Synthesis Of Systems Research Articles

Application of a Cell-Free Synthetic Biology Platform for the Reconstitution of Teleocidin B and UK-2A Precursor Biosynthetic Pathways.

We report the successful cell-free reconstitution of two natural product biosynthetic pathways of divergent complexity and structural classes. We first constructed the teleocidin biosynthetic pathway using our BY-2 (tobacco) cell-free protein synthesis (CFPS) system. We discovered a direct interaction between TleA and MbtH, and showed that the BY-2 system is capable of producing more than 80 mg/L teleocidin B-3 with cofactor supplementation and ∼20 mg/L with no cofactors supplemented, demonstrating the high metabolic activity of the system. We then extended our methodology and report the first successful cell-free biosynthesis of UK-2 diol (precursor to the commercially valuable secondary metabolite UK-2A) from simple building blocks by refactoring a complex pathway of 10 proteins in the wheat germ CFPS system. We show that plant CFPS systems are suitable for reconstructing pathways and identifying the functions of uncharacterized genes linked to biosynthetic gene clusters and rate-limiting biosynthetic steps.

Machine Learning as a “Catalyst” for Advancements in Carbon Nanotube Research

The synthesis, characterization, and application of carbon nanotubes (CNTs) have long posed significant challenges due to the inherent multiple complexity nature involved in their production, processing, and analysis. Recent advancements in machine learning (ML) have provided researchers with novel and powerful tools to address these challenges. This review explores the role of ML in the field of CNT research, focusing on how ML has enhanced CNT research by (1) revolutionizing CNT synthesis through the optimization of complex multivariable systems, enabling autonomous synthesis systems, and reducing reliance on conventional trial-and-error approaches; (2) improving the accuracy and efficiency of CNT characterizations; and (3) accelerating the development of CNT applications across several fields such as electronics, composites, and biomedical fields. This review concludes by offering perspectives on the future potential of integrating ML further into CNT research, highlighting its role in driving the field forward.

Electric-Field Catalysis on Carbon Nanotubes in Electromicrofluidic Reactors: Monoterpene Cyclizations.

The control over the movement of electrons during chemical reactions with oriented external electric fields (OEEFs) has been predicted to offer a general approach to catalysis. Recently, we suggested that many problems to realize electric-field catalysis in practice under scalable bulk conditions could possibly be solved on multiwalled carbon nanotubes in electromicrofluidic reactors. Here, we selected monoterpene cyclizations to assess the scope of our system in organic synthesis. We report that electric-field catalysis can function by stabilizing both anionic and cationic transition states, depending on the orientation of the applied field. Moreover, electric-field catalysis can promote reactions which are barely accessible by general Brønsted and Lewis acids and field-free anion-π and cation-π interactions, and drive chemoselectivity toward intrinsically disfavored products without the need for pyrene interfacers attached to the substrate to prolong binding to the carbon nanotubes. Finally, interfacing with chiral organocatalysts is explored and evidence against contributions from redox chemistry is provided.

Sustainable methods for producing food-derived bioactive compounds

Background: Pursuing alternative starting materials has gained significant importance in the food and drug industries, driven by the global shift towards sustainable and renewable resources. Terpenes, naturally occurring hydrocarbons derived from plants, present a promising solution due to their abundant availability and range of physiological activities, including antioxidant, anti-inflammatory, antimicrobial, and anticancer properties. Objective: This study aims to develop a waste-free synthesis methodology for functionalizing limonene and myrcene, two notable terpenes, through Atom Transfer Radical Reactions (ATRR). The objective is to achieve high yields and purities while enhancing the therapeutic potential of these compounds and adhering to green chemistry principles. Methods: A catalytic complex of Cu(I)Br and dimethyl sulfoxide (DMSO) was used to facilitate the coupling of limonene and myrcene with various trichloroacetic acid derivatives. Reactions were performed at 80°C for 90 minutes, and products were purified by column chromatography. The synthesized compounds were characterized using Gas Chromatography-Mass Spectrometry (GC-MS) and Nuclear Magnetic Resonance (NMR). Microbiological analyses were conducted using the disk-diffusion method, and statistical analyses were performed to assess the reliability of the experiments. Results: The ATRR methodology enabled the efficient functionalization of limonene and myrcene, achieving high yields and purities. The best yield for limonene was 70%, and for myrcene, 82%, both using trichloroacetonitrile. The Cu(I) catalytic system showed dualistic biological effects, with 3% concentration exhibiting strong bactericidal and fungicidal activity, while 0.03% promoted growth. These results demonstrate the potential of the methodology and the catalytic system for bioactive compound synthesis and agricultural applications. Conclusion: This study demonstrates the viability of ATRR for efficiently functionalizing limonene and myrcene, achieving high yields and purities. The synthesized compounds show promise for pharmaceutical development. The dual antibacterial and antifungal activities of the Cu(I) catalytic system also suggest potential applications in wastewater reutilization. Future work should explore the biological effects and environmental impact of this process. Keywords: Terpenes, ATR reactions, Green Chemistry, Bioactive compounds, Therapeutic potential.

