A novel static cultivation of bacterial cellulose production from sugar beet molasses: Series static culture (SSC) system.

In bacterial cellulose (BC) production, we developed a new static cultivation system named series static culture (SSC) to eliminate air limitation problem encountered in conventional static culture (CSC). In SSC system, the fermentation broth at the bottom of BC pellicle produced in initial culture medium is transferred to the next empty sterile culture medium at the end of a certain fermentation period. This procedure was performed until BC production ceased. Fermentation experiments were carried out using Gluconacetobacter xylinus NRRL B-759 and sugar beet molasses at 30 °C and initial pH 5. Also, some quality parameters of produced BC pellicles were determined. Final pH at the stages of SSC system was higher that of the initial pH due to sugar content (sucrose) of molasses and microorganism used. Total BC production increased with increasing sugar concentration in SSC. As a result, an increase of 22.02 % in BC production was achieved using developed SSC. FT-IR spectra of all BC pellicles produced were typical spectra. The absorption bands at the relevant wavenumbers identify the mode of vibrations of the created chemical bonds arising at the BC surface such as OH, CH, H-O-H, C-O-C, and C-OH. XRD analyses showed that the crystallinity index values of BC obtained from CCS and SSC were high. The form of produced all BC pellicles is generally Cellulose I. Removal of surface moisture and depolymerisation of carbon skeleton were determined from TGA-DTA thermograms. SEM images showed that the BC samples produced had nano-sized cellulose fibrils which were aggregated in fermentation media containing molasses. Finally, the BC samples, especially in molasses media, having high mechanical strength and WHC were found.

Metabolic and Proteomic Analysis of Chlorella sorokiniana, Chloroidium saccharofilum, and Chlorella vulgaris Cells Cultured in Autotrophic, Photoheterotrophic, and Mixotrophic Cultivation Modes.

Chlorella is one of the most well-known microalgal genera, currently comprising approximately a hundred species of single-celled green algae according to the AlgaeBase. Strains of the genus Chlorella have the ability to metabolize both inorganic and organic carbon sources in various trophic modes and synthesize valuable metabolites that are widely used in many industries. The aim of this work was to investigate the impact of three trophic modes on the growth parameters, productivities of individual cell components, and biochemical composition of Chlorella sorokiniana, Chloroidium saccharofilum, and Chlorella vulgaris cells with special consideration of protein profiles detected by SDS-PAGE gel electrophoresis and two-dimensional gel electrophoresis with MALDI-TOF/TOF MS. Mixotrophic conditions with the use of an agro-industrial by-product stimulated the growth of all Chlorella species, which was confirmed by the highest specific growth rates and the shortest biomass doubling times. The mixotrophic cultivation of all Chlorella species yielded a high amount of protein-rich biomass with reduced contents of chlorophyll a, chlorophyll b, carotenoids, and carbohydrates. Additionally, this work provides the first information about the proteome of Chloroidium saccharofilum, Chlorella sorokiniana, and Chlorella vulgaris cells cultured in molasses supplementation conditions. The proteomic analysis of the three Chlorella species growing photoheterotrophically and mixotrophically showed increased accumulation of proteins involved in the cell energy metabolism and carbon uptake, photosynthesis process, and protein synthesis, as well as proteins involved in intracellular movements and chaperone proteins.

Long term performance of a pilot scale anaerobic membrane bioreactor treating beet molasses based industrial wastewater.

