JPS60109536A - Production of polyprenyl compound or mixture thereof - Google Patents

Abstract

PURPOSE:To separate the titled substance useful as an intermediate for producing mammalian dolichol from an extract, by extracting a leaf of a plant belonging to the genus Picea or Abies of the family Pinaceae with an organic solvent, and obtaining the extract. CONSTITUTION:A leaf of a plant belonging to the genus Picea or Abies of the family Pinaceae is extracted with an organic solvent, e.g. a hydrocarbon, halogenated hydrocarbon, ether, ester or ketone, and a polyprenyl compound of the formula (A is OH or acyloxy; n is an integer 12-18) or a mixture thereof is obtained from the resultant extract. The aimed substance is separated from the extract by subjecting the extract to the separation method, e.g. chromatography, fractional dissolution method, and isolating a fraction exhibiting 0.18-0.25 and/ or 0.50-0.55 Rf value under conditions as to give 0.40-0.45 Rf value of solanesyl acetate using a thin layer chromatographic plate manufactured by Merck & Co., Inc., with a mixed solvent of n-hexane and ethyl acetate as a developing solvent.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a method for producing polyprenyl compounds or mixtures thereof. More specifically, the present invention relates to a polyprenyl compound represented by the following general formula (wherein A represents a hydroxyl group or an acyloxy group, and n is an integer from 12 to 18) or a mixture thereof. Regarding the manufacturing method.

The compound represented by the general formula (1) is disclosed in JP-A No. 57-9193.
As described in Publication No. 2, the general formula % (%) (in the formula, -CI(r-C=C-CHz- represents a trans isoprene unit, and j is an integer from 12 to 18) It is useful as an intermediate for the production of mammalian dolichol, which is represented by 58-96032), and it was also shown that mammalian dolichol could be synthesized from this product (see Japanese Patent Application Laid-Open No. 58-83643). However, when viewed as plant resources, Ichi, Tsu, and Himalayan are limited in terms of area and quantity, so we will search for plants that produce the compound represented by general formula (I) in addition to these. This is extremely important.

W., 5asak et al. isolated a mixture of polyprenylene esters containing 11 to 19 isoprene units from leaves of German spruce and European fir, but the type of esters and the number of cis and trans forms of isoprene units were unknown. The detailed structure has not yet been determined (FEBS Le
tters, 64.55 (1976)]. As described in JP-A-58-83643, it is essential that a polyprenyl compound that can be used as a raw material for the synthesis of mammalian dolichol has a polyprenyl μ chain structure represented by the general formula (I). For this purpose, it is at least necessary to correctly determine the number and arrangement order of the cis-type and trans-type isoprene units.

From this point of view, the present inventors extensively examined plants belonging to the Pinaceae family, the genus Abies and the genus Abies. As a result, the organic solvent extracts from the leaves of these plants contained the above general formula (I).
Polyprenol 1 (A-OH) and/or its acetate ether (A=O
CCH3) a polyprenyl fraction containing a relatively high concentration of a mixture of congeners is obtained, the polyprenyl fraction desirably being relatively easily separated into its constituent individual polyprenyl congeners; Therefore, it has been found that the polyprenyl fraction and each polyprenyl homolog separated therefrom are highly suitable as synthetic intermediates for mammalian Dolicho/I/classes, and the invention has been completed.

Therefore, according to the present invention, leaves of a plant belonging to the Pinaceae family, Picea genus, Abies genus are extracted with an organic solvent, and a polyprenyl compound or a mixture thereof is separated from the obtained extract,
The polypreny/L before or after separation as necessary
'A method for producing a polyprenylene μ compound represented by the general formula (I) or a mixture thereof, which comprises subjecting the compound or a mixture thereof to hydrolysis, esterification or transesterification, or two or more reactions thereof. provided. After extracting the leaves of the above-mentioned plants with an organic solvent and hydrolyzing the resulting extract as necessary, chromatography, fractional dissolution method, fractional cryoprecipitation method, molecular distillation method, or two or more of these methods are used. For the separation method consisting of the above combination, a plate for thin layer chromatography manufactured by Muri Co., Ltd. [Silica gel 60F254 coated @ (pre-co
Thin layer chromatography (10 cM development) using a mixed solvent of hexane and ethyl acetate in a volume ratio of 9:1 using a layer thickness of 0.25 m.
The Rf of solanesyl acetate as a standard substance in
o,is~0. under the condition that the value is 0.40~0.45.
25 and/or Rf within the range of 0.50 to 0.55
By isolating and collecting both components showing the value, polyprenylene μ consisting essentially of a mixture of the compound of general formula (I) where A is a hydroxyl group and/or the compound of general formula (I) where A is an acetoxy group is obtained. A composition can be obtained. This composition may contain one or more polyprenyl analogues in which the value of n in general formula (I) is 11 or less and/or 19 or more. It can be cut by chromatography or column chromatography.

