JP5704544B2 - Immunostimulants, cancer cell growth inhibitors, anti-inflammatory agents, or antioxidants containing Cordyceps as an active ingredient - Google Patents

Description

  The present invention relates to an immunostimulant, a cancer cell growth inhibitor, an anti-inflammatory agent, or an antioxidant containing Cordyceps as an active ingredient.

Cordyceps has been prized as a secret medicine since ancient China. In recent years, the effect and component analysis of Cordyceps as a secret medicine have been performed. In addition, the use as a health food is also advancing due to changes in lifestyle and increased health consciousness.
Cordyceps is often used as a collective term for mushrooms that parasitize insects, but is positioned as a genus of Ascomycetes, Bacchiaceae, and Bacchiaceae.
It is said that the classification system has not been established yet, but the host that parasitizes differs depending on the fungus species.For the identification of cordyceps, not only the form of the fruiting body that is formed, but also the type of insect that becomes the host It has been helpful.
For example, the black-bellied semi-bamboo (scientific name Cordyceps, sinclairii, Kobayasi) is called the black-headed bovine, and the red-spotted bamboo (scientific name Torrubiella sp.
In addition, there are cordyceps, which are mainly dragonflies, bees, spiders and mites.
For example, Cordyceps militaryis is known to have an immunostimulatory effect using cordycepin as an index, and Futakuso bamboo, which is parasitic on Tsukutsukuboushi, is known to have an immunosuppressive effect of FTY720. Different activities have been found.
In Non-patent Document 1, the report was made on the active ingredients of the Japanese bamboo shoots, the black-bellied semi-bamboo shoots, the Japanese bamboo shoots, the Marmino Aritake (Cordyceps formicarum), and the Ususanagitake (Cordyceps takaomontana) in a medium that does not contain parasite components. ing.
Further, Patent Document 1 discloses that the extract of cultured mycelia of Sanagitake has a cardiotonic action and bronchodilating action, and Patent Document 2 proposes a method for extracting Cordycepin by paying attention to Sanagitake.
In Patent Documents 3, 4, and 5, effective substances are found by paying attention to the bamboo shoot.
Cordyceps sinensis, which is prized as a Chinese herbal medicine, has not reached fruiting body formation by artificial culture, and as a substitute, many artificial culture methods for Cordyceps sinensis have been proposed. ing.
In Patent Document 6, the mycelia of Cordyceps or Aspergillus spores are sterilized in a culture solution in which garlic, soy sauce, and sugar, which are existing foods, are mixed, and liquid culture is performed by a conventional method, or this liquid culture solution is cultivated in rice, wheat, There has been proposed a method of solid culture by adsorbing to cereals such as corn or adults, moths and larvae of insects of arthropods such as moths and cicadas.
In Patent Document 7, mycelia of Cordyceps sinensis are mainly composed of one or several types of sugars, protein substances, vitamins, nucleic acids, etc., and grains are added to these main components, transplanted to a solid solid medium and cultured. The mycelium cultured in this three-dimensional solid medium is mainly composed of one or several kinds of glycoprotein substances, vitamins, nucleic acids, etc., and one or several kinds of amino acids are added thereto, and water is used as a substrate. There has been proposed a method of transplanting to a liquid medium having a pH of 4.0 to 7.0 and culturing it statically to form bacterial loci on the surface of the liquid medium.
In Patent Document 8, the mycelium of Cordyceps sinensis is directly infected and inoculated into living insects, or the mycelium of Cordyceps sinensis is put on a pure isolation medium using agar with a third amount of insect tissue added. It is a method of inoculation. In addition, when using a living insect, the cocoon just before forming a cocoon is used.
Patent Document 9 proposes a method in which cordyceps present in the field is collected, mycelia and conidia are grown on an agar medium and liquid medium, and final mass growth is performed in field cultivation.
Patent Document 10 proposes a method for culturing persimmon persimmon itself as a medium, and Patent Document 11 proposes a culture method that is closer to nature by using live persimmon as a medium.
Moreover, in patent document 12, it proposes manufacturing the caterpillar with a high effect by making the culture medium which manufactures the cordyceps edible vinegar contain vinegar and vinegar mash.

