Obtaining fatty acid esters from native oils and fats
Process and device for obtaining fatty acids or fatty acid esters from native oils and fats by their enzymatic hydrolysis and optional simultaneous esterification with alcohols, especially n- and iso-alcohols Process and device for obtaining fatty acids or fatty acid esters from native oils and fats, in which lipases, as biocatalysts for hydrolysis of oils or fats, are caused to act on a mixture of an oil or fat, water, and optionally an oil- or fat-soluble alcohol to hydrolyze the oil or fat and optionally to form an ester, where the reaction mixture thus formed is placed in an self-discharging centrifuge for separation into a glycerol-containing aqueous phase and an organic phase, the centrifuge is adjusted so that a lipase-enriched intermediate phase collects in the centrifuge between the aqueous that is drained off and the organic phase that is drained off, and the centrifuge is emptied at specified times and the discharged drum contents from the centrifuge are returned to the combined hydrolysis or optionally combined hydrolysis/esterification process.
A process and device for obtaining fatty acids or fatty acid esters from native oils and fats by their enzymatic hydrolysis and optional simultaneous esterification with alcohols, especially n- and iso-alcohols
The invention concerns a process and a device for obtaining fatty acids or fatty acid esters from native oils and fats by their enzymatic hydrolysis and optional simultaneous esterification using alcohols, especially n- and iso-alcohols.
Enzymatic hydrolysis of oils and fats has indeed been known for a long time, but has not been able to compete with hydrolysis under pressure because of the high enzyme consumption.
The fatty acid esters from native fatty acids contained in fats and oils, and medium-chain (chain length less than 6 carbons) to long-chain (generally up to a 24-carbon chain length) n- and iso-alcohols are of high economic importance in numerous applications, especially in the lubricant field.
It is quite difficult to prepare esters of these alcohols with unsaturated fatty acids, especially oleic acid esters, by classical chemical routes such as acidic esterification. Enzymatic preparation of these esters from fatty acids and alcohols has not been carried out in the past because of the high enzyme requirement.
The objective of this invention is to provide an economically feasible enzymatic process for producing fatty acids and fatty acid esters from native oils and fats, and a corresponding device.
This objective is realized in the subject matter of the independent claims. Preferred further embodiments are defined in the subclaims.
The inventors have developed an economically efficient enzymatic process for hydrolyzing fats or oils, and a corresponding device to accomplish the process. They have also found that this process and device together constitute an outstanding solution to the previous problems of enzymatic hydrolysis and enzymatic production of the esters of interest in one operation.
The process of the present invention for the enzymatic preparation of fatty acid esters is characterized by the fact that lipases, as biocatalysts for the hydrolysis of oils or fats and for esterification, are caused to act on a mixture of an oil or fat, water, and a fat- or oil-soluble alcohol, especially n- and/or iso-alcohols. The resulting reaction mixture is transferred to a self-discharging centrifuge in order to separate the glycerol-containing aqueous phase formed in the combined hydrolysis-esterification process from the organic phase which contains the fatty acid esters. The centrifuge is adjusted so that an intermediate layer enriched in lipase (enzyme) that forms between the aqueous and organic phase collects in the centrifuge drum. The centrifuge drum is emptied at specified times and the discharged contents are returned to the combined hydrolysis and esterification process. The contents of the drum are also available to be used in another, separate hydrolysis and esterification process, in which is incorporated into the present process or just made available for a later process.
Without addition of the alcohol, enzymatic hydrolysis occurs without simultaneous esterification, and the hydrolysis makes possible an extremely economical production of free fatty acids and glycerol. The basic procedure is otherwise identical.
The above-mentioned intermediate layer is enriched in the plate pack of self-desludging separators, which are suitable as self-discharging centrifuges. However, in principal ribbed inserts or other internal structures such as blades and the like may be used in an equivalent manner in place of the plate pack. It is important that the centrifuge be self-discharging, making it possible for the intermediate layer that collects in the centrifuge to be discharged from time to time by emptying the centrifuge drum.
The above-mentioned intermediate layer forms during the enzymatic fat hydrolysis even when alcohol is not added, along with the separation of the two phases that form: the glycerol-containing aqueous phase and the organic phase that contains the free fatty acids formed in the hydrolysis. The problem of this emulsion-like intermediate layer is known from “Continuous Uses of Lipases in Fat Hydrolysis,” M. Bühler and Chr. Wandrey, Fat Science Technology 89, December 1987, pages 598 to 605; “Enzymische Fettspaltung” [Enzymatic Fat Hydrolysis], M. Bühler and Chr. Wandrey, Fett Wissenschaft Technologie [Fat Science and Technology] 89, No. 4, 1987, pages 156 to 164; and “Oleochemicals by Biochemical Reactions?” M. Bühler and Chr. Wandrey, Fat Science Technology 94, No. 3, 1992, pages 82 to 94.
In “Continuous Use of Lipases in Fat Hydrolysis,” oil is hydrolyzed continuously in a first stirred reactor. The reaction product, which in addition to free fatty acids contains water, glycerol, and mono- and diglycerides, unhydrolyzed oil and enzyme, is transferred to a solid wall plate centrifuge which is adjusted so that the intermediate layer between the aqueous glycerol phase and the organic phase is discharged with the organic phase. The organic phase containing the intermediate layer is delivered to a second stirred reactor, to which a fresh water/enzyme mixture is also added. The reaction product from the second reactor is again transferred to a solid wall plate centrifuge, which in this case is adjusted so that the intermediate layer is discharged with the glycerol-containing aqueous phase, so that the free fatty acids produced will be discharged without any intermediate emulsion layer. The aqueous phase is returned to the first reaction, so that the enzyme portion contained in the emulsion intermediate layer is resupplied to that process. Moreover, as is usually the case with self-discharging centrifuges, the solids that deposit on the drum wall are discharged discontinuously when the drum is emptied.
