JPH0358276B2 - - Google Patents

Description

[Detailed description of the invention]

[Industrial Field of Application] The present invention relates to a method for transesterification of fats and oils using lipase (lipid degrading enzyme). More specifically, the present invention relates to a method for transesterification of fats and oils using a lipase preparation pre-wetted with a lipase activator. Transesterification of fats and oils is an important technology, along with hydrogenation, in the production of processed fats such as margarine and shortening. [Prior Art] Transesterification reactions of fats and oils have conventionally been carried out chemically, ie, by using alkaline substances such as alkali metal alcoholates, alkali metals, and alkali metal hydroxides as catalysts.
However, in this method, indiscriminate exchange occurs in the positions of fatty acids in fats and oils, so no specificity is observed in the positions of fatty acids in the resulting fats and oils. That is, the disadvantage of conventional chemical transesterification is that it is non-selective regarding the position of fatty acids in fats and oils. Recently, methods for site-specific transesterification of fats and oils have been developed in place of conventional non-selective methods. That is, a typical example is a method of transesterifying fats and oils using lipase, which is oxygen that hydrolyzes fats and oils (Japanese Unexamined Patent Publication No. 104506/1983). According to this method, the presence of water in the reaction system is an essential requirement for activating lipase. Although this water content is small at 0.2 to 1.0%, lipase is originally an oxygen that hydrolyzes fats and oils in the presence of water, so as long as a small amount of water is present, diglycerides etc. are produced by hydrolyzing fats and oils. By-products or a decrease in the yield of exchange fat cannot be avoided. Diglycerides and the like, which are by-products, greatly impair the properties of the fats and oils that are intended for exchange fats, so a complicated separation and purification process is required to remove them. Thus, the known methods are not satisfactory. Under these circumstances, various methods have been proposed for overcoming the drawbacks of the above-mentioned known methods, suppressing hydrolysis of fats and oils, and efficiently performing transesterification. Specifically, they are as follows. (a) A method of suppressing the hydrolysis of fats and oils by using a lower polyhydric alcohol as a lipase activator in place of water during transesterification of fats and oils (Japanese Patent Publication No. 57-6480). (b) Focusing on the fact that the transesterification reaction of oils and fats occurs at the interface of a heterogeneous system consisting of oil and water (lipase is water-soluble), we added a surfactant (emulsifier) to this heterogeneous reaction system. A method of efficiently bringing oil and fat into contact with lipase at an interface by adding a lipase (Japanese Unexamined Patent Publication No. 198798/1983). (c) A method of increasing the rate of transesterification by controlling the water content using a water-absorbing resin that has the characteristic of absorbing several hundred times its own weight in water (Japanese Patent Laid-Open No. 116689/1989). (d) In transesterification of fats and oils, instead of using a fatty acid with a high melting point, a lower alcohol ester of the fatty acid with a low melting point is used to conduct the reaction more uniformly (Japanese Patent Publication No. 57-27159
Publication No.). (e) A method for increasing the rate of transesterification and at the same time suppressing the hydrolysis of fats and oils by controlling the water content in the reaction system by dry circulation of solvent vapor during transesterification of fats and oils (Special Publication No. 58
-500638). However, each of these known methods has several drawbacks, so none of them can be said to be fully satisfactory. Specific examples of these drawbacks are as follows. The feature of method (a) is that a lower polyhydric alcohol such as glycerin is used instead of water as a lipase activator. However, according to the study results of the present inventors, although the effect of suppressing the hydrolysis reaction to some extent is recognized, the transesterification reaction rate is extremely slow, and it takes nearly a week to obtain the desired reaction rate. A shortcoming was discovered: In the method (b), the addition of a surfactant (emulsifier) is said to effectively bring the fat and oil into contact with the lipase at the interface between the oil layer and the water layer, thereby selectively promoting transesterification. In other words, the formation of reverse micelles on the surface of the enzyme protein provides a condition in which it is easy to form a complex between the lipase and the substrate, resulting in good transesterification reactivity. It is considered. However, as disclosed in the examples in the publication, the suppression of the hydrolysis reaction is insufficient;