Baker's yeast industries (BYI) generate highly polluted effluents, especially vinasse from yeast separators, with very high chemical oxygen demand (COD), nitrogen, sulphate and salts, mainly potassium and calcium. Anaerobic treatment is the most commonly applied method for treating BYI wastewaters. However, it is quite challenging to obtain a high performance due to the difficulties in biomass retention. Moreover, it does not provide compliance with COD and color discharge limits when used as a sole treatment process. In this context, a pilot scale anaerobic membrane bioreactor, which provides excellent biomass retention, was operated to investigate its treatment performance for vinasse from a BYI. The reactor achieved a COD removal between 48% and 92% up to a volumetric load of 10 kg COD m3 d-1. A specific methane production of 0.37 m3 CH4 kg-1 CODremoved was observed in the study. On the other hand, passage of inert organic compounds through membrane deteriorated permeate quality and treatment efficiency. High alkalinity and pH led to the accumulation of calcium precipitates, which reduced volatile solids fraction of sludge and biomass activity in the reactor. The present study showed the operational challenges and potential drawbacks of AnMBR systems for BYI wastewater treatment. The experience gained in the pilot system can be utilized in the design and operation of full scale AnMBRs for high strength industrial effluents.

Preparation and characterization of cellulose nanocrystals from bacterial cellulose produced in sugar beet molasses and cheese whey media.

Bacterial cellulose (BC) is gaining considerable attention due to its unique physicochemical and mechanical properties. In this study, BC production by Gluconacetobacter xylinus PTCC 1734 in sugar beet molasses, cheese whey and standard Hestrin-Schramm (HS) media was evaluated. The synthesized BC was hydrolyzed by sulfuric acid to prepare bacterial cellulose nanocrystals (BCNC). The results showed that treated sugar beet molasses led to the highest BC concentration and productivity, followed by treated cheese whey. Structural analysis of BC and BCNC was carried out by Fourier Transform Infrared (FTIR) spectroscopy. The crystallinity index of the BCNC determined by X-ray diffraction (XRD) was higher than BC. The morphological analysis carried out by FE-SEM showed that microfibrils diameter decreases with acid treatment. TEM images confirmed the formation of rod like cellulose nanocrystals having an average diameter and length of 25 ±â€¯5 and 306 ±â€¯112 nm, respectively. In conclusion, food industrial byproducts can be used as cost-effective culture media to produce BC for large-scale industrial production and isolated cellulose nanocrystals are useful in the fabrication of bio-nanocomposite films for food packaging applications.

Engineered E. coli W enables efficient 2,3-butanediol production from glucose and sugar beet molasses using defined minimal medium as economic basis.

BACKGROUND: Efficient microbial production of chemicals is often hindered by the cytotoxicity of the products or by the pathogenicity of the host strains. Hence 2,3-butanediol, an important drop-in chemical, is an interesting alternative target molecule for microbial synthesis since it is non-cytotoxic. Metabolic engineering of non-pathogenic and industrially relevant microorganisms, such as Escherichia coli, have already yielded in promising 2,3-butanediol titers showing the potential of microbial synthesis of 2,3-butanediol. However, current microbial 2,3-butanediol production processes often rely on yeast extract as expensive additive, rendering these processes infeasible for industrial production. RESULTS: The aim of this study was to develop an efficient 2,3-butanediol production process with E. coli operating on the premise of using cost-effective medium without complex supplements, considering second generation feedstocks. Different gene donors and promoter fine-tuning allowed for construction of a potent E. coli strain for the production of 2,3-butanediol as important drop-in chemical. Pulsed fed-batch cultivations of E. coli W using microaerobic conditions showed high diol productivity of 4.5 g l-1 h-1. Optimizing oxygen supply and elimination of acetoin and by-product formation improved the 2,3-butanediol titer to 68 g l-1, 76% of the theoretical maximum yield, however, at the expense of productivity. Sugar beet molasses was tested as a potential substrate for industrial production of chemicals. Pulsed fed-batch cultivations produced 56 g l-1 2,3-butanediol, underlining the great potential of E. coli W as production organism for high value-added chemicals. CONCLUSION: A potent 2,3-butanediol producing E. coli strain was generated by considering promoter fine-tuning to balance cell fitness and production capacity. For the first time, 2,3-butanediol production was achieved with promising titer, rate and yield and no acetoin formation from glucose in pulsed fed-batch cultivations using chemically defined medium without complex hydrolysates. Furthermore, versatility of E. coli W as production host was demonstrated by efficiently converting sucrose from sugar beet molasses into 2,3-butanediol.