An emulsion-like intermediate layer also forms during the phase separation in the hydrolysis carried out according to the present invention, with or without simultaneous esterification. This intermediate layer contains substantial quantities of enzyme, along with the organic phase that contains the free fatty acids or their esters which are formed, and with the glycerol- containing aqueous phase. However, the objective is accomplished successfully with a centrifuge having exceptionally high capability, and an efficient and nearly loss-free recycling of the enzyme is achieved. This makes it possible to perform the fat hydrolysis with a high cleavage rate and the esterification in high yield, at high enzyme concentration and consequently short reaction time, without significant loss of enzyme.
A self-discharging centrifuge is used, preferably a self-discharging separator with a plate pack. The centrifuge is adjusted so that significant amounts of the enzyme-containing interfacial emulsion are not included in the ester-containing organic phase, nor in the aqueous phase. Instead, this emulsion accumulates in the centrifuge, preferably in the region of the separation zone in the plate pack of a separator. This is an unusual operating configuration to the extent that generally the accumulation of large quantities of an intermediate layer in the plate package during liquid/liquid phase separation is specifically avoided with the known self-discharging separators or self-desludging separators. On the contrary, one generally takes care that the smallest possible amount of this intermediate layer is produced, and it is generally discharged with the phase which is not the principal one to be recovered. This is also essentially what was done in the above publication, in which the intermediate layer was discharged with the aqueous phase from the second continuously operating solid wall centrifuge and resupplied to the first reactor. Furthermore, with self-discharging centrifuges, generally the solids that deposit on the drum wall are discharged discontinuously when the drum is emptied.
For one skilled in the art, adjustment of the centrifuge according to the present invention means applying the measures known to the art for adjusting the weir and/or adjusting the back pressure at the exit port to insure that both the organic phase and the aqueous glycerol-containing phase discharge as clear and free of emulsion as possible.
In the process of the present invention, the presence of alcohols does not result in either significantly higher enzyme consumption or substantially extended reaction time. The recovery of the esters from the reaction mixture in a simple manner is also possible.
The alcohols are added in the stoichiometrically required proportion for ester formation, but it is advantageous to use an excess of 2% to 100%, preferably 5% to 20%, based on the stoichiometric requirement for the corresponding oils or fats. An excess of alcohol accelerates the fat hydrolysis and unexpectedly causes complete hydrolysis of all glycerides in a short time.
The glycerol formed in the reaction moves into the water phase because of its extremely high solubility in water and very low solubility in the hydrophobic organic phase. Since the medium-chain to long-chain alcohols are very poorly soluble in water but quite soluble in the organic phase, while water on the other hand is very poorly soluble in the organic phase, these alcohols are converted enzymatically to fatty acid esters according to the chemical equilibrium. This occurs either by esterification of the fatty acid derived from the fat hydrolysis with the loss of water, where this latter product migrates to the aqueous phase, or by transesterification of the oils or fats with loss of glycerol, where this latter product likewise moves into the aqueous phase. Thus, the chemical equilibrium in the organic phase lies completely-on the side of the ester.
The addition of more than the stoichiometric amount of alcohol will shift the equilibrium further toward the ester side, and the rate of reaction will be increased significantly. In many cases the enzymatic fat hydrolysis of the present invention is nearly complete even with the addition of a slight excess of 5% over the stoichiometric amount of alcohol. This has been demonstrated for a series of n- and iso-alcohols from C8 to C24. The hydrophobic organic phase contains no mono-, di- or triglycerides. It consists solely of the fatty acid esters of the added alcohol, a small amount of the free fatty acids along with the corresponding amount of alcohol, and the excess alcohol. It is possible to remove the excess alcohol, including the unreacted alcohol portion, and the free fatty acids from the organic phase, in order to obtain the pure esters.
The methods used to recover the pure esters depend on the nature of the fatty acids and the alcohols. For instance, if esters of C18 fatty acids and C18 alcohols are produced from corresponding oils or fats and alcohols, it is possible to remove the free C18 fatty acids along with the excess C18 alcohol from the end product of the hydrolysis/esterification reaction by distillation, because the fatty acid esters produced have a substantially lower vapor pressure than the free fatty acids and alcohols.
It is possible for the fatty acids and unreacted alcohol removed by distillation to be recycled to the combined hydrolysis/esterification reaction, so that they will likewise be converted to esters without loss. In continuous operation, then, only the stoichiometric amount of the starting oil is added, while the residual amounts of alcohol and the free fatty acids are recycled.
In some cases, the free fatty acids and the resulting esters may have approximately the same vapor pressure. This occurs, for example, in the esters of C18 fatty acids with C 13 iso-alcohols. In these cases, it is possible to remove the excess amounts of C13 iso-alcohol by distillation and to separate the free C18 fatty acids from the ester/fatty acid mixture in the distillation pot by base extraction, followed by neutralization, so that these are available for recycling.
