JPH11131089A - Purification of oil or fat - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a method for purifying an oil or fat, capable of efficiently removing phospholipids contained in the oil or fat. SOLUTION: A crude raw fatty oil or a gum-removed fatty oil is treated with an acidic phospholipase A1 to efficiently convert phospholipids contained in the fatty oil into oil-insoluble lysophospholipids, which can easily be removed from the fatty oil. The immobilization of the acidic lysophospholipid on a carrier as a means for treating the crude fatty oil or gum-removed fatty oil with the acidic lysophospholipid A1 permits the repeated employment of the lysophospholipid, and improves the production of the lysophospholipids.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to a method for purifying fats and oils. For more information, see
The fat A / F (hereinafter referred to as crude oil) or the crude oil is forcibly hydrated by the action of ZeA1 to remove various suspended substances from the oil / fat immediately after oil extraction from the oil / fat raw material. The present invention relates to a method for purifying fats and oils which efficiently decomposes and removes phospholipids in fats and oils (hereinafter referred to as degummed oils) obtained by precipitating and centrifuging.

[0002]

2. Description of the Related Art Crude crude oil derived from plants includes polar lipids mainly composed of phospholipids, glycolipids, sugars, free fatty acids, metal salts (C
a, Mg, Fe, etc.), impurities such as pigments and odorous components, etc., are removed and used in each step of degumming, deacidification, decolorization, dewaxing, deodorization, etc. It is necessary to finish refined oil of high quality with high value, and this is called refinement of fats and oils. The most noticeable behavior of fats and oils is the presence of a phospholipid-based gum. This water-soluble or oil-soluble gum is collectively referred to as phosphatides. Table 1 shows the phosphatide contents in various crude oils.

[0003]

[Table 1] (Source: Takaaki Miyagawa, Actual cooking oil production, p81 (1988)).

[0004] Since gums mainly composed of phospholipids remaining in fats and oils cause coloring, generation of off-flavors and deterioration of stability due to heating, appropriate separation and removal of phospholipids is performed by eliminating metal salts. It is also an important step that determines the success or failure of oil and fat refining. This process is called a degumming step.

In the degumming step, warm water is added to the crude oil to hydrate and precipitate phosphatides (phospholipids) and the like, and the gum is removed by a centrifuge to obtain a degummed oil. Table 2 shows changes in the components of soybean oil before and after the degumming treatment. The components that have an unfavorable effect on the quality of fats and oils have been greatly reduced by the degumming treatment.

[0006]

[Table 2] Source JC Seegers, Fette, Seifen, Anstrichmittel 87, 541 (1985).

In the step of refining fats and oils, it is essential to neutralize and remove free fatty acids with caustic soda after the above-mentioned degumming treatment. This process is called a deacidification step.

[0008] In this step, there has been a problem that viscous substances, phosphatides (phospholipids), metal salts, coloring components and the like present in the fats and oils are generated as industrial waste containing waste water and a large amount of fats and oils. . In addition, in crude crude oil or degummed oil, in addition to a hydrophilic gum that precipitates and separates by simple hydration, it binds to metal salts such as Ca, Mg, and Fe, and dissolves in oils and fats as an oil-soluble gum and removes it. There are phospholipids that are difficult to perform, and it has been an issue in oil and fat refining to remove as much oil-soluble gum as possible from crude crude oil or degummed oil.

[0009] On the other hand, there has been proposed a method for removing phospholipids in fats and oils by treatment with enzymes derived from animals or microorganisms. For example, a method of treating a crude oil with an enzyme having phospholipase A activity (JP-A-2-153997)
), A method of treating degummed oil with an enzyme having phospholipase A1, A2, and B activities (EP-A-05137).
09), an enzymatic treatment of edible oil (US Pat. No. 5,264,367).
No.), Unilever's super-degaming method, Vandemoort
Ele's total degumming method, Lurgi's Alcon method and EnzyMax Degumming method.

However, in these methods, a conventional gum hydration removal method is incorporated for the purpose of removing hydratable phospholipids, or the method is connected as a previous step, or the productivity of the enzyme used is reduced. Low or enzyme chemistry (substrate specificity, optimal pH of action, temperature, temperature stability, p
H stability, etc.) were not optimal for refining fats and oils, and the refining method did not make full use of the enzymatic chemical properties of each enzyme, so that the phospholipid content in the fats and oils was not sufficiently reduced.