Single-stage photofermentative biohydrogen production from sugar beet molasses by different purple non-sulfur bacteria.

Biohydrogen production via fermentative routes offers considerable advantages in waste recycling and sustainable energy production. This can be realized by single-stage dark or photofermentative processes, or by a two-stage integrated process; the latter offering the higher production yields due to complete conversion of sugar substrates into H2 and CO2. However, problems arising from the integration of these two processes limit its scale-up and implementation. Hence, high efficiency one-step fermentative biohydrogen production processes from sugar-rich wastes are preferable. In this study, different strains of purple non-sulfur bacteria were investigated for their biohydrogen production capacity on pure sucrose and sugar beet molasses, and the feasibility of single-stage photofermentative biohydrogen production was evaluated. A single-stage photofermentation process was carried out using four different strains of purple non-sulfur bacteria (Rhodobacter capsulatus DSM 1710, R. capsulatus YO3, Rhodobacter sphaeroides O.U.001, and Rhodopseudomonas palustris DSM 127) on different initial sucrose concentrations. The highest hydrogen yield obtained was 10.5 mol H2/mol of sucrose and the maximum hydrogen productivity was 0.78 mmol/L h by Rp. palustris on 5 mM sucrose. A hydrogen yield of 19 mol H2/mol sucrose, which represents 79% of theoretical yield, and a maximum hydrogen productivity of 0.55 mmol/L h were obtained by Rp. palustris from sugar beet molasses. The yield was comparable to those values obtained in two-stage processes. The present study demonstrates that single-stage photofermentation using purple non-sulfur bacteria on sucrose-based wastes is promising.

Cytotoxicity and Apoptotic Effects of Polyphenols from Sugar Beet Molasses on Colon Carcinoma Cells in Vitro.

Three polyphenols were isolated and purified from sugar beet molasses by ultrasonic-aid extraction and various chromatographic techniques, and their structures were elucidated by spectral analysis. Cytotoxicity and the molecular mechanism were measured by methyl thiazolyl tetrazolium (MTT) assay, flow cytometry, caspase-3 activity assay and Western blot assay. The results showed that gallic acid, cyanidin-3-O-glucoside chloride and epicatechin have cytotoxicity to the human colon, hepatocellular and breast cancer cells. Cyanidin-3-O-glucoside chloride showed its cytotoxicity against various tumor cell lines, particularly against colon cancer Caco-2 cells with half maximal inhibitory concentration (IC50) value of 23.21 ± 0.14 µg/mL in vitro. Cyanidin-3-O-glucoside chloride may be a potential candidate for the treatment of colon cancer. In the mechanism study, cyanidin-3-O-glucoside chloride increased the ratio of cell cycle at G0/G1 phase and reduced cyclin D1 expression on Caco-2 cells. Cyanidin-3-O-glucoside chloride decreased mutant p21 expression, and increased the ratio of Bax/Bcl-2 and the activation of caspase-3 to induce apoptosis.

Sugar beet molasses as an ingredient to enhance the nutritional and functional properties of gluten-free cookies.

Sugar beet molasses is a raw material with high potential to be a functional ingredient in baked goods. This paper investigated the nutritional and functional properties of gluten-free cookies enriched with sugar beet molasses. At all enrichment levels and forms tested (liquid and dry), the addition of beet molasses improved the micronutrient pattern and antioxidative status of gluten-free cookies. The cookies prepared with molasses were significantly higher in potassium, magnesium, calcium, iron, betaine, total phenolics and DPPH radical scavenging abilities. Molasses contributed to wider spectra of phenolic compounds. The dominating phenolic compounds in the molasses-enriched cookies were catechin, ferulic, syringic and vanillic acid. Molasses also contributed to the presence of p-hydroxybenzoic acid in the cookies. Addition of molasses increased the content of hydroxymethyfurfural in the cookies, but not above values commonly reported for this product type. Molasses addition improved the overall acceptance of gluten-free cookies up to 30% enrichment level.

Enhancing L-Lysine Production of Beet Molasses by Engineered Escherichia coli Using an In Situ Pretreatment Method.