In addition, the polypreny lvl [! ii can be separated into its respective homologs by, for example, high performance liquid chromatography.

The plants of the genus Spruce or the genus Abies (hereinafter sometimes referred to as raw material plants) used as raw materials in the present invention are both plants of the phylum Pullinphyta, subphylum Gymnosperta, subphylum Coniferae, class Coniferae, order Pinaceae. It is a plant belonging to Plants of the genus Picea are widely distributed from the temperate zone to the subarctic zone of the Northern Hemisphere, and approximately 40 species are known.
omorih), Japanese pine (Pioaa glee)
hnii), Abies fir (preferably used in the box). Plants of the genus Abies are widely distributed in the Northern Hemisphere and Central America, and about 5
0 types are known, but are especially used.

The leaves of the raw material plants may be subjected to the extraction treatment according to the present invention after drying, or they may be used undried. Dried leaves are generally preferred, and the degree of drying is advantageously such that the moisture content is generally less than about 30%, preferably less than 10%, based on the weight of the dry leaves. Furthermore, it is preferable to extract the leaves after crushing them, thereby increasing the contact area with the extraction solvent and increasing the extraction efficiency.

Polyprenyl congeners represented by the general formula (I) are generally contained in fairly high concentrations in the form of free alcohol and/or in the form of acetic acid NTE.
In order to effectively extract the polyprenyl analogue from the leaves, an oil-soluble organic solvent that well dissolves the polyprenyl analogue is preferably used. As such oil-soluble organic solvents, those having a dielectric constant (ε) of 32.7 or less, preferably 25.0 or less, and more preferably 20.7 or less are suitable. Each of the MKs can be used alone or as a mixed solvent of two or more.

(a) Hydrocarbons: for example, petroleum ether, pentane,
Hexane, heptane, benzene, toluene, xylene, etc.

(bl Halogenated hydrogen oxides: For example, chloroform, methylene chloride, carbon tetrachloride, ethane tetrachloride, perchlorethylene, trichlorethylene, etc.

(el Este/L's: For example, methyl acetate, ethyl acetate, ethyl propionate, etc.

fdl Ether/L/s: For example, diethyl pether, diisopropyl ether, tetrahydrofuran, dioxane, etc.

(e) Ketones: For example, acetone, methyl ethyl ketone, diethyl ketone, diisopropyl ketone, etc.

+f) Apco/l's: For example, methyl alcohol, ethyl alcohol-μ, propyl alcohol, butyl alcohol-μ, etc.

When selecting the solvent to be used, it is desirable that the target polyprenyl compound of general formula (I) be selectively extracted with high efficiency, and other substances should be extracted as little as possible. Among them, hydrogen ash, halogenated hydrocarbons, ethers with low polarity such as ester ivg, diethyl ether μ, and diisopropyl ether, and ketones are particularly preferred.

The number of extraction solvents used is not critical and can vary widely depending on the type of solvent used and the type and condition of the leaves to be extracted, but in general, 1 part by weight (dry) of the leaves of the raw material plant is used. (on a weight basis) It is advantageous to use within the range of about 1 to about 100 parts by weight, preferably 5 to 50 parts by weight, and more preferably 10 to 30 parts by weight.

Extraction can be carried out by immersing the leaves of the raw material plant in the above solvent and stirring continuously or intermittently as necessary. The temperature during extraction is also not critical and can be varied over a wide range depending on conditions such as the type and amount of solvent used, but generally a temperature from about 0°C to the reflux temperature of the solvent can be used. and room temperature is usually sufficient. It is advantageous to carry out the extraction under such conditions, usually for a period of 1 hour to 10 days.

The immersion liquid after the extraction process is made into a concentrated liquid by removing the leaves and other solid content, and then removing the solvent as necessary. The extract thus obtained is then subjected to chromatography, fractional lysis, fractional cryoprecipitation, molecular distillation, or two of these methods.
The target polyprenyl fraction can be recovered by subjecting it to a separation step consisting of one or more species.

Confirmation of the content of the polyprenyl compound in the above separation process was performed using a thin layer chromatography plate (Silikage/L' 60 Fzsapre-coat manufactured by Merck & Co., Ltd.).
Solanesil as a standard substance in thin-layer chromatography (101 development) using a mixed solvent of n-hexane and ethyl acetate in a volume ratio of 9:1 as a developing solvent. The Rf value of acetate is 0.4
Under the conditions of 0 to 0.45, 0.18 to 0.25[:
When A represents a hydroxyl group in the general formula 0)] and/or Rf within the range of 0.50 to 0.55 (when A represents an acetoxy group in the preceding general formula (1))
This can be done depending on whether or not a Nupot exists at the value. Therefore, when referring to the Rf value of thin layer chromatography in the following explanation, it should be understood that unless otherwise specified, it means the value measured under the above conditions.

