were 36.950 g/L, 35.470 g/L, 0.064 h−1, 0.036 [g pullulan/(g mo.sa)], and 54.480%, respectively, as a result of validation experiments. EPS samples were characterized by Fourier-transform infrared spectroscopy and scanning electron microscope analyses. The optimization process caused more than a 50% increase in EPS and pullulan concentrations. This study showed that the color-variant A. pullulans AZ-6 strain could become a significant industrial strain if it is explored further in the future to produce pullulan on a larger scale. [ABSTRACT FROM AUTHOR]

Due to its unique physicochemical properties, Pullulan is an exopolysaccharide with many applications in the food, biomedical, and pharmaceutical industries. Aiming to reduce its production cost, an interesting alternative is to consider other possibilities of raw materials, including the production of this biopolymer in a lignocellulosic biorefinery concept. Xylose is the main sugar of hemicellulosic hydrolysates obtained from different biomasses, and it is a sugar still not extensively exploited regarding its potential for pullulan production. This study aimed to evaluate the production of pullulan from sugarcane bagasse hemicellulosic hydrolysate by cultivating Aureobasidium pullulans ATCC 42023 in a bubble column reactor. The hemicellulosic hydrolysate was obtained through dilute acid treatment carried out in a stirred tank reactor before being detoxified to remove microbial growth inhibitors. The maximum concentration of 28.62 ± 1.43 g/L of pullulan was obtained after 120 h of fermentation in a bubble column reactor in batch mode. Analysis of spectroscopic properties through FTIR of the obtained pullulan revealed α-(1→6)-linked maltosyl units, similar to those of commercial samples of the biopolymer. XRD analysis showed that the prepared pullulan is amorphous, and a homogeneous morphology with a smooth surface of the pullulan was observed in SEM analysis. This study showed the potential of the production of pullulan from sugarcane bagasse hemicellulosic hydrolysate in a bubble column bioreactor, an alternative strategy for the industrial production of this biopolymer. [ABSTRACT FROM AUTHOR]

A high-yielding microbial polysaccharide-producing strain, named RM1603, was isolated from rhizosphere soil and identified by morphological and phylogenetic analysis. The extracellular polysaccharides (EPS) were identified by thin-layer chromatography and infrared spectroscopy. The fermentation conditions were optimized by single factor experiments in shake flasks and a 5-L fermentor. The results of morphological and phylogenetic tree analysis showed that RM1603 was a strain of Aureobasidium pullulans. Its microbial polysaccharide was identified as pullulan, and the EPS production capacity reached 33.07 ± 1.03 g L−1 in shake flasks. The fermentation conditions were optimized in a 5-L fermentor, and were found to encompass an initial pH of 6.5, aeration rate of 2 vvm, rotor speed of 600 rpm, and inoculum size of 2 %. Under these conditions, the pullulan yield of RM1603 reached 62.52 ± 0.24 g L−1. Thus, this study contributes RM1603 as a new isolation with high-yielding pullulan and potential application value in biotechnology. [ABSTRACT FROM AUTHOR]

Melanin is a high value bioproduct generated through the fermentation of Aureobasidium pullulans, playing a crucial role in various fields, including food, medicine, environmental protection, and materials science. However, its high production costs and low synthetic yields significantly limit its applications. Therefore, it is essential to identify high-yield strains, reduce production costs, and optimize fermentation strategies. In this study, a high melanin-yielding Aureobasidium pullulans 53LC7 was screened and identified, and the fermentation process was optimized based on melanin yield, color value, and pullulan yield. The results indicated that the melanin yield peaked at an initial pH of 6.0, temperature of 27 °C, fermentation time of 6.5 d, and inoculation quantity of 2.5%, achieving a melanin yield of 16.33 g/L. Subsequently, huangjiu lees, a byproduct of huangjiu production, was incorporated into the fermentation medium, resulting in a melanin yield of 5.91 g/L. This suggests that the Aureobasidium pullulans was not effectively utilizing huangjiu lees. To address this, we employed an adaptive evolution strategy, which increased the melanin yield to 8.72 g/L. The enhanced production was correlated with the expression of key genes, including FKS, PKS, and Cmr1. Finally, cellulase was utilized to convert the crude fibers in huangjiu lees, which were difficult to utilize, into usable substrates, while pullulanase was employed to minimize byproduct formation in the fermentation system, resulting in a melanin yield of 19.07 g/L. This study not only provides promising strains for further research but also offers valuable insights for resource production technologies.

