JPH08154688A - Production of composition containing trans-2-hexenal and perfume composition containing the same composition - Google Patents

Abstract

PURPOSE: To efficiently obtain the subject composition useful as a perfume composition, etc., by bringing a composition containing trans-2-hexenol into contact with a microbial cell of Candida boidinii or its treated substance and producing trans-2-hexenal. CONSTITUTION: A composition, obtained by distilling a mint oil and containing trans-2-hexenol and a 6C acyclic alcohol other than that is brought into contact with a microbial cell of Candida boidinii [e.g. Candida boidinii SA051 strain (FERM BP-4893)] cultured in a culture medium containing methanol or a mixture thereof with glycerol as a carbon source or its treated substance to convert the trans-2-hexenol into trans-2-hexenal. Thereby, the trans-2-hexenol having an aroma unfavorable as a green note is selectively oxidized and converted into the trans-2-hexenal to efficiently afford the objective composition such as a perfume composition.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a method for producing a trans-2-hexenal-containing composition having a preferred green note, a method for producing trans-2-hexenal, and a perfume composition containing the composition.

[0002]

2. Description of the Related Art Trans-2-hexenal is a very strong green note [= fragrant scent, edited by The Japan Fragrance Association, "Encyclopedia of Fragrances" (issued by Asakura Shoten Co., Ltd. on June 25, 1989).
It is a compound having “a term indicating the quality of fragrance” on page xi] and is widely used as a flavoring agent for imparting green notes to apple flavors, grape flavors and the like. Fragrances are roughly classified into two, depending on their raw materials or manufacturing methods: chemically synthesized flavors (so-called "synthetic flavors") and non-chemically synthesized flavors (so-called "natural flavors"). Those who tend to avoid "synthetic flavors" and prefer "natural flavors". But transformer-2
-Hexenal is present in trace amounts in nature, and those extracted from natural products are very expensive. Therefore, in most cases, those produced by chemical synthetic means are used.

On the other hand, as a natural fragrance having a green note, for example, a composition containing an acyclic alcohol having 6 carbon atoms, which is obtained by distilling mint oil derived from Mentha, is used. There is. This composition is mainly composed of 30-55% by weight of trans-2-hexenol, 15-50% by weight of cis-3-hexenol and 15-30% by weight of n-hexanol, and among them, cis-3-hexenol is excellent. I have a green note. However, since the fragrance of trans-2-hexenol is not preferable as a green note, when the composition is used for giving a green note, trans-2 hexenol is not preferable.
-Hexenol content should be as low as possible.

However, trans-2-hexenol and cis-
Since it is extremely difficult to separate it from 3-hexenol, it is also very difficult to separate only trans-2-hexenol from the above composition. It is also possible to selectively oxidize only trans-2-hexenol in the composition to convert it to trans-2-hexenal having a preferable odor, but an effective method for such selective oxidation is known. Has not been done. Therefore, under the present circumstances, a fragrance having a more preferable green note has not been obtained from the above-mentioned composition containing an acyclic alcohol having 6 carbon atoms, which is derived from mint oil.

By the way, Candida Boydini (Ca
ndida boidinii) AOU-1 strain is capable of oxidizing various lower alcohols such as methanol, ethanol, n-propanol and allyl alcohol in a medium containing methanol or methanol and glycerin to convert them into corresponding aldehydes. [Yoshiki Tani et al., Ag
ric.Biol.Chem., 49 (9) pp.2699-2706 (1985) and Yasu
yoshi Sakai et al., Agric. Biol. Chem., 51 (9) pp.2617-26.
20 (1987)]. According to the fermentation method using the Candida voidini AOU-1 strain, aldehydes can be easily obtained by oxidizing alcohols without a coenzyme due to the reaction of enzymes produced by the cells. Moreover, the reaction stops when it is oxidized to aldehyde, and the further oxidation reaction does not proceed.
For the purpose of obtaining an aldehyde, it has an advantage that a step such as stopping the reaction on the way is not required.

However, according to reports to date,
In this method using the Candida voidini AOU-1 strain, alcohols other than the above-mentioned lower alcohols, for example, alcohols having 5 to 8 carbon atoms are not oxidized at all or are trace amounts even if oxidized. Also transformer
There have been no reports of selective oxidation of alcohols with a specific configuration, such as -2-hexenol.

[0007]

Therefore, an object of the present invention is, for example, to selectively oxidize trans-2-hexenol contained in a trans-2-hexenol-containing composition to trans-2-hexenal. Another object of the present invention is to provide a method for producing a composition having a more preferable green note, and a method for producing trans-2-hexenal. Another object of the present invention is to provide a perfume composition having a more preferable green note.