Furthermore, surfactants (emulsifiers) remain in the transesterified fat, which may impair the physical properties of the fat. Therefore, it is necessary to remove these emulsifiers from the transesterified fat, which poses an obstacle in terms of industrial implementation, as it requires complicated processing steps. Regarding the method (c), it is not possible to sufficiently suppress the hydrolysis of fats and oils, and furthermore, there is a risk that the raw material monomers present as impurities in the resin may be eluted into the fats and oils. Further, according to additional tests conducted by the present inventors, the absorbent resin swells when it comes into contact with water and does not adhere to the walls of the reaction vessel. This results in loss of lipase when recovery and reuse of lipase is considered. In method (d), a fatty acid ester must first be separately produced prior to transesterification of fats and oils. Therefore, this causes the process to become complicated. In method (e), until the water in the reaction system is removed from the system by solvent circulation drying, there is a risk that the lipase will be inactivated due to the large amount of water. this is,
This is a major obstacle when collecting and reusing lipase. As mentioned above, all of the conventionally known methods have several drawbacks, and these drawbacks become an obstacle when considering industrial application. In addition to the above-mentioned known methods, various methods have been proposed, but a specific method for suppressing hydrolysis of fats and oils and performing only transesterification has not yet been found. Under these circumstances, the present inventors have conducted intensive studies on a method for suppressing the hydrolysis of oils and fats and efficiently performing only transesterification. ), and filed a patent application (Japanese Patent Application No. 110334/1982). [Problems to be Solved by the Invention] However, the transesterification reaction of fats and oils using the lipase preparation described above has drawbacks such as a long reaction time. The long reaction time is disadvantageous when considering implementation on an industrial scale, and furthermore, there is a risk that the lipase will deteriorate if the enzyme catalyst is used for a long time. [Means for Solving the Problems] From this perspective, the present inventors have conducted intensive studies on methods for efficiently carrying out transesterification reactions in a short period of time, and have found an effective and simple method for achieving the objective. We discovered a method for activating lipase in lipase preparations and completed the present invention. That is, the present invention can be obtained by adding fats and oils to a mixture consisting of a lipase activator, lipase, and a carrier and reacting them to decompose the fats and oils, and then removing the fats and oils from the decomposition product by filtration or the like. In the presence of a lipase preparation that has been moistened with a lipase activator before being used in the transesterification reaction, the transesterification reaction between fats and oils or between fats and oils or between fats and oils is carried out. The present invention relates to a characteristic method for transesterification of fats and oils. According to the present invention, the reaction time of the ester exchange reaction can be greatly shortened, and the hydrolysis of fats and oils can also be suppressed. The method of the present invention will be explained in more detail as follows. First, the manufacturing method for the lipase preparation is as follows. lipase activator (e.g. water or dihydric or trihydric lower alcohol),
