KR102143203B1 - Method for improving efficiency of enzymatic esterification by adjustment of peroxide value of reaction mixture - Google Patents

Abstract

The present invention relates to a method of improving the efficiency of enzymatic esterification, and more particularly, in performing a reaction of exchanging an acyl group of a triacyl glyceride with another acyl group using an enzyme, by controlling the peroxide value of the reaction mixture It relates to a method that can further improve the overall process efficiency and economy by increasing the reaction efficiency.

Description

{Method for improving efficiency of enzymatic esterification by adjustment of peroxide value of reaction mixture}

The present invention relates to a method for improving the efficiency of enzymatic esterification, and more particularly, in performing a reaction of exchanging an acyl group of triacylglyceride with another acyl group using an enzyme, by adjusting the peroxide value of the reaction mixture It relates to a method that can further improve the overall process efficiency and economy by increasing the reaction efficiency.

Triacylglyceride fats and oils are widely used in the food industry. Among them, 1,3-dioleoyl-2-palmitoyl glyceride (OPO) is a breast milk fat. It is an important ingredient in nutritional research.

Fat compositions containing the major fatty acids that mainly make up human milk fat are found in oils and fats of plant origin, but almost all glycerides of plant origin do not have saturated fatty acids in the 2-position. In contrast, in human milk fat, a significant amount of saturated fatty acid, palmitic acid, occupies the 2-position of glycerides.

Recently, as people's interest in infant health has increased, the demand for foods that have similar ingredients to breast milk is increasing.

Innis (Adv. Nutr., 2:275-283, 2011) describes the structural role of triacylglycerol in infant diet.

However, conventional powdered milk is a mixture of powdered milk and vegetable oil and hardly contains palmitic acid in the 2-position when the oil component of the powdered milk is analyzed.

Accordingly, the application of high-quality breast milk fat substitute paper to add 1,3-dioleoyl-2-palmitoyl glyceride (OPO) containing palmitic acid at the 2-position, which has a similar component to breast milk fat. Is increasing.

OPO is a kind of USU (Unsaturated-Saturated-Unsaturated) triglyceride, and Mohamad, et al (Lipase-catalyzed interesterification reactions for human milk fat substitutes production, 2013, 115, 270-285) is a tree when using lipase. It has been shown that studies of synthesizing 1,3-dioleoyl-2-palmitoyl glyceride by substituting the 1,3-position of acyl glyceride have been mainly conducted. The provision of a glyceride backbone containing a saturated fatty acid at the 2-position of the triacylglyceride fat due to the 1,3-position-specific reaction property of the triacylglyceride of lipase is 1,3-dioleoyl-2-palmi It is important in synthesizing soil glyceride (OPO).

These prior arts use an enzymatic esterification reaction, but the enzyme activity decreases as the reaction proceeds, and as a result, the efficiency of the entire process decreases. Therefore, development of a method capable of producing OPO with higher efficiency than the conventional 1,3-dioleoyl-2-palmitoyl glyceride (OPO) production method is requested.

An object of the present invention is to provide a method capable of improving overall process efficiency and economy by increasing reaction efficiency in an enzymatic esterification reaction.

In order to achieve the above technical problem, the present invention is characterized in that the peroxide value (POV) of the reaction mixture is adjusted to 50 or less in performing the reaction of exchanging the acyl group of triacylglyceride with another acyl group using an enzyme. As described above, an enzymatic esterification method is provided.

According to an aspect of the present invention, (1) an enzymatic esterification reaction is performed on a reaction mixture comprising a triacyl glyceride as a starting material and a fatty acid having an acyl group different from the acyl group in the triacyl glyceride as a reactant. A method for producing triglyceride is provided, comprising the step of exchanging the acyl group of the triacyl glyceride with the acyl group of the fatty acid, wherein the peroxide value of the reaction mixture obtained in the step is adjusted to 50 or less.

According to the present invention, in performing a reaction of exchanging an acyl group of a triacyl glyceride with another acyl group using an enzyme, the reaction efficiency of the enzymatic esterification reaction can be increased, thereby improving overall process efficiency and economy. Accordingly, according to an embodiment of the present invention, dioleoyl monopalmitoyl glyceride can be produced efficiently and economically from tripalmitoyl glyceride through an enzymatic esterification reaction.

1 is a graph showing the results of C52 concentration analysis performed in an example of the present invention.
2 is a graph showing the results of USU concentration analysis performed in an example of the present invention.

Hereinafter, the present invention will be described in detail.

Triacyl glyceride used as a starting material for the enzymatic esterification reaction in the present invention may be edible fat or oil.

As the edible fats and oils, any suitable edible material such as liquid or solid vegetable fats or animal fats, hydrogenated or transesterified oils of the fats and oils, liquid oils obtained by fractionating the fats and oils, and solid fats can be used.

