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Levan, a polyfructan which consists of D-fructofuranosyl residues linked predominantly by β-(2,6) linkage as a core chain with some β-(2,1) branch chains have potential applications in the pharmaceutical, food, and cosmetic industries. The present work reports on characterization of levan produced using Pseudomonas fluorescens by Fourier transform...
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... by one-factor-at-a-time The levan synthesizing strain identified as P. fluorescens is known to utilize oxygen and/or nitrate as hydrogen acceptor. Hence, the production profile of levan under agitating and sodium nitrate added static culture condition was studied. The results of this study are shown in Fig. 4. It was observed that the levan production was slightly higher in static culture condition (6.72 g/L after 6 days) than agitating culture condition (5.08 g/L after 6 days). After 6 days of incubation no significant change was observed in accumulation of levan. In both culture conditions dry cell weight was increased exponentially up to ...
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... Levan generated by Pseudomonas fluorescens was examined, and it was found that the ideal medium composition was 60 g/L sucrose, 1.5 g/L ammonium chloride, 2.0 g/L sodium nitrate, and 15.0 g/L casein peptone. Under these conditions, the concentration of levan significantly increased from an initial value of 5.27 g/L to 15.42 g/L (Jathore et al. 2012). A novel strain, SK 21.002, capable of producing intracellular Lsc, was isolated from beet sugar-growing soil and identified as Bacillus methylotrophicus, with optimal conditions for levan production using Lsc, revealing that the presence of Mg 2+ increased production to approximately 100 g/L (Zhang et al. 2014). ...
Levan, a β-(2,6)-linked fructose polymer, exhibits diverse properties that impart versatility, rendering it a highly sought-after biopolymer with various industrial applications. Levan can be produced by various microorganisms using sucrose, food industry byproducts and agricultural wastes. Microbial levan represents the most potent cost-effective process for commercial-scale levan production. This study reviews the optimization of levan production by understanding its biosynthesis, physicochemical properties and the fermentation process. In addition, genetic and protein engineering for its increased production and emerging methods for its detection are introduced and discussed. All of these comprehensive studies could serve as powerful tools to optimize levan production and broaden its applications across various industries.
... An endosialidase, Endo92, from phage phi92 was capable of digesting K1 and K92 capsules of E. coli and is uniquely able to cleave both the α-2,8and α-2,9-linkages of sialic acid [151]. Levanases are predominately found in bacterial species such as Bacillus and Pseudomonas and can hydrolyze the β-2,6-linked D-fructofuranosyl residues of levan [152,153]. Levan is an important structure in the development of a robust biofilm for Bacillus spp.; however, it is not a necessity. It plays a role in the stability of floating biofilms, can provide a nutritional reserve, and was found to be the major polysaccharide present in the EPS matrix [154]. ...
Healthcare faces a major problem with the increased emergence of antimicrobial resistance due to over-prescribing antibiotics. Bacteriophages may provide a solution to the treatment of bacterial infections given their specificity. Enzymes such as endolysins, exolysins, endopeptidases, endosialidases, and depolymerases produced by phages interact with bacterial surfaces, cell wall components, and exopolysaccharides, and may even destroy biofilms. Enzymatic cleavage of the host cell envelope components exposes specific receptors required for phage adhesion. Gram-positive bacteria are susceptible to phage infiltration through their peptidoglycan, cell wall teichoic acid (WTA), lipoteichoic acids (LTAs), and flagella. In Gram-negative bacteria, lipopolysaccharides (LPSs), pili, and capsules serve as targets. Defense mechanisms used by bacteria differ and include physical barriers (e.g., capsules) or endogenous mechanisms such as clustered regularly interspaced palindromic repeat (CRISPR)-associated protein (Cas) systems. Phage proteins stimulate immune responses against specific pathogens and improve antibiotic susceptibility. This review discusses the attachment of phages to bacterial cells, the penetration of bacterial cells, the use of phages in the treatment of bacterial infections, and the limitations of phage therapy. The therapeutic potential of phage-derived proteins and the impact that genomically engineered phages may have in the treatment of infections are summarized.