A lipase preparation can be obtained by adding fats and oils to a mixture consisting of lipase and a carrier and reacting them to decompose the fats and oils, and then removing the fats and oils separately from the decomposition product. The obtained lipase preparation can be used as it is or, if necessary, after being washed with a solvent (hydrocarbon) that does not impair lipase activity and further subjected to a drying treatment, it can be used in a wet treatment. In addition to the above-mentioned lipase preparations, lipase preparations that have been used at least once in transesterification of fats and oils can also be used as lipase preparations. Alternatively, it can be used for wet treatment after washing with a solvent (hydrocarbons) that does not impair lipase activity and further drying, if necessary. A lipase activator (water, divalent or trivalent lower alcohol, etc.) used in the transesterification reaction of fats and oils was added to the lipase preparation obtained as described above, and a wetting treatment was performed by stirring or standing still. After that, this is added to a reaction mixture consisting of fats and oils, fatty acids, and solvents (hydrocarbons) and reacted, or added to a mixture consisting of fats and oils (fats and oils) and solvents (hydrocarbons) and reacted. Depending on the situation, transesterification of fats and oils can be carried out. From the exchanged fat obtained by transesterification, liquid
Purified exchange fat can be obtained by removing fatty acids, small amounts of monoglycerides, diglycerides, etc. using conventionally known separation and purification means such as liquid extraction, alkali neutralization, vacuum or molecular distillation, alone or in combination as appropriate. . Regarding the lipase used in the present invention, if the selectivity in the transesterification reaction by lipase is poor, it will not have any particular superiority over the conventional transesterification reaction using an alkali metal catalyst. For example, it is preferable to have selectivity for the position where it binds to glyceride or selectivity for the type of fatty acid. Specifically, lipases with excellent regioselectivity include, for example, Rhizopus, Aspergillus, and Mucor lipases, pancreatic lipase, and the like. When specifically transesterifying fatty acid groups at the 1 and 3 positions of glycerides, lipases having properties that meet the purpose include, for example, Rhizopus delemar , Rhizopus japonicus , and Mucor japonicus. ( Mucor japonicus ) may be used, and these lipases are available as commercial products. As the lipase activator, water or divalent or trivalent lower alcohol is suitable, and among these, water or glycerin is particularly effective. The carrier can be selected from known carriers, such as celite, diatomaceous earth, kaolinite, perlite, silica gel, glass fiber, activated carbon, cellulose powder, calcium carbonate, etc.
It can be used as long as it is insoluble in the lipase preparation production system and the transesterification reaction system and does not adversely affect lipase activity. There are various forms of the carrier, such as powder, granules, and fibers, and any of them can be used. The oils and fats used in the present invention include general vegetable oils, animal oils, processed oils and fats, and mixtures thereof. Specific examples include soybean oil, cottonseed oil, rapeseed oil, olive oil,
Examples include corn oil, coconut oil, safflower oil, beef tallow, lard, and fish oil. Furthermore, if the purpose is to use a cocoa butter substitute fat in the transesterification reaction,
Oils and fats containing a large amount of oleic acid at the 2-position of glycerides, such as palm oil, olive oil, camellia oil,
Sasanqua oil, monkey oil, iritupe butter, kokum butter, shea butter, mora butter, furwara butter, Borneo tallow butter, or fractionated fats and oils thereof can be used.
Note that the fats and oils used in the production of the lipase preparation and the fats and oils used in the transesterification reaction can be arbitrarily selected independently of each other, but the fats and oils used in the transesterification or the fats and oils with a composition similar to these may be selected independently. It is desirable to use it when manufacturing lipase preparations. Transesterification of fats and oils is carried out by reacting fats and oils with fatty acids, or reacting fats and oils with fats and oils. As the fatty acid, those that are straight chain having 8 to 22 carbon atoms and normally exist in nature are used. For example, palmitic acid, stearic acid, oleic acid, etc. During transesterification, in addition to the above fatty acids,
Alcohol esters of fatty acids can be used. Alcohol esters of fatty acids include the above-mentioned fatty acids (straight chain fatty acids having 8 to 22 carbon atoms) and 1 carbon atom.
