On-column capping of poly dT media-tethered mRNA accomplishes high capping efficiency, enhanced mRNA recovery, and improved stability against RNase.

The messenger RNA (mRNA) 5'-cap structure is indispensable for mRNA translation initiation and stability. Despite its importance, large-scale production of capped mRNA through in vitro transcription (IVT) synthesis using vaccinia capping enzyme (VCE) is challenging, due to the requirement of tedious and multiple pre-and-post separation steps causing mRNA loss and degradation. Here in the present study, we found that the VCE together with 2'-O-methyltransferase can efficiently catalyze the capping of poly dT media-tethered mRNA to produce mRNA with cap-1 structure under an optimized condition. We have therefore designed an integrated purification and solid-based capping protocol, which involved capturing the mRNA from the IVT system by using poly dT media through its affinity binding for 3'-end poly-A in mRNA, in situ capping of mRNA 5'-end by supplying the enzymes, and subsequent eluting of the capped mRNA from the poly dT media. Using mRNA encoding the enhanced green fluorescent protein as a model system, we have demonstrated that the new strategy greatly simplified the mRNA manufacturing process and improved its overall recovery without sacrificing the capping efficiency, as compared with the conventional process, which involved at least mRNA preseparation from IVT, solution-based capping, and post-separation and recovering steps. Specifically, the new process accomplished a 1.76-fold (84.21% over 47.79%) increase in mRNA overall recovery, a twofold decrease in operation time (70 vs. 140 min), and similar high capping efficiency (both close to 100%). Furthermore, the solid-based capping process greatly improved mRNA stability, such that the integrity of the mRNA could be well kept during the capping process even in the presence of exogenously added RNase; in contrast, mRNA in the solution-based capping process degraded almost completely. Meanwhile, we showed that such a strategy can be operated both in a batch mode and in an on-column continuous mode. The results presented in this work demonstrated that the new on-column capping process developed here can accomplish high capping efficiency, enhanced mRNA recovery, and improved stability against RNase; therefore, can act as a simple, efficient, and cost-effective platform technology suitable for large-scale production of capped mRNA.

Simultaneous Optimization and Integration of Multiple Process Heat Cascade and Site Utility Selection for the Design of a New Generation of Sugarcane Biorefinery.

Biorefinery plays a crucial role in the decarbonization of the current economic model, but its high investments and costs make its products less competitive. Identifying the best technological route to maximize operational synergies is crucial for its viability. This study presents a new superstructure model based on mixed integer linear programming to identify an ideal biorefinery configuration. The proposed formulation considers the selection and process scale adjustment, utility selection, and heat integration by heat cascade integration from different processes. The formulation is tested by a study where the impact of new technologies on energy efficiency and the total annualized cost of a sugarcane biorefinery is evaluated. As a result, the energy efficiency of biorefinery increased from 50.25% to 74.5% with methanol production through bagasse gasification, mainly due to its high heat availability that can be transferred to the distillery, which made it possible to shift the bagasse flow from the cogeneration to gasification process. Additionally, the production of DME yields outcomes comparable to methanol production. However, CO2 hydrogenation negatively impacts profitability and energy efficiency due to the significant consumption and electricity cost. Nonetheless, it is advantageous for surface power density as it increases biofuel production without expanding the biomass area.

Continuous integrated manufacturing for biopharmaceuticals: A new paradigm or an empty promise?

Continuous integrated bioprocessing has elicited considerable interest from the biopharma industry for the many purported benefits it promises. Today many major biopharma manufacturers around the world are engaged in the development of continuous process platforms for their products. In spite of great potential, the path toward continuous integrated bioprocessing remains unclear for the biologics industry due to legacy infrastructure, process integration challenges, vague regulatory guidelines, and a diverging focus toward novel therapies. In this article, we present a review and perspective on this topic. We explore the status of the implementation of continuous integrated bioprocessing among biopharmaceutical manufacturers. We also present some of the key hurdles that manufacturers are likely to face during this implementation. Finally, we hypothesize that the real impact of continuous manufacturing is likely to come when the cost of manufacturing is a substantial portion of the cost of product development, such as in the case of biosimilar manufacturing and emerging economies.

