DE10105289A1 - Porous polymeric bodies, for use in separation of e.g. low-density lipoproteins from whole blood, are obtained by radical polymerization of porogen-containing dispersion including hydrophobic monomers - Google Patents

Abstract

Production of a porous polymeric body involves washing and drying a product obtained by polymerization of a dispersion comprising: (i) a hydrophobic linear monomer; (ii) a hydrophobic crosslinking monomer; (iii) a radical initiator; (iv) an organic solvent; (v) a porogen; (vi) an emulsifier; and (vii) water. Production of a porous polymeric body involves washing and drying a product obtained by polymerization of a dispersion comprising: (i) a hydrophobic linear monomer; (ii) a hydrophobic crosslinking monomer; (iii) a radical initiator; (iv) an organic solvent; (v) a porogen; (vi) an emulsifier; and (vii) 10-60 wt.% water.

Description

The present invention relates to a porous polymeric molded body. More accurate The invention relates to a porous polymeric shaped body which is used for the separation of Mixtures of substances is suitable, in particular for the adsorptive separation of Substances from physiological liquids and solutions. Furthermore, the Invention a method for producing a housing for the porous polymer ren molded body, and its use in separation processes.

In conventional column separation processes, the mixture of substances to be separated by means of pressure or gravity over or through a bed of a particulate Separation material, the so-called stationary phase. With a suitable choice of the separating material interact in the mixture of substances to be separated contained components with the release material and are due for example a different molecular size, charge or hydrophobicity and a resulting different degree of interaction with the Separation material delayed to different extents and separated into zones, which leave the column postponed and can be collected separately. It can also separate one or more components from the The mixture of substances completely adhere to the separating material and thereby from the Separated mixture of substances and optionally isolated.

One is usually used to pack or manufacture a conventional column tubular container filled with the particulate separation material, and the Particles are e.g. B. by shaking or tapping as evenly as possible compacted. However, even with careful packing of the column, it cannot excluded that the column is not packed completely evenly. By an uneven column packing or operating errors during the separation process, such as a so-called "dry running" of the column, it can  to crack and / or channel formation in the column bed, from which one Zone broadening and thus a poorer separation effect or even that complete loss of the separating effect can result.

So that the particulate separation material can be packed well, the Particles have a size that is as uniform as possible. The particulate Separating material can, for example, by suspension polymerization or Emul ion polymerization can be produced. However, since the particles obtained in the Rule polydisperse is a complex and repeated fractionation of the Particles by sieving the desired particle size (s) necessary to Obtain particles of a uniform size.

The preparation of the particulate column material and the subsequent one Packing the column is therefore time consuming and costly.

To increase the effectiveness of the column, it should be packed so that the Intermediate grain volume is as small as possible, and the accessible surface of the Release material should be as large as possible. Therefore, it is advantageous to use the middle one To reduce the particle size of the separating material. With the downsizing of the Intermediate grain volume also reduces the channels for the convective flow, which results in an increased back pressure. to Compensation in such cases, shorter columns are used, from which however, a lower separation capacity results. Alternatively, at Separation higher pressures are used, in which case the column is like this must be constructed so that it can withstand these higher pressures. A Increasing the separation effect of the column by reducing the middle part Chen size is also associated with disadvantages.

Particular problems arise when separating mixtures, which are relative contain large particles such as protein aggregates, micelles or nucleic acids. Such large particles can be separated on a conventional one during separation Column through the packed in the spaces of a particulate Separating material occurring shear forces are destroyed or reduced. If only smaller particles are to be removed from such mixtures  Another danger is that the large particles can be pore grain volume are prevented from flowing freely and therefore unintentionally can also be separated. Such problems occur with the Separation of lipoproteins with low density ("low density lipoproteins", "LDL"), such as LDL cholesterol, from whole blood.

To summarize are the use of particulate release materials some disadvantages have been linked and therefore has been in the prior art tried to connect one instead of a particulate separating material de, porous matrix to use as a separating material.

For example, WO 90/07965 relates to a chromatography column in the form of a water Insoluble plug made of a cross-linked polymer, which is sufficient has the degree of crosslinking and sufficient hydrophilicity to close channels form that are permeable to macromolecular eluents. The stopper is for suitable for chromatographic separation by gravity, but not for Separation by means of overpressure, since it has been shown that the stopper at Pressurization collapses. The channels are large enough to hold a hydro to allow dynamic flow. The stopper is made by the polymerization of water-soluble monomers prepared in aqueous solution, from which the ent standing water-insoluble polymer fails.