[0008]

In order to solve the above-mentioned problems, the present inventors have conventionally known to oxidize a lower alcohol such as methanol to produce a corresponding aldehyde, Candida voidini. Various mutant strains of The Candida voidini SA051 strain, which is a mutant strain whose parent strain is the Candida voidini AOU-1 strain described in JP-A-61-67480, can produce formaldehyde by culturing in methanol. Although known, [Yasuyoshi Sakai et al., Agric.Bio
l.Chem., 51 (8) pp.2177-2184 (1987)], the present inventors have confirmed that the bacterium of Candida voidini SA051 strain or a treated product thereof has a double bond at the 2-position carbon. It has been found that a trans-type acyclic alcohol having about 4 to 10 carbon atoms having ## STR7 ## is oxidized to be converted into a corresponding aldehyde (trans-2-alkenal). Moreover, when a mixture of alcohols having the same number of carbon atoms is contacted with the bacterial cells of Candida voidini SA051 strain or a treated product thereof, the oxidation rate of trans-2-alkenol is remarkably large as compared with other alcohols. It was found that the trans-2-alkenal can be obtained before the alcohol of the above is oxidized. That is, by utilizing such characteristics of the bacterial cells of the Candida voidini SA051 strain or a processed product thereof, a plurality of trans-2-hexenol, cis-3-hexenol, n-hexanol and the like obtained from mint oil and the like can be obtained. It was found that it is possible to selectively oxidize trans-2-hexenol in a composition containing an acyclic alcohol having 6 carbon atoms into trans-2-hexenal to obtain a composition having a preferable green note, The present invention has been completed.

The present invention provides a composition containing trans-2-hexenol containing Candida boidin.
Contact the microbial cell of ii) or a treated product thereof,
Provided is a method for producing a trans-2-hexenal-containing composition, which comprises converting -2-hexenol to trans-2-hexenal. The present invention also provides a method for producing trans-2-hexenal, which comprises isolating trans-2-hexenal from a trans-2-hexenal-containing composition obtained by the method for producing the above composition. To do.

Furthermore, the present invention provides a perfume composition characterized by containing a trans-2-hexenal-containing composition obtained by the method for producing the above composition. The trans-2-hexenol-containing composition, which is the starting material for the method for producing the composition of the present invention, may be any as long as it contains trans-2-hexenol, but the method of the present invention is It is suitable when it is a composition containing trans-2-hexenol and an alcohol other than trans-2-hexenol, and in particular, trans-2-hexenol and other acyclic alcohols having 6 carbon atoms (for example, cis-3- It is suitable for a composition containing hexenol, n-hexanol, etc.). A typical example of a composition containing trans-2-hexenol and other acyclic alcohol having 6 carbon atoms is a composition obtained by distilling a mint oil derived from the genus Mentha. Can be mentioned. In addition, for example, the genus (Vi
essential oil of fruit of tis); orange oil, mandarin oil, bergamot oil and their water-soluble fractions obtained by pressing the peel of Rutaceae (Rutaceae); water-soluble fraction of apple juice; geranium (Pelargonium inquinan)
s) leaves obtained by steam distillation of geranium oil and its water-soluble fraction; parsley (Petroselinum sativu)
m) leaves obtained by steam-distilling parsley relief oil; legume (Legminosae) Gene (also known as Enida or Bloom. Cytisus scoparius or Genistetinctoria) Absolute obtained by solvent extraction of flowers; narcissus (Na)
A composition derived from a natural product such as absolute obtained by solvent extraction of rcissusTazetta) can also be mentioned.

The composition obtained by distilling the above-mentioned mint oil is obtained by steam-distilling Western plants of peppermint, spearmint, etc., and whole plants of the genus (Mentha) of Japanese genus, such as Japanese mackerel. About 0.08 in mint oil
~ 0.3 wt% is the alcohol content contained in the ratio,
Preferably, a mint oil (a commercially available product can be directly used as the mint oil) is distilled to obtain the first fraction [distillation temperature: 70 to 80 ° C. (13 to 18 mmHg), about 10 to mint oil]. Wt%] can be obtained by further refining by precision distillation or column chromatography. Such a composition consists mainly of an acyclic alcohol having 6 carbon atoms, especially a primary alcohol, and specifically, mainly contains about 30-55% by weight of trans-2-hexenol,
It is composed of about 15 to 50% by weight of cis-3-hexenol and about 15 to 30% by weight of n-hexanol, but if trans-2-hexenol is contained, the other components and the content ratio of each component are not particularly limited. It is not something that will be done. For example, in addition to the above components, trace components such as isomers of acyclic alcohols having 6 carbon atoms such as cis-2-hexenol and trans-3-hexenol may be contained.