JP-A-8-12588 JP 2004-8938 A JP 2002-272267 A JP 2003-128515 A JP 2005-239585 A JP-A-54-80486 Japanese Patent Publication No. 61-53033 JP 62-107725 A JP-A-6-233627 Japanese Patent No. 2676502 Japanese Patent No. 3593429 JP 2003-116342 A

Nobuo Yabuchi “Culture and Physiological Activity of Cordyceps”, March 29-31, 2005, 125th Annual Meeting of the Pharmaceutical Society of Japan

As shown in Non-Patent Document 1, Patent Document 1 to Patent Document 5, the difference in pharmacological action according to the type of Cordyceps fungus is currently focused on and research is ongoing.
And even in the case of Cordyceps sinensis, whose host is the same Lepidoptera, the difference in pharmacological action has been studied for different types of Cordyceps species such as Sanatake and Sasatake.
In addition, as shown in Patent Document 6 to Patent Document 12, attention has been paid to the difference in pharmacological effect due to the difference in culture medium, but it is only a difference in medium components, a difference between insects and cereals, This was due to the difference in the growth stage, or whether the insect body was alive or dead, and there was no idea focusing on the host moth species, and no research was conducted.
The inventors of the present invention have various characteristics in the parasitic species Cordyceps, and there are various varieties and strains in the host moth. Therefore, the inventors conducted research focusing on the combination of the parasitic species and the host. It was.
In particular, Koishimaru, which is used for silk fabrics in Shosoin in Japan, has a sufficient historical basis for differentiation from a global perspective as well as in Japan. Breeding continues. Furthermore, another characteristic of “Koishimaru” is that the produced raw silk has a fineness and a good feel, and larvae are considered to have excellent disease resistance. Therefore, from the historical value and biological evaluation of “Koishimaru”, we thought that “Koishimaru” was highly significant as a parasitic species of cordyceps.

  An object of the present invention is to find a cordyceps that shows excellent activity depending on the moth species even with cordyceps of the same bacterial species.

The immunostimulant of the present invention according to claim 1 is characterized by comprising, as an active ingredient, Cordyceps sinensis or an extract of Cordyceps sinensis, which is obtained by using Koishimaru as the culm seed and mainly cultivating the Koishimaru.
The cancer cell growth inhibitor of the present invention according to claim 2 is characterized by comprising, as an active ingredient, Cordyceps sinensis or an extract of Cordyceps sinensis, which is obtained by using Koishimaru as a soy seed and mainly cultivating the Koishimaru.
The anti-inflammatory agent of the present invention according to claim 3 is characterized by comprising, as an active ingredient, Cordyceps or an extract of Cordyceps cultivated by using Koishimaru as a cultivar.
The antioxidant of the present invention according to claim 4 is characterized by comprising, as an active ingredient, Cordyceps or an extract of Cordyceps cultivated by using Koishimaru as the culm seed.

  According to the present invention, even with the same kind of Cordyceps fungus, excellent activity was recognized when Koishimaru was used as the culm species.