WO 93/07945 describes a chromatography column which contains a separation medium in Contains form of a continuous macroporous polymer plug. The stopper is by polymerization of a polyvinyl monomer, preferably one Mixture of a polyvinyl monomer and a monovinyl monomer in Presence of an initiator, a porogen and, if appropriate, macroporous produced polymer particles. Due to the presence of low molecular weight and / or polymeric porogens form large pores with a diameter <600 nm and small pores with a diameter of <200 nm, the large pores make up at least 10% of the total pore volume. This Separation material has the disadvantage that small particles can be adsorbed is.  

WO 97/19347 describes a continuous macroporous matrix made of a crosslinked organic polymer, which is produced by polymerizing a "High Internal Phase Emulsion" (HIPE) system, ie a water-in-oil emulsion of organic monomers, the water phase of the emulsion is at least 75% by weight, preferably at least 90% by weight, and the polymerization results in an open porous structure which allows convective flow through the macroporous matrix. However, the method for producing a macroporous matrix using such a HIPE system has the disadvantage, among other things, that considerable amounts of a surfactant have to be added to the emulsion (according to WO 97/19347 up to 30% by weight, based on the oil phase) to stabilize the emulsion. In addition to the economic disadvantage, such surface-active agents, in particular ionic surface-active agents, cannot be completely removed from the resulting polymeric matrix, as a result of which the separation properties of the matrix can be adversely affected. In particular, functionalization of the surface is only possible to a limited extent if there are surfactants in the interface. If the surface is functionalized by oxirane, reactions of the oxirane with the surfactant residues R-SO ₃ H, for example, interfere. The surface of the matrix also changes because micelle lysis can occur.

An object of the present invention is therefore to provide a porous polymer To provide moldings which have neither the disadvantages of a particulate Separating material, still that of the previously known matrix-shaped separating materials having. In particular, a porous molded body should be provided, in on the one hand, the smallest possible particle with a correspondingly large total surface for the adsorptive separation are present and on the other hand large interconnected pores or cavities between the particles so that the free flow of larger components of the material to be separated mixture is hampered as little as possible, the particles to ensure sufficient mechanical stability covalently linked are.  

Another object of the invention is to provide a method for producing a such porous polymeric molded body and a method for training a form-fitting housing to indicate the molded body.

These tasks are characterized by the Aus in the claims leadership forms solved.

According to the invention, there is provided a method for producing a porous polymeric molded body, which comprises the steps:

• a) preparing a mixture which is at least one hydrophobic, linear polymerizing monomer, at least one hydrophobic, crosslinking polymerizing monomer, at least one radical initiator, at least one organic solvent, at least one porogen and at least one emulsifier and further 10 to 60 wt .-% water, bezo gene on the total mixture, contains
• b) preparing a dispersion from that prepared in step (a) Mixture,
• c) polymerizing the linear and crosslinking monomers (at the time the reaction is initiated a clear solution) and
• d) washing and drying the porous polymeric molding obtained pers.

The invention is based on the finding that by polymerization of the above defined dispersion, a porous polymeric molded body is formed, the one extraordinary structure, which is small, covalent with each other Binding linked, approximately globular particles, which in the porous moldings are arranged so that they form pores which are larger are the same as the intermediate grain volume of a particle bed of particles Size. Due to the covalent linkage of the approximately globular particles these large pores are fixed and therefore mechanically stable.  

According to the invention comprises the step (a) of the method according to the invention produced mixture at least one hydrophobic, linear polymerizing Monomer, at least one hydrophobic, crosslinking polymerizing monomer, at least one radical initiator, at least one organic solution medium, at least one porogen, at least one emulsifier and 10 to 60% by weight % Water, based on the total amount of the mixture. The components this mixture form a two-phase system, the hydrophobic or lipophilic phase in the following as the oil phase and the hydrophilic or lipophobic Phase is referred to below as the water phase.

The oil phase comprises the hydrophobic components, i.e. H. the at least one hydrophobic, linearly polymerizing monomer which contains at least one hydrophobic, crosslinking polymerizing monomer, the at least one radical Initiator, the at least one organic solvent and at least one Porogen.

