JPH066032B2 - Method for producing enzyme-modified lecithin - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION In the present invention, soybean lecithin (1,2-diacylglycerophospholipid) is partially hydrolyzed with PLA _{2 in the} presence of a fat-soluble solvent to be modified to lysolecithin (1-monoacylglycerophospholipid). The total phospholipids contained in the total phospholipids in an amount of 95% by weight or more, and the total phospholipids contained in the total phospholipids in an amount of 65% by weight or more. The present invention relates to a method for producing an enzyme-modified lecithin containing:

(Industrial field of application) Soybean lecithin is not only used as a natural emulsifier and surfactant for various foods, cosmetics, paints, and other industrial applications, but also has a physiologically active effect. Although it is also used in the field, its application is limited in terms of emulsification characteristics and functionality because it is easily influenced by temperature, pH, salt concentration and the like.

INDUSTRIAL APPLICABILITY The present invention improves the emulsifying property and functionality of lecithin by enzymatic modification, and provides a novel use of lecithin.

(Prior art) Conventional lecithin is used in the form of pasty lecithin containing neutral lipids such as triglycerides derived from raw materials, high-purity lecithin obtained by degreasing neutral lipids with acetone, and specific phospholipid components. Fractionated lecithin and the like (Yamano, Tsuru et al .: Nissho Kogaku, 29, 137, (1982)) are known, but in order to impart more excellent functionality to lecithin, lecithin is chemically or enzymatically added. Reforming technology was devised and reformed by hydrogenation (Japanese Patent Application No. 59-167).
10), a method of preparing a complex of lecithin and starch to enhance emulsifying power (Japanese Patent Application No. 59-69188), and a method of obtaining lysolecithin from PLA ₂ derived from snake venom (Anael).
1, G.I. B. & Hawthome, J. et al. N. , Phos
pholipids (1964)), a method of partially hydrolyzing pancreatin to produce lysolecithin (Japanese Patent Application No. 53-71517), and the like.

(Problems to be Solved by the Invention) The main phospholipid components in soybean lecithin are phosphatidylcholine (hereinafter referred to as PC), phosphatidylethanolamine (hereinafter referred to as PE), phosphatidylinositol (hereinafter referred to as PI), and phosphatidylserine. Although it is (hereinafter referred to as PS), since PI is an anionic substance, there is almost no effect on the surfactant activity due to pH change. However, PC, PE, and PS have an isoelectric point because they are zwitterionic substances, and the vicinity of that isoelectric point, that is, pH
PC, PE and PS under acidic conditions of 1.5 to 3.5
Has a reduced affinity for water and accordingly a reduced surface activity. In addition, when the salt concentration increases, the hydration property decreases, and similarly the surface activity decreases.

In order to catch the above-mentioned drawbacks, it is necessary to modify lecithin itself to make it highly hydrophilic. As a method,
A method of partially hydrolyzing one of the fatty acids, which is the hydrophobic group of the component phospholipid, or a method of introducing a new hydrophilic group into the phospholipid can be considered, but the modification by the chemical synthesis method is a by-product. It is not preferable from the viewpoint of prevention of mixture, and it cannot be said that the formation of a complex with a water-soluble polymer such as starch or protein is a fundamental modification, and the use is further limited.
Compared with the above-mentioned modification method, the enzyme modification method is carried out under mild conditions, so that it has an advantage of suppressing the deterioration of the product, but when PLA ₂ used is derived from a snake venom, the enzyme is expensive. However, it has a drawback that it is difficult to handle due to its strong skin erosion property, and crude enzymes such as pancreatin do not exert their original potency due to the effect of lipase and protease which are contaminated, and the hydrolysis rate is not improved. Therefore, the lysolecithin content of the conventional enzyme-modified lecithin was only about 15 to 35% by weight in the total phospholipid, and it did not exhibit a remarkable effect by competing with the coexisting unmodified lecithin component. Furthermore, in the conventional enzyme-modified lecithin, free fatty acid is not treated due to deacylation, which is a factor of deteriorating the surface activity and stability of lysolecithin.

(Means for solving the problem) As a modification method utilizing the original characteristics of soybean lecithin,
It is best to convert to lysolecithin (1-monoacyl glycerophospholipid) under mild conditions by an enzymatic reaction in the presence of a fat-soluble solvent. In this case, the lipophilic solvent improves the viscosity reduction and contact with the enzyme, and consequently the lysolecithin content in the total phospholipids. The fat-soluble solvent may be appropriately selected or mixed such as hexane and ethyl acetate. As the enzyme used in this reaction, PLA ₂ derived from pancreatin is the most suitable, and it is preferable that the mixture of revertase, protease and the like is 0.1% or less per total activity unit.

