JP3159551B2 - Method for producing lignans and fats and oils containing the same - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for producing a lignan glycoside, which is an oil-soluble lignan or its precursor, by allowing an enzyme preparation to act on pulverized sesame seeds, defatted lees or a hydroalcoholic extract thereof. The present invention relates to a method and a method for producing fats and oils containing them.

[0002]

2. Description of the Related Art Sesame seeds, which have been widely used for food since ancient times, contain a large amount of lipid components (oil content: 45 to 55%).
It has been prized as an excellent oil and fat resource in the food industry. Despite its high content of unsaturated fatty acids such as oleic acid and linoleic acid, its lipid components are rich in long-term storage in seeds and are not found in the fats and oils of other raw materials. It has excellent stability against oxidative deterioration. For this reason, there have been many reports on studies on antioxidant components in sesame seeds (for example, edited by Mitsuo Namiki and Sadasaku Kobayashi, "Sesame Science", Asakura Shoten, 1989).

A characteristic component of sesame oil is sesamin (tetrahydro-1,4-di [3,3] derived from sesame seeds.
4- (methylenedioxy) phenyl] -1H, 3H-furo [3,4-C] furan), sesamoline (tetrahydro-1- [3,4- (methylenedioxy) phenoxy]-
4- [3,4- (methylenedioxy) phenyl] -1
Lignans such as H, 3H-furo [3,4-C] furan) are known. Further, P-1 (tetrahydro-1- (3-methoxy-
4-hydroxyphenyl) -4- [3,4- (methylenedioxy) phenyl] -1H, 3H-furo [3,4-
C] furan), sesamolinol (tetrahydro-1-)
[3-methoxy-4-hydroxyphenoxy] -4-
[3,4- (methylenedioxy) phenyl] -1H, 3
H-furo [3,4-C] furan), pinoresinol (tetrahydro-1,4-di (3-methoxy-4-hydroxyphenyl) -1H, 3H-furo [3,4-C] furan), sesaminol (Tetrahydro-1- [6-hydroxy-3,4- (methylenedioxy) phenyl] -4-
[3,4- (methylenedioxy) phenyl] -1H, 3
So-called phenolic lignans such as H-furo [3,4-C] furan) have been found to have superior antioxidant activity over conventional natural antioxidants. However, many of these lignans are present as sugar compounds (lignan glycosides) in sesame seeds or their defatted meals.

[0004] As a method for producing such lignans, sesame seeds or defatted lees are extracted with hydroalcoholic alcohol, β-glucosidase is allowed to act on the extract, and sugar chains are cleaved.
Separation using a solvent such as ethyl acetate (Japanese Patent Publication Sho 62
No. 4,479,793) and a method of allowing a microorganism such as Bacillus natto to act on the same raw material (JP-A-59-1570).
No. 86) has been reported.

However, in the method for producing sesame lignans using β-glucosidase as described above,
Due to the substrate specificity of β-glucosidase, only a part of the lignan glycoside in sesame seed is hydrolyzed,
When sesame seeds or its defatted lees are used as a raw material, coexisting cell tissue residues such as cellulose prevent sufficient extraction and separation of sesame lignans from the enzyme reaction solution. for that reason,
As a pretreatment, it is necessary to extract the raw material with hydroalcohol, etc., which consumes a large amount of solvent.Because β-glucosidase is a relatively expensive enzyme agent, it is used alone to produce large amounts of sesame lignans on an industrial scale. Production has problems such as an increase in the cost of the product, and there are many points to be improved.

[0006] Thus, despite the desire for a stable supply of excellent natural antioxidants, to date, the production of lignan analogs from sesame seeds or defatted meal has been carried out on an industrial scale. The current situation is that it has not been
Development of a more efficient manufacturing method has been desired.

[0007]

In view of this situation, the present inventors have made intensive studies on an efficient method for producing lignans derived from sesame seeds, and have completed the present invention. That is, the present invention relates to a method for producing a large amount and easily of oil-soluble lignans by causing a specific enzyme preparation to act on ground sesame seeds or a processed product thereof, and a method for producing oils and fats containing the lignans.

[0008]

SUMMARY OF THE INVENTION The first aspect of the present invention is as follows.
A process for producing oil-soluble monoglucoside lignans or fats and oils containing the same, characterized in that a crushed sesame seed, defatted cake or a hydroalcoholic extract thereof is used as a raw material and a cellulase preparation is allowed to act thereon.