According to the invention, the term “hydrophobic monomer” means that one of the like monomer is essentially immiscible with water and in the essentially does not dissolve in the aqueous phase.

The at least one hydrophobic, linearly polymerizing monomer can be one Compound with a radical polymerizable double bond and is preferably an acrylic, methacrylic or vinylic monomer. For example, as hydrophobic, linearly polymerizable monomers Acrylic acid and its esters, methacrylic acid and its esters, such as Methyl methacrylate (MMA), and glycidyl methacrylate (GMA) can be used. In particular, the linearly polymerizing monomers in addition to radical polymerizing double bond further functional groups contain, so that after the conversion to the porous polymeric molded body Functionalization of the inner surface of the porous molded body is possible. For example, the monomers can be a strained heterocyclic system included as a functional group. This is preferably tense heterocyclic system a three ring, selected from the group of epoxides, Aziridines and episulfides. The aforementioned groups can do this  unsubstituted or with alkyl, aryl, aralkyl or corresponding silicon organic compounds, such as trialkylsilyl or triarylsilyl radicals, be substituted. A particularly preferred linear polymerizing monomer is Glycidyl methacrylate, as well as easily hydrolyzable esters of acrylic acid or allylic compounds. The monomers described can be used alone or as Mixture can be used.

In addition to the at least one linearly polymerizing monomer, at least a hydrophobic, "crosslinking polymerizing monomer", which in The following is also called crosslinker. Such a crosslinker comprises at least two radically polymerizable groups per molecule and Examples include divinylbenzenes, divinylnaphthalenes, alkanediol acrylates or methacrylates, such as ethylene glycol dimethacrylate, butanediol dimethacrylate, hexane diol dimethacrylate and trimethylolpropane trimethacrylate, and at least bifunk tionalized polyethylene glycols or hydrophilic, functionalized polyvinyl alcohols to be named. Divinylbenzene and ethylene glycol are particularly preferred dimethacrylate.

Linear polymerizing monomers and crosslinking polymerizing monomers are preferably in a weight ratio of <50%, especially preferably <20% crosslinking polymerizing monomers, based on the Sum of linear polymerizing monomers and cross-linking polymerizing monomers used. That or the networking polymerizing monomer (s) should be present in at least 10% by weight his.

As free-radical or free-radical initiators, customary ones can be in the oil phase soluble compounds forming free radicals can be used. Are preferred Azo compounds and peroxides. Peroxodicarbonate is particularly preferred. Such initiators can be used alone or as a mixture. virtue they are used in a proportion of <0.5 to about 2% by weight, based on the polymerizable monomers used. The decomposition temperature of the Initiator is preferably below the boiling point in the mixture existing monomers and solvents.  

The oil phase further comprises at least one organic solvent in which the polymer is only soluble to a limited extent and the amount thereof is preferably from 20 to 40 wt .-%, based on the oil phase. The solubility of a polymer in a certain solvent is not only dependent on the type of Solvent, but also on the molecular weight of the polymer. So can low molecular weight polymers in a particular solvent still be soluble, whereas higher molecular weight polymers are not are more soluble in the solvent and it occurs after exceeding one a certain molecular weight a precipitation of the polymer from the solution. The Size of the globular particles in the porous shaped body can be chosen of the solvent can be controlled. The worse the resulting Polymer dissolves in the solvent, the smaller the shaped body build-up particles, since a precipitation of the polymer already at lower Molecular weights are carried out.

As organic solvents for the monomer (s), customary Solvents are used and preferably cyclohexanone or other cyclic, linear or branched, short chain hydrocarbons, i.e. H. Hydrocarbons with up to 8 carbon atoms.

Furthermore, according to the invention, the reaction mixture is a non-solvent for the resulting polymer, a so-called porogen added, which a Precipitation of the polymer with smaller molecular weights and thus a low one Particle size of the polymer particles and the formation of the pores in the poly mer particles. The non-solvent for the resulting polymer is also not water soluble and is preferably a longer chain alcohol. Alcohols with a chain length of 8 to 18 are particularly preferred Carbon atoms such as octanol, decanol, dodecanol and Octadecanol.

The water phase comprises water and at least one emulsifier or stabilizer sator, such as a protective coloid and / or a surface or surface active Substance.  