The operations of chromatography, fractional dissolution, fractional cryoprecipitation, and molecular distillation that can be used in the extract separation step described above are known per se, and the present invention is also carried out in accordance with the known methods. Because you can
Details of each method will be explained with reference to literature, and only points to be particularly noted will be described here.

(4) Chromatography [e.g. H, Heftm
an.

“Chromatograpby”, Ra1nhol
d Publish Co., New York (
(1961)] Thin layer chromatography and liquid chromatography are suitable for small amounts of extracts, but column chromatography is suitable for processing large amounts of extracts. Examples of carriers for chromatography that can be used include silica gel, alumina, florizi μ, celite, cellulose, and cellulose, with silica gel being preferred.

When performing a separation operation using a silica gel column, the developing solvent may be, for example, hexane/ethyl acetate/I/(
Capacity ratio 9B: 2 to 80: 20), hexane/
Diisopropyl ether/L/(volume ratio 95:5~B6
:20), petroleum ether/L//methyacetate/I/(volume ratio 98:2 to 80:20), petroleum 1-te IV/I'77''.

Bil alcohol (volume ratio 99:1 to 90:10)
, benzene/diethyl ether/I/(total ratio 95:5~
Examples include mixed solvent systems such as benzene/ethyl acetate/I/(solubility ratio 98:2 to go: 20), chloroform, methylene chloride, etc.

(B) Fractionation m solution [e.g. L, CoCraig.

“Teehnique of Org & nic Che
mistry”, Vol, 3゜Interscience
ee, (1951)] The polyp Vt-W compound of the general formula (1) is easily soluble in nonpolar solvents such as pentane and hexane, but is dissolvable in polar solvents such as methanol and water. Therefore, by utilizing this difference in solubility, it can be purified by the molecule 11 dissolution method. For example, by dissolving a crude product such as an extract concentrate in the above non-polar solvent and then washing with a polar solvent that is immiscible with the polar solvent, impurities that are easily soluble in the polar solvent are largely removed. can do. Examples of the It-polar solvent preferably used in this method include petroleum ether, hydrocarbon solvents such as pentane, hexane, heptane, benzene, and toluene, and halogenated hydrocarbons such as methylene chloride and chloroform. It is. In addition, examples of polar solvents that are immiscible with such non-polar solvents include water or methanoic acid.
/L' is suitable.

(C) Fractional cryoprecipitation method [e.g., E + W, −
Berg.

“Physical and Chemical Me
thods of 5eparation"Chapt
er 14, 15, McGraw-Hill
, New York (1963)] The polypreny compound of general formula (I) solidifies at about -10°C or lower. Therefore, impurities that do not solidify at such temperatures can be removed by leaving the extract under cooling at -10°C or below, preferably about -15 to about -30°C, solidifying the target product, and then performing solid-liquid separation. Can be purified. However, since the polyvinyl compound becomes a waxy solid with very good crystallinity, it is difficult to completely purify it by this method, so it is removed by combining it with other purification methods. ■) Molecular distillation method [e.g. G, Burrow+s
.

“Mo1ecular Diatillation”
, C1arendon Press, Oxfor
(1960)] Since the compound of the general formula (I) has a large molecular weight, low molecular weight impurities can be removed by using a molecular distillation method. For example, by molecular distillation under heating conditions of 100 to 250°C at a vacuum degree of 10 to 10 wn,
It is divided into a low molecular fraction and a high molecular fraction. At this time, the target substance is retained in the high molecular fraction, and low molecular weight impurities can be largely removed.

Polyprene μ of sufficiently high purity can be obtained by each of the above separation methods.
If a fraction cannot be obtained, a combination of two or more of these separation methods can also be used. for example,
Chromatography and fractional dissolution; Chromatography and fractional cryoprecipitation and fractional dissolution; Chromatography and fractional cryoprecipitation and molecular distillation; Molecular distillation and fractional dissolution; Molecular distillation and fractional dissolution and fractional cryoprecipitation A combination of methods can be used.