Possibilities of using secondary plant metabolites as antitumor agents

The review summarized the literature data of recent years on the antitumor effect of secondary plant metabolites, as well as their immunotropic and anti-inflammatory effects as components of the antitumor response. The biological basis for the action of secondary plant metabolites was characterized in the form of influence on potential targets: transcription factors, signaling pathways and receptors responsible for proliferation and apoptosis. The ways of increasing the bioavailability of secondary plant metabolites to enhance the effectiveness and possibility of their medicinal use were considered, the effects of berberine, curcumin and their derivatives were described. The search for scientific publications was conducted in foreign (PubMed) and domestic (eLibrary) electronic libraries. It was found that the multiplicity of molecular targets of secondary plant metabolites and the pleiotropy of their effects suggest the possibility of their use for the regulation of various processes in tumor and normal cells. There was a connection between the antitumor effect of secondary plant metabolites and their anti-inflammatory and immunomodulatory action. However, a significant limitation of their use was the fact that most studies were conducted on cell cultures, which was insufficient to judge the antitumor effect. Clinical trials were few and their results were contradictory. In addition, a certain contradiction has been noted between the idea of a more effective action when using a pure substance or a complex composition of various plant components. An important problem was the low bioavailability of most secondary plant metabolites, for which various methods have been proposed. Despite the long history of phytotherapy in oncology, the development of new derivatives of secondary plant metabolites with high water solubility remains relevant, including modified molecules of known secondary plant metabolites and the search for new ones with unexplored biological activity. Modern methods of chemical synthesis and delivery systems of derivatives of secondary plant metabolites, as well as the study of their effects in model experiments, seem to be promising scientific directions for the creation of new drugs with antitumor activity.

Ionic liquid supported hydrogel–lipase biocatalytic systems in asymmetric synthesis of enantiomerically pure S-ibuprofen

Novel hydrogel biocatalysts with immobilized lipase, stabilized by ionic liquids (ILs) of different hydrophobicity, were synthesized and evaluated. Variations of the time of immobilization and ratio of substrates during hydrogel synthesis were considered to obtain the most stable biocatalyst with the highest activity. Physicochemical characterization proved the success of the hydrogel synthesis and enzyme deposition on the surface of the support. Nevertheless, the key objective was to produce a biocatalyst for further application in ibuprofen methyl ester resolution, with the aim of obtaining an enantiomerically pure product. The hydrogel biocatalysts obtained in the presence of 5 wt% ILs after 8 h of immobilization achieved the highest activity recovery of 62 %. After 10 reaction cycles, enzymatic activity was still above 60 %, and the negative effect of pH and temperature on the activity of immobilized lipase was much lower than in the case of the free enzyme. After application of the catalyst in the resolution of ibuprofen methyl ester, the enantiomeric excess and conversion rate of the process were obtained for the dynamic kinetic resolution in isooctane. A conversion rate of 95 % was achieved due to the stabilization of the biocatalyst with IL and its resulting high catalytic activity. The study thus provides the pharmaceutical industry with a new potential approach with a strong scientific foundation.

