ANTI-INFLUENZA VIRUS COMPOSITION FOR MUCOUS MEMBRANES

Abstract

The present invention relates to an anti-influenza virus composition for mucous membranes, comprising a fermented ginseng extract or fermented red ginseng extract as an active component.

Claims

1. A method for preventing or treating an influenza virus-caused disease, wherein the method includes a step of mucosal administering a pharmaceutical composition comprising a fermented ginseng or fermented red ginseng as an active component to a subject in need thereof.

2. The method according to claim 1, wherein the fermented ginseng or fermented red ginseng is prepared by means of a two-step fermentation of lactic-acid fermentation and enzymatic fermentation.

3. The method according to claim 2, wherein the lactic acid bacteria are at least one lactic acid bacterium selected from the group consisting of Lactococcus, Lactobacillus, Leuconostoc, Propionibacterium, Enterococcus, Bifidobacterium, Streptococcus and Pediococcus.

4. The method according to claim 3, wherein the lactic acid bacteria are at least one lactic acid bacterium selected from the group consisting of Lactobacillus alimentarius, Lactobacillus sakei, Lactobacillus acidophilus, Lactobacillus casei, Lactobacillus gasseri, Lactobacillus delbrueckii, Lactobacillus fermentum, Lactobacillus bulgaricus, Lactobacillus helveticus, Leuconostoc mesenteroides, Streptococcus thermophilus, Streptococcus lactis, Enterococcus faecium, Enterococcus faecalis, Bifidobacterium bifidum, Bifidobacterium infantis, Bifidobacterium brave and Bifidobacterium longum.

5. The method according to claim 4, wherein the lactic acid bacteria are Lactobacillus alimentarius M-2 strain (KCTC 11054 BP) and Leuconostoc mesenteroides M-3 strain (KCTC 11055 BP).

6. The method according to claim 2, wherein the enzyme is at least one enzyme selected from the group consisting of pectinase, cellulase, hemicellulase, xylanase, pectolyase, pectinesterase and laminarinase.

7. The method according to claim 1, wherein the fermented ginseng or fermented red ginseng is contained in the composition in an amount of 0.1 to 50 w/v %.

8. The method according to claim 1, wherein the influenza virus is an influenza virus type A.

9. The method according to claim 8, wherein the influenza virus type A is at least one selected from the group consisting of H1N1, H5N1 and H3N2.

10. The method according to claim 1, wherein the mucous membranes are nasal mucous membranes, oral mucous membranes or airway mucous membranes.

11. The method according to claim 1, wherein the composition is in a form of spray, powder, gel, ointment or drop.

12. The method according to claim 1, wherein the subject is animal.

13. The method according to claim 1, wherein the disease is at least one selected from the group consisting of cold, flu, cough, sneeze, runny nose, muscle pain, sore throat, rhinocleisis, laryngitis, neckache, hoarseness, headache, pain in paranasal sinuses, rhinitis, pharyngitis, bronchitis, asthma, fever, dyspnea, general lethargy and chill.

14-15. (canceled)

16. A method for preparing fermented ginseng or fermented red ginseng, wherein the method includes: a step of performing a lactic-acid fermentation for a ginseng or red ginseng by using Lactobacillus alimentarius M-2 strain (KCTC 11054 BP) and Leuconostoc mesenteroides M-3 strain (KCTC 11055 BP); and a step of performing an enzymatic fermentation by using pectinase, cellulase and hemicellulase.

17. A hygiene product, which contains an anti-influenza virus composition for mucous membranes comprising fermented ginseng or fermented red ginseng as an active component, or is coated therewith.

18. The hygiene product according to claim 17, wherein the hygiene product is at least one selected from the group consisting of soap, wet tissue, tissue, shampoo, mouth freshener, air freshener, cleansing gel and anti-influenza virus spray.

19. The hygiene product according to claim 18, the hygiene product is the anti-influenza virus spray, which is filled with the anti-influenza virus composition to spray to mucous membranes.

20. The hygiene product according to claim 19, wherein the mucous membranes are nasal mucous membranes, oral mucous membranes or airway mucous membranes.