One interesting variant of fat hydrolysis with an integrated esterification or transesterification occurs with use of oils in which the fatty acids are not randomly bound to the alcohol groups of the glycerol but rather systematically, as for example the erucic acid in crambe oil. In this case, with proper selection of the hydrolyzing enzyme, it is possible to target the production of only the erucic acid esters of the added alcohols and monoglycerides of the other fatty acids of the crambe oil, or erucic acid diglycerides and fatty acid esters of the non-erucic acids. The resulting mixtures are separated by methods known to the art, preferably by distillation. For example, it is possible to separate by distillation the mixture of erucic acid diglycerides, excess C13 iso-alcohol, and C18 fatty acid—C13 iso-alcohol ester obtained, after using a chain-length specific hydrolyzing enzyme and iso—C13 alcohol as the alcohol component. Particularly after the separation of a component formed selectively by a specific enzyme (e.g., the C18 fatty acid—C13 iso-alcohol ester), it is advantageous to subject the mixture of substances resulting from the enzymatic hydrolysis/esterification to a subsequent, second enzymatic hydrolysis process using another enzyme. This method allows one to obtain the fatty acids or fatty acid esters for which this latter enzyme exhibits selective catalytic action in the hydrolysis and esterification process. For example, it is possible to convert an erucic acid diglyceride, free of C 18 fatty acids as explained later, to glycerol and erucic acid, or to an erucic acid ester in the presence of an alcohol, in a second enzymatic hydrolysis process using an enzyme which is not chain-length specific.
The process of enzymatic fat hydrolysis starting from oils and fats of the present invention thus allows the targeted production of fatty acids and fatty acid esters which previously could be obtained only at substantially higher cost. This process utilizes the known action of lipases as biocatalysts that establish a chemical equilibrium between esters, alcohols, water and acids. They act particularly on fats and oils. The latter are glycerol esters, primarily triglycerides, of medium- to long-chain and generally unbranched fatty acids. Lipases operate at the phase boundary of two-phase systems having oil or fatty acids as the hydrophobic phase and water as the hydrophilic phase, and establish a chemical equilibrium in both phases. The position of the chemical equilibrium is determined by the concentrations of the particular materials in the particular phases.
In the phase system of water and oil or native fatty acid, the water concentration is dominant in the water phase but very slight in the oil phase. As glycerol is very highly soluble in water, but hardly soluble at all in the hydrophobic phase of oil or hydrophobic fatty acids, the glycerol concentration at equilibrium must be substantially higher in the water phase than in the oil phase. Glycerol present or formed in the oil phase passes almost completely into the water phase. Native fatty acids with medium to long fatty acid chains are hydrophobic and almost insoluble in water. Their concentration in the water phase is consequently very low. On the other hand, they are quite soluble in the hydrophobic phase, and they themselves will occasionally constitute the hydrophobic phase. When lipases act on the oil at the water/oil interface, the oil is hydrolyzed with the consumption of water to diglycerides and monoglycerides, and finally to fatty acids and glycerol. At equilibrium, the law of mass action applies to both phases:
For the system of reaction equations for hydrolysis of triglyceride to fatty acids:
Then, from the law of mass action:
K = K1· K2· K3 = {[triglyceride] · [H2O]3}/{[glycerol] · [fatty acid]3}
The water concentration [H2O] in the organic phase is low and constant. As glycerol goes predominantly into solution in the water phase, the glycerol concentration, [glycerol], in the organic phase is also low and thus quasi-constant. Thus:
Because of the differences in the solubilities of the individual components in the hydrophilic and hydrophobic phases, lipases hydrolyze fats and oils almost completely to glycerol and fatty acids when the water concentration in the hydrophilic phase is high. The glycerol that is formed thus dissolves in the water, and the fatty acids initially dissolve in oil, but later form a separate hydrophobic fatty acid phase.
If the enzymatic fat hydrolysis of the present invention is carried out in the presence of added alcohols other than the glycerol contained in the fats and oils, the lipases will likewise produce a chemical equilibrium in the hydrophilic and hydrophobic phases. Exceptions to this are found for alcohols and alcohol concentrations that inhibit the activity of the enzyme, or which are incompatible with it and inactivate it. Even in this case, the position of the chemical equilibrium is determined by the distribution coefficients of the individual components between the hydrophilic and hydrophobic phases. The calculation or estimation of the chemical equilibrium distribution in two phases is naturally more complex than for simple fat or oil hydrolysis. That is particularly true for multifunctional alcohols which are water-soluble, for example trimethylolpropane and pentaerythritol.
The behavior of hydrophobic alcohols that are practically insoluble in water (both monohydric and polyhydric alcohols) is considerably easier to calculate. The medium-chain (starting with a C6 chain length) to long-chain n- and iso-alcohols of the present invention are included in this group. Such alcohols when added to a two-phase system will in practice be soluble only in the organic phase. The glycerol-containing aqueous phase will contain very little alcohol as a consequence of its low saturation concentration. Addition of such alcohols shifts the hydrolysis equilibrium toward hydrolysis because of esterification and consumption of free fatty acids. This effect is increased by the addition of excess alcohol, so that there will no longer be any tri-, di- or monoglycerides in the organic phase at equilibrium. For such alcohols, which are quite soluble in the organic phase, the above reaction in the presence of alcohol is described as follows:
This is essentially independent of the nature of the alcohol:
In contrast to water-soluble alcohols, for alcohols which are quite soluble in the organic phase: [alcohol] is saturated in H20, and so is constant. Thus:
The process of the present invention is carried out very successfully with alcohols whose water solubility is less than 5 wt.%, based on the aqueous phase. The yield and conversion rate decrease with increasing water solubility of the alcohol. For instance, when TMP was used as the alcohol in the process of the present invention, the conversion rate decreased by about 50%.