The phospholipase conventionally used for the purification of fats and oils is porcine pancreatic phospholipase A2 or bee venom phospholipase A2, and their optimal pH is
Are around pH 8 and pH 8.9, respectively. According to Sigma's reagent catalog, the optimum pH of other commercially available phospholipases A2 and B is bovine pancreatic phospholipase A2: pH 8.0, Streptomyces v
It is iolaceoruber phospholipase A2: pH 8.0, Vibrio species phospholipase B: pH 8, etc., and each of the optimum pH is in a neutral to weakly alkaline region.

On the other hand, when the phospholipases A1, A2 or B act, the pH of the reaction system is lowered by the liberated fatty acids and the reaction system is deviated from the neutral or alkaline range. In order to efficiently purify fats and oils, it was necessary to always pay attention to the shift of the reaction system pH to the acidic side during the enzyme treatment, and to allow the enzyme reaction to proceed at around the optimum pH. Furthermore, an enzyme treatment method for edible oil (US526)
No. 4367), Removal of phosphatide from vegetable oil by enzyme catalysis (Fat Sci. Technol., 95. Jahrgang Nr. 8 1993)
In the above, the optimum pH for the enzymatic reaction is around pH 5.

[0013]

SUMMARY OF THE INVENTION The present inventors have solved the above-mentioned problems by efficiently decomposing and removing phospholipids in crude crude oil or degummed oil under mild conditions.
As a result of intensive studies to develop a method for purifying oils and fats of excellent quality, the use of acidic phospholipase A1 in the stage of refining crude crude oil or degummed oil allows the use of a small amount of equipment under moderate conditions. It has been found that fats and oils can be efficiently purified by adding a small amount of an enzyme at a low cost without changing the pH during the action of the enzyme.

When the immobilization method is used, the acid phospholipase A1 is immobilized on an inorganic or organic carrier, and then a lysolation reaction is carried out using an immobilized carrier which has been cross-linked and reinforced with a polyfunctional reagent. The enzyme can be used repeatedly. Further, after the lysation reaction is completed, the immobilized carrier is separated by filtration to prevent the enzyme from being mixed into the product. This has led to the completion of the present invention.

[0015]

According to the present invention, fats and oils are degummed,
When deacidifying, after adjusting the pH of the fat or oil to pH 3 to 6,
Acidic phospholiper dispersed and dissolved in water or an appropriate aqueous solution
The enzyme solution containing ZeA1 is dispersed in fats and oils containing impurities mainly composed of phospholipids to be removed, and the temperature is 30 to 80 ° C.
To act while stirring.

The fats and oils to be subjected to the refining method of the present invention are generally unrefined oils such as crude crude oil or degummed oil containing about 100 to 10,000 ppm of phospholipids, and their origin is soybean, rapeseed, Sunflower seeds, flax seeds, cottonseed, peanut seeds, corn, etc., regardless of the type.

The enzyme may be added to fats and oils at any stage of purification, but it is preferable to add the enzyme to crude crude oil or degummed oil and react it. Further, when the target fat or oil is soybean oil, rapeseed oil or the like having a high phospholipid content, it is more preferable to perform a degumming treatment by an ordinary method before the enzyme treatment.

In the enzymatic reaction, a high speed stirrer, a homomixer, a colloid mill,
The emulsification may be performed using an appropriate emulsifier such as a pipeline mixer, an ultrasonic disperser, a high-pressure homogenizer, a vibrator, or a film emulsifier.

When the immobilization method is used, specifically, an insoluble carrier is added to a solution containing acidic phospholipase A1,
Ethylene glycol to reinforce crosslinking on the carrier
By adding diglycidyl ether and immobilizing by crosslinking, a stable reaction rate is observed.

The first advantage of the method of the present invention is that, by immobilizing acidic phospholipase A1, it can be used repeatedly, and a reduction in the cost contribution rate is expected.

Secondly, at the end of the reaction, the immobilized enzyme and the reaction solution can be easily separated, the enzyme is not mixed into the product, and the quality is expected to be improved.

