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INTRODUCTION: Astragaloside IV (AS-IV) is an index for the quality evaluation of the traditional Chinese medicine Astragalus and an important material basis for Astragalus to exert its medicinal effects, and it is difficult to obtain a single AS-IV by ordinary separation methods. OBJECTIVE: To find a new isolation method that can prepare AS-IV quickly and efficiently. METHODOLOGY: AS-IV was isolated from Astragalus membranaceus extract by high-speed countercurrent chromatography using a two-phase solvent system consisting of ethyl acetate/n-butanol/water (4.2:0.8:5, v/v) at a speed of 950 rpm at a flow rate of 2 mL/min using one of the high-speed countercurrent chromatographic sequential injection models developed during the previous study. RESULTS: Compared with the common countercurrent chromatographic separation, this separation method increased the injection volume and yield by 4-fold and 4.47-fold, respectively, with only about 1.2-fold increase in solvent consumption and separation time, and the purity was basically not reduced, and 55.9 mg of AS-IV, with a purity of 96.95%, was finally prepared from 400 mg of the crude extract in 240 min. CONCLUSION: The continuous injection mode of high-speed countercurrent chromatography was able to successfully prepare a large amount of AS-IV with high purity at one time.

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We introduce magnetophoresis-based microfluidics for sorting biological targets using positive Magnetophoresis (pM) for magnetically labeled particles and negative Magnetophoresis (nM) for label-free particles. A single, externally magnetized ferromagnetic wire induces repulsive forces and is positioned across the focused sample flow near the main channel's closed end. We analyze magnetic attributes and separation performance under two transverse dual-mode magnetic configurations, examining magnetic fields, hydrodynamics, and forces on microparticles of varying sizes and properties. In pM, the dual-magnet arrangement (DMA) for sorting three distinct particles shows higher magnetic gradient generation and throughput than the single-magnet arrangement (SMA). In nM, the numerical results for SMA sorting of red blood cells (RBCs), white blood cells (WBCs), and prostate cancer cells (PC3-9) demonstrate superior magnetic properties and throughput compared to DMA. Magnetized wire linear movement is a key design parameter, allowing device customization. An automated device for handling more targets can be created by manipulating magnetophoretic repulsion forces. The transverse wire and magnet arrangement accommodate increased channel depth without sacrificing efficiency, yielding higher throughput than other devices. Experimental validation using soft lithography and 3D printing confirms successful sorting and separation, aligning well with numerical results. This demonstrates the successful sorting and separating of injected particles within a hydrodynamically focused sample in all systems. Both numerical and experimental findings indicate a separation accuracy of 100% across various Reynolds numbers. The primary channel dimensions measure 100 µm in height and 200 µm in width. N52 permanent magnets were employed in both numerical simulations and experiments. For numerical simulations, a remanent flux density of 1.48 T was utilized. In the experimental setup, magnets measuring 0.5 × 0.5 × 0.125 inches and 0.5 × 0.5 × 1 inch were employed. The experimental data confirm the device's capability to achieve 100% separation accuracy at a Reynolds number of 3. However, this study did not explore the potential impact of increased flow rates on separation accuracy.

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Spontaneous separation of immiscible organic droplets has substantial research implications for environmental protection and resource regeneration. Compared to the widely explored separation of oil-water mixtures, there are fewer reports on separating mixed organic droplets on open surfaces due to the low surface tension differences. Efficient separation of mixed organic liquids by exploiting the rapid spontaneous transport of droplets on open surfaces remains a challenge. Here, through the fusion of inspiration from the fast droplet transport capability of Sarracenia trichome and the asymmetric wedge channel structure of shorebird beaks, this work proposes a spine with hierarchical microchannels and wedge channels (SHMW). Due to the synergistic effect of capillary force and asymmetric Laplace force, the SHMW can rapidly separate mixed organic droplets into two pure phases without requiring additional energy. In particular, the self-spreading of the oil solution on the open channel surface is utilized to amplify the surface energy difference between two droplets, and SHMW achieves the pickup of oil droplets floating on the surface of the organic solution. The maximum separation efficiency on 3-SHMW can reach 99.63%, and it can also realize the antigravity separation of mixed organic droplets with a surface tension difference as low as 0.87 mN·m-1. Furthermore, SHMW performs controllable separation, oil droplet pickup, and continuous separation and collection of mixed organic droplets. It is expected that this cooperative structure composed of hierarchical microchannels and wedge channels will be realized in resource recovery or chemical reactions in industrial production processes.