Some lipases, called specific lipases, are unable to hydrolyze all the fatty acid ester glyceride bonds. In particular, it is not possible for certain lipases to hydrolyze the central fatty acid bonded to the glycerol C2. Such lipases are used to target the production of monoglycerides and fatty acids, for example. If the starting oils contain certain fatty acids that are not bound in the glyceride of the oil or fat randomly, but rather are systematically distributed, then it is possible to obtain fatty acids or their esters which do not correspond to the fatty acid pattern present in the triglyceride though the use of specifically acting lipases.
For instance, it is known that long-chain fatty acids with chain lengths >20, such as erucic acid, are always bound to the outer hydroxyl groups of glycerol in native oils and fats, and not to the central hydroxyl group. In oils rich in erucic acid such as crambe oil, for instance, with more than 60% by weight erucic acid (and about 6% by weight of fatty acids >C22), practically all the fatty acids having chain lengths >C20 are bound to the glycerol C1 and C3, while the remaining 33.33 mole-percent of C18 fatty acids are bound to the glycerol C2. In this case, a specific lipase is used to target the cleavage of only the terminal acids and produce their esters. Then, the erucic acid esters are isolated by fractional crystallization, for example. In this case, the enzyme also acts at the phase boundary layer, and it is possible for it to be recycled efficiently according to the present invention.
It is also known that the activities of particular lipases often depend on the chain length of the particular fatty acids or on the degree of and the configuration (cis, trans, conjugated or not conjugated, etc.) of unsaturation of the fatty acids. The process of the present invention also utilizes these effects for targeted production of fatty acids or their esters.
In this case, too, the enzyme catalysis occurs in the boundary layer between the hydrophilic and hydrophobic phases. Recycling of the enzyme and removal of components enable the targeted production of the desired products. This is explained with the above-mentioned example of crambe oil.
The nonspecific lipase from Candida rugosa cleaves long-chain fatty acids such as erucic acid considerably more slowly than C16 and C18 fatty acids. When carried out according to the present invention, the enzymatic fat hydrolysis using Candida rugosa lipase yields the 1,3-diglyceride of erucic acid along with the fatty acids cleaved from the glycerol C2. These latter are C 18 fatty acids, and these fatty acids and their esters are separated advantageously from the diglyceride as the distillate when short-path distillation is used. It is possible to realize similar results for many other fatty acids such as omega-3- and omega-6-fatty acids by using suitable enzymes.
As noted above, the separation of the fatty acids from the reaction products of the hydrolysis reaction or, alternatively, the separation of the fatty acid esters from the reaction products of the hydrolysis/esterification reaction, is preferably carried out by vacuum distillation, and especially by gentle short-path distillation.. If the fatty acid esters produced have lower vapor pressure than the free fatty acids and alcohols, then the distillate contains the excess amount of n- or iso-alcohols, the unreacted alcohol, and the free fatty acids. The distillate is preferably recycled back to the hydrolysis/esterification process. It is possible to use adsorptive separation (such as column chromatography) as an alternative to the separation of the fatty acid esters or fatty acids by distillation.
Vacuum thin-film evaporators such as falling film evaporators or short-path stills are gentler than batch distillation, and above all are operated continuously, and so their use is preferred. In any case, these distillation techniques also require a liquid residue of at least 5% to 10%, as otherwise the distillation film will break. This condition is satisfied in the present invention for both the hydrolysis and the combined hydrolysis/esterification processes.
As previously discussed, the residue from the distillation in the combined hydrolysis/esterification process contains either the desired ester, or the ester and unreacted free fatty acids. In the case of a reaction involving only hydrolysis, the distillate contains the hydrolyzed fatty acids. The pot, or distillation residue, contains the unreacted triglycerides, and is recycled back to the hydrolysis process.
The emulsion-like intermediate layer collected in the centrifuge according to the present invention, in the plate pack of a separator, according to the preferred embodiment, is discharged discontinuously by completely emptying the drum periodically, and the enzyme-containing intermediate layer obtained is reused. In this case it is convenient always to empty the drum when the discharged organic phase and/or the glycerol-containing aqueous phase begins to become turbid with the discharged intermediate layer. It is also possible to empty the drum more frequently at specified times, but this is not preferred. The fact that both the aqueous and organic phases are also discharged when the drum is emptied fully is not disadvantageous because all the phases are reused by recycling them back into the reactor. It is also possible for the drum to be emptied partially instead of fully, and these procedures should be arranged so that the intermediate layer is discharged as completely as possible.
The losses of enzyme through discharge with the aqueous and organic phases are reduced drastically in the manner shown. The discharge losses in the process of the present invention are very slight compared with time-dependent enzyme consumption (enzyme aging).
A technically interesting supplement to the invention for further reducing the enzyme loss in the separated organic phase consists of using an additional self-discharging polishing separator. This additional separator is configured immediately after the separator for the single or last hydrolysis or hydrolysis/esterification step and receives the organic phase discharged from it. This added separator is preferably a self-discharging centrifuge with a plate pack adjusted so that the solids undergoing sedimentation, the enzyme in this case, and the remnants of the aqueous phase which are still removable will separate at the drum wall. Then the quantities of enzyme centrifuged off in this manner are discharged discontinuously again and recycled back to the hydrolysis or hydrolysis/esterification process.