The enzyme is immobilized by contacting powdered acidic phospholipase A1 with a water-insoluble carrier in water to perform an adsorption treatment on the carrier, and after washing and filtering the carrier, 1 to 1
Cross-linking is strengthened in an aqueous solution containing 0% of a cross-linking agent for 2 to 5 hours, and the carrier is filtered and washed, and the cross-linking agent is continued until the absorbance (280 nm) in the washing solution becomes 0.01 or less. Can be

Any carrier can be used as long as it does not affect the activity of phospholipase A1 and is physically and chemically stable from the viewpoint of operation.

The immobilized carrier is selected from the group consisting of chitopal BCW-2610 (trade name; manufactured by Fuji Boseki Co., Ltd.) or chitopal BCW-2620, an ion exchange resin and Celite as an insoluble carrier. It is characterized by that.

There are various shapes of the carrier such as powder and granule, and any of them can be used. Especially the process,
From the viewpoint of operation, it is preferable to use a porous carrier having a particle size of 300 to 2500 μm and a specific surface area of 20 to ²⁰⁰ m ² / g.

Particularly preferred carriers include commercially available BCW-2610 and BCW-262 manufactured by Fuji Spinning Co., Ltd.
And a porous resin having a pore diameter of 0.1 to 0.2 μm such as 0.

The temperature for immobilization may be a temperature at which deactivation does not occur, and is 0 to 70 ° C., preferably 20 to 4 ° C.
5 ° C is good.

Regardless of the usual enzyme treatment or immobilization method, depending on the type of enzyme, a factor essential for the expression of the activity or a factor for enhancing the activity, for example, calcium or the like is added as necessary. Further, in order to promote the separation between the oil system and the aqueous system after the reaction, salts such as salt may be added in an amount of about 5% by weight or less of the washing water.

If pH adjustment is required, an acid (eg,
Phosphoric acid, acetic acid, citric acid, oxalic acid, maleic acid, tannic acid, tartaric acid, etc., alkali (eg, caustic soda), or buffer (eg, acetate buffer) can be used.

The enzyme treatment condition depends on the optimum temperature of the enzyme, but is usually 30 to 80 ° C., preferably 30 to 70 ° C. for about 30 minutes to 7 days. The same applies when the method is used. The acidic phospholipase A1 used in the present invention desirably has low lipase activity regardless of the usual enzyme treatment or immobilization method. The origin of the enzyme may be derived from animals, plants, or microorganisms, and the purity of these enzymes is not limited.

Regardless of the usual enzymatic treatment or immobilization method, the optimum pH for the action of the acidic phospholipase A1 used in the present invention is determined by a conventional activity measurement system (Triton X-1).
(Reaction at 37 ° C. for 10 minutes in the presence of 00), the pH is 3 to 6.5, and most preferably 3.2 to 5.0.

For example, in the method for purifying fats and oils, Aspergillus
Acid phospholipase A1 derived from oryzae or Aspergillus niger is industrially available, and details of these enzymes are described in phospholipase A1 and its use (Japanese Patent Application Laid-Open No. Hei 7-31472). .

When acid phospholipase A1 acts on phospholipids, 2-acyl lysophospholipids are formed, and 1-acyl lysophospholipid, which is a product of phospholipase A2, is defined as a fatty acid binding site, constituent fatty acid composition and The composition of the lysed phospholipids is different, and the difference in the physical properties of the enzymatic reaction products based on these differences has a positive effect on the removal of phospholipids and metal salts from fats and oils.

In particular, Aspergillus oryzae or Aspergill
The acid phospholipase A1 from us niger efficiently releases the fatty acid at position 1 of phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol, and phosphatidic acid, and has a substrate specificity and an optimal action. The pH / temperature, temperature stability, pH stability, productivity of the enzyme preparation and the like are also suitable for purification of fats and oils as compared with phospholipase A1 derived from Rhizopus arrhizus described in US Pat. No. 5,264,367.

Specifically, when a crude crude oil or a degummed oil is treated with an enzyme, the activity of the acidic phospholipase A1 used is not particularly limited.

For example, in the case of treatment with a small amount of added water, the amount may be about 1.5 to 30,000 units per liter or kg of crude crude oil or degummed oil, and in the case of ordinary enzyme treatment, In the case of the method, crude crude oil or degummed oil 1
An immobilized product of about 3,000 to 300,000 units per 1 or 1 kg can be used repeatedly.

