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Therapeutic oligonucleotides (ONs) have great potential to treat many diseases due to their ability to regulate gene expression. However, the inefficiency of standard purification techniques to separate the target sequence from molecularly similar variants is hindering development of large scale ON manufacturing at a reasonable cost. Multicolumn Countercurrent Solvent Gradient Purification (MCSGP) is a valuable process able to bypass the purity-yield tradeoff typical of single-column operations, and hence to make the ON production more sustainable from both an economic and environmental point of view. However, operating close to the optimum of MCSGP can be challenging, resulting in unstable process performance and in a drift in product quality, especially when running a continuous process for extended periods where process parameters such as temperature are prone to variation. In this work, we demonstrate how greater process robustness is introduced in the design and execution of MCSGP for the purification of a 20mer single-stranded DNA sequence through the implementation of UV-based dynamic control. With this novel approach, the cyclic steady state was reached already in the third cycle and disturbances coming from fluctuations in the feed quality, loading amount and temperature were effectively compensated allowing a stable operation close to the optimum. In response to the perturbations, the controlled process kept the standard deviation on product recovery below 3.4%, while for the non-controlled process it increased up to 27.5%.

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Spent Coffee Grounds (SCG) is an agricultural residue obtained in a large quantity from local cafes in Thailand. In order to handle this waste effectively, the valorization of SCG is essential. SCG consists of beneficial phenolic compounds with antioxidative properties and caffeine, which can be recovered through extraction followed by separation and purification processes. In this work, water extraction of SCG was carried out. The volumetric composition of the liquid extract of SCG was then adjusted with an organic solvent, and the obtained mixture was used as the feed for subsequent separation. For the separation method of the SCG extract, a single chromatographic column was employed to separate a group of phenolic compounds (represented by chlorogenic acid) and a group of contaminants (represented by caffeine). The volumetric composition of the mobile phase was varied to determine the condition suitable for the separation of chlorogenic acid and caffeine in a C18 column. Adsorption parameters were determined and used to formulate the mathematical models describing the adsorption dynamics of those two bioactive compounds in the experimental breakthrough curves of standard solutions and the liquid extract of SCG. Furthermore, the three-zone simulated moving bed system (TZ-SMB) was designed to continuously separate fractions of chlorogenic acid and caffeine in the liquid extract of SCG. The adsorption parameters were employed in the optimization of TZ-SMB operating conditions using triangle theory, conducted via computer simulation. The experimental result of water extraction revealed that the yields of chlorogenic acid and caffeine were 0.292 and 0.583 mg/g dried SCG, respectively, using solid-to-liquid ratio of 1 g: 30 mL and temperature of 75 °C. The separation result in a single chromatographic column showed that the mobile phase consisting of acetonitrile, water, and formic acid (10: 90: 1.5 vol%) provided the linear adsorption isotherms for both chlorogenic acid and caffeine, and the chromatographic peaks of all compounds in the liquid extract of SCG were well separated. The simulated results of TZ-SMB at the optimal point revealed that the flow rates of desorbent, feed, extract product, and raffinate product were 0.626, 0.115, 0.081, and 0.593 mL/min, respectively, with the switching time of 20 min. At this point, the relative purities of caffeine in the extract product and chlorogenic acid in the raffinate product were 99.45 % and 98.88 %, respectively, with the maximum productivity of 0.045 mg/mL⋅h. In addition, for demonstration purposes, the lab-scale TZ-SMB experiment was conducted to show the separation of chlorogenic acid and caffeine in the liquid extract of SCG. The operating point from the triangle separation region was chosen based on the sensitivity of flow rate that ensured the criteria of purity. The experimental results showed that the relative purities of caffeine in extract product and chlorogenic acid in raffinate product were both 100 %, verifying the successful separation.