When a self-discharging centrifuge is used to separate the organic phase containing the fatty acid esters, it is particularly advantageous to carry out the hydrolysis reaction in loop reactors, or in other words, intermittently or batchwise, not continuously in flow reactors. Thus, for instance, a reactor is filled with oil, so that its loop is not active for circulating the oil or fat, water, alcohol and enzyme contained in the reactor. At the same time a second reactor carries out the hydrolysis or hydrolysis/esterification reaction with an active loop. Also at this time, a third reactor is being emptied through an self-discharging centrifuge, so that the reaction mixture is separated into the glycerol-containing aqueous phase, the organic phase containing the separated fatty acids or fatty acid esters, and the enzyme-containing emulsion boundary phase which forms as an intermediate layer. The emulsion boundary phase is discharged discontinuously from the separator from time to time, replenished with fresh enzyme, and returned to the reactor.
It is possible to run the reaction at a high enough enzyme concentration as to produce an unusually large phase boundary due to the circulation and the shear fields produced in the circulation pumps during the loop reactor operation. Thus it is possible to keep the amount of added water low and to obtain an aqueous phase with a considerably higher glycerol concentration than was previously possible in the hydrolysis processes mentioned. In this case, there is no significant lengthening of the reaction time even with more than 30 wt.% glycerol in the discharged aqueous phase. It has not previously been considered feasible to obtain such high glycerol concentrations. Furthermore, the high glycerol concentrations will drastically reduce the loss of enzyme.
An advantage of the present invention is that the amount of added water is low, both in the case involving only the hydrolysis process, as well as for the combined hydrolysis/esterification process. In the latter case, a minimum of 5% water by weight is added, based on the organic phase, including the oil or fat and alcohol used. It is possible to add more than 200% by weight water, but that just complicates the entire process unnecessarily. In order to take advantage of the high glycerol concentration in the range of 10 to 35% by weight of the resulting glycerol-containing aqueous phase made possible in the present invention, the amount of added water should preferably be in the range of 20 to 30% by weight based on the organic phase used, and not over 50% by weight. Even in the process involving only hydrolysis, one works preferably in the range of 5 to 200% added water based on the organic phase used, preferably 20 to 30% and maximally 50% by weight.
It is possible to run the process of the present invention at a high enzyme level without consuming much enzyme. According to the present invention, even when the enzyme activity is high, the amount of lipase added to the reactor as an effective amount is generally at least 0.01 wt.%, based on the oil or fat used. The present preferred range for the amount of lipase used in the working examples is between 0.1 and 0.5% by weight, based on the oil or fat used. This high enzyme level greatly accelerates the hydrolysis as well as the hydrolysis/esterification process. In addition, the actual enzyme consumption is very low because the enzyme is recycled, so that only fractions of the initial amounts need to be added later. In actual runs, the amounts of lipase added later were less than ten percent of the active amount placed in the reactor. One skilled in the art knows that the optimal enzyme level is selected not only for the specific enzyme, but also depends on the activity of the particular enzyme preparation. Finally, one must consider that the process becomes steadily slower as the enzyme level in the process decreases. It is also known, and has previously been stated in the publications mentioned above, that increasing the enzyme level above certain values gives no advantage with respect to the process technology or economics. It is possible for one skilled in the art to determine the optimum enzyme level for the particular starting materials with a few experiments by considering these facts.
According to the present invention, it is possible for the hydrolysis or hydrolysis/esterification reaction to be carried out in just one stage in the presence of alcohol with complete release of the glycerol, with recycling of the enzyme in this stage.
However, the hydrolysis or hydrolysis/esterification reaction of the present invention is preferably carried out in multiple stages with circulating reactors, for example in two stages. The aqueous glycerol phase obtained in the second stage is then recycled back to the first stage as the aqueous phase, and fresh water as the water aqueous phase as along with the organic phase obtained from the first stage are also cycled to the second stage. Such a two-stage or multi-stage reaction is advantageous because the enzyme loss is further minimized.
The process of the present invention is also suitable for hydrolyzing the mono-, di-, and triglycerides from soap stock obtained from the alkali refining of feed oils, and converting them to fatty acid esters. For this purpose it is preferable to release the fatty acids bound in the soaps by adding acid before the hydrolysis/esterification.
The invention will now be exemplified using working examples and the accompanying drawings, in which
The exit port of the separator for the second hydrolysis stage, from which the organic phase is discharged with the fatty acid esters, is connected to a vacuum short-path still, in which a short-path distillation is effected to separate the free fatty acids and the alcohols from the fatty acid esters formed, for the case presented above in which the latter have a lower vapor pressure than the former.
The residue from the distillation will contain the desired esters if the unreacted free fatty acids and the alcohol are more volatile than the desired fatty acid ester end product. This corresponds to the process shown in
If, instead, only the alcohol is obtained as the distillate, as in
The reactor is charged with a buffer solution, the triglyceride to be hydrolyzed, the particular ester-forming alcohol, and the enzyme, i.e., lipase. More enzyme is obtained in each time the separator is emptied intermittently, and is returned to the reactor of the same stage that is being filled, with each particular separator routed to a particular reactor. Thus the enzyme remains in circulation in one stage, along with the discharged proportions of free fatty acids, unhydrolyzed triglycerides, etc. This prevents the partial mixing of starting materials of different quality from the two stages. This also reduces the risk of reverse reactions. Finally, it should also be mentioned that a glycerol solution is removed from the first-stage separator as the separated heavier liquid phase and is ready for further processing. The glycerol solution from the second-stage separator is returned to the first-stage reactor that is being filled, as shown in the figures.
