Systems metabolic engineering of Escherichia coli for hyper-production of 5­aminolevulinic acid.

BACKGROUND: 5-Aminolevulinic acid (5-ALA) is a promising biostimulant, feed nutrient, and photodynamic drug with wide applications in modern agriculture and therapy. Although microbial production of 5-ALA has been improved realized by using metabolic engineering strategies during the past few years, there is still a gap between the present production level and the requirement of industrialization. RESULTS: In this study, pathway, protein, and cellular engineering strategies were systematically employed to construct an industrially competitive 5-ALA producing Escherichia coli. Pathways involved in precursor supply and product degradation were regulated by gene overexpression and synthetic sRNA-based repression to channel metabolic flux to 5-ALA biosynthesis. 5-ALA synthase was rationally engineered to release the inhibition of heme and improve the catalytic activity. 5-ALA transport and antioxidant defense systems were targeted to enhance cellular tolerance to intra- and extra-cellular 5-ALA. The final engineered strain produced 30.7 g/L of 5-ALA in bioreactors with a productivity of 1.02 g/L/h and a yield of 0.532 mol/mol glucose, represent a new record of 5-ALA bioproduction. CONCLUSIONS: An industrially competitive 5-ALA producing E. coli strain was constructed with the metabolic engineering strategies at multiple layers (protein, pathway, and cellular engineering), and the strategies here can be useful for developing industrial-strength strains for biomanufacturing.

CNT-TiO2 coating bonded onto stainless steel wire as a novel solid-phase microextraction fiber.

A novel solid-phase microextraction (SPME) fiber based on carbon nanotubes-titanium oxide (CNT-TiO2) composite coating bonded onto stainless steel wire was prepared via electroless plating and sol-gel techniques. The SPME coating was characterized by scanning electron microscopy and Raman microscopy. Coupled to gas chromatography (GC), the fiber was investigated with seven polycyclic aromatic hydrocarbons (PAHs) in direct-immersion mode. The SPME-GC analytical method was evaluated under optimized extraction conditions. Compared with other reports, higher sensitivity (LODs, 0.002-0.004 µg L(-1)) and better linear range (0.01-100 and 0.01-200 µg L(-1)) were obtained by the proposed method. The fiber exhibited high thermal stability to 300 °C and excellent durability in HCl and NaOH solutions. The as-established SPME-GC method was used to analyze the real water samples and satisfactory results were obtained.

A solid-phase microextraction fiber with carbon nanoparticles as sorbent material prepared by a simple flame-based preparation process.

A novel carbon nanoparticles-coated solid-phase microextraction (SPME) fiber was prepared via a simple and low-cost flame-based preparation process, with stainless steel wire as support. Surface characteristic of the fiber was studied with scanning electron microscope. A nano-scaled brushy structure was observed. Coupled to gas chromatography (GC), the fiber was used to extract phthalate esters (PAEs) and polycyclic aromatic hydrocarbons (PAHs) in aqueous samples. Analytical performances of the proposed method were investigated under the optimum extraction conditions (extraction temperature, 40°C; content of KCl, 30% (w/v); extraction time, 50min for PAEs and 40min for PAHs) and compared with other reports for the same analytes. Calibration ranges were 0.06-500µgL(-1) for di-n-butyl phthalate (DBP), and 0.1-300µgL(-1) for di-cyclohexyl phthalate (DCHP) and di-(2-ethyl-hexyl) phthalate (DEHP). For the eight PAHs, good linearity was obtained ranging from 0.01 to 150µgL(-1). Limits of detection were 0.005µgL(-1) for three PAEs and 0.001-0.003µgL(-1) for eight PAHs. The fiber exhibited excellent stability. It can be used for 100 times with RSDs of extraction efficiency less than 22.4%. The as-established SPME-GC method was applied to determine PAEs in food-wrap and PAHs in cigarette ash and snow water, and satisfactory results were obtained. The carbon nanoparticles-coated SPME fiber was efficient for sampling of organic compounds from aqueous samples.

Adsorbent for hydroquinone removal based on graphene oxide functionalized with magnetic cyclodextrin-chitosan.