... Maximum yield of levan was 15.42 g L −1 under optimized conditions, which were (in g L −1 ) sucrose 60; ammonium chloride 1.5, sodium nitrate 2.0 and casein peptone 15.0. 66 Table 3 highlights some of the levan-producing microorganisms. ...
Polymers have been used in various industries over the past few decades due to their tremendous applications. Among these, polyhydroxyalkanoates and poly(lactic acid) are easily biodegradable biopolymers derived from bacteria, including recombinant Escherichia coli, Alcaligenes eutrophus, Alcaligenes latus, Azotobacter vinelandii, methylotrophs and Pseudomonas. Conventional petroleum‐derived polymers have become potentially harmful to the environment due to their complex degradation process. The nonbiodegradability of synthetic polymers has become a global issue of concern. There is an urgent need for a substitute to tackle the increasing environmental stress. Microorganisms are small factories for producing different types of polymers during their growth cycle. Various features like biodegradability, biocompatibility, nontoxicity and wide substrate spectrum make such microbial polymers highly reliable. Biopolymers such as alginate, cellulose, cyanophycin, levan, polyhydroxyalkanoates, xanthan, poly(lactic acid) and poly(γ‐glutamic acid) can be obtained from different microorganisms like Aureobasdium pullulans, Acetobacter xylinum, Bacillus thermoamylovorans and Cupriavidusnecator. These are extensively used in various fields like food, medicine, wastewater treatment, biofuel production, packaging and cosmetics. Despite being advantageous in several ways, the biopolymer market still faces several hurdles. This review mainly emphasizes the different types of biopolymers, production by microorganisms and various applications of these biopolymers in different fields. The main drawback limiting the development of these polymers is the high production cost and low efficiency of the microbial strains. Genetic recombination is an efficient technique to enhance the microbial yield and to expand the biopolymer market size. © 2023 Society of Chemical Industry (SCI).
... FTIR spectra were captured in the 4000-400 cm − 1 range. [44]. ...
... Based on the spectra in the previous studies [44,54,57,58], it was determined that the polymer obtained at different temperatures was levan. The absence of a one-to-one match between levan polymers obtained at different temperatures reveals the differences between levan structures. ...
Levan is a biopolymer with many different uses. Temperature is an important parameter in biopolymer synthesis. Herein, levan production was carried out from Bacillus haynesii, a thermophilic microorganism, in the temperature range of 4 o C-95 o C. The highest levan production was measured as 10.9 g/L at 37 o C. The synthesized samples were characterized by FTIR and NMR analysis. The particle size of the levan samples varied between 153-824.4 nm at different temperatures. In levan samples produced at high temperatures, the water absorption capacity is higher in accordance with the particle size. Irregularities were observed in the surface pores at temperatures of 60 o C and above. The highest emulsion capacity of 83.4% was measured in the sample synthesized at 4°C. The antioxidant activity of all levan samples synthesized at different temperatures was measured as 84% on average. All synthesized levan samples showed antibacterial effect on pathogenic bacteria. In addition, levan synthesized at 45 o C showed the highest antimicrobial effect on E.coli ATCC 35218 with an inhibition zone of 21.3±1.82 mm. Antimicrobial activity against yeast sample C.albicans, was measured only in levan samples synthesized at 80 o C, 90 o C, 95 o C temperatures. Levan synthesized from Bacillus haynesii at low and high temperatures showed differences in characterization and bioactivity.
... The peaks at 924 and 808 cm −1 revealed symmetric stretching vibration and C-H bending vibration of furanose, respectively [62]. FTIR spectrum of levan biopolymer produced in this study was found to be quite compatible with the ones that were reported in the literature and obtained via various types of microbial species [62][63][64]. 13 C NMR and 1 H NMR analyzes were carried out to confirm the FTIR results of levan biopolymer (Fig. 9). In 13 C NMR spectrum, a distinct peak was observed at 104.16 ppm, which was related with C2 signal of β-fructose. ...