Atmospheric Pressure Portable Catalytic Thermal Plasma System for Fast Synthesis of Aqueous NO3 and NO2 Fertilizer from Air and Water

AbstractMeaningful deployment of plasma water-based nitrogen fixation in agricultural application is hindered primarily due to its poor synthesis rate in compact systems. The study reports a directly deployable thermal plasma based portable catalytic compact system, offering typical synthesis rate as high as 1035 mg/min for nitrate and 635 mg/min for nitrite directly from naturally abundant atmospheric air and water. Developed technology is clean, sustainable, easily decentralizable, and completely free from fossil fuels and harmful intermediates like ammonia. The system avoids safety hazards and costs related to the requirements of continuous energy resources, pressurized environment for synthesis, regulated storage, refrigeration need, transportation of raw materials and distribution of fertilizer, as may be required by other competing technologies. Described system, consisting of air plasma torch, reaction chamber, water injection manifold and catalytic bed creates a unique nascent reactive plasma environment at ambient pressure that auto activates the catalyst in the field of thermal plasma for highly efficient fixation of nitrogen. Presented results indicate that use of combination catalysts with mechanically enhanced surface area allows drastic enhancement in the nitrogen fixation. Possible reaction chemistries, results of trials with different catalysts, time evolution of concentration, auto-conversion from nitrite to nitrate in aqueous media, time stability of concentration of the synthesized nitrate and observed remarkable effectiveness in the actual field trials are presented. Achieved synthesis rates are compared with those reported in literature in the area of thermal and non-thermal plasma.

Assessment of the effects of cannabidiol and a CBD-rich hemp extract in Caenorhabditis elegans.

Consumer use of cannabidiol (CBD) is growing, but there are still data gaps regarding its possible adverse effects on reproduction and development. Multiple pathways and signaling cascades involved in organismal development and neuronal function, including endocannabinoid synthesis and signaling systems, are well conserved across phyla, suggesting that Caenorhabditis elegans can model the in vivo effects of exogenous cannabinoids. The effects in C. elegans on oxidative stress response (OxStrR), developmental timing, juvenile and adult spontaneous locomotor activity, reproductive output, and organismal CBD concentrations were assessed after exposure to purified CBD or a hemp extract suspended in 0.5% sesame oil emulsions. In C. elegans, this emulsion vehicle is equivalent to a high-fat diet (HFD). As in mammals, HFD was associated with oxidative-stress-related gene expression in C. elegans adults. CBD reduced HFD-induced OxStrR in transgenic adults and counteracted the hypoactivity observed in HFD-exposed wild-type adults. In C. elegans exposed to CBD from the onset of feeding, delays in later milestone acquisition were irreversible, while later juvenile locomotor activity effects were reversible after the removal of CBD exposure. CBD-induced reductions in mean juvenile population body size were cumulative when chronic exposures were initiated at parental reproductive maturity. Purified CBD was slightly more toxic than matched concentrations of CBD in hemp extract for all tested endpoints, and both were more toxic to juveniles than to adults. Dosimetry indicated that all adverse effect levels observed in C. elegans far exceeded recommended CBD dosages for humans.

Formation kinetics and equilibrium thermodynamics study on the Bi2O3-TiO2 system for synthesis of the bismuth titanate ceramics

Formation kinetics and equilibrium thermodynamics are fundamental knowledge for predicting microstructure and properties during materials processing and utilization. This work prepared high purity Bi12TiO20, Bi4Ti3O12 and Bi2Ti4O11 compounds and characterized these compounds by in-suit X-ray diffraction, scanning electron microscope, energy dispersive spectrometer as well as calorimetry. Based on the experimental data obtained in this work and the literature, a thermodynamic description of the Bi2O3–TiO2 system was carried out, a thermodynamic database as well as the calculated phase diagram of the system was provided. The discrepancy between the present calculation with the experimental data were discussed, the formation kinetics of the compounds was explained and the bismuth titanate ceramics with tailored structure and specific physical properties were sintered.

The Potential of Artificial Cells Functioning under In Situ Deep-Sea Conditions.

Artificial cells with reconstructed cellular functions could serve as practical protocell models for studying the early cellular life on the Earth. Investigating the viability of protocell models in extreme environments where life may have arisen is important for advancing origin-of-life research. Here, we tested the survivability of lipid membrane vesicles in deep-sea environments. The vesicles were submerged in the deep-sea floor with a human-occupied vehicle. Although most of the vesicles were broken, some vesicles maintained a spherical shape after the dives. When a cell-free protein synthesis system was encapsulated inside, a few vesicles remained even after a 1,390 m depth dive. Interestingly, such artificial cells could subsequently synthesize protein in a nutrient-rich buffer solution. Together with on shore experiments showing artificial cells synthesized protein under high pressure, our results suggest artificial cells may be able to express genes in deep-sea environments where thermal energy is available from hydrothermal vents.

