JP2000312598A - Synthesis of new capsaicinoid-like substance - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain a capsaicinoid substance not having hot taste at a low cost in a short time and in large quantities by reacting vanillyl alcohol with a fatty acid ester, a triacylglyceride and/or a fatty acid utilizing an inverse reaction with a lipase. SOLUTION: By reacting vanillyl alcohol as a substrate with at least one selected from the group consisting of a fatty acid ester of the formula R'-OCO-R (wherein R and R' are each an aliphatic hydrocarbon group), a triacylglyceride of formula I (wherein R1 to R3 are each an aliphatic hydrocarbon group and at least one of R1 to R3 is identical to R) and a fatty acid of the formula RCOOH as substrate, utilizing an inverse reaction with an enzyme such as a lipase, the objective capsaicinoid substance of formula II, useful e.g. for a food additive or medicaments and not having hot taste is obtained at a low cost in a short time and in large quantities.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to a method for synthesizing a novel capsaicinoid-like substance.

[0002]

2. Description of the Related Art Pepper (Capsicum annuum L.)
It is a plant widely used worldwide as a food, spice and pharmaceutical ingredient, and its pungent component is capsaicin ((E) -N-[(4-hydroxy-3-methoxyphenyl) -methyl] -8-m
ethyl-6-nonenamide), dihydrocapsaicin, ie
And 6,7-dihydro derivatives of capsaicin. However, despite the fact that these capsaicins and the like have physiological activities such as an obesity-suppressing action, their use is limited due to their strong pungency and invasiveness, and their use as food additives or pharmaceuticals is quite limited. Met. In recent years, a non-pungent fixed cultivar "CH-19 sweet" containing a large amount of capsaicinoid-like substances without pungency has been reported (Yazawa,
S .; Suetome, N .; Okamoto, K .; Namiki, T., J. Japa
n Soc. Hort. Sci., 1989, 58, 601-607). This non-pungent capsaicinoid-like substance has the following chemical formula:

[0003]

Embedded image

[0004]

These novel non-pungent capsaicinoid-like substances have the potential to be used as pharmaceuticals or food additives because of their lack of pungency, but when obtained from plants, their production is limited. was there.

[0005] In order to solve the above problems, the present invention has been made to develop a synthesis method for mass-producing a novel capsaicinoid-like substance at a low cost in a short time.

[0006]

Means for Solving the Problems The present inventors have solved the above problems by establishing a biochemical synthesis method using enzymes, particularly lipase, which is a kind of esterase, instead of chemical synthesis.
The present invention has been completed. That is, the present invention uses an enzyme such as lipase, and is characterized by general formula (I):

[0007]

Embedded image

(Wherein, R represents an aliphatic hydrocarbon group)
The present invention relates to a method for synthesizing a novel capsaicinoid-like substance represented by the formula: The present invention particularly uses lipase as the enzyme and has the general formula (II):

[0009]

Embedded image

Wherein R, R1, R2, and R3, and R ′ each represent an aliphatic hydrocarbon group;
At least one of the three is equal to R. The present invention relates to a method for synthesizing the above capsaicinoid-like substance, characterized in that at least one of the fatty acid ester, triacylglyceride and fatty acid represented by the formula (1) and vanillyl alcohol are used as substrates and a reverse reaction of lipase is utilized. In the above general formulas (I) and (II), R, R1, R
2 and R3 are linear or branched and are a saturated or unsaturated aliphatic hydrocarbon group. R is preferably a linear saturated aliphatic hydrocarbon or a branched unsaturated aliphatic hydrocarbon group. Further, the number of carbon atoms contained in these aliphatic hydrocarbon groups is not particularly limited, and may be, for example, in the range of 5 to 17. R ′ of the fatty acid ester in the general formula (II)
Is preferably an alkyl group such as methyl and ethyl. As a substrate other than vanillyl alcohol, a fatty acid ester is preferable in terms of yield and purification of a capsaicinoid-like substance. The fatty acid used as a substrate in the synthesis method of the present invention may be one generated in situ by hydrolysis of a fatty acid ester or triacylglyceride by esterase.