A slightly acidic standard solution selected and adjusted according to the conditions specified by the enzyme producer for use as the buffer solution. An aqueous solution with sodium acetate and acetic acid, adjusted to be slightly acidic, is used in the working examples. The optimal pH of the buffer solution is adjusted for the particular enzyme.
This also applies for the process control temperature. Temperatures between 25° C. and 45° C. have been tested in experiments. Here it should be noted that the temperature is slightly elevated in general because of the exothermic reaction. In the cases tested, though, it was easily possible to ensure that the aqueous and organic phases were kept liquid and that no crystallization occurred in the liquid phases.
In principle, it is possible for any type of enzyme presented in the above-mentioned Bühler and Wandery publications to be used. Various Candida rugosa enzymes were tested thoroughly. It is also suitable to use enzymes obtained from oil seeds in the process of the present invention (e.g., from castor oil). They will naturally have the advantage of a particularly selective activity for certain fatty acids. There has been scant use of such enzymes for fat hydrolysis, however, and they are generally more costly than other enzymes prepared by industrial fermentation of yeasts, molds, bacteria and the like.
Oleyl alcohol and stearyl alcohol were tested as the n-alcohols in the experimental series. Experimental series were carried out with iso-C8, iso-C10, iso-C13, iso-C16, iso-C18, iso-C20 and iso-C24 as iso-alcohols.
The esters were prepared from high-oleic sunflower oil. Depending on the application, it is possible to start with dewaxed and refined, or crude oil or fat.
It is also possible to carry out the invention with less available and certain longer-chain alcohols, from greater than C26 up to C36, and with other oils and fats.
EXAMPLES1. Enzymatic Esterification
An 80 liter stirrer vessel was charged with 20 kg of dewaxed and refined high-oleic sunflower oil 90plus®, and 22.3 kg (10% stoichiometric excess) of Isofol 20 (C20 alcohol from Fuchs Petrolub), 10.6 kg buffer solution (0.1 N sodium acetate/acetic acid, pH 4.6) and 40 g of OF Enzyme 360 (from the Meito Sangyo company) was blended in. This mixture was circulated with a centrifugal pump for 3 hours at about 40° C.
Then the combined phase mixture was moved directly by gravity feed at a rate of about 30 kg/hr to a plate centrifuge (SA 1-01, Westfalia Separator AG, Oelde) and separated continuously. The acid value of the discharged organic phase, as determined by titration with 0.1 N KOH in alcoholic solution, according to DIN 53169 and DIN 53402, passed through a maximum of about 55, and decreased to about 15 at the end.
The enzyme was discharged from the centrifuge together with small amounts of the organic phase and glycerol-water. It showed hardly any loss of activity and could be reused.
A clear oil-ester phase and a clear glycerol solution as the aqueous phase were taken from the centrifuge. The aqueous phase contained 17% by weight of glycerol as expected.
No tri-, di- or monoglyceride could be detected in the oil-ester phase by thin-layer chromatography. After a final vacuum distillation of a 4 kg aliquot, a residue of the corresponding C20 iso-ester was obtained in 95% yield, based on the amount of oil used.
The distillate contained the excess amount and the residual unreacted portions of iso-alcohol and free fatty acids.
In a laboratory experiment, a stoichiometric mixture of oil, iso-alcohol and buffer solution was added to part of the distillate, so that the alcohol excess, based on the new amount of oil used, was again 10%.
The yield likewise reached 95% over the same time and with the same amount of enzyme. Here, too, comparable quantities of unreacted iso-alcohol and free fatty acids could be distilled out of the organic phase that was centrifuged off after termination of the reaction, so that it was possible to circulate the distillate without loss. It was also shown that the enzyme used (OF 360) catalyzes the esterification of free fatty acids and iso-alcohol in the presence of an aqueous phase.
The effectiveness of the process of the present invention was further tested for the n-alcohols oleyl alcohol and stearyl alcohol. For example, an experiment was carried out with equivalent success using oleyl alcohol (MW =268.49) and the lipase used in the example above. Iso-C8, iso-C10, iso-C13, iso-C16, iso-C18, iso-C20 and iso-C24 were used as iso-alcohols. Experiments were also done successfully with crambe oil, whereby the other possibilities presented above were demonstrated. A mixture of branched C16/C18 fatty alcohols (MW =286) was also converted successfully with the lipase above according to the present invention.
In principle the process of the present invention is also applicable to synthetic fatty acid esters, such as synthetic triglycerides and other polyol esters.
2. Enzymatic Hydrolysis
Thus the exit port of the second hydrolysis stage separator, from which the organic phase with the fatty acids (instead of the fatty acid ester) is discharged, is again connected to a vacuum short-path distillation system in which short-path distillation is used to separate the free fatty acids. The residue from the distillation was sent to a stirred crystallizer in which the waxes crystallized. In connection with that, the residual oil in which the waxes and other higher-boiling components had crystallized out as solids was pumped into a filter assembly, where it was freed of those concomitants. The oil purified in that manner was then returned to the first stage for hydrolysis.
