Welan Gum Production Research Articles

Effect of an inorganic nitrogen source (NH4)2SO4 on the production of welan gum from Sphingomonas sp. mutant obtained through UV-ARTP compound mutagenesis

As one of the most expensive extracellular polysaccharides, welan gum is widely used in biomedicine, food products, and petroleum because of its unique structure and excellent rheological properties. To reduce the cost of welan gum fermentation, together with (NH4)2SO4, which served as the sole nitrogen source, a high-welan-gum-producing mutant, B-8, screened through UV-ARTP compound mutagenesis was used. Under optimum conditions (C:N ratio 25:1, sucrose 50 g/L, (NH4)2SO4 4 g/L, and adding 8 mM NaCl at 32 h fermentation), the yield of welan gum and sucrose conversion were 18.86 g/L and 0.38 g/g, respectively, which were 98.95% and 137.50% higher than those achieved with the parent strain FM01, respectively. After the same treatment process, IN-welan (obtained with (NH4)2SO4) consumed less 95% ethanol, had higher molecular weight, and exhibited better rheological properties than ON-welan (obtained with beef extract). Transcriptome analysis revealed that (NH4)2SO4 could affect the synthetic pathway and monosaccharide content of welan gum by increasing bacterial chemotaxis and the availability of key intermediates. The fermentation performance of Sphingomonas sp. mutants could further be improved by providing several target genes to the mutants through metabolic engineering.

Sustainable production and characterization of medium-molecular weight welan gum produced by a Sphingomonas sp. RW

Welan gum is widely used in food, concrete, and oil recovery for its excellent stability and unique rheological properties. This study describes an engineered Sphingomonas sp. RW strain deficient in the lyase gene (welR) and outlines its application in an industrial by-product fermentation strategy to produce welan gum. The yield of the engineered strain was enhanced to 25.11 ± 0.05 g/L, whereas the broth viscosity (2110 mPas) increased by 281.81%. The welan gum produced by Sphingomonas sp. RW was named medium-molecular weight welan gum (MMWG, ~350 kDa). FT-IR and monosaccharide composition analysis revealed that the MMWG was composed of l-mannose, l-rhamnose, glucuronic acid, and glucose (with mole ratios of 1:4.46:1.26:2.82). Significantly, MMWG had favorable hydroxyl radical scavenging activity and emulsifying capacity. These results provide a strategy for cost-effective welan gum production and enrich the properties of welan gum with different molecular weight.

Effect of NaCl addition on the production of welan gum with the UV mutant of Sphingomonas sp

Because of its excellent stability, non-toxicity, biodegradability and unique rheology, welan gum can be widely used in various fields, such as petroleum, biomedicine and food products. In this study, a high-yield mutant strain FM01-S09 was screened through two rounds of UV mutagenesis. Remarkably, the production of welan gum could be further increased by adding 4 mM NaCl at 32 h fermentation, reaching 30.12 ± 0.25 g/L (28.66% higher than no adding), and the NaCl-WG solution had stronger structural, impact resistance, and temperature resistance than H2O2-WG and WG solutions. Furthermore, the mechanism by which NaCl promotes welan gum synthesis was also investigated. It was found that cell membrane characteristics, intracellular microenvironment makeup, and key enzyme gene expression levels were significantly altered in different fermentation stages. Therefore, the addition of NaCl could effectively promote the growth and fermentation performance of Sphingomonas sp., providing a novel strategy for cost-effective welan gum production.

A strategy for the synthesis of low-molecular-weight welan gum by eliminating capsule form of Sphingomonas strains

Welan gum is widely used in food, concrete additives, and oil recovery. Here we changed the capsule form of Sphingomonas strains by knocked out the sortase gene (srtW). The obtained welan gum was mainly composed of mannose, glucose, rhamnose, and glucuronic acid at a molar ratio of 4.0:5.8:1.6:1, respectively. Meanwhile, the molecular weight of welan gum decreased sharply (about 68 kDa). Moreover, the low molecular weight (LMW) welan gum was characterized by FT-IR and NMR spectroscopy. The rheological results revealed that the LMW welan gum solution is a pseudoplastic fluid with a lower apparent viscosity. Furthermore, the oscillation test illustrated stable dynamic viscoelasticity within the temperature range of 5–68 °C and frequency range of 0.01–15 rad/s. To the best of our knowledge, this is the first report of LMW welan gum production and characterization. These results provide references for LMW welan gum applications, and likely applicable for other biopolymers production.

