The limited availability of high-cost nucleotide sugars is a significant constraint on the application of their downstream products (glycosides and prebiotics) in the food or pharmaceutical industry. ...To better solve the problem, this study presented a one-pot approach for the biosynthesis of UDP-Gal using a thermophilic multienzyme system consisting of GalK, UGPase, and PPase. Under optimal conditions, a 2 h reaction resulted in a UTP conversion rate of 87.4%. In a fed-batch reaction with Gal/ATP = 20 mM:10 mM, UDP-Gal accumulated to 33.76 mM with a space-time yield (STY) of 6.36 g/L·h–1 after the second feeding. In repetitive batch synthesis, the average yield of UDP-Gal over 8 cycles reached 10.80 g/L with a very low biocatalyst loading of 0.002 genzymes/gproduct. Interestingly, Galk (Tth0595) could synthesize Gal-1P using ADP as a donor of phosphate groups, which had never been reported before. This approach possessed the benefits of high synthesis efficiency, low cost, and superior reaction system stability, and it provided new insights into the rapid one-pot synthesis of UDP-Gal and high-value glycosidic compounds.
•Fermentation with continuous feeding SASL improved cellulase production.•The maximum FPase reached 17.66U/mL using a multi-stage feeding strategy.•Comparative proteomic analysis showed that SASL ...induced cellulases synthesis.•The SASL-induced cellulase could hydrolyze ammonium sulphite pulp efficiently.
Spent ammonium sulphite liquor (SASL) is the main effluent from the ammonium sulphite pulping process, and contains amounts of lignocellulosic oligomers, monosaccharides and ammonium salts. The effect of continuous SASL-feeding on cellulase production by Penicillium oxalicum was studied. With a rate-varying feeding strategy, the maximal filter paper enzyme (FPase) activity reached 17.66U/mL at 144h, and the specific FPase activity increased from 1.74U/mg (without SASL) to 2.40U/mg. Accordingly, the glucan hydrolysis conversion of delignified corn cob residue by the enzymes from continuous SASL-feeding fermentation was significantly higher than that without SASL at equal protein loadings. Comparative proteomic analysis demonstrated that the proteins involved in lignocellulose degradation were specifically up-regulated in the crude cellulase with SASL-feeding. The obtained crude enzyme was efficient in the hydrolysis of pulping products, with a glucan conversion of 81.87% achieved after 72h saccharification of ammonium sulphite pulp.
Co‐cultures for simultaneous production of ethanol and xylitol were studied under different operation bioreactor modes using Candida tropicalis IEC5‐ITV and Saccharomyces cerevisiae ITV01‐RD in a ...simulated medium of sugarcane bagasse hydrolyzates. Xylitol and ethanol tolerance by S. cerevisiae and C. tropicalis, respectively, was evaluated. The results showed that C. tropicalis was sensitive to ethanol concentrations up to 30 g/L, while xylitol had no effect on S. cerevisiae viability and metabolism. The best condition found for simultaneous culture was S. cerevisiae co‐culture and C. tropicalis sequential cultivation at 24 h. Under these conditions, productivity and yield for ethanol were QEtOH = 0.72 g L−1 h−1 and YEtOH/s = 0.37 g/g, and for xylitol, QXylOH = 0.10 g L−1 h−1 and YXylOH/S = 0.31 g/g, respectively; using fed‐batch culture, the results were QEtOH = 0.87 g L−1 h−1 and YEtOH/s = 0.44 g L−1 h−1, and QEtOH = 0.27 g L−1 h−1 and YEtOH/s = 0.57 g/g, respectively. Maximum volumetric productivity in continuous multistep cultures of ethanol and xylitol was at dilution rates of 0.131 and 0.074 h−1, respectively. Continuous multistep production, QEtOH increased up to 50% more than in fed‐batch culture, even though xylitol yield remained unchanged.
Lutein is a metabolite produced by plants or microalgae, and offers various health benefits. Chlorella sorokiniana FZU60 is considered a hyper-lutein producing microalga. In this study, the effects ...of temperature and light intensity on cell growth and lutein accumulation by FZU60 under mixotrophic conditions were investigated. Maximal lutein productivity and production were obtained at 33 °C, and high light intensity enhanced cell concentration but decreased lutein content when nitrate was replete. Further, two fed-batch strategies with gradient (fed-batch strategy I) or constant (fed-batch strategy II) NaAc feeding concentrations were compared. Results showed that the second strategy was more desirable for lutein production. Moreover, two types of mixotrophy/photoautotrophy two-stage strategies with a light intensity of 750 μmol/m2/s (two-stage strategy I) or 150 μmol/m2/s (two-stage strategy II) in stage 2 were explored. Under the two-stage strategy I, high lutein content (11.22 mg/g), productivity (8.25 mg/L/d), and production (65.96 mg/L) were achieved. The lutein content, productivity, and production obtained in this study are higher than those reported in most related studies, indicating that the proposed strategy can be used in commercial production.