When the enzyme treatment is carried out in an emulsified state using a highly active acidic phospholipase A1 and increasing the amount of added water to increase the contact efficiency between the oil phase and the aqueous phase, it takes 30 minutes to 5 minutes.
The processing can be completed in about time.

When the reaction is carried out using a column by the column method, the treatment liquid containing added water is circulated to the fats and oils, crude crude oil or degummed oil by the reverse flow method (up-flow method), whereby the column is prepared. The method of repeatedly sending the solution into the inside is preferable.
The flow rate is adjusted to 3 times or less of the total processing solution per minute.

In order to carry out a lysination reaction containing 10% and 20% of lecithin by the immobilization method, it is necessary to repeatedly use approximately 3,000 to 4.5 million units immobilized per liter or 1 kg of lecithin. it can.

In the method of the present invention, the amount of water used in the ordinary enzymatic treatment and immobilization method is approximately 0.05 to 20 parts by weight with respect to 100 parts by weight of crude oil when the amount of treated water is suppressed. More optimally, 10 parts by weight may be used for 100 parts by weight of crude oil. When a high-activity acidic phospholipase A1 is used and the amount of treated water is increased to carry out a rapid normal enzymatic treatment and immobilization method in an emulsified state, crude crude oil 1
About 50 to 2,000 parts by weight,
Or more treated water can be used.

For example, when fats and oils having a high phospholipid content, such as soybean oil, are degummed using acidic phospholipase A1, in order to remove gums, these fats and oils are firstly pre-washed with water. Degumming is a good idea. The phosphorus content in the above fats and oils is adjusted to 50 to 500 p
It is preferable to reduce it to the range of pm. In this case, the degumming process itself may be based on a conventional technique.

According to the method of the present invention, phospholipids in crude crude oil or degummed oil can be efficiently decomposed and removed.

In the method of the present invention, chemical refining can be replaced with physical refining, thereby making it possible to purify fats and oils by omitting the steps of deoxidation and waste disposal.

That is, the phospholipids in the crude crude oil or the degummed oil are converted into lysophospholipids by the action of acidic phospholipase A1, and the hydrophilicity is increased, so that the phospholipids are easily transferred to the aqueous phase. In order to change to a state easily adsorbed by the adsorbent, after enzymatic treatment or, if necessary, after dehydration treatment by a centrifugal separator or the like, decolorization and deodorization steps are carried out by a conventional method to improve the quality of fats and oils. Phospholipid content can be reduced to trace amounts without adverse effects.

More specifically, acid phospholipase A
The reaction product 2-acyl lysophospholipid when using No. 1 is
1-acyl lysophospholipid which is a reaction product of phospholipase A2 is different from the 1-acyl lysophospholipid in terms of fatty acid binding site, constituent fatty acid composition and lysophospholipid composition, so that phospholipid is more efficiently removed from fats and oils than expected. In addition to this, the content of metal salts such as Ca, Mg, and Fe in fats and oils can be reduced.

10% and 20% using immobilized enzyme
When lecithin undergoes a lysination reaction, it is converted into lysolecithin by the action of the immobilized acidic phospholipase A1. The lysation reaction proceeds quickly when 0.1 to 0.5 part by weight of the surfactant Triton X-100 is added to 1 part by weight of the raw material lecithin. Desirably, 0.2 to 0.3 parts by weight is good. The remaining parts by weight are purified water.

After the treatment with the immobilized enzyme, the immobilized carrier is separated by filtration through a 32 mesh wire mesh and used for the next reaction. Lysolecithin can be obtained from the filtrate through a conventional method.

[0048]

EXAMPLES The present invention will be described with reference to Examples and Comparative Examples.

The following acidic phospholipase A1 activity or phospholipase A2 activity were measured at pH 4 and pH 4, respectively.
The activity at 8 was shown. The analysis of the phospholipid was carried out in accordance with Japanese standard fat and oil analysis method 2.22.8.1-71.