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This work is to design and optimize the 4column-simulated moving bed unit to separate and recovery alkali and lignin from black liquor. Since both alkali lignin and NaOH are the main products, we proposed the 'extended separation volume' methodology to construct a four-dimensional separation region (QI×QII×QIII×QIV) to design the operating conditions. Special attention was paid to investigate the influence of flow-rates in each zone on the performance parameters of these two products. The results show besides QII and QIII, QI and QIV can affect the concentrations and productivities significantly and have to be taken into account. Afterwards, the Response Surface Methodology, conducted by Design-Expert® was chosen to deal with the multi-objective optimization problem. The unique optimal operating conditions were finally obtained with the concentration of 74.7% and 48.1% of feed solution, and the productivity of 3.9 and 0.7 g/(L·h)-1 for alkali lignin and NaOH. Considering the actual operation, the productivity of 2.5 and 0.446 g/(L·h)-1 were obtained by experiment of internal actual operation point.

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Conventional Simulated Moving Bed Chromatography (C-SMBC) characterized by a single operating point comprising of the internal flow rates and switching period, yields a single fraction of the extract and raffinate, at cyclic steady state. In the current work, we deal with two-fraction yielding Dual Switch SMBC (DS-SMBC) characterized by two operating points implemented during alternate switches. The higher degrees of freedom in Dual Switch SMBC give it the potential of obtaining a higher average extract purity, for one of its two fractions, compared to conventional SMBC, at a certain minimum feed flow rate and minimum average extract recovery. This becomes possible with a trade-off in the average purity of the other extract fraction of Dual Switch SMBC, without any addition to the conventional SMBC equipment. We solve a multi-objective optimization problem wherein we maximize the average purity of one of the extract fractions of Dual Switch SMBC, with the requirement of certain minimum feed flow rate, minimum average extract recovery and minimum average purity of the non-objective extract fraction. One of the aims of the current work is deduction of the working principle of Dual Switch SMBC, that facilitates superior average extract purity. Next, we analyze the trade-offs due to the said process bounds on the objective i.e. average purity of objective extract fraction of Dual Switch SMBC. This has been done for the two case studies of separation: 1) linear isotherm based separation of fructose-glucose in deionized water on cation exchange resin 2) non-linear isotherm based separation of enantiomers of Tröger's base in ethanol on microcrystalline cellulose triacetate. The findings from the trade-offs in the multi-objective optimization problem, further corroborate our understanding of the working principle.

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The separation of p-xylene (PX) and m-xylene (MX) isomers with near boiling points is a worldwide problem. The metal-organic framework material is an ideal stationary phase for chromatographic separation because of its high porosity, homogeneous pore diameter and good chemical stability. In this paper, a simulated moving bed (SMB) chromatography system with MIL-53(Fe) as the stationary phase and petroleum ether-dichloromethane as the mobile phase was designed to separate PX and MX at ambient temperature. Firstly, according to the elution curves of a single column, nonlinear competitive Langmuir adsorption isotherm equation was confirmed by equilibrium dispersive chromatography model. Then, the SMB separation zone was determined based on triangle theory, and the SMB operating conditions were optimized. Finally, the purity, recovery and productivity of PX reached 100.0%, 99.1% and 93.1 g/L/h, respectively; the purity, recovery and productivity of MX reached 96.4%, 100.0% and 23.5 g/L/h, respectively; the solvent consumption was 0.42 L/g.

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In the modeling of a simulated moving bed, several assumptions are considered, the key assumption is there are no radial concentration gradients based on perfect mixing. However, it is difficult to achieve perfect mixing because the injected flowrate of the bed is periodically changed in the process. In this study, the performance of the simulated moving bed process was analyzed when the injected flow such as the feed or desorbent stream was unevenly distributed. To this end, the distribution function of the injected flow was calculated and applied to the model. Two types of distribution functions were obtained using the experimental results of a previous study, and the simulation results were compared with classical modeling assuming perfect mixing. In the base case simulation, the purity was similar in all cases, the productivity was higher more than 5% in the even distribution case compared to the most uneven distribution case. The effect of distribution was analyzed through sensitivity analysis by changing the overall flow rate, switching time, bed length, and flow rate of sections 2 and 3. As a result, regardless of the distribution applied, the trends of the performance parameters were the same. However, the more uneven the distribution, the greater the difference in productivity, recovery, and desorbent consumption compared to the even distribution case. It was confirmed that the design that distributes the injected flow more evenly has a better performance.