The enzyme-modified lecithin of the present invention contains naturally-derived 1-monoacylglycerophospholipids, that is, lysophosphatidylcholine, lysophosphatidylethanolamine, lysophosphatidylinositol, and lysophosphatidylserine as main components, and contains 95% or more of total phospholipids, and It is characterized in that the total phospholipid content is 65% by weight or more.

The surface-active abilities of 1-monoacyl glycerophospholipids and diacyl glycerophospholipids are greatly different, and the property of the emulsion formed is determined by the abundance ratio of the two. It is necessary that 65 wt% or more of the lysolecithin component is present in the total phospholipid.

Also, lysophosphatidylcholine, lysophosphatidylethanolamine, lysophosphatidylinositol,
The abundance ratio of lysophosphatidylserine depends on the composition of the natural product as a raw material, but other phospholipids, for example, complex phospholipids such as sphingomyelin, cardiolipin, and plasmalogen should be removed from the viewpoint of improving the surface activity. Is desirable.

Fatty acids liberated during the enzymatic reaction adversely affect the stability of lecithin and enzyme-modified lecithin and impair the taste, odor and color tone of the product during application, and therefore need to be removed. In order not to impair the taste, odor and color of the product, the total phospholipid content (dry weight ratio) must be 95% or more. Conventionally, acetone was mainly used for degreasing and refining of lecithin, but in the case of acetone treatment, an acetone adduct of lecithin is produced to generate a unique bitterness and odor, and the dye component derived from the raw material is liable to remain. there were. The organic solvent used for degreasing purification in the present invention, methanol, ethanol, propanol, because it is selected from the low molecular weight aliphatic alcohol group of isopropanol, there is no bitterness and offensive odor production, the purity of the phospholipid to be purified is very high. Get higher The above organic solvent may be appropriately selected according to the application field.

(Function) The enzyme-modified lecithin of the present invention comprises lysolecithin (1-monoacylglycerophospholipid), that is, lysophosphatidylcholine, lysophosphatidylethanolamine, lysophosphatidylinositol, and a mixture of one or more lysophosphatidylserine as main components. For this reason, the electrochemical properties of polar groups are the same as those of conventional lecithin, but the hydrophilicity is improved by deacylation, so the hydration power is increased, and the emulsion stability at high temperature, acid and high salt concentration is increased. Is significantly improved compared to conventional lecithin.

Further, the conventional emulsion system using lecithin is stable in the oil-in-water type (O / W type), but it is stable in the water-in-oil type (W / O type).
Type) is known to be difficult to stabilize. It is considered that the two fatty acid groups of the diacylglycerophospholipid in the conventional lecithin repel each other due to the repulsive force between them in the W / O type system to make the micelle structure sparse. In the case of the enzyme-modified lecithin, not only the O / W type emulsifying property is improved because a regular micelle structure can be formed like the known monoglyceride, but the W / O type emulsifying property is also improved. Moreover, the enzyme-modified lecithin significantly reduces the iodine color-reactivity of starch. This indicates that the ability to form a complex with starch is high, and when it is used for improving the functionality of starch, it exhibits remarkable effects on water retention, gel strength and the like.

Removal of free fatty acids is important in order to fully exert the functionality of lysolecithin as described above.

The present invention will be specifically described in the following examples and comparative examples, but the present invention is not limited thereto.

Example. 1 Pasty lecithin (lecithin AY: phospholipid content 65
%, Manufactured by Hosei Oil Co., Ltd.) 20 kg of n-hexane 10
Was added and dissolved with stirring, and then 1.0 × 10 ⁶ U of PLA ₂ was added.
0.1M tris (hydroxymethyl) aminomethane-hydrochloric acid buffer containing 0.05 mol of calcium chloride (pH
7.3) PLA ₂ solution prepared by dispersing and dissolving in 5.0 was added, and reacted at 40 ° C. for 48 hours under continuous stirring.

After the reaction, n-hexane and water were distilled off under reduced pressure to obtain a paste-like reaction product, which was dispersed and dissolved in nm-hexane 10 and suction-filtered through a mixture of Celite and fine cellulose powder. Evaporated under reduced pressure,
The obtained paste-like product was degreased with 8 times or more volume of ethanol and then dried under reduced pressure to obtain 6.50 kg of enzyme-modified lecithin.