Example 1 Phospholipid content: 3,000 pp
100 g of crude oil derived from soybeansAspergillus oryzae
(FERM BP-3887) derived acidic phospholippers
ZeA1 solution [0.2 units / ml (pH5), 5 mM chloride
Calcium, 10 mM citric acid and 0.073 g NaO
H)], and with stirring with a stirrer,
The reaction was performed at 60 ° C. for 24 hours. After the reaction, centrifuge
(1,500 G, 10 minutes) to remove the enzyme solution.
A degummed oil having a lipid content of 100 ppm was obtained.

[Comparative Example 1] Phospholipase A2 was added to the enzyme solution.
Solution [manufactured by Novo, trade name recitase, 0.2 units / ml
(PH 6) containing 5 mM calcium chloride, 10 mM citric acid and 0.087 g NaOH]. 300 ppm in degummed oil
Of phospholipids remained. Example 2 Aspergillus niger (ATCC 964) was added to 100 g of soybean-derived degummed oil having a phospholipid content of 130 ppm.
2) 0.5 ml of an enzyme solution (containing pH 5, 10 mM citric acid and 0.00023 g NaOH) having an activity of 50 units of acidic phospholipase A1 was added, and the mixture was reacted at 60 ° C. for 24 hours while stirring with a stirrer. To this oil was added 1.0% of activated clay [manufactured by Mizusawa Chemical Industry Co., Ltd., trade name NV], and the oil was subjected to a decolorizing treatment at 105 ° C., 30 Torr and 20 minutes. The phospholipid content in the fat was reduced to 25 ppm.

[Comparative Example 2] Phospholipase A2 was added to the enzyme solution.
Enzyme solution having an activity of 50 units [manufactured by Novo, trade name: Lecitase, pH 6, 10 mM citric acid and 0.00028 g
NaOH] was used in the same manner as in Example 2. 45 ppm of phospholipid remained in the fat and oil.

The results of Example 1 and Comparative Example 1, and Examples 2 and Comparative Example 2 are summarized. The optimal pH of the action of phospholipase A2 used in Comparative Examples 1 and 2 is 8 to 9, but in this reaction system, a strong emulsion is formed when the pH exceeds 8, and the reaction system has a slightly acidic pH of 5.5. It is practical to perform the steps from 6.5 to 6.5.

The pH of water in contact with ordinary unrefined oil is 5.
The pH is 5 to 6.5, but if the pH is not adjusted, the pH of the reaction system shifts to the acidic side as the free fatty acids increase due to the enzymatic reaction.

Further, the reaction product of acid phospholipase A1 differs from the reaction product of phospholipase A2 in the composition of fatty acid binding sites, constituent fatty acids, and lysophospholipids. Can be removed.

The residual amount of phospholipids in the fats and oils in Examples 1 and 2 was smaller than that in Comparative Examples 1 and 2, respectively. In addition, problems such as coloring due to heating and generation of an unusual odor are reduced.

Example 3 3 g of a commercially available chitosan resin (Chitopal BCW-2620; manufactured by Fuji Boseki Co., Ltd.)
Filtration and washing were performed three times with 0 ml of ion-exchanged water to remove adhering water. Acid phospholipase A1 derived from Aspergillus oryzae (FERM BP-3887) (19,000 units / g)
Add 150 mg to 50 ml of ion-exchanged water, and add
Stirred for 0 minutes. Add all the chitosan resin to this solution,
The mixture was kept at 30 ° C. for 3 hours while stirring at 150 rpm using a rotary shaker.

Next, the chitosan resin was filtered off from the suspension,
Washed with water. Next, the entire amount of the acidic phospholipase A1 adsorption resin was added to 50 ml of a 2.8% aqueous solution of ethylene glycol diglycidyl ether, and the mixture was kept at 30 ° C. for 2.5 hours while stirring at 150 rpm using a rotary shaker. . Thereafter, the operation of washing the resin with water and filtering off the resin was repeated until the absorbance (280 nm) of the filtrate became 0.01 or less, to obtain an acidic phospholipase A1-immobilized chitosan resin.