Optimized and nonTi-site doped synthesis of lithium titanate by mechanochemical method

The Li2CO3-ammonia-ballmilling synthesis system of Li4Ti5O12 (LTO) was optimized and doped Li and O by theirs adjacent elements Mg and F respectively. By adjusting the ballmilling parameter, the distribution of Li and Ti sources, the hydrolysis rate and different nucleophilic / electrophilic hydrolysis path of Ti, and the interaction between Li and Ti species can be effectively controlled. The temperature programmed calcination is beneficial to the formation of the middle state (Li2TiO3), obtaining the high quality LTO. Mg and F doping can further optimize the hydrolysis and condensation degree of Ti source, the number of crystal nucleus and the particle size. Therefore, the initial first discharged capability of Mg doped LTO and F doped LTO reach to 152.4 mAh/g and 163.1 mAh/g at 5 C respectively, corresponding 32.4 % and 41.7 % enhancing compared to LTO (115.1 mAh/g). Moreover, the discharge voltage of LTO-Mg decreases from 1.5 V to 1.3 V.

A cell-free bacteriophage synthesis system for directed evolution

Efficient phage production has always been an urgent need in fields such as drug discovery, disease treatment, and gene evolution. To meet this demand, we constructed a robust cell-free synthesis system for generating M13 phage by simplifying its genome, enabling a three-times faster efficiency compared with the traditional method in vivo. We further developed a cell-free directed evolution system in droplets, comprising a modified helper plasmid (ΔPS-ΔgIII-ΔgVI) and the simplified M13 genome-carrying gene mutation library. This system was greatly improved when coupled with fluorescence-activated droplet sorting (FADS). We successfully evolved the T7 RNA polymerase (RNAP), achieving a twofold higher activity to read through the T7 terminator. Moreover, we evolved the tryptophan tRNA into a suppressor tRNA with an eightfold increase in activity to read through the stop codon UAG.

Thermodynamic and Economic Analysis of the Green Ammonia Synthesis System Driven by Synergistic Hydrogen Production Using Alkaline Water Electrolyzers and Proton Exchange Membrane Electrolyzers

Green ammonia and hydrogen from renewable energy sources have emerged as crucial players during the transition of the chemical industry from a fossil energy‐dominated economy to one that is environmentally friendly. This work proposes a green ammonia synthesis system driven by synergistic hydrogen generation using alkaline water electrolyzers (AWE) and proton exchange membrane electrolyzers (PEMEC). The effects of hydrogen‐production ratios of PEMEC and AWE on the thermodynamic and economic performance of the system are compared and analyzed via multi‐objective optimization. The findings showed that an increase in the amount of hydrogen produced by PEMEC improves the system's energy efficiency, but the payback period is delayed because of the PEMEC high initial investment cost. The techno‐economic performance of the system at a 1:1 ratio of PEMEC to AWE hydrogen production are investigated considering the system level heat integration based on the pinch point analysis method to maximize the heat recovery. The results show that increasing the operational temperature, the pressure of the electrolyzer, and the ammonia synthesis pressure will enhance the system's thermal performance. Economic analysis shows that reducing electricity prices and electrolyzer investment costs will be the key to achieving the economic feasibility of the green ammonia system.

Regulating the Scaling Relations in Ammonia Synthesis through a Light‐Driven Bendable Seesaw Effect on Tailored Iron Catalyst

AbstractAdvancing the energy‐intensive Haber–Bosch process faces significant challenges due to the intrinsic constraints of scaling relations in heterogeneous catalysis. Herein, we reported an approach of bending the “seesaw effect” to regulate the scaling relations over a tailored α‐Fe metallic material (α‐Fe‐110s), realizing highly efficient light‐driven thermal catalytic ammonia synthesis with a rate of 1260 μmol gcatalyst−1 h−1 without additional heating. Specifically, the thermal catalytic activity of α‐Fe‐110s was significantly enhanced by the novel stepped {110} surface, exhibiting a 3.8‐fold increase compared to the commercial fused‐iron catalyst with promoters at 350 °C. The photo‐induced hot electron transfer further accelerates the dinitrogen dissociation and hydrogenation simultaneously, effectively overcoming the limitation of scaling relation over identical sites. Consequently, the ammonia production rate of α‐Fe‐110s was further enhanced by 30 times at the same temperature with irradiation. This work designs an efficient and sustainable system for ammonia synthesis and provides a novel approach for regulating the scaling relations in heterogeneous catalysis.