A hydrolysis of high-oleic sunflower oil was carried out. After a starting phase, 30.0 kg of a crude, unrefined high-oleic sunflower oil 90 plus (registered trademark) from the Dr. Frische GmbH company, having an acid value of 4 as determined by titration with alkaline potassium hydroxide according to DIN 53169 and DIN 53402, was charged to one of the first-stage reactors along with 7.0 kg of a buffer solution consisting of a 12% glycerol/water solution buffered with 3.0 g sodium acetate. The charged mixture of oil and buffer solution was circulated by means of a centrifugal pump with stirring and maintained at 35 - 40° C. Next, 2 kg of enzyme-enriched discharge product from the first separator stage of the start-up phase was added, along with 3 g fresh lipase from Candida rugosa (lipase in the form of a powdered solid from Meito Sangyo, Japan, 360,000 units/gram) which had also been used in the start-up phase. After 60 minutes reaction time with stirring and then being allowed to settle, the reaction mixture was separated in the first stage separator (self-discharging plate centrifuge SA1-01 from Westfalia Separator AG Co.) into an organic phase containing fatty acids and an aqueous glycerol phase. The reactor content was fed to the separator at 40 kg/hour, and the centrifuge drum was completely emptied hydraulically every 15 minutes. The discharge product from the drum was transferred to the particular first-stage reactor that was being filled. The second-stage reactor to be filled received, along with the oleic acid-containing organic phase from the first stage separator, 7.0 kg of the above-described buffer solution, 2 kg of the discharge product from the second-stage separator from the start-up phase, and replenished with 5 g fresh lipase of the type noted above. Otherwise, this process was carried out as in the first-stage fat hydrolysis, with the result that the lighter phase discharged from the second-stage separator was a clear crude oleic acid phase with an acid value of 184, corresponding to a 93% conversion for the hydrolysis (calculated as the measured acid value divided by the theoretical acid value for this mixture). The heavy phase separated by the centrifuge constituted a 12% by weight solution of glycerol in aqueous buffer solution. The crude fatty acid thus obtained in the second stage was collected in a 200 liter vessel serving as an intermediate container. This was subjected to short-path distillation in a short-path vacuum still, Type KD, from UIC Co., Alzenau-Hörstein, with an initial degassing stage to separate any traces of water. For example, in a distillation at 191° C. and system pressure of 0.014 mbar, about 8% by weight of the fatty acid used came out continuously under these conditions as a residue of non-boiling components. The other 92% by weight of the crude product distilled as oleic acid having an acid value of 199-200, with only the slightest amounts of minor constituents in the form of fatty acids with a lower vapor pressure. This was consistent with the theoretically calculated acid value, within the accuracy of the measurement. The value was somewhat higher than the theoretical acid value because of the small proportion of more volatile, shorter fatty acids.
The residual product was dewaxed, in which the higher fatty acids (chain lengths of C22 and higher), waxes, and other higher-boiling components contained in the oil crystallized out as solids and were separated by filtration. The filtered residual product was returned to the hydrolysis process at the first stage by a pump as indicated in
Some other examples were run in the same manner, including some with higher proportions of enzyme and correspondingly shorter hydrolysis times; with beef tallow, and with crambe oil. In the example with crambe oil, which contains 60% erucic acid, a mixture of the non-specific enzyme OF 360 noted above and the 1,3-specific enzyme Novozym 388 was used. The latter specifically hydrolyzes off the fatty acids bound to the 1- and 3-positions of the triglyceride structure, erucic acid in this case.
Claims
1. A process for the enzymatic hydrolysis of oils and/or fats with simultaneous enzymatic formation of fatty acid esters using lipases acting as biocatalysts and alcohols, especially n- and iso-alcohols, said process comprising:
- causing lipases, as biocatalysts for hydrolysis of oil or fat and formation of fatty acid esters in a fat hydrolysis/esterification, to act on a mixture of triglycerides, water, and an alcohol soluble in oil or fat to create a reaction mixture formed in the fat hydrolysis/esterification;
- transferring the reaction mixture to a drum of a self-discharging centrifuge for separation into a glycerol-containing aqueous phase and an organic phase that contains the fatty acid esters which have formed;
- adjusting the centrifuge so that a lipase-enriched intermediate layer formed in the centrifuge between the aqueous phase that is drained off and the organic phase that is drained off is accumulated in the centrifuge; and
- emptying contents of the drum of the centrifuge, the drum contents including the accumulated intermediate layer, at specified times for recycling to the combined hydrolysis and esterification process or are made ready for a further hydrolysis and esterification process.
2. The process of claim 1, wherein the alcohol is used in an excess of 2 to 100%, preferably 5% to 20%, based on the stoichiometric amount required for esterification.
3. The process of claim 1, wherein the amount of water added is at least 5% by weight based upon the organic phase employed that is comprised of oil or fat and alcohol.
4. The process of claim 1, wherein the alcohol used is an alcohol which is quite soluble in the organic phase formed, but is considerably less soluble in water, and especially medium-chain to long-chain n- and iso-alcohols.