This study addresses the production enhancement of pullulan, an extracellular polysaccharide with various applications. Pullulan is primarily produced by Aureobasidium pullulans (A. pullulans), and genetic modification is commonly used to increase its yield. However, there is a need for a more efficient and safer method. To achieve this, we designed a high-throughput screening system utilizing a unique fluorescent protein specific to pullulan. Ultraviolet (UV) mutagenesis was applied to create a pool of mutant strains, and flow cytometry allowed for single-cell screening. Our approach yielded strain M1-B3, which exhibited a substantial increase in pullulan production from 26.5 g/L to 76.88 g/L. Additionally, the molecular weight of the produced pullulan significantly increased, expanding its potential commercial application. This study demonstrates an efficient and safe method to enhance pullulan production in A. pullulans. The UV mutagenesis and flow cytometry based on screening not only increased yield but also improved pullulan's molecular weight. The adaptability of this method to other polysaccharides and its potential for genomic analysis and broader applications make it a valuable tool in bioproduction. [ABSTRACT FROM AUTHOR]

Due to its unique physicochemical properties, Pullulan is an exopolysaccharide with many applications in the food, biomedical, and pharmaceutical industries. Aiming to reduce its production cost, an interesting alternative is to consider other possibilities of raw materials, including the production of this biopolymer in a lignocellulosic biorefinery concept. Xylose is the main sugar of hemicellulosic hydrolysates obtained from different biomasses, and it is a sugar still not extensively exploited regarding its potential for pullulan production. This study aimed to evaluate the production of pullulan from sugarcane bagasse hemicellulosic hydrolysate by cultivating Aureobasidium pullulans ATCC 42023 in a bubble column reactor. The hemicellulosic hydrolysate was obtained through dilute acid treatment carried out in a stirred tank reactor before being detoxified to remove microbial growth inhibitors. The maximum concentration of 28.62 ± 1.43 g/L of pullulan was obtained after 120 h of fermentation in a bubble column reactor in batch mode. Analysis of spectroscopic properties through FTIR of the obtained pullulan revealed α-(1→6)-linked maltosyl units, similar to those of commercial samples of the biopolymer. XRD analysis showed that the prepared pullulan is amorphous, and a homogeneous morphology with a smooth surface of the pullulan was observed in SEM analysis. This study showed the potential of the production of pullulan from sugarcane bagasse hemicellulosic hydrolysate in a bubble column bioreactor, an alternative strategy for the industrial production of this biopolymer.

Pullulan is a polymer produced by Aureobasidium spp. The yield of pullulan production can be impacted by the cellular differentiation of Aureobasidium spp., which changes with alterations in the growth environment. To improve pullulan yield, identifying key factors that regulate cellular differentiation is crucial. In this study, the main form of pullulan synthesis in Aureobasidium pullulans NG was through swollen cells (SC). The results showed that citric acid (CA) can regulate the cellular differentiation of Aureobasidium pullulans NG by accumulating higher levels of CA in the cells to maintain growth in SC form and increase pullulan production. The addition of 1.0% CA to Aureobasidium pullulans NG for 96 h resulted in a significant increase in pullulan production, producing 18.32 g/l compared to the control group which produced 10.23 g/l. Our findings suggest that controlling cellular differentiation using CA is a promising approach for enhancing pullulan production in Aureobasidium pullulans. Key points: • The regulation of cell differentiation in Aureobasidium pullulans NG is demonstrated to be influenced by citric acid. • Intracellular citric acid levels in Aureobasidium pullulans NG have been shown to support the growth of swollen cells. • Citric acid has been found to increase pullulan production in Aureobasidium pullulans NG. [ABSTRACT FROM AUTHOR]

Pullulan is a commercially available exopolymer biosynthesized by Aureobasidium pullulans supplemented with nitrogen, carbon and other vital components through submerged and solid-state fermentation. These nutrients are very expensive and it raises the cost for the production of pullulan. Hence, the need of alternative cost-effective raw materials for its production is a prerequisite. Owing to its unique physicochemical features, pullulan has various applications in the food, pharmacological, and biomedical domains. Food industrial wastes generate a considerable number of by-products which accumulates and has a negative influence on the environment. These by-products are made up of proteins, carbohydrates, and other components, can be employed as substrates for the production of pullulan. The present review briefs on the pullulan production using food processing waste and by-products and the elements that impact it. It provides an insight into versatile applications of pullulan in food industries. Various challenges and future prospects in the field of research on pullulan production have been uncovered. [ABSTRACT FROM AUTHOR]

Pullulan is a linear exopolysaccharide, produced in the fermentation media of Aureobasidium pullulans, with a variety of applications in the food and pharmaceutical industries. Pullulan derivatives have growing potential for biomedical applications, but the high cost of pullulan biofabrication currently restricts its commercial use. Better control over pullulan yield, molecular weight and melanin production by altering fermentation conditions could improve the economics. In this study, the effects of sugar and mineral salt stresses on the pullulan production of A. pullulans ATCC 42023 were examined in batch processes. The chemical structure of the recovered pullulan was characterized by FTIR and NMR spectroscopy, and the molecular weight distribution was obtained via SEC. Pullulan yield and melanin production varied when the conditions were adjusted, and pullulans with different molar masses were obtained. Higher-yield pullulan production and a lower polydispersity index were observed when CuSO4 was added to the fermentation in comparison with the control and with the addition of sugars and other salts. Biofabrication of pullulan under stress conditions is a promising strategy to enhance biopolymer yield and to obtain pullulan with a targeted molecular weight. [ABSTRACT FROM AUTHOR]

