JP3130381B2 - Tissue culture method of Saffron Mesbe - 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 effectively cultivating a pigment which is one of the active ingredients in edible and medicinal saffron.

[0002]

2. Description of the Related Art Saffron is a perennial herb of the family Iridaceae, Crocus satius L .; (Crocus sativus L.)
The dried stigma is used as a crude drug. Spain,
It is produced in Hiroshima, Kagawa, Oita and Iwate prefectures in France, Italy and Japan. It contains about 2% of carotenoid pigment, crocin (glycoside of crocetin), and bitter glycoside (picrocrocid).
n) About 2%, containing 0.4-1.3% of essential oils. The main component of the essential oil is safranal (fragrance component peculiar to crude drugs).

[0003] This saffron is used as a spice, food coloring, or as a medicinal product. However, since it uses a dried product of mesibet, it cannot be obtained in a natural product.
The unit price is also very high. In addition, natural products are hard to obtain products of a certain quality due to climate change. Therefore, various attempts have been made for tissue culture.

[0004] For example, Japanese Patent Application Laid-Open No. 62-275617 discloses Crocus-Satius L. That is, a method is disclosed in which a saffron pistil or an organ corresponding thereto is grown and hypertrophically matured using cytokinin or an artificial culture medium containing cytokinin and auxin. Cytokinin has been known for a long time as a substance that induces cell division of cultured plant cells, and it is also known that callus induction and cell growth are promoted when used in combination with auxin.

[0005] JP-A-63-109788 discloses Crocus-Satius L. Of the female organ of the present invention, cultured in an artificial culture medium containing cytokinin, and selected from the group consisting of safranal, picrocrosin and analogs thereof, crocin, crocetin and analogs thereof
It produces more than one kind of substance. However, it has long been known that saffron pistils contain these substances.

JP-A-63-160580 discloses a method of culturing callus of a plant belonging to the genus Crocus in an MS medium supplemented with plant hormones. The use of plant hormones such as auxin and cytokinin for culturing plant tissues has been well known for a long time, and Murashige-Skoog medium is also a commonly used medium.

[0007] JP-A-63-258574 discloses that the stigma, style, ovary, ovule, and petals of saffron are excised and cut and placed in a LS medium or a B5 medium supplemented with cytokinin and auxin. It is transplanted and subcultured. Extraction, cutting and cultivation of plant parts have been performed for a long time, and plant hormones such as cytokinin and auxin are usually used, and Linsmeier Skoog's medium and Gamborg's medium are also commonly used media.

Tests of any of these methods do not significantly increase the rate of growth of the active ingredient of saffron and do not significantly increase the production of the active ingredient.

[0009]

SUMMARY OF THE INVENTION An object of the present invention is to provide a method for more efficiently obtaining pigments of crocin and crocetin, one of the active ingredients of saffron.

[0010]

Means for Solving the Problems The present inventors have conducted intensive studies to solve the above-mentioned problems, and as a result, the present invention has been achieved by adding a special additive to a conventionally known medium and / or additive. The present invention has been completed by finding out what can be done.
That is, the present invention relates to salicylic acid, salicylic acid ester,
A tissue culture method for saffron messibe, characterized by culturing using a medium containing at least one selected from the group consisting of benzoic acid, benzoic acid esters, and acetylsalicylic acid.

At least one compound selected from the group consisting of salicylic acid, salicylic acid ester, benzoic acid, benzoic acid ester and acetylsalicylic acid has a good result when 1 to 1000 ppm, preferably 10 to 100 ppm is added to the medium. Was obtained.

The medium used for these is not particularly limited, but is Murashige and Skoog's medium, White's medium, Gamborg's medium, Nitsch's medium, Heller's medium. ) Medium, Schenk and Hildebra
ndt), Nitsch and Nitsc
h) Medium, Kohlenback and Schmidt
and Schmidt's medium, Knop's medium, Linsmaier and Skoog's medium, and the like.

If necessary, cytokinins and auxins known as plant growth regulators may be added to these media. Examples of cytokinins include benzyladenine, zeatin, kinetin, etc., which vary depending on the type.
Add 1 to 20 ppm. Examples of the auxin include naphthalene acetic acid, indole acetic acid, 2,4-dichlorophenoxy acetic acid, and the like. The auxin varies depending on the type, but when added, 0.01 to 20 ppm is added.

[0014] If necessary, other medium-forming agents can also be added. In addition, the cells are cultured in a dark place at a temperature of 20 to 25 ° C. It was found that large amounts of pigment could be produced by inoculating mesibes at appropriate times and culturing under these conditions.

[0015]

The present invention will be described in more detail with reference to the following examples, but the present invention is not limited to these examples. (Examples 1 to 14) (Comparative Example 1) Peel off the outer skin of a saffron bulb with flower buds extending to about 10 cm
After removing the discolored portion, the plate was washed with running water for 3 hours. 1
3-4 seconds in 0% benzalkonium chloride, 1-2 seconds in 70% ethanol, 20-40 in 5% sodium hypochlorite
Minutes. After that, it was performed aseptically. The flower buds were cut off leaving the bulbs, and washed three times with sterile water. Next, the buds were taken out from the flower buds, incised, and the mesibes were taken out with the style long, and inoculated on the medium. The medium was Murashige and Skoog's medium, 4% saccharose, 0.1% agar powder.
9% was added to adjust the pH to 5.8. To this was added 10 ppm of naphthalene acetic acid and 1 ppm of benzyl adenine. The additives shown in Table 1 were further added thereto. Table 2 shows the culture results. After culturing, the whole was cut into small pieces and dried at 80 ° C for 1 hour.
1% with 50 ml of 4% sodium hydroxide 50% ethanol solution
After stirring in a dark place for 8 hours, take 10 ml of this solution and add 1 ml of phosphoric acid.
And the volume was adjusted to 50 ml with ethanol. Add this solution to 0.
After filtration through a 45 μm membrane, the absorbance at 425 nm was measured.

[0016]

[Table 1]

[0017]

[Table 2]

[0018]

According to the present invention, at least one selected from the group consisting of salicylic acid, salicylic acid ester, benzoic acid, benzoic acid ester and acetylsalicylic acid is added to the culture medium of saffron radix, so that it is not added as shown in the examples. The amount of the pigment was greatly increased as compared with the case.

────────────────────────────────────────────────── ─── Continued on the front page (58) Field surveyed (Int. Cl. ⁷ , DB name) C12N 5/00 A01H 4/00 A23L 1/275 C12P 7/44 BIOSIS (DIALOG) JICST file (JOIS) WPI ( DIALOG)

Claims (1)

(57) [Claims] 1. A method for cultivating saffron messibe, comprising culturing using a medium containing at least one selected from the group consisting of salicylic acid, salicylic acid ester, benzoic acid, benzoic acid ester, and acetylsalicylic acid.