21. The hygiene product according to claim 19, wherein the spray is an aerosol spray, push-to-type spray or nebulizer.

22. (canceled)

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0097] FIG. 1 shows an anti-viral protective effect of fermented red ginseng specimens A and B on H5N1 influenza virus in a mouse infection model (CD4 C57BL/6). FIG. 1 shows a change in body weights of a mouse group (n=5), which was given a mixture of the fermented red ginseng specimen A (or B) (250 μg, 1 w/v % aqueous solution) and H5N1 influenza virus. A “non-fermented red ginseng sample” means a control group, which was given a mixture of a non-fermented red ginseng extract (500 μg, 1 w/v % aqueous solution) and H5N1 influenza virus.

[0098] FIG. 2 shows a high anti-viral activity of the fermented red ginseng specimen A against H5N1 and H3N2 influenza viruses. FIG. 2 shows a change in body weights of mice (n=5, CD4 C57BL/6), which were given a mixture of influenza virus (H5N1 or H3N2) and different amounts of the fermented red ginseng (specimen A or B).

[0099] FIGS. 3A and 3B show an inhibitory effect of the fermented red ginseng specimen on a replication of H5N1 influenza virus. FIGS. 3A and 3B show a change in body weights of a mouse group (n=5, CD4 C57BL/6), which was given the mixture of H5N1 influenza virus and the fermented red ginseng specimen A or B, as well as results of analyzing a virus concentration in lung on the 6th day. “H5N1 only” means a control group, which was given the virus only without the red ginseng specimen.

[0100] FIGS. 4A to 4C show an inhibitory effect of the fermented red ginseng specimen A on a production of inflammatory cytokines caused by H5N1 influenza virus. FIGS. 4A to 4C show results of analyzing inflammatory cytokines in lung and bronchoalveolar lavage fluid (BALF) of a mouse group (n=5, CD4 C57BL/6), which was given the mixture of H5N1 influenza virus and the fermented red ginseng specimen A or B, on the 6th day. “H5N1 only” means a control group, which was given the virus only without the red ginseng specimen.

[0101] FIG. 5 shows an inhibitory effect of the fermented red ginseng specimen A on an induction of lung histopathology caused by H5N1 influenza virus. A histopathological analysis was performed by extracting a lung of a mouse group (n=5, CD4 C57BL/6), which was given the mixture of H5N1 influenza virus and the fermented red ginseng specimen A or B, on the 6th day. “Red ginseng A” and “Red ginseng B” show results of mice, which were given the mixture of the fermented red ginseng specimen A and H5N1 influenza virus, and the mixture of the fermented red ginseng specimen B and H5N1 influenza virus, respectively; “H5N1” means a control group, which was given H5N1 influenza virus only; and a “non-dosed group” is a control group, which was not given the fermented ginseng and virus.

[0102] FIGS. 6A and 6B show a protective effect of the fermented red ginseng specimen A on treatment before or after a virus infection. FIG. 6A shows a change in body weights of mice (CD5 C57BL/6) with regard to treatment with the fermented red ginseng specimen A before or after the infection with H5N1 influenza virus. FIG. 6B shows a pre-treatment effect of the fermented red ginseng specimen A on H1N1 pandemic influenza virus.

[0103] FIG. 7 shows an anti-viral effect of the fermented red ginseng specimen A on H5N1 influenza virus in an immunodeficient mouse. A μMT mouse is a mouse, which is deficient in B cells for producing antibodies. “CD4 KO 250 μg” means a CD4-deficient mouse, which was given 250 μg of the fermented red ginseng specimen A by means of 1 w/v % aqueous solution.

[0104] FIG. 8 shows a view of illustrating an example of an aerosol spray for mucous membranes according to the present invention.

[0105] FIG. 9 shows a view of illustrating an example of a push-to-type oral spray according to the present invention.

[0106] FIG. 10 shows a view of illustrating an example of a push-to-type nasal spray according to the present invention.

MODE FOR INVENTION

[0107] Hereinafter, the present invention will be described in detail through preferred Examples for better understanding of the present invention. However, the following Examples are provided only for the purpose of illustrating the present invention, and thus the present invention is not limited thereto.