Thus, the Rf value in thin layer chromatography is 0.
．． 18-0.25' and/or 0.50-0-55
A fraction of the amount is isolated and collected. Rf value is 0.18-0.25
, consisting essentially of a mixture of homologues of the general formula (I) when human represents a hydroxyl group; on the other hand, Rf
Both values of 0.50 to 0.55 consist essentially of a mixture of homologs in the above general formula (I) when the group represents an acetoxy group. These fractions may contain one or more polyprenyl analogues in which the value of n in general formula (I) is 11 or less and/or 19 or more;
These can be cut by, for example, liquid chromatography or column chromatography, if necessary.

The individual homologue components can also be isolated by further subjecting the two fractions thus obtained, for example, to partition 5 high performance liquid chromatography.

In addition, in the above separation step, it is possible to convert the extract in advance from the above-mentioned homologue in which the setoxy group is represented to the corresponding homologue in the case where A represents a hydroxyl group. This may simplify the subsequent separation operation. However, such hydrolysis can of course be carried out on both components containing components having an Rf value of 0.50 to 0.55 after the separation operation. This hydrolysis can be carried out using any conventional method known for hydrolyzing fatty acid esters/I/s, such as sodium hydroxide or About 100 parts by weight of a solution in which potassium hydroxide is dissolved (the potassium metal hydroxide concentration can preferably be about 0.1 to 30% by weight) is added to about 100 parts by weight of the above extract or both. 5 to 50 parts by weight plus about 25 to 90 parts by weight
The reaction may be carried out at a temperature of about 0.5 to 5 hours.

In the polyprene μ fraction isolated and recovered by the above-mentioned method, the fraction with an Rf value of 0.18 to 0.25 is a polyprene ρ homolog in the general formula (1) where human represents a hydroxyl group. consisting essentially of a mixture and R
Both the f-values of 0.50 to 0.55 consist essentially of a mixture of a plurality of polyprenylacetate analogues in which A in the general formula (summer) represents an acetoxy group. The ratio of the former to the latter in the extract is approximately 9.
:1 to 1:999, and the distribution order (distribution pattern) of polypreno/l' or polyprenylacetate-F congeners in each division varies depending on the type of raw material plant, and the same species of raw material The distribution of substances obtained from plants is generally the same, regardless of factors such as the youth of the leaves used as raw materials, the time of collection, and the region.

When introducing the polyprenyl μ composition obtained by the method of the present invention into Dolicho/I/class mammals, the composition may be used as it is, or if necessary, each polyprenyl compound constituting the composition may be isolated. You can also react afterwards. Furthermore, the composition or the isolated polyprenyl compound, depending on whether it is in the free alcohol or in the acetate form, can be subjected to hydrolysis, esterification or transesterification reactions or their 28
Polyprenol may be converted into polyprenyl ester, or polyprenyl acetate may be converted into other polyprenyl aceters or polyprenol μ shown by the general formula (1) by subjecting it to reactions f or more.

As an example of the polyprenyl ester represented by the general formula (1) produced by the method of the present invention, when A in the general formula (1) is represented by RCOO-, R is a hydrogen atom, and the number of carbon atoms is 1 to 1.
18 alkyl groups (e.g. methyl 1ethyl/l/, n-
Propyl, isopropyl! , n-butyl, 5ea-butyl, isobutyl, member rL-butyμ, n-pentyl, isoamide A/, n-hexyl, n-octyμ, n-deciμ
, n-unde V71/, n-dodecyl 1-stearyl μ, etc.), carbon number 2-18 aμ, 3-butenyl μ, 3-bentenyl p, 4-pentenyl, geranyl, geranyl p1 farnesi 〃, farnesyl methyl, etc.)
, cyclohexyl having 5 to 7 carbon atoms (e.g., cyclopentyl, cyclohexyl, methylcyclohexyl, cyclohebutyp, etc.), aryl group having 6 to 10 carbon atoms, phenyl, tolyl, xylyl, naphthyl, etc.), carbon number 7 to 11 afuruki/L' groups (e.g. benzyl, phenethyl, methylbenzyl, dimethylbenziμ, α- or β-naphthylmethyl, etc.) or methyl groups substituted with 1 to 3 fluorine or chlorine atoms ( For example, monofluoromethyl, trifluoromethyl, monochloromethyl, dichloromethane, etc.) can be mentioned. The esterification or transesterification reaction of the polypreno μ represented by the general formula (I) or a mixture thereof to obtain these polyprenyl esters or mixtures thereof can be carried out using a known esterification or transesterification reaction method to obtain the enethers. This can be done using conditions. For example, esterification can be carried out using the general formula (1): acid, linoleic acid, lylunic acid, farnesylic acid, farnesyl acetic acid, benzoic acid, 3,5-dimethylbenzoic acid, 4-ethylbenzoic acid, etc. with the desired strength μ acid or their acid nitride or acid anhydride. This can be easily done by mixing and heating and stirring if necessary. Preferably, the polypreno/l' or a mixture thereof is dissolved in a solvent such as hexane, benzene, methylene chloride, chloroform, diethyl ether, etc., and 1 to 5 molar equivalents of the acid halide relative to the polypreno p are added thereto. In addition, the reaction is carried out by stirring for 1 to 24 hours at an appropriate temperature from room temperature to the reflux temperature of the solvent in the presence of 1 to 5 molar equivalents of pyridine relative to the polyprenol.

