ANTRODIA CINNAMOMEA FERMENTED BROWN RICE, PREPARING METHOD AND COMPOSITION THEREOF

Abstract

The present invention relates to Antrodia cinnamomea fermented brown rice, which is characterized by being produced by inoculating an A. cinnamomea strain in a medium composed of brown rice as cultivating substrate for fermentation. The A. cinnamomea fermented brown rice produced according to the present invention contains the special nutrients of brown rice and the unique active ingredients of A. cinnamomea. The A. cinnamomea fermented brown rice may be used as food material directly, and for the preparation of brown rice-based health products such as brown rice powder, brown rice milk, brown rice tea, and nutritious additives of other foods.

Claims

1. A method for preparing an A. cinnamomea fermented brown rice, comprising steps of: inoculating A. cinnamomea strain into a sterilized culture substrate consisting of brown rice and water; fermenting the A. cinnamomea strain and brown rice at a temperature of 21˜23° C. and a humidity of 65˜70% for 1-2 months until the brown rice is completely covered with fruiting bodies of the A. cinnamomea; and harvesting the A. cinnamomea fermented brown rice.

2. The method of claim 1, wherein the culture substrate is a mixture consisting of brown rice and water at ratio of 1:1.1.

3. The method of claim 1, wherein the A. cinnamomea strain is inoculated into the culture substrate with a cube form of about 2 mm×3 mm, a liquid form or a solid form.

4. The method of claim 1, wherein the step of harvesting the A. cinnamomea fermented brown rice is carried out after 40 days of fermentation.

5. The method of claim 1, wherein the harvested A. cinnamomea fermented brown rice comprises nutrients of brown rice including tocopherols, tocotrienols, γ-oryzanol, and active ingredients of A. cinnamomea fruiting bodies including 1,4-dimethoxy-2,3-methylenedioxy-5-methyl-benzene, antcin C, 2,2′,5,5′-tetramethoxy-3,4,3′,4′-bimethylenedioxy-6,6′-dimethyl-biphenyl, antcin H, dehydrosulphurenic acid, antrocamphin C, antcin B, antcin antcin A, 15-acetyldehydrosulfurenic acid and dehydroeburicoic acid.

6. The method of claim 1, further comprising a step of drying the harvested A. cinnamomea fermented brown rice under 70˜80° C. for 20˜30 hours or lyophilizing the harvested A. cinnamomea fermented brown rice to obtain the dried A. cinnamomea fermented brown rice.

7. The method of claim 6, wherein the harvested A. cinnamomea fermented brown rice is dried with hot air under 70˜80° C. for 24 hours.

8. The method of claim 6, further comprising a step of grinding the dried Antrodia cinnamomea fermented brown rice to obtain the A. cinnamomea fermented brown rice powder.

9. The method of claim 1, further comprising a step of extracting the harvested A. cinnamomea fermented brown rice with water and alcohol, followed by filtration to obtain an A. cinnamomea fermented brown rice extract.

10. The method of claim 9, further comprising a step of concentrating the A. cinnamomea fermented brown rice extract to obtain a concentrate of A. cinnamomea fermented brown rice extract.

11. The method of claim 10, further comprising a step of spray drying and grinding the concentrate of A. cinnamomea fermented brown rice extract to obtain an A. cinnamomea fermented brown rice extract powder.

12. An A. cinnamomea fermented brown rice, which is characterized in that the brown rice is produced by the method of claim 1.

13. The A. cinnamomea fermented brown rice of claim 12, further comprising a step of drying the harvested A. cinnamomea fermented brown rice under 70˜80° C. for 20˜30 hours or lyophilizing the harvested A. cinnamomea fermented brown rice to obtain the dried A. cinnamomea fermented brown rice.