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Organelle size varies with normal and abnormal cell function. Thus, size-based particle separation techniques are key to assessing the properties of organelle subpopulations differing in size. Recently, insulator-based dielectrophoresis (iDEP) has gained significant interest as a technique to manipulate sub-micrometer-sized particles enabling the assessment of organelle subpopulations. Based on iDEP, we recently reported a ratchet device that successfully demonstrated size-based particle fractionation in combination with continuous flow sample injection. Here, we used a numerical model to optimize the performance with flow rates a factor of three higher than previously and increased the channel volume to improve throughput. We evaluated the amplitude and duration of applied low-frequency DC-biased AC potentials improving separation efficiency. A separation efficiency of nearly 0.99 was achieved with the optimization of key parameters-improved from 0.80 in previous studies (Ortiz et al. Electrophoresis, 2022;43;1283-1296)-demonstrating that fine-tuning the periodical driving forces initiating the ratchet migration under continuous flow conditions can significantly improve the fractionation of organelles of different sizes.

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A novel and meaningful theoretical model was established with counter-current chromatography based on the elution-extrusion mode for efficient continuous separation. For the experimental verification of the theory, the separation of the binary mixture luteolin/baicalein was studied. The velocity model and volume model of the chromatographic separation behavior of the target compounds in the separation process were given by theoretical analysis. The results showed that this method had obvious advantages in the separation of binary mixtures. In addition, the established model was used to predict and isolate oleuropein from olive leaves. A two-phase solvent system of n-butanol/ethyl acetate/methanol/water (1:19:1:19, v/v/v/v) was chosen for the continuous separation of oleuropein. After optimizing the conditions in this way, a large amount of sample loading was achieved; the volume of injections can reach 48 ml, approximately 35.29% of the volume of the counter-current chromatography column, and oleuropein with a purity of 86.42% was obtained within 80 min. The model provided technical support for the prediction of chromatographic behavior and operating parameters during continuous separation and preparation of counter-current chromatography. It has great application prospects and significance in separation preparation, especially in large-scale industrial preparation.

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Isolation of exosome from culture medium in an effective way is desired for a less time consuming, cost saving technology in running the diagnostic test on cancer. In this study, we aim to develop an inertial microfluidic channel to separate the nano-size exosome from C666-1 cell culture medium as a selective sample. Simulation was carried out to obtain the optimum flow rate for determining the dimension of the channels for the exosome separation from the medium. The optimal dimension was then brought forward for the actual microfluidic channel fabrication, which consisted of the stages of mask printing, SU8 mould fabrication and ended with PDMS microchannel curing process. The prototype was then used to verify the optimum flow rate with polystyrene particles for its capabilities in actual task on particle separation as a control outcome. Next, the microchip was employed to separate the selected samples, exosome from the culture medium and compared the outcome from the conventional exosome extraction kit to study the level of effectiveness of the prototype. The exosome outcome from both the prototype and extraction kits were characterized through zetasizer, western blot and Transmission electron microscopy (TEM). The microfluidic chip designed in this study obtained a successful separation of exosome from the culture medium. Besides, the extra benefit from this microfluidic channels in particle separation brought an evenly distributed exosome upon collection while the exosomes separated through extraction kit was found clustered together. Therefore, this work has shown the microfluidic channel is suitable for continuous separation of exosome from the culture medium for a clinical study in the future.

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Forward osmosis (FO) driven by osmotic pressure difference has great potential in water treatment. However, it remains a challenge to maintain a steady water flux at continuous operation. Herein, a FO and photothermal evaporation (PE) coupling system (FO-PE) based on high-performance polyamide FO membrane and photothermal polypyrrole nano-sponge (PPy/sponge) is developed for continuous FO separation with a steady water flux. The PE unit with a photothermal PPy/sponge floating on the surface of draw solution (DS) can continuously in situ concentrate DS by solar-driven interfacial water evaporation, which effectively offsets the dilution effect due to the injected water from FO unit. A good balance between the permeated water in FO and the evaporated water in PE can be established by coordinately regulating the initial concentration of DS and light intensity. As a consequence, the polyamide FO membrane exhibits a steady water flux of 11.7 L m-2 h-1 over time under FO coupling PE condition, effectively alleviating the decline in water flux under FO alone. Additionally, it shows a low reverse salt flux of 3 g m-2 h-1. The FO-PE coupling system utilizing clean and renewable solar energy to achieve a continuous FO separation is significantly meaningful for practical applications.

