Metabolic engineering of E. coli for producing phloroglucinol from acetate.

Phloroglucinol is a three-hydroxyl phenolic compound and has diverse physiological and pharmacological activities such as antivirus and anti-inflammatory activities. Chemical synthesis of phloroglucinol suffered from many drawbacks such as high cost and environmental pollution. To avoid the above issues, microbial phloroglucinol biosynthesis was successfully accomplished in this study, while the abundant and low-cost acetate was used as the main carbon source. Firstly, the toxicity of phloroglucinol was tested, and E. coli BL21(DE3) could tolerate 5 g/L phloroglucinol. The ability of phloroglucinol synthase (PhlD) for catalyzing malonyl-CoA to phloroglucinol was confirmed, and E. coli BL21(DE3) expressing PhlD and acetyl-CoA carboxylase (ACCase) could produce 1107 ± 12 mg/L phloroglucinol from glucose. Then, E. coli BL21(DE3) was engineered to utilize acetate to produce 228 ± 15 mg/L phloroglucinol. Then, the endogenous citrate synthase (GltA) which could catalyze oxaloacetate and acetyl-CoA generated from acetate to citrate was knocked down by CRISPRi system in order to enhance the carbon flux for phloroglucinol production, and the titer was improved to 284 ± 8 mg/L. This work demonstrated that acetate could be used as low-cost substrate to achieve the biosynthesis of phloroglucinol and provided an example of effective utilization of acetate.

Processing tactics for low-cost production of pure nuciferine from lotus leaf.

Nuciferine is an important drug candidate for the treatment of many diseases. However, there is no general method for its low - cost production. In this work, a feasible method for the production of nuciferine from lotus leaf, using ultrasonic-assisted extraction-solid phase extraction (UAE-SPE) as extraction and cleanup procedure, was developed. Petroleum ether and silica gel have been successfully used as extraction solvent and adsorbent to integrate UAE with SPE, respectively. Except for filtration, no treatment (e.g. concentration and redissolution, etc) was needed on UAE extract before SPE and the effluents obtained in the loading process of SPE could be used as UAE extraction solvent without purification. No obvious decline in the extraction efficiency of UAE and adsorption capacity of SPE was observed at least for 5 runs, which provides a feasible way for the continuous production of nuciferine in industry, i.e. Cyclic UAE-SPE. Moreover, SPE column could be conveniently regenerated and reused without significant decline in its adsorption capacity at least for 5 cycles, which can be used to reduce the cost of the whole system further. In comparison with other cleanup procedures, Cyclic UAE-SPE showed apparent advantages in energy conservation and emission reduction. LLE and crystallization were applied to separate nuciferine from other impurities further. Under optimum conditions, the total recovery rate of nuciferine with a purity over 90.0% from lotus leaf reached 50.1%. All in all, the developed method has advantages in convenient operation, low cost, and high efficiency, thus, is fitting for the production of high purity nuciferine.

Optimization of a food industry-waste-based medium for the production of the plant growth promoting microorganism Pseudomonas oryzihabitans PGP01 based on agro-food industries by-products.

In this study, three wastes based on potato peels and pulps, tomato seeds and wheat bran were used as basis for the preparation of a cheap medium to produce the bacterium P. oryzihabitans PGP01. In flasks experiments, P. oryzihabitans PGP01 growth at 25 °C in a medium based on frozen potato peels and pulp (FPP) with tryptone as a nitrogen source resulted in the maximum production compared to the commercial TSB medium. In the scale-up to 2 L bioreactors, FPP supplemented with tryptone, molasses, NaCl and K2HPO4 allowed to reach similar biomass production than in the TSB medium. A maximum growth of 4.4 × 109 CFU mL-1 after setting the agitation and the air flux conditions at 400 rpm and 0.75 vvm. Finally, P. oryzihabitans PGP01 growing in this optimized medium conserved its biological activity showing the expected effect in root development previously reported for this microorganism.

Photothermal and Moisture Actuator Made with Graphene Oxide and Sodium Alginate for Remotely Controllable and Programmable Intelligent Devices.

Functional materials with energy storage and conversion properties have been useful for actuating devices. Here, a new kind of torsional fiber-based actuator including graphene oxide (GO) and natural sodium alginate was prepared by traditional wet spinning and twisting methods, during which the fiber structure was reconstructed, and the mechanical energy was prestored. When the twisted GO/SA (graphene oxide/sodium alginate) fiber was stimulated by infrared light or moisture, the torsional structure of the fiber was activated instantaneously to generate rapid and reversible rotational motion, thus realizing the automatic release and re-storage process of rotational kinetic energy. In addition, the full revolutions of the twisted GO/SA fiber have no attenuation after 100 reversible rotations when stimulated by moisture, which proves the excellent rotational stability. Due to its excellent flexibility and wettability, the twisted GO/SA fiber can be woven into a network or prepared into a series of programmable intelligent devices, which is of great significance for future flexible intelligent electronic devices.

Efficient biosynthesis of (2S)-pinocembrin from d-glucose by integrating engineering central metabolic pathways with a pH-shift control strategy.

Microbial fermentations promise to revolutionize the conventional extraction of (2S)-pinocembrin from natural plant sources. Previously an Escherichia coli fermentation system was developed for one-step (2S)-pinocembrin production. However, this fermentation platform need supplementation of expensive malonyl-CoA precursor malonate and requires morpholinopropane sulfonate to provide buffering capacity. Here, a clustered regularly interspaced short palindromic repeats interference was constructed to efficiently channel carbon flux toward malonyl-CoA. By exploring the effects of different culture pH on microbial fermentation, it was found that high pH values favored upstream pathway catalysis, while low pH values favored downstream pathway catalysis. Based on this theory, a two-stage pH control strategy was proposed. The pH was controlled at 7.0 during 0-10h, and was shifted to 6.5 after 10h. Finally, the (2S)-pinocembrin titers increased to 525.8mg/L. These results were attained in minimal medium without additional precursor supplementation, thus offering opportunities for industrial scale low-cost production of flavonoids.

Uniform nanoparticles by flame-assisted spray pyrolysis (FASP) of low cost precursors.

A new flame-assisted spray pyrolysis (FASP) reactor design is presented, which allows the use of inexpensive precursors and solvents (e.g., ethanol) for synthesis of nanoparticles (10-20 nm) with uniform characteristics. In this reactor design, a gas-assisted atomizer generates the precursor solution spray that is mixed and combusted with externally fed inexpensive fuel gases (acetylene or methane) at a defined height above the atomizing nozzle. The gaseous fuel feed can be varied to control the combustion enthalpy content of the flame and onset of particle formation. This way, the enthalpy density of the flame is decoupled from the precursor solution composition. Low enthalpy content precursor solutions are prone to synthesis of non-uniform particles (e.g., bimodal particle size distribution) by standard flame spray pyrolysis (FSP) processes. For example, metal nitrates in ethanol typically produce nanosized particles by gas-to-particle conversion along with larger particles by droplet-to-particle conversion. The present FASP design facilitates the use of such low enthalpy precursor solutions for synthesis of homogeneous nanopowders by increasing the combustion enthalpy density of the flame with low-cost, gaseous fuels. The effect of flame enthalpy density on product properties in the FASP configuration is explored by the example of Bi(2)O(3) nanoparticles produced from bismuth nitrate in ethanol. Product powders were characterized by nitrogen adsorption, X-ray diffraction, X-ray disk centrifuge, and transmission electron microscopy. Homogeneous Bi(2)O(3) nanopowders were produced both by increasing the gaseous fuel content and, most notably, by cutting the air entrainment prior to ignition of the spray.