14. The A. cinnamomea fermented brown rice of claim 12, which is characterized in the method of claim 1, wherein the harvested A. cinnamomea fermented brown rice comprises nutrients of brown rice including tocopherols, tocotrienols, γ-oryzanol, and active ingredients of A. cinnamomea fruiting bodies including 1,4-dimethoxy-2,3-methylenedioxy-5-methyl-benzene, antcin C, 2,2′,5,5′-tetramethoxy-3,4,3′,4′-bimethylenedioxy-6,6′-dimethyl-biphenyl, antcin H, dehydrosulphurenic acid, antrocamphin C, antcin B, antcin antcin A, 15-acetyldehydrosulfurenic acid and dehydroeburicoic acid.

15. A food composition, which is characterized in that the composition comprising the A. cinnamomea fermented brown rice of the method of claim 1.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0016] FIG. 1 shows an analysis results of the special nutrients of the brown rice in the A. cinnamomea fermented brown rice prepared by an example of the invention.

[0017] FIG. 2A shows an analysis results of the active ingredients of the A. cinnamomea in the A. cinnamomea fermented brown rice prepared by an example of the invention.

[0018] FIG. 2B shows a sample of fermented brown rice without inoculation of A. cinnamomea strain (as a control, the fermentation conditions of the fermented brown rice were the same as those of the A. cinnamomea fermented brown rice in FIG. 2A).

[0019] FIG. 2C shows a sample of A. cinnamomea strain inoculated brown rice without fermentation (as a control, the fermentation conditions of the A. cinnamomea strain-inoculated brown rice were the same as those of the A. cinnamomea fermented brown rice in FIG. 2A).

[0020] FIG. 2D shows a blank sample.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0021] As used herein, the term “brown rice” (also referred Genmi) means the parts of bran and germ retained after the shell of a rice grain being removed therefrom. The term “special nutrients of brown rice” refers to the functional ingredients beneficial to the body existing in brown rice, mainly including tocopherols (Toc), tocotrienols (T3), γ-oryzanol, γ-aminobutyric acid (GABA), ferulic acid, polyphenols, dietary fiber, and the like.

[0022] As used herein, the term “unique active ingredients of A. cinnamomea” refers to the components with specific medicinal physiological activity uniquely existing in A. cinnamomea, which mainly contains polysaccharides, triterpenes, diterpenes, sesquiterpenes, sesquiterpene lactones, benzenoids, benzoquinone derivatives, superoxide dismutases, polyacetylenes, adenosine, derivatives of maleic acid and succinic acid, proteins (including immunoproteins), phenolic compounds, steroids, and vitamins (e.g., vitamin B, nicotinic acid, etc.), trace elements (such as calcium, phosphorus, germanium, etc.), nucleic acids, amino acids, lignans, γ-aminobutyric acid (GABA), blood pressure stabilizing substances (such as antrodia acid), and the like.

[0023] The other characteristics and advantages of the present invention will be further illustrated and described in the following examples. The examples described herein are using for illustrations, not for limitations of the invention.

EXAMPLE 1

Preparation of Antrodia cinnamomea Fermented Brown Rice

[0024] Culture medium: brown rice was mixed with water at a ratio of 1:1.1 and placed in a jar. Each jar with the medium weighs about 100-120 g. The medium was sterilized at 121° C. for 1 hour. Inoculation: a 2 mm×3 mm cube form or liquid from and/or solid form of A. cinnamomea KJ-HL strain was inoculated into the above-mentioned sterilized medium. Culture conditions: temperature: 21-23° C., humidity: 65 to 70%, ventilation frequency: 5 min/12 hours, carbon dioxide concentration: 1000 to 2000 ppm, culture without light for 1 to 2 months

[0025] After 40 days of fermentation, the medium was well covered with the light reddish yellow fruiting bodies of A. cinnamomea. The A. cinnamomea fermented brown rice was harvested and dried with hot air at 70˜80° C. for 24 hours, or lyophilized for 48 hours to obtain the dried A. cinnamomea fermented brown rice. The obtained dried A. cinnamomea fermented brown rice may be directly used as a food material for cooking, or to be processed into cereal pieces, or can be used for making Genmi tea. The dried fermented brown rice may also be further ground into A. cinnamomea fermented brown rice powder and thus can be added to other ingredients or beverages as a nutrient blending agent, and made into healthy food composition such as healthy brown rice syrup, brown rice formulated milk, and brown rice cake, etc.