The enzymes used in the synthesis method of the present invention include:
For example, it is preferable to use esterases, especially lipases. Lipase is a type of esterase, an enzyme involved in the hydrolysis of fats, which are glycerin esters of fatty acids, and is widely distributed in animals, plants and microorganisms.
In the synthesis method of the present invention, there is no particular limitation on the biological source of the enzyme to be used. For example, a lipase derived from fungi, animals and plants, and the like can be suitably used. Furthermore, in the synthesis method of the present invention, it is preferable to use an immobilized enzyme obtained by binding such an enzyme to an appropriate carrier from the viewpoint of reaction efficiency and economical efficiency. Various techniques for immobilizing these enzymes are known to those skilled in the art,
In addition, since various immobilized enzymes, for example, immobilized lipase are commercially available, those skilled in the art can easily obtain them.

The type and amount of the enzyme used in the synthesis method of the present invention, the biological source, and the type and amount of the substrate can be appropriately selected by those skilled in the art. Further, the reaction temperature, time, pH, reaction solvent, etc. Those skilled in the art can appropriately set the various conditions described above. After the synthesis, the obtained capsaicinoid-like substance can be purified by a suitable known means such as a silica gel column.

[0013]

DESCRIPTION OF THE PREFERRED EMBODIMENTS Hereinafter, the present invention will be described with reference to examples, but the present invention is not limited to these examples. Incidentally, in the following examples, the structure of the synthesized compound has a high-resolution mass spectrum, an infrared absorption spectrum,
It was identified by an ultraviolet absorption spectrum and a nuclear magnetic resonance spectrum.

[0014]

EXAMPLES Example 1 Synthesis of Capsiate 1 gram of capsaicin (Sigma, M-2028, 98% purity)
The mixture was refluxed for 45 hours with 750 ml of 4N aqueous methanolic hydrochloric acid (a mixture of 250 ml of concentrated hydrochloric acid and 500 ml of methanol). Extract three times with petroleum ether, wash twice with 0.1 N hydrochloric acid, dehydrate with sodium sulfate, evaporate the solvent,
0 mg was obtained. The yield was as high as 92.9%. The thus obtained fatty acid methyl ester 560 mg (3.04 mmol, final concentration; 40 mM) and 11.7 g of vanillyl alcohol (76.0 mmo)
l) was dissolved in 76 ml of 1,4-dioxane, and 3.0 g of immobilized lipase (Novozym435: manufactured by Novo Nordisk) was added.
The reaction was performed for 30 minutes. After the reaction is completed,
Excluding vozym435, hexane was added to the filtrate to remove excess vanillyl alcohol by precipitation, and the compound was recovered.
The solvent was distilled off, and the weight of the compound was measured.
Met. Next, silica gel 30
The capsiate was purified using a column packed with g in a column having an inner diameter of 3.5 cm. The obtained compound was purified with an eluent of hexane and ethyl acetate (10: 1) to finally obtain 561 mg of capsaicin, a capsaicinoid-like substance.
The yield was as high as 60.1% and the purity was as high as 95%.
The yield and purity were determined by HPLC measurement under the following conditions, with the area obtained using the standard substance as 100%.
It was calculated and calculated from the ratio of the area obtained from the purified material. HPLC conditions: Column: 5C18 wakosil AR (4.6 mm diameter x 15
0 mm) Eluent: 85% methanol Flow rate: 0.5 ml / min Detection: fluorescence EX280 nm EM320 nm

Example 2 Investigation of reaction time and reaction temperature Using vanillyl alcohol 7.7 mg (50 μmol) and methyl nonanoate 1.0 μl (5 μmol) as a substrate, 1,4-dioxane 500 μl
Was used as a reaction solvent, and enzymatic synthesis of a capsaicinoid-like substance, vanillyl nonanoate, was performed using 20 mg of immobilized lipase Novozym435, and the reaction time and reaction temperature were examined. The purified compound was purified in the same manner as in Example 1. As a result, as shown in FIG. 1, the reaction at 60 ° C. for 30 minutes was the highest in terms of yield, with a yield of 60% or more.

Example 3 Capsaicinoid-like substance using various fatty acid methyls
Mass Synthesis of Methyl Nonanoate of Example 2 (R is an aliphatic hydrocarbon group)
(Carbon number contained in R which is an aliphatic hydrocarbon group: 7) and methyl caprate (decanoic acid) methyl (aliphatic hydrocarbon group) And the number of carbon atoms contained in methyl hexanoate (an aliphatic hydrocarbon group R):
5), methyl laurate (the number of carbon atoms contained in R as the aliphatic hydrocarbon group: 11), methyl stearate (the number of carbon atoms contained in R as the aliphatic hydrocarbon group: 17), and methyl myristate (the fat) An enzymatic synthesis of a capsaicinoid-like substance was performed using the number of carbon atoms contained in R which is a group hydrocarbon group: 13). Further, in this example, the scale was raised, and using 7.7 g (50 mmol) of vanillyl alcohol and each of the above fatty acid methyl (2.5 mmol) as substrates, 50 ml of 1,4-Dioxane was used.
Was used as a reaction solvent, and 2 g of immobilized lipase Novozym435 was shaken at 60 ° C. for 30 minutes to react. The purified compound was purified in the same manner as in Example 1. As a result, as shown in Table 1, various capsaicinoid-like substances contained about 30 to 77%
And a high yield was obtained.