According to the invention, the mixture comprises water in an amount of 10 to 60% by weight, preferably 15 to 50% by weight, more preferably 20 to 40% by weight and most preferably 25 to 35% by weight based on the total mixture.

The emulsifier or stabilizer used according to the present invention is preferably at least one water-soluble polymer. Examples are phenyl dextran, allyl dextran, agarose and its derivatives, water-soluble cellulose derivatives and water-soluble synthetic polymers, such as polyvinyl alcohol, poly hydroxyethyl vinyl ether, polyethylene glycol, polyhydroxyethyl acrylate, polyacrylic acids, polyvinylpyrrolidone, polyacrylamides and their derivatives. Especially water-soluble or water-emulsifiable polyvinyl alcohols are preferred. The Emulsifier is preferably used in an amount of 1 to 10% by weight, based on the total mixture used.

In step (b) of the method according to the invention is preferably carried out by mechanical agitation, such as intensive stirring, a sufficiently stable disper sion or emulsion of the ingredients. "Sufficiently stable" means in Connection with the present invention that until the formation of the Matrix structure of the porous polymeric molded body from covalently to one another connected, approximately globular polymer particles of the desired size no phase separation due to the polymerization, d. H. no "breaking" of the disper sion takes place.

According to step (c) of the method according to the invention, those in the stable Dispersion contained monomers by increasing the temperature, preferably polymerized beyond the decomposition point of the radical initiator. The polymerization is preferably carried out in a protective gas atmosphere, for example wise under nitrogen or argon. It can be the usual conditions conditions of a solution, dispersion and / or emulsion polymerization be applied.

After the polymerization is complete, the solid porous polymer molded body are removed by from the polymerization vessel. For example  of the at least one porogen and unreacted mono from residues The porous shaped body can according to step (d) of the invention Process be washed with suitable solvents. suitable Solvents are, for example, methanol, ethanol, isopropanol, benzene, toluene, Acetone, diethyl ether, tetrahydrofuran and dioxane. The washing will preferably by pumping the solvent through the molded body carried out. After washing, the porous molded body is dried.

The present invention further relates to one by the invention Process available, porous polymeric molded body.

The porous polymeric molded body according to the invention has a structure in the small globular or approximately globular particles covalently with each other are connected. The small globular particles preferably have one Diameter from 1 to 50 microns, particularly preferably from 1 to 10 microns. This small globular particles are arranged in the porous shaped body such that they form large pores, which are larger than the intermediate grain volume Particle bed of particles of the same particle size are. Preferably the pores have a diameter of 0.5 to 10 µm. Because the little ones Particles firmly connected to each other by covalent bonds the large pores are mechanically stable and fixed and do not change by mechanical and / or thermal treatment of the porous Molding.

The porous polymeric molded body according to the invention preferably has a Pore volume of 1 to 2 ml / g.

The pictures show:

Fig. 1 shows a scanning electron micrograph of the surface of a porous polymeric body according to the invention obtained in Example 1. The surface has a regular structure of small, interconnected, globular particles with a diameter of about 1 to 10 µm, between which there are larger cavities.

Fig. 2 shows a scanning electron micrograph of a fractured surface of a porous polymeric molded body according to the invention obtained in Example 1, by which it is confirmed that the structure of the surface of the porous molded body already shown in Fig. 1 has formed over the entire cross section of the molded body ,

Surprisingly, it was found that the adsorption capacity of the Shaped body according to the invention essentially independent of the applied is flow rate and so high flow rates can be realized. The reason this is probably due to the special structure of the porous according to the invention polymeric molded body. The relatively large voids between the approximately globular, covalently linked particles even allow a high flow rate Mixtures of substances that are loaded with coarser constituents, the hollow space me because of the covalent linkage of the polymer particles forming them remain stable when higher pressures occur. On the other hand, the Polymer matrix has a very large surface because of its composition approximately globular particles with a small diameter, the most of the particle surfaces can be reached by convective flow. The Diffusive flow occurring in the small pores of the particles therefore occurs their importance, so that the kinetics of the adsorption is not diffusion-controlled mass transport is determined.

Therefore, the porous polymer molded body is excellent for use, for. B. in suitable for adsorption therapy, for example to remove specific ones Plasma components such as LDL cholesterols, from whole blood.

Furthermore, the porous molded body has the property that it is steam sterilized is cash.