EXAMPLE 2

Preparation of Concentrate of the Antrodia cinnamomea Fermented Brown Rice Extract

[0026] The A. cinnamomea fermented brown rice obtained in Example 1 was extracted with water and alcohol at 95° C. for 5 hours to obtain A. cinnamomea fermented brown rice extract solution. The A. cinnamomea fermented brown rice extract solution was filtrated on a stainless steel mesh (150 mesh).

[0027] Then, the filtered A. cinnamomea fermented brown rice extract solution was transferred to a vacuum concentrator and concentrated at a temperature of 52±2° C. After the filtrate was concentrated to a concentration standard line, the concentrate was taken and placed into a fluidized-bed spray dryer to perform spray drying (temperature: 62±2° C., pump rotation speed: 35 rpm, wind turbine rotation speed: 40 rpm). The dried A. cinnamomea fermented brown rice extract was taken, ground (mesh size: 90 mesh) and sieved (mesh size: 60 mesh) to obtain the A. cinnamomea fermented brown rice extract powder.

EXAMPLE 3

Analysis of Special Nutrients of Brown Rice in the Antrodia cinnamomea Fermented Brown Rice

[0028] In order to determine whether the special nutrients are retained after fermentation with A. cinnamomea, an analysis for the active ingredients such as tocopherols, tocotrienols and γ-oryzanol in the A. cinnamomea fermented brown rice was conducted.

[0029] Extraction of Active Ingredients

[0030] Weigh 0.5 g of the A. cinnamomea fermented brown rice sample and mixed with 3 mL of n-hexane, the mixture was extracted by vortex vigorously for 20 minutes in a 60° C. water bath and then centrifuging at 2000 rpm for 15 min. After collecting the supernatant, the residue was further extracted in the same manner twice. Finally, the supernatants from the three separate extractions were combined and filtered through a 0.45 μM filter. The filtrate was collected for HPLC analysis (cf. Huang & Ng, Journal of Chromatography A 1218: 4709-4713, 2011).

[0031] Analysis of Active Ingredients

[0032] The High Performance Liquid Chromatography (HPLC) used in this example is a Hitachi system device, including: chromatographic pump (Model L-2100/2130), UV-visible spectroscopic detector (Model U-1800), and fluorescence detector (Model L-2485). The analysis of the active ingredients of the brown rice was performed using the fluorescence detector set at wavelengths of 290 nm (excitation wavelength) and 330 nm (emission wavelength). The analysis conditions are as follows: mobile phase: containing n-hexane, isopropanol, ethyl acetate, and acetic acid; analytical column: Inertsil SiL 100A, 5 μm, 4.6×250 mm; the flow rate of mobile phase: 0.7 mL/min.

[0033] As shown in FIG. 1, the A. cinnamomea fermented brown rice of present invention retains the special nutrients of brown rice as analyzed by the HPLC after fermentation, and extraction with n-hexane. The special nutrients include tocopherols (α-tocopherol represented as α-Toc, β-tocopherol represented as β-Toc, γ-tocopherol represented as γ-Toc, and δ-tocopherol represented as δ-Toc in the figure), tocotrienols (α-tocotrienol represented as α-T3, β-tocotrienol represented as β-T3, γ-tocotrienol represented as γ-T3, and 6-tocotrienol represented as δ-T3 in the figure) and γ-oryzanol.

EXAMPLE 4

Analysis of Active Ingredients of Antrodia cinnamomea in the Antrodia cinnamomea Fermented Brown Rice

[0034] In order to determine whether the unique active ingredients are included, an analysis for the active ingredients such as triterpenes, non-triterpenoids like antcin C and phenols in the A. cinnamomea fermented brown rice was conducted.