Production of welan gum from cane molasses by Sphingomonas sp. FM01

As an extracellular exopolysaccharide, welan gum could serve as a suspending agent, stabilizer, emulsifier and thickener with characteristics that include bio-adhesion, non-toxicity, biocompatibility and biodegradability. In this study, Sphingomonas sp. FM01 was applied to effectively produce welan gum using cane molasses as the carbon source. After pretreatment with phosphoric acid and polyacrylamide, 60 g/L cane molasses combined with 6.0 g/L beef extract, 3 g/L KH2PO4, 0.5 g/L MgSO4, 15 mmol/L H2O2 and 4 mg/L niacin significantly improved the fermentation performance of Sphingomonas sp. FM01, increasing the production of welan gum to 37.65 g/L. Investigation of the rheological behavior of the welan gum obtained from the molasses-welan gum mixture showed it had an acceptable molecular weight and similar rheological properties and better viscosity stability compared to that obtained from sucrose, indicating that cane molasses could be explored as a suitable and inexpensive substrate for cost-effective welan gum production.

Significantly improved production of Welan gum by Sphingomonas sp. WG through a novel quorum-sensing-interfering dipeptide cyclo(L-Pro-L-Phe)

A variety of physiological functions such as exopolysaccharide synthesis, antibiotic production, and primary metabolism are tightly controlled by quorum sensing in microorganisms. In this study, a marine-derived bacterium Sphingomonas sp. WG was found to possess a cyclo(L-Pro-L-Phe)-mediated quorum sensing mechanism. The cyclo(L-Pro-L-Phe) produced by Sphingomonas sp. WG functions as a signalling molecule in this quorum sensing system. It is the first attempt to characterize cyclic dipeptides as quorum sensing signalling molecules in Sphingomonas sp. and the results effectively make the classical quorum sensing theory more perfect. Furthermore, the supplementation with isolated cyclo(L-Pro-L-Phe) resulted in a 15% increase in the production of welan gum secreted by Sphingomonas sp. WG in the submerged fermentation. The data presented in this study will provide evidences for exploring the role of cyclic dipeptides in regulating the production of welan gum.

Optimal experimental conditions for Welan gum production by support vector regression and adaptive genetic algorithm.

Welan gum is a kind of novel microbial polysaccharide, which is widely produced during the process of microbial growth and metabolism in different external conditions. Welan gum can be used as the thickener, suspending agent, emulsifier, stabilizer, lubricant, film-forming agent and adhesive usage in agriculture. In recent years, finding optimal experimental conditions to maximize the production is paid growing attentions. In this work, a hybrid computational method is proposed to optimize experimental conditions for producing Welan gum with data collected from experiments records. Support Vector Regression (SVR) is used to model the relationship between Welan gum production and experimental conditions, and then adaptive Genetic Algorithm (AGA, for short) is applied to search optimized experimental conditions. As results, a mathematic model of predicting production of Welan gum from experimental conditions is obtained, which achieves accuracy rate 88.36%. As well, a class of optimized experimental conditions is predicted for producing Welan gum 31.65g/L. Comparing the best result in chemical experiment 30.63g/L, the predicted production improves it by 3.3%. The results provide potential optimal experimental conditions to improve the production of Welan gum.

Two-step economical welan gum production by Sphingomonas sp. HT-1 from sugar industrial by-products

A two-step fermentation strategy using glucose mother liquor (GML) for cell growth and xylose mother liquor (XML) for welan gum synthesis was used to alleviate uneconomic welan gum fermentation. This study revealed: (1) optimal initial GML concentration was 11.7g/L (10g/L sugars contained); (2) optimal XML feeding strategy was pseudo-exponential fed-batch and feeding time was 12thh–54thh, amounting to 25.7g/L XML (20g/L sugars contained); and (3) in a 7.5-L bioreactor, welan gum concentration was 22.68±0.50g/L and its yield reached 0.756g/g sugars with trace residual sugars. Compared with the cost of batch fermentation using glucose as sole carbon source, the final carbon source costs decreased by 61.40% and the welan gum yield increased by 50%. GML and XML can be used as inexpensive carbon sources for welan gum production with higher yield, giving them industrial application potential to produce value-added chemicals.

Different nitrogen sources change the transcriptome of welan gum-producing strain Sphingomonas sp. ATCC 31555.