Magnetic cyclodextrin-chitosan/graphene oxide (CCGO) with high surface area was synthesized via a simple chemical bonding method. The characteristics results of FTIR, SEM, TEM and XRD showed that CCGO was prepared. The large saturation magnetization (22.35 emu/g) of the synthesized nanoparticles allows fast separation of the CCGO from liquid suspension. These composites could efficiently remove hydroquinone from simulated wastewater with a facile subsequent solid-liquid separation because of their large area, abundant hydroxyl and amino groups with handy operation, and hydrophobicity. The hydroquinone removal process was found to obey the Freundlich adsorption model and its kinetics followed pseudo-second-order rate equation. The hydroquinone removal mechanism of CCGO might be attributed to the electrostatic adsorption of hydroquinone in the form of negatively charged hydroquinone by positively charged chitosan, accompanying hydroquinone absorbed by cavities of the cyclodextrin, and forming hydrogen bonds between hydroquinone and the hydroxyl groups on the surface of CCGO. The used CCGO could be recovered with ethanol. This study provides a promising nanostructured adsorbent with easy separation property for heavy metal ions removal.

Adsorbent for chromium removal based on graphene oxide functionalized with magnetic cyclodextrin-chitosan.

A simple chemical bonding method to synthesize magnetic cyclodextrin-chitosan/graphene oxide (CCGO) was reported. The adsorption behaviors of Cr(VI) in aqueous solution on CCGO were systematically investigated. As the results shown that, with the advantage of high surface area, abundant hydroxyl and amino groups of CCGO, and the magnetic property of Fe3O4, the Cr(VI) can be easily and rapidly extracted from the water by magnetic attraction under investigation. The adsorption equilibrium of CCGO for Cr(VI) corresponded with Langmuir isotherm, and the novel adsorbent exhibited better Cr(VI) removal efficiency in solutions with low pH. It was found that the Cr(VI) adsorption performance of CCGO strongly depends on their surface charge concentration and specific surface area. These results provide evidences for estimating and optimizing the removal of metal ions from the wastewater by using of CCGO composites in the future.

A microflow chemiluminescence sensor for indirect determination of dibutyl phthalate by hydrolyzing based on biological recognition materials.

A microflow chemiluminescence (CL) sensor for determination of dibutyl phthalate (DBP) based on magnetic molecularly imprinted polymer (MMIP) as recognition element was fabricated. Briefly, a hydrophilic molecularly imprinted polymer layer was produced at the surface of Fe3O4@SiO2 magnetic nanoparticles (MNPs) via combination of molecular imprinting and reversible stimuli responsive hydrogel. In this protocol, the initial step involved co-precipitation of Fe²âº and Fe³âº in an ammonia solution. Silica was then coated on the Fe3O4 nanoparticles using a sol-gel method to obtain silica shell magnetic nanoparticles. The MMIP was synthesized using methacrylic acid (MAA) as functional monomer and ethylene glycol dimethacrylate (EGDMA) as cross-linker and 2,2-azobisisobutyronitrile (AIBN) as initiator in chloroform. Then the synthesized MMIP and magnetic non-molecular imprinted polymers (MNIP) were employed as recognition by packing into lab-made straight shape tubes, connected in CL analyzer for establishing the novel sensor with a single channel syringe pump. And a mixer for hydrolyzing of DBP was followed. Based on this experiment principle, DBP was determined indirectly. And the MMIP showed satisfactory recognition capacity to DBP, resulting to the wide linear range of 3.84 × 10⁻8 to 2.08 × 10⁻5 M and the low detection limit of 2.09 × 10⁻9 M (3σ) for DBP. The relative standard deviation (RSD) for DBP (3.20 × 10⁻6 M) was 1.40% (n=11). Besides improving sensitivity and selectivity, the sensor was reusable. The proposed DBP-MMIP-CL sensor has been successfully applied to determine DBP in drink samples.

Highly selective adsorption of lead ions by water-dispersible magnetic chitosan/graphene oxide composites.

Magnetic chitosan/graphene oxide (MCGO) materials were fabricated through a facile and fast process and their application as excellent adsorbents for metal ions was also demonstrated. The characteristics results of FTIR, SEM, TEM, VSM and XRD showed that MCGO was successfully prepared. The SEM and TEM revealed that magnetic chitosan had been assembled on the surface of graphene oxide layers with a high density. The XRD and VSM indicated the MCGO had enough magnetic response to meet the need of magnetic separation. The magnetic chitosan grafted with graphene oxide sheets showed an increased surface area. The MCGO was used as sorbents for the removal of Pb(II) ions from large volumes of aqueous solutions. The effects of pH, contact time, and concentration on Pb(II) ions sorption were investigated. The results indicated that Pb(II) ions sorption on MCGO was strongly dependent on pH. The abundant functional groups on the surfaces of MCGO played an important role on Pb(II) sorption. Equilibrium studies showed that the data of Pb(II) adsorption followed the Langmuir model. The maximum adsorption capacity for Pb(II) was estimated to be 76.94 mg/g. The MCGO was stable and easily recovered.