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The use of magnetic nanoparticles shows a steadily increasing technical importance. Particularly in medical technology disciplines such as cancer treatment, the potential of these special particles is increasing rapidly. Magnetic nanoparticles are particles with a submicron size, and consist mostly of magnetite-containing composites. An important quality parameter of such particles is a particle size distribution as narrow as possible, which can only be obtained to a certain degree by synthesis. Apart from ultracentrifugation, there are so far only methods on an analytical scale to narrow the size distribution as a post-processing step. We present a method based on magnetic chromatography, by which high separation efficiencies at yields of up to 99.9% are achieved. The novel technique is based on a competition between the magnetic interaction of the nanoparticles and the separation matrix, as well as the hydrodynamic forces. Furthermore, the method is extended using a continuous mode, namely simulated moving bed chromatography, to obtain potent space-time yields of up to 2.94 g/(L*h). For those reasons, this novel continuous magnetic chromatography method offers high potential for large-scale refinement of magnetic nanoparticles while fulfilling sophisticated quality criteria for high-technology applications.

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The VariCol and ModiCon processes are two variants of the simulated moving bed (SMB) process, characterized by the modulation of the length of zones of the chromatographic column train and the feed concentration. These features give more flexibility than the conventional operation, leading to essential improvements in the separation and purification of mixtures. The optimal performance comparison of these two variants, the hybrid formed by their combination, and the conventional SMB process are scarce in the literature. This comparison helps discover new characteristics of each single and combined operation mode and creates guidelines to select the appropriate operation mode for possible real applications. In this work, the performance comparison of the ModiCon, VariCol, ModiCon+VariCol, and SMB processes is carried out in terms of maximal throughput for specific product purity values. Particular emphasis is placed on both the ModiCon and the hybrid ModiCon+VariCol processes characteristics. A strategy for combining and optimizing the ModiCon and the VariCol processes was determined. As a case study, the enantioseparation of guaifenesin was considered. In the ModiCon process, more than two modulation subintervals did not improve the performance in the separation. The optimal pattern, based on two subintervals, has zero feed concentration in the first subinterval and the maximal concentration in the second one. The best result for the hybrid operation (ModiCon+VariCol) was reached when the feed port moves simultaneously as the SMB process switching period. The optimal throughput of the ModiCon and the ModiCon+VariCol processes was almost doubled than that of the SMB process. These performances were based on larger zones I and II and not in zones II and III as occur with the SMB and VariCol process. The throughput in the hybrid operation increases more significantly than the ModiCon process when 5 columns were considered instead of 6. The hybrid operation could be more attractive for a system with a few numbers of columns.

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Spent sulfite liquor (SSL), a waste stream from wood pulp production, has great potential as carbon source for future industrial fermentations. In the present study, SSL was separated into a hemicellulose derived sugar syrup (HDSS) and a lignosulfonic fraction by simulated moving bed chromatography. The recovery of SSL sugars in the HDSS was 89% and the fermentation inhibitors furfural, 5-hydroxymethylfurfural and acetic acid were removed by 98.7%, 60.5% and 75.5%, respectively. The obtained sugars have been converted to L-lactic acid, a building block for bioplastics, by fermentation with the lactic acid bacterium Enterococcus mundtii DSM4838. Batch fermentations on HDSS produced up to 56.3 g/L L-lactic acid. Simultaneous conversion of pentose and hexose sugars during fed-batch fermentation of wildtype E. mundtii led to 87.9 g/L optically pure (>99%) L-lactic acid, with maximum productivities of 3.25 g/L.h and yields approaching 1.00 g/g during feeding phase from HDSS as carbon source.