[0035] Preparation of the extract for analysis: 20 mg of the A. cinnamomea fermented brown rice sample was placed in a 1.5 mL Eppendorf tube, followed by adding 0.8 mL of 95% alcohol (1:40, w/v) and mixed thoroughly. The mixture was shaken in an ultrasonic water bath at room temperature for 1 hour, and then centrifuged at 6000 rpm for 5 minutes. The supernatant was filtered through a 0.45 μm filter into a 1.5 mL Eppendorf tube, and then made into volume of 1 mL with 95% alcohol. The extract was stored at −20° C. until HPLC analysis (cf. Lin et al., Journal of Agricultural and Food Chemistry 59:7626-7635, 2011).

[0036] Conditions of HPLC analysis comprise of analytical column: Luna C18(2) (5 μm, 4.6×250 mm, Phenomenex); detection wavelength: 254 nm; mobile phase containing A: 0.1% acetic acid (H.sub.2O with 0.1% (v/v) acetic acid), B: methanol (MeOH), and C: acetonitrile (ACN). The flow rate of mobile phase is shown in the following table.

TABLE-US-00001 Time A: H.sub.2O Flow rate (min) (0.1% Acetic acid) B: MeOH C: ACN (mL/min) 0 40 30 30 0.5 5 40 30 30 0.5 95 10 10 80 0.5 95.1 1.0 105 0 0 100 1.0 115 0 0 100 1.0 118 40 30 30 1.0 120 40 30 30 1.0

[0037] The analysis results were shown in FIG. 2A to 2D. FIG. 2A showed (in FIG. 2A, a: 1,4-Dimethoxy-2,3-methylenedioxy-5-methylbenzene, b: Antcin C, c: 2,2′,5,5′-Tetramethoxy-3,4,3′,4′-bimethylenedioxy-6,6′-dimethylbiphenyl), d: Antcin H, e: Dehydrosulphurenic acid, f: Antrocamphin C, g: Antcin B, h: Antcin Antcin A, j: 15-Acetyldehydrosulfurenic acid, and k: Dehydroeburicoic acid. That is, the A. cinnamomea fermented brown rice of the invention was confirmed to contain the unique active chemical markers of A. cinnamomea that includes 1,4-dimethoxy-2,3-methylenedioxy-5-methyl-benzene, antcin C, 2,2′,5,5′-tetramethoxy-3,4,3′,4′-bimethylenedioxy-6,6′-dimethylbiphenyl, antcin H, dehydrosulphurenic acid, antrocamphin C, antcin B, antcin antcin A, 15-acetyldehydrosulfurenic acid and dehydroeburicoic acid. Regarding the fermented brown rice (as a control, showed in FIG. 2B) and the A. cinnamomea inoculated brown rice (as a control, showed in FIG. 2C), the HPLC profiles of these controls and the blank sample indicated that there are no peaks of unique active ingredients of A. cinnamomea. The results suggest that, the unique active ingredients of A. cinnamomea contained in the A. cinnamomea fermented brown rice of the invention has to be produced by the fermentation process composing of brown rice and A. cinnamomea.

[0038] The present invention first utilizes brown rice as a nutrient substrate, to obtain the special nutrients of brown rice and unique active ingredients of A. cinnamomea through fermentation. The A. cinnamomea fermented brown rice can be directly used for cooking as food, since the medium used for fermenting A. cinnamomea fermentation is composed of brown rice that results in simple composition thereof. Furthermore, brown rice itself is a well-known food material having high nutritional value. After going through the fermentation process with A. cinnamomea, the A. cinnamomea fermented brown rice not only retain the special nutrients originally present in the brown rice, but also has the unique active ingredients of A. cinnamomea, thereby further enhancing its industrial value and breadth in the preparation of healthy products and nutraceuticals.