[0009] In the present invention, sesame seeds can be used irrespective of their type and production area, and irrespective of roasting or moulting. Further, it may be coarsely or finely pulverized by a general-purpose pulverizer, or may be in a pressed state. There are various methods for obtaining defatted cake, but it is obtained as a residue obtained by squeezing using a normal pressing device, an expeller, or the like. It cannot be used as a raw material for the invention. Further, in the present invention, an extract obtained by extracting the crushed sesame seeds or defatted lees with hydroalcohol may be used as a raw material. Here, as the water-containing alcohol, a linear or side chain lower alcohol having 1 to 4 carbon atoms, for example, methanol, ethanol, propanol, isopropanol, butanol or the like is mixed with water, and the alcohol concentration is adjusted to 30 to 100 vol / vol. % (Hereinafter,% (v /
v)), preferably adjusted to 50 to 100% (v / v). An alcohol concentration of less than 30% (v / v) is not preferable because a large amount of the water-soluble polysaccharide not containing the target substance is contained in the extract. 1 to 10 times volume / weight (hereinafter, times (v / w)
The extraction procedure is repeated if necessary, and the insoluble matter is removed by a conventional method to obtain an extract. The extract may be appropriately concentrated, but it is necessary to remove at least the alcohol content in order to avoid inactivation of the enzyme during the enzyme treatment described below. The extract is a mixture containing various sugar chain compounds in addition to the lignan glycoside.

Next, the object of the present invention is achieved by allowing a cellulase preparation to act on the above-mentioned sesame seed pulverized matter, defatted lees or a hydroalcoholic extract thereof (hereinafter, these may be simply referred to as raw materials). Oil-soluble monoglucoside lignans or fats and oils containing the same can be obtained. The cellulase preparation can be used irrespective of its origin and purification degree as long as it has an activity of hydrolyzing long-chain and short-chain sugar bonds, and a commercially available microorganism-derived product can be suitably used. Also included are those in which the cellulase is immobilized on a suitable carrier so that it can be used repeatedly. The treatment with such a cellulase preparation may be performed in accordance with a general enzyme reaction method, and is not particularly limited, but is usually performed as follows. That is, the raw material is dispersed or dissolved in 1 to 20 times (v / wt) water or a buffer solution (pH 2 to 6), and 0.1 to 30% (wt / wt) with respect to the raw material,
Preferably, a cellulase preparation of 1 to 10% (wt / wt) is added, and the sugar chain is hydrolyzed at 10 to 50 ° C for 1 to 24 hours, preferably with gentle stirring. When crushed sesame seeds or defatted cake is used as a raw material, cell tissue debris remains in the reaction solution, which is removed by appropriate means such as centrifugation and filtration, and ethyl acetate is added to the reaction solution. The desired monoglucosidic lignans can be obtained by adding a solvent that is hardly soluble in water, such as butanol or hexane, followed by extraction. In addition, in order to obtain fats and oils containing the monoglucosidic lignans, a raw material containing fats and oils such as crushed sesame seeds or pressed lees may be treated in the same manner.

The monoglucosidic lignan thus obtained is a mixture of sesaminol as a main component and lignans such as sesamolinol, P-1, pinoresinol and glucose, and ethyl acetate, butanol, hexane, edible oils and fats. Oil-soluble and water-insoluble. In addition, if necessary, it can be fractionated and purified into individual components such as monoglucoside sesaminol and monoglucoside sesamolinol using an adsorbent such as silica gel and ODS. Unlike the lignan glycosides originally present in sesame seeds, these monoglucosidic lignans are oil-soluble and can be used for lipid materials, and have no antioxidant activity per se. As a precursor of an oxidized component, it is a component that undergoes hydrolysis of a sugar bond, for example, is expected to specifically exhibit its activity in a living body.

The second aspect of the present invention is to provide an oil-soluble method characterized by using a crushed sesame seed, defatted lees or a hydroalcoholic extract thereof as a raw material, and allowing a cellulase preparation and a β-glucosidase preparation to act on this. A method for producing lignans or fats and oils containing the same.

