solid phase peptide synthesis

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Aqueous solid-phase peptide synthesis (ASPPS): A novel concept of peptide synthesis

Aqueous solid-phase peptide synthesis (ASPPS): A novel concept of peptide synthesis

During the time elapsed since the first oligopeptide has been assembled by chemical means, scientists constantly improve the methods of peptide synthesis in view of optimized reaction conditions, protecting schemes, tailor-made coupling agents, and solvent systems. The introduction of Merrifield’s solid-phase approach in early 1960s became a major breakthrough that has designated the vector of peptide synthesis for the following decades. To date, production of peptides has gained almost optimal efficiency in terms of coupling yield and reaction times. However, solvent consumption, that has been recognized as an economic and environmental issue since a decade, makes solid-phase peptide synthesis one of the most inefficient synthetic approaches known. Indeed, DMF and NMP which are currently the solvents of choice in SPPS, are classified by the European REACH Regulation as substances of very high concern due to their carcinogenic, mutagenic or toxic for reproduction (CMR) properties. Therefore, sustainable, green peptide synthesis that does not require or minimizes application of highly hazardous solvents is currently in research focus.
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Solid Phase Peptide Synthesis by Four Component Condensation: Peptide Formation on an Isocyano Polymer Support 

Solid Phase Peptide Synthesis by Four Component Condensation: Peptide Formation on an Isocyano Polymer Support 

Coupling of various protected amino acids or small peptides on an isocyano polymer and in the presence of 1 -methyl-3-formylindole dem- onstrates the feasibility of peptide formation b[r]

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Secondary structure mimetic polymer and peptide conjugates : synthesis and structural investigations

Secondary structure mimetic polymer and peptide conjugates : synthesis and structural investigations

β-Turn mimetic peptides were synthesized by solid phase peptide synthesis (SPPS) as illustrated in Scheme 21. As SPPS is started from the carboxyl end (C-terminus) of the peptide, a copolymer resin linked to Fmoc-protected valine was used as solid support. After deprotection of the amine using piperidine in DMF, followed by a washing step using DMF, coupling of the neighboring Fmoc-protected amino acid is performed using N,N-diisopropylcarbodiimide (DIC) and oxyma at 90 °C, followed again by washing with DMF. These steps of deprotection, activation and coupling were repeated with single amino acids until the first peptide sequence was finished. Then, coupling with the Fmoc-protected β-turn mimetics was performed using HOBt and DIC and DMF as solvent and gentle stirring at room temperature for twelve hours. This adapted method assured complete modification, which was shown by MALDI-TOF-MS. Further elongation of the peptide was performed again via SPPS as described above. After the final step, the peptide is cleaved off from the solid resin using TFA, purified via preparative HPLC and analyzed via analytical HPLC and MALDI-TOF-MS.
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Solid Phase-Supported Synthesis of Muraymycin Analogues

Solid Phase-Supported Synthesis of Muraymycin Analogues

It is known from literature precedent that a final global de- protection step under acidic conditions furnishes muraymycins and their analogues in a reliable and robust manner. [10,11] Hence, the overall strategy for the protection of amino acid side chains and the cleavage of the peptide unit from the solid support was conceived accordingly. For the envisioned solid phase-supported synthesis, we have chosen a trityl resin in combination with an Fmoc protecting group strategy. As Fmoc is a standard protecting group used in SPPS, well-established protocols are available. [14] The trityl resin is cleavable under mild acidic conditions and can thus be orthogonally used in combination with acid-sensitive side chain protecting groups such as Boc (which requires harsher conditions for cleavage). In contrast to standard SPPS, we intended to prepare a peptide- linker aldehyde 14 that would undergo reductive amination after its cleavage from the resin. Konno et al. reported the prep- aration of similar peptides on solid support, [15] but obtained epimers for the amino acid bearing the aldehyde functionality. However, this problem was impossible for our synthetic route because of the propyl linker between the first amino acid and the aldehyde functionality.
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Alternative methods for the liquid phase synthesis of peptide nucleic acids and polyphosphazene based reagents for the halogenation of alcohols / Author Dipl.-Ing. Michael König