-6 esterified products of linear saturated monohydric alcohols are used. For example, methyl palmitate, ethyl palmitate, methyl stearate, ethyl stearate, etc. can be used. Oils and fats are
Any oil or fat can be selected from the above-mentioned oils and fats (general vegetable oils, animal oils, processed oils and fats, or mixtures thereof) depending on the purpose. When carrying out the transesterification reaction of the present invention in a solvent, an organic solvent inert to lipase can be used as the solvent. Examples of this type of organic solvent include n-hexane, industrial hexane, petroleum ether, petroleum benzene, and the like. Incidentally, the solvent used in transesterification can be used as the solvent used in producing the lipase preparation. A more specific method of the present invention is as shown below. First, a lipase preparation is manufactured as follows. That is, a lipase preparation is obtained from fats and oils, a lipase activator (water or divalent or trivalent lower alcohol), lipase, and a carrier. This is done by adding 0.01 to 10 parts by weight of lipase (commercially available), 0.1 to 20 parts by weight of a lipase activator, and 1 to 50 parts by weight of a carrier to 100 parts by weight of fat and oil, and adding 1 to 10 parts by weight of a carrier at 20 to 80°C. Fats and oils are decomposed by reacting for 24 hours. Although the decomposition temperature is within the range mentioned above, it is desirable to select a temperature suitable for the action of lipase. A lipase preparation is obtained by separately removing the fat portion from the fat decomposition product. This lipase preparation may optionally contain an inert organic solvent that does not impair the activity of lipase, such as the above-mentioned n-hexane,
After washing the lipase preparation with hydrocarbons such as petroleum ether, it is subjected to drying treatment (heat drying, etc. is undesirable as it impairs enzyme activity). The lipase formulation prepared as described above is wet treated with a lipase activator. The transesterification reaction using the infiltrated lipase preparation is carried out as follows. That is, 25 to 100 parts by weight of fats and oils
300 parts by weight of fatty acids (or alcohol esters of fatty acids or other fats and oils), lipase preparation 0.1~
100 parts by weight (the lipase preparation consists of lipase and a carrier, and lipase is used in a range of 0.01 to 10 parts by weight), 0.01 to 10 parts by weight of a lipase activator (water or divalent or trivalent lower alcohol) and 1 part by weight. 0.01 to 100 parts by weight of an inert organic solvent that has been wet-treated for more than an hour (preferably several hours or more) at a temperature that does not inhibit lipase activity (preferably 25 to 30°C), and further 0 to 100 parts by weight if necessary. It is carried out by stirring a mixture consisting of at 20-80°C. The transesterification reaction of the present invention is normally carried out within the above temperature range, but it is desirable to select a temperature suitable for the action of lipase. Fatty acids, small amounts of monoglycerides, diglycerides, etc. are extracted from the reaction solution after the transesterification reaction by liquid-liquid extraction, alkali neutralization, or by using conventional separation and purification methods such as vacuum or molecular distillation alone or in combination as appropriate. It can be easily removed.
In this way, purified exchange fat can be obtained. [Effects of the Invention] The effects or advantages of the present invention are as follows: Using a lipase preparation with high transesterification activity obtained by a simple method, before using it in the transesterification reaction,
By performing a wet treatment, the activity is further increased, and only the desired transesterification can be carried out efficiently in a short period of time. At the same time, the hydrolysis reaction of fats and oils can be suppressed, making it an extremely productive method. It provides: Another effect of the present invention is that the deactivation of lipase during the reaction is reduced due to the shortening of the reaction time, and it is possible to effectively reuse the lipase preparation recovered after the reaction. Improve its economy in implementation. Further, according to the method for transesterification of fats and oils according to the present invention, by using a regioselective lipase, for example, an expensive cocoa butter substitute can be effectively produced from inexpensive palm oil. [Example] Hereinafter, the present invention will be explained in detail with reference examples, examples, comparative examples, etc. Reference example (Manufacturing example of lipase preparation) 100 g of soft palm oil, 10 g of Celite, 1.0 g of ion-exchanged water, and 8.7 g of commercially available lipase (manufactured by Tanabe Seiyaku Co., Ltd., lipase originating from Rhizopus deremer) were heated at 40°C for 18 hours. The enzymatic reaction (hydrolysis) was carried out by stirring in a closed container. After the reaction was completed, the insoluble material (a mixture of Celite and lipase) was separated and washed three times with 5 ml of n-hexane to completely remove fats and oils. Then under reduced pressure,
A lipase preparation was obtained by drying at 20-30°C for 1 hour. Example 1 Transesterification reaction using a lipase preparation (when subjected to wet treatment) The lipase preparation obtained in Reference Example (0.87 g of lipase,
(consisting of 1.00g of Celite) and 1.87g of ion-exchanged water.