Synthesis of magnetic ß-cyclodextrin-chitosan/graphene oxide as nanoadsorbent and its application in dye adsorption and removal.

Magnetic ß-cyclodextrin-chitosan/graphene oxide materials (MCCG) were fabricated through a facile chemical route and their application as excellent adsorbents for dye removal were also demonstrated. The characteristics results of FTIR, SEM, TEM and XRD showed that MCCG was successfully prepared. The results showed that, benefiting from the surface property of graphene oxide, hydrophobicity of ß-cyclodextrin, the abundant amino and hydroxyl functional groups of chitosan, and from the magnetic property of Fe(3)O(4), the adsorbent possesses quite a good and versatile adsorption capacity to the dye under investigation, and can be easily and rapidly extracted from water by magnetic attraction. Most importantly, the adsorbent can be easily and efficiently regenerated for reuse with hardly any compromise of the adsorption capacity. The adsorption kinetics, isotherms and thermodynamics were investigated to indicate that the kinetics and equilibrium adsorptions were well-described by pseudo-second-order kinetic and Langmuir isotherm model, respectively. The thermodynamic parameters suggested that the adsorption process was spontaneous and endothermic in nature. The inherent advantages of the nano-structured adsorbent, such as adsorption capacity, easy, handy operation, rapid extraction, and regeneration, may pave a new, efficient and sustainable way towards highly-efficient dye pollutant removal in water and wastewater treatment.

A chemiluminescence array sensor based on graphene-magnetite-molecularly imprinted polymers for determination of benzenediol isomers.

A chemiluminescence (CL) array sensor for determination of benzenediol isomers simultaneously using the system of luminol-NaOH-H(2)O(2) based on a graphene-magnetite-molecularly imprinted polymer (GM-MIP) is described. Use of graphene in the GM-MIP thus prepared is helpful to improve the adsorption capacity, while use of magnetite nanoparticles can facilitate the isolation of GM-MIP at end of their synthesis, and rendering easier the use of the polymers in the array sensor. The adsorption performance and properties were characterized. The GM-MIP was used to increase the selectivity in CL analysis. In addition, the sensor was reusable and of good selectivity and adsorption capacity. The array sensor was finally used for the determination of hydroquinone, resorcinol and catechol in waste water samples simultaneously.

Determination of L-tryptophan based on graphene oxide-magnetite-molecularly imprinted polymers and chemiluminescence.

A new method for determination of L-tryptophan (L-try) using the flow injection chemiluminescence (FI-CL) system of KMnO(4)-SnCl(2)-CHOH based on a graphene oxide-magnetite-molecularly imprinted polymer (GM-MIP) is described. The L-try GM-MIP was synthesized using graphene oxide (G) which improved the adsorption capacity as carrier, and magnetite nanoparticles which made the polymers easier to use in the sensor. The adsorption performance and properties were characterized. The GM-MIP was used in CL analysis to increase the selectivity and the possible mechanism was also discussed. The CL sensor responded linearly to the concentration of L-try over the range from 2.10×10(-7) to 7.09×10(-4) M with a detection limit of 2.11×10(-8) M (3σ). The relative standard deviation (RSD) for the determination of 3.0×10(-5) M L-try was 2.40% (n=11). On the basis of speediness and sensitivity, the sensor is reusable and shows a great improvement in selectivity and adsorption capacity over other sensors. The sensor has been used for the determination of L-try in drug samples.

Flow injection chemiluminescence sensor using core-shell molecularly imprinted polymers as recognition element for determination of dapsone.

This paper reports the preparation of dapsone (DDS) imprinted polymer layer-coated silica submicron particles (SiO(2)) combined with chemiluminescence (CL) toward analysis of tracing DDS in practical samples. To induce the selective occurrence of surface polymerization, the amino groups were first grafted at the surface of SiO(2) by the (3-aminopropyl)triethoxysilane (APTES). The molecularly imprinted polymers (MIP) were coated at the surface of modified SiO(2) by the graft copolymerization. After the removal of templates, recognition sites of DDS were exposed in the polymer layers. The DDS-imprinted products were characterized by FT-IR, SEM, TEM, dynamic adsorption, and static adsorption tests. The proximity between the thickness of MIP layer and the spatial size of DDS indicated that the imprinted sites almost situated at the surface of MIP, leading to rapid adsorption saturation within 90 min. The apparent maximum binding amount of MIP toward DDS was evaluated as 14.98 mg·g(-1), which was much higher than that of non-molecularly imprinted polymers. The CL sensor provided a wide linear range for DDS within 1.0 × 10(-6) to 1.0 × 10(-4) mol·L(-1) with a detection limit of 5.27 × 10(-7) mol·L(-1) and the relative standard deviation of 1.8 % (n = 11) by determinations of 5.0 × 10(-6) mol·L(-1) DDS. This method was applied to determine DDS in urine samples and satisfactory results were obtained.