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Efficient and rapid removal of p-arsanilic acid (p-ASA) in water is very important in environmental protection and human health, however it is still a severe challenge in actual engineering. Herein, a novel sorbent (CF-PEI) was successfully fabricated by simply modifying the amphiphilic skin collagen fiber (CF) substrate with Polyethylenimine (PEI). The as-prepared CF-PEI exhibits high-efficiency adsorption for negatively charged p-ASA with aromatic rings due to the introduction of amino groups and the existence of hydrophobic bands, and the maximum adsorption capacity of CF-PEI for p-ASA was high up to 285.71 mg g-1. In addition, the adsorption mechanism of CF-PEI on p-ASA mainly includes electrostatic interaction, hydrogen bond and amphiphilicity. The multi-level all-fiber structure of CF makes it mainly focus on surface mass transfer with short mass transfer distance, and its capillary drainage effect can realize large flow and rapid separation. CF-PEI based on CF can realize the ability to separate low-concentration p-ASA with high flow rate and high efficiency. The effective processing volume was 12.5 L g-1 when the separation flux reached as high as 9931.27 L m-2 h-1. Notably, the p-ASA adsorbed on CF-PEI was almost completely eluted by NaOH (0.5 mol L-1). The adsorbent is convenient to prepare, recyclable, high in efficiency, and has a great application prospect in removing organic micro-pollutants.

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Heterogeneity in organelle size has been associated with devastating human maladies such as neurodegenerative diseases or cancer. Therefore, assessing the size-based subpopulation of organelles is imperative to understand the biomolecular foundations of these diseases. Here, we demonstrated a ratchet migration mechanism using insulator-based dielectrophoresis in conjunction with a continuous flow component that allows the size-based separation of submicrometer particles. The ratchet mechanism was realized in a microfluidic device exhibiting an array of insulating posts, tailoring electrokinetic and dielectrophoretic transport. A numerical model was developed to elucidate the particle migration and the size-based separation in various conditions. Experimentally, the size-based separation of a mixture of polystyrene beads (0.28 and 0.87 µ$\umu $ m) was accomplished demonstrating good agreement with the numerical model. Furthermore, the size-based separation of mitochondria was investigated using a mitochondria mixture isolated from HepG2 cells and HepG2 cells carrying the gene Mfn-1 knocked out, indicating distinct size-related migration behavior. With the presented continuous flow separation device, larger amounts of fractionated organelles can be collected in the future allowing access to the biomolecular signature of mitochondria subpopulations differing in size.

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Magnetic nanoparticles are researched intensively not only for biomedical applications, but also for industrial applications including wastewater treatment and catalytic processes. Although these particles have been shown to have interesting surface properties in their bare form, their magnetisation remains a key feature, as it allows for magnetic separation. This makes them a promising carrier for precious materials and enables recovery via magnetic fields that can be turned on and off on demand, rather than using complex (nano)filtration strategies. However, designing a magnetic separator is by no means trivial, as the magnetic field and its gradient, the separator dimensions, the particle properties (such as size and susceptibility), and the throughput must be coordinated. This is showcased here for a simple continuous electromagnetic separator design requiring no expensive materials or equipment and facilitating continuous operation. The continuous electromagnetic separator chosen was based on a current-carrying wire in the centre of a capillary, which generated a radially symmetric magnetic field that could be described using cylindrical coordinates. The electromagnetic separator design was tested in-silico using a Lagrangian particle-tracking model accounting for hydrodynamics, magnetophoresis, as well as particle diffusion. This computational approach enabled the determination of separation efficiencies for varying particle sizes, magnetic field strengths, separator geometries, and flow rates, which provided insights into the complex interplay between these design parameters. In addition, the model identified the separator design allowing for the highest separation efficiency and determined the retention potential in both single and multiple separators in series. The work demonstrated that throughputs of ~1/4 L/h could be achieved for 250-500 nm iron oxide nanoparticle solutions, using less than 10 separator units in series.

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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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In this work, a continuous high-speed countercurrent chromatography method has been developed on the basis of elution-extrusion mode and this method was successfully applied to the separation of maslinic and oleanolic acid from the extract of olive pulp. In the process of 'elution', the sample solution was continuously loaded into the column and the maslinic acid was steadily eluted out in this step while highly retained oleanlic acid always stayed in the column. In the process of 'extrusion', the oleanlic acid was pushed out of the column with the stationary phase. In this way, we achieved a large sample loading. A total of 120 mL sample solution (about 89.55% of the column volume) which contains 600 mg olive pulp extract was pumped in the apparatus by a constant-flow pump and the maslinic and oleanolic acids were largely separated within 120 min. Both of these two compounds presented high yields and high purities (271.6 mg for maslinic acid with 86.7% and 83.9 mg oleanolic acids with 83.4%).

