JP2012213358A - Production method of anthocyanin - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a production method of anthocyanin which safely and easily increase the anthocyanin in a plant body.SOLUTION: There is provided the production method of the anthocyanin which is characterized in that an anthocyanin-producing plant is an exposed to green leaf volatile component, and the anthocyanin-producing plant is exposed to methyl jasmonate. There is also provided the anthocyanin-producing plant. In the production method anthocyanin, the green leaf volatile component is one of n-hexanol, Z-3-hexenol, Z-3-hexenal, E-2-hexenol, n-hexyl acetate, Z-3-hexenyl acetate, n-hexanal, E-2-hexanal, n-nonanol and n-nonanal.

Description

  The present invention relates to a method for producing anthocyanins present in plants such as fruits and vegetables.

  Anthocyanins are glycoside components in which anthocyanidins are linked to sugars and sugar chains, among anthocyans, which are widely present in the plant world. This anthocyanin has been used as a coloring agent for various foods for a long time because it is relatively safe even when eating and drinking and has a natural color. Anthocyanins are said to have a wide range of effects from vision improvement to antioxidant effects, and those extracted from plants are added to various health foods.

  As a method for producing the anthocyanin, a method of extracting and obtaining anthocyanin from an anthocyanin-containing plant cultivated by sowing has been conventionally performed. However, since anthocyanins are produced only from specific parts of specific plants and have seasonality and regional characteristics depending on the plants, it has been difficult to secure the required amount in the market.

  Thus, attempts have been made to produce plants with high anthocyanin content for the purpose of efficiently obtaining anthocyanins. As a method for producing a plant containing a high amount of anthocyanin, for example, γ-rays are applied to seeds of at least one plant selected from the group consisting of red radish, purple corn, perilla and red cabbage and sown. A method for increasing anthocyanins produced in plants by cultivation is known (for example, see Patent Document 1 and Claim 1). However, the method for producing a plant containing a high amount of anthocyanin in Patent Document 1 irradiates the seeds of the plant with γ rays, which may damage the plant gene and is dangerous.

  Moreover, as a method for producing a plant containing a high amount of anthocyanins, for example, seeding root germs containing anthocyanins in the hypocotyl or root, or seeds collected in the following step 4 to germinate and grow, Cut off the hypocotyl or part of the root part of the root vegetable obtained in Step 1 and measure the amount of anthocyanin contained in the slice; and select the root vegetable derived from the slice with a high anthocyanin content and grow it again. Step 3 for growing until the flowering period, and flowering root vegetables obtained in Step 3 are crossed or pollinated between different individuals, and seeds are collected, and the seeds obtained in Step 4 are collected, Or what has the process 5 which is provided to the said process 1, and the process of sowing and germinating the seed obtained at process 5 is known (for example, patent documents 2) Irradiation).

  However, the method for producing a plant containing a high amount of anthocyanin described in Patent Document 2 is to measure the amount of anthocyanin contained in the root vegetable cultivated by sowing, select a root vegetable having a high anthocyanin content, grow it again, and flowering period. It is necessary to cultivate twice since seeds are collected by mating different roots with different individuals and seeded, and the seeds obtained are further sown and cultivated. For this reason, the method for producing a plant containing a high amount of anthocyanins described in Patent Document 2 has a problem that it takes time and effort.

JP 2006-166788 A JP 2007-06092 A

  Then, the subject of this invention is providing the production method of the anthocyanin which increases the anthocyanin in a plant body safely and easily.

  The invention according to claim 1 has been made to solve the above-mentioned problems of the prior art, wherein an anthocyanin-producing plant is exposed to a green leaf volatile component, and the anthocyanin-producing plant is exposed to methyl jasmonate. The present invention relates to a characteristic method for producing anthocyanins.

  The invention according to claim 2 relates to the method for producing anthocyanins according to claim 1, wherein the anthocyanin-producing plant is Arabidopsis thaliana.

  The invention according to claim 3 is characterized in that the green leaf volatile component is n-hexanol, Z-3-hexenol, Z-3-hexenal, E-2-hexenol, n-hexyl acetate, Z-3-acetic acid hexenyl, n-hexanal. The method for producing anthocyanins according to claim 1, wherein the method is any one of E-2-hexenal, n-nonanol, and n-nonanal.

  The invention according to claim 4 relates to a method for producing anthocyanins according to claim 1 or 2, wherein the green leaf volatile component has an aldehyde group.