Preparation of novel magnetic chitosan/graphene oxide composite as effective adsorbents toward methylene blue.

A novel magnetic composite bioadsorbent composed of magnetic chitosan and graphene oxide (MCGO) was prepared as the magnetic adsorbent toward methylene blue. The magnetic composite bioadsorbent was characterized by SEM, FTIR and XRD measurements. The effect factors including pH, contact time and temperature on the adsorption properties of methylene blue onto MCGO were investigated. The resulting shows extraordinary adsorption capacity and fast adsorption rates for removal of methylene blue. The kinetics are well-described by pseudo-second-order kinetic. The experimental data of isotherm followed the Langmuir isotherm model and the Freundlich model, respectively. This work shows that the MCGO could be utilized as an efficient, magnetically separable adsorbent for the environmental cleanup.

Fabrication of magnetic chitosan nanoparticles grafted with ß-cyclodextrin as effective adsorbents toward hydroquinol.

The adsorption characteristics of hydroquinol from aqueous solutions onto the ß-cyclodextrin modified magnetic chitosan nanoparticles (CMCN) had been investigated. The characteristics results of FTIR, SEM and XRD showed that CMCN was successfully prepared. The influences of the pH of the solution and the contact time on the adsorption amounts had been discussed, and the appropriate process conditions for the adsorption of hydroquinol had been obtained. Equilibrium experiments fitted well with the Freundlich isotherm model, and the maximum adsorption capacity of the CMCN at 303 K was determined to be 1.75 mmol/g for hydroquinol at the concentration of 9.0 mmol/L, much higher than some conventional adsorbents. The CMCN was stable and easily recovered. Moreover, the adsorption capacity was about 90% of the initial saturation adsorption capacity after being used four times.

Fabrication of novel magnetic chitosan grafted with graphene oxide to enhance adsorption properties for methyl blue.

A novel magnetic composite bioadsorbent composed of magnetic chitosan and graphene oxide (MCGO) was prepared as the magnetic adsorbent. The morphology, chemical structure and magnetic property of the MCGO were characterized by Fourier transform infrared spectrometer (FT-IR), X-ray diffraction (XRD) and Scanning electronic microscope (SEM), respectively. Adsorption of methyl blue (MB) onto MCGO was investigated with respect to pH, adsorption time, initial MB concentration and temperature. Kinetics data and adsorption isotherm, obtained at the optimum pH 5.3, were better fitted by pseudo-second-order kinetic model and by Langmuir isotherm, respectively. The values of activation parameters such as free energy (ΔG, -0.74∼-1.46kJmol(-1)), enthalpy (ΔH, -10.28kJmol(-1)) and entropy (ΔS, -36.35Jmol(-1)K(-1)) were determined, respectively, indicating that the adsorption was spontaneous, favorable and exothermic process in nature. Moreover, the MCGO was stable and easily recovered, the adsorption capacity was about 90% of the initial saturation adsorption capacity after being used four times.

Flow injection chemiluminescence sensor based on core-shell magnetic molecularly imprinted nanoparticles for determination of sulfadiazine.

A novel flow injection chemiluminescence (FI-CL) sensor for determination of sulfadiazine (SDZ) using core-shell magnetic molecularly imprinted polymers (MMIPs) as recognition element is developed. Briefly, a hydrophilic MMIPs layer was produced at the surface of Fe(3)O(4)@SiO(2) magnetic nanoparticles (MNPs) via combination of molecular imprinting and reversible stimuli responsive hydrogel. And it provided the MMIPs with excellent adsorption capacity and rapid adsorption rate due to the imprinted sites mostly situated on the surface of MMIPs. Then the prepared SDZ-MMIPs were packed into flow cell to establish a novel FI-CL sensor. The sensor provided a wide linear range for SDZ of 4.0×10(-7) to 1.0×10(-4) mol L(-1) with a detection limit of 1.54×10(-7) mol L(-1). And the relative standard deviation (RSD) for the determination of 1.0×10(-6) mol L(-1) SDZ was 2.56% (n=11). The proposed method was applied to determine SDZ in urine samples and satisfactory results were obtained.

Synthesis and characterization of magnetic ß-cyclodextrin-chitosan nanoparticles as nano-adsorbents for removal of methyl blue.