To reveal effects of different nitrogen sources on the expressions and functions of genes in Sphingomonas sp. ATCC 31555, it was cultivated in medium containing inorganic nitrogen (IN), organic nitrogen (ON), or inorganic-organic combined nitrogen (CN). Welan gum production and bacterial biomass were determined, and RNA sequencing (RNA-seq) was performed. Differentially expressed genes (DEGs) between the different ATCC 31555 groups were identified, and their functions were analyzed. Welan gum production and bacterial biomass were significantly higher in the ON and CN groups compared with those in the IN group. RNA-seq produced 660 unigenes, among which 488, 731, and 844 DEGs were identified between the IN vs. ON, IN vs. CN, and ON vs. CN groups, respectively. All the DEGs were related significantly to metabolic process and signal transduction. DEGs between the IN vs. CN and ON vs. CN groups were potentially associated with bacterial chemotaxis. Real-time PCR confirmed the expressions of selected DEGs. Organic nitrogen led to higher bacterial biomass and welan gum production than inorganic nitrogen, which might reflect differences in gene expression associated with metabolic process, signal transduction, and bacterial chemotaxis induced by different nitrogen sources.

Bioconversion of Welan Gum from Kitchen Waste by a Two-Step Enzymatic Hydrolysis Pretreatment.

Kitchen waste (KW) is a worldwide issue, which can lead to environment pollution. Nevertheless, it is also a low-cost and sustainable resource for bio-production. Meanwhile, welan gum (WG) is one kind of the most important exopolysaccharide but with high material cost. The aim of this study was to adopt two-step enzymatic hydrolysis to improve the release and recovery of both sugar and protein in KW for subsequent WG production. As the results, the recovery rates of sugar and protein reached 81.07 and 77.38%, which were both satisfactory. After the conditions optimized in flasks, the welan fermentation was conducted in a 5-L fermentor, and the WG yield, utilization rates of reducing sugar and KDN, respectively, reached 5.57gL-1, 94.25% and 61.96%. Moreover, the kinetic analyses demonstrated that the WG fermentation in KWH was a partly growth-associated process. The KW was successfully treated by fermentation for the bioconversion to WG.

Production and Rheological Properties of Welan Gum Produced by Sphingomonas sp. ATCC 31555 with Different Nitrogen Sources

This study aimed to investigate the effect of nitrogen sources on the production and rheological properties of welan gum produced by Sphingomonas sp. ATCC 31555. Six different nitrogen sources were used for ATCC 31555 fermentation, and 2 of these were further analyzed due to their more positive influence on welan gum production and bacterial biomass. Bacterial biomass, welan gum yield, welan viscosity, molecular weight, monosaccharide composition, acyl content, and welan structure were analyzed. Welan gum production and the biomass concentration of ATCC 31555 were higher in media containing NaNO₃ and beef extract. Welan viscosity decreased at higher temperatures of 30-90°C, and it increased with a higher welan concentration. In the media containing NaNO₃ (3 g·L^-1), welan viscosity was higher at 30-70°C and a welan solution concentration of 6-10 g·L^-1. With a reduced NaNO₃ concentration, the molecular weight of welan gum and the molar ratio of mannose decreased, but the molar ratio of glucuronic acid increased. With different nitrogen sources, the acetyl content of welan gum differed but its structure was similar. NaNO₃ and beef extract facilitated welan production. A reduced NaNO₃ concentration promoted welan viscosity.

Biological production of welan gum

Biological production of welan gum

Enhancement of welan gum production in Sphingomonas sp. HT-1 via heterologous expression of Vitreoscilla hemoglobin gene

Welan gum is a microbial polysaccharide produced by Sphingomonas sp. Its production is limited by the dissolved oxygen levels in the highly viscous fermentation. A strategy of heterologous expression of the Vitreoscilla hemoglobin gene in Sphingomonas sp. HT-1 was investigated to alleviate oxygen limitation and improve the yield of welan gum. Ultimately, the welan gum production increased from 25.3g/L to 34.6g/L, whereas the rheological behavior of welan gum solutions remained virtually unchanged. The transcriptional levels of the key genes in the electron transfer chain, TCA cycle and welan gum synthesis pathway, as well as ATP level revealed that the VHb expression in Sphingomonas sp. HT-1 enhanced welan gum biosynthesis by improving respiration and ATP supply. This study would pave the genetic manipulation way for enhancing welan gum yield, and it’s also of great importance for the industrial applications of welan gum under harsh conditions.

Improved production of carotenoid-free welan gum in a genetic-engineered Alcaligenes sp. ATCC31555

To improve the production of welan gum and obtain a carotenoid-free strain while reducing the fermentation and post-treatment costs. The vitreoscilla globin (vgb) gene combined with the β-galactosidase (lacZ) promoter was inserted into the phytoene synthase (crtB) gene region of the chromosome in Alcaligenes sp. ATCC31555. When the recombinant strain was grown in a 5l fermentor, welan gum was produced at 24±0.4gl(-1) compared to 21g±0.4gl(-1) in the wild type. Furthermore, the carotenoid-free welan gum produced using Alcaligenes sp. ATCC31555 VHb strain was less expensive with improved properties. Alcaligenes sp. ATCC31555 VHb strain was a better neutral welan-producing strain with a higher production than the wild-type strain.