The microorganism used in the production method of the present invention may be any strain as long as it belongs to Candida voidini and selectively oxidizes trans-2-hexenol. For example, Candida voidini SA051 Stocks. This Candida Boydini SA051 strain is, as described above, Agric.Biol.Chem., 51 (8) pp.2177-
2184 (1987), a known mutant strain of Candida voidini, which is stored in the Faculty of Agriculture, Kyoto University. The present inventors internationally deposited the Candida Boydini SA051 strain as FERM BP-4893 on November 15, 1994 at the Institute of Biotechnology, Institute of Biotechnology, AIST.

The Candida voidini SA051 strain is finally cultivated using a medium containing methanol alone or a mixture of methanol and glycerin as a carbon source. Of course, in that case, it may be cultured with a substance that promotes the growth of bacterial cells, such as ethanol, glucose, xylose, and mannitol, as well as methanol and glycerin. However, when the bacterial cells cultured in a medium containing no methanol, for example, the bacterial cells cultured in a medium containing only glucose as a carbon source or a treated product thereof, are used as they are in the production method of the present invention, the property of oxidizing alcohol Is not sufficiently extracted, it may be difficult to oxidize trans-2-hexenol.

The medium used for the culture and pre-culture includes, for example, assimilable inorganic nitrogen compounds such as ammonium chloride, ammonium sulfate, ammonium phosphate and urea; potassium phosphate, potassium dihydrogen phosphate, sodium phosphate,
Inorganic salts such as magnesium sulfate, calcium chloride, ferric chloride, ferric sulfate, manganese chloride, manganese sulfate and trace amounts of inorganic salts such as sodium molybdate and potassium iodide; such as disodium ethylenediaminetetraacetate Organic salts; salts of trace elements such as copper, zinc and cobalt; inorganic acids such as boric acid; additives such as vitamins such as biotin and thiamine hydrochloride are appropriately used for culturing general microorganisms. It can be added in a concentration range. A liquid medium is prepared by dissolving these additives and the above carbon source in water. The addition of organic substances such as yeast extract, meat extract and corn steep liquor is not preferable because the production of alcohol oxidase for producing the target trans-2-hexenal is reduced.

For the cultivation of the Candida voidini SA051 strain, the pH of the medium is adjusted to about 4 to 7 at the start of the cultivation, and then about 4 thereafter.
Maintained at ~ 6, under aerobic conditions at a temperature of about 20 ~ 30 ° C,
It is carried out for 50 to 180 hours, preferably by shaking culture or stirring culture using a rotary shaker, a jar fermenter or the like. When using a jar fermenter, an antifoaming agent such as polypropylene glycol may be added to the medium in advance.

Further, in the culture, if the operation of further adding methanol (and glycerin) before the methanol (and glycerin) added as the carbon source is completely consumed is repeated, the bacterial cells grow linearly. Subsequently, it is possible to increase the amount of dry cells in the culture solution to a high density of about 100 g / liter. The addition amount of methanol or a mixture of methanol and glycerin is preferably in the range of about 0.5 to 3% by weight based on the total weight of the medium.

The method for producing a trans-2-hexenal-containing composition of the present invention is carried out by the culture solution containing the cells of the Candida voidini SA051 strain obtained as described above, as it is, or by centrifugation from this culture solution. The suspension obtained by suspending the microbial cells obtained by separating by such means or a treated product thereof, and the trans-2-hexenol-containing composition are mixed, and the microbial cells are shaken or stirred. Alternatively, the treated product and the trans-2-hexenol-containing composition are brought into sufficient contact with each other.

The concentration of the cells in the reaction solution is preferably in the range of about 5 to 60 g / liter in terms of dry cells. Further, the concentration of the trans-2-hexenol-containing composition in the reaction solution is such that the concentration of trans-2-hexenol contained is about 5 to 150 g /
The concentration is about 10 liters and the total composition is about 10
It is preferably in the range of to 300 g / liter. The pH of the reaction solution is usually in the range of 6 to 10, preferably in the range of 7 to 9, and any buffer solution may be used as long as it is in such a range, and an alkali such as sodium hydroxide may be used. However, it is most preferable to use a phosphate buffer. The reaction temperature is usually in the range of about 4-30 ° C. The reaction is carried out under aerobic conditions, but may be carried out by injecting air or concentrated oxygen, preferably in a closed system under a pure oxygen atmosphere, or by injecting pure oxygen while applying pressure. . The reaction time is usually in the range of about 30 minutes to 2 hours, but it may vary depending on the component composition of the raw material composition. Therefore, the amount of trans-2-hexenal produced in the composition is sampled to obtain trans-hexenal. It may be determined while observing the conversion rate of 2-hexenol to trans-2-hexenal. However, if the reaction is carried out for a too long time, other alcohols may be oxidized, and unnecessary substances may be generated in the composition.