Ordered versus Non-Ordered Mesoporous CeO2-Based Systems for the Direct Synthesis of Dimethyl Carbonate from CO2.

In this work, non-ordered and ordered CeO2-based catalysts are proposed for CO2 conversion to dimethyl carbonate (DMC). Particularly, non-ordered mesoporous CeO2, consisting of small nanoparticles of about 8 nm, is compared with two highly porous (635-722 m2/g) ordered CeO2@SBA-15 nanocomposites obtained by two different impregnation strategies (a two-solvent impregnation method (TS) and a self-combustion (SC) method), with a final CeO2 loading of 10 wt%. Rietveld analyses on XRD data combined with TEM imaging evidence the influence of the impregnation strategy on the dispersion of the active phase as follows: nanoparticles of 8 nm for the TS composite vs. 3 nm for the SC composite. The catalytic results show comparable activities for the mesoporous ceria and the CeO2@SBA-15_SC nanocomposite, while a lower DMC yield is found for the CeO2@SBA-15_TS nanocomposite. This finding can presumably be ascribed to a partial obstruction of the pores by the CeO2 nanoparticles in the case of the TS composite, leading to a reduced accessibility of the active phase. On the other hand, in the case of the SC composite, where the CeO2 particle size is much lower than the pore size, there is an improved accessibility of the active phase to the molecules of the reactants.

Construction and characterization of Fe3O4@SiO2-Dop/Amide-BTA-pd(0) nanocomposite as a novel and efficient nanomagnetic recoverable catalyst for synthesis of 5-substituted 1h-tetrazoles in water

The development of efficient, and environmentally friendly catalytic systems for synthesis of 5-substituted 1H-tetrazoles is an important challenge in organic synthesis because tetrazole derivatives are well known in medicinal chemistry with many biological activities such as anti-bacterial, anti-fungal, anti-allergic, anti-viral, anti-cancer, reducing blood pressure, and so on. In this research, we fabricated a palladium(0) complex stabilized on magnetic Fe3O4 nanoparticles modified with dopamine and benzo[d]thiazole-2-carbonyl chloride [Fe3O4@SiO2-Dop/Amide-BTA-Pd(0)] (1) and evaluated its catalytic behavior in the synthesis of 5-substituted 1H-tetrazoles through one-pot tandem reactions of aryl halides with K4[Fe(CN)6] and sodium azide under eco-friendly conditions. One-pot tandem reactions of aryl halides with K4[Fe(CN)6] and sodium azide were conducted using a catalytic amount of 1 in the presence of potassium carbonate in water at 100 °C for 2 h the 5-substituted 1H-tetrazole products were obtained in high to excellent yields. At the end of the reaction, 1 can be easily recycled and the products obtained had high purity.

Efficient separation of HEPES and CO2-derived fructose by nanofiltration: Optimizing pH for improved separation and proposing a potential mechanism

Utilization of an in vitro multi-enzyme catalytic system for the synthesis of fructose from carbon dioxide is a promising approach towards achieving carbon neutrality. However, the lack of effective separation methods for the buffer HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) and the product fructose within the system has hindered the industrialization of in vitro multi-enzyme catalytic synthesis systems. This study aimed to separate and recover HEPES and fructose using nanofiltration technology. We investigated the influence of various parameters, including total solution concentration, solute concentration ratio, pH, and temperature, on the nanofiltration separation performance. Experimental results revealed that increasing the pH to 9.0 significantly decreased the fructose retention rate while maintaining a nearly complete HEPES retention rate. The HEPES-fructose mixed solution was further subjected to diafiltration using a spiral-wound membrane, ultimately yielding fructose and HEPES solutions with purities of 90.22 % and 92.93 %, respectively. Combining separation test results with models such as DSPM-DE, and using characterization methods such as ATR-FTIR and pore size distribution analysis, we observed pore swelling during nanofiltration of the HEPES-fructose solution at pH 9.0. A mechanism for this pore swelling, induced by the dissociation of HEPES under the influence of pH, was proposed: Dissociated HEPES in the solution interacts with the membrane surface at the liquid-membrane interface, causing reversible swelling of the membrane pores at the membrane surface, thereby reducing the retention rate of fructose. The nanofiltration process developed in this study effectively separates HEPES and fructose, enabling resource recycling and cost reduction. Furthermore, this study provides theoretical foundations and practical guidance for the design of nanofiltration separation strategies for similar systems using HEPES as a buffer, holding significant scientific and practical value.