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Optimal control of a simulated moving bed (SMB) process is challenging because the system dynamics is represented as nonlinear partial differential-algebraic equations combined with discrete events. In addition, product purity constraints are active at the optimal operating condition, which implies that these constraints can be easily violated by disturbance. Recently, artificial intelligence techniques have received significant attention for their ability to address complex problems, involving a large number of state variables. In this study, a data-based deep Q-network, which is a model-free reinforcement learning method, is applied to the SMB process to train a near-optimal control policy. Using a deep Q-network, the control policy of a complex dynamic system can be trained off-line as long as a sufficient number of data is provided. These data can be efficiently generated by performing numerical simulations in parallel on multiple machines. The on-line computation of the control input using a trained Q-network is fast enough to satisfy the computational time limit for the SMB process. However, because the Q-network does not predict the future state, it is not possible to explicitly impose state constraints. Instead, the state constraints are indirectly imposed by providing a relatively large penalty (negative reward) when the constraints are violate. Furthermore, logic-based switching control is utilized to limit the ranges of the extract and raffinate purities, which helps to satisfy the state constraints and reduce the regions in the state space for reinforcement learning to explore. The simulation results demonstrate the advantages of applying deep reinforcement learning to control the SMB process.

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In downstream processing, large-scale chromatography plays an important role. For its development, screening experiments followed by pilot-plant chromatography are mandatory steps. Here we describe fast, simple, and inexpensive methods for establishing a preparative chromatography for the separation of complex protein mixtures, based on sample displacement batch chromatography. The methods are demonstrated by anion-exchange chromatography of a human plasma protein fraction (Cohn IV-4), including the screening step and upscaling of the chromatography by a factor of one hundred. The results of the screening experiments and the preparative chromatography are monitored by SDS-PAGE electrophoresis. In summary, we provide a protocol, which should be easily adaptable for the chromatographic large-scale purification of other proteins, in the laboratory as well as in the manufacturing of biopharmaceuticals. These protocols cover the initial piloting steps for establishing a large-scale sample batch chromatography. The results from the piloting steps may also be applied for packed columns for performing simulated-moving-bed (SMB) chromatography rather than batch chromatography.

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The VariCol process is a variant of the conventional simulated moving bed (SMB) process, distinguished by the asynchronous shifting of the inlet and outlet ports of the chromatographic column train. This feature allows for a more flexible operation in column utilization and can also achieve higher separation performances. However, to take full benefit out of it, the operating parameters, such as the strategy for port switching, must be optimal. in this paper, a novel methodology for optimizing those parameters, based on a single NLP (non-linear programming), is proposed. The main advantage of this approach is that it significantly reduces the complexity of the original MINLP (mixed-integer non-linear programming) formulation currently discussed in the literature. The proposed optimization problem is built, considering that the average column configuration of three zones provides the necessary and sufficient information to describe the VariCol process. Several optimization scenarios for the enantioseparation of 1,1´-bi-2-naphthol and aminoglutethimide were considered to evaluate the proposed methodology and to compare the performance of VariCol and SMB processes. The results have shown that with the single NLP approach, it is possible to explore the optimal solution in all the VariCol process domains with less computational effort than other optimization strategies reported in the literature. That is a great advantage, especially in the context of real-time applications.

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The separation of the proteins Bovine Serum Albumin (BSA) and Myoglobin (Mb) was achieved by Size-Exclusion Simulated Moving Bed (SE-SMB) and performed experimentally in the FlexSMB® unit, an SMB unit designed and built in the Laboratory of Separation and Reaction Engineering. Before accomplishing the separation experiments in the mentioned unit, separation regions were computed by simulation based on a phenomenological mathematical model to determine appropriate operating conditions. The developed model was validated in advance, against fixed-bed dynamic adsorption experimental results, for pure component and binary mixtures. Then the SMB experiments were carried out, and purities of the Mb on the extract and BSA on the raffinate streams were 98% and 96%, respectively. The achieved recoveries were 80% of Mb on the extract and 94% of BSA on the raffinate. Lastly, productivities of 6.4 gprotein⋅lads-1⋅day-1 for the extract and 28.8 gprotein⋅lads-1⋅day-1 for the raffinate were obtained.