Specific examples of the vegetable oils include those selected from corn oil, rapeseed oil, soybean oil, cottonseed oil, palm oil, palm oil, rice bran oil, sesame oil, cacao fat, olive oil, and combinations thereof, and specific examples of the animal oils include fish oil, What is selected from pork fat, beef tallow (beef oil), chicken fat, and combinations thereof. These edible fats and oils may be used alone or in combination of two or more.

In one embodiment, the triacyl glyceride may be palm fractionated oil obtained by fractionating palm oil among vegetable oils.

In one embodiment, the acyl group present in the triacylglyceride is independently saturated or unsaturated having 4 to 24 carbon atoms, more specifically 12 to 22 carbon atoms, and more specifically 14 to 20 carbon atoms It may be selected from linear aliphatic acyl groups. For example, the acyl group present in the triacylglyceride may be independently selected from a palmitoyl group, a stearoyl group, an oleoyl group, and a linoleoyl group, and preferably may be a palmitoyl group, but is not limited thereto.

In one embodiment, at least one of the acyl groups present in the triacylglyceride is a palmitoyl group, preferably two, more preferably all three palmitoyl groups. That is, in the most preferred embodiment, the starting material triacylglyceride is tripalmitoyl glyceride (PPP).

The fatty acid used as a reactant for the enzymatic esterification reaction in the present invention has an acyl group different from the acyl group in the triacylglyceride.

In one embodiment, the fatty acid is selected from saturated or unsaturated linear fatty acids having 4 to 24 carbon atoms, more specifically 12 to 22 carbon atoms, and even more specifically 14 to 20 carbon atoms. You can use more than one. More specifically, the fatty acid may be selected from the group consisting of palmitic acid, stearic acid, oleic acid, linoleic acid, and combinations thereof, and preferably oleic acid, but is not limited thereto.

The mixing ratio of the starting material triacylglyceride to the reactant fatty acid may be 1:1 to 1:7 by weight, more specifically 1:1 to 1:5, and even more specifically 1:1 To 1:3.

In the present invention, in order to increase the efficiency of the enzymatic esterification reaction, the peroxide value of the mixture containing the triacylglyceride and the fatty acid (also referred to as the'reaction mixture') increases with the course of the reaction time, that is, the reaction proceeds, In order to solve the problem of lowering the efficiency of the esterification reaction, it is characterized in that the peroxide value of the reaction mixture is adjusted to be 50 or less during the reaction time. Specifically, the peroxide value of the reaction mixture may be 47.5 or less, more specifically 45 or less, even more specifically 40 or less, and even more specifically 35 or less.

The lower limit of the peroxide value of the reaction mixture is not particularly limited, but may be, for example, 0 or more, specifically 0.01 or more.

Here, the'peroxide value' means the milli-equivalent of oxygen peroxide converted from the milli-equivalent of free iodine per 1 kg of oil and fat.

There is no particular limitation on the method of adjusting the peroxide value of the reaction mixture, and within the range to achieve the desired effect, a method commonly known in the art depending on the type and ratio of the starting/reacting materials, the type of enzyme used, etc. Can be used appropriately. In one embodiment, the peroxide value can be lowered by adding an appropriate antioxidant (e.g., tocopherol, Fortium, ascorbyl palmitate, or a mixture of two or more thereof) to the reaction mixture prior to the enzymatic reaction. It is not limited.

In one embodiment, the amount of antioxidants added based on the total 100% by weight of the reaction mixture may be 0.01 to 0.5% by weight. More specifically, the added amount of the antioxidant based on the total 100% by weight of the reaction mixture may be 0.02% by weight or more, 0.03% by weight or more, 0.04% by weight or more, or 0.05% by weight or more, and also 0.4% by weight or less, 0.3% by weight. Hereinafter, it may be 0.2% by weight or less or 0.1% by weight or less, but is not limited thereto.

In the present invention, the reaction temperature of the enzymatic esterification reaction may be determined according to the type of enzyme used, and specifically, it may be 40°C or higher, 45°C or higher, 50°C or higher, 55°C or higher, or 60°C or higher, and It may be 85°C or less, 80°C or less, 75°C or less, 70°C or less, or 65°C or less, but is not limited thereto. In order to optimize the effect of the reaction and the efficiency of the process, the reaction temperature may be preferably 40°C to 80°C, and more preferably 45°C to 70°C.

In the present invention, known enzymes may be used without particular limitation as enzymes for the enzymatic esterification reaction. Specifically, Candida Antarctica B , Thermomyces lanuginose , Rhizomucour miehei , Rhizopus delemar , Mucormiehei , alkaline agent ( Alcaligenes sp. ), Aspergillus niger , Candida cylindraces , Burkholderia cepacia , Candideum ( Geotricum candidum ), and a combination thereof. Derived esterification enzyme can be used, but is not limited thereto.

