JP3226851B2 - Oncovirus activation inhibitor - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to an oncovirus activation inhibitor, and more particularly, to a plant extract that suppresses oncogenic virus, and a carotenoid substance obtained from the plant extract.

[0002]

BACKGROUND OF THE INVENTION Cancer is the number one cause of death in Japanese people.
Prevention of carcinogenesis has become a very important issue for the health of the population. It is generally believed that cancer arises through two distinct phases, initiation and promotion. That is, initiators such as ultraviolet rays, radiation, and mutagens cause damage that cannot repair the DNA of normal cell genes (initiation), and the resulting potential cancer cells are repeatedly stimulated by chemicals called promoters. Being (promotion) cancerous. Therefore, cancer can be prevented by blocking either the initiation or promotion process.

However, it is impossible to completely remove the initiator because ultraviolet rays, radiation, and various mutagens which act as the initiator are present in the environment. In addition, since it is considered that an adult already has an initiated cell, inhibition of the initiation process is not effective from the viewpoint of preventing carcinogenesis. On the other hand, since the promotion process is long-term, inhibiting this process is considered to be an effective method for preventing carcinogenesis.

[0004]

In view of the above, attempts have been made to search for an anti-tumorigenic promoter, and it has been reported that many plant extracts such as Huangbai act as anti-tumorigenic promoters. It has been desired to provide a safe plant extract originating from an edible plant having an activity of inhibiting the formation of a plant. In addition, examples have been reported in which crude extracts of many plants such as Huangbai can suppress experimental carcinogenesis,
Its active ingredients are being revealed.

[0005] An object of the present invention is to provide a plant extract having an inhibitory effect on the activation of oncogenic virus as an anti-cancer promoter and an oncovirus activation inhibitor containing the plant extract as an active ingredient.

[0006]

As a primary screening test for an anti-tumor promoter, there is an Epstein-Barr virus genome activation suppression test. This test method uses tetradecanoyl phorbol acetate (hereinafter abbreviated as TPA) as a promoter in a culture system of Raji cells, which are cultured cells derived from Parkin's lymphoma containing the Epstein-Barr virus genome. Inhibition of the expression of the virus genome is used as an index. This method is rapid and quantitative, and allows for the detection of active substances in trace amounts.

[0007] Epstein-Barr virus is a virus belonging to the family Herpesviridae, and is considered to be a cause of Perkit lymphoma and nasopharyngeal carcinoma. The virus is not only detected in these cancer patients, but is extremely ubiquitous and universal in humans worldwide, and it is said that almost all adults are infected with the Epstein-Barr virus. ing. Epstein-Barr virus has been shown to blast into normal B lymphocytes of humans as an infection target and confer an infinite proliferative capacity on them, and it has been shown that Epstein-Barr virus imparts infinite proliferative capacity to humans. Defined as resident virus factor.

The inventor has conducted systematic studies and experiments on a large number of microorganisms and plants using the Epstein-Barr virus genome activation-suppressing action as an index, and as a result, extracts extracted from Paradicham paprika, The present inventors have obtained the remarkable finding that the carotenoid obtained from the above has a strong Epstein-Barr virus genome activation inhibitory effect, and completed the present invention.

[0009] The invention of claim 1 is an agent for suppressing the activation of oncogenic virus, which uses an extract of paradicom paprika. The invention of claim 2 is a carcinogenesis virus activation inhibitor comprising a carotenoid extracted from paradicham paprika as an active ingredient. The invention according to claim 3 is a plant extract having a carcinogenic virus activation inhibitory action extracted from paradicham paprika.

In the present invention, paradice paprika (pa)
radicsom paprika) is a vegetable native to Hungary. Its original fruit is clover-shaped, deep red,
The surface is glossy and waxy, and has the property that the color does not come off when heated. In addition, the pulp is thick, has a high sugar content, and has no green odor. As an example of the component analysis, the edible portion 10
Vitamin A potency (IU) per 0g is 780, Vitamin B
2 0.23 mg, Vitamin C 189 mg, Iron 0.
62 mg. The term "paradicsom paprika" as used in the present invention refers to this original species of paradicom paprika. In the present invention, among the original species, those that give a specific fruit are repeatedly selected, fixed, and fixed. Varieties (hereinafter referred to as pure varieties) are defined. In addition, the pure type is scientific name Capsicum annuum L.var.grossum
are categorized. In addition, a species obtained by crossing such a pure paradigm paprika with any of the large bell type, the Pimento type, the Hungarian paprika type, or the large neapolitan type peppers (hereinafter referred to as F1 type) and F1 type comrades It is defined to include the crossed species (hereinafter referred to as quaternary hybrids) and the reverted species of these hybrids (hereinafter referred to as F2 species). In other words, the term "paradicom paprika" as used in the present invention means, in addition to the original species, pure strains, F1 species, quaternary hybrids, F2 species, and hybrids of paradicham paprika thereafter.

[0011]

DESCRIPTION OF THE PREFERRED EMBODIMENTS In the present invention, an extract obtained by extracting paradicom paprika with acetone or methanol,
Further, the extract is preferably extracted with hexane. However, the extraction solvent is not limited to these.