What MEMS Research and Development Can Learn from a Production Environment.

The intricate interdependency of device design and fabrication process complicates the development of microelectromechanical systems (MEMS). Commercial pressure has motivated industry to implement various tools and methods to overcome challenges and facilitate volume production. By now, these are only hesitantly being picked up and implemented in academic research. In this perspective, the applicability of these methods to research-focused MEMS development is investigated. It is found that even in the dynamics of a research endeavor, it is beneficial to adapt and apply tools and methods deduced from volume production. The key step is to change the perspective from fabricating devices to developing, maintaining and advancing the fabrication process. Tools and methods are introduced and discussed, using the development of magnetoelectric MEMS sensors within a collaborative research project as an illustrative example. This perspective provides both guidance to newcomers as well as inspiration to the well-versed experts.

An integrated and continuous downstream process for microbial virus-like particle vaccine biomanufacture.

In this study, we present the first integrated and continuous downstream process for the production of microbial virus-like particle vaccines. Modular murine polyomavirus major capsid VP1 with integrated J8 antigen was used as a model virus-like particle vaccine. The integrated continuous downstream process starts with crude cell lysate and consists of a flow-through chromatography step followed by periodic counter-current chromatography (PCC) (bind-elute) using salt-tolerant mixed-mode resin and subsequent in-line assembly. The automated process showed a robust behavior over different inlet feed concentrations ranging from 1.0 to 3.2 mg ml-1 with only minimal adjustments needed, and produced continuously high-quality virus-like particles, free of nucleic acids, with constant purity over extended periods of time. The average size remained constant between 44.8 ± 2.3 and 47.2 ± 2.9 nm comparable to literature. The process had an overall product recovery of 88.6% and a process productivity up to 2.56 mg h-1 mlresin-1 in the PCC step, depending on the inlet concentration. Integrating a flow through step with a subsequent PCC step allowed streamlined processing, showing a possible continuous pathway for a wide range of products of interest.

Integration of thermochemical water splitting with CO2 direct air capture.

Renewable production of fuels and chemicals from direct air capture (DAC) of CO2 is a highly desired goal. Here, we report the integration of the DAC of CO2 with the thermochemical splitting of water to produce CO2, H2, O2, and electricity. The produced CO2 and H2 can be converted to value-added chemicals via existing technologies. The integrated process uses thermal solar energy as the only energy input and has the potential to provide the dual benefits of combating anthropogenic climate change while creating renewable chemicals. A sodium-manganese-carbonate (Mn-Na-CO2) thermochemical water-splitting cycle that simultaneously drives renewable H2 production and DAC of CO2 is demonstrated. An integrated reactor is designed and fabricated to conduct all steps of the thermochemical water-splitting cycle that produces close to stoichiometric amounts (∼90%) of H2 and O2 (illustrated with 6 consecutive cycles). The ability of the cycle to capture 75% of the ∼400 ppm CO2 from air is demonstrated also. A technoeconomic analysis of the integrated process for the renewable production of H2, O2, and electricity, as well as DAC of CO2 shows that the proposed scheme of solar-driven H2 production from thermochemical water splitting coupled with CO2 DAC may be economically viable under certain circumstances.

Clustering and optimising regional segregated resource allocation networks.