According to the present invention, it is possible to effectively prevent a decrease in enzyme activity during enzymatic esterification, so that a large amount of starting materials can be continuously treated without replacement of the enzyme. In one embodiment, 100 parts by weight to 3000 parts by weight of the reaction mixture per 1 part by weight of the enzyme, or 100 parts by weight to 2000 parts by weight, or 100 parts by weight to 1000 parts by weight may be continuously treated without replacement of the enzyme, but the activity of the enzyme is As long as it is maintained, more quantities may continue to be processed. For example, if the enzyme activity is appropriate, the reaction mixture per 1 part by weight of the enzyme can be continuously treated without replacement of 500 parts by weight or more, 1000 parts by weight or more, 2000 parts by weight or more, or 3000 parts by weight or more of the enzyme. None, and even up to 10000 parts by weight or more can be processed. According to a preferred embodiment, the treatment amount of the reaction mixture per 1 part by weight of the enzyme may be 1000 parts by weight to 3000 parts by weight.

In one embodiment, the triglyceride prepared according to the above manufacturing method may be dioleoyl monopalmitoyl glyceride, and more specifically 1,3-dioleoyl-2-palmitoyl glyceride (OPO).

Hereinafter, the present invention will be described in more detail through examples. However, this is only provided to aid understanding of the present invention, and the scope of the present invention is not limited thereto.

[ Example ]

A 1:2 weight ratio mixture (=peroxide value (POV) 54.5) of tripalmitoyl glyceride (PPP, superstearin) to oleic acid (OA) was prepared.

For each experimental set, 25 g of the mixture was added to a 100 mL Erlenmeyer flask, and the peroxide values of the mixture were adjusted to 50.0, 47.5, 27.0 and 24.2, respectively, using tocopherol.

Enzyme (Set 1: Novozym 40086, Set 2: Lipozyme TL IM) was separately weighed in an amount of 2.5 g (10% by weight of the substrate) and added to each of the reaction mixtures in which the peroxide was adjusted immediately before the reaction.

The enzymatic esterification reaction was carried out under a stirring speed of 150 rpm in a shaking incubator at which the temperature (Set 1: 55°C, Set 2: 60°C) was adjusted for each experimental set, and a certain period of time (0.5, 1, 1.5, respectively) after the reaction was started. 2, 3, and 4 hours), 1 mL of the upper layer of the reaction mixture was taken and analyzed. The analysis method is as follows.

1) neutral lipid Carbon number analysis

Neutral lipids are separated by carbon number using gas chromatography, and neutral lipids having a total carbon number of 52 (eg, C18-C16-C18, C16-C18-C18, etc.), such as SPS, SSP, OPO, and OOP. , LPL, LLP, OPL, etc.) were analyzed. The results are shown in Table 1 and FIG. 1 below.

2) Analysis of the ratio of neutral lipid structure

At 4 hours after the initiation of the reaction, HPLC (detector: ELSD, column: silver ion column) was used to analyze the ratio of the neutral lipid by structure and the concentration of USU (OPO). The results are shown in Table 2 and FIG. 2 below.

Measurement and calculation of peroxide value

About 10 to 20 g of a fat or oil sample was placed in a 250 mL Erlenmeyer flask, 10 mL of chloroform was added to completely dissolve it, and 20 mL of glacial acetic acid was added. After adding 1 mL of saturated KI solution to this, the stopper was capped, shaken for 1 minute, and left in a cool dark place for 10 minutes. Thereafter, 75 mL of distilled water was added, and 1 mL of a 1% starch solution was added thereto, followed by titration with a 0.01N Na ₂ S ₂ O ₂ solution, and the point from dark brown to colorless was set as the end point. The peroxide value was calculated by the following equation.

Peroxide value =

[(Appropriate amount for this test- Blank test Appropriate amount)(mL)× 0.01N Na ₂ S ₂ O ₂ Solution Titer ]/[Sample weight (g)]×10

From Tables 1 and 2, it can be seen that when the peroxide value of the mixture of the enzymatic esterification reaction is adjusted within the range according to the present invention, the efficiency of the enzymatic reaction can be increased, thereby improving the overall process efficiency and economy. Accordingly, according to the present invention, it was confirmed by this that dioleoyl monopalmitoyl glyceride can be efficiently and economically produced from tripalmitoyl glyceride through enzymatic esterification.

Claims (8)

As a method for producing 1,3-dioleoyl-2-palmitoyl glyceride from tripalmitoyl glyceride by an enzyme-based esterification reaction,
The enzyme is Novozym 40086, an esterification enzyme derived from Rhizomucour miehei ,
The esterification reaction is carried out by adjusting the peroxide value of the reaction mixture, which is a mixture of tripalmitoyl glyceride and oleic acid, to 35 or less, and then introducing the enzyme thereto and reacting tripalmitoyl glyceride and oleic acid.
Method for producing 1,3-dioleoyl-2-palmitoyl glyceride. The method for producing 1,3-dioleoyl-2-palmitoyl glyceride according to claim 1, wherein the treatment amount of the reaction mixture per 1 part by weight of the enzyme is 1000 parts by weight to 3000 parts by weight. delete delete delete delete delete delete

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