Reducing the viscosity of molasses environmentally and selectively removing the harmful ingredients for microbes are the keys to promoting the bioavailability of molasses. A simple and environmental in situ pretreatment method integrating surfactants and alkali was developed to reduce the viscosity of molasses prior to L-lysine production using Escherichia coli ZY0217. Adding activated carbon and modified orange peel based on the in situ pretreatment process effectively removed pigments and excessive zinc in the molasses and also significantly increased the cell growth and L-lysine yield from E. coli ZY0217. The experimental results showed that a mixture of secondary alkane sulfonate, an anionic surfactant, and HodagCB-6, a non-ionic surfactant, effectively reduced the viscosity of the molasses more so than any single surfactant. When the surfactant mixture was added at a concentration of 0.04 g/L to the molasses, the ω value was 0.4, and when ammonia was added at 0.6 %, the lowest viscosity of 705 mPa · s was obtained. Further, 91.5 % of the color and 86.68 % of the original levels of zinc were removed using an activated carbon and modified orange peel treatment on the molasses with the lowest viscosity, which further promoted cell growth and L-lysine production. In the fed-batch cultivation process, the L-lysine concentration achieved using a constant-speed feeding strategy was 45.89 g/L, with an L-lysine yield of 27.18 %, whereas the L-lysine yield from untreated molasses was only 10.13 %. The increase in L-lysine yield was related to the reduced viscosity and the detoxification of the molasses. Lastly, the pretreatment was found to significantly enhance the conversion of sugars in the molasses to L-lysine.

Structural analysis of novel kestose isomers isolated from sugar beet molasses.

Eight kestose isomers were isolated from sugar beet molasses by carbon-Celite column chromatography and HPLC. GC-FID and GC-MS analyses of methyl derivatives, MALD-TOF-MS measurements and NMR spectra were used to confirm the structural characteristics of the isomers. The (1)H and (13)C NMR signals of each isomer saccharide were assigned using COSY, E-HSQC, HSQC-TOCSY, HMBC and H2BC techniques. These kestose isomers were identified as α-D-fructofuranosyl-(2- > 2)-α-D-glucopyranosyl-(1 < ->2)-ß-D-fructofuranoside, α-D-fructofuranosyl-(2- > 3)-ß-D-fructofuranosyl-(2 < ->1)-α-D-glucopyranoside, α-D-fructofuranosyl-(2- > 4)-ß-D-fructofuranosyl-(2 < ->1)-α-D-glucopyranoside, ß-D-fructofuranosyl-(2- > 4)-ß-D-fructofuranosyl-(2 < ->1)-α-D-glucopyranoside, ß-D-fructofuranosyl-(2- > 3)-α-D-glucopyranosyl-(1 < ->2)-ß-D-fructofuranoside, α-D-fructofuranosyl-(2- > 1)-ß-D-fructofuranosyl-(2 < ->1)-α-D-glucopyranoside, α-D-fructofuranosyl-(2- > 6)-α-D-glucopyranosyl-(1 < ->2)-ß-D-fructofuranoside, and α-D-fructofuranosyl-(2- > 6)-ß-D-fructofuranosyl-(2 < ->1)-α-D-glucopyranoside. The former five compounds are novel saccharides.

Structural confirmation of novel oligosaccharides isolated from sugar beet molasses.

Eleven oligosaccharides were isolated from sugar beet molasses using carbon-Celite column chromatography and HPLC. The constituent sugars and linkage positions were determined using methylation analysis, MALDI-TOF-MS, and NMR measurements. The configurations of isolated oligosaccharides were confirmed based on detailed NMR analysis. Based on our results, three of the 11 oligosaccharides were novel.

A Novel Production Method for High-Fructose Glucose Syrup from Sucrose-Containing Biomass by a Newly Isolated Strain of Osmotolerant Meyerozyma guilliermondii.