  The invention according to claim 5 relates to the method for producing anthocyanins according to claim 4, wherein the green leaf volatile component is n-hexanal or E-2-hexenal.

  The invention according to claim 6 relates to an anthocyanin-rich plant obtained by exposing an anthocyanin-producing plant to a green leaf volatile component secreted from the plant and exposing the anthocyanin-producing plant to methyl jasmonate.

  According to the invention according to claim 1, since the anthocyanin-producing plant is exposed to the green leaf volatile component, and the anthocyanin-producing plant is exposed to methyl jasmonate, the anthocyanin in the plant body increases. For the purpose of increasing the anthocyanins, it is not necessary to irradiate the seeds of the plants with γ rays or to cultivate them twice. Therefore, the anthocyanins in the plant can be increased safely and easily, and the required amount in the market can be easily secured by taking out the anthocyanins from the increased anthocyanin-producing plants.

  According to the invention which concerns on Claim 2, since the said anthocyanin production plant is Arabidopsis thaliana Col-0, the anthocyanin in a plant body can be increased further easily. This is because Arabidopsis thaliana (L.) Heynh. Is distributed all over the world, and it has the advantage that it takes a short time to sow seeds, bloom, and harvest the seeds.

  According to the invention of claim 3, the green leaf volatile component is n-hexanol, Z-3-hexenol, Z-3-hexenal, E-2-hexenol, n-hexyl acetate, Z-3-hexenyl acetate, n -The increase rate of the anthocyanin in a plant body can be improved by being either hexanal, E-2-hexenal, n-nonanol, or n-nonanal.

  According to the invention which concerns on Claim 4, when the said green leaf volatile component has an aldehyde group, the increase rate of the anthocyanin in a plant body can be improved more.

  According to the invention which concerns on Claim 5, the increase rate of the anthocyanin in a plant body can further be improved because a green leaf volatile component is n-hexanal and E-2-hexenal.

  According to the invention of claim 6, since an anthocyanin-producing plant is exposed to green leaf volatile components, and this anthocyanin-producing plant is exposed to methyl jasmonate, an anthocyanin-rich plant with increased anthocyanins in the plant body can be obtained. For the purpose of increasing the anthocyanins in the plant body as in the prior art, it is not necessary to irradiate the seeds of the plants with γ rays or to cultivate them twice. Therefore, an anthocyanin-rich plant can be obtained safely and easily. By taking out anthocyanin from this anthocyanin-rich plant, the required amount in the market can be easily secured.

The figure which shows the effect of this invention

  Hereinafter, the anthocyanin production method and the anthocyanin-rich plant according to the present invention will be described in detail.

  The method for producing anthocyanins according to the present invention is to increase anthocyanins in plants safely and easily by exposing anthocyanin-producing plants to green leaf volatile components and exposing the anthocyanin-producing plants to methyl jasmonate. It is.

  As the anthocyanin-producing plant, a plant cultivated with plant cells such as those sown and cultivated or callus producing anthocyanin may be used. As anthocyanin-producing plants, red radish (Perilla frutescens (L.) Britton var. Crispa (Thunb.) H. Deane f. Purpurea (Makino) Makino), red cabbage (Brassica oleracea L. var. Capitata L), red radish (Raphanus sativus L. var. Sativus), black beans (Glycine max (L.) Merr. Subsp. Max), black currant (Ribes nigrum L.), purple corn (Zea mays L.), purple sweet potato (Ipomoea batatas L.) , Grapes (Vitis vinifera L.), blueberries (Vaccinium spp.), Arabidopsis thaliana (L.) Heynh., Any plant that produces anthocyanins in the plant body may be used.

  Akajiso (Perilla frutescens (L.) Britton var. Crispa (Thunb.) H. Deane f. Red cabbage (Brassica oleracea L. var. Capitata L) is a Brassicaceae, Brassica genus, and contains mainly anthocyanins in the leaves. Japanese radish (Raphanus sativus L. var. Sativus) is a Brassicaceae and radish genus, and contains mainly anthocyanins in the roots. Black beans (Glycine max (L.) Merr. Subsp. Max) are legumes and soybeans, and mainly contain anthocyanins in the outer skin. Black gooseberry (Ribes nigrum L.) is a family of gourdaceae and gooseberry, and contains mainly anthocyanins in fruits.