In this study, levan biopolymer (a fructose-based exopolysaccharide) was produced by Paenibacillus polymyxa HCT33-3, which was immobilized onto the composite of silica-coated magnetite nanoparticles (Fe3O4@SiO2 NPs). For this purpose, firstly, synthesis of magnetite nanoparticles (Fe3O4 NPs) was followed by Fe3O4@SiO2 composite formation to enhance its stability. The characterization studies were investigated with the scanning electron microscopy and energy dispersive X-ray analyzes, transmission electron microscopy, X-ray diffraction analyses. Then, the microorganism Paenibacillus polymyxa HCT33-3, immobilized onto Fe3O4@SiO2 and levan fermentation was carried out in the media including molasses. Effects of molasses percentage in the growth media, initial pH, temperature and fermentation period parameters were tested on levan production capability. The highest levan concentration of 35.8 g/L was obtained at the 54th hour in the medium including 20% molasses (v/v) with the initial pH and temperature values of 7.0 and 37 °C, respectively. The characterization studies clearly confirmed that the produced exopolysaccharide was levan, which showed an antimicrobial effect against all microorganisms used in the study, besides its effectiveness on the biofilm formed by Pseudomonas aeruginosa. This is the first study focusing on high-value levan biopolymer production by the immobilized microorganisms onto Fe3O4@SiO2 nanocomposite in the growth media including molasses as the sole carbon source. This environmentally friendly process, which can potentially enable the repeated use of the cells, seemed to be significantly advantageous in terms of both cost and sustainability.
... After the extraction of levan, precipitates were hydrolyzed with 1 mL of 0.1 M HCl and the fructose content was estimated by the DNS method. The quantity of fructose found in the sample was divided by the factor 1.11 to determine the quantity of levan produced [22]. Using Fourier-transform ionization radiation (FTIR) spectroscopy (Shimadzu/Prestige-21), the chemical bonds and functional groups of the levan synthesis were determined. ...
Levan is a homopolysaccharide of fructose units that repeat as its structural core. As an exopolysaccharide (EPS), it is produced by a great variety of microorganisms and a small number of plant species. The principal substrate used for levan production in industries, i.e., sucrose, is expensive and, hence, the manufacturing process requires an inexpensive substrate. As a result, the current research was designed to evaluate the potential of sucrose-rich fruit peels, i.e., mango peels, banana peels, apple peels, and sugarcane bagasse, to produce levan using Bacillus subtilis via submerged fermentation. After screening, the highest levan-producing substrate, mango peel, was used to optimize several process parameters (temperature, incubation time, pH, inoculum volume, and agitation speed) employing the central composite design (CCD) of response surface methodology (RSM), and their impact on levan production was assessed. After incubation for 64 h at 35 °C and pH 7.5, the addition of 2 mL of inoculum, and agitation at 180 rpm, the highest production of levan was 0.717 g/L of mango peel hydrolysate (obtained from 50 g of mango peels/liter of distilled water). The F-value of 50.53 and p-value 0.001 were calculated using the RSM statistical tool to verify that the planned model was highly significant. The selected model’s accuracy was proven by a high value (98.92%) of the coefficient of determination (R2). The results obtained from ANOVA made it clear that the influence of agitation speed alone on levan biosynthesis was statistically significant (p-value = 0.0001). The functional groups of levan produced were identified using FTIR (Fourier-transform ionization radiation). The sugars present in the levan were measured using HPLC and the levan was found to contain only fructose. The average molecular weight of the levan was 7.6 × 106 KDa. The findings revealed that levan can be efficiently produced by submerged fermentation using inexpensive substrate, i.e., fruit peels. Furthermore, these optimized cultural conditions can be applied on a commercial scale for industrial production and commercialization of levan.
... Levan polysaccharide is present in many microbial products and plants [12]. Microbial levan production was performed using numerous kinds of microorganisms such as Pseudomonas, Zymomonas, Bacillus, Xanthomonas, and Streptococcus [13]. Levansucrase, an extracellular enzyme, it is predominantly produced by the bacteria Bacillus sp., a Gram-positive bacterium [5]. ...