One osmotolerant strain from among 44 yeast isolates was selected based on its growth abilities in media containing high concentrations of sucrose. This selected strain, named SKENNY, was identified as Meyerozyma guilliermondii by sequencing the internal transcribed spacer regions and partial D1/D2 large-subunit domains of the 26S ribosomal RNA. SK-ENNY was utilized to produce high-fructose glucose syrup (HFGS) from sucrose-containing biomass. Conversion rates to HFGS from 310-610 g/l of pure sucrose and from 75-310 g/l of sugar beet molasses were 73.5-94.1% and 76.2-91.1%, respectively. In the syrups produced, fructose yields were 89.4-100% and 96.5-100% and glucose yields were 57.6-82.5% and 55.3-79.5% of the theoretical values for pure sucrose and molasses sugars, respectively. This is the first report of employing M. guilliermondii for production of HFGS from sucrose-containing biomass.

Optimisation of ultrasonic-assisted extraction of phenolic compounds, antioxidants, and anthocyanins from sugar beet molasses.

Response surface methodology was used to optimise experimental conditions for ultrasonic-assisted extraction (UAE) of functional components from sugar beet molasses. The central composite design (CCD) was used for the optimisation of extraction parameters in terms of total phenolic contents, antioxidant activities and anthocyanins. Result suggested the optimal conditions obtained by RSM for UAE from sugar beet molasses were as follows: HCl concentration 1.55-1.72 mol/L, ethanol concentration 57-63% (v/v), extraction temperature 41-48 °C, and extraction time 66-73 min. In the optimal conditions, the experimental total phenolic contents were 17.36 mg GAE/100mL, antioxidant activity was 16.66 mg TE/g, and total anthocyanins were 31.81 mg/100g of the sugar beet molasses extract, which were well matched the predicted values. Teen compounds, i.e. gallic acid, vanillin, hydroxybenzoic acid, syringic acid, cyanidin-3-O-rutinoside, cyanidin-3-O-glucoside, catechin, delphinidin-3-O-rutinoside, delphinidin-3-O-glucuronide and ferulic acid were determined by HPLC-DAD-MS/MS in sugar beet molasses.

Improved ethanol production from cheese whey, whey powder, and sugar beet molasses by "Vitreoscilla hemoglobin expressing" Escherichia coli.

This work investigated the improvement of ethanol production by engineered ethanologenic Escherichia coli to express the hemoglobin from the bacterium Vitreoscilla (VHb). Ethanologenic E. coli strain FBR5 and FBR5 transformed with the VHb gene in two constructs (strains TS3 and TS4) were grown in cheese whey (CW) medium at small and large scales, at both high and low aeration, or with whey powder (WP) or sugar beet molasses hydrolysate (SBMH) media at large scale and low aeration. Culture pH, cell growth, VHb levels, and ethanol production were evaluated after 48 h. VHb expression in TS3 and TS4 enhanced their ethanol production in CW (21-419%), in WP (17-362%), or in SBMH (48-118%) media. This work extends the findings that "VHb technology" may be useful for improving the production of ethanol from waste and byproducts of various sources.

Evaluation of baker's yeast strains exhibiting significant growth on Japanese beet molasses and compound analysis of the molasses types.

Cane molasses, most of which is imported, is used as a raw material for production of baker's yeast (Saccharomyces cerevisiae) in Japan. On the other hand, beet molasses is scarcely used for this purpose, but it can be of great advantage to cane molasses because it is domestically produced in relatively high amounts as a by-product of beet sugar processing. However, the yield of baker's yeast is sometimes low with Japanese beet molasses compared to imported cane molasses. For the production of baker's yeast with Japanese beet molasses, we evaluated S. cerevisiae strains, including industrial and laboratory strains, to group them according to the growth profile on beet and cane molasses. To discuss the factors affecting growth, we further analyzed the major compounds in both types of molasses. Beet molasses seems to contain compounds that promote the growth of beet molasses-favoring strains rather than inhibit the growth of cane molasses-favoring strains. It was assumed that α-amino acid was one of the growth promotion factors for beet molasses-favoring strains.