Table showing the yield of extract from each cordyceps of the present invention The graph which shows the growth inhibitory activity with respect to the cancer cell by Example 1 (Konasanagitake (Koishimaru)) of this invention. The graph which shows the growth inhibitory activity with respect to the cancer cell by Example 2 (Hanasanagitake (Koishimaru)) of this invention. The graph which shows the growth inhibitory activity with respect to the cancer cell by the comparative example 1 (Konasanagitake (practical variety)) The graph which shows the growth inhibitory activity with respect to the cancer cell by the comparative example 2 (Hanasanagitake (practical variety)) The graph which shows the immunostimulatory activity with respect to the lymphocyte by Example 1 (Konasanagitake (Koishimaru)) of this invention. The graph which shows the immunostimulatory activity with respect to the lymphocyte by Example 2 (Hanasanagitake (Koishimaru)) of this invention. Graph showing immunostimulatory activity against lymphocytes by Comparative Example 1 (Konasanagitake (practical variety)) The graph which shows the immunostimulatory activity with respect to the lymphocyte by the comparative example 2 (Hanasanagitake (practical variety)) The graph which shows the antioxidant activity by Example 1 (Konasanagitake (Koishimaru)) of this invention The graph which shows the antioxidant activity by Example 2 (Hanasanagitake (Koishimaru)) of this invention Graph showing the antioxidant activity of Comparative Example 1 (Konasanagitake (practical variety)) The graph which shows the antioxidant activity by the comparative example 2 (Hanasanagitake (practical variety)) The graph which shows the antioxidant activity by Example 1 (Konasanagitake (Koishimaru)) of this invention The graph which shows the antioxidant activity by Example 2 (Hanasanagitake (Koishimaru)) of this invention

1. Sample In the present invention, the host cultivar, Koishimaru, was used as a comparative example, which was a practical variety (Kinkiaki x Kanwa) popular in Japan. As parasitic species, Paecilomyces farinosus and Paecilomyces tenuipes were used.
Cordyceps sinensis (Rhizopus japonica) and pickled cicada take out the cocoon after the cocoon has formed a cocoon, and inoculate the taken cocoon with the ascospore or conidia formed on the fruiting body of the cordyceps so that the fruiting body grows completely What was cultured until was used. The Cordyceps sinensis in which the fruiting bodies were completely grown was dried at a temperature of about 60 ° C., then pulverized with a blender (WB-1, φ134 × 270 mm, ASONE Co., Ltd.) and used in the experiment. The dried cordyceps was stored at 4 ° C., and the powdered one was immediately extracted.
In the following explanation, Konasanagitake, which uses the host as the practical variety, is Konasanagitake (practical variety), Konasanatake, which uses the host as Koishimaru, Konasanagitake (koishimaru), and The hanasanagitake that is called Koishimaru is called the hanasanagitake (Koishimaru).

2. Extraction method of physiologically active fractions 200 g was used for Konasa Gitake (practical variety), 168 g was used for Prunus charcoal (practical variety), and 100 g was used for each Peach bamboo (Koishimaru) and Hanasa Nagatake (Koishimaru). After confirming that the moisture content in the sample was around 10% with a weighing bottle, sequential extraction was performed. For extraction, 3 times the amount of solvent was added to the sample, left to stand at 25 ° C. for 24 hours, and the supernatant obtained by filtration was used as the extract. According to the sequential extraction method, n-hexane extraction, ethyl acetate extraction, 70% acetone (30% water) extraction, and water extraction were performed in the order of low polarity, and each was divided into n-hexane fraction, ethyl acetate fraction, 70% Acetone fraction and water fraction were used for various bioassays. The extraction was performed twice.
The n-hexane extract and the ethyl acetate extract are concentrated using a rotary evaporator (N-1000, EYELA), further air-dried with nitrogen and concentrated using a centrifugal evaporator (C-100, EYELA), and then freeze-dried (FDU810). , EYELA) and lyophilized to determine the yield. The 70% acetone extract was concentrated by a rotary evaporator, then concentrated by a centrifugal evaporator, and freeze-dried to obtain a yield. The aqueous extract was freeze-dried to determine the yield. Hereinafter, these lyophilized products were used as crude extracts for each extraction.

3. Bioassay method 1) Cancer cell growth inhibitory activity test Cancer cell growth inhibitory activity was examined by the MTT method, which is a reduced coloring dye. MTT {3- (4,5-Dimethyl-2-thiazolyl) -2,5-diphenyl-2H-tetrazole bromide} is reduced by intracellular dehydrogenase to produce formazan. The number of cells and the amount of formazan produced are linearly proportional, and the number of viable cells can be measured by measuring the absorbance of formazan (Oka et al., 1992). According to this method, an experiment was performed according to the following procedure.