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Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are essential fatty acids for human body, which are widely used in the field of healthy food and medicine. Meanwhile, there are some differences in their physiological functions, such as "scavenger for blood vessel" of EPA and "brain protector" of DHA. In order to make full use of EPA and DHA, it is necessary to prepare their high-purity component. In this paper, EPA and DHA were separated and purified by three-zone simulated moving bed (SMB) chromatography with C18 used as stationary phase and ethanol-water as mobile phase. For the single column experiment, a separation unit of SMB, the effects of the ratio of ethanol to water, pH value and temperature on the separation were investigated. The equilibrium dispersion (ED) model was used to obtain the adsorption parameters of EPA and DHA by inverse method and genetic algorithm, and the accuracy of the adsorption parameters was verified by fitting the overloaded elution curves under different conditions. Based on the acquired nonlinear adsorption isotherms the complete separation region was found according to triangle theory. The effects of sample concentration, flow ratios of adsorption zone and rectification zone, and column distribution mode of SMB on the separation were investigated. Under the optimized SMB conditions, the experimental result was that without regard to the other components, the chromatographic purity and recovery values of EPA and DHA exceeded 99% with the productivity of 4.15 g/L/h, and the solvent consumption of 1.11 L/g.

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Process streams of agro-food industries are often large and viscous. In order to fractionate such a stream the viscosity can be reduced by either a high temperature or dilution, the former is not an option in case of temperature sensitive components. Such streams are diluted prior to chromatographic fractionation, resulting in even larger volumes and high energy costs for sub-sequential water removal. The influence of feed viscosity on the performance of simulated moving bed chromatography has been investigated in a case study of the recovery of a γ-aminobutyric acid rich fraction from tomato serum. This work addresses the chromatographic system design, evaluates results from a pilot scale operation, and uses these to calculate the productivity and water use at elevated feed concentration. At the two higher feed viscosities (2.5 and 4 mPa·s) water use is lower and productivity higher, compared to the lowest feed viscosity (1 mPa·s). The behavior of the sugars for different feed viscosities can be described well by the model using the ratio of feed to eluent as dilution factor. The behavior of γ-aminobutyric acid is highly concentration dependent and the recovery could not be accurately predicted.

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Black chokeberry (Aronia melanocarpa) fruits are rich in anthocyanins, which are vital secondary metabolites that possess antioxidative properties. The aim of this study was to isolate and purify the anthocyanins from black chokeberry by simulated moving bed (SMB) chromatography, and to investigate the neuroprotective effect of SMB purified anthocyanin against Aß-induced memory damage in rats. The parameters of the SMB process were studied and optimized. Anthocyanin extracts were identified by HPLC and UPLC-QTOF-MS, and antioxidant abilities were evaluated. The Aß-induced animal model was established by intracerebral ventricle injection in rat brain. Through the SMB purification, anthocyanins were purified to 85%; cyanidin 3-O-galactoside and cyanidin 3-O-arabinoside were identified as the main anthocyanins by UPLC-QTOF-MS. The SMB purified anthocyanins exhibited higher DPPH and ABTS free radical scavenging abilities than the crude anthocyanins extract. Furthermore, rats receiving SMB purified anthocyanins treatment (50 mg/kg) showed improved spatial memory in a Morris water maze test, as well as protection of the cells in the hippocampus against Aß toxicity. These results demonstrate that anthocyanins could serve as antioxidant and neuroprotective agents, with potential in the treatment of Alzheimer's disease.