Alternative methods for the liquid phase synthesis of peptide nucleic acids and polyphosphazene based reagents for the halogenation of alcohols / Author Dipl.-Ing. Michael König

After this addition, the pH was set to a basic value (around 8.5 to 9 in our case) with additional DIEA. While in the original paper, the pH of the reaction solution was set to three with 20 % citric acid, it was found to be more convenient, when the DMF-reaction solution was added to a pH 3 citric acid buffer. The pH of the buffer which exceeded 3.5 in some reaction runs was then set back to pH 3 with 20 % citric acid and the turbid solution was extracted with EtOAc. The organic phase was further washed with citric acid and brine and the extract evaporated to an oily residue. By heating this oil with EtOAc the product could be obtained as solid in 66 % yield. For the cytosine monomer and its acetic acid derivative (see Figure 68 for full synthesis sequence), we started from the nucleobase itself and alkylated it with methyl bromoacetate and sodium hydride according to the procedure of Schwergold et al. [178] . The product CT01 could be obtained in 36 %. Following the alkylation, the exocyclic amine was then protected with the ClBhoc group according to procedures from Winssinger [47] which is basically indentical with the one that Coull and others had used for the Bhoc protection in their seminal patent [64] . They reacted a cytosine-1-acetic acid ester with CDI, which first formed the isocyanate. This isocyanate was then transformed to the carbamate by adding the benzhydrol derivative. In our case, the use of 4,4’-dichlorobenzhydrol lead to the formation of product CT02 in 78 % yield. For the removal of the methyl ester, it was first tried to use 1M NaOH in THF which surprisingly was not able to fully deprotect the methyl ester. Thus, also for this step Winssingers procedure was followed and LiOH was used in a rather complex solvent mixture. The final (N 4 -(4,4’- dichlorobenzhydryloxycarbonyl)cytosine-1-yl)acetic acid (CT03) could be obtained from the deprotection mixture by setting the pH to three with citric acid which caused the product to precipitate. After purification through recrystallization from EtOH and drying, the product could be obtained in 78 % yield.
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Synthesis and Solid-Phase Application of a 9-Xanthenyl Handle 

Synthesis and Solid-Phase Application of a 9-Xanthenyl Handle 

sole:D M S, 1:1:4) at room tem p e ra tu re (Schem e 2). D iffe re n t cleavage tim es (5, 15, 30, and 60 m in) w ere used to check the peptide com position of the cleavage m ixture. H P L C analysis indicated th at 15 m in cleavage proved to be the optim al one, yielding 45% sulfated peptide and 15% of unsul- fa te d C C K - 8 , though tert-butyl p ro tectio n of the side chain of the aspartates was still p resen t to the e x te n t of -4 0 % during this period. D esulfation b e ­ cam e p ro m in en t (74% ) and only 14% of sulfate e ste r could be detected using 60 min cleavage (Fig. 1). It should be noticed th a t these findings are significantly different from those re p o rte d by
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Gutenberg Open Science: Biomimetic Identification of Phase-Selective Peptide-Additives

Gutenberg Open Science: Biomimetic Identification of Phase-Selective Peptide-Additives

investigation of these presumed adsorption rates. For this purpose, the diazo-dye (15) was selected and successfully utilised in the synthesis of an Fmoc SPPS compatible dye-label (16). Along with the additionally obtained spacer amino acid (14), the dye-label (16) was employed in the solid phase synthesis of the dye- labelled peptide (20), a derivative of peptide (8). This newly accessed dye-labelled peptide (20) was intended for the demonstration of the individual adsorptions towards the respective anhydrous calcium carbonate polymorphs in a first stage. Moreover, it was planned to apply the dye-labelled peptide (20) for the determination of the respective adsorptions via photometry at a later stage. However, during the first adsorption tests the solution of peptide (20) solidified into a hydrogel in the presence of the calcium carbonate crystallites and thus prohibited further characterisation. Further experiments at lower concentrations of peptide (20) yielded similar results and simultaneously indicated a comparably low critical gelation concentration below 0.5 ‰ (w/v). Even lower concentrations of peptide (20) were not expected to allow the photometric determination of the individual adsorptions, let alone the simultaneous demonstration of the individual adsorptions by light-microscopy. Hence, it was refrained from further applications of peptide (20). Regardless, the general approach of demonstrating and subsequently determining the adsorption rates still appears viable. In this regard, it was suggested that a different, more polar dye compound could be applied to circumvent the impeding hydrogel formation. Furthermore, an additional alternative approach had been considered, which proposed the application of a fluorine- labelled derivative of peptide (8) in combination with 19 F-NMR experiments, to
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Sol Gel vs Solid State Synthesis of the Fast Lithium-Ion Conducting Solid State Electrolyte Li7La3Zr2O12 Substituted with Iron