After completion of the reaction, known means, for example, hydrocarbons such as n-hexane, organic solvents such as esters such as ethyl acetate (the organic solvent may be used alone or in a mixture of 2
It can be used as a mixed solvent of one or more kinds. ), Concentrated, and distilled, most of the trans-2-hexenol in the starting material is selectively trans-converted.
The desired composition converted to 2-hexenal can be obtained. Since this composition has a strong and preferable green note, it can be used by being added to a perfume composition for the purpose of imparting a green note. In particular, when the obtained trans-2-hexenal-containing composition is used by blending it with a flavor composition, the content of trans-2-hexenol remaining in the trans-2-hexenal-containing composition is 25% by weight or less. Is preferable, and more preferably 20% by weight or less.

Furthermore, when the composition obtained by the production method of the present invention is separated into an aldehyde component and an alcohol component by means such as column chromatography, the aldehyde component has a purity of about 90 to 95% trans-2-. You can get hexenal. If a small amount of n-hexanal is removed by means such as further precision distillation,
Further, highly pure trans-2-hexenal can be obtained. This isolated trans-2-hexenal can also be used by blending it in the fragrance composition.

The alcohol content obtained by removing the aldehyde content as described above from the composition obtained by the production method of the present invention has a reduced content of trans-2-hexenol, which has an unpleasant aroma as a green note. Is
It is a component having a preferable green note in which the content ratio of cis-3-hexenol and n-hexanol having a preferable aroma is increased. This alcohol content can also be used by blending it with the fragrance composition. Further, the alcohol content of which the content ratio of trans-2-hexenol is reduced, based on the production method of the present invention, is repeatedly contacted with the cells of Candida voidini or a treated product thereof, and remains in the alcohol content. It is also possible to convert trans-2-hexenol into trans-2-hexenal to obtain a composition having a more preferable green note.

The perfume composition of the present invention is characterized by containing the trans-2-hexenal-containing composition obtained as described above. The fragrance composition of the present invention is a trans
-2- It can be obtained by mixing the composition containing hexenal as it is or with other commonly used compounded flavors, that is, synthetic flavors, natural flavors, essential oils and the like. The fragrance composition of the present invention can be added to foods and drinks, cosmetics and the like to enhance its commercial value.

Trans-2 to be incorporated into the fragrance composition of the present invention
-As the hexenal-containing composition, it is preferable to use a composition having a higher content ratio of trans-2-hexenal and a lower content ratio of trans-2-hexenol, specifically, as a main component, Containing about 20-45% by weight of trans-2-hexenal, 0-20% by weight of trans-2-hexenol, about 15-45% by weight of cis-3-hexenol, and about 15-30% by weight of n-hexanol. It is preferred to use a composition which The content ratio of the trans-2-hexenal-containing composition in the fragrance composition of the present invention is preferably in the range of about 0.01 to 1% by weight.

Further, the fragrance composition of the present invention includes, for example, synthetic fragrances and natural fragrances having a fruity aroma such as apple, strawberry, orange, grapefruit and grape; for example, almond oil, neroli oil and orris oil. , Essential oils such as rose oil, mandarin oil, orange essence, grapefruit essence, grape essence; for example, absolute products such as Gene Absolute, and processed products of essential oils such as terpeneless mandarin oil; Alcohol extracts of perfume raw materials such as tinctures and infusions such as strawberry infusion; alcohols such as ethyl alcohol and octanal; esters such as ethyl butyrate and ethyl isovalerate; fusel oil It can be formulated. The content ratio of these is not particularly limited and can be appropriately changed depending on the application.

The blending amount of the obtained flavor composition in foods and drinks, cosmetics and the like varies depending on the application and is not particularly limited. For example, when blending it in a beverage, It is preferable that the flavor composition is blended in the beverage in an amount of about 0.03 to 1% by weight.

The transformer obtained as described above
The trans-2-hexenal isolated from the 2-hexenal-containing composition can also be used as a flavor composition by blending it with the other compounded flavor and the like as described above. In this case, the blending ratio of trans-2-hexenal is preferably in the range of about 0.002 to 0.45% by weight with respect to the perfume composition.