1) Cell and medium composition Activity was measured using rat hepatoma cells (dRLh84), and the increase in the number of cells was used as an index. As the medium, Dulbecco's modified Eagle medium (DMEM, Nippon Pharmaceutical Co., Ltd.) containing 10% newborn calf serum (NBS), 4 mM glutamine, 50 U / ml penicillin, 50 μg / ml streptomycin, 100 μg / ml kanamycin was used. Incubation was performed under humid conditions at 37 ° C. in the presence of 5% CO ₂ .

2) Preparation of cell suspension The cells were cultured in a tissue culture dish (φ100 × 20 mm, Greiner). After removing the culture solution in the logarithmic growth phase, PBS (−) {phosphate-buffered saline (−)} (137 mM) NaCl, 2.7 mM KCl, 2 mM Na ₂ HPO ₄ · 12H ₂ O, 1.5 mM KH ₂ PO ₄ ). Thereafter, 2.5% trypsin was added to suspend cells attached to the dish. In order to make the cell concentration in the dish 5 × 10 ⁴ cells / mL, 10 ml of a medium was added to obtain a cell suspension.

3) Measurement of cancer cell growth inhibitory activity The cell suspension was diluted 20-fold, and 200 μl (microliter) was added to each well of a 96-well microplate (Asahi Techno Glass Scitech). The cells were cultured for 24 hours under humid conditions at 37 ° C. in the presence of 5% CO ₂ . Thereafter, 1 μl of each adjusted crude extract was added to the experimental group, and 1 μl of solvent was added to the control group instead of the sample. After culturing under the same conditions for 48 hours after the addition of the sample, the medium was replaced with 100 μl of a medium containing 0.55 mg / ml MTT and cultured for 4 hours. Thereafter, the medium containing MTT was removed, 200 μl of Dimethylsulfoxide (DMSO) was added, and the absorbance was measured at two wavelengths of 590 nm and 620 nm with a microplate reader (Immuno-Mini NJ-2300, Intermed). All reagents and instruments used were sterilized. The experiment was performed in a clean bench to prevent contamination by other microorganisms.

2) Immunostimulatory activity test The immunostimulatory activity was determined using the activity of lymphocyte cells derived from mouse spleen and changes in the number of cells as indicators. For the measurement of the activity and the number of cells, the amount of formazan salt produced by the reducing action of the intracellular mitochondria was measured using the WST-1 method, which is a reducing chromogenic reagent.
The WST-1 method is reduced by intracellular dehydrogenase to produce water-soluble formazan. In this experiment, the activity of the cells was measured by measuring the absorbance of this water-soluble formazan.

1) Experimental Animal and Medium Composition As experimental animals, 5-7 week-old ICR mice (female, Japan SLC) were used.
The lymphocyte medium was RPMI 1640 medium (Nissui Pharmaceutical Co., Ltd.) containing 10% fetal bovine serum (FBS), 2 mM glutamine, 50 μM 2-mercaptoethanol, 100 U / ml penicillin, 100 μg / ml streptomycin. Culturing was performed under humid conditions at 37 ° C. in the presence of 5% CO ₂ .