Sol Gel vs Solid State Synthesis of the Fast Lithium-Ion Conducting Solid State Electrolyte Li7La3Zr2O12 Substituted with Iron

gel synthesis is much lower than for the solid state synthesis and was therefore further used after the measurement and needed to be kept away from humidity. In Fig. 2 SEM images of the calcined powders for the sol gel and solid state method are shown. Both samples contain similar mi- crostructural features. For both materials sintered areas and branches can be observed. The sintered areas of the solid state powder exhibit sizes up to 50 μm and branches with sizes ranging from 5–10 μm. The sol gel powder exhibits sintered areas up to 30 μm and branches around 5 μm building up a coral like structure which can be seen in a magnification of 1000 in the left corner at the top. Furthermore, in the sol gel powder grains which are in good contact to each other with distinct grain boundaries are visible in the image with a magnification of 5000. These grains range from 1 μm to 10 μm. The grain bound- aries are mainly strong curved. A curved grain boundary compared to a straight one is energetically unfavored as grain boundaries contain a large amount of defects and the shortest distance is preferred. As the main driving force of a sintering process is the reduction of the free energy this curved boundaries show a strong tendency to further grow. 30
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Sound Velocities of Lennard-Jones Systems
Near the Liquid-Solid Phase Transition

Sound Velocities of Lennard-Jones Systems Near the Liquid-Solid Phase Transition

The longitudinal and transverse sound velocities of the Lenard-Jones system have been evaluated at the liquid–solid coexistence. Two methods have been employed, one uses the additivity principle and the other uses relations between sound velocities and excess energy and pressure. The first method is simple but approximate, while the second is exact, but requires the knowledge of the excess energy and pressure. The agreement between the two methods is rather good, the deviations are only observable near the triple point. This is not surprising, because in this region the repulsive and attractive contributions are comparable in magnitude, so that even a small relative inaccuracy in each of these terms can result in a much greater relative inaccuracy of their difference. Nevertheless, even near the triple point the results based on the additivity principle demonstrate acceptable accuracy.
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From Synthesis to Battery Cell Integration of Al-doped Li7La3Zr2O12 Solid Electrolyte

From Synthesis to Battery Cell Integration of Al-doped Li7La3Zr2O12 Solid Electrolyte

Nebulized spray pyrolysis (NSP) is an aerosol based synthesis method that allows for the synthesis of nanocrystalline materials. The principle of NSP is depicted in Figure 3-1. The precursors, usually water-soluble nitrates, are dissolved to prepare a stable solution. This is then injected into a glass nebulizer by means of a syringe pump. Underneath the solution a membrane performing periodic mechanical vibration of ultrasound frequency destabilizes the liquid to a point of break-up and the formation of droplets (aerosol) takes place [37]. The droplets are carried through the hot-wall reactor by means of the carrier and/or reaction gas, where the synthesis temperature can be controlled. The pressure of the system as well as the gas flow can be controlled using a pump and a mass flow controller allowing for the variation of the residence time in the reactor. The processes occurring in the reactor are presented in Figure 3-1. The aerosol droplets first undergo solvent evaporation and precursor precipitation upon heating. The individual droplets of the aerosol can act as segregated, micron scale reaction vessels for synthesis. As the aerosol droplets proceed further into the heated reaction zone, precursor decomposition or solid state reactions occur to produce particles of the synthesized material [37]. The crystallization process takes place depending on the solubility of each material system, and nanocrystallites are synthesized forming spherical agglomerates according to the droplet size at the solubility limit. In the subsequent step sintering of the nanocrystallites takes place. The nanocrystalline powder is then collected using a glass fiber filter (Sartorius, Germany) collector, which is moderately heated (120 °C) to eliminate residual moisture using a heating tape (HBS 450, Horst, Germany).
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HF-free solid-state synthesis of the oxyfluoride phosphor K3MoOF7:Mn4+