The surface of the polymer is used to improve the adsorption properties Matrix of the porous polymeric shaped body according to the invention according to a preferred embodiment modified. When using one or more Monomers with functional groups, such as glycidyl methacrylate,  can after the polymerization and training of the Invention According to the porous molded body, these functional groups are polymer-analogous be implemented. When using glycidyl methacrylate as a monomer for example the resulting porous molded body with at least one Polyelectrolytes are implemented, the at least one polyelectrolyte while opening the epoxy ring by direct, covalent bond to the Polymer surface of the porous molded body is bound.

The outer shape of the porous molded body according to the invention is not be limits. For example, the shape of the shaped body can be selected appropriately of the reaction vessel can be determined, since the polymerizing mixture naturally completely adapts to the contours of the vessel. So there is the possibility ability to produce the porous polymeric molded body already in the vessel, the shape of which is adapted for later intended use, for example as a chromatography column.

However, the molded body can also be mechanically changed into any desired one afterwards Formed and inserted into an appropriate housing. there the outer dimensions of the molded body should generally be somewhat larger than those of the housing so that the molded body against the housing wall is pressed to avoid voids and / or unwanted channels.

According to a preferred embodiment of the present invention, the the porous molded body enclosing a housing on the molded body applied or from a film of a polymeric material formed thereon educated. Such a film is more flexible than rigid columns, so that the shaped body, for example when changing solvents or sterilizing with Superheated steam, can expand freely within limits. Furthermore, the molded body of the film fits snugly and adheres better than, for example, one Glass surface, because the film is the surface structure of the molded body adapts or slightly in the areas of the molded body near the surface penetrates. This effect can be intensified when using shrink film become.  

The porous can be used to produce the housing around the porous molded body Shaped body with a film made of a suitable polymer and this for Formation of a closed housing to be welded. But it can also a melt of the polymer with a suitable viscosity onto the surface of the molded body are applied and solidified by cooling.

However, the housing is preferably on the molded body by Polymerisa tion formed in situ. Are for the production of a film by polymerization Monomers particularly suitable, which by polyaddition or poly polymerize condensation reactions, but it can also radical poly merizable monomers are used. For the production of such Films or housings can, for example, be polyurethanes, epoxy resins and Polyester reactive resin mixtures are used, which as a film or as Coating have sufficient strength to withstand the separation to provide sufficient resistance to the occurring pressures. For the Separation of physiological liquids, pharmaceutical agents or In addition to the porous molded article, foods are also such coatings heißdampfsterilisierbar. With appropriate selection of the monomer or poly Transparent housings can also be formed, provided that this is the case with certain Use cases is an advantage. Adhesion of the polymeric film is also included the surface of the adsorber material is desired.

It is also possible to use dispersions of prepolymers which crosslink to form films be, or highly viscous solutions or dispersions of a plastic with high molecular weight, which after drying the solution for the Medium flowing through column packing is insoluble to form a film the porous molded body can be used.

The thickness of such a polymeric film can be varied as desired but for economic reasons at most 5 mm. Furthermore, the polymer Film have sufficient strength for subsequent use, so that its thickness is preferably at least 0.2 mm.  

The porous molded body according to the invention can function depending on the selection len groups of the monomer or subsequent functionalization of the porous Shaped body can be used in every separation process. For example Applications in (bio) affinity chromatography, ion exchange Chromatography, hydrophobic chromatography or as a molecular sieve possible.

As already mentioned, the molded body according to the invention is particularly suitable for the selective elimination of components such as pathogenic Biopolymers from physiological liquids, especially from whole blood, Plasma and serum, e.g. B. in the so-called adsorption therapy. For example, after suitable functionalization, the inner surface of the porous molded body selectively lipoproteins with a specific density <1.063 kg / l, can be separated from blood, plasma or serum. Enrichment of such low-density lipoproteins, such as LDL cholesterols, in the Blood is suspected of causing atherosclerosis and thus causes an increased Heart attack risk.