Hazelnut husk is one of the most abundant agricultural residue in Turkey. Valorization of this lignocellulosic biomass would provide a promoting alternative for economical production of pullulan. In this study, dried hazelnut husk hydrolysate was used directly as fermentation medium for pullulan production by a domestic strain of Aureobasidium pullulans. The aim of this work was the optimization of some fermentation medium parameters by central composite design using response surface methodology (RSM). The effects of (NH4)2SO4 concentration, the volume of concentrated H2SO4 and the amount of ground hazelnut husk on pullulan production were optimized by RSM. The optimum levels of the fermentation parameters defined as 7.2 gL−1, 2.5 mL and 20 g, respectively. The maximum pullulan and exopolysaccharide concentrations were determined as 74.39 and 75.95 gL−1, respectively in the optimum conditions. Specific growth rate of the strain was found as 0.097 h−1. FTIR spectral attributes confirmed the structure of pullulan. Thermal decomposition temperature of synthesized pullulan was found as 247.15 °C. This study showed that hazelnut husk was one of the novel substrate for production of the pullulan by A. pullulans AZ-6. No previous work was found to utilize dried hazelnut husk as fermentation medium for pullulan production by A. pullulans. [ABSTRACT FROM AUTHOR]

Introduction: Biopolymers, such as pullulan, a natural exopolysaccharide from Aureobasidium pullulans, and their nanocomposites are commonly used in the food, pharmaceutical, and medical industries due to their unique physical and chemical properties.Methods: Pullulan was synthesized by the A. pullulans ATCC 201253 strain. Nanocomposite films based on biosynthesized pullulan were prepared and loaded with different concentrations of silver nanoparticles (AgNPs) synthesized by the Fusarium culmorum strain JTW1. AgNPs were characterized by transmission electron microscopy, Zeta potential measurements, and Fourier-transform infrared spectroscopy. In turn, the produced films were subjected to physico-chemical analyses such as goniometry, UV shielding capacity, attenuated total reflection–Fourier-transform infrared spectroscopy, scanning electron microscopy, and X-ray photoelectron spectroscopy, and their mechanical and degradation properties were assessed. The antibacterial assays of the nanoparticles and the nanocomposite films against both food-borne and reference pathogens, including Listeria monocytogenes, Salmonella infantis, Salmonella enterica, Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, and Klebsiella pneumoniae, were performed using standard methods.Results: AgNPs were small (mean 15.1 nm), spherical, and displayed good stability, being coated with protein biomolecules. When used in higher concentrations as an additive to pullulan films, they resulted in reduced hydrophilicity and light transmission for both UV-B and UV-A lights. Moreover, the produced films exhibited a smooth surface. Therefore, it can be concluded that the addition of biogenic AgNPs did not change the morphology and texture of the films compared to the control film. The nanoparticles and nanocomposite films demonstrated remarkable antibacterial activity against both food-borne and reference bacteria. The highest activity of the prepared films was observed against L. monocytogenes.Discussion: The obtained results suggest that the novel nanocomposite films prepared from biosynthesized pullulan and AgNPs can be considered for use in the development of medical products and food packaging. Moreover, this is the first report on pullulan-based nanocomposites with mycogenic AgNPs for such applications.

This study addresses the production enhancement of pullulan, an extracellular polysaccharide with various applications. Pullulan is primarily produced by Aureobasidium pullulans (A. pullulans), and genetic modification is commonly used to increase its yield. However, there is a need for a more efficient and safer method. To achieve this, we designed a high-throughput screening system utilizing a unique fluorescent protein specific to pullulan. Ultraviolet (UV) mutagenesis was applied to create a pool of mutant strains, and flow cytometry allowed for single-cell screening. Our approach yielded strain M1-B3, which exhibited a substantial increase in pullulan production from 26.5 g/L to 76.88 g/L. Additionally, the molecular weight of the produced pullulan significantly increased, expanding its potential commercial application. This study demonstrates an efficient and safe method to enhance pullulan production in A. pullulans. The UV mutagenesis and flow cytometry based on screening not only increased yield but also improved pullulan’s molecular weight. The adaptability of this method to other polysaccharides and its potential for genomic analysis and broader applications make it a valuable tool in bioproduction.