HF-free solid-state synthesis of the oxyfluoride phosphor K3MoOF7:Mn4+

Before examining the luminescence properties, the PXRDs before and after the ball milling experiment were compared. The PXRDs revealed no obvious phase change of the material by the ball milling process (Figure S2). Phase analysis of the powder sample after ball milling via the Rietveld technique de- termined the powder composition to 79 wt.-% K 3 MoOF 7 and

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Solid-phase microextraction for bioconcentration studies according to OECD TG 305

Solid-phase microextraction for bioconcentration studies according to OECD TG 305

An important aim of the European Community Regulation on chemicals and their safe use is the identification of (very) persistent, (very) bioaccumulative, and toxic substances. In other regulatory chemical safety assessments (pharmaceuticals, biocides, pesticides), the identification of such (very) persistent, (very) bioaccumulative, and toxic substances is of increasing importance. Solid-phase microextraction is especially capable of extracting total water concentrations as well as the freely dissolved fraction of analytes in the water phase, which is available for bioconcentration in fish. However, although already well established in environmental analyses to determine and quantify analytes mainly in aqueous matrices, solid-phase microextraction is still a rather unusual method in regulatory ecotoxicological research. Here, the potential benefits and drawbacks of solid-phase microextraction are discussed as an analytical routine approach for aquatic bioconcentration studies according to OECD TG 305, with a special focus on the testing of hydrophobic organic compounds characterized by log K OW > 5.
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Two phase flow solver for solid particles in hypersonic Martian entry flows

Two phase flow solver for solid particles in hypersonic Martian entry flows

flows into dusty atmosphere or the user defined geometrical shapes like circle or square may also be used for solid particle injection into the flowfield (This may be required for powder injection into plasma generators or solid particle simulation in nozzle flows). The task that is necessary for the Euler-Lagrangian modeling is to locate the particles in the flowfield after injection. A mathematical model based on the vector products of all the sides of the volume cell is developed and implemented in the code and it works perfectly and efficiently for the location of the particles. The translational motion of the particle is governed by Newtons second law. The particle movement is characterized by transitional and rarefied flow properties due to the low gas densities and small particle sizes. The Henderson’s drag correlation is implemented in the code because it has the capability to compute the drag coefficient not only in the continuum region but also it predicts drag values very good in the transitional and rarefied regions. Due to hypersonic entry flows, a strong bow shock is developed in front of the vehicle and the gas behind the shock heated up to the very high temperatures. Because of this effect, the particles entering the flowfield also experiences very high temperatures. An adequate heat transfer model is also included for particle temperature computation in high temperature gas flows. The rarefaction correction is also included in the Nusselt number computation for convective heat transfer model. It is also worthy to mention here that Nusselt number is the key parameter for measure of convective heat transfer. Many correlation for the computation of Nusselt number are available. However all of them are developed for subsonic flows. It cannot be directly used for solid particle simulation in high temperature gas flows because of relative supersonic flow between the gas and particles after the shock. Usually this ef- fect is ignored however the results may vary significantly if this phenomenon is ignored. A stagnation point shock correlation is also introduced to take care of this effect. In the end of this chapter, particle-wall interaction is discussed and different models being implemented in the code are presented.
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Prediction of secondary-phase formation and degradation in solid oxide fuel cell anodes

Prediction of secondary-phase formation and degradation in solid oxide fuel cell anodes

