JPH03187385A - Production of monoglyceride - Google Patents

Abstract

PURPOSE:To increase the monoglyceride content in decomposed oil and fat and to obtain a monoglyceride on an industrial scale at a low cost by adding water and alkaline lipase to fist oil and decomposing the oil and fat in the presence of a bicarbonate, etc. CONSTITUTION:A decomposition product rich in the objective monoglyceride can be produced by adding 0.5-200wt.% of water (based on oil and fat), 5-1,000 unit of an alkaline lipase (based on 1g of oil and fat), 0.1-30mol of a bicarbonate, a monohydrogen phosphate or an alkaline-earth metal salt (based on 1mol of oil and fat) and 5wt.% of glycerol to an oil and fat such as soybean oil or beef tallow and stirring the mixture while keeping the pH to 8.0-9.0. The monoglyceride can be separated from the product by neutralizing the decomposition product with hydrochloric acid, etc., precipitating with n-hexane, etc., and recovering the precipitate.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a process for producing monoglycerides, and in particular to a process for obtaining monoglyceride-rich degradation products from fish oil.

That is, the present invention adds water and alkaline lipase to fish oil or fish oil and glycerin, and performs fat decomposition in the presence of bicarbonate, lhydrogen phosphate, or alkaline earth metal salt to obtain a decomposition product rich in monoglycerides. This is a method for producing monoglyceride characterized by the following.

The manufacturing method of monoglyceride has been researched for a long time, but currently it is mainly done by adding around 0.1% metal catalyst to a mixture of fats and oils and glycerin and heating it at 200 to 230''C while stirring.
The so-called glycerolysis method in which a monoglyceride is synthesized by a reaction [Tsuda 1. ``Production and Application of Monoglyceride'', Maki Shoten, p. 141 (1964)] is used.

However, zinc, tin, etc. added as reaction catalysts remain in the mixed product of treey and monoglyceride obtained by the above synthesis method.

Therefore, in order to use the monoglyceride obtained in this way in foods, cosmetics, and medicines, it is necessary to remove the metal catalyst. However, even when the metal catalyst is removed, although it is possible to reduce these harmful metals, it is difficult to completely remove them.

Therefore, recently, a method of directly producing monoglycerides from natural oils and fats using enzymes without using harmful heavy metals as catalysts has been attracting attention. It is already widely known that when lipase is applied to fats and oils, a certain amount of monoglyceride accumulates in the process of decomposing the fats and oils [Shigeru Tsuda: "Monoglycerides"
"Manufacturing and Application" Maki Shoten, p. 113-117 (196
4) ]. However, a method for industrially obtaining large amounts of monoglyceride by making Reliever act on oils and fats has not yet been successful. The reason for this is that even though lipase acts on fats and oils to produce monoglycerides, further decomposition proceeds in parallel, so the monoglycerides are not sufficiently accumulated and most of them are converted into fatty acids and glycerin. When using a lipase that has a low accumulation amount from 1 mol of oil or fat, the amount is about 0.055 mol, and even when using pancreatic lipase, which is known to have a large accumulation amount, it is 0.6 mol. This is because monoglycerides accumulate only in molar amounts.

In order to compensate for the drawback of lipase in monoglyceride accumulation, the present inventors investigated a method for promptly accumulating a large amount of monoglyceride produced during the decomposition of fats and oils. As a result, alkaline lipase was allowed to act on fats and oils in the presence of various alkaline agents such as caustic alkali salts such as sodium and potassium, carbonates, phosphates, and silicates, as well as hydroxides of calcium and magnesium. They discovered that the amount and speed of monoglyceride accumulation differed depending on the type of alkaline agent.

This will be explained using an experimental example.

50g of olive oil and 15.25d of water 500111
Place it in an Erlenmeyer flask with a baffle, and add Na as an alkali agent.
HCOz is 4.75 g, NaJPO-42HzO
is 10.12 g, Ca (OH) z is 6.6 g
, 4.9 g of Mg(OH)z was added, and other alkaline agents were added gradually so that the pH of the reaction system was always 8.0 to 9.0. -266 bacteria (Feikoken Bacteria No. 3187) or the name tJ! PL-679 bacterium (Feikoken Bacteria No. 3783)
Alkaline lipase produced by No. 1) was added and the reaction was carried out at 30°C for 4 hours with shaking and stirring.