In addition to trans-2-hexenol, the production method of the present invention can be widely applied to a method for producing trans-2-alkenal by oxidizing trans-2-alkenol. That is, a branched chain or linear chain trans-2-alkenol having about 4 to 10 carbon atoms is converted into a corresponding trans-2- by bringing it into contact with a bacterium of Candida voidini SA051 strain or a treated product thereof. It can be efficiently converted into alkenal. When trans-2-alkenol is present as a mixture with other alcohols having the same number of carbon atoms, the oxidation rate of trans-2-alkenol in the mixture is significantly higher than that of other alcohols, so other alcohols Before being oxidized
Trans-2-Alkenol is selectively oxidized to trans-
2-Alkenal can be obtained.

[0028]

EXAMPLES Next, the present invention will be specifically described by way of examples, but the present invention is not limited to these examples. Example 1 Preparation of cell suspension I Glucose 20 g, ammonium sulfate 3 g, potassium dihydrogen phosphate 4 g, magnesium sulfate 0.4 g, calcium chloride
10 mg, manganese chloride 2 mg, zinc sulfate 5 mg, biotin 0.05
mg and thiamin hydrochloride 5 mg were dissolved in ion-exchanged water 1000 ml, and an 8 wt% concentration sodium hydroxide aqueous solution was added to the resulting solution to adjust the pH to 6.0. Thus, the liquid medium was prepared. Next, 100 ml of the above liquid medium was added to a 500 ml capacity Sakaguchi flask with folds, the flask was placed in an autoclave and heated at 121 ° C. for 10 minutes for sterilization. Then, in the medium in the flask,
Apply the Bodini SA051 strain twice with a platinum loop, and use a rotary shaker under aeration at a rotation speed of 140 rpm and 28 ° C.
Culture was carried out with shaking for 48 hours to obtain a preculture liquid.

Then, a liquid medium having the same composition as the above liquid medium except that glucose was removed was prepared and sterilized in the same manner as above, and then 1.5 ml of methanol was added to 100 ml of the liquid medium, and the above preculture was added thereto. 5 ml of the liquid was added, and the mixture was shake-cultured at 140 rpm and 28 ° C. for 65 hours using a rotary shaker. The cells were separated from the obtained culture broth by centrifugation, washed with 0.1 M (= mol / liter) phosphate buffer of pH 7.5, and then the dry cells were adjusted to 60 g / liter.
Resuspended in 0.1M phosphate buffer. Thus, the bacterial cell suspension I was obtained.

Of the composition I containing trans-2-hexenol
Preparation Commercially available spearmint oil (manufactured by AM TODD) was distilled to obtain the first fraction [distillation temperature: 70-80 ° C (13-18mmHg)
, Ratio to spearmint oil: about 10% by weight) was collected and the fraction was again subjected to precision distillation [distillation temperature: 70-80 ° C (13-
18 mmHg)] to remove azeotropic contaminants during the first distillation. Subsequently, silica gel 60 (manufactured by Merck & Co., Inc.) as a stationary phase and n-hexane: ethyl acetate = 98: as a mobile phase.
What was eluted by column chromatography using the mixed solution of 2 was removed, and then the stationary phase was passed through a mixed solution of n-hexane: ethyl acetate = 80: 20 as a mobile phase to separate the eluted ones. The purified trans-2-hexenol-containing composition I was obtained by removing n-hexane and ethyl acetate in the collected eluate (yield based on spearmint oil: 0.08%). This trans-2-hexenol-containing composition I was trans-2-hexenol 32.
It contained 6% by weight, 45.3% by weight of cis-3-hexenol, 16.2% by weight of n-hexanol and other acyclic alcohol isomers having 6 carbon atoms.

Of the composition I containing trans-2-hexenal
Preparation 10 ml of the above-mentioned bacterial cell suspension I and 0.3 g of the above-mentioned trans-2-hexenol-containing composition I were placed in an Erlenmeyer flask having a capacity of 100 ml, the inside of the flask was replaced with pure oxygen and the flask was sealed, and then a reciprocal shaker was placed. It was shaken for 1 hour at 170 rpm and 25 ° C.

The obtained reaction solution was extracted with 10 ml of n-hexane, and then n-hexane was removed using a KD concentrator and a rotary evaporator. Thus, trans-2-hexenal 20.8% by weight, trans-2-hexenol 8.0% by weight, cis-3-hexenol 36.3% by weight, n-
Hexanol 15.8% by weight, n-hexanal 1.2% by weight
A trans-2-hexenal-containing composition I 0.
28 g (yield 93.3%) was obtained. This trans-2-hexenal containing composition I was strong and had a pleasing green note.