Policymakers and officials worldwide are making more stringent environmental norms and waste disposal policies to encourage industries to move towards cleaner production. One of the main challenges that industries face moving towards cleaner production is the adoption of different strategies for optimising their resource utilisation and waste reduction economically. This is particularly challenging for large-scale industries or a group of industrial plants located in an industrial region. This paper presents a novel approach to economic resource optimisation focussed mainly on large-scale industries, different industrial plants located in the vicinity of each other, or an industrial symbiosis network. In this work, a clustering algorithm is developed to segregate the given plants into different clusters based on the concept of load deficits and surpluses of each plant. The concept ideally allows only the plants with surpluses to send out their unused sources and plants with deficits to only receive external sources/resources. The clusters are formed based on the distances between plants, which in turn helps in saving transportation and communication costs. The clustered plants are then easy to optimise and manage for resource and cost optimality. The applicability of the proposed clustering algorithm is demonstrated using two case studies from the domain of water recycling networks containing multiple contaminants with detailed network design, highlighting the importance of clustering in an industrial symbiosis network. It is observed that directing the excess flows from one plant to other plants in the same cluster can save a considerable amount of fresh resources. It implies that in the broader aspect, the developed methodology can address the optimisation of economic resources and can aid in the better management of overall resources for a large-scale industrial symbiosis network.

Antibody capture process based on magnetic beads from very high cell density suspension.

Cell clarification represents a major challenge for the intensification through very high cell density in the production of biopharmaceuticals such as monoclonal antibodies (mAbs). The present report proposes a solution to this challenge in a streamlined process where cell clarification and mAb capture are performed in a single step using magnetic beads coupled with protein A. Capture of mAb from non-clarified CHO cell suspension showed promising results; however, it has not been demonstrated that it can handle the challenge of very high cell density as observed in intensified fed-batch cultures. The performances of magnetic bead-based mAb capture on non-clarified cell suspension from intensified fed-batch culture were studied. Capture from a culture at density larger than 100 × 106 cells/ml provided an adsorption efficiency of 99% and an overall yield of 93% with a logarithmic host cell protein (HCP) clearance of ≈2-3 and a resulting HCP concentration ≤≈5 ppm. These results show that direct capture from very high cell density cell suspension is possible without prior processing. This technology, which brings significant benefits in terms of operational cost reduction and performance improvements such as low HCP, can be a powerful tool alleviating the challenge of process intensification.

Resilience, self-esteem, self-efficacy, social support, depression and ART adherence among people living with HIV in Sichuan, China.

ABSTRACTAIDS has had physical, psychological and social consequences on People living with HIV (PLWH) with the result that the challenges and adversity they face have significantly increased. Resilience helps individuals cope with these adversities and difficulties. For PLWH to face increased challenges and setbacks created by AIDS, they are required to have resilience. This paper presents research carried out in China aiming to examine the relationships among resilience, self-esteem, self-efficacy, social support, depression and antiretroviral therapy (ART) adherence in PLWH. A cross-sectional study using a convenience sample was conducted and 223 PLWH were recruited from 2 hospitals and 1 Center of Disease Control in Sichuan, China, from May to August 2018. The present research found that resilience was positively affected by self-esteem, self-efficacy and social support, and negatively predicted depression and positively predicted ART adherence. Resilience plays a mediating role between influential factors (self-esteem, self-efficacy and social support) and adaptive outcomes (depression and ART adherence). It suggests that resilience should be considered as a factor in intervention designed to reduce PLWH's depression and improve ART adherence. Improving self-esteem, self-efficacy and social support could enhance resilience.

The role of ozone combined with UVC/H2O2 process for the tertiary treatment of a real slaughterhouse wastewater.

The main goal of this work is to evaluate the usage of ozone (O3) as a pre-treatment or simultaneously combined with UVC/H2O2 process for the polishing stage treatment of real bio-treated slaughterhouse wastewater. Two different treatment strategies were tested: i) pre-ozonation of the wastewater followed by an UVC/H2O2 process (two-step treatment); ii) simultaneous application of O3/UVC/H2O2 combined process (one-step treatment). For the two-step strategy, the pre-treatment with 30 mg O3/min for 10 min reduces significantly total suspended solids (TSS), turbidity and colour, reducing light filtering effects and increasing the efficiency of the following UVC/H2O2 process. In turn, the one-step treatment strategy (O3/UVC/H2O2) allows a more efficient use of injected O3 by reducing the amount of O3 required (from 273 to 189 mg O3/Leffluent) to achieve similar mineralization levels. The real bio-treated slaughterhouse wastewater treated by O3/UVC/H2O2 process achieved final colour values of 20 Pt/Co, TSS of 35 mg/L and COD of 61 mg O2/L, allowing its direct discharge into water compartments according to European Council Directive 91/271/EEC.