After 4 hours, the reaction was stopped by acidifying with hydrochloric acid, and diethyl ether was added to extract the oil. After washing with water and dehydration, the ether was removed under reduced pressure to recover the oil.

The monoglyceride content and fat decomposition rate in this oil were analyzed by the method described below. The results are shown in Table 1.

(Margins below this page) As can be seen from the above experimental examples, when alkaline lipase is applied to fats and oils in the presence of bicarbonate, monohydrogen phosphate, or alkaline earth metal salts, the reaction is most rapid. It has been found that the most suitable method for increasing the amount of monoglyceride accumulated is to allow alkaline lipase to act on fats and oils in the presence of these alkaline salts. The present invention has been completed based on the above discovery.

As the alkaline lipase used in the present invention, any alkaline lipase can be used, for example, the strain No. 1iPL-266 (Feikoken Bibori No. 3187) described in Japanese Patent Publication No. 58-36953. or,
Name 1iPL-6 described in Japanese Patent Application Laid-Open No. 53-59093
This can be carried out using alkaline lipase produced by Bacterium No. 79 (Feikoken Bacteria No. 3783).

In addition, as the form of alkaline lipase, a culture solution of alkaline lipase-producing bacteria or a processed product thereof, such as an acetone-precipitated enzyme powder, a purified enzyme, and an enzyme obtained by supporting and immobilizing these on a suitable immobilization carrier are used. I can do it.

Next, as the oil or fat, vegetable oil, animal oil, fish oil, etc. can be used, and specific examples thereof include soybean oil, palm oil, coconut oil, beef tallow, lard, whale oil, etc.

The amount of alkaline lipase used may be increased or decreased as appropriate depending on the strength of the fat-decomposing power of the enzyme preparation, but it may be 5 to 1000 units, preferably 5 to 100 units, per 1 g of fat or oil. The unit of alkaline lipase is measured according to the Journal Mondial de Pharmacier.
1 Mondil de Pharmacie) No. 3, pp. 349-352, 1968.

Examples of the hydrogen carbonate or lhydrogen phosphate used in the present invention include sodium salt, potassium salt, and ammonium salt, and examples of the alkaline earth metal salt include calcium salt and magnesium salt. The amount is 0.1 to 30 mol per mol of oil or fat, preferably 0.5
It is preferable to add up to 5 molar amount. The entire amount of hydrogen carbonate, lhydrogen phosphate, or alkaline earth metal salt may be added at the beginning, or a portion may be added in portions during the reaction.

In the present invention, water and alkaline lipase are added to fats and oils,
Fats and oils are decomposed in the presence of bicarbonate, alkaline earth metal salts, and monohydrogen phosphate to obtain a decomposition product rich in monoglycerides, from which monoglycerides are collected, but the amount of water added to fats and oils is , 0.5 to fat
It is preferable to set it to 200% (W/W).

In addition, at this time, glycerin is further added to the fat, preferably 5% (W/W) or more based on the fat and oil to decompose the fat, and a re-esterification reaction is performed between the free fatty acid and the added glycerin by the action of lipase. By doing so, it is possible to increase the monoglyceride content in the decomposed fats and oils, and at the same time significantly reduce the free fatty acid content.

It is preferable to select an appropriate decomposition temperature depending on the fats and oils and alkaline lipase used, and stir if necessary.

If it is necessary to collect and use monoglyceride from the decomposition product, first neutralize it with hydrochloric acid, etc., and then use
- Monoglycerides may be recovered by precipitation with hexane, petroleum ether, etc., or they may be separated from fatty acids by distillation.

Decomposition rate measurement and monoglyceride quantification were performed as follows.

After the decomposition product was acidified with hydrochloric acid, the oil was extracted with diethyl ether, washed with water, dehydrated, and the ether was distilled off to collect the oil.

The acid value and saponification value of this oil were determined, and the ratio of the two was expressed as a percentage to determine the decomposition rate. In addition, the total monoglycerides in the oil were measured by the following method.