In addition, the transesterification reaction is carried out by combining the polyprenol represented by the general formula (I) or a mixture thereof with a lower acid p-ester A/
(preferably methyl ester μ, methyl ester μ, etc.) in the optional presence of a transesterification reaction. Preferably, the polyprenol or the mixture thereof and the desired carboxylic acid ester are dissolved in a solvent such as benzene, toluene, xylene, etc., and 0.01 to 0.17μ equivalent of sodium hydride is added to the polyprenol. In addition, the reaction is conducted under refluxing conditions of the solvent for 2 hours to 5 days.

The polyprenyl compound represented by the general formula (1) or a mixture thereof obtained by the method of the present invention is, for example,
According to the method described in Japanese Patent No. 8-83643, mammalian dolichols can be easily derived.

In addition to the above, the polyprenyl compounds and mixtures thereof obtained by the method of the present invention can be used, for example, as cosmetic base materials.
It is also useful as an ointment base material or a raw material for their production.

EXAMPLES The present invention will be explained below with reference to Examples, but the present invention is not limited by these Examples.

Note that in the Examples and Reference Examples, IR analysis was measured using a liquid film, and NMR was measured in CDCl3 using TMS as an internal standard. The values of FD-MASS analysis (field ionization mass spectrometry) are values corrected as 'H, 12C, 'O, and 'Br.

Example 1 10 European fir leaves collected in Hokkaido in late July
kg (undried weight) was dried with hot air at about 50°C for 24 hours, and then extracted by immersing it in a mixed solvent 1001 of acetone/n-hexane = 171 (volume ratio) at room temperature (about 30°C) for about 6 days. Thereafter, it was filtered, the residue was divided into 10 equal parts, the above-mentioned mixed solvent 31 was added to each part, homogenized and extracted for 3 minutes, and then filtered.

The concentrated 55of obtained by combining all these filtrates and distilling off the solvent was divided into 10 equal parts, and each of them was mixed in an n-hexane/methanol/water = 10/9/1 (volume ratio) mixed bath [21 Distributed extraction was performed four times each. all n
- After concentrating the hexane layers, they were separated using a silica gel column using a mixed solvent of n-hexane/ethyl acetate = 97/3 to 90/10 (volume ratio) as a concentration gradient eluent. Silica gel thin layer chromatography using a mixed solvent of ethyl acetate-9/1 (volume ratio) [
Merck thin layer chromatography plate (silica gel 60
F>Coating; layer thickness 0.25m+) is used, 10CM
Fractions containing components with Rf values of 0.51 and 0.20 (referred to as fractions 1 and 2, respectively) were separated to obtain oils of approximately 489 and approximately 6'3' f, respectively. In addition, in the above thin layer chromatography, solanesyl acetate showed an Rf value of 0.40. Fraction 1
and 2 oils separately into 700 ml of methanol, 11 water, and 700 ml of water.
〇- and sodium hydroxide 5011 for 2 hours 65
After heating to °C, methanol was distilled off, and the residue was extracted with 700% of Ni-diethyl ether. The solvent was distilled off to obtain liquid products. These liquids were then mixed with about 2 kg of silica gel and n-hexane/diisopropyl ether: 9
A liquid mixture of 21.59 and 0.21 (fraction 1a and 2a) was obtained. Fractions 1a and 2a were both polyprenols with a purity of 90% or more, and the molecular weight distribution measured for them was that of a queen. This value is calculated using a semi-preparative high performance liquid chromatography column Li manufactured by Merck Co., Ltd. for the above liquid material.
Area of chromatogram obtained by high performance liquid chromatography using Chrosorb RP-18-5 (C+s type) with acetone/methanol-90/10 mixed solvent as eluent and differential refractometer as detector It is calculated from the ratio.