Total Site Heat Integration benefiting from geothermal energy for heating and cooling implementations.

Geothermal energy is a promising renewable energy source that has been developed by many countries in recent years. It can be utilised to meet various energy demand. This paper studies the performance of integrating geothermal energy in the Locally Integrated Energy Sector (LIES). The heating and cooling demand of various processes should be satisfied, and heat among processes should be recovered. This is done by using Grand Composite Curves and Total Site Profiles to visually illustrate how much load is required for utility systems. The geothermal utility system and steam utility system are compared. The integration plan for geothermal energy under different temperatures are studied. An illustrative case shows that by using this type of renewable energy under a specific and favourable condition, above 70% of steam utility load can be saved. The working cycle of using a geothermal utility system is studied by using the Time Slice model. The heat recovery plan for normal operation, mineral scaling, and cleaning periods are optimised. The minimum temperature for heat storage can also be identified.

Urban and industrial symbiosis for circular economy: Total EcoSite Integration.

The paper presents an extension of Pinch Analysis and namely, Total Site Process Integration. It benefits from up to date developments and introduction of Total EcoSite Integration for urban and industrial symbiosis. An important development is Pinch Analysis for Solid Waste Integration which is a crucial step for the symbiosis in a circular economy. As the potential EcoSites are usually extensive and cover various units, a methodology based on clusters has been used. The solution has been supported by graphical tools using the analogy with already implemented extensions of Pinch Analysis. The results of a demonstration case study revealed the potential of the novel approach. The identified integrated design increased the energy recovered from the solid waste by 11.39 MWh/d and diverted 2 t/d of the waste from the landfill, benefiting both the urban and industrial site. The proposed approach is also capable of minimising the requirement of energy-intensive thermal drying for waste whenever the process allowed, subsequently offer a solution with lower environmental footprint and cost. For future work, a even more comprehensive case study can be conducted by considering the other forms of the waste, recovery process and drying approaches.

A Heat and Power Pinch for Process Integration targeting in hybrid energy systems.

Hybrid energy systems have been widely used for residential and industrial purposes. In this system, the total energy requirement of each unit can be met with heat and electricity. Pinch Analysis becomes a widely used tool for Process Integration, and using Pinch Analysis for Heat Integration is well-established. However, for the combined heat and power system, the theory and the corresponding tool deserve some more development. This paper extended the Pinch Analysis concept and proposed a Heat and Power Pinch Analysis to target the amount of heat that should be recovered from the hybrid energy system. Heat and Power Composite Curve (HPCC) is developed to visualise the total energy and the separated heat and power (electricity) requirement of a hybrid energy system in a working time period. The amount of outsourced electricity that should be purchased, and stored electricity at the startup period, and the extra electricity generated by the system at the end of the working period can be demonstrated. A case is studied to illustrate the steps of using this tool, two scenarios are discussed, and the targets are shown.

Anaerobic Digestion for Producing Renewable Energy-The Evolution of This Technology in a New Uncertain Scenario.

Anaerobic digestion is a well-known technology with wide application in the treatment of high-strength organic wastes. The economic feasibility of this type of installation is usually attained thanks to the availability of fiscal incentives. In this review, an analysis of the different factors associated with this biological treatment and a description of alternatives available in literature for increasing performance of the process were provided. The possible integration of this process into a biorefinery as a way for producing energy and chemical products from the conversion of wastes and biomass also analyzed. The future outlook of anaerobic digestion will be closely linked to circular economy principles. Therefore, this technology should be properly integrated into any production system where energy can be recovered from organics. Digestion can play a major role in any transformation process where by-products need further stabilization or it can be the central core of any waste treatment process, modifying the current scheme by a concatenation of several activities with the aim of increasing the efficiency of the conversion. Thus, current plants dedicated to the treatment of wastewaters, animal manures, or food wastes can become specialized centers for producing bio-energy and green chemicals. However, high installation costs, feedstock dispersion and market distortions were recognized as the main parameters negatively affecting these alternatives.