Copyright of Food Research & Development is the property of Food Research & Development Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Efficient pullulan production has long been a central research focus. This study used maltodextrin as the carbon source for pullulan production by Aureobasidium pullulans fermentation. A hybrid optimization approach, integrating orthogonal experimental design (OED), backpropagation artificial neural network (BP-ANN), and elite strategy non-dominated sequential genetic algorithm-II (NSGA-II), was developed. Range analysis based on OED revealed that MgSO 4 ·7H 2 O significantly affects production but less impacts molecular weight, while pH notably influences molecular weight with a lesser effect on production, underscoring the need for multi-objective optimization. The BP-ANN model showed strong predictive capabilities, with goodness-of-fit values of 0.984 and 0.980 for production and molecular weight, respectively. Using this model as the fitness function for the optimization algorithm enhanced efficiency. Taking cost factors into account, the BP-ANN-NSGA-II algorithm identified the optimal fermentation medium conditions, resulting in a 6.89 % increase in production, a 368.97 % increase in molecular weight, and a 42.49 % reduction in cost. The maximum comprehensive optimization efficiency is 63.73 %, and the multi-objective optimization is better than the single objective optimization. This method significantly guides the improvement of pullulan fermentation optimization efficiency. [ABSTRACT FROM AUTHOR]

Pullulan has many potential applications in the food, pharmaceutical, cosmetic and environmental industries. However, the yield and molecular properties of pullulan produced by various strains still need to be promoted to fit the application needs. A novel yeast-like strain Aureobasidium pullulans BL06 producing high molecular weight (Mw) pullulan (3.3 × 106 Da) was isolated and identified in this study. The remarkable Mw of pullulan produced by A. pullulans BL06 was the highest level ever reported thus far. To further regulate the biosynthesis of pullulan in A. pullulans BL06, three gene knockout strains A. pullulans BL06 ΔPMAs, A. pullulans BL06 Δmel, and A. pullulans BL06 ΔPMAsΔmel, were constructed. The results showed that A. pullulans BL06 ΔPMAs could produce 140.2 g/L of moderate Mw (1.3 × 105 Da) pullulan after 120 h of fermentation. The highest yield level of pullulan to date could vastly reduce its production cost and expand its application scope and potential. The application experiments in food preservation showed that the moderate-Mw pullulan obtained in this work could reduce the weight loss of celery cabbages and mangos by 12.5% and 22%, respectively. Thus, the novel strains A. pullulans BL06 and A. pullulans BL06 ΔPMAs possessed unlimited development prospects in pullulan production at various Mw ranges and pullulan applications in multiple fields.

To investigate the role of the sugar transporter MAL31 on pullulan biosynthesis, the coding gene mal31 was respectively disrupted and overexpressed in the parental strain A. pullulans CCTCC M 2012259 to construct mutants of A. pullulans Δ mal31 and A. pullulans Mal31. Batch pullulan production significantly decreased by 69.1 % in A. pullulans Δ mal31 but increased by 15.9 % in A. pullulans Mal31 , as compared to the parental strain. We performed kinetics analysis, assays of key enzymes, determination of intracellular UDPG, NADH, and ATP contents, and measurement of transcriptional levels of genes associated with pullulan biosynthesis and excretion. The results confirmed that the mal31 disruption decreased the glucose consumption rate, decreased the formation rate and titer of pullulan, but increased the intracellular UDPG supply for β -glucan accumulation. In contrast, the mal31 overexpression increased the transcriptional levels of genes associated with pullulan biosynthesis, and accelerated the rates of glucose consumption and pullulan formation, thereby increased pullulan production. Our findings revealed that MAL31 is involved in the transport of precursors for pullulan biosynthesis. This study provides an accurate operating site for genetic modification of A. pullulans for improving pullulan production and also presents a feasible technique route for the overproduction of other polysaccharides. • The mal31 disruption decreased pullulan production but favored β -glucan formation. • The mal31 overexpression increased both UDPG supply and pullulan production. • MAL31 is involved in transmembrane transport of precursors for pullulan biosynthesis. [ABSTRACT FROM AUTHOR]

This review examines the immobilization of A. pullulans cells for production of the fungal polysaccharide pullulan. Pullulan is a water-soluble gum that exists structurally as a glucan consisting primarily of maltotriose units, which has a variety of food, non-food and biomedical applications. Cells can be immobilized by carrier-binding or entrapment techniques. The number of studies utilizing carrier-binding as a method to immobilize A. pullulans cells appears to outnumber the investigations using cell entrapment. A variety of solid supports, including polyurethane foam, sponge, diatomaceous earth, ion-exchanger, zeolite and plastic composite, have been employed to immobilize pullulan-producing A. pullulans cells. The most effective solid support that was used to adsorb the fungal cells was polyurethane foam which produced polysaccharide after 18 cycles of use. To entrap pullulan-producing fungal cells, agents such as polyurethane foam, polyvinyl alcohol, calcium alginate, agar, agarose, carrageenan and chitosan were investigated. Polysaccharide production by cells entrapped in polyurethane foam, polyvinyl alcohol or calcium alginate was highest and the immobilized cells could be reutilized for several cycles. It was shown that the pullulan content of the polysaccharide synthesized by cells entrapped in calcium alginate beads was low, which limits the method's usefulness for pullulan production. Further, many of the entrapped fungal cells synthesized polysaccharide with a low pullulan content. It was concluded that carrier-binding techniques may be more effective than entrapment techniques for A. pullulans cell immobilization, since carrier-binding is less likely to affect the pullulan content of the polysaccharide being synthesized. [ABSTRACT FROM AUTHOR]