Example 1: Preparation of a Fermented Red Ginseng

[0108] Red ginseng (Panax ginseng) provided from Ginseng Nonghyup (national agricultural cooperative federation) was ground through a 20-30 mesh, then put into a fermentation tank with an addition of purified water in an amount equivalent to 20 times more than a weight of the red ginseng, then mixed well, then sterilized by pressurizing at a high temperature of 120° C. and at 1.5 atmospheric pressure for 15 minutes. Then, a temperature of the fermentation tank was cooled down to 37° C. and kept at the same temperature, during which a sterilized mixture of red ginseng powder was inoculated with 1% (v/v) Lactobacillus alimentarius M-2 strain (KCTC11054BP) and Leuconostoc mesenteroides M-3 strain (KCTC11055BP), which are the strains of the Korean Patent Registration No. 10-0856790. After inoculation, fermentation was performed for a pre-determined period of time (12 days for the fermented red ginseng specimen A and 5 days for the fermented red ginseng specimen B), while a temperature of the fermentation tank was kept at 37° C. Then, a temperature of the fermentation tank was raised up to 50° C., then kept at the same temperature, during which a complex enzyme (Citrozym Cloudy, Novozyme) of pectinase, cellulase and hemicellulase, which are the enzymes of the Korean Patent Registration No. 10-0877489, was added 5% (v/v) thereinto, and then was subjected to reaction at 50° C. for a pre-determined time (72 hours for the fermented red ginseng specimen A and 24 hours for the fermented red ginseng specimen B). After finishing the fermentation, the mixture of red ginseng powder was sterilized in a fermentation culture tank at 95° C. for two hours, then put into an extractor with an addition of 70% ethyl alcohol by such an amount as to be five times more than the mixture, and then an extraction was performed at a temperature of 70° C. for eight hours repeatedly three times. A resulting extracted solution was filtered through a 10-50 microfilter, and a filtered extracted solution was vacuum-concentrated at a temperature of 60° C. and under a reduced pressure of 600-700 mmHg such that a solid content may reach 55%, and thus a concentrated solution of the fermented red ginseng (fermented red ginseng specimens A and B) was prepared.

Examples 2-1 to 2-9: Preparation of a Liquid Composition for a Spray Dosage Form of the Fermented Red Ginseng

[0109] The fermented red ginseng specimen A, which was prepared in Example 1, as well as propolis and purified water were weighed to meet the contents described in a following table 1, then put into a ready preparation tank, then stirred for 20 to 60 minutes, and thus a homogenized liquid composition for a spray dosage form for mucous membranes was prepared.

TABLE-US-00001 TABLE 1 Content (in total amount of 100 mL) Example Example Example Example Example Example Example Example Example Component 2-1 2-2 2-3 2-4 2-5 2-6 2-7 2-8 2-9 Fermented 0.1 g 5 g 10 g 30 g 50 g 5 g 5 g 5 g 5 g red ginseng Propolis — — — — — 0.1 g 3 g 10 g 20 g Purified Drop Drop Drop Drop Drop Drop Drop Drop Drop water dose dose dose dose dose dose dose dose dose Total 100 mL 100 mL 100 mL 100 mL 100 mL 100 mL 100 mL 100 mL 100 mL

Examples 3-1 to 3-9: Preparation of an Aerosol Spray Container Including the Fermented Red Ginseng

[0110] As shown in FIG. 8, a spray container including: a container capable of filling; a push-to-type operation button; and an injection nozzle was filled with a liquid composition of Examples 2-1 to 2-9, and thus an aerosol spray container including the fermented red ginseng was prepared.

Examples 4-1 to 4-9: Preparation of a Push-to-Type Oral Spray Container Including the Fermented Red Ginseng

[0111] As shown in FIG. 9, a spray container including: a container capable of filling; a push-to-type operation button; and an injection nozzle was filled with the liquid composition of Examples 2-1 to 2-9, and thus a push-to-type oral spray container including the fermented red ginseng was prepared.

Examples 5-1 to 5-9: Preparation of a Push-to-Type Nasal Spray Container Including the Fermented Red Ginseng

[0112] As shown in FIG. 10, a spray container including: a container capable of filling; a push-to-type operation button; and an injection nozzle was filled with the liquid composition of Examples 2-1 to 2-9, and thus a push-to-type nasal spray container including the fermented red ginseng was prepared.