Recent development of unconventional aqueous biphasic system: characteristics, mechanisms and applications.

Aqueous biphasic system (ABS) is widely used in the recovery, extraction, purification and separation of proteins, enzymes, nucleic acids and antibodies. The ABS with high water content and low interfacial tension offers a biocompatible environment for the recovery of labile biomolecules. Process integration can be achieved using ABS by incorporating multiple-steps of purification, concentration and purification of biomolecules in a single-step operation which often results in high product recovery yield and purity. Conventional ABS is usually formed by aqueous solutions of two polymers or a polymer and a salt above a critical concentration. The high viscosity of polymer-based ABS causes slow phase separation and hinders the mass transfer of biomolecules, whereas polymer/salt ABS is characterized by high ionic strength resulting in the loss of bioactivity of recovered biomolecules. These limitations have encouraged the development of novel ABS which is more cost-effective for various biotechnological applications. This review discusses the characteristics and mechanisms of several types of emerging unconventional ABS using phase-forming components such as hyperbranched polymers, special salts, surfactants, magnetic fields, the addition of nanoparticles and incorporation of various solvent. Moreover, several novel applications of ABS for different separation purposes such as microfluidic-based ABS, ABS bioreactors, application of ABS as an analytical tool, and ABS micropatterning are discussed in this review. In the last section of this review, a comprehensive summary of process integration using ABS for extractive fermentations, bioconversion, crystallization and precipitation is also supplemented for the comprehensive review of various types and applications of ABS in recent years.

Recent advances and comparisons of conventional and alternative extraction techniques of phenolic compounds.

Phenolic compounds are a group of secondary metabolites produced by plants under stressful conditions. Phenolic compounds play an important role in the prevention and treatment of certain illnesses and are exploited by the food and pharmaceutical industries. Conventional methods are commonly used as models to compare the efficiencies of alternative extraction methods. Among alternative extraction processes, microwave-assisted extraction (MAE), pressurized liquid extraction (PLE), supercritical fluid extraction (SFE) and ultrasonic-assisted extraction (UAE) are the most studied. These methods produce extracts rich in phenolic compounds using moderate temperatures, short extraction times, and solvents generally recognized as safe. The combination of extraction time and temperature plays a critical role in the stability of the compounds. Solvents of higher polarity enhance the extraction of phenolic compounds. The use of the ethanol-water mixture for MAE, PLE, and UAE is recommended. MAE and UAE involve shorter extraction times than do PLE and SFE. SFE requires a low average temperature (40 °C). MAE produces the highest total phenolic content [227.63 mg GAE/g dry basis (d.b.)], followed by PLE (173.65 mg GAE/g d.b.), UAE (92.99 mg GAE/g d.b.) and SFE (37 mg GAE/g d.b.). Extraction yields and recovery rates of the phenolic compounds can be enhanced by combining and integrating extraction methods.

Xylitol bioproduction: state-of-the-art, industrial paradigm shift, and opportunities for integrated biorefineries.