Assay method for monoglycerides: [Reagent] Periodic acid reagent dimetetaperiodate sodium 5
．． Dissolve 0g in 200cc of distilled water and add glacial acetic acid to make 1
Set to 1.

[Method] Precisely weigh the sample into a 250cc stoppered Erlenmeyer flask,
Add 15 cc of a solvent of chloroform glacial acetic acid 1:2 to dissolve. Add 0.045 cc of 56% perchloric acid, stir for 1 minute, and leave for 9 minutes. Add 25 cc of periodic acid reagent and let it stand for 15 to 30 minutes, then add 20 cc of 10% potassium iodine solution, and after 1 minute, satisfy with N/10 sodium thiosulfate solution. A 1% starch solution is used as the indicator. A blank test will be conducted at the same time.

[Calculation]

B: Sodium thiosulfate required for blank test CCC: Sodium thiosulfate required for this test ccN: Normality of sodium thiosulfate M: Molecular weight of monoglyceride (356, as monoolein) S: Weight of sample collected in this test g Note that C/B is preferably about 80 to 90%. For this purpose, the amount of sample collected should be adjusted, and the total amount of monoglycerides should be 1
Approximately 0.00 g is appropriate.

Next, examples of the present invention will be shown, but the present invention is not limited to these examples.

Example 1 Put 50 g of olive oil, 9.49 g of sodium bicarbonate, and 14 m of water into a 500 m Erlenmeyer flask, and heat at 30°C for 2 hours.
The mixture was stirred using a rotary shaker at 20 revolutions/minute. When everything is thoroughly mixed, Meito PL-266 bacteria (@
1 rtrl (containing 2500 units) of an aqueous solution of alkaline lipase powder obtained by drying the precipitate obtained by adding acetone to the culture filtrate of Koken Bacterium No. 3187) was added.

Stirring was continued, and after 4 hours, 15 d of 4N hydrochloric acid was added to stop the reaction, and diethyl ether ■0011I1. was added to extract the oil, and after washing with water and dehydration, the ether was distilled off to obtain 48.7 g of oil. The monoglyceride content in this oil was measured to be 31.0%, and the fat and oil decomposition rate at this time was 66.0%.

Example 2 Name I! 1PL-679 bacterium (Feikoken Bacteria No. 3783)
Using an aqueous solution (containing 2,500 units) of alkaline lipase powder obtained by adding acetone to the culture filtrate and drying the precipitate, use 19.67 g of potassium monohydrogen phosphate instead of sodium bicarbonate. Except for this, the same procedure as in Example 1 was carried out to obtain 48.5 g of oil. The molar glyceride content contained in this oil is 29%,
The fat and oil decomposition rate at this time was 63%.

Example 3 50 g of palm oil, 11.8 g of potassium bicarbonate, and 15 ml of water were stirred in the same manner as described in Example 1, and then 1 ml of an aqueous solution of the same alkaline lipase powder as used in Example 1 was added. (containing 1500 units) was added.

Continue stirring, and after 4 hours, heat to 70℃,
15 d of N-hydrochloric acid was added and 47.5 g of oil was obtained in the same manner as described in Example 1 below. The monoglyceride content in this oil is 32.3%, and the fat decomposition rate at this time is 6.
It was 4.5%.

10 ml of petroleum ether was added to 40 g of this oil and dissolved by heating while stirring. The solution was left at 22° C. for 24 hours, the resulting precipitate was collected by filtration, and the petroleum ether was removed under reduced pressure to obtain 22 g of an oil.

The monoglyceride content of the main island was 58.6%.

Example 4 50 g of olive oil, 4.3 g of sodium hydrogen phosphate,
After adding 50 g of glycerin and 3.0 ml of water and stirring in the same manner as described in Example 1, the name tJ! PL-266
1 d (contains 2500 units) of an aqueous suspension of alkaline lipase adsorbed on bentonite obtained by adding bentonite to the culture filtrate of No. 3 bacteria (Feikoken Bacteria No. 3187) and filtering it.
was added and continued stirring, and after 3 days, 4N-5W hydrochloric acid was added.
The reaction was stopped by adding t1, and 49.5 g of oil was obtained in the same manner as described in Example 1 below.