The present inventor separated the components by liquid chromatography and analyzed the components in order to clarify the components of the extract. The main components of the hexane extract were carotenoids, and capsanthin, capsanthin monoester, and capsanthin. It was found that the diester was high. That is, the invention of claim 2 is a carcinogenesis virus activation inhibitor containing a carotenoid extracted from paradicham paprika as an active ingredient.

An extract obtained by extracting paradicom paprika with acetone, and a further extract obtained by extracting the extract with hexane exhibited a strong oncogenic virus activation-inhibiting activity almost equal to or higher than that of Huangbai extract as a pharmaceutical. Therefore, these extracts, or carotenoids obtained from the extracts, inhibit the expression of the Epstein-Barr virus genome, thereby exhibiting a carcinogenesis preventive effect and an antiviral effect. Therefore, they can be widely used in the fields of pharmaceuticals, cosmetics, and health foods from the viewpoint of cancer prevention. The invention of claim 3 is a plant extract having an inhibitory effect on the activation of oncogenic virus, which is extracted from paradicom paprika.

The Epstein-Barr virus
The effect of inhibiting the expression of the genome was observed by the following method. That is, the above-mentioned raji cell culture system is cultured by adding TPA, which is a carcinogenesis promoter, to n-butyric acid, which acts as a synergistic effect for the activity expression, and a non-detectable substance, and activated by TPA to be expressed on the cell surface. Epstein-Barr virus early antigens were detected by indirect immunofluorescence using antibodies from nasopharyngeal carcinoma patients.

[0015]

EXAMPLES The plant extract of the present invention is extracted by the following procedure. In the extraction of this embodiment, the pure species (scientific name)
Varieties classified as Capsicum annuum L.var.grossum)
Was used. That is, 1 kg of the edible portion of paradicom paprika was cut, immersed in acetone, stopped in a dark place at room temperature, and sometimes permeated to obtain a red acetone extract. Acetone was added again to the extraction residue, and the same operation was repeated three more times to collect an acetone extract. The extract was concentrated under reduced pressure to dryness to obtain an acetone extract. Further, methanol was used as an extraction solvent to prepare a methanol extract in the same manner as in this method. Hexane was added to these and dissolved, and the hexane-soluble fraction was collected to obtain a hexane extract. This was concentrated under reduced pressure to produce a hexane extract.

Using the acetone extract and the hexane extract, the expression inhibitory activity of the Epstein-Barr virus genome was measured under the following conditions. Epstein-
A culture obtained by adding fetal serum and antibiotics to PRMI1640 was used as a culture solution of the human lymphoblastoid cell line latently infected with the burr virus and Raji cells. Under these culture conditions, the spontaneous incidence of Epstein-Barr virus early antigen is less than 0.1%. Raji cells adjusted to a concentration of 1 × 10 ⁶ cells / ml were mixed with 4 mM n-butyric acid, 20 ng / ml.
Epstein-Barr virus early antigen was expressed by an indirect immunofluorescence method using nasopharyngeal carcinoma patient serum cultured at 37 ° C. for 48 hours in the above-mentioned culture medium to which TPA and 100 μg / ml of a test substance were added. Cells were detected, and the ratio of positive cells was calculated relative to a control to which no test substance was added, and the result was defined as the activity of inhibiting the reproduction of the virus genome. Further, the same activity was measured by changing the concentration of the test substance to 10 μg / ml and 1 μg / ml. Table 1 shows the results.

[0017]

[Table 1]

The extract obtained by extracting paradicom paprika with acetone and the extract obtained by extracting the extract with hexane showed a strong inhibitory effect on the activation of oncogenic virus. Its inhibitory action was almost the same as that of Huangbai extract used as a pharmaceutical, and its value as an oncogenic virus activity inhibitor was recognized. The radio cell survival rate is 70%.
, And it was found that there was almost no toxicity to cells.

Further, in order to analyze the extract, the hexane extract was saponified with 5% KOH / methanol and then analyzed by high performance liquid chromatography (hereinafter abbreviated as HPLC). The column used a silica gel adsorbent. As a carrier, a 65:35 mixture of n-hexane and tetrahydrofuran (hereinafter abbreviated as THF) was used. FIG. 1 shows the separation chart. The HPLC is Hitachi L-6200, and the detector is L-
Detection was performed at 450 mm using 4250. As a result, 10
The carotenoid of the component could be detected. Table 2 shows the composition of carotenoids obtained from these data.

[0020]

[Table 2]

The extract according to the present invention contains various carotenoids, in particular, as much as 37.1% of capsanthin (including capsanthin monoester and capsanthin diester), and was found to have an anticancer virus inhibitory effect.

Further, in order to clarify the effect of suppressing carcinogenesis,
A skin cancer suppression test using mice was performed. That is,
The mouse skin two-stage carcinogenesis inhibition test was performed under the following conditions.

15 ICR female mice per group (6 weeks old)
24 hours after shaving the back body hair of 7,12-dimethylbenz [α] anthracene (hereinafter abbreviated as DMBA) dissolved in acetone (0.1 ml) on the back skin.
(100 μg, 390 nmol) was applied for initiation, and one week later, each experimental group was treated as follows.