Experimental Example 1: Analysis of Ginsenoside in the Fermented Red Ginseng

[0113] Methanol was added into a vacuum-dried matter, then filtered, and then analyzed by means of HPLC. The HPLC analysis was performed by using Waters HPLC (600 controller, 717 plus Autosampler, 2487 dual absorbance detector) and a column of ZORBAX Edipse XDB-C18 (4.6×250 mm, 5 micron), wherein an oven temperature of the column was 30° C. and a loading amount of a sample was 10 μl. A mobile phase was obtained by applying a concentration gradient of 100% water and 100% acetonitrile for 75 minutes. A flow rate was 2.5 ml per minute and a detection was made at 203 nm.

[0114] As a result, it might be identified that the fermented red ginseng specimen A and fermented red ginseng specimen B contain ginsenoside F1, F2, protopanaxatriol (PPT), compound K, Rh2 and protopanaxadiol (PPD) components, as shown in a table 2. Such components of the fermented red ginseng specimens A and B are the components specific to the fermented red ginseng, which are not detected from a conventional red ginseng concentrated solution (Cheongkwanjang, Republic of Korea) (Table 2).

TABLE-US-00002 TABLE 2 Fermented Fermented Red ginseng red ginseng red ginseng concentrated Ginsenoside specimen A (mg/g) specimen B (mg/g) solution (mg/g) Rg1 0.60 1.27 1.36 Re 0.49 0.87 1.43 Rf 0.29 0.13 1.02 Rb1 0.95 2.74 6.86 Rc 0.04 0.05 2.76 Rg2 0.65 0.25 1.03 Rh1 0.58 0.19 1.02 Rb2 0.02 0.26 2.46 Rb3 0.01 0.02 0.57 F1 0.28 0.17 — Rd 0.62 0.75 0.82 F2 0.55 0.47 — Rg3 0.62 0.54 1.51 PPT 0.88 0.33 — CK 4.19 1.69 — Rh 1.49 0.43 — PPD 2.13 0.93 — Total 14.39 11.09 20.84  ginsenoside

Experimental Example 2: Anti-Viral Activity of the Fermented Red Ginseng—Identification of Weight and Survival Rate

[0115] <Preparation of Virus and Administration Thereof into a Nasal Cavity of a Mouse>

[0116] As for virus, H5N1, A/Puerto Rico/8/1934 (H1N1; A/PR8) and A/Philippines/82 (H3N2 subtype) were cultured in embryonated hen's eggs by means of a known method (Quan F S et al. (2007) J Virol 81: 3514.524; Song J M et al. (2011) Proc Natl Acad Sci USA 108: 757.61; Song J M et al. (2011) PLoS One 6: e14538; Kim M C et al. (2013) Mol Ther 21: 485.92).

[0117] As for a mouse, a female BALB/c mouse (six weeks old, Harlan Laboratories) was used at a ratio of 5-6 mice per group. For an intranasal administration, the mice were anesthetized with isoflurane and given the fermented red ginseng specimen A or B, and H5N1, A/PR8 H1N1 virus (2.5 LD50) or A/Philippines/82 H3N2 virus (2.5 LD50). The mice infected with virus were observed daily and their body weights and survival rates were recorded. As for a control group, a non-fermented red ginseng extract was used.

[0118] As a result, the fermented red ginseng specimens A and B showed an anti-viral effect on rgH5N1 bird flu virus, as might be identified in FIG. 1. Particularly, as a result of administering a low dose (250 μg/mouse) of the fermented red ginseng specimens A and B into a nasal cavity of each mouse by means of 1 w/v % aqueous solution, mice given a mixture of rgH5N1 virus and the fermented red ginseng specimen A did not show a weight loss, but showed a complete inhibition against a lethal dose of rgH5N1 virus. On the other hand, mice given the fermented red ginseng specimen B showed a slight weight loss, but regained a weight again in nine days later. However, mice given a non-fermented ginseng sample showed a severe weight loss and eventually died all, even if a dosage thereof doubled the fermented red ginseng specimen.

[0119] Also, a degree of anti-viral activity of the fermented red ginseng specimens A and B was identified in FIG. 2. Particularly, in mice given H5N1 and H3N2, a group of the fermented red ginseng specimen A (500 μg or 250 μg, 1 w/v % aqueous solution) did not show a weight loss, and a group of the fermented red ginseng specimen B (500 μg or 250 μg, 1 w/v % aqueous solution) showed a slight weight loss within a range of 5-15%. On the other hand, mice given H5N1 and H3N2 influenza viruses only showed a severe weight loss all.