11 1.8 2.3 12 4.8 6.5 13 21.6 23.4 14 36.5 37.0 15 1B, 7 17.2 16 6.0 5.3 17 4.3 3.4 18 3.2 2.6 19 2.1 1.7 20 1.0 0.6 The molecular weight distributions of fraction IIL and both fractions 2a are in good agreement within a range of ±2%, and are substantially equal distributions. You can say that. fraction 1a using high performance liquid chromatography as described above.
Components with n values from 12 to 18 were separated from the liquid material of 2a and subjected to mass spectrometry, infrared absorption Subekle V, 'H
-NMR spectrum and 13C-NMR spectrum [mu] confirmed that these components were polyprenol in which human is a hydroxyl group in general formula (I). FD-MASS results and 'H-N for each component of fraction 1a
The δ values of MR are summarized in Table 1, and the δ values of '3C-NMH are summarized in Table 2. The values of D for each component of fraction 2a were also substantially equal to those of the corresponding components of fraction 1a.

Example 2 In Example 1, 500 r of acetone was added to the oily substance of fraction 1.
The resulting mixture was filtered, and the oil obtained by concentrating the p solution was directly converted to about 2 kg without saponification reaction with sodium hydroxide. υ column chromatography was performed using silica gel and chloroform, and both fractions showed Rf=0.51 in the thin layer chromatography described above.22.
A liquid of 1fI was separated. This liquid is polyprenylacetate with a purity of 90% or more, and the molecular weight distribution measured using the same method as in Example 1 is as follows. Good agreement was shown within ±1% with that for turtle polyprenol.

i i 2.5 12 5・4 13 22.5 14 36.7 15 18.2 16 5.4 17 3.7 18 3.0 19 1.7 20 0.9 High performance liquid chromatography same as Example 1 Then, each component with an n value of 12 to 18 was separated from the above liquid material, and subjected to mass spectrometry, infrared absorption spectrum /l/, 'H-N
It was confirmed by MR spectrum and -NMR spectrum that these components were polyprenylacetate in general formula (I) where A is an acetoxy group. Table 3 shows the results of FD-MASS analysis for each component.

Table 3 Furthermore, polyprenols obtained by similarly hydrolyzing each component according to the hydrolysis reaction performed in Example 1 are the same as those obtained in Example 1 with the same value of n. 'H-NMR Nuvector,' which is exactly the same as polyprenols.
3C-NMR spectrum and infrared absorption spectrum were given.

Examples 3 and 4 Instead of the iK of European fir in Example 1, Serbian spruce leaves collected in Hokkaido in late July and German spruce leaves collected in Kyoto City in late December were used as raw materials. By the same operation and conditions as in Example 1, a polyprenol in which A is a hydroxyl group in the general formula (I) was obtained. Regarding the structure of the polyprenol, the results of mass spectrometry, IR analysis, and NMR analysis of each component fractionated by high-performance liquid chromatography in the same manner as in Example 1 substantially match those obtained in Example 1. This was confirmed by doing so. The yield of the polypreno/L/ (referring to the combined fractions 1a and 2a equivalents in Example 1), the molecular weight distribution (chain length distribution) measured in the same manner as in Example 1, etc. are summarized in Table 4. It was shown.

Table 4 Examples 5 to 16 The leaves of the raw material plants were dried with hot air at about 55° C. for 30 hours, then divided into 100′I portions, immersed in 11 solvents shown in Table 5, and extracted at room temperature for 7 days. The extraction solvent was distilled off from the obtained extract, and the weight of the residue was measured and summarized in Table 5 as the total amount of extract. These extracts were mixed with n-hexane 20
0- and the solution was dissolved in methanol-W/water = 971
After washing three times with a mixed solvent (volume ratio) of 100-, it was dried over anhydrous magnesium sulfate, and the solvent was distilled off to obtain an oil. Add this oil to MegnotV 501717'
After heating at 65°C for 2 hours with 1f of potassium hydroxide, methanol was distilled off, and the residue was diluted with diethyl ether/
' (100 FFIA was added for extraction, and the ether layer was washed three times with about 50 ml of saturated saline, dried over anhydrous magnesium sulfate, and the solvent was distilled off to obtain an oil. using n-hexane/
Separate with a mixture of ethyl acetate = 9/1 (volume ratio), and perform plating by thin layer chromatography in the same manner as in Example 1.
Both fractions (polyprenol mixture) showing an Rt value of 0.19 were obtained. Table 5 shows the weight of both parts as the polyprenol content.
are summarized in. The composition of the obtained polyprenol mixture was the same as that of Example 1.3, regardless of the type of solvent used.
and 4 were substantially identical to that of the polyprenol mixtures obtained from the same source plants.

Table 5 It was used in this case.

Example 17 Polypreno/lz1.24F of general formula (I) with n=15 and A=OH obtained from leaves of European fir in the same manner as in Example 1 and pyridine 1. In a solution of Of in dry diethyl ether was added 1.21 g of acetic anhydride at room temperature.
was added dropwise, and after the addition was completed, the mixture was stirred overnight at room temperature. The resulting reaction mixture was washed with saturated brine, then dried over anhydrous magnesium sulfate, diethyl ether was distilled off, and a pale yellow viscous liquid was obtained.