The enzyme-modified lecithin had a benzene-insoluble content of 0.01% and an acetone-insoluble content of 98.80%. As a result of high performance liquid chromatography and thin layer chromatography analysis, the lysolecithin content was 85 mol% (about 70% by weight)
Met.

Test example. 1 Example. In the same manner as in 1, paste-like lecithin (lecithin AY: phospholipid content 65%, manufactured by Hosei Oil Co., Ltd.) 20
n-Hexane 10 was added to kg and dissolved by stirring, and then P
PLA ₂ prepared by dispersing and dissolving 1.0 × 10 ⁶ U of LA ₂ in 5.0 of 0.1 M tris (hydroxymethyl) aminomethane-hydrochloric acid buffer solution (pH 7.3) containing 0.05 mol of calcium chloride. Add the solution and under continuous stirring at 40 ℃
After reacting for 15 hours, the mixture was subjected to degreasing and purification to prepare enzyme-modified lecithin having a lysolecithin content of 35% by weight (benzel insoluble content 0.01%, acetone insoluble content 99.05%).
Example. The emulsifying properties of 1 and the enzyme-modified lecithin were compared.

Both are decomposed and dissolved in water to prepare a 0.5 wt% aqueous solution,
To 50 parts by weight of this aqueous solution, 50 parts by weight of rapeseed white squeezing oil was added, emulsified with a homomixer, transferred to a calibrated test tube and allowed to stand at 80 ° C. for 8 hours, and then the ratio of the emulsified layer to all solution layers. Was measured to examine the emulsion stability. The emulsion stability of the enzyme-modified lecithin No. 1 was 80%, but it was 27% with the enzyme-modified lecithin having a lysolecithin content of 35% by weight. As a control, defatted lecithin measured by the same method as above (manufactured by Central Soya)
The emulsion stability of was 10%.

Test example. 2 Example. The heat resistance test of the emulsion composition prepared with the enzyme substance lecithin of 1 was conducted.

The enzyme-modified lecithin is dispersed and dissolved in water to prepare a 0.5 wt% aqueous solution. To 50 parts by weight of this lecithin aqueous solution, 50 parts by weight of rapeseed white squeezing oil was added, emulsified with a homomixer, and transferred to a graduated test tube at 5, 25, 40 and 80 ° C.
After standing still for 8 hours, the ratio of the emulsion layer to the total solution layer was measured to examine the emulsion stability. In addition, heat-resistant emulsion compositions were prepared in the same manner as above, using defatted lecithin (manufactured by Central Soya Co., Ltd.) and crude enzyme-modified lecithin (undefatted product, lysolecithin content of about 70% by weight / total phospholipid, commercial product) as controls. A sex test was conducted. The results obtained are shown in Fig. As shown in 1, the emulsified composition with the enzyme-modified lecithin exhibits remarkable heat resistance, but the crude enzyme-modified lecithin containing free fatty acid exhibits the original emulsion stability even if the phospholipid content is the same. Was not done.

Test example. 3 Example. The acid resistance test of the emulsion composition prepared with the enzyme-modified lecithin of No. 1 was conducted.

The enzyme-modified lecithin was dispersed and dissolved in a citrate buffer solution of pH 3, 5, and 7 to prepare 0.5% by weight solutions, and 50 parts by weight of rapeseed white squeezing oil was added to 50 parts by weight of each solution. Emulsify with a homomixer, transfer to a graduated test tube, and 80 ℃
After leaving still for 4 hours, test example. The emulsion stability was measured by the same method as in 2. In addition, defatted lecithin (manufactured by Central Soya) and crude enzyme-modified lecithin (undefatted product, lysolecithin content of about 70% by weight / total phospholipid, commercial product)
As a control, the emulsion composition was subjected to the acid resistance test in the same manner as above.

The acid resistance of each emulsion composition is shown in Fig. As shown in FIG. 2, it was found that the enzyme-modified lecithin exhibited remarkable acid resistance, but the crude enzyme-modified lecithin did not exhibit a theoretical titer.

Test example. 4 Example. The salt resistance test of the emulsion composition prepared with the enzyme-modified lecithin of No. 1 was conducted.