Experimental Example 3: Anti-Viral Activity of the Fermented Red Ginseng—Analysis of Virus Concentration

[0120] To figure out a protective efficacy of the fermented red ginseng specimens A and B on anti-viral activity, a virus concentration of mouse lungs was analyzed on the 6th day after being infected with H5N1 virus.

[0121] The virus concentration of the lungs was analyzed with MDCK cells by means of a known method (Quan F S et al., J Virol (2008) 82:1350-1359). The lungs were extracted in six days after being infected with influenza virus. Briefly, lung extracts were inoculated by serial dilution into a 6-well plate, which was seeded with MDCK cells in a single layer, and infected at 37° C. for one hour.

[0122] An overlay medium comprising DEAE dextran, non-essential amino acid, glutamine and trypsin was added thereinto, and cultured for two or three days. The cells were fixed with 0.25% glutaraldehyde, then dyed with 1% crystal violet, and then plaque was counted.

[0123] As a result, a pattern of weight changes was similarly observed in the fermented red ginseng specimens A and B (FIG. 3A). On average, 7.4×10.sup.5 PFU (particle forming units) of virus was detected from a lung of the mouse infected with H5N1 virus (FIG. 3B). Mice given the mixture of the fermented red ginseng specimen B and virus showed a significantly low virus concentration of 1.5×10.sup.5 PFU (FIG. 3B). Surprisingly, a mouse group given the fermented red ginseng specimen A showed a virus concentration in lung, which was less than or equal to a detection limit (FIG. 3B). It means that the fermented red ginseng specimen A completely inhibits a virus replication in the lung against H5N1 influenza virus.

Experimental Example 4: Anti-Viral Activity of the Fermented Red Ginseng—Analysis of Cytokine Concentration

[0124] To figure out a protective effect of the fermented red ginseng specimen A on an inflammatory disease, a degree of pro-inflammatory cytokines in a lung extract of mice was analyzed on the 6th day after being infected with H5N1 influenza virus. A cytokine ELISA was performed by means of a known method (Quan F S et al., Vaccine (2007) 25:72-28). Cytokines were detected in the lung extract according to a manufacturer's recommended procedure by using Ready-Set-Go TNFα and IL-6 (eBioscience, San Diego, Calif.).

[0125] As a result, mice given H5N1 influenza virus only showed a high concentration of inflammatory cytokines (IL-6, TNF-α) in the lung and bronchoalveolar lavage fluids (BALF) (FIGS. 4A to 4C). A mouse group given the mixture of H5N1 influenza virus and the fermented red ginseng specimen B showed a low concentration of inflammatory cytokines in the BALF (FIG. 4B). Both the fermented red ginseng specimens A and B showed an effect of reducing inflammatory cytokines in the lung compared to a dosed group with H5N1 influenza virus only (FIGS. 4A and 4C). In particular, a mouse group given the mixture of the fermented red ginseng specimen A and H5N1 influenza virus showed that inflammatory cytokines were almost completely inhibited at an almost similar degree to a non-dosed mouse group (FIGS. 4A to 4C). Thus, such results mean that the fermented red ginseng specimen inhibits a production of inflammatory cytokines caused by H5N1 influenza virus.

Experimental Example 5: Anti-Viral Activity of the Fermented Red Ginseng—Analysis of Histopathological Effect

[0126] An infection with influenza virus causes severe lung inflammations along with highly invasive cells in a respiratory tract and parenchymal tissue (H5N1 of FIG. 5). A histopathological analysis was performed on a lung tissue section of each group on the 6th day after being infected with H5N1 influenza virus (FIG. 5). The mouse group given the mixture of the fermented red ginseng specimen B and H5N1 influenza virus showed that a degree of lung inflammations was alleviated more than in the mouse group given H5N1 influenza virus only. In particular, on the 6th day after being given the mixture of the fermented red ginseng specimen A and H5N1 influenza virus, the mice showed a lung histopathology at a similar degree to the lungs of non-dosed mice (FIG. 5).