5. The process o f one of claim 1, wherein the fat hydrolysis/esterification is carried out discontinuously in reactors that are run in loop operation with the reactor contents circulated by pumps, with multiple reactors being provided in parallel for a single, or for each added, reaction stage, with one of the reactors being filled with the circulating loops not active, with a hydrolysis/esterification operation being run in a second reactor having active circulating loops, with a third reactor having its circulating loops not active and being emptied through a centrifuge which separates the glycerol-containing aqueous phase formed in the hydrolysis process from the organic phase containing the fatty acid esters prior to when the fatty acid esters are separated from the organic phase.
6. The process of claim 1, wherein the lipases is selected from a group consisting of non-specific lipases, specific lipases, or mixtures of non-specific and specific lipases.
7. The process of claim 1, wherein the free fatty acids and alcohol from the organic phase are separated by distillation from the fatty acid esters that have formed and these separated free fatty acids and alcohol are returned to the hydrolysis/esterification process.
8. The process of claim 1, wherein the organic phase drained out from the self-discharging centrifuge is transferred to another self-discharging centrifuge, which is likewise emptied intermittently to recover lipase residues that have collected as a sediment on the centrifuge wall for reuse in the hydrolysis process.
9. A device for carrying out the process of the enzymatic hydrolysis of oils and/or fats with simultaneous enzymatic formation of fatty acid esters using lipases acting as biocatalysts and alcohols, especially n- and iso-alcoholsone, said device comprising:
- at least one hydrolysis/esterification reactor for use in a hydrolysis and esterification process;
- at least one self-discharging centrifuges in which a lipase-enriched phase formed in the at least one hydrolysis/esterification reactor between an aqueous phase that is drained off and an organic phase that is drained off, accumulates in a drum of the at least one centrifuge which is emptied at specified times;
- a feedback system for returning contents of the intermittently emptied drum including said lipase-enriched phase from the at least one centrifuge to the combined hydrolysis and esterification process; and a
- a means for separating alcohol, free fatty acids and fatty acid esters formed from the organic phase that is supplied from the at least one centrifuge.
10. The device of claim 9, wherein the means for separation is a distillation apparatus.
11. A process for the enzymatic hydrolysis of oils and/or fats using lipases acting as biocatalysts to obtain fatty acids and glycerol, said process comprising:
- causing lipases to act as biocatalysts on a mixture of an oil or fat and water to hydrolyze the oil or fat to produce a reaction mixtures;
- transferring the reaction mixture thus produced to a self-emptying centrifuge for separation into a glycerol-containing aqueous phase and an organic phase that contains free fatty acids that have been hydrolyzed off in the preceding hydrolysis;
- adjusting the centrifuge so as to accumulate a lipase-enriched intermediate phase that forms in the centrifuge between the aqueous phase that is drained off and organic phase that is drained off; and
- emptying the centrifuge of drum contents at specified times and the centrifuge drum contents that have been emptied from the centrifuge are returned to the hydrolysis process or are prepared for a further hydrolysis process.
12. The process of claim 11, wherein the fat hydrolysis is carried out discontinuously in reactors that are run in loop operation with the reactor contents circulated by pumps, with multiple reactors provided in parallel for a single or for each of numerous reaction stages, with one of the reactors being filled with the circulation loops not active, with a hydrolysis operation being run in a second reactor having active circulation loops, with a third reactor having its circulating loops not active and being emptied through a centrifuge which separates the glycerol-containing aqueous phase formed in the hydrolysis process from the organic phase containing the free fatty acids prior to when the fatty acids are separated from the organic phase.
13. The process of claim 11, wherein the free fatty acids and alcohol are separated by distillation out of the organic phase from the fatty acid esters that have formed, and are returned to the hydrolysis/esterification process.
14. The process of claim 11, wherein the organic phase flowing out of the self-discharging centrifuge is transferred to another self-discharging centrifuge, which is likewise emptied intermittently to recover residues of lipase that have collected as a sediment on the centrifuge wall for reuse in the hydrolysis process.
15. A device for carrying out the process for the enzymatic hydrolysis of oils and/or fats using lipases acting as biocatalysts to obtain fatty acids and glycerolany, said device comprising:
- at least one hydrolysis reactor for use in a hydrolysis process to form a reaction mixture;
- at least one self-discharging centrifuges in which the reaction mixture is separated into an aqueous phase that is drained off and an organic phase that is drained off, wherein a lipase-enriched intermediate layer being formed between said aqueous phase and organic phase is accumulated in the centrifuge, and which is emptied at specified times;
- a feedback system for transporting contents of the intermittently emptied at least one centrifuge including the intermediate layer from the centrifuge to the hydrolysis process; and
- a means for separating free fatty acids from the organic phase that is supplied from the centrifuge.
16. The device of claim 15, wherein the means for separation is a distillation apparatus.
17. The device of claim 16, wherein said distillation apparatus is selected from a group consisting of a short-path still and a falling film evaporator.
18. The device of claim 10, wherein said distillation apparatus is selected from a group consisting of a short-path still and a falling film evaporator.
19. The process of claim 14, wherein the another self-discharging centrifuge is a polishing centrifuge.
20. The process of claim 8, wherein the another self-discharging centrifuge is a polishing centrifuge.
Type: Application
Filed: Jun 4, 2002
Publication Date: Apr 5, 2007
Inventors: Karlheinz Brunner (Grosskrotzenburg), Rainer Frische (Frankfurt), Rainer Ricker (Dietzenbach), Corinna Kaske (Florsbachtal), Dirk Kilian (Maintal)
Application Number: 10/531,658
International Classification: C12P 7/64 (20060101);