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Detection and analysis of circulating tumor cells (CTCs) have emerged as a promising way to diagnose cancer, study its cellular mechanism, and test or develop potential treatments. However, the rarity of CTCs among peripheral blood cells is a big challenge toward CTC detection. In addition, in cases where there is similar size range between certain types of CTCs (e.g. breast cancer cells) and white blood cells (WBCs), high-resolution techniques are needed. In the present work, we propose a deterministic dielectrophoresis (DEP) method that combines the concept of deterministic lateral displacement (DLD) and insulator-based dielectrophoresis (iDEP) techniques that rely on physical markers such as size and dielectric properties to differentiate different type of cells. The proposed deterministic DEP technology takes advantage of frequency-controlled AC electric field for continuous separation of CTCs from peripheral blood cells. Utilizing numerical modeling, different aspects of coupled DLD-DEP design such as the required applied voltages, velocities, and geometrical parameters of DLD arrays of microposts are investigated. Regarding the inevitable difference and uncertainty ranges for the reported crossover frequencies of cells, a comprehensive analysis is conducted on applied electric field frequency as design's determinant factor. Deterministic DEP design provides continuous sorting of CTCs from WBCs even with similar size and has the future potential for high throughput and efficiency.

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In this work, separation of multicomponent mixtures containing components with the same and different electrophoretic mobility by using orthogonal pressurised planar electrochromatography is studied theoretically. Additionally, a simple way for determination of a maximum amount of mixture causing volume overload of flat/planar columns used in this technique is presented. In the next stage, effects of change in different parameters on process performance by simulation case studies are investigated. A comparative study of separation productivity of orthogonal pressurised planar electrochromatography with the continuous and periodic modes of mixture delivery and column chromatography is carried out.

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If a multi-component monosugar mixture including fucose was used as the substrates for the Klebsiella oxytoca fermentation, it could offer the following two benefits simultaneously; (i) the removal of all monosugars other than fucose, and (ii) the acquisition of 2,3-butanediol (BD). To utilize such two benefits in favor of the economical efficiency of the fucose production process, it is essential to accomplish a high-purity separation between fucose and BD on the basis of a highly-economical mode. To address this issue, we aimed to develop a simulated moving bed (SMB) process for continuous-mode separation of fucose and BD with high purities. It was first found that an Amberchrom-CG71C resin could become a suitable adsorbent for the separation of interest. The intrinsic parameters of fucose and BD on such proven adsorbent were determined, and then applied to the optimal design of the fucose-BD separation SMB. The capability of the designed SMB in ensuring high purities and high yields was experimentally verified. Finally, we devised two potential strategies to make a further improvement in product concentrations and/or desorbent usage while keeping the purities and yields of fucose and BD almost unchanged. The first strategy was based on partial extract-collection and partial extract-discard, which was found to result in 33% higher BD product concentration. The second strategy was based on partial extract-collection, partial extract-recycle, and partial desorbent-port closing, which could lead to 25% lower desorbent usage, 33% higher BD product concentration, and 7% higher fucose product concentration.

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The ability to separate RBCs from the other components of whole blood has a number of useful clinical and research applications ranging from removing RBCs from typical clinical blood draw, bone marrow transplants to transfusions of these RBCs to patients after significant blood loss. Viewed from a mechanistic/process perspective, there are three routine methodologies to remove RBCs: 1) RBCs lysis, 2) separation of the RBCs from the nucleated cells (i.e., stem cells) based on density differences typically facilitated through centrifugation or sedimentation agents, and 3) antibody based separation in which a targeted RBC is bound with an affinity ligand that facilitates its removal. More recently, several microfluidic based techniques have also been reported. In this report, we describe the performance of continuous RBC separation achieved by the deflection of intrinsically magnetic, deoxygenated RBCs as they flow through a magnetic energy gradient created by quadrupole magnet. This quadrupole magnetic, with aperture of 9.65 mm, has a maximum field of B0 = 1.36 T at the pole tips and a constant field gradient of B0 /r0 = 286 T/m. The annular flow channel, contained within this quadrupole magnet, is 203 mm long, has an inner radius of 3.98 mm, and an inner, outer radius of 4.36 mm, which corresponds to an annulus radius of 380 micrometer. At the entrance and exit to this annular channel, a manifold was designed which allows a cell suspension and sheath fluid to be injected, and a RBC enriched exit flow (containing the magnetically deflected RBCs) and a RBC depleted exit flow to be collected. Guided by theoretical models previously published, a limited number of operating parameters; total flow rate, flow rate ratios of flows in and flow out, and ratios of RBC to polystyrene control beads was tested. The overall performance of this system is consistent with our previously presented, theoretical models and our intuition. As expected, the normalized recovery of RBCs in the RBC exit fraction ranged from approximately 95% down to 60%, as the total flow rate through the system increased from 0.1 to 0.6 ml/min. At the cell concentrations studied, this corresponds to a flow rate of 1.5 × 106 -9 × 106 cells/min. While the throughput of these pilot scale studies are slow for practical applications, the general agreement with theory, and the small cross-sectional area in which the actual separation is achieved, 77 mm2 (annulus radius times the length), and corresponding volume of approximately 2 mls, suggests the potential to scale-up a system for practical applications exists and is actively being pursued.