This product was purified by silica gel column chromatography (using hexane/ethyl acetate as a developing solution).
．． A pale yellow liquid of 07F was obtained. When this material was subjected to IR analysis, the absorption caused by the OH groups of the raw material polyprenol of about 3,300 c1j' disappeared, and the absorptions of 1745 ci'o and 1255ai'' caused by 0cOCHs appeared.In addition, NMR analysis When this was carried out, the signal /L' (double, δ = 4.08) assigned to -CI'hQH of the raw material polyprenol disappeared, and a new signal assigned to CHxOCOCHx (
doublet, δ=,i,ss) was observed. −
The signal to be assigned to CHzOCOCHi was observed to overlap with the signal/L/(δ=2.04) assigned to -Cdan2-C-. Further, p mle = 1284 was obtained by FD-MASS analysis. From the above, it was confirmed that this liquid was polyprenylacetate of general formula (I) where n = 15 and A = 0COC)h. Polyprenyl acetates with n other than 15 and polyprenyl acetate mixtures with n arbitrarily distributed between 12 and 18 could also be synthesized by similar operations.

Example 18 Polypreno Az 1.24 f of general formula (I) with n=15 and A=OH obtained from leaves of Scots fir in the same manner as in Example 1, methi ivo, sy oleate and sodium hydride 0.01f was dissolved in 50% of toluene and heated at 110°C for 24 hours under a nitrogen gas atmosphere. After the reaction solution was cooled to room temperature, it was washed with saturated brine, dried over anhydrous magnesium sulfate, and the solvent was distilled off to obtain a yellow liquid. This product was purified by silica gel column chromatography (using hexane/ethyl acetate as a developing solution) to obtain a colorless viscous liquid of 0.469. When this liquid was analyzed by IR, absorption of approximately 3,300 ci' due to the OH groups of the raw material polyprenol had disappeared. In addition, by FD-MASS analysis j) ml
gave e = 1506. From the above, this one has n = 15 and A = 0COC1? It was confirmed that it was a polyprenyl oleate of the general formula (I). Polyprenyl oleates with n other than 15 and polyprenyl oleate mixtures with n arbitrarily distributed between 12 and 18 could also be synthesized by similar operations.

Example 19 0 benzoyl chloride was added at room temperature to a mixture of polypreno IV 1.11f of n=13, A=OI (general formula 〇) obtained from Serbian spruce leaves in the same manner as in Example 5 and 101 nl of pyridine. .289 was added and stirred overnight at room temperature. The reaction mixture was then poured into about 150 m/m of water, extracted with diethyl ether, and the resulting ether layer was mixed with saturated brine, diluted hydrochloric acid, saturated sodium hydrogen oxide,
After washing with saturated brine again, it was dried over anhydrous magnesium sulfate, and diethyl ether μ was distilled off to obtain a yellow liquid. This was purified by silica gel column chromatography (using hexane/ethyl acetate as a developing solution) to obtain a slightly yellow liquid of 0.76F. When this product was analyzed by IR, the absorption caused by the OH group of the raw material polyprenol disappeared, and 17 s s , -;' and 127
Absorption due to ester bonds appeared in 0Ci''.Furthermore, FD-MASS analysis revealed that mle = 1210
gave.

From the above, this liquid substance has n = 13, A −O
It was confirmed to be polyprenyl benzoate of general formula (I), which is COCgHs. Polyprenyl benzoates with n other than 13 and polyprenyl benzoate mixtures with n arbitrarily distributed between 12 and 18 could also be synthesized by similar operations.

Reference Example: In a three-layer argon-substituted Rofuranuco, 0.316 f (13 mmol) of magnesium strips and 0.0.0 of anhydrous tetrahydrofuran and 0.5- and 1.2-dibromoethane were added.
8- was added and heated with a hair dryer until it foamed violently. Then 2-[4-bromo-3-methylbutoxy]-
Tetrahydro-2H-pilaf 2.51 f (10 mmo
1) in anhydrous tetrahydrofuran (3, omz) was added dropwise to the activated magnesium at such a rate that the solvent just boiled. After completing the dropping, the mixture was heated to 70 ml.
The mixture was stirred at ℃ for 15 minutes. To this was added 60 ml of anhydrous tetrahydrofuran to obtain a Grignard solution.