In this paper, the pullulan produced by A. pullulans ATCC 15233 was characterized and chemically modified. The pullulan derivative was synthesized with an acetylation reaction with a degree of substitution (DS) of 2.90. Its structural elucidation was performed using infrared (IR) and nuclear magnetic resonance (NMR) spectroscopy. The results showed signs of α (1–4) and α (1–6) glycosidic bonds, typical of pullulan. The molecular weight (MW) of pullulan was 93.47 kDa. The thermal analysis conducted indicated increased thermal stability (Td) and glass transition temperature (Tg) due to acetylation. Homogeneous morphologies of the samples under analysis were obtained using scanning electron microscopy (SEM). The mechanical properties of the pullulan film exhibited tensile strength (TS) values of 1.70 ± 0.48 MPa and an elongation at break (% E) of 11.89%. Likewise, the pullulan acetate film recorded a TS of 1.23 ± 0.09 MPa and a % E of 1.25%. The results suggest that the physicochemical properties of biosynthesized pullulan and pullulan derivatives can be considered in the development of biomaterials.

Copyright of Food Research & Development is the property of Food Research & Development Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

This review highlights the applications of pullulan in biomedical field, focusing on drug delivery. Pullulan is a microbial exo-polysaccharide produced by yeast like fungus Aureobasidium pullulans and it has been declared safe by FDA in United States and has GRAS status. Pullulan has biocompatible, biodegradable, non-mutagenic, non-toxic, noncarcinogenic, non-immunogenic properties, as well as other functional properties. Furthermore, pullulan can be easily derivatized by several chemical reactions such as etherification, amidification, esterification, oxidation and copolymerization in order to widen its applications. Due to its unique features pullulan and its derivative is being explored for various biomedical applications like drug and gene delivery, tissue engineering, wound healing, diagnostic imaging, etc. This research was supported through Nucleu project PN 1941-04 01. [ABSTRACT FROM AUTHOR]

Pullulan is a microbial polymer, commercially produced from Aureobasidium pullulans. Downstream processing of pullulan involves a multi-stage process which should be efficient, safe and reproducible. In liquid-liquid separations, firstly cell free extract is separated. Cell biomass can be separated after fermentation either by centrifugation or filtration. Due to practically insolubility of pullulan in organic solvents, ethanol and isopropanol are the most commonly used organic solvents for its recovery. Pullulan can also be purified by chromatographic techniques, but these are not cost effective for the purification of pullulan. Efficient aqueous two-phase system can be used for the purification of pullulan. The current review describes the methods and perspectives used for solid-liquid separation, liquid-liquid separations and finishing steps for the recovery of pullulan. Techniques used to determine the structural attributes of pullulan have also been highlighted. • Overview of downstream processing of pullulan has been elaborated. • Organic solvents and precipitation conditions for pullulan recovery have been discussed. • Conventional techniques and aqueous-phase system used for pullulan recovery has also been discussed. • Structural attributes determination methods of pullulan have been described. [ABSTRACT FROM AUTHOR]

Bacterial cellulose (BC) exhibits a unique combination of porosity, tensile strength, reticulated crystal structure and biocompatibility useful for many applications in the food, biomedical and other industries. Polysaccharide addition has been shown to improve the production and the mechanical properties of BC nanocomposites. This study examined the effect of pullulan on BC fermentation as well as the co-culturing of the BC producer with Aureobasidium pullulans, a fungal strain that produces pullulan as an exopolysaccharide. Results showed that a 1% pullulan addition improved Young's modulus of BC pellicles for sixfold. Addition of pullulan at 1.5% and 2% levels could increase the BC production from 0.447 to 0.814 and 1.997 g/L, respectively. The co-culture fermentation demonstrated a mixed effect on the aggregation and bundling of BC while resulting in a significant improvement in mechanical properties. The study provided a polysaccharide additive and a novel fermentation method to produce BC with improved properties. [ABSTRACT FROM AUTHOR]

Bacterial cellulose (BC11 Uncommon abbreviation used: BC – Bacterial Cellulose) is a biomaterial produced by various strains of microorganisms. BC has improved strength and unique structural properties as compared to plant cellulose, thus has many usages in the food and pharmaceutical industries. In our previous study, a novel co-culture agitated fermentation of Komagataeibacter hansenii, a BC producer, with Aureobasidium pullulans, a producer of pullulan polysaccharide, had been demonstrated where the BC produced exhibited improved mechanical properties. Therefore, this study is undertaken to analyze BC production under different medium composition using response surface methodology (RSM) in shake-flasks and benchtop bioreactors. A verified local high point provided 22.4% higher BC production and 4.5- to 6- folds higher elastic moduli in shake-flasks and bioreactors compared to the baseline media. Overall, the study had revealed the potential of the co-culturing method to enhance BC production while maintaining the desired mechanical properties of BC produced in shake-flasks and larger scale bioreactors.