The wet treatment was carried out for 24 hours in a closed container to which 0.015 g was added. This and palm oil medium melting point (iodine value 3
4. diglyceride content 1%) 10g, stearic acid
Stir 10 g and 40 ml of n-hexane in a closed container at 40℃ for 1 day for enzymatic reaction (ester exchange reaction)
I went there. After the reaction was completed, insoluble substances such as lipase preparations were removed separately, and n-hexane was distilled off from the solution under reduced pressure. The obtained exchange fat was subjected to column chromatography to obtain a diglyceride fraction and a triglyceride fraction. Regarding the triglyceride fraction, the stearic acid content was measured by gas chromatography. Regarding the stearic acid content and diglycerin content, the results are shown in Table 1. Comparative example 1 Transesterification reaction using a lipase preparation (without wet treatment) 1.87 g of the lipase preparation obtained in the reference example, 10 g of palm oil medium melting point, 10 g of stearic acid, ion-exchanged water
0.015 g and 40 ml of n-hexane were stirred in a closed container at 40°C for 2 days to perform an enzymatic reaction (ester exchange reaction). After the reaction was completed, the stearic acid content and diglyceride content in the produced triglyceride were determined in the same manner as in Example 1, and the results are shown in Table 1. Comparative Example 2 Transesterification reaction without using a lipase preparation (when subjected to wet treatment) 0.87 g of commercially available lipase (used in reference example),
0.1 g of Celite was mixed, 0.015 g of water was added, and a wet treatment was performed in a closed container for 24 hours. Total amount of this, 10g of palm oil medium melting point part, 10g of stearic acid and n-
Enzyme reaction (ester exchange reaction) was carried out by stirring 40 ml of hexane in a closed container at 40°C for 3 days. After the reaction was completed, the stearic acid content and diglyceride content in the produced triglyceride were determined in the same manner as in Example 1, and the results are shown in Table 1. Comparative Example 3 Transesterification reaction without using a lipase preparation (without wet treatment) 0.87 g of commercially available lipase (used in reference example),
1.0 g of Celite, 10 g of medium melting point part of palm oil, 10 g of stearic acid, 0.015 g of ion-exchanged water, and 40 ml of n-hexane were stirred in a closed container at 40° C. for 4 days to perform an enzymatic reaction (ester exchange reaction). After the reaction was completed, the stearic acid content and diglyceride content in the produced triglyceride were determined in the same manner as in Example 1, and the results are shown in Table 1.

[Table] Example 2 Transesterification reaction by repeated use of lipase preparation (when subjected to humid treatment) 18.7 g of the lipase preparation obtained in the reference example was added with 0.3 g of ion-exchanged water and subjected to humid treatment for 24 hours in a closed container. I went. Total amount of this, palm oil medium melting point 200g,
200g of stearic acid and 800ml of n-hexane at 40℃
The enzyme reaction was carried out by stirring in a closed container for 2 days. After the reaction was completed, insoluble substances such as the lipase preparation were separated, and n-hexane was distilled off from the solution under reduced pressure. The stearic acid content of the obtained exchange fat was determined in the same manner as in Example 1. The separated insoluble substances such as lipase preparations were dried under reduced pressure at 20° C. for 1 hour, then 0.3 g of ion-exchanged water was added to perform a wet treatment for 24 hours, and the mixture was used again for the transesterification reaction. The stearic acid content in each triglyceride obtained is shown in Table 2.

【table】

Claims (1)

[Scope of Claims] 1. Adding fats and oils to a mixture consisting of a lipase activator, lipase, and a carrier and reacting them to decompose the fats and oils, and then removing the fats and oils from the decomposition product by filtration or the like. Before using the lipase preparation obtained in the transesterification reaction, the transesterification reaction between fats and oils or the transesterification reaction between fats and oils is carried out in the presence of a lipase preparation that has been wet-treated with a lipase activator in advance. A method for transesterification of oils and fats, characterized by carrying out the transesterification reaction. 2 The lipase activator is water or divalent or trivalent
2. The method for transesterification of oils and fats according to claim 1, wherein the transesterification reaction is carried out using one type or a mixture of two or more types of lower alcohols.