Experimental Example 6: Anti-Viral Activity According to a Nasal Mucosal Administration of the Fermented Red Ginseng

[0127] To verify preventive and therapeutic effects of the fermented red ginseng on an influenza virus infection, a protective effect thereof before and after the virus infection was determined. To test the preventive effect of the fermented red ginseng, the fermented red ginseng specimen A (500 μg) was intranasally administered into mice by means of 1 w/v % aqueous solution 4, 4.5, 12 or 24 hours before the virus infection (FIG. 6). To test the therapeutic effect of the fermented red ginseng, the fermented red ginseng specimen A (500 μg) was intranasally administered into the mice infected with influenza virus by means of 1 w/v % aqueous solution in 3.5 hours after the virus infection (FIG. 6).

[0128] As a result, it might be identified that a treatment with the fermented red ginseng has the protective and therapeutic effects in both pre-treatment and post-treatment with the H5N1 influenza virus infection. In particular, an excellent protective effect might be observed in the pre-treatment (FIG. 6A).

[0129] Also, the mice treated with the mixture of virus and the fermented red ginseng specimen showed a complete inhibition against H1N1 influenza virus (A/California/2009 pandemic virus) compared to the mice infected with virus only (FIG. 6B).

[0130] From the results, it was identified that the fermented red ginseng shows preventive and therapeutic effects on the influenza virus infection and thus may be also applied to actual treatment.

Experimental Example 7: Effect in an Immunodeficient Mouse

[0131] It is known that a production of antibodies is closely associated with an immunized protection against influenza virus. In other words, it means that mice deficient in B cells for producing antibodies are hardly protected from an influenza virus infection. It was investigated if the fermented red ginseng specimen A has an anti-viral protective effect on the H1N1 influenza virus infection in the B-cell-deficient mice. A μMT mouse model was used after modifying the previously mentioned protocol. As might be identified in FIG. 7, as a result of infecting the μMT mice with the mixture of the fermented red ginseng specimen A and H5N1 influenza virus (A/California/2009 pandemic virus), a complete protection was shown as observed in the other mice (CD4 C57BL/6, TLR4 C57BL/6). Such protective effect in the B-cell-deficient μMT mice is consistent with the result that the fermented red ginseng specimen may inhibit a virus replication. The fermented red ginseng specimen showed an excellent anti-viral activity even in mice without B cells for producing antibodies.

Experimental Example 8: Evaluation of Anti-Viral Activity of a Spray Dosage Form of the Fermented Red Ginseng

[0132] To identify an anti-viral activity against H5N1 bird flu virus, the fermented red ginseng or the fermented red ginseng and propolis were administered into mice infected with H5N1 virus by using a push-to-type oral spray container of Example 4-2 or 4-7. As a result, the mice, which were given the fermented red ginseng or the fermented red ginseng and propolis after being given H5N1 virus, did not show a weight loss, but showed an inhibitory effect on a lethal dose of H5N1 virus. On the other hand, the mice, which were not given the fermented red ginseng after being given H5N1 virus, showed a severe weight loss and eventually died all.

Experimental Example 9: Evaluation of Inhibitory Activity of the Spray Dosage Form of the Fermented Red Ginseng Against Virus Replication in the Body

[0133] To evaluate an anti-viral inhibitory efficacy of the spray dosage form of the fermented red ginseng on virus replication, the fermented red ginseng was administered into the mice infected with H5N1 virus by using a push-to-type oral spray container of Examples 4-2 and 4-7, and a virus concentration of mouse lungs was determined on the 6th day after the infection.

[0134] In case of the mice infected with H5N1 virus, a considerable amount of viruses was detected from lungs thereof. On contrary, a considerably little amount of viruses was detected from the mouse group given the fermented red ginseng or the fermented red ginseng and propolis. It means that the fermented red ginseng provided through the spray dosage form of the fermented red ginseng remarkably inhibits a virus replication in the lung against H5N1 influenza virus.

Experimental Example 10: Histopathological Effect of the Spray Dosage Form of the Fermented Red Ginseng

[0135] An infection with influenza virus causes severe lung inflammations along with highly invasive cells in a respiratory tract and parenchymal tissue. To figure out an effect of the spray dosage form of the fermented red ginseng on lung histopathology, a histopathological analysis was performed on a lung tissue section of each group, on the 6th day after being infected with H5N1 influenza virus, by using a push-to-type oral spray container of Examples 4-2 and 4-7.

[0136] As a result, the group of mice given H5N1 influenza virus only showed considerable inflammations in the lung, but the mice showed a fair lung histopathology on the 6th day after being given the fermented red ginseng or the fermented red ginseng and propolis.