Insufficient lifetime is a major factor impeding the large-scale commercialization of solid oxide fuel cell (SOFC) technology. The formation of secondary phases in the porous electrodes can occur via various chemical mechanisms and represents a major cause of degradation due to pore clogging and deactivation of active surfaces. We present a modeling and simulation study of secondary-phase formation in porous Ni/YSZ-based anodes. Specifically, we investigate the formation of solid nickel oxide (reoxidation) in the case of high fuel utilization or low cell voltages, as well as the formation of solid carbon (coking) in the case of external and internal reforming.
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Solid catalysts for methanol and ammonia synthesis investigated by in-situ neutron diffraction

Solid catalysts for methanol and ammonia synthesis investigated by in-situ neutron diffraction

6.3 Results and Discussion 57 two peaks, which due to their high Q and often low intensity are only rarely recorded in XRD patterns of fcc nanomaterials, show an anisotropic broadening. The 331 at lower scattering angle is significantly broader compared to the neighbouring 420 peak. The Rietveld routine applied in this study tries to fit these peaks with similar widths and with interdependent positions given by the Cu lattice parameter. As a result, the intensity of the sharper 420 peak is not properly described in the calculated pattern. Additionally, only the outer tails of the doublet are adequately fitted while the inner tails are underestimated. As a consequence, the calculated spacing of the peaks is larger than the experimentally observed one. Naturally, the model-free pattern decomposition approach with its higher numbers of parameters and less restrictions allows fitting this feature more satisfactorily. It is noted, however, that this method generally deals much worse with peak overlapping and some instrumental artefacts like the zero-shift of the Q-scale that can be treated much better with the Rietveld method. For the analysis of the catalysts in this study, a combination of Rietveld and pattern decomposition was used to mitigate these problems. The zero-shift and the peak profiles of the ZnO phase (if present in crystalline form) were determined by Rietveld refinement and used for correction of the raw peak profiles of the Cu phase extracted from pattern decomposition. Furthermore, the instrumental contribution to the peak profiles was deconvoluted from the experimental data as described in the Appendix A.
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Molecular dynamics study of interactions between nano crystals and solid-liquid phase equilibria

Molecular dynamics study of interactions between nano crystals and solid-liquid phase equilibria

energy of the solid phase: the extended Einstein crystal method supplied me with the free energy pathway while I utilized the Einstein molecule method to restrain center of mass movement of the crystal. In combination with the force constant as obtained from unrestrained equilibrium simulations, this constitutes a numerically stable and efficient way to compute the absolute free energy of a solid. In addition, I studied different free energy methods, i.e. a method based on overlap sampling, thermodynamic integration as well as a nonequilibrium method, and assessed their ease of applicability, convergence and statistical uncertainty. Phase coexistence lines were constructed in two steps. First, I reused simulation data conducted to determine an initial melting point to make an estimate of additional melting conditions via reweighting. And second, I conducted ad- ditional, short simulations in the isothermal-isobaric ensemble at the estimated phase equilibrium states in order to refine the initial estimate through simulations conducted in parallel. For the systems studied in this work, I found that a single set of additional simulations was sufficient to trace the coexistence line for a wide range of pressures (or temperatures). This work included absolute free energies and phase coexistence lines for three systems of different complexity – argon, methanol and water. For all systems my results showed good agreement with literature data. The method worked very well for rigid molecules and while I did not test it for fully flexible molecules it should be directly applicable as the original extended Einstein crystal study was presented for fully flexible molecules.
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Detection of Constant Phase Shifts in Filters for Sound Field Synthesis

Detection of Constant Phase Shifts in Filters for Sound Field Synthesis

Filters with constant phase shift in conjunction with 3/6 dB amplitude decay per octave frequently occur in sound field synthesis and sound reinforcement applications. These ideal filters, known as (half) differentiators, exhibit zero group delay and 45/90 degree phase shift. It is well known that certain group delay distortions in electro-acoustic systems are audible for trained listeners and critical audio stimuli, such as transient, impulse-like and square wave signals. It is of interest if linear distortion by a constant phase shift is audible as well. For that, we conducted a series of ABX listening tests, diotically presenting non-phase shifted references against their treatments with different phase shifts. The experiments revealed that for the critical square waves, this can be clearly detected, which generally depends on the amount of constant phase. Here, -90 degree (Hilbert transform) is comparably easier to detect than other phase shifts. For castanets, lowpass filtered pink-noise and percussion the detection rate tends to guessing for most listeners, although trained listeners were able to discriminate treatments in the first two cases based on changed pitch, attack and roughness cues. Our results motivate to apply constant phase shift filters to ensure that also the most critical signals are technically reproduced as best as possible. In the paper, we furthermore give analytical expressions for discrete-time infinite impulse response of an arbitrary constant phase shifter and for practical filter design.
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Significance of Posttransplant HLA Antibody Monitoring by Solid-Phase Immunoassays in Renal Allograft Recipients