2) Preparation of lymphocyte suspension The method of Fujiwara and Sakurai (1996) was used to adjust the lymphocyte suspension.
Mice anesthetized with diethyl ether were dislocated from the cervical spine and the spleen was removed. The spleen was ground in a dish containing PBS (−), filtered through a cell strainer (pore size 100 μm, FALCON), and the cell suspension was collected in a 50 ml conical tube (FALCON). After centrifugation (1,100 rpm, 10 min), the supernatant is removed, and a hemolysis buffer {0.14 M ammonium chloride, 1.7 mM Tris-HCl buffer (pH 7.56)} is used to remove red blood cells from the lymphocyte-containing precipitate. Was added and incubated for 5 minutes. Thereafter, centrifugation (1,100 rpm, 10 min) was performed, and the supernatant was removed. This was repeated twice to remove erythrocytes by hemolysis. Further, the supernatant was removed, 30 ml of PBS (−) was added for washing, and centrifugation (1,100 rpm, 1 min) was performed twice. Thereafter, a medium was added to the precipitate and incubated for 2 hours in a CO ₂ incubator to remove cells other than lymphocytes.
After incubation, the cells were stained with 0.4% trypan blue solution, and the number of cells was counted using a Bürkerturk hemocytometer. And it adjusted with the culture medium so that the cell number might be set to 5x10 < ⁶ > cells / ml.

3) Measurement of immunostimulatory activity The prepared lymphocyte suspension 200 μl was added only to the well having the highest concentration in each test section. To other wells including the control group, 100 μl of lymphocyte suspension prepared so that the concentration of the solvent was 0.5% was added to each well. Each crude extract was added only to the portion with the highest assay concentration, and 100 μl was taken from the well with one step higher concentration to other wells and diluted by adding to the next well. After each crude extract was added, the cells were cultured for 48 hours in the presence of 5% CO ₂ at 37 ° C. wet conditions. Then, WST-1 solution {2- (4-lodophenyl) -3- (4-nitrophenyl) -5- (2,4-disulfophenyl) -2H-tetrazolium, monosodium salt} (Wako Pure Chemicals) and 1-Methyl PMS 10 μl of WST-1 solution (6.5 mg WST-1, 0.7 mg 1-methyl PMS, PBS (−) 10 ml) consisting of (1-Methoxy-5-methylphenylmethylsulfate) (Wako Pure Chemicals), PBS (−) After addition and incubation in a CO ₂ incubator for 4 hours, the absorbance of each well at 450 nm was measured with a microplate reader. All reagents and instruments used were sterilized in an autoclave (120 ° C., 20 minutes). The experiment was performed in a clean bench to prevent contamination by other microorganisms.

3) Antioxidant activity test In the test for measuring antioxidant activity, DPPH (1,1-Diphenyl-2-picrylhydryl) spectroscopy (Yamaguchi et al., 2000) was used, and the amount of residual radicals in the DPPH solution was measured in black. The radical scavenging ability was measured by spectroscopic analysis of purple fading.
Since DPPH (1,1-Diphenyl-2-picrylhydrazyl) is a stable free radical and exhibits a black-purple color, the amount of radicals remaining in the DPPH solution is analyzed spectroscopically. , Radical scavenging ability can be measured.
As a positive control, ascorbic acid, which has been confirmed to have a high antioxidant activity, was used (Arabshahi-Delouee and Uroj, 2006).
In each well of a 96-well microplate, 20 ml of a sample adjusted to each concentration, 80 ml of 0.1 M Tris-HCl buffer (pH 7.4) and 100 ml of 500 mM DPPH solution dissolved in ethanol are dispensed, and 20 minutes at room temperature. After standing, the absorbance at a wavelength of 517 nm was measured by a multipurpose microplate reader Power scan HT (Dainippon Sumitomo Pharma Co., Ltd.) to quantify the amount of residual DPPH radicals. The radical scavenging activity was calculated by the following formula using each absorbance.
DPPH radical scavenging activity (%) = (1− (absorbance of sample−blank value)) / (absorbance of control−blank value) × 100

4) Anti-inflammatory activity test For the anti-inflammatory activity test, a method (Colorometric COX Inhibitor Screening Assay) introduced by Okudera (2005) was used.
There are two isomers of cyclooxygenase (COX). COX-1 is constantly expressed in various cells, particularly expressed in the gastrointestinal tract, kidneys and platelets, and maintains normal tissue metabolism. On the other hand, COX-2 produces prostaglandin E2 (PGE2) and the like, and expands the inflammatory reaction. Therefore, in this study, in order to search for a selective inhibitor of COX-2, an experiment was performed using the Corimetric Cox (ovine) Inhibitor Screening Assay kit. This method uses COX oxidation ability as an index, and oxidizes the chromogenic substrate N, N, N ′, N ′,-tetramethyl-p-phenylenediamine (TMPD) and measures the absorbance at 590 nm.

