The present invention relates to a process of an enzymatic reaction in a low water content condition (a system containing a very small amount of water) wherein the reaction is carried out with adjusting the water concentration in the reaction system to a suitably very low value in order to effectively take advantage of the activity of enzyme and in order to suppress the formation of by-product, in an interesterification of a fat or oil for obtaining desired useful products by utilizing a enzymatic reaction.
As an use of technique of enzymatic reaction in a low water content condition, a production of substitutional fats for cacao butter known as a raw material of chocolates is exemplified. Further, by choosing kind of substrate and of lipase, various highly purified products having optional constitution of fatty acids can be produced.
In recent years, active studies on a process for preparing valuable a fat or oil by interesterification using lipase have been made.
The interesterification by lipase is generally considered to be caused by a reversible reaction of a hydrolysis in which triglyceride (TG) is hydrolyzed into diglyceride (DG), monoglyceride (MG) or glycerin and fatty acid (FA), and of synthesizing triglycerde again from the hydrolysates. Thus existence of water is necessary in an interesterification system, and therefore some amount of water is added to the reaction system.
However, it is impossible to inhibit the proceeding of the hydrolysis if the amount of water added is excess, and therefore by-products such as released fatty acid, monoglyceride and diglyceride are formed in a large amount and the yield of the desired triglyceride is lowered. And consequently, there arises a problem that the quality of triglyceride as the product is Therefore, in order to effectively carry out an interesterification, the reaction system should be maintained in a low water content condition constantly.
In case, however, the amount of water in a reaction system is too little, there arises a that the catalytic activity of enzyme cannot be sufficiently invigorated and the rate of reaction is lowered.
As a means of solving the above problem in the conventional process of interesterification, there is disclosed, for instance, an addition of a water-soluble polyhydroxy compound, and a protein or a peptide in an emulsion system (Japanese Unexamined Patent Publication No. 111398/1982), or an addition of a surface active agent (Japanese Unexamined Patent Publication No. 198798/1982) to increase the rate of reaction.
On the other hand, some processes for industrial production, in which an immobilized enzyme or a microorganism is used, have been proposed. For instance, a process in which an enzyme is immobilized onto an adsorbent such as Celite (Japanese Unexamined Patent Publication No. 71797/1980, Japanese Unexamined Patent Publication No. 104506/1977, Japanese Unexamined Patent Publication No. 84397/1980 and Japanese Unexamined Patent Publication No. 78496/1982); a process in which an enzyme is immobilized onto various water-absorptive materials such as saccharides (Japanese Unexamined Patent Publication No. 116688/1983, Japanese Unexamined Patent Publication No. 116689/1983, Japanese Unexamined Patent Publication No. 187195/1983 and Japanese Unexamined Patent Publication No. No. 28482/1984); a process in which a dried microbial cell is used as a catalyst (Japanese Unexamined Patent Publication No. 34189/1985); and the like are exemplified.
For industrially carrying out an interesterification using an immobilized enzyme or a microorganism, however, in both cases of batch reaction system and continuous reaction system, it is indispensable -to-. constantly adjust the water concentration in the vicinity of the enzyme to a suitably very low concentration in order to maintain a high yield. In an interesterification, since water is consumed by a hydrolysis, it is necessary to supply water properly. For instance, a process in which the reaction is proceeded with supplying a reaction liquid in which water is dissolved in an amount of 40 to 70% by weight of the saturated amount is disclosed (Japanese Unexamined Patent Publication No. 154951/1981 and Japanese Unexamined Patent Publication No. 500649/1984). In such a process, however, it is difficult to highly precisely adjust the water concentration in the vicinity of the enzyme to a suitably very low concentration, and therefore the industrialization of this process is obstructed.
As described above, an interesterification is caused by a combination of a hydrolysis and a synthesis. In a reaction system of interesterification, when the water concentration is high, water is consumed by a hydrolysis. That is, the water concentration in the reaction system is changing every moment with the proceeding of the reaction. Therefore, the determination of the water concentration in the beginning of the reaction is meaningless, and the constant adjustment of water concentration to a suitable concentration during the reaction is the important subject in an invention.
Moreover, in a process using an immobilized enzyme or microorganism (hereinafter referred to as "immobilized enzyme catalyst"), there is required a technique for constantly adjusting the water content held in the immobilized enzyme catalyst (hereinafter referred to as "water content of catalyst") to the optimum.
In almost all the conventional processes, however, attention is paid only to the water concentration at the beginning of the reaction and there is no suggestion about a method of adjustment of the water concentration to a suitable during the reaction. Such adjustment has been said to be technically difficult.
Since the starting materials of interesterification such as a triglyceride, a fatty acid and a fatty acid ester with a lower alcohol or solvents for dilution contains a small amount of water, a diglyceride or a monoglyceride is produced by a hydrolysis, when the starting material or the solvent is used without taking any measures. It is known that diglyceride forms an eutectic mixture with triglyceride to suppress the crystallization of triglyceride. And consequently, there arise problems that the yield of triglyceride as product is lowered, and that the quality of product is deteriorated. Therefore, a technique for carrying out the reaction with lowering the water concentration in the reaction system to a lower value was required.
For an industrialization of interesterification, it is an inevitable problem that the rate of reaction is increased by utilizing a high-cost immobilized enzyme catalyst as effectively as possible, and at the same time the yield and quality of the product are improved by suppressing the proceeding the hydrolysis. And there is desired a technique for determining the water concentration in catalyst suitable for a desirable reaction and for carring out a reaction efficiently with adjusting the water concentration to the determined suitable value constantly.
The object of the present invention is to provide a process for efficiently preparing valuable a fat or oil as described above.