The monoglyceride content in this oil was 46.0%, and the fat and oil decomposition rate at this time was 11%.

Next, this oil is purified by distillation under reduced pressure, resulting in a purity of 9.
24.2 g of 3.2% monoglyceride were obtained.

Example 5 Sardine oil (manufactured by Riken Vitamin Co., Ltd.) 50 g, calcium hydroxide 6 g, and lipase PL-679 (100,000 units/g) 3
12 d of distilled water in which 5.5.

Collect 2 g of the reaction product, add 2 d of 5N hydrochloric acid and 2 d of petroleum ether.
After adding 0- and shaking thoroughly to extract the oil, 1.000
The petroleum ether layer was recovered by centrifugation at g. After washing the petroleum ether layer with 20 d of distilled water, the petroleum ether was distilled off using a rotary evaporator to obtain an oil component.

Regarding this oil content, Standard Oil and Fat Analysis Methods 2 and 4 edited by Japan Oil Chemists' Association
- The acid value was measured according to 1-83. In addition, the saponification value of sardine oil was measured according to the same analytical method 2, 4, 3-71. The decomposition rate of sardine oil is calculated by dividing the acid value by the saponification value, 1
When calculated by multiplying by 00, it was 84%. Also,
5cm of this oil was added to a silica gel plate (manufactured by Merck & Co., No.
13895) at a position of 1 cm from the lower end, and after developing for 10 cm with a mixed solvent of petroleum ether, ether, and acetic acid (70:30:1°V/ν), monoglyceride (abbreviated as MG) was applied under ultraviolet irradiation with a wavelength of 254 nm. Detect the spot, scrape it into a test tube with a stopper, and apply the above analysis methods 2, 4, and 20.
- Fatty acid methyl ester was prepared according to 2.77. 1
Using a glass column packed with 0% EGS (manufactured by Gas Kuro Kogyo Co., Ltd., inner diameter 3 m, length 2 m, NαAA45095), heating gas chromatography (analysis temperature 170-2 m) was used.
The composition of the fatty acid methyl ester was analyzed using a heating rate of 2°C/min (hereinafter abbreviated as GC). Similarly, fatty acids contained in the oil were separated by TLC, and the composition was analyzed by GC as methyl esters. These results are shown in Table 2.

The fatty acid composition of sardine oil is also shown in Table 2.

Table 2 MG fatty acid group 1 (area %) EPA, DPA, and DHA represent eicosapencitric acid, docosapenecitric acid, and docosahexaenoic acid, respectively.

As can be seen from Table 2, compared to the raw material sardine oil, MG
EPA, DHA, etc. P [JFA is 2.2 times, 70
% and concentrated to a high concentration.

Furthermore, 5 μg of the recovered oil was applied to Chromarod S-■ (manufactured by Yatron), and benzene, chloroform, acetic acid (
50: 30: 1, v/v) 10 cm with mixed solvent
Expand and use Iatoroscan Tl-10 (manufactured by Yatron)
When the MG content in the oil was measured, it was found to be 16.5% (
area%).

Example 6 Distilled water in which 50 g of sardine oil (manufactured by Hyozan Yakuhin Co., Ltd.), 12 g of sodium hydrogen carbonate, and lipase PL-266125 were dissolved was collected in a 500-flask with a baffle, and heated at 35°
A shaking reaction was carried out at C for 24 hours.

The decomposition rate measured by the above method was 58%, and the MG content was 3.
It was 2.3%. The PUFA content of MG is shown in Table 3.

Table 3 PUFA in MG The results in Table 3 show that PUFA was concentrated 2.1 times in MG compared to sardine oil.

Claims (1)

[Claims] 1. Adding water and alkaline lipase to fish oil and decomposing the oil in the presence of bicarbonate, monohydrogen phosphate or alkaline earth metal salt to obtain a decomposition product rich in monoglycerides. A method for producing monoglyceride characterized by: 2. The method according to claim 1, wherein 5.0% (W/W) or more of glycerin is added to the fish oil.