Example 2 Preparation of Cell Culture Solution II Glucose 1 was added to a jar fermenter having a volume of 70 liters.
20g, ammonium chloride 305.2g, potassium dihydrogen phosphate 112.4g, magnesium sulfate 23.6g, calcium chloride
2.2 g, ferric chloride 1.5 g, manganese sulfate 0.68 g, ethylenediaminetetraacetic acid disodium 18 g, zinc sulfate 0.88
g, copper sulfate 0.16 g, cobalt chloride 0.11 g, sodium molybdate 0.1 g, boric acid 0.16 g, potassium iodide 24 m
g, biotin 2 mg, thiamine hydrochloride 200 mg and polypropylene glycol 1.6 g were added, and 40 liters of ion-exchanged water was added and dissolved. The obtained solution was adjusted to pH 6.0 by adding an aqueous 8% by weight sodium hydroxide solution. In this way, a liquid medium was obtained. This liquid medium is stored at 121 ° C for 10
Sterilization was performed by heating for minutes. Subsequently, 800 ml of Candida voidini SA051 strain culture medium precultured in a liquid medium in the jar fermenter in the same manner as the preculture method of Example 1 in a liquid medium having the same composition as the liquid medium. After the addition, stirring culture was carried out at 28 rpm at 200 rpm under aeration of 20 l / min.

After 16 hours had elapsed from the start of the culture, the pH was adjusted to 5.0 by adding an aqueous 8 wt% sodium hydroxide solution, and then 400 ml of methanol was added to continue the culture while maintaining the pH at 5.0. Before the added methanol was completely digested, the addition of 1.2% by weight of methanol to the culture solution was repeated, and the culture was performed for 105 hours in total from the start of the culture. increased to g / l. The changes in the concentration of methanol and the amount of cells were examined over time. The result is shown in FIG. The cells were separated from the obtained culture broth by centrifugation, washed with 0.1 M phosphate buffer of pH 7.5, and then reconstituted with 0.1 M phosphate buffer to a dry cell volume of 60 g / liter. Suspended. Thus, the bacterial cell suspension II was obtained.

Of trans-2-hexenol containing composition II
Preparation Commercially available spearmint oil (manufactured by AM TODD) was distilled to obtain the first fraction [distillation temperature: 70-80 ° C (13-18mmHg)
, Ratio to spearmint oil: about 10% by weight] was collected, and the fraction was purified by the same method as in the preparation of the composition of trans-2-hexenol-containing composition I of Example 1.
Thus, trans-2 hexenol 50.8% by weight, cis-3-hexenol 19.9% by weight, n-hexanol 25.8% by weight and other acyclic alcohol isomers having 6 carbon atoms, trans-2 -Hexenol-containing composition II was obtained (yield to spearmint oil: 0.08
%).

Of the composition containing trans-2-hexenal II
Preparation 3 L of the above-mentioned bacterial cell suspension II and 150 g of the above trans-2-hexenol-containing composition II were placed in a jar having a volume of 5 L, and 400 r.pm, while supplying pure oxygen at a rate of 150 ml / min. The mixture was stirred at 25 ° C for 1 hour. The resulting reaction solution was steam distilled under normal pressure to obtain a fraction of 600 ml. 30 g of sodium chloride was added to this fraction and the mixture was allowed to stand and the upper layer was separated. Thus, trans-2-hexenal 31.5% by weight, trans-2-hexenol 19.1% by weight, cis-3-hexenol 1
141 g of trans-2-hexenal-containing composition II containing 6.3% by weight, 26.0% by weight of n-hexanol and 1.6% by weight of n-hexanal (yield 94.0%) was obtained. This trans-2-hexenal containing composition II was strong and had a favorable green note.

Isolation of trans-2-hexenal By column chromatography using 700 g of silica gel 60 (manufactured by Merck) as a stationary phase and 2 liters of a mixed liquid of n-hexane: ethyl acetate = 98: 2 as a mobile phase,
The aldehyde component contained in the trans-2-hexenal-containing composition II was separated together with the solvent. When the solvent was removed by distilling the solution containing the separated aldehyde, 93.0% by weight of trans-2-hexenal and n
Aldehyde content 45.0 containing 4.7% by weight of hexanal
g (yield 30.0%) was obtained. This component was very strong and had a pleasant green note. In addition, precision distillation of this component (distillation temperature: 45 ~ 55 ℃ (13 ~ 18mmHg)
By this, 40.0 g of trans-2-hexenal having a purity of 97% (yield 26.7%) was obtained.