The invention is illustrated by the following examples.

example 1

6 parts by weight of glycidyl methacrylate, 3 parts by weight of ethylene glycol dimethacrylate, 9 parts by weight of cyclohexanone, 9 parts by weight of dodecanol, 0.1 part by weight of peroxodicarbonate, 0.4 part by weight of polyvinyl alcohol (from Fluka; Prod. No. 81386, 100,000 g / mol, viscosity 35-45 mPas (4%, 20 ° C), degree of hydrolysis 86-89%) and 12 parts by weight of water and homogenized by intensive stirring. As soon as the mixture has been homogenized, the stirring is stopped, the stirrer is removed and the stable dispersion is heated to 50 ° C. under a nitrogen atmosphere, causing the initiator to decompose and the polymerization to be initiated. After the polymerization is complete, the shaped body is removed from the mold and washed with a solvent such as acetone and ethanol (methanol) or isopropanol and dried.

The result was a porous polymer molded body, the structure of which is shown in Fig. 1 and Fig. 2.

Example 2

Example 1 is with styrene as a linear polymerizing monomer and divinylbenzene repeated as crosslinker.

Example 3 (Forming a Housing Around the Shaped Body)

The porous molded body obtained in Example 1 was placed in a container with a larger diameter than the porous molded body and Elastocoat (the Elastogran, subsidiary of BASF AG), from component A (C 6947 V 0536) and component B (C 6947) were mixed in the space filled between the container wall and the porous molded body. The emerging Polyurethane cures after a few minutes and the one with polyurethane (PU) coated porous moldings can be removed from the container.

The PU coating adheres well to the porous molded body and is mechanical stable, transparent and steam sterilisable.

The good adhesion of the PU coating can be partly due to covalent bonds hydrolyzed areas of the molded body with the isocyanate groups of Re action mixture and by a superficial penetration of the polymerization mixture can be explained in the porous molded body. Respond in particular Oxirane groups with bases and acids to diols, with isocyanates to uretha can react further.

Claims (20)

1. A process for producing a porous polymeric shaped body, which comprises the steps:

• a) the preparation of a mixture which comprises at least one hydrophobic, linearly polymerizing monomer, at least one hydrophobic, crosslinking polymerizing monomer, at least one radical initiator, at least one organic solvent, at least one porogen, at least one emulsifier and further 10 to 60% by weight. % Water, based on the total mixture,
• b) preparing a dispersion from the mixture prepared in step (a),
• c) polymerizing the linear and crosslinking monomers contained in the dispersion and
• d) washing and drying the porous polymeric shaped body obtained.

2. The method of claim 1, wherein the mixture 15 to 50 wt .-% water contains. 3. The method of claim 1, wherein the mixture 20 to 40 wt .-% water contains. 4. The method of claim 1, wherein the mixture 25 to 35 wt .-% water contains.   5. The method according to one or more of the preceding claims, wherein the at least one organic solvent is a cyclic, linear or branched, short chain hydrocarbon. 6. The method according to one or more of the preceding claims, the organic solvent being cyclohexanone. 7. The method according to one or more of the preceding claims, wherein the porogen has at least one longer-chain alcohol and / or one longer chain carboxylic acid. 8. The method according to one or more of the preceding claims, the porogen is dodecanol. 9. The method according to one or more of the preceding claims, wherein the at least one linearly polymerizing monomer is an acrylic cal, methacrylic or vinylic monomer. 10. The method according to one or more of the preceding claims, wherein the linearly polymerizing monomer is glycidyl methacrylate. 11. The method according to one or more of the preceding claims, wherein the at least one crosslinking polymerizing monomer Ethy Lenglycoldimethycrylat or divinylbenzene. 12. The method according to one or more of the preceding claims, wherein the at least one linearly polymerizing monomer and / or the at least one crosslinking polymerizing monomer at least one has another functional group. 13. The method according to one or more of the preceding claims, wherein the at least one emulsifier is a water-soluble polymer.   14. The method according to one or more of the preceding claims, the emulsifier being polyvinyl alcohol. 15. The method according to one or more of the preceding claims, wherein the at least one radical initiator is a peroxide or Azo compound is. 16. The method according to one or more of the preceding claims, the free radical initiator being peroxodicarbonate. 17. Porous molded body, obtainable by a process according to one of the An sayings 1 to 16. 18. Shaped body according to claim 17, wherein the surface of the polymer matrix of the molded body is modified. 19. Shaped body according to one of claims 17 or 18, wherein the shaped body is enclosed by a housing made of a polymer film. 20. Use of the porous polymeric shaped body according to one of the An Proverbs 17 to 19 in separation processes.