  Purple corn (Zea mays L.) is a genus of the family Gramineae and corn, and mainly contains a flower shaft and anthocyanins. Purple sweet potato (Ipomoea batatas L.) is a convolvulaceae family and a sweet potato genus, and anthocyanins are mainly contained in tuberous roots. Grapes (Vitis vinifera L.) are from the vine family, grape genus, and mainly contain anthocyanins in the pericarp. Blueberry (Vaccinium spp.) Is an azalea family and the genus Vaccinium, and anthocyanins are mainly contained in fruits and leaves. Arabidopsis thaliana (L.) Heynh. Is a member of the Brassicaceae family, Arabidopsis, and contains anthocyanins mainly in leaves and stems.

  Among the anthocyanin-producing plants, Arabidopsis thaliana is distributed all over the world, and has the advantage that it takes a short time until seeds are sown and flowers bloom and the seeds are harvested, so that the object of the present invention is achieved. Most preferred above.

  Anthocyanins are glycoside components in which anthocyanidins are linked to sugars and sugar chains, among anthocyans, which are widely present in the plant world. This anthocyanin is useful for expressing the color of flowers and fruits, is a kind of flavonoid, and is known as an antioxidant. Examples of anthocyanidins include pelargonidin, cyanidin, delphinidin, aurantidine, luteolinidine, peonidin, malvidin, petunidin, europeinidine, rosinidine and the like having a chemical structure represented by the following formula (I). The bonding positions of the sugar chains are mainly the hydroxy group at the 3-position of the bicyclic structure on the left side of the structural formula and the 5-position of the bicyclic structure on the left side of the structural formula.

  The green leaf volatile component is carbon 6 or carbon that is secreted through oxidative degradation from 18 fatty acids such as α-linolenic acid and linoleic acid in the chloroplast membrane when the plant is damaged by insects. A general term for volatile compounds of formula 9, for example, n-hexanol (molecular weight 102.17) (Chemical formula 2), Z-3-hexenol (molecular weight 100.16) (Chemical formula 3), Z-3-hexenal (molecular weight) 98.14) (Chemical formula 4), E-2-hexenol (molecular weight 100.16) (Chemical formula 5), n-hexyl acetate (molecular weight 144.21) (Chemical formula 6), Z-3-hexenyl acetate (molecular weight 142. 2) (Chemical formula 7), n-hexanal (molecular weight 100.16) (Chemical formula 8), E-2-hexenal (molecular weight 98.14) (Chemical formula 9), n-nonanol (molecular weight 144.25) (Chemical formula 10) , N-nonal (min The amount 142.24) (Formula 11) include, but are not limited to. The structural formulas of these green leaf volatile components are shown in the order described above.

  Methyl jasmonate is a physiologically active substance derived from jasmonic acid that is secreted from α-linolenic acid in the chloroplast membrane through oxidative degradation and cyclization processes when plants are damaged by insects. It plays an important role in defense responses against insect damage. The structural formula of methyl jasmonate (molecular weight 224.29) (Chemical Formula 12) is shown below.

  Here, the priming process which exposes an anthocyanin production plant to a green leaf volatile component is demonstrated. The priming treatment can be performed, for example, by a method in which a green leaf volatile component is allowed to flow into the space in a state where an anthocyanin-producing plant is placed in a closed or semi-closed space such as a plant factory. Any method may be used as long as the anthocyanin-producing plant can be exposed to the green leaf volatile component.

  Next, methyl jasmonate treatment in which an anthocyanin-producing plant is exposed to methyl jasmonate will be described. The methyl jasmonate treatment can be performed by the same method as the priming treatment, but is not limited thereto, and any method can be used as long as the anthocyanin-producing plant can be exposed to methyl jasmonate. Good.

  Anthocyanins can be extracted from the plant body by extraction, for example, but are not limited thereto. As the extraction, for example, solvent extraction or supercritical extraction can be used. In order to increase the extraction efficiency of anthocyanin, it is preferable to use a grated state, a pulverized state, or a squeezed juice.

  The anthocyanin-rich plant according to the present invention is obtained by exposing an anthocyanin-producing plant to a green leaf volatile component and exposing the anthocyanin-producing plant to methyl jasmonate.

  Hereinafter, the effect of the present invention will be clarified by showing examples relating to the anthocyanin production method according to the present invention. However, the present invention is not limited to the following examples.

(1) Seeding culture First, Arabidopsis seeds are sterilized with 5% hypochlorous acid, then seeds of Arabidopsis seeds are sown on 1 / 2MS agar medium, and vernalization is performed in a dark place at 4 ° C for 2 days. . Next, seeds are cultivated while performing an alternating operation of light period 16h and dark period 8h for 9 days in a state where the artificial weather device is maintained at 23 ° C. At this time, it was confirmed that seeds germinated after 2 days. Thereafter, the agar medium is transplanted into a cylindrical glass tube having an inner diameter of 3.4 cm and a height of 12.5 cm.