A colour-variant strain of Aureobasidium pullulans FB-1 was used for the production of pullulan in a stirred tank reactor in a medium composed of sucrose, ammonium sulphate, yeast extract, K2HPO4, NaCl and MgSO4. Maximum pullulan production (4.8%, w/v) was obtained at an agitation rate of 300 rpm and aeration of 0.75 vvm in a stirred tank reactor (1 L) with a fermentation time of 6 days. Fourier transformed infrared and NMR spectrum of crude pullulan from A. pullulans FB-1 revealed solely α-(1→6) linked maltosyl units, in accord with the generally accepted structure of pullulan. Hydrolysis of crude pullulan, i.e. digestion with pullulanase also confirmed the identity as well as homogeneity of the produced pullulan. XRD analysis of crude pullulan verified its amorphous nature. Thermal analysis showed melting range of crude pullulan from 245.99–295.71 °C.

Copyright of Shipin Kexue/ Food Science is the property of Food Science Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

In this study, cost-effective substrates such as cassava starch, corn steep liquor (CSL) and soybean meal hydrolysate (SMH) were used for pullulan production by Aureobasidium pullulans CCTCC M 2012259. The medium was optimized using response surface methodology (RSM) and artificial neural network (ANN) approaches, and analysis of variance indicated that the ANN model achieved higher prediction accuracy. The optimal medium predicted by ANN was used to produce high molecular weight pullulan in high yield. SMH substrates increased both biomass and pullulan titer, while CSL substrates maintained higher pullulan molecular weight. Results of kinetic parameters, key enzyme activities and intracellular uridine diphosphate glucose contents revealed the physiological mechanism of changes in pullulan titer and molecular weight using different substrates. Economic analysis of batch pullulan production using different substrates was performed, and the cost of nutrimental materials for CSL and SMH substrates was decreased by 46.1% and 49.9%, respectively, compared to the control using glucose and yeast extract as substrates, which could improve the competitiveness of pullulan against other polysaccharides in industrial applications. • Cassava starch, CSL, and SMH were used for pullulan production by A. pullulans. • The medium components were optimized and predicted by RSM and ANN models. • ANN had higher prediction accuracy than RSM for medium optimization. • SMH increased pullulan titer, while CSL favored high Mw pullulan production. • The cost of pullulan production from CSL and SMH substrates reduced by about 50%. [ABSTRACT FROM AUTHOR]

To improve β-1,3–1,6-D-glucan (β-glucan) production by Aureobasidium pullulans, an Agrobacterium tumefaciens–mediated transformation method was developed to screen a mutant A. pullulans CGMCC 19650. Based on thermal asymmetric–interlaced PCR detection, DNA sequencing, BLAST analysis, and quantitative real-time PCR assay, the T-DNA was identified to be inserted in the coding region of mal31 gene, which encodes a sugar transporter involved in pullulan biosynthesis in the mutant. The maximal biomass and β-glucan production under batch fermentation were significantly increased by 47.6% and 78.6%, respectively, while pullulan production was decreased by 41.7% in the mutant, as compared to the parental strain A. pullulans CCTCC M 2012259. Analysis of the physiological mechanism of these changes revealed that mal31 gene disruption increased the transcriptional levels of pgm2, ugp, fks1, and kre6 genes; increased the amounts of key enzymes associated with UDPG and β-glucan biosynthesis; and improved intracellular UDPG contents and energy supply, all of which favored β-glucan production. However, the T-DNA insertion decreased the transcriptional levels of ags2 genes, and reduced the biosynthetic capability to form pullulan, resulting in the decrease in pullulan production. This study not only provides an effective approach for improved β-glucan production by A. pullulans, but also presents an accurate and useful gene for metabolic engineering of the producer for efficient polysaccharide production. Key points: • A mutant A. pullulans CGMCC 19650 was screened by using the ATMT method. • The mal31 gene encoding a sugar transporter was disrupted in the mutant. • β-Glucan produced by the mutant was significantly improved. [ABSTRACT FROM AUTHOR]

The interest in pullulan produced from industrial wastes has substantially increased due to its industrial applications. The major constraint in pullulan production is expensive and low yield. In this study, attempts were made on the evaluation of media components for maximum pullulan production from Aureobasidium pullulans MTCC 1991 using cassava waste in solid-state fermentation. A statistical approach was scrutinized to evaluate the effects of yeast extract, K2HPO4, (NH4)2SO4, MgSO4⋅7H2O, and NaCl on pullulan production. The optimal conditions eliciting the maximum pullulan yield (6.45 g pullulan/L of culture broth) were found to be 0.6 g/L yeast extract, 3 g/L K2HPO4, 0.2 g/L (NH4)2SO4, 0.3 g/L MgSO4⋅7H2O, and 1 g/L NaCl. The produced pullulan was extracted using the solvent precipitation method and characterized by Fourier transform infrared spectroscopy and proton nuclear magnetic resonance spectroscopy spectroscopic analysis that confirmed pullulan with linear α (1–6) linkage. The results revealed that cassava waste could be used as an effective carbon source for the production of pullulan. [ABSTRACT FROM AUTHOR]

