JPH09220037A - Method for producing patchouli mutant plant and modified patchouli oil - Google Patents

Abstract

(57) Abstract: A patchouli plant-derived callus or cell is obtained by subculturing to recover individual mutant cells, and the total content of patchouli alcohol and norpachorenol is 35. A patchouli mutant plant that produces essential oil in an amount of at least wt%, and a modified patchouli oil production method using the same. [Effect] A high-quality patchouli oil free of contaminating odor can be stably obtained.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a patchouli mutant plant which produces a modified patchouli oil useful as a perfume ingredient for perfumery and cosmetics, and a modified patchouli oil using the same, which has an excellent odor with no odor. Regarding manufacturing method.

[0002]

2. Description of the Related Art Patchouli ( Pogostemon cablin ) is a fragrance plant of the Labiatae family, and the essential oil (patchouli oil) obtained from the foliage has a woody and earthy odor that is unique to the cosmetic industry. Is one of the important plant essential oils. This patchouli oil is a multi-component mixed system centered on sesquiterpenes, but the main components that produce patchouli-like aroma are two types of sesquiterpene alcohols, patchouli alcohol and norpacholenol.

However, in patchouli oil obtained from natural patchouli, these two components account for 20 parts of the entire patchouli oil.
The sesquiterpene hydrocarbons, which occupy only about 30% by weight and are the main components other than the sesquiterpene alcohol, are odorless or mere odor. Therefore, various refining means have been tried in order to obtain high-quality patchouli oil with a high ratio of patchouli alcohol and norpachorenol, but it is still industrially adopted because of its close physical properties or production cost. No purification method has been found.

It is also possible to produce new patchouli plants that produce such high-quality patchouli oil. However, patchouli plants are sterile, so breeding by conventional breeding methods such as crossbreeding is impossible. is there.

[0005]

SUMMARY OF THE INVENTION Therefore, an object of the present invention is to provide a plant producing high-quality patchouli oil having an excellent odor without contaminated odor, and a method for producing high-quality patchouli oil using the plant. is there.

[0006]

[Means for Solving the Problems] Therefore, the present inventors created a mutant using cell culture technology and modified patchouli oil components, and as a result, the contents of patchouli alcohol and norpachorenol were high and excellent. We succeeded in breeding a mutant plant that produces essential oil having aroma, and completed the present invention.

That is, the present invention is obtained by subculturing callus or cells derived from patchouli plants to restore individual cells from mutant cells induced, and the total content of patchouli alcohol and norpachorenol is 35% by weight. It is intended to provide a patchouli mutant plant that produces the above essential oil and a method for producing a modified patchouli oil using the mutant plant.

[0008]

BEST MODE FOR CARRYING OUT THE INVENTION The mutant plant of the present invention is characterized by producing an essential oil (modified patchouli oil) having a total content of patchouli alcohol and norpachorenol of 35% by weight or more, and other properties It may be almost the same as natural patchouli, or may have some morphological changes such as leaf tip winding and leaf shape rounding. Such a mutant plant of the present invention can be obtained by subculturing a callus or cell derived from a natural patchouli plant and individually reconstituting the induced mutant cell.

More specifically, somatic mutations are induced by subculture of callus or cultured cells of natural patchouli plants in the presence of plant hormones for a long period of time, and plants are sequentially reconstructed from the obtained cultured mutants. And then grown to a sufficient size.

To explain the above method in more detail, first,
Explants such as leaves and stems of patchouli plants are placed on a callus induction medium. As a callus induction medium, saccharides,
Examples include those to which a growth regulator, pH stabilizer, gelling agent and the like have been added. As the basic medium used, any medium commonly used in plant tissue culture can be applied, and for example, a medium such as MS, LS, B5, White or the like can be used. Particularly preferred as a basal medium is LS medium. In addition, examples of the saccharides added to the medium include sucrose and glucose, and the concentration thereof is 0.1 to 10
%, Especially 0.5 to 5% by weight is preferred. Further, it is preferable to add various plant hormones as growth regulators. Plant hormones that can be used include auxins,
Cytokinins are used. As auxins, 2,4-dichlorophenoxyacetic acid (hereinafter abbreviated as 2,4-D), 1-naphthalene acetic acid (hereinafter abbreviated as NAA),
Indole-3-acetic acid (hereinafter abbreviated as IAA), indole-3-butyric acid (hereinafter abbreviated as IBA) and the like,
Examples of cytokinins include 6-benzyladenine (hereinafter abbreviated as BA) and kinetin (hereinafter KIN).
Abbreviated), zeatin (hereinafter abbreviated as ZEA), and the like. One or more kinds of these plant hormones are usually added, but the concentration of auxins is preferably 0.01 to 40 ppm, particularly preferably 0.02 to 10 ppm, and the cytokinins are 0.01 to 20 ppm, particularly 0.02 to 10 ppm. Is preferred. The pH of the medium is 4.0 to 7.0, especially 5.5 to 6.
0 is preferred. 2- (N-morpholino) ethanesulfonic acid (hereinafter abbreviated as MES) or the like is used as a pH stabilizer.
1 wt% may be appropriately added. It is preferable to use a solid medium in which agar, agarose, gellite or the like is added to these as a gelling agent. Callus can be obtained by placing patchouli explants on this DNA doubling induction medium and aseptically culturing under conditions of low light or darkness of 0 to 500 lux and temperature of 20 to 30 ° C., for example.