(2) Priming treatment and methyl jasmonate treatment Next, a priming treatment is performed in which Arabidopsis thus cultivated is exposed to volatile components of green leaves. The priming process is performed by closing the upper end of the glass tube with cotton and impregnating the cotton with a volatile component of green leaves dissolved in methanol. Hereinafter, each example will be described in detail.

Example 1
The seed-cultured Arabidopsis thaliana was exposed to n-hexanol by impregnating 50 μl of n-hexanol dissolved in methanol to 1 mM with the cotton at the upper end of the glass tube. This state was maintained for 7 days. At this time, the initial concentration of n-hexanol in the glass tube was 0.5 μM. Next, Arabidopsis thaliana after the priming treatment was exposed to methyl jasmonate by impregnating 50 μl of methyl jasmonate dissolved in methanol to 1 mM with cotton at the upper end of the glass tube, and this state was maintained for 7 days ( Treatment with methyl jasmonate). At this time, the initial concentration of E-2-hexenal in the glass tube was 0.5 μM.

(Example 2)
In the same manner as in Example 1, Z-3-hexenol was subjected to priming treatment and methyl jasmonate treatment.
(Example 3)
In the same manner as in Example 1, n-hexanal was subjected to priming treatment and methyl jasmonate treatment.
Example 4
In the same manner as in Example 1, E-2-hexenal was subjected to priming treatment and methyl jasmonate treatment.
(Comparative Example 1)
The seeded Arabidopsis thaliana was exposed to methanol only by impregnating 50 μl of methanol only with cotton at the upper end of the glass tube. This state was maintained for 7 days. Next, methyl jasmonate treatment was performed in the same manner as in Example 1.

(Comparative Example 2)
In place of the methyl jasmonate treatment performed in Examples 1 to 4 and Comparative Example 1, Arabidopsis thaliana after the priming treatment was exposed to methanol alone by impregnating 50 μl of methanol alone into the cotton at the upper end of the glass tube.

(3) Measurement of anthocyanin content Next, the above ground fresh weight of each of Examples 1 to 4 and Comparative Examples 1 and 2 was immersed in a 1% HCl-MeOH aqueous solution to obtain anthocyanins from the fresh ground weight of Arabidopsis thaliana. And the amount of the extracted anthocyanins was measured by spectroscopic analysis. The results are shown in FIG.

  As shown in FIG. 1, it can be seen that the amount of anthocyanin extracted in Examples 1 to 4 is higher than the amount of anthocyanin extracted in Comparative Example 2. Therefore, it was confirmed that when Arabidopsis thaliana was exposed to green leaf volatile components and then exposed to methyl jasmonate, anthocyanins in the plant were increased. For this reason, by exposing an anthocyanin-producing plant to a green leaf volatile component and exposing this anthocyanin-producing plant to methyl jasmonate, a plant with high anthocyanin content in which the anthocyanin in the plant body is increased can be obtained.

  In addition, among Examples 1 to 4, it is confirmed that Examples 3 and 4 have a large anthocyanin increase rate. It is considered that n-hexanal and E-2-hexenal have an aldehyde group that is more easily involved in the redox reaction than the hydroxy group.

Claims (6)

  An anthocyanin-producing plant is exposed to a green leaf volatile component, and the anthocyanin-producing plant is exposed to methyl jasmonate.   The method for producing anthocyanins according to claim 1, wherein the anthocyanin-producing plant is Arabidopsis thaliana (L.) Heynh.   The green leaf volatile component is n-hexanol, Z-3-hexenol, Z-3-hexenal, E-2-hexenol, n-hexyl acetate, Z-3-hexenyl acetate, n-hexanal, E-2-hexenal, n The method for producing anthocyanins according to claim 1 or 2, wherein the method is either nonanol or n-nonal.   The method for producing anthocyanins according to claim 1 or 2, wherein the green leaf volatile component has an aldehyde group.   The method for producing anthocyanins according to claim 4, wherein the green leaf volatile component is n-hexanal or E-2-hexenal.   An anthocyanin-rich plant obtained by exposing an anthocyanin-producing plant to green leaf volatile components and exposing the anthocyanin-producing plant to methyl jasmonate.

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