... Among the different found local fruit wastes (peels), the production of levansucrase from lemon peels, orange peels, and banana peels were reported to yield 0.65 Umg/ml, 0.58 Umg/ml, and 0.50 Umg/ml levan, respectively [5]. All these peels are rich in sucrose; it is proved from experiments that sucrose is considered as the direct carbon source for the levan using less water content [13]. ...
... In solid state fermentation (SSF), the growth of microorganisms is done on a solid support which makes it easy to handle [13] while in submerged fermentation microorganisms are grown in a liquid media, using free flowing liquid substrates [17]. SSF uses solid support for the production of various metabolites by using microorganisms, and it has great advantage over submerged fermentation because it has less water output, simple equipment for cultivation, capital investment is low, less energy requirement, no formation of foam, and lesser chances of bacterial contamination [13]. ...
Levan is a naturally occurring polysaccharide, found in many microorganisms and plants. The polymer is composed of fructose molecules. Food processing industries generate a lot of sugar rich waste consisting of different fruit peels that could be used for levan biosynthesis of levan. The present research presents levan production by Bacillus subtillis via solid state fermentation (SSF) using various sucrose-rich fruit peels. Peels of mango, banana, apple, and sugarcane bagasse were screened and maximum levan yield (1.87 mg/g) was found with mango peels after 24 h at pH 7 and 30 °C. Levan production from mango peel was further optimized for physical parameters using central composite design (CCD) of response surface methodology (RSM). The maximum levan yield (7.586 mg/g) from mango peels was achieved at 120 h incubation, 60% moisture content, 7 pH, and 32 °C. The functional groups of levan were confirmed by FTIR analysis. Sugar content was done by HPLC analysis, which confirmed levan has only fructose sugar. Mango peel could be a potential source for levan production.
... Strong absorption was observed at 1014 cm -1 , corresponding to stretching vibrations of the glycosidic bond (C-O-C). Finally, two typical signals were observed at 989 cm and 829 cm -1 , indicating the presence of the furanoid ring of the levan sugar units (Jathore et al., 2012;Dahech et al., 2013). ...
... In turn, FTIR characterization of levan showed signs of -OH vibration, C-H vibrational stretching, glycosidic bond elongations (C-O-C), and typical signs of furanosidic rings of sugar units (Dahech et al., 2013). All absorption signals were identical to other levans described in the literature (Jathore et al., 2012;Dahech et al., 2013;Xu et al., 2016). ...
The simultaneous production of microbial polymers levan and poly[3-hydroxybutyrate] (PHB), a type of polyhydroxyalkanoates, was investigated in this work. The study involved the fermentation of sucrose and molasses by H. smyrnensis AAD6T (BAE2 strain) to produce PHB (intracellular) and levan (extracellular). Both polymers were isolated and characterized by FTIR. Levan was also characterized by thin-layer chromatography (TLC) and viscosimetric analysis. The amount of biomass was 25 g until the end of fermentation. The PHB rate was 0.015 g in both media and the average PHB productivity was 6.0 x 10-4 g PHB/g biomass. The highest rate of levan was 9 g/L in the range of 72–80 h, in the molasses-based medium. The FTIR spectra showed specific signals for each of the polymers, such as the peak at 1700 for the carbonyl group of esters for the PHB and signals at 900 and 800, which are typical signals for levan fructose rings. Furthermore, acid hydrolysis of levan revealed that it was formed only by fructose, as confirmed by TLC With this study, H. smyrnensis AAD6T BAE2 co-produced PHB and levan using a low-cost carbon source, showing great potential in reducing biopolymer manufacturing costs.
... Levan, one of the most important EPSs, is formed of fructose molecules with a glucose residue at the end linked by β-2,6 glycosidic linkages and is generated with the process of a secreted levansucrase (EC 2.4.1.10) that directly transforms sucrose into the polymer [5] by microorganisms such as Zymomonas [6,7] Erwinia [8] Bacillus [9] Lactobacillus [10] Acetobacter [11] Gluconobacter [12] Streptococcus [13] and Pseudomonas [14,15]. Levan distinguishes itself from other water-soluble, biocompatible, and film-forming biopolymers due to an unusual mix of properties such as low intrinsic viscosity, health benefits, high adhesive strength, and the capacity to gel. ...