A novel nano-adsorbent, ß-cyclodextrin-chitosan (CDC) modified Fe(3)O(4) nanoparticles (CDCM) is fabricated for removal of methyl blue (MB) from aqueous solution by grafting CDC onto the magnetite surface. The characteristics results of FTIR, SEM and XRD show that CDC is grafted onto Fe(3)O(4) nanoparticles. The grafted CDC on the Fe(3)O(4) nanoparticles contributes to an enhancement of the adsorption capacity because of the strong abilities of CDCM, which includes the multiple hydroxyl, carboxyl groups, amino groups and the formation of an inclusion complex due to the ß-CD molecules through host-guest interactions, to adsorb MB. The adsorption of MB onto CDCM is found to be dependent on pH and temperature. Adsorption equilibrium is achieved in 50 min and the adsorption kinetics of MB is found to follow a pseudo-second-order kinetic model. Equilibrium data for MB adsorption are fitted well by Langmuir isotherm model. The maximum adsorption capacity for MB is estimated to be 2.78 g/g at 30°C. The CDCM was stable and easily recovered. Moreover the adsorption capacity was about 90% of the initial saturation adsorption capacity after being used four times.

Determination of L-phenylalanine on-line based on molecularly imprinted polymeric microspheres and flow injection chemiluminescence.

A novel molecular imprinting-chemiluminescence (MIP-CL) sensor for the determination of L-phenylalanine (Phe) using molecularly imprinted polymer (MIP) as recognition element is reported. The Phe-MIP was synthesized using acrylamide (AM) as functional monomer and ethylene glycol dimethacrylate (EGDMA) as cross-linker, 2,2-azobisisobutyronitrile (AIBN) as initiator and the polymers' properties were characterized. Then the synthesized MIP was employed as recognition element by packing into flow cell to establish a novel flow injection CL sensor. The CL intensity responded linearly to the concentration of Phe in the range 1.3 × 10(-6) to 5.44 × 10(-4) mol/L with a detection limit of 6.23 × 10(-7) mol/L (3σ), which is lower than that of conventional methods. The sensor is reusable and has a great improvement in sensitivity and selectivity for CL analysis. As a result, the new MIP-CL sensor had been successfully applied to the determination of Phe in samples.

Removal of alizarin red from water environment using magnetic chitosan with Alizarin Red as imprinted molecules.

A novel, chitosan coating on the surface of magnetite (Fe(3)O(4)) (MIMC) was successfully synthesized using alizarin red (AR) as a template for adsorption and removal of AR from aqueous solutions. Characterization of the obtained MIMC was achieved by FTIR spectra, SEM micrographs and XRD. Batch adsorption experiments were performed to investigate the adsorption conditions, selectivity and reusability. The results showed that the maximum adsorption capacity was 40.12 mg/g, observed at pH 3 and temperature 30°C. Equilibrium adsorption was achieved within 50 min. The kinetic data, obtained at the optimum pH 3, could be fitted with a pseudo-second-order equation. Adsorption process could be well described by Langmuir adsorption isotherms and the maximum adsorption capacity was calculated as 43.08 mg/g. The selectivity coefficient of AR and other dyes onto MIMC indicated an overall preference for AR, which was much higher than non-imprinted magnetic chitosan beads. Moreover, the sorbent represented high stability and good repeatability.

Removal of Ag+ from water environment using a novel magnetic thiourea-chitosan imprinted Ag+.

A novel, thiourea-chitosan coating on the surface of magnetite (Fe(3)O(4)) (Ag-TCM) was successfully synthesized using Ag(I) as imprinted ions for adsorption and removal of Ag(I) ions from aqueous solutions. The thermal stability, chemical structure and magnetic property of the Ag-TCM were characterized by the scanning electron microscope (SEM), Fourier transform infrared spectrometer (FT-IR) and vibrating sample magnetometer (VSM), respectively. Batch adsorption experiments were performed to evaluate the adsorption conditions, selectivity and reusability. The results showed that the maximum adsorption capacity was 4.93 mmol/g, observed at pH 5 and temperature 30°C. Equilibrium adsorption was achieved within 50 min. The kinetic data, obtained at the optimum pH 5, could be fitted with a pseudo-second order equation. Adsorption process could be well described by Langmuir adsorption isotherms and the maximum adsorption capacity calculated from Langmuir equation was 5.29 mmol/g. The selectivity coefficient of Ag(I) ions and other metal cations onto Ag-TCM indicated an overall preference for Ag(I) ions, which was much higher than non-imprinted thiourea-chitosan beads. Moreover, the sorbent was stable and easily recovered, the adsorption capacity was about 90% of the initial saturation adsorption capacity after being used five times.