[0013] In the present method, the method for producing oil-soluble monoglucoside lignans by the above-mentioned cellulase preparation comprises the step of producing an oil-soluble lignan or an oil or fat containing the same by carrying out an enzyme reaction in combination with a cellulase preparation and a β-glucosidase preparation. Is the law. In this method, the same raw materials as those used in the treatment with the cellulase preparation alone are used. The origin and activity of the β-glucosidase preparation are not limited, and it is convenient to use a general-purpose commercial product. Also immobilized β
-A glucosidase preparation may be used. Such a β-glucosidase preparation and the cellulase preparation are in a weight ratio of 10 to 90:90 to 10, preferably 10 to 40: 9.
Mix at a ratio of 0 to 60, and add 0.1 to
30% (wt / wt), preferably 1 to 10% (wt / wt) may be used. The reactions using both enzyme preparations are performed simultaneously or, if necessary, the reaction using the cellulase preparation and then the β-glucosidase preparation are separately performed.
In addition, the enzyme reaction conditions (temperature, pH, time) and the method of treating the reaction product in this method may be the same as those of the above-described treatment using only the cellulase preparation, and thus the desired oil-soluble lignans can be obtained. If a raw material containing fats and oils such as ground sesame seeds or pressed lees is used, fats and oils containing the lignans can be obtained.

According to this method, the decomposition of impurities (polysaccharides such as cellulose) in the raw material is promoted by the cellulase preparation, the lignans can be efficiently extracted from the reaction solution, and the cellulase preparation and the β-glucosidase preparation are used in combination. The single or continuous enzymatic reaction has an effect that a larger amount of lignans can be produced as compared with the conventional β-glucosidase alone method. Furthermore, since cellulase preparations are versatile and inexpensive compared to β-glucosidase preparations, it is possible to reduce the production cost of lignans.

The lignans obtained by the present method are a mixture of sesaminol as a main component, sesamolinol, P-1, pinoresinol and the like, and these are known as oil-soluble antioxidant components. Such lignans can be used as they are, or if necessary, subjected to a conventional adsorbent treatment or fractionation treatment to be highly purified, and can be used as antioxidants for oils and fats, foods, cosmetics, agricultural chemicals, pharmaceuticals and the like.

[0016]

【Example】

Example 1 After crushing sesame seeds with an expeller, the remaining oil was extracted with hexane, and 100 g of defatted lees was extracted with 100 g of 1N acetic acid buffer (pH 4.5), 1 liter, cellulase preparation (Boehringer Mannheim Yamanouchi, 500 units / g). 10
g was added and shaken at 40 ° C. for 10 hours. Then, 500 ml of ethyl acetate was added to the reaction solution, and extraction was performed twice.
After separating the aqueous layer and the residue, the ethyl acetate layer was dried over anhydrous sodium sulfate to remove the desiccant, and the solvent was distilled off to obtain 3 g of an extract. This extract was treated with octadecyl silica (YMC, YMC-GEL ODS-A 60-2).
(00/60) as a carrier and purified. Elution with a mixed solvent of water / methanol
The major fraction eluted with 0% (v / v) aqueous methanol was collected. The measurement results of ¹³ C-NMR spectrum and mass spectrum of this product, and GC-MS of the hydrolyzate of the fraction with β-glucosidase
The isolated substance was identified as monoglucoside sesaminol from the result of spectrum measurement (identification of sugar chain). ¹³ C-NMR (CD ₃ OD) 53.3, 53.8, 60.7, 69.5, 70.6,
72.3, 73.2, 76.4, 76.5, 81.1,
85.2, 98.3, 100.7, 100.9, 10
1.8, 104.4, 105.9, 107.3, 11
9.1, 123.8, 134.9, 142.6, 14
7.1, 147.8, 149.1 Mass spectrum (relative intensity) 442 (14.3), 361 (20.3), 271
(7.2), 243 (7.6), 217 (23.6),
169 (13.5), 147 (29.1), 129 (2
7.0), 103 (21.9), 73 (100.0) From similar analysis results of other fractions obtained by the column chromatography, sesamolinol, P-1, Pinoresinol was observed. Furthermore, the solubility of the above extract and monoglucoside sesaminol in various solvents (100 ml solvent /
1g sample), both samples showed ethyl acetate, n
-Soluble in butanol, hexane, soybean oil and insoluble in water.

Example 2 100 g of sesame defatted lees as in Example 1, 1 liter of 0.1 N acetate buffer (pH 4.5), 10 g of cellulase preparation (same as in Example 1) and β-glucosidase preparation (10000 units manufactured by Funakoshi) / G) and shaken at 40 ° C. for 20 hours. 500 ml of ethyl acetate was added to the reaction solution, and extraction was performed twice. Thereafter, the same treatment as in Example 1 was performed to obtain 2 g of an extract. From the results of analysis and identification of the extract by gas chromatography and mass spectroscopy, the ratio of lignans in the extract was as follows: sesaminol: sesamolinol: P-1: pinoresinol = 1
0.3: 1.8: 1.8: 2.7.