Curcumin is a multitargeting nutraceutical with numerous health benefits, however, its efficacy is limited due to poor aqueous solubility and reduced bioavailability. While nano-formulation has emerged as an alternative to encounter such issues, it often involves use of toxic solvents. Microbial synthesis may be an innovative solution to address this lacuna. Present study, for the first time, reports exploitation of Aureobasidium pullulans RBF4A3 for production of nano-curcumin. For this purpose, Aureobasidium pullulans RBF4A3 was inoculated in YPD media along with curcumin (0.1 mg/mL) and incubated for 24 h, 48 h, and 72 h. Subsequently, residual sugar, biomass, EPS concentration, curcumin concentration, and curcumin nanoparticle size were measured. As a result, nano-curcumin with an average particle size of 31.63 nm and enhanced aqueous solubility was obtained after 72 h. Further, investigations suggested that pullulan, a reducing polysaccharide, played a significant role in curcumin nano-formulation. Pullulan-mediated nano-curcumin formulation, with an average particle size of 24 nm was achieved with conversion rate of around 59.19 %, suggesting improved aqueous solubility. Additionally, the anti-oxidant assay of the resulting nano-curcumin was around 53.7 % per μg. Moreover, kinetics and thermodynamic studies of pullulan-based nano-curcumin revealed that it followed first-order kinetics and was favored by elevated temperature for efficient bio-conversion. Also, various physico-chemical investigations like FT-IR, NMR, and XRD reveal that pullulan backbone remains intact while forming curcumin nanoparticle. This study may open up new avenues for synthesizing nano-polyphenols through a completely green and solvent free process with plausible diverse applications. • First report of microbial fermentation based production of nano-curcumin. • Pullulan might be the key biomolecule responsible for nanonization of curcumin. • Bio-nanoconversion of curcumin was found to follow first order reaction kinetics. • Nano-formation enhanced aqueous solubility and antioxidant activity of curcumin. [ABSTRACT FROM AUTHOR]

The content of pullulan and melanin in 500 mutants of Aureobasidum pullulans obtained by ultraviolet mutagenesis were examined and statistically analyzed, and a strong positive correlation was found between them. The result was further confirmed by culturing wild type strain As3.3984 in different media. Then we constructed melanin-deletion mutant As- Δalb1 and pullulan-deletion mutant As- Δpul. As- Δalb1 was a melanin-free strain with the yield of pullulan decreased by 41.01%. The supplementation of melanin in the culture of As- Δalb1 increased the production of pullulan. As- Δpul synthesized neither pullulan nor melanin and recovered melanin synthesis by adding pullulan to the medium. The results suggested that high concentration- of pullulan induced morphological transformation and synthesis of melanin, and melanin promoted the synthesis of pullulan. The pullulan biosynthetic genes, upt , pgm , ugp , and pul , were down-regulated, while the negative regulatory gene of pullulan synthesis, creA , was up-regulated by melanin deficiency. • The synergistic correlation exists between the synthesis of pullulan and melanin. • High concentrations of pullulan induced the synthesis of melanin. • Melanin promotes the synthesis of pullulan. [ABSTRACT FROM AUTHOR]

Sesame seed oil cake (SSOC) was applied as a novel substrate for production of high-added value pullulan by Aureobasidium pullulans KY767024 (a melanin-deficient strain). The optimization of production conditions by central composite design showed that the maximum production yield (54.50 g pullulan/kg substrate) was obtained under optimum conditions (Liquid-solid ratio (LSR) of 1.7 v/w, inoculum size of 7 mL, and temperature of 23 °C), which was confirmed by the experimental data (52.50 ± 0.73 g/kg). The obtained pullulan had an average-molecular weight of 358.736 kDa. The present peaks in the FT-IR spectrum validated that the obtained purified precipitate was composed of pullulan. Also, the XRD pattern and TGA analysis showed that the produced pullulan had a fully amorphous structure with high thermal stability. In addition, the pullulan solutions had a different flow behavior in different concentrations and with an increase in concentration, their behavior changed from Newtonian to pseudoplastic.