Separately, in a third flask purged with argon, 6.42 y (s mmol
) solution of anhydrous tetrahydrofufurn (15 ml) and Li
A solution of zCuCβ4 in anhydrous tetrahydrofuran (0.1 molar solution, 20 m/) was charged. The previously prepared Grignard solution was added dropwise to this at 0°C over 1 hour, and then at 0°C.
Extraction was carried out for 2 hours. Wash the ether layer with saturated saline,
After drying over anhydrous magnesium sulfate, the solvent was distilled off using a rotary evaporator to obtain a pale yellow liquid of 7.69y. This product was determined by silica gel thin layer chromatography (using hexane/ethyl acetate IV-=9'l/3 as a developing solution) to have p Rr = o, with the main neupot at as.

Furthermore, when this pale yellow liquid was analyzed by FD-MASS, m/e =
1284 was not detected at all, and the formula (wherein -CHx-C
=C-CH2- represents a trans isoprene unit (CHx H1), and -CHr-C=C-CH2- represents a cis isoprene unit. ) was detected as a main peak with m/e = 1396, which represents a compound represented by:

Next, this pale yellow liquid was dissolved in 40-hexane,
To this, 0.13 f of p-toluene-nulphonic acid pyridine
(0.5 mmol) and ethanol-1v20- were added.

This solution was heated and stirred at 55° C. for 3 hours. After cooling, 0.219 g of sodium carbonate was added to neutralize, and the solvent was distilled off using a rotary evaporator. The obtained concentrate was dissolved in ether, washed with saturated sodium hydrogen oxide solution, then saturated brine, and dried over anhydrous magnesium sulfate. The solvent was removed on a rotary evaporator and the remaining oil was heated to 0.5 Torr, 1
The mixture was heated at 50° C. for 30 minutes to remove low-boiling components. The remaining liquid portion was purified by silica gel column chromatography (hexane/ethyl acetate; 9/1 was used as a developing solution) to obtain a colorless and transparent liquid 5.47F. This product was determined by silica gel thin layer chromatography (using hexane/ethyl acetate = 971 as a developing solution) to have Rr = 0.
A single spot was given to 19. Also, according to the analysis results shown below, this liquid has j=15 in general formula (II).
It was confirmed that there is a compound +.

FD-MASS analysis m7e = 1312 (calculated value 1312) IR analysis (ci'); 830, 1
060, 1376, 1440°2850.2920
．． 3320 "C-NMR analysis (ppm/intensity) x: as, 36s
/43o.

135.229/3567.135.005/349
.

134.937/290.131.210/213゜1
25.07115242, 124.993/499゜1
24.4481505, 124.282/463.

124.214/445.61.2411551.

40.0291541.39.757/683°37,
No. 481582, 32.24515500゜32.02
1/456.29.3161528.

26°825/492, 26.6991548.

26.43615166.25.6771542゜25
．． 3081567.23.430/6330.

19.5571548, 17.679/353.

16.006/640.

'H-NMR analysis (ppm, signal type, proton ratio) 5.10 (b, 18H), 3.66 (m, 2H), 2
．． 03 (b, 70H), 1.68 (8, 48H), 1.
60 (s, 9)f), 1.80 1.10 (
m, 5H), 0.91 (a, 31EI) Patent applicant: RiRa Co., Ltd. Agent: Ken Honda, patent attorney

Claims (3)

[Claims] (1) Extract the leaves of plants belonging to the Pinaceae spruce genus or Abies genus with an organic solvent, separate the polyprenyl compound or its mixture from the resulting extract, and if necessary, remove the polyprenyl compound or its mixture before or after the separation. A general formula characterized by subjecting a mixture to hydrolysis, esterification or transesterification, or two or more of these reactions (wherein A represents a hydroxyl group or an AV/L/oxy group and represents a soglene unit; , n is an integer from 12 to 18.) A method for producing a polyprenyl compound or a mixture thereof. (2) The organic solvent is a hydrocarbon, a halogenated hydrocarbon,
The manufacturing method according to claim 1, wherein the ether is selected from ethers, esters, and ketones. (3) Extract the purple color of plants belonging to the Pinaceae spruce or Abies genus with an organic solvent, hydrolyze the resulting extract as necessary, and then perform chromatography, fractional dissolution, fractional cryoprecipitation, and molecular distillation. For a separation method consisting of a method or a combination of two or more of these methods, a plate for thin layer chromatography (Silikage/I/6
Thin layer chromatography (1
Soranesi as a standard material in 01 development)! 0.0 under the condition that the Rf value of acetate is 0.40 to 0.45.
2. The manufacturing method according to claim 1, wherein the civolibrenyl composition is obtained by isolating and collecting both components exhibiting an Rf value in the range of 18 to 0.25 and/or 0.50 to 0.55.