Draft Genome Sequence of Sphingomonas sp. WG, a Welan Gum-Producing Strain.

We report the draft genome sequence of Sphingomonas sp. WG, a high welan gum-producing strain with a yield of 33 g/L. The core of wel cluster for welan gum biosynthesis contained 24 coding sequences in the genome, which will provide the genetic information on welan gum production.

Improved welan gum production by Alcaligenes sp. ATCC31555 from pretreated cane molasses

Welan gum production by Alcaligenes sp. ATCC31555 from cane molasses was studied in batch fermentation to reduce production costs and enhance gum production. The pretreatment of cane molasses, agitation speed and the addition of supplements were investigated to optimize the process. Sulfuric acid hydrolysis was found to be the optimal pretreatment, resulting in a maximum gum concentration of 33.5g/L, which is 50.0% higher than those obtained from the molasses’ mother liquor. Agitation at 600rpm at 30°C and addition of 10% n-dodecane following fermentation for 36h increased the maximum gum production up to 41.0±1.41g/L, which is 49.1% higher than the greatest welan gum concentration in the literature so far. The welan gum product showed an acceptable molecular weight, similar rheological properties and better thermal stability to that obtained from glucose. These results indicate that cane molasses may be a suitable and inexpensive substrate for cost-effective industrial-scale welan gum production.

Enhanced welan gum production using cane molasses as substrate by Alcaligenes sp. ATCC31555

Enhanced welan gum production using cane molasses as substrate by Alcaligenes sp. ATCC31555

Screening and characterization of Sphingomonas sp. mutant for welan gum biosynthesis at an elevated temperature

The optimal temperature for the microbial polysaccharide fermentation is no higher than 30°C, which is economically undesirable due to additional cooling cost. To solve this problem in the case of welan gum production, we obtained the high-temperature-tolerant-producing strain, Sphingomonas sp. HT-1 by atmospheric and room-temperature plasma-induced mutation. Using HT-1, we obtained a concentration and 1% aqueous viscosity of 26.8±0.34g/L and 3.50±0.05Pas at a comparatively higher optimal temperature (37°C). HT-1 was further characterized to understand the mechanism by which these properties are improved. Results indicated that high yield could be attributed to the following: (1) enhanced intracellular synthesis, demonstrated by an increase in the activities of key enzymes, and (2) accelerated cross-membrane substrate uptake and product secretion, indicated by improved membrane fluidity and permeability. Temperature tolerance could be attributed to the overexpression of the investigated heat shock proteins and oxidative stress proteins.

Improvement of welan gum production and redistribution of metabolic flux under pH control process in Alcaligenes sp. CGMCC2428

Batch fermentations of welan gum from Alcaligenes sp. CGMCC2428 at pH values of 5.5 ∼ 7.0 were studied. Based on the kinetic analysis, a pH control process for improving welan production was developed. By maintaining the culture pH at 7.0, the process significantly improved the maximal welan concentration and productivity to reach 25.1 ± 0.65 g/L and 0.42 ± 0.003 g/L/h, respectively, compared with those in native pH processes where pH value would decrease from 7.0 to 5.1 (18.5 ± 0.72 g/L and 0.28 ± 0.002 g/L/h). This improvement may be due to the increased metabolic flux of glucose-1-phosphate to welan induced by pH control process. Furthermore, scale-up fermentation under controlled pH was implemented using 300-L fermentor. The highest welan concentration of 27.5 ± 0.97 g/L was obtained. This simplified process proved effective in industry fermentation for high welan production.

Optimization of exopolysaccharide welan gum production by Alcaligenes sp. CGMCC2428 with Tween-40 using response surface methodology

Abstract The effect of additives on welan gum production produced by fermentation with Alcaligenes sp. CGMCC2428 was studied. Tween-40 was the best additive for improving welan gum production and welan gum displayed better rheological properties than that obtained by control fermentation without additives. Response surface methodology was employed to optimize the culture conditions for welan gum production in the shake flask culture, including Tween-40 concentration, pH and culture temperature. The optimal conditions were determined as follows: Tween-40 concentration 0.94 g/l, pH 6.9 and temperature 29.6 °C. The corresponding experimental concentration of welan gum was 23.62 ± 0.60 g/l, which was agreed closely with the predicted value (23.48 g/l). Validation experiments were also carried out to prove the adequacy and the accuracy of the model obtained. The welan gum fermentation in a 7.5 l bioreactor reached 24.90 ± 0.68 g/l.