1) Measurement of anti-inflammatory activity Assay Buffer (10 ×) was diluted 10-fold with MQ, and 160 μl was added to each well of the 96-well microplate for the negative control group for cell culture and 150 μl for the positive control group and experimental group. Noted. Heme was diluted to 88 μl at a ratio of 1.912 ml of the prepared Assay Buffer, and 10 μl was dispensed into each well. COX has two active sites, a cyclooxygenase active site and a peroxygenase active site, and Heme is required to activate the latter. COX-1 and COX-2 were each diluted at a ratio of 400 μl of the prepared Assay Buffer to 200 μl of the enzyme, and 10 μl was dispensed into each well of the positive control group and the experimental group. The sample was adjusted to a final concentration of 100 μg / ml, 10 μl was added to each well of the experimental group, a solvent was added to the negative / positive control group, the plate was gently shaken horizontally, and 5 ° C. at 25 ° C. Left to stand. Meanwhile, 100 μl of potassium hydroxide was added to 100 μl of arachidonic acid and mixed by inversion, and MQ was added at a rate of 1.8 ml. 20 μl of TMPD solution and 20 μl of prepared arachidonic acid were added to each well in this order. After leaving still at 25 degreeC for 5 minutes, the light absorbency in 590 nm of each well was measured with the microplate reader.

Yield of Each Crude Extract from Cordyceps FIG. 1 is a table showing the yield of extract from each Cordyceps of the present invention.
As shown in FIG. 1, from 200 g of Konasanagitake hosted with a practical variety, 5.8% (11.645 g) in the n-hexane extract fraction, 2.1% (4.175 g) in the ethyl acetate fraction, The recovery rate was 13% (25.977 g) for the 70% acetone fraction and 18% (35.992 g) for the water fraction. In addition, from 100 g of the same Kosanagitake with Koishimaru as the host, it was 3% in the n-hexane extract fraction, 0.5% in the ethyl acetate fraction, 13.5% in 70% acetone, and 25.9% in the water fraction. Met. In addition, from 168 g of hanasanagitake, which is the main varieties, from n-hexane extracted fraction, 13.98% (23.3837 g), ethyl acetate fraction, 0.81% (1.3645 g), and 70% acetone fraction. The recovery rate was 12% (20.2655 g), and the water fraction was 33.4% (56.122 g). Furthermore, in the case of the Japanese bamboo shoots with Koishimaru as the host, n-hexane extract fraction was 6.3%, ethyl acetate fraction was 0.6%, 70% acetone was 9.4%, and water fraction was 33.5%. Met.
From the results of this experiment, the ethyl acetate fraction was 4 to 5 times more recovered than Koishimaru in practical varieties, and conversely, the water fraction was 1.6 times more recovered than Koishimaru. That is, it is considered that the yield of each extracted fraction differs depending on the host silkworm cultivar rather than the species of cordyceps.