Example 3 The same sesame defatted cake as in Example 1 was extracted by adding 5% (v / wt) of an 80% (v / v) aqueous ethanol solution to 1 kg of the sesame defatted lees, and the insoluble material was filtered off. The residue was dried under reduced pressure to obtain 40 g of an extract. The extract was then added to a 0.1N acetate buffer (pH 5.
0) 400 ml, 4 g of the cellulase preparation used in Example 2
And 2 g of a β-glucosidase preparation were added and shaken at 40 ° C. for 20 hours to cause a hydrolysis reaction. The reaction solution was sampled with time and used in Example 1 using ethyl acetate.
An extract was obtained in the same manner as described above. The extract was analyzed in the same manner as in Example 2, and the content of sesaminol, a main component, was determined. The change with time is shown in FIG. In this method, β-
About 7 of the conventional method using only glucosidase (Comparative Example 1)
Double the amount of sesaminol was obtained.

Example 4 In Example 3, the reaction was carried out using only the cellulase preparation, and the extract was similarly analyzed for the sesaminol content in the extract. The time-dependent changes are shown in FIG. Compared with the conventional method (Comparative Example 1), it became clear that the amount of sesaminol production increased as the reaction time was increased.

Comparative Example 1 In Example 3, the reaction was carried out using only the β-glucosidase preparation, and the extract was similarly analyzed for the sesaminol content in the extract, and the change with time is shown in FIG. The amount of sesaminol produced is lower when the cellulase preparation and the β-glucosidase preparation are used in combination (Example 3) than when only the cellulase preparation is used (Example 4).

Example 5 Example 2 was repeated except that sesame seeds were used as a raw material instead of sesame defatted meal in a blender, and ethyl acetate / hexane (1: 1) was used as an extraction solvent for the reaction solution. The same operation was performed. The obtained extract was subjected to separation of oil and fat using a silica gel column and hexane, and lignans in the residue were analyzed.
Lignans: 0.25% (including sesaminol: 0.15%, sesamolinol: 0.025) with respect to 100%
%, P-1: 0.025%, Pinoresinol: 0.05
%) Was included.

[0022]

According to the present invention, pulverized sesame seeds, defatted lees or a hydroalcoholic extract thereof are used as raw materials,
By using an inexpensive cellulase preparation, oil-soluble monoglucoside lignans, which are precursors of lignans containing sesaminol as a main component, and fats and oils containing the same can be produced by an easy operation. By using the preparation and the β-glucosidase preparation together, lignans (sesaminol, sesamolinol, P-1, pinoresinol, etc.) known as antioxidant active substances, and fats and oils containing the same are easier than conventional methods. In large quantities and at low cost.

[Brief description of the drawings]

FIG. 1 shows a cellulase preparation (Ce) prepared using an extract obtained by extracting defatted sesame seeds with 80% (v / v) ethanol as a substrate.
ll), β-glucosidase preparation (Glu) or cellulase preparation and β-glucosidase preparation (Cell + Glu)
FIG. 5 is a plot of the amount of sesaminol produced in a reaction product subjected to the reaction with the reaction time. The horizontal axis shows the enzyme reaction time, and the vertical axis shows the relative value of the amount of sesaminol produced.

──────────────────────────────────────────────────続 き Continued on the front page (58) Fields surveyed (Int. Cl. ⁷ , DB name) C12P 19/58 C12P 17/18 C12P 7/64 CA (STN) REGISTRY (STN) JICST file (JOIS) BIOSIS ( DIALOG) WPI (DIALOG)

Claims (4)

(57) [Claims] 1. A method for producing oil-soluble monoglucoside lignans or fats and oils containing the same, wherein a cellulase preparation is allowed to act on pulverized sesame seeds, defatted lees or a hydroalcoholic extract thereof. 2. The process according to claim 1, wherein the monoglucoside lignan is monoglucoside sesaminol. 3. A method for producing oil-soluble lignans or fats and oils containing the same, wherein a cellulase preparation and a β-glucosidase preparation are allowed to act on pulverized sesame seeds, defatted cake, or an aqueous alcohol extract thereof. 4. The method according to claim 3, wherein the lignan is sesaminol.