[0017]

[Table 1]

Example 4 Examination of Substrate Vanillyl nonanoate was synthesized in the same manner as in Example 2 except that trinonanoin was used instead of methyl nonanoate. The purified compound was purified in the same manner as in Example 1. As a result, as shown in Table 2, under the optimal conditions, the case where trinonanoin was used (a yield of about 46 and about 50% in a reaction for 90 minutes) was better than the case where methyl nonanoate was used (15 minutes). Yield of about 40%). But,
In the case of trinonanoin, the reaction time had to be extended.

[0019]

[Table 2]

The excess amount of vanillyl alcohol used as a substrate is reduced to 90% by precipitation with hexane.
About% was recoverable and could be reused. Furthermore, the immobilized lipase (Novozym435) did not show a remarkable decrease in activity even when repeatedly used, and could be reused at least 10 times or more.

[0021]

According to the present invention, a novel capsaicinoid-like substance can be mass-produced inexpensively and in a short time and in high yield.

[Brief description of the drawings]

FIG. 1. Relationship between reaction time and yield in the synthesis of vanillyl nonanoate at various reaction temperatures.

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[Procedure amendment]

[Submission date] April 26, 2000 (200.4.2
6)

[Procedure amendment 1]

[Document name to be amended] Statement

[Correction target item name] Claims

[Correction method] Change

[Correction contents]

[Claims]

Embedded image A method for synthesizing a novel capsaicinoid-like substance represented by the general formula (II):

Embedded image (In the above formulas (I) and (II), R1, R2 and R
3, and R ′ represent an aliphatic hydrocarbon group; R represents an aliphatic hydrocarbon group having 5 to 17 carbon atoms;
2, and at least one of R3 is equal to R. )) Wherein at least one of the fatty acid ester, triacylglyceride and fatty acid represented by formula (1) and vanillyl alcohol are used as substrates, and the reverse reaction of lipase is used.

 ────────────────────────────────────────────────── ─── Continued on the front page (72) Inventor Masatake Imai 2-1-1 Shimosueyoshi, Tsurumi-ku, Yokohama, Kanagawa Prefecture Inside Morinaga Confectionery Co., Ltd. (72) Inventor Masatoshi Kato 2- Shimosueyoshi, Tsurumi-ku, Yokohama-shi, Kanagawa 1-1 Morinaga Confectionery Co., Ltd. (72) Inventor Shuichi Hashizume 2-1-1 Shimosueyoshi, Tsurumi-ku, Yokohama, Kanagawa Prefecture Morinaga Confectionery Co., Ltd. F-term (reference) 4B064 AD64 AD68 CA21 CA31 CD06 CD07 CD12 CD22 CE10 DA16

Claims (7)

[Claims] 1. A compound of the general formula (I), wherein an enzyme is used. (Wherein R represents an aliphatic hydrocarbon group). A method for synthesizing a novel capsaicinoid-like substance represented by the formula: 2. The method according to claim 1, wherein a reverse reaction by an esterase is used. 3. A compound of the general formula (II): (Wherein, R, R1, R2, and R3, and R ′ each represent an aliphatic hydrocarbon group, and at least one of R1, R2, and R3 is equal to R). The method according to claim 2, wherein at least one of acylglycerides and fatty acids and vanillyl alcohol are used as substrates, and a reverse reaction of lipase is used. 4. In general formulas (I) and (II),
Is a linear saturated aliphatic hydrocarbon group, The synthesis method according to claim 3, wherein 5. In general formulas (I) and (II),
Is a branched unsaturated aliphatic hydrocarbon group, The synthesis method according to claim 3, wherein 6. In general formulas (I) and (II),
The synthesis method according to claim 4, wherein the number of carbon atoms contained in the compound is 5 to 17. 7. The synthesis method according to claim 3, wherein an immobilized lipase is used.