Further, 700 g of silica gel 60 (manufactured by Merck & Co., Inc.) was used as it was as a stationary phase, and 2 g of a mixed liquid of n-hexane: ethyl acetate = 80: 20 was used as a mobile phase. The alcohol component contained in the component excluding the aldehyde component from the 2-hexenal-containing composition II was separated together with the solvent. When the solution containing the collected alcohol was distilled to remove the solvent, trans-2-hexenol 30.8% by weight, cis-
85.6 g of alcohol content (yield 57.1%) containing 26.2% by weight of 3-hexenol and 42.0% by weight of n-hexanol was obtained. This component had an improved green note over the starting trans-2-hexenol containing composition II.

Example 3 Preparation of Apple Flavor No. 1 Trans-2 obtained in Example 1 according to the formulation shown in Table 1.
-Apple Flavor No. 1 containing hexenal-containing composition I was prepared. Preparation of Apple Flavor No. 2 (Reference Example) According to the formulation shown in Table 1, apple flavor No. 2 blended with trans-2-hexenal having a purity of 97% obtained in Example 2 was prepared. Preparation of Apple Flavor No. 3 (Reference Example) According to the formulation shown in Table 1, the transformer in Example 2 was used.
-2- Apple-flavor No. 3 containing an alcohol content obtained by removing the aldehyde content from the hexenal-containing composition II was prepared. Preparation of Apple Flavor No. 4 (Comparative Example) According to the formulation shown in Table 1, an apple flavor containing the trans-2-hexenol-containing composition II used as the starting material in Example 2 was prepared.

[0040]

[Table 1]

Obtained Apple Flavor No. 1 to No. 4
Is added to commercially available fruitless apple juice and apple juice containing 10% fruit juice at a ratio of 500 ppm each, and a professional flavorist (creator of food spices. Genichi Indo "Actual knowledge of spices" (March 1975) 25th) Toyo Keizai Inc. 205
(See page), its aroma was evaluated, and the apple juice added with Apple Flavor No. 1 containing the trans-2-hexenal-containing composition I obtained by the production method of the present invention was Than apple juice to which apple flavor No. 4 containing the starting material trans-2-hexenol-containing composition II is added,
The characteristics of natural apples had a very pronounced odor.
Further, as a reference example, apple juice added with apple flavor No. 2 containing trans-2-hexenal, and apple juice added with apple flavor No. 3 containing an alcohol content excluding aldehyde content are also comparative examples. Compared with apple juice added with No. 4 apple flavor, the characteristics of natural apples had a distinctive aroma.

Example 4 Preparation of Strawberry Flavor A strawberry flavor containing the trans-2-hexenal-containing composition II obtained in Example 2 was prepared according to the following formulation. The strawberry flavor obtained had an aroma very characteristic of natural strawberry juice. (Prescription) Strawberry infusion: 99.9 parts by weight (Strawberry alcohol extract, manufactured by Takasago International Corporation) Trans-2-hexenal-containing composition II: 0.1 parts by weight

Example 5 Preparation of Flavor for Orange Juice A flavor for orange juice containing the trans-2-hexenal-containing composition II obtained in Example 2 was prepared according to the following formulation. The obtained orange juice flavor had an aroma very characteristic of natural orange juice. (Prescription) Natural orange essence: 99.9 parts by weight (essential oil, manufactured by Takasago International Corporation) Trans-2-hexenal-containing composition II: 0.1 parts by weight

Example 6 Preparation of Grapefruit Flavor A grapefruit flavor containing the trans-2-hexenal-containing composition II obtained in Example 2 was prepared according to the following formulation. The grapefruit flavor obtained had an aroma very characteristic of natural grapefruit juice. (Prescription) Natural grapefruit essence: 99.5 parts by weight (essential oil, manufactured by Takasago International Corporation) Natural octanal: 0.2 parts by weight (Octanol derived from natural products, manufactured by Takasago International Corporation) Natural Noot Katong: 0.2 parts by weight (natural products) Origin Nute Katong, manufactured by Takasago International Corporation) Trans-2-hexenal-containing composition II: 0.1 part by weight

Example 7 Preparation of Grape Flavor A grape flavor containing the trans-2-hexenal-containing composition II obtained in Example 2 was prepared according to the following formulation. The grape flavor obtained had an aroma very characteristic of natural grape juice. (Prescription) Natural grape essence: 99.3 parts by weight (essential oil, manufactured by Takasago International Corporation) Natural ethyl butyrate: 0.5 parts by weight (Ethyl acetate derived from natural products, manufactured by Takasago International Corporation) Terpenless mandarin oil: 0.1 parts by weight ( Essential oil excluding terpenes, manufactured by Takasago International Corporation) Trans-2-hexenal-containing composition II: 0.1 part by weight

Reference Example Trans-2-alkenol and other alcohol mixture having the same number of carbon atoms were used as Candida Boydini SA.
The selective oxidation of trans-2-alkenol was examined by contacting the strain with strain 051.