Recent advances in biomass conversion technologies have shown a promising future toward fermentation during xylitol production. Xylitol is one of the top 12 renewable added-value chemicals that can be obtained from biomass according to US Department of Energy (USDOE). Currently, xylitol accounts for approximately US$823.6 million of annual sales in the market, and this amount is expected to reach US$1.37 billion by 2025. This high demand has been achieved owing to the chemical conversion of hemicellulosic hydrolysates from different lignocellulosic biomasses, which is a costly and non-ecofriendly process. Xylose-rich hemicellulosic hydrolysates are the major raw materials for xylitol production through either chemical or biotechnological routes. Economic production of a clean hemicellulosic hydrolysate is one of the major bottlenecks for xylitol production on the commercial scale. Advancements in biotechnology, such as the isolation of novel microorganisms, genetic manipulation of xylose metabolizing strains, and modifications in the fermentation process, can enhance the economic feasibility of xylitol production on the large scale. Furthermore, xylitol production in integrated biorefineries can be even more economic, given the readily available raw materials and the co-use of steam, electricity, and water, among others. Exploring new biotechnology techniques in integrated biorefineries would open new markets and opportunities for sustainable xylitol production to fulfill the market's growing demands for this sugar alcohol. This article is a review of the advancements reported in the whole biotechnological process for xylitol production, and involve pretreatment technologies, hemicellulosic hydrolysate preparation, xylose conversion into xylitol, and product recovery. Special attention is devoted to current metabolic engineering strategies to improve this bioprocess, as well as to the importance of xylitol production processes in biorefineries.

A hybrid method for synthesis of integrated water and regeneration networks with variable removal ratios.

This work presents a systematic optimization framework that integrates graphical insights with mathematical modelling to reduce both model complexity and computational time. The graphical technique adopted in this work is Composite Table Algorithm (CTA), which is improved to determine an optimal regenerator removal ratio (RR) that simultaneously minimizes the freshwater requirement and wastewater generation within the water network. The improved CTA is demonstrated using literature examples for both fixed load and fixed flowrate problems. It is further adapted to solve a multiple contaminant problem using the reference contaminant approach. The mathematical model developed includes a detailed design of a reverse osmosis (RO) unit to allow for simultaneous optimization of water and energy used by the regenerator. This provides accurate cost estimation of the water network rather than the linear cost functions associated with the use of blackbox representations in graphical targeting. Upon integrating the graphical and mathematical techniques in this study, results show that there was a reduction of about 85% in CPU time. This implies that the model converges faster and therefore favours the use of insight-based techniques as a preprocessing step for mathematical modelling.

Pre-disinfection columns to improve the performance of the direct electro-disinfection of highly faecal-polluted surface water.

This work presents the design and evaluation of a new concept of pre-disinfection treatment that is especially suited for highly polluted surface water and is based on the combination of coagulation-flocculation, lamellar sedimentation and filtration into a single-column unit, in which the interconnection between treatments is an important part of the overall process. The new system, the so-called PREDICO (PRE-DIsinfection Column) system, was built with low-cost consumables from hardware stores (in order to promote in-house construction of the system in poor countries) and was tested with a mixture of 20% raw wastewater and 80% surface water (in order to simulate an extremely bad situation). The results confirmed that the PREDICO system helps to avoid fouling in later electro-disinfection processes and attains a remarkable degree of disinfection (3-4 log units), which supplements the removal of pathogens attained by the electrolytic cell (more than 4 log units). The most important sizing parameters for the PREDICO system are the surface loading rate (SLR) and the hydraulic residence time (HRT); SLR values under 20 cm min-1 and HRT values over 13.6 min in the PREDICO system are suitable to warrant efficient performance of the system.

Process integration of crude oil distillation with technological and economic restrictions.

The petrochemical industry is one of the most important industries in the world economy. In the largest oil-producing countries, more than half of GDP is generated by hydrocarbons production and refining. Reduction of oil prices and new environmental regulations are forcing petrochemical companies to improve their energy efficiency. Improvement of the energy efficiency of Crude oil distillation process at atmospheric vacuum distillation unit (AVDU) with a capacity of 3.3 million ton per year is considered in this paper. The amount of fuel spent for reprocessing of one ton of crude oil has been defined and it is 3.79 kg of natural gas. This paper shows the ways to achieve the objectives of retrofit in the context of administrative and technical restrictions. The retrofit goal was achieved by the retrofit of the heat exchange network, which allowed reducing gas consumption by 0.98 t/h (natural gas). The provided case studies show the pathway for efficient retrofit of crude oil distillation and most profitable ways for investment taking into account various administrative and technical constraints. The results of this work allow achieving reduction of energy consumption by 26%.