Pullulan 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. • Imperative role of pullulan in drug delivery and gene delivery has been elaborated. • Application of pullulan as a scaffold for tissue engineering has been described. • The use of pullulan as blood plasma substitute has been explained. • Action of pullulan as a molecular chaperone has been described. • Potential role of pullulan in vaccination has been discussed. [ABSTRACT FROM AUTHOR]

The 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. Key points: • 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. [ABSTRACT FROM AUTHOR]

The effects of zinc sulfate at various concentrations on β -1,3-D-glucan (β -glucan) and pullulan production were investigated in flasks, and 0.1 g/L zinc sulfate was found to be the optimum concentration favoring increased β -glucan production. When batch culture of Aureobasidium pullulans CCTCC M 2012259 with 0.1 g/L zinc sulfate was carried out, the maximum dry biomass decreased by 16.9% while β -glucan production significantly increased by 120.5%, compared to results obtained from the control without zinc sulfate addition. To reveal the mechanism underlying zinc sulfate improved β -glucan production, both metabolic flux analysis and RNA-seq analysis were performed. The results indicated that zinc sulfate decreased carbon flux towards biomass formation and ATP supply, down-regulated genes associated with membrane part and cellular components organization, leading to a decrease in dry cell weight. However, zinc sulfate increased metabolic flux towards β -glucan biosynthesis, up-regulated genes related to glycan biosynthesis and nucleotide metabolism, resulting in improved β -glucan production. This study provides insights into the changes in the metabolism of A. pullulans in response to zinc sulfate, and can serve as a valuable reference of genetic information for improving the production of polysaccharides through metabolic engineering. • Zinc sulfate decreased biomass but improved β -glucan production by A. pullulans. • Metabolic flux towards biomass decreased whereas towards β -glucan increased. • Transcriptome analysis of A. pullulans was carried out by RNA-seq. • Zinc sulfate up-regulated genes related to β -glucan biosynthesis. [ABSTRACT FROM AUTHOR]

In 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 (M w) of 2.07 × 105 g.mol−1 based on gel permeation chromatography results. The decomposition temperature (T d) of the produced pullulan was ~300 °C and also, the resulting pullulan had a Newtonian flow behavior in a wide range of concentrations. • A new strain of A. pullulans was isolated from trees leaves. • Structural analysis confirmed the presence of chemical structure of pullulan. • 51.4 ± 0.50 g/L of pullulan were obtained under optimum condition. • Pullulan had a high decomposition temperature (~300 °C) based on TGA. • Pullulan solution had a Newtonian behavior in a wide range of concentrations. [ABSTRACT FROM AUTHOR]

The 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. [ABSTRACT FROM AUTHOR]

Pullulan 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. [ABSTRACT FROM AUTHOR]

Pullulan production from Aureobasidium pullulans was explored to increase yield. Non-linear hybrid mathematical tools for optimization of process variables as well as the pullulan yield were analyzed. The one variable at a time (OVAT) approach was used to optimize the maximum pullulan yield of 35.16 ± 0.29 g/L. The tools predicted maximum pullulan yields of 39.4918 g/L (genetic algorithm coupled with artificial neural network (GA-ANN)) and 36.0788 g/L (GA coupled with adaptive network based fuzzy inference system (GA-ANFIS)). The best regression value (0.94799) of the Levenberg-Marquardt (LM) algorithm for ANN and the epoch error (6.1055 × 10-5) for GA-ANFIS point towards prediction precision and potentiality of data training models. The process parameters provided by both the tools corresponding to their predicted yield were revalidated by experiments. Among the two of them GA-ANFIS results were replicated with 98.82% accuracy. Thus GA-ANFIS predicted an optimum pullulan yield of 36.0788 g/L with a substrate concentration of 49.94 g/L, incubation period of 182.39 h, temperature of 27.41 °C, pH of 6.99, and agitation speed of 190.08 rpm. [ABSTRACT FROM AUTHOR]

• An integrated bio-refinery process was used to valorize eggplant peel waste (EPW). • Simultaneous recovery of pectin (26.1%) and phenolics (20.2%) was optimized. • EPW hydrolysates were used as carbon source in production of pullulan (16.8 g/L). • Physicochemical analysis suggested good purity for pectin and pullulan sample. • Antiradical activity of extract sample was comparable to ascorbic acid and BHA. Based on a bio-refinery concept, the valorization of eggplant peel wastes (EPW) in the production of multiple value-added products was aimed. The acid-free extraction process was applied in the simultaneous recovery of pectin and phenolic compounds. The extraction variables were optimized by response surface methodology using a Box-Behnken design and the maximum yield of pectin (26.1%) and phenolic compounds (20.2%) was obtained in the extraction temperature of 90 °C, time of 90 min and liquid/solid ratio of 40 mL/g. After recovery of pectin and phenolic compounds from EPW, the solid leftovers were enzymatic hydrolyzed and the hydrolysates were used as a carbon source in the microbial production of pullulan by Aureobasidium pullulans. The produced pectin and pullulan were characterized through the chemical and structural features. The results of FT-IR and H-NMR analysis approved the predominant presence of these two polysaccharides in the isolated samples. On the other hand, the antioxidant activity of the recovered phenolic compounds extract was evaluated by DPPH and ABTS radical scavenging activity and reducing power assay. [ABSTRACT FROM AUTHOR]