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The exploitation of separation materials with high selectivity for oil pollutants is of great importance due to severe environmental damage from oil spillages and industrial discharge of oils. A facile in situ growth process for creating superhydrophobic-superoleophilic fabrics for oil-water separation is developed. This proposed method is based mainly on the deposition Cu nanoparticles and subsequent hydrophobic modification. Compared with the hydrophilicity of original fabric, the water contact angle of the modified fabric rises to 154.5°, suggesting its superhydrophobicity. The as-prepared fabrics also exhibit wonderful oil-water selectivity, excellent recyclability, and high separation efficiency (>94.5%). Especially, via pumping the fabric rolled into a multilayered tube, various types of oils on water surface can be continuously separated in situ without any water uptake. Furthermore, the superhydrophobic fabrics show excellent superhydrophobic stability, and can resist different chemicals, such as salty, acidic, and alkaline solutions, oils, and hot water. After the abrasion of 400cycles, the broken fabric still possesses highly hydrophobicity with water contact angle of 145°. Therefore, due to simple fabrication steps, low cost, and scalable process, the as-prepared fabrics can be applied in the separation of oils and other organic solvents from water.

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The economical efficiency of valine production in related industries is largely affected by the performance of a valine separation process, in which valine is to be separated from leucine, alanine, and ammonium sulfate. Such separation is currently handled by a batch-mode hybrid process based on ion-exchange and crystallization schemes. To make a substantial improvement in the economical efficiency of an industrial valine production, such a batch-mode process based on two different separation schemes needs to be converted into a continuous-mode separation process based on a single separation scheme. To address this issue, a simulated moving bed (SMB) technology was applied in this study to the development of a continuous-mode valine-separation chromatographic process with uniformity in adsorbent and liquid phases. It was first found that a Chromalite-PCG600C resin could be eligible for the adsorbent of such process, particularly in an industrial scale. The intrinsic parameters of each component on the Chromalite-PCG600C adsorbent were determined and then utilized in selecting a proper set of configurations for SMB units, columns, and ports, under which the SMB operating parameters were optimized with a genetic algorithm. Finally, the optimized SMB based on the selected configurations was tested experimentally, which confirmed its effectiveness in continuous separation of valine from leucine, alanine, ammonium sulfate with high purity, high yield, high throughput, and high valine product concentration. It is thus expected that the developed SMB process in this study will be able to serve as one of the trustworthy ways of improving the economical efficiency of an industrial valine production process.

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A highly hydrophobic and highly oleophilic sponge was synthesized by simple vapor-phase deposition followed by polymerization of polypyrrole followed by modification with palmitic acid. The prepared sponge shows high absorption capacity in the field of separation and removal of different oil spills from water surface and was able to emulsify oil/water mixtures. The sponge can be compressed repeatedly without collapsing. Therefore, absorbed oils can be readily collected by simple mechanical squeezing of the sponge. The prepared hydrophobic sponge can collect oil from water in both static and turbulent conditions. The proposed method is simple and low cost for the manufacture of highly oleophilic and highly hydrophobic sponges, which can be successfully used for effective oil-spill cleanup and water filtration.

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The economically-efficient separation of galactose, levulinic acid (LA), and 5-hydroxymethylfurfural (5-HMF) in acid hydrolyzate of agarose has been a key issue in the area of biofuel production from marine biomass. To address this issue, an optimal simulated moving bed (SMB) process for continuous separation of the three agarose-hydrolyzate components with high purities, high yields, and high throughput was developed in this study. As a first step for this task, the adsorption isotherm and mass-transfer parameters of each component on the qualified adsorbent were determined through a series of multiple frontal experiments. The determined parameters were then used in optimizing the SMB process for the considered separation. Finally, the optimized SMB process was tested experimentally using a self-assembled SMB unit with four zones. The SMB experimental results and the relevant computer simulations verified that the developed process in this study was quite successful in the economically-efficient separation of galactose, LA, and 5-HMF in a continuous mode with high purities and high yields. It is thus expected that the developed SMB process in this study will be able to serve as one of the trustworthy ways of improving the economic feasibility of biofuel production from marine biomass.

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