A Self-Assembling γPFD-SpyCatcher Hydrogel Scaffold for the Coimmobilization of SpyTag-Enzymes to Facilitate the Catalysis of Regulated Enzymes.

In this study, a γPFD-SpyCatcher hydrogel scaffold with the capacity for spontaneous assembly was established. With a maximum loading capacity of a 1:1 molar ratio with SpyTag-enzymes, the immobilized proteins can not only rapidly provide pure enzymes but also exhibit improved thermal and pH stability. The results of the transmission electron microscopic analysis and the traits they present indicated that SpyCatcher promotes the aggregation of γPFD and the formation of hydrogels. In the cell-free pyruvate synthesis system, the γPFD-SpyCatcher coimmobilized SpyTag-hexokinase (HK), SpyTag-phosphofructokinase (PFK) and SpyTag-pyruvate kinase (PK) were employed, and the production of pyruvate increased by 43, 78 and 47% respectively. In in vitro experiments, the oxidative deamination activity of glutamate dehydrogenase (GDH) coimmobilized with γPFD-SpyCatcher was 38% higher than that of purified enzymes. These findings indicate that the γPFD-SpyCatcher-based hydrogels play an important role in breaking the barrier of regulatory enzymes and will provide more strategies for the development of synthetic biology.

Interface Nucleus Complementarity: An Iterative Process for the Discovery of Modular Intermetallics Guided by Chemical Pressure.

We present an iterative process for the discovery of modular intermetallic structures based on a recently developed interface nucleus approach. The process begins with the proposal of a suitable geometric motif that may serve as an interface nucleus. We then screen crystallographic databases for structures containing this motif as potential intergrowth partners. The extent to which pairs of these structures are likely to combine into more complicated assembles is then assessed with a new chemical pressure-based metric, interface nucleus complementarity (INC). Promising combinations of structures are translated into systems for synthesis, with new compounds providing either support for the importance of the original interface nucleus or new geometrical motifs for the next round of analysis. We demonstrate this process using a fragment derived from the σ-phase structure as a starting point, leading to the synthesis of PrMg4.13Zn10.20 and a new motif to seed the next cycle.

Efficient metabolic pathway modification in various strains of lactic acid bacteria using CRISPR/Cas9 system for elevated synthesis of antimicrobial compounds

Lactic acid bacteria (LAB) are known to exhibit various beneficial roles in fermentation, serving as probiotics, and producing a plethora of valuable compounds including antimicrobial activity such as bacteriocin-like inhibitory substance (BLIS) that can be used as biopreservative to improve food safety and quality. However, the yield of BLIS is often limited, which poses a challenge to be commercially competitive with the current preservation practice. Therefore, the present work aimed to establish an optimised two-plasmid CRISPR/Cas9 system to redirect the carbon flux away from lactate towards compounds with antimicrobial activity by disrupting lactate dehydrogenase gene (ldh) on various strains of LAB. The lactic acid-deficient (ldhΔ) strains caused a metabolic shift resulting in increased inhibitory activity against selected foodborne pathogens up to 78 % than the wild-type (WT) strain. The most significant effect was depicted by Enterococcus faecalis-ldh∆ which displayed prominent bactericidal effects against all foodborne pathogens as compared to the WT that showed no antimicrobial activity. The present work provided a framework model for economically important LAB and other beneficial bacteria to synthesise and increase the yield of valuable food and industrial compounds. The present work reported for the first time that the metabolism of selected LAB can be manipulated by modifying ldh to attain metabolites with higher antimicrobial activity.