Significance of Posttransplant HLA Antibody Monitoring by Solid-Phase Immunoassays in Renal Allograft Recipients

Without a doubt, solid organ TX is a history of resounding success (119). However, the perpetual enigma of late renal allograft dysfunction is still unsolved. In this thesis we addressed this issue from a diagnostical point of view and could clearly demonstrate that (i) HLAab as detected by Luminex ® SAB posttransplant are in fact valuable biomarkers to predict long-term outcome (Figure 29). Consequently, we established an effective posttransplant HLAab monitoring scheme currently in place at our institution which will be presented in one of the following sections. (ii) We successfully modified the standard Luminex ® assay to facilitate the discrimination between complement-fixing and non- complement-fixing HLAab. Thereby, we could further improve the prediction of the antibodies’ pathogenicity much better than using the standard approach of MFI (Figure 31). (iii) Epitope analyses and longitudinal follow-up of patients helped gaining more insight into the natural history of the humoral alloimmune response after TX. HLAab can appear de novo at any time point posttransplant (Figure 34) and are directed against immunogenic HLA epitopes resulting in a broad reactivity against multiple antigens with shared epitopes.
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Mechanochemical synthesis, phase transformation and surface functionalization of metal oxides

Mechanochemical synthesis, phase transformation and surface functionalization of metal oxides

Thermogravimetric analysis (Supplementary Materials, Figure S4) between 25 and 1000 °C shows a mass loss up to the temperature range of 575 to 600 °C (attributed to Li 2 CO 3 ). An exothermic signal arises in the calculated differential thermal analysis (DTA) curve, which correlates to the complete crystallization of β-Li 2 TiO 3 as observed in the PXRD measurements. At 600 °C a composition with equal amounts of α- and β-Li 2 TiO 3 was refined. Fitting with only the monoclinic phase led to an insufficient refinement (GOF = 4.39) while inclusion of the cubic phase results in a greatly improved description (GOF = 1.42). At 700 °C an abrupt increase to >90% β-Li 2 TiO 3 occurs, with only minor amounts of the cubic phase remaining. Fitting with only the monoclinic structure results in a similar description (GOF = 2.09) as by using both structures (GOF = 1.93). Though the improvement of the description is small, this may still be attributed to remaining portions of the cubic phase. This leads to an overall interpretation that an ordering phenomenon takes place in the cubic phase at lower temperatures with a discrete structure change between 600 and 700 °C.
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A Fluid Dynamics Approach for Solid-Liquid Phase Change in Binary and Elemental Melts

A Fluid Dynamics Approach for Solid-Liquid Phase Change in Binary and Elemental Melts

In this thesis, a simulation model is developed which couples the Navier-Stokes equa- tions as well as transport equations for the temperature and composition field with an independent phase change model based on the phase-field method in a thermodynamic consistent manner. In particular, a phenomenological free energy functional is constructed and the governing equations for the phase volume fractions and their coupling with the other transport equations are obtained by using the formalism of irreversible thermody- namics. A dynamic calculation procedure based on free energy minimization is proposed for a parameter appearing in the free energy functional, which characterizes the width of the smooth transition regions of the phase volume fractions between pure phases. The final set of coupled evolution equations can describe convective binary eutectic alloy solid- liquid phase change with sharp as well as diffuse interfaces and contains the representation of elemental materials as a special case. In particular, the model is able to describe the detailed solidification microstructure if it is applied on sufficiently small length scales. The model equations have been implemented in the computational fluid dynamics toolbox OpenFOAM and the code is verified by comparisons with analytical predictions as well as with neighbouring modelling approaches in suitable scenarios.
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