Further, although callus is transplanted to a liquid medium and shake-cultured, cultured cells containing free cells and small cell clusters can be obtained. In the present invention, callus is preferably used.

The callus obtained as described above is further subcultured in the above-mentioned medium to induce somatic mutation. A mutagen such as methylethanesulfonic acid may be added to the medium in order to positively increase the mutation frequency. Also, the mutation frequency increases even when cultured in a medium having a high plant hormone concentration (for example, 2,4-D is 100 to 400 ppm or NAA is 40 to 80 ppm). The subculture for mutagenesis is preferably performed 3 times or more. Culture conditions are preferably, for example, 20 to 30 ° C. under low light or darkness of 0 to 500 Lux.

Restoration of the obtained mutant cells into a plant individual causes foliage or root differentiation by reducing the auxin concentration or transplanting into a hormone-free medium.

[0014] Among the restoration processes of the plant individual, those that have lost the ability to reproduce the individual are culled in the process of regeneration.

The plant individual thus reconstructed is further culled by a pot test outside a test tube, and a strain having low viability and a strain having no environmental adaptability are removed. This pot test
For example, it is performed in a phytotron or a glass chamber kept at 20 to 30 ° C. and a humidity of 60% or more, and planted in vermiculite or the like. Select a strain that survives after operations such as lowering the humidity.

Furthermore, it is desirable to select an optimum strain by growing these patchouli mutant plants having environmental adaptability in the field by means such as cuttings and examining the essential oil composition by gas chromatography or the like.

To produce a modified patchouli oil from the patchouli mutant plant thus obtained, for example, the harvested foliage may be dried and steam-distilled by a conventional method. The modified patchouli oil extracted from the patchouli mutant plant of the present invention has a total content of patchouli alcohol and norpacholenol of 3
Since it accounts for 5% by weight or more, it has an excellent strong aroma with less contaminating odor. In addition, compared to the high contents of these two components, the content of δ-guaien, which is a major contaminant, is not more than 3% by weight or is undetectable.

[0018]

The present invention will be described in more detail with reference to examples, but the present invention is not limited to these examples.

Example 1 (1) Callus Culture Pogostemon calbin cultural
r I (Sugimura et al, Flavou
r and Fragrance Journal
5: 109-114 (1990)) was sterilized in a 2% sodium hypochlorite solution for 10 minutes and washed with sterile water. These leaf pieces were placed in LS basal medium in a solid medium in which sucrose was 1% as a saccharide, 2,4-D 2 ppm as a plant hormone, gellite was 0.18% as a gelling agent, and the pH was adjusted to 6 before autoclaving. 26 ° C in the dark
A large number of callus was induced after 3 days to 2 weeks when left still for 3 days.

(2) Subculture of callus The obtained callus was further subcultured every 3 weeks in the above-mentioned medium for 5 weeks.
Subculture was performed.

(3) Restoration of individual plant The callus that was repeatedly subcultured was replaced with the above medium, and sucrose was 1%, NAA was 0.2 ppm as a plant hormone, and BA was used.
Transfer to a medium containing 0.2 ppm of gellite 0.2% as a gelling agent and adjusting the pH to 6 before autoclaving.
A large number of adventitious buds were differentiated by culturing under the conditions of lux and 26 ° C for 2 to 8 days. Those shoots that have grown to 2 to 4 cm are mowed and LS containing no plant hormone as a medium is cut off.
It was cut to a medium. When cultured at 26 ° C. under 3000 lux light illumination, rooting was observed one after another within 3 to 14 days, and the plant was regenerated into a complete plant. Further cultivated in a test tube, the plant height is 10
The cells were cultured until they reached about cm.

(4) Selection of Mutant Plants Plants sufficiently grown in a test tube were transplanted together with roots into a pot containing vermiculite and acclimated and cultured in Phytotron (3000 lux, 27 ° C., humidity 70).
%). At the end of acclimation, reduce the humidity to 40%,
The surviving strains were grown in the field in the field.

The foliage of the resulting patchouli mutant plant was collected, dried and steam-distilled (under normal pressure, 5 to dry foliage).
The upper layer obtained by passing 0% steam was used as an essential oil fraction. The obtained essential oil was analyzed by gas chromatography. Table 2 shows the results. The gas chromatography conditions are as shown below.

[0024]

[Table 1]

[0025]

[Table 2]

Further, when the aroma of patchouli oil obtained from the mutant strain of the present invention was evaluated by a specialized panelist, it had woody, earthy, and herbal aromas, had less blue odor than conventional patchouli oil, and had aroma. It was strong.

Further, when the viability of the mutant strain of the present invention after 3 months of cutting was examined, it was the same as that of the normal variety as shown in Table 3.

[0028]

[Table 3]

[0029]

EFFECTS OF THE INVENTION According to the present invention, patchouli oil having an excellent strong scent without any odor such as blue odor can be stably produced.

Claims (3)

[Claims] 1. Essential oil obtained by subculturing patchouli plant-derived callus or cells by individual restoration from mutant cells induced and having a total content of patchouli alcohol and norpacholenol of 35% by weight or more. A patchouli mutant plant that produces. 2. The patchouli mutant plant according to claim 1, wherein the δ-guaien content in the essential oil is 3% by weight or less. 3. A method for producing a modified patchouli oil having a total content of patchouli alcohol and norpacholenol of 35% by weight or more, which is extracted from the patchouli mutant plant according to claim 1.