Cancer Cell Growth Inhibitory Activity Test FIG. 2 is a graph showing the growth inhibitory activity against cancer cells according to Example 1 of the present invention (Konasanagitake (Koishimaru)), and FIG. 3 is according to Example 2 of the present invention (Hanasanagitake (Koishimaru)). FIG. 4 is a graph showing growth inhibitory activity against cancer cells, FIG. 4 is a graph showing growth inhibitory activity against cancer cells according to Comparative Example 1 (Konasanatake (practical variety)), and FIG. 5 is cancer according to Comparative Example 2 (Hanasanatake (practical variety)). It is a graph which shows the growth inhibitory activity with respect to a cell.
As shown in the figure, the ethyl acetate fraction showed the strongest cancer cell growth inhibitory activity in any combination.
Compared with Konasanagitake, the cancer cell growth inhibitory activity is superior to that of Example 1, in which the IC ₅₀ value of the ethyl acetate section of Comparative Example 1 is about 80 μg / ml, which is about 45 μg / ml. . Further, even when compared with Japanese bamboo shoots, Comparative Example 2 is about 50 μg / ml, while Example 2 is about 35 μg / ml, indicating superiority.
Therefore, it can be seen that when Koishimaru is the host, the activity is superior to that of the practical variety. In addition, when the host is the same, it can be seen that the activity of the bamboo shoot is stronger than that of the bamboo shoot.

Immunostimulatory activity test FIG. 6 is a graph showing immunostimulatory activity against lymphocytes according to Example 1 of the present invention (Konasanagitake (Koishimaru)), and FIG. 7 is a graph showing lymphocytes according to Example 2 of the present invention (Ohanasanatake (Koishimaru)). FIG. 8 is a graph showing the immunostimulatory activity, FIG. 8 is a graph showing the immunostimulatory activity against lymphocytes of Comparative Example 1 (Konasanatake (practical variety)), and FIG. 9 is the immunity against lymphocytes according to Comparative Example 2 (Practus varieties). It is a graph which shows activation activity.
As shown in the figure, immunostimulatory activity was confirmed in Example 1 and Example 2, whereas immunostimulatory activity was not confirmed in Comparative Example 1 and Comparative Example 2. Comparing the active water extracts, when the concentration of the extract was 500 μg / ml, it was about 140% for Japanese bamboo shoots and about 220% for Japanese bamboo shoots.
From this result, it can be considered that the water extract of the pine needles of Koishimaru has the most excellent immunostimulatory activity.

Antioxidant Activity Test FIG. 10 is a graph showing the antioxidant activity according to Example 1 of the present invention (Konasanagitake (Koishimaru)), and FIG. 11 is a graph showing the antioxidant activity according to Example 2 of the present invention (Ohanasanagitake (Koishimaru)). FIG. 12 is a graph showing the antioxidant activity according to Comparative Example 1 (Konasanatake (practical variety)), and FIG. 13 is a graph showing the antioxidant activity according to Comparative Example 2 (Pterocarpus (practical variety)).
Antioxidant activity was confirmed in the 70% acetone extract fraction and the water extract fraction. When Example 1 was compared with Comparative Example 1, it was confirmed that the activity with both the 70% acetone extract and the water extract was higher when the host was Koishimaru.

Anti-Inflammatory Activity Test FIG. 14 is a graph showing the antioxidant activity of Example 1 of the present invention (Konasanagitake (Koishimaru)), and FIG. 15 is a graph of the antioxidant activity of Example 2 of the present invention (Ohanasanatake (Koishimaru)). It is.
Anti-inflammatory activity was observed in the ethyl acetate extract fraction, and higher activity was found in the bamboo shoot than in the bamboo shoot.

  INDUSTRIAL APPLICABILITY The present invention is suitable for cordyceps used as foods, additives, skin external preparations, and bath preparations.

Claims (4)

  An immunostimulant characterized by comprising, as an active ingredient, Cordyceps edible grass, which is obtained by using Koishimaru as a seed, and mainly cultivating the Pebbles maru.   A cancer cell growth inhibitor comprising, as an active ingredient, a cordyceps or an extract of cordyceps, which is obtained by cultivating the above-mentioned koishi-maru as a seed.   An anti-inflammatory agent comprising, as an active ingredient, cordyceps or an extract of cordyceps that has been cultivated mainly using koishimaru as a cocoon seed.   An anti-oxidant characterized in that it uses Koishimaru as a pod seed and contains, as an active ingredient, Cordyceps sinensis or an extract of Cordyceps sinensis cultivated mainly by Koishimaru.