Group 1: Promotion is carried out by applying TPA (1 μg, 1.7 nmol) dissolved in acetone (0.1 ml) twice a week for 20 weeks. At this time, T
One hour before PA application, acetone (0.1 ml) is applied to the same site (positive control group). Group 2: As in Group 1, twice weekly for 20 weeks
One hour prior to the application of PA (1 μg, 1.7 nmol), 50 μg of a test sample paradicom paprika extract (extracted with methanol) dissolved in acetone (0.1 ml) is applied.

Start of promotion by TPA application 20
Until the week after, papillomas generated on the backs of the mice were observed every week, and the ratio of mice having papillomas generated and the average number of papillomas generated per mouse were evaluated in comparison with the positive control group. The results are shown in FIGS.

As a result of the test, as shown in FIG. 2, in the positive control group, the first tumor was formed 7 weeks after the start of the promotion, and the tumor was formed in all mice at the 11th week. In each of the two groups (methanol extract-treated group), tumor formation was observed for the first time at 9 weeks, and it was revealed that tumor formation was delayed. In addition, even after 20 weeks at the end of the test, no tumor formation was observed in 20% of the mice in the second group.

As is apparent from FIG.
The average number of tumors per mouse was 9.1 in the positive control group after 20 weeks, whereas it was 4.5 in the second group, indicating a 50% tumor-suppressing effect.

[0028]

According to the present invention, as shown in the embodiments,
An excellent anti-Epstein-Barr virus genome inactivator derived from nature can be provided. Therefore, the novel anti-Epstein-Barr virus genome inactivator according to the present invention can be expected to have anti-carcinogenic and anti-viral effects, and can be used in a wide variety of applications depending on the fields of pharmaceuticals, cosmetics, and health foods. Becomes

[Brief description of the drawings]

FIG. 1 is a separation chart of liquid chromatography.

FIG. 2 is a graph showing the relationship between the elapsed weeks after the start of the experiment and the percentage of papilloma-bearing mice.

FIG. 3 is a graph showing the relationship between the elapsed weeks after the start of the experiment and the number of papillomas generated in mice.

[Explanation of symbols]

1. HPL against β-carotene in the extract of the present invention
C peak 2 HPLC peak for cryptoxanthin in the extract of the present invention 3 HPLC peak for lutein in the extract of the present invention 4 HP for zeaxanthin in the extract of the present invention
LC peak 5 H to Cryptocapsin in extract of the present invention
PLC peak 6 HPLC peak for anthraxanthin in the extract of the present invention 7 HPLC peak for trans-capsanthin in the extract of the present invention 8 HPLC peak for cis-capsanthin in the extract of the present invention 9 In the extract of the present invention HPLC peak for cis-capsorbin in the extract of the present invention 10 HPLC peak for trans-capsorbin

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification code FI A61P 35/00 A61P 35/00 (72) Inventor Haruhuni Tokuda 3 Shimogamo Kitazonocho, Sakyo-ku, Kyoto-shi, Kyoto (72) Inventor Yoshihiro Ito 37-3 Takaya Takeyacho, Sakyo-ku, Kyoto-shi, Kyoto (56) References Agric. Food Chem. , Vol. 40, no. 11, pp. 2072-2076, 1992 J.P. Agric. Food Chem. , Vol. 44, no. 3, pp. 711-716, 1996 Biol. Pharm. Bull. , Vol. 18, No. 2, pp. 227-233, 1995 Cancer Letters, Vol. 100, pp. 211-214, 1996 (58) Fields investigated (Int. Cl. ⁷ , DB name) A61K 35/78 A61K 7/00 BIOSIS (DIALOG) CA (STN)

Claims (11)

(57) [Claims] 1. An oncogenic virus activation inhibitor using an extract of paradicham paprika. 2. An oncovirus activation inhibitor comprising, as an active ingredient, an extract extracted from acetone extracted from paradicom paprika. 3. A plant extract having an inhibitory effect on the activation of oncogenic virus, which is extracted from Paradigm paprika. 4. A carcinogenesis inhibitor using an extract of paradicham paprika. 5. A carcinogenesis inhibitor comprising, as an active ingredient, an extract of paradicom paprika extracted with acetone. 6. A plant extract having a carcinogenesis-suppressing activity, which is extracted from paradicham paprika. 7. A carcinogenesis inhibitor comprising, as an active ingredient, a hexane extract of paradichompaprika. 8. A carcinogenesis inhibitor comprising, as an active ingredient, a methanol extract of paradicom paprika. 9. Paradigm paprika is a paradigm paprika original, pure strain, F1, quaternary hybrid, F2
Claim 1, wherein said species is any of the following species:
9. The carcinogenesis inhibitor according to 2, 4, 5, 7 or 8. 10. The paradise paprika is a paradise paprika original, pure strain, F1, quaternary hybrid, F
4. A hybrid of any of two or more hybrids.
Or the plant extract according to 6. 11. A source contained in paradigm paprika.
Cosmetics containing an extract component for cancer suppression .