•Cell growth and lutein accumulation had similar trend at different temperatures.•Sufficient CO2 and light illumination were essential for lutein production.•High light enhanced cell growth but decreased lutein content under replete nitrate.•Fed-batch strategy II with constant NaAc feeding enhanced lutein production.•Two-stage strategy I with high light in stage 2 improved lutein production.
In industrial microbial biotechnology, fed-batch processes are frequently used to avoid undesirable biological phenomena, such as substrate inhibition or overflow metabolism. For targeted process ...development, fed-batch options for small scale and high throughput are needed. One commercially available fed-batch fermentation system is the FeedPlate
, a microtiter plate (MTP) with a polymer-based controlled release system. Despite standardisation and easy incorporation into existing MTP handling systems, FeedPlates
cannot be used with online monitoring systems that measure optically through the transparent bottom of the plate. One such system that is broadly used in biotechnological laboratories, is the commercial BioLector. To allow for BioLector measurements, while applying the polymer-based feeding technology, positioning of polymer rings instead of polymer disks at the bottom of the well has been proposed. This strategy has a drawback: measurement requires an adjustment of the software settings of the BioLector device. This adjustment modifies the measuring position relative to the wells, so that the light path is no longer blocked by the polymer ring, but, traverses through the inner hole of the ring. This study aimed at overcoming that obstacle and allowing for measurement of fed-batch cultivations using a commercial BioLector without adjustment of the relative measurement position within each well.
Different polymer ring heights, colours and positions in the wells were investigated for their influence on maximum oxygen transfer capacity, mixing time and scattered light measurement. Several configurations of black polymer rings were identified that allow measurement in an unmodified, commercial BioLector, comparable to wells without rings. Fed-batch experiments with black polymer rings with two model organisms, E. coli and H. polymorpha, were conducted. The identified ring configurations allowed for successful cultivations, measuring the oxygen transfer rate and dissolved oxygen tension, pH, scattered light and fluorescence. Using the obtained online data, glucose release rates of 0.36 to 0.44 mg/h could be determined. They are comparable to formerly published data of the polymer matrix.
The final ring configurations allow for measurements of microbial fed-batch cultivations using a commercial BioLector without requiring adjustments of the instrumental measurement setup. Different ring configurations achieve similar glucose release rates. Measurements from above and below the plate are possible and comparable to measurements of wells without polymer rings. This technology enables the generation of a comprehensive process understanding and target-oriented process development for industrial fed-batch processes.
Microbial ammonia oxidation is the first and usually rate limiting step in nitrification and is therefore an important step in the global nitrogen cycle. Ammonia-oxidizing archaea (AOA) play an ...important role in nitrification. Here, we report a comprehensive analysis of biomass productivity and the physiological response of
to different ammonium and carbon dioxide (CO
) concentrations aiming to understand the interplay between ammonia oxidation and CO
fixation of
. The experiments were performed in closed batch in serum bottles as well as in batch, fed-batch, and continuous culture in bioreactors. A reduced specific growth rate (μ) of
was observed in batch systems in bioreactors. By increasing CO
gassing μ could be increased to rates comparable to that of closed batch systems. Furthermore, at a high dilution rate (
) in continuous culture (≥ 0.7 of μ
) the biomass to ammonium yield (Y
) increased up to 81.7% compared to batch cultures. In continuous culture, biofilm formation at higher
prevented the determination of
. Due to changes in Y
and due to biofilm, nitrite concentration becomes an unreliable proxy for the cell number in continuous cultures at
towards μ
. Furthermore, the obscure nature of the archaeal ammonia oxidation prevents an interpretation in the context of Monod kinetics and thus the determination of
. Our findings indicate that the physiological response of
might be regulated with different enzymatic make-ups, according to the ammonium catalysis rate. We reveal novel insights into the physiology of
that are important for biomass production and the biomass yield of AOA. Moreover, our study has implications to the field of archaea biology and microbial ecology by showing that bioprocess technology and quantitative analysis can be applied to decipher environmental factors affecting the physiology and productivity of AOA.
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•Conversion of agro-industrial waste residue into high value-added biomaterial.•First study on K. europaeus SGP37 utilizing sweet lime pulp waste for BNC production.•SLPW extracts as ...a sole nutrient source yielded significant amounts of BNC.•Static intermittent fed-batch cultivation greatly enhanced BNC production.•SLPWE-HS(w) under IFB-48h yielded commendable amount of BNC(∼38gL−1).
Herein, sweet lime pulp waste (SLPW) was utilized as a low- or no-cost feedstock for the production of bacterial nanocellulose (BNC) alone and in amalgamation with other nutritional supplements by the isolate K. europaeus SGP37 under static batch and static intermittent fed-batch cultivation. The highest yield (26.2±1.50gL−1) was obtained in the hot water extract of SLPW supplemented with the components of HS medium, which got further boosted to 38±0.85gL−1 as the cultivation strategy was shifted from static batch to static intermittent fed-batch. BNC obtained from various SLPW medium was similar or even superior to that obtained with standard HS medium in terms of its physicochemical properties. The production yields of BNC thus obtained are significantly higher and fit well in terms of industrial scale production.