... RSM is generally used in biotechnology to improve fermentation medium conditions and other process parameters that are critical for the generation of a variety of microbial metabolites [25][26][27][28]. This approach has been used to successfully improve the yield of microbial polysaccharides as well as pullulan from Aureobasidium pullulans [29,30] glucan from Leuconostoc dextranicum [31] or levan from Zymomonas mobilis [6,7,32] and Pseudomonas fluorescens [14]. Species belonging to Halomonas genus are gram-negative, aerobic, moderately halophilic bacteria thrives at saline and hypersaline environments and also that synthesize high-value products such as levan, polyhydroxybutyrates (PHB) and ectoin etc. [33]. ...
Halophilic organisms are a novel attractive option as cell factories for the production of industrially valuable bioproducts. Halomonas elongata is the cell factory of choice for ectoine production, but its levan production has not been well researched. Based on this scientific motivation, in this study, we evaluated the chemical and biological properties of levan produced by the halophilic extremophile Halomonas elongata 153B (HeL). First, the central composite design was used to determine the optimal process variables for maximum levan biosynthesis. Then, the levan produced from HeL was purified, quantified, and chemically characterized with FTIR, ¹H-NMR, and GPC analyses. This was followed by antioxidant, anti-inflammatory, antibiofilm, and antimicrobial activity tests to assess its biological activities as well as a cytotoxcity assay. Maximum levan yields of 5.13 ± 0.38 g/L were achieved after dialysis at the optimum levels of process variables. The ¹H-NMR spectrum of HeL revealed characteristic signals. It showed a strong antioxidant activity of 67.88% and the best radical scavenger. At a concentration of 400 µg/mL, HeL showed the most anti-inflammatory efficacy. Also, at all indicated concentrations (250, 500, 750, and 1000 μg/mL) HeL, acted against biofilms formed by Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 6538, Pseudomonas aeruginosa ATCC 11778, Candida albicans ATCC 10231. Furthermore, HeL displayed antimicrobial activities against all strains tested. Finally, HeL showed high Cell viability in all dosages and no cytotoxicity was observed. In light of these results, HeL may have high potential in the medical, pharmaceutical and dermo-cosmetics industries.
Graphical Abstract
... -1 for the model, the adjustable frequency of the C-H group appeared for the two samples, respectively. Jathore et al. (2012) studied the characterization of Levan produced from Pseudomonas fluorescens and, based on the FTIR analysis, found that the structure of Levan was homologous to the standard Levan sample which the broad stretching peak of O-H stretching around 3319.26cm -1 , C-H vibration was observed at about 2935.48cm -1 , carbonyl C=O stretching noted at 1722.31cm -1 . ...
Few bacterial strains have been found to produce Levan, a valuable and versatile fructose homopolymer. Although Levan products have a variety of unique uses, their application and promotion were constrained by the cost and production capacity. Although Bacillus lichniformans MJ8 is a strain that can synthesize Levan, its production is insufficient for use. Halophilic bacteria are halotolerant microorganisms commonly found in natural environments. Bacillus lichniformans, the isolate sequence, was recorded in the NCBI database Accession number OM672244.1, a moderately halophilic bacterium isolated from soil of rhizosphere Sativa plant, Baghdad-Iraq, with a Levansucrase activity of 529.8 U/ml. In this study, we amplified the Levansucrase (SacB) gene isolated from Bacillus lichniformans MJ8, encoding the enzyme as a clone. It contained 1449 nucleotides, encoding a 482 amino-acid protein with a predicted 29 amino-acid signal peptide, the sequence of the gene Accession number ON81164.1.The levansucrase (SacB) gene was cloned to plasmid pTG19-T, and the recombinant plasmids were transferred to E. coli DH5 competent cells, and then the expression vector pet-28a+ into E. coli BL21 (DE3), For the production of the (SacB) protein, we induced by 1mM IPTG. The levansucrase activity after transformation had 989.1 U/ml. The Levan was identified after isolating and purification using chemical structures and was analyzed by Fourier Transform Infrared Spectroscopy (FTIR) and High-Performance Liquid Chromatography (HPLC) technique.