Experiment No. 1 bacterial cell was dried to 0.1 M so that the dry bacterial cell amount became 33 g / liter.
3 ml of bacterial cell suspension III prepared by the same method as the bacterial cell suspension I prepared in Example 1 except that it was suspended in a phosphate buffer.
And trans-2-hexenol and other C6 alcohol mixtures (trans-2-hexenol,
n-hexanol, cis-2-hexenol, trans-3-
90 mg of a mixture containing hexenol and cis-3-hexenol in equal weights) is mixed and reacted at 27 ° C., and aldehydes corresponding to the respective alcohols (trans-2
-Hexenal, n-hexanal, cis-2-hexenal, trans-3-hexenal, and cis-3-hexenal) were examined over time in the content concentration. The result is shown in FIG.

Experiment No. 2 As the alcohol mixture, trans-2-heptenol and the other alcohol mixture having 7 carbon atoms (mixture containing trans-2-heptenol and n-heptanol by the same weight) were used. The reaction was carried out in the same manner as in Experiment No. 1 to examine the change over time in the content concentration of the aldehydes (trans-2-heptenal and n-heptanal) corresponding to the respective alcohols in the reaction solution. The result is shown in FIG.

Experiment No. 3 except that trans-2-octenol and other C8-containing alcohol mixture (mixture containing trans-2-octenol and n-octanol by the same weight) were used as the alcohol mixture. The reaction was performed in the same manner as in Experiment No. 1 to examine the change over time in the content concentration of the aldehydes (trans-2-octenal and n-octanal) corresponding to the respective alcohols in the reaction solution. FIG. 4 shows the results. From these experiments, it was confirmed that trans-2-alkenol was selectively and significantly oxidized in both cases to produce trans-2-alkenal.

[0050]

According to the manufacturing method of the present invention, the transformer-2 is used.
-Trans-2-hexenol, which has an unpleasant odor as a green note in a hexenol-containing composition, is selectively oxidized to convert into trans-2-hexenal, whereby trans having a preferable green note is obtained.
-2-A hexenal-containing composition can be obtained. Also, trans-2-hexenal having a preferable green note can be obtained. Since the composition obtained by the method of the present invention has a preferable green note, a fragrance composition containing the composition can be added to foods and drinks and cosmetics to enhance its commercial value.

[Brief description of drawings]

FIG. 1 is a diagram showing a graph showing changes with time in methanol concentration and bacterial cell amount in Example 2.

FIG. 2 is a diagram showing a graph showing changes over time in the content concentration of aldehydes (trans-2-hexenal etc.) in a reaction solution.

FIG. 3 is a diagram showing a graph showing changes over time in the content concentration of an aldehyde (trans-2-heptenal etc.) in a reaction solution.

FIG. 4 is a diagram showing a graph showing changes over time in the concentration of aldehyde (trans-2-octenal etc.) contained in a reaction solution.

─────────────────────────────────────────────────── ─── Continuation of the front page (51) Int.Cl. ⁶ Identification code Office reference number FI technical display location C12R 1:72)

Claims (6)

[Claims] 1. A trans-2-hexenol-containing composition is contacted with Candida boidinii cells or a treated product thereof to convert the trans-2-hexenol into trans-2-hexenal. A method for producing a trans-2-hexenal-containing composition, comprising: 2. The trans-2-hexenal composition according to claim 1, wherein the trans-2-hexenol-containing composition contains trans-2-hexenol and other acyclic alcohol having 6 carbon atoms. The manufacturing method of a containing composition. 3. The method for producing a trans-2-hexenal-containing composition according to claim 1, wherein the trans-2-hexenol-containing composition is obtained by distilling mint oil. 4. The Candida bodini
The trans-2 according to any one of claims 1 to 3, wherein the microbial cell of idinii) or a treated product thereof is cultivated in a medium containing methanol or a mixture of methanol and glycerin as a carbon source. -Method for producing a composition containing hexenal. 5. Trans-2-hexenal is isolated from the trans-2-hexenal-containing composition obtained by the production method according to any one of claims 1 to 4. Method for producing hexenal. 6. A perfume composition comprising a trans-2-hexenal-containing composition obtained by the method according to any one of claims 1 to 4.