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For the high-purity production of acetoin or 2,3-butanediol (BD) from related fermentation processes, it is essential to accomplish a detailed separation between acetoin and BD in an economical mode. To address this issue, we aimed to develop a highly-efficient simulated-moving-bed (SMB) process for the continuous-mode separation of acetoin from BD with high purity and small loss. As a first step for this task, the adsorption and mass-transfer parameters of acetoin and BD on a proven adsorbent were estimated while assuming that BD isomers (meso-BD and DL-BD) would be identical in adsorption and mass-transfer behaviors. The resultant parameters from such estimation were applied to the optimal design of the acetoin-BD separation SMB. The designed SMB was then experimentally investigated, which revealed that some sign of BD isomerism occurred in the SMB column-profile data and thus had an adverse effect on the SMB separation performance. To resolve this problem, the individual parameters of BD isomers were determined on the basis of the SMB column-profile data and an inverse-method principle. The resulting parameters of BD isomers were used in the re-design of the target SMB, which was then experimentally checked for its separation performance. It was confirmed that such SMB re-designed in consideration of BD isomerism was quite effective in the continuous-mode separation of acetoin from BD with high purity (> 99.2%) and small loss (< 1.52%).

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A new approach for determining optimal operating conditions for simulated moving bed chromatographic processes is presented. The method is based on recursive online estimation and requires only rough initial estimates. It is based on a simple foot point model of the moving concentration fronts and an online measurement of the corresponding retention times in the different zones of the plant. A mathematical representation of the adsorption isotherms is not required. The method is validated experimentally for the separation of bicalutamide enantiomers.

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For a simulated moving bed (SMB) chromatographic process, it has been common knowledge that as desorbent usage increases, the level of separation capability (i.e., the purity and recovery of a target product) continues to improve and then eventually reaches almost a constant level. To check whether there are any exceptional cases concerning such a generally accepted relationship, the effect of desorbent usage on the recovery of a target product under the condition of high purity was investigated using a three-zone SMB process for separation of xylobiose (X2) from a ß-xylosidase reaction output. It was found that the considered SMB represented some unusual relationship between desorbent usage and X2 recovery, which also took quite a different pattern according to the searching region for optimal SMB operation parameters. If the optimal operation parameters are determined in the region of positive flow-rate-ratios ("m+ approach"), the use of a larger amount of desorbent than required to make X2 recovery reach a constant level can rather lead to a little reduction in X2 recovery. If the optimal operation parameters are selected in the region of negative flow-rate-ratios ("m- approach"), there exists an optimal desorbent usage, beyond which a further increase in desorbent usage can bring about a significant reduction in X2 recovery. Comparison of the two design approaches reveals that the m- approach can lead to higher X2 recovery and much lower desorbent usage than the m+ approach. Furthermore, such merits of the m- approach over the m+ approach became greater with increasing the SMB throughput.

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The pseudobinary preparative separation of nadolol stereoisomers is performed by simulated moving bed chromatography (SMB). Using the Chiralpak IA adsorbent, a new 25:75:0.1 (v/v/v) methanol-acetonitrile-diethylamine solvent composition was selected to perform the experimental SMB separation and compare it with the previous results obtained using pure methanol. Using a 2 g L-1 total feed concentration of an equimolar mixture of the four stereoisomers of nadolol, the more retained component was fully recovered (100% purity and 100% recovery), with a system productivity of 0.77 g L-1  hour-1 and a solvent consumption of 9.62 L g-1 . Comparing these results with the ones previously reported using 100:0.1 methanol-diethylamine solvent composition, this work shows that the 25:75:0.1 methanol-acetonitrile-diethylamine is a better alternative for the preparative separation of nadolol stereoisomers by SMB chromatography. These results are confirmed by simulation of the SMB operation for higher feed concentrations, by comparing the performances of the two solvent compositions using the data obtained experimentally through the measurement of the adsorption equilibrium isotherms and the kinetic data obtained for both solvents. The new experimental and simulation results stress out that the performance of the preparative separation can be improved by a careful selection of the solvent composition.