In the case of the lysination reaction of lecithin, 2 g of lecithin (10%), 0.6 g of Triton X-100,
After mixing 17.4 ml of water at 50 ° C. for 15 minutes, the pH was adjusted to 3.8 to 5 with acetic acid. After stirring at 50 ° C. for 20 minutes, the mixture is cooled to 40 ° C. and 3 g of acidic phospholipase A1 immobilized resin is added. The lysation reaction was carried out at 200 to 400 rpm and 40 ° C. using a rotary shaker.
For 1 to 7 days. After the completion of the reaction, the acidic phospholipase A1-immobilized resin was separated by filtration through a 32 mesh (Tyler-standard sieve) wire mesh, collected, and lecithin and a pH-adjusted solution were newly added to carry out the reaction under the above conditions. . The above operation was repeated 25 times.

In the test of the reaction solution, oil was extracted from the sample by the Bligh-Dyer method, and the acetone-insoluble matter of the obtained extracted oil was subjected to thin-layer chromatography to determine the phosphorus composition ratio of each phospholipid by a colorimetric method. I asked for it. Next, the phospholipid in the sample was quantified by a colorimetric method, and the amount of each phospholipid was calculated by calculation.
In particular, the lysification rate was determined by quantification of phosphatidylcholine and lysophosphatidylcholine, typically. As a result, the 10th lysolation rate was 97.52%, the 15th lysolation rate was 100%, and the 25th lysolation rate was 94.
31% / 48h.

Further, the use of acidic phospholipase A1 having an optimum pH of 3 to 6 is not limited to the pH in this enzyme treatment step.
This means that no adjustment is required, and workability is greatly improved. Example 4 3 g of acidic phospholipase A1 immobilized resin (BCW-2620) prepared in the same manner as in Example 3
To crude oil 10 g, caustic soda 4.3 mg, citric acid 1
0 mg and 1 ml of water were added to the reaction solution adjusted to pH 3.2 to 6.5, and the mixture was reacted at 50 ° C. for 2 to 6 days while stirring at 200 rpm using a rotary shaker.

[0062] The phospholipid (phosphatidylcholine) in the crude oil was 360 ppm. After completion of the reaction, the acidic phospholipase A1 immobilized resin was recovered in the same manner as in Example 3 and repeatedly used. The above operation was repeated 21 times.

The phospholipids in the reaction solution were quantified in the same manner as in Example 3, and the lysation rate was calculated. As a result, the 15th lysolation rate was 100% and the 20th lysolysis rate was 100%.

As a result, the phospholipid content in the fat and oil was reduced to zero (not detected).

[Test for phospholipase A1 activity] The activity of acidic phospholipase A1 used in the examples was measured by the following method. 2.0 (w / v)% soy lecithin (phospholipid) (manufactured by True Lecithin Industries, trade name: SLP-White) and 4 (v / v)% Triton X
-100 aqueous solution 0.5 ml, 0.1 M calcium chloride 0.05 m, 10.2 M acetate buffer (pH 4.0)
0.25 ml was added. Next, 0.1 ml of the enzyme solution was added to make the mixture uniform, and the mixture was allowed to stand at 37 ° C. to carry out an enzyme reaction for 10 minutes. Thereafter, 0.1 ml of 1N hydrochloric acid was added to stop the enzyme reaction. Take 0.02 ml of the reaction solution and use a free fatty acid determination reagent (trade name: Determinator NE, manufactured by Kyowa Medix Co., Ltd.)
FA) was used to quantify free fatty acids. Enzyme activity that produces 1 μmol of fatty acid per minute of enzymatic reaction was defined as 1 unit.

[0066]

According to the present invention, the phospholipids in crude crude oil or degummed oil are efficiently converted to oil-insoluble lysophospholipids by the action of acidic phospholipase A1, and the removal of phospholipids from fats and oils is facilitated. It is an excellent purification method.

Claims (6)

[Claims] 1. A method for purifying fats and oils, wherein the fats and oils are treated with acidic phospholipase A1. 2. A method for purifying an oil or fat, comprising treating a crude oil with an acidic phospholipase A1. 3. A method for purifying fats and oils, wherein the degummed oil is treated with acidic phospholipase A1. 4. A method for purifying an oil or fat, comprising fixing an acid phospholipase A1 to a carrier and treating the oil or fat, crude crude oil or degummed oil. 5. The method according to claim 1, wherein the acid phospholipase A1 is phospholipase A1 produced by Aspergillus oryzae . 6. The method according to claim 1, wherein the acid phospholipase A1 is phospholipase A1 produced by Aspergillus niger .