Enzyme modified lecithin 0.5% with 1% sodium chloride solution
% Dissolved in each of the calcium chloride aqueous solution and dissolved in water.
Prepare a 5 wt% solution. 50 parts by weight of rapeseed white squeezing oil was added to 50 parts by weight of each aqueous solution, emulsified with a homomixer, transferred to a test tube with a scale, and allowed to stand at 5, 25, 40 and 80 ° C. for 4 hours, and then tested. Example. The salt resistance of the emulsified composition was examined in the same manner as in 2. In addition, the acid-resistant emulsion composition was treated in the same manner as described above, using defatted lecithin (manufactured by Central Soya Co., Ltd.) and crude enzyme-modified lecithin (undefatted product, lysolecithin content of about 70% by weight / total phospholipid, commercially available product) as the control. A sex test was conducted.

The obtained results are as shown in FIG. 3, and the defatted lecithin markedly decreases the emulsion stability in the presence of calcium ions, but the emulsion composition containing the enzyme-modified lecithin shows remarkable salt tolerance even under the same conditions. However, the effect of free fatty acids was not so significant under these conditions.

Test example. 5 Example. A water retention test was performed on starch of the enzyme-modified lecithin of No. 1.

Enzyme modified lecithin 0.25, 0.5, 1.0 respectively
A starch paste solution is prepared by dispersing and dissolving it in a 5% flour starch suspension so that the weight% of the product is obtained, and then gelatinizing the mixture in a hot water bath at 80 ° C. This starch paste solution was transferred to a graduated test tube and allowed to stand at 25 ° C, and the water retention property was examined by measuring the length of the water separation layer from the starch paste solution layer. In addition, defatted lecithin (manufactured by Central Soya), crude enzyme-modified lecithin (undefatted product, lysolecithin content of about 70% by weight / total phospholipid, commercial product), sucrose fatty acid ester (HLB: 1)
Using Mitsubishi Kasei Co., Ltd. as a control, a water retention test with flour starch was performed in the same manner as above.

The results obtained are shown in Fig. As shown in 4, the water retention by the enzyme-modified lecithin was remarkable, and the crude enzyme-modified lecithin containing free fatty acids did not exhibit the original water retention even if the phospholipid content was made equal. Further, the water retention of the sucrose fatty acid ester having almost the same HLB was much lower than that of the enzyme-modified lecithin.

(Effects of the Invention) The present invention dramatically expands the range of use by improving the functionality of lecithin, which is a natural surfactant, and is effective in the chemical industry, diet or medical field, and safe. It provides a surfactant.

[Brief description of drawings]

Fig. 1 is enzyme-modified lecithin, crude enzyme-modified lecithin,
The heat resistance of an emulsion composition prepared from defatted lecithin and sucrose fatty acid ester is expressed using the emulsion stability as an index. Fig. 2 is enzyme-modified lecithin, crude enzyme-modified lecithin,
The acid resistance of an emulsified composition prepared with defatted lecithin and sucrose fatty acid ester is expressed using the emulsion stability as an index. Fig. 3 is enzyme-modified lecithin, crude enzyme-modified lecithin,
The salt resistance of an emulsified composition prepared from defatted lecithin and sucrose fatty acid ester is represented by the emulsion stability as an index. Fig. 4 is enzyme-modified lecithin, crude enzyme-modified lecithin,
It shows the water retention of starch paste to which defatted lecithin and sucrose fatty acid ester have been added.

Claims (3)

[Claims] 1. Soybean lecithin (1,2-diacylglycerophospholipid) is partially hydrolyzed by phosphalipase A ₂ (hereinafter abbreviated as PLA ₂ ) in the presence of a fat-soluble solvent to give lysolecithin (1-monoacylglycerophospholipid). The total phospholipid contains 95% by weight or more of total phospholipids that have been purified by separating and removing free fatty acids and other fat-soluble components derived from raw materials that have been modified during the reaction with an organic solvent. A method for producing an enzyme-modified lecithin, which comprises 65% by weight or more. 2. The production of the enzyme-modified lecithin according to claim 1, wherein the organic solvent used for purification is selected from the group of low molecular weight aliphatic alcohols of methanol, ethanol, propanol and isopropanol. Law. 3. The enzyme modification according to claim 1, wherein the lysolecithin component is one or a mixture of two or more of lysophosphatidylcholine, lysophosphatidylethanolamine, lysophosphatidylinositol and lysophosphatidylserine. Of producing high quality lecithin.