TW565430B - Incubation method for obtaining solid culture of Zang Zhi, solid culture obtained therefrom - Google Patents

Abstract

The present invention provides an incubation method for obtaining solid culture of Zang Zhi (Antrodia camphorata) comprising the steps of: incubating a stock strain of Zang Zhi in a PP bag for growth of mycelia, followed by an incubation in the air for fruiting the Zang Zhi body. The invention also provides the solid culture of Zang Zhi obtained by the incubation method.

Description

565430

1 BACKGROUND OF THE INVENTION Zang Zhi 'Its scientific name is Antrodia camphorata. kanelurai), the inner wall of the hollow material, the fruit body has a strong camphor tree aroma, and its shape is plate-shaped or bell-shaped. Folklore has soothing effects, timidity, stasis and blood circulation, detoxification in the middle temperature, detoxification and swelling, and analgesic effects. It is a good antiseptic and antidote, and is regarded as the most expensive wild fungus on the Taiwan market. What's more, it is different from the indirect effect of Ganoderma lucidum to improve immunity and anti-cancer effect. Antrodia cinnamomea has always been a mechanism that directly inhibits or kills cancer cells. At the same time, it has strong heart, immune regulation, anti-parasympathetic effect, and anti-serotonin efficacy. In particular, Antrodia camphorata can treat gastrointestinal pain, diarrhea, vomiting, diabetes, gout, nephritis, arthritis, inflammation, allergies, excessive urinary protein, uremia, cirrhosis, liver cancer, and influenza. Numerous research units have invested human and material resources to research the active ingredients of Antrodia cinnamomea. The special research plan of the National Science Council of the Executive Yuan uses chemical methods, nuclear magnetic resonance spectroscopy (NMR) and comparison with known compounds aq 'to establish The structural characteristics of the active ingredients of Antrodia cinnamomea and the research results of the active ingredients of Antrodia cinnamomea indicate that the water extract of Antrodia cinnamomea citrifolia has an inhibitory effect on the growth of Staphylococcus aureus and P. acnes. Anthocyanin A (zhank U icacid), a methanol extract of Antrodia cinnamomea, has significant effects on lymphoma cells of p-3 8 8 mice: O: \ 71 \ 7l526.DOCWVCK · 4 · · This paper is applicable to the standard @ 国 标准 (CNS ) A4 specification (21Q χ Norwegian Gongfu)-565430 V. Description of the invention (A7 B7: first use and platelet texture; while cinnamolic acid B has weak anticholinergic and brain-stimulating amines (ser 〇t 〇ni η) Function ^ Gao Xiaowei's master thesis aimed at the three beneficial ingredients of the new species of Ganoderma lucidum in Taiwan ^ ^ (Gano derma c〇mphoratum Zang e''Su) & 190 A new species of Ganoderma lucidum published in 190, extracted with acetone : Body to obtain a crude extract, continued separated Chromatography (LC, [mu])

Human recrystallization purification; thin layer liquid chromatography (TLC)

Seannmg) and high performance liquid chromatography (Hp via mass spectrometry, infrared spectroscopy, UV wells, ττ $ b, 'Quanxian spectrum, nuclear magnetic resonance spectroscopy (H-NMR and C-NMR) for structural determination. Of which 3 η × 〇-8,23-dlen-26-olc acid For r rni, For churnamic acid-pyrogluconate transaminase in the serum of mice infected with acute hepatitis by carbon tetrachloride (cCl4) (GPT) value has a decreasing effect. Because Antrodia camphorata is mostly distributed in the broad-leaved forest area of the Taitung coast mountain range in Taiwan, according to the Forestry Bureau's Angustifolia Conservation Announcement, this area has been classified as ^ Antrodia cinnamomea is very small in quantity, and the Uyo Cup gathers. Natural wild antrodia cinnamomea can no longer be obtained on the market. Summary of the Invention _ The main purpose of the present invention is to provide a π ^ I ancient silkworm method of Antrodia spp. Cultivation of Antrodia camphorata solids with the same effect as natural wild _ 〃 、 by artificial cultivation: It is another object of the present invention to provide a solid culture of Antrodia camphorata (J: \ 7I \ 71526.DOC \ WCK -5- 565430 A7 B7 V. Description of the invention () 4 〇. 〇1). Values are average + standard Error representation. 1 Figure 5 shows the effect of different concentration factors of Antrodia camphorata on carbon tetrachloride in liver function biochemical values of glutamic acid-oxaacetate transaminase (G 〇 T) enzyme activity changes. # Represents normal group and injury There is a statistical difference between the groups (P < 0 · 0 1); * represents a statistical difference between the feeding group and the injury group (P < 0. 0 1). Values are expressed as mean + standard error. Figure 6 Represents the effect of different concentration multiples of Antrodia camphorata on the macroscopic changes of liver tissue caused by carbon tetrachloride (Table A normal group, Table B carbon tetrachloride injury group, Table C fed Antrodia camphorata triple concentrated group and Table D fed Antrodia camphorata Eight-fold concentrated group). Figure 7 represents the effect of different concentration multiples of Antrodia cinnamomea on liver tissue changes caused by carbon tetrachloride. Mice were fed with different concentration multiples of Antrodia cinnamomea for five weeks, and their livers were taken for sectioning and staining. Observe liver tissue changes (Table A normal group, Table B carbon tetrachloride injury group, Table C fed with Antrodia camphorata three-times concentrated group and Table D fed Antrodia camphora eight-times concentrated group). Figure 8 shows the Antrodia camphorata powder extract on the intestines The effect of the growth of cancer cells (COLO 320HSR). The present invention provides a solid cultivation method of Antrodia camphorata, which aims to artificially cultivate a large amount of natural Antrodia camphorata fruit components and active Antrodia camphorata fruiting bodies. The present invention The mycelium is cultivated in mycelium with space bag, and the fruit body cultivation is continued in the air. O: \ 71 \ 71526.DOC \ WCK-7-This paper size is applicable to China National Standard (CNS) A4 (210X297 mm) ) 565430 A7 ---- —_ B7 V. Description of the invention () Before the mycelium culture stage, it can usually go through the multiplication step. For example, the following steps can be used to prepare a large number of bacteria for space pack culture, which can include three steps.骡: (1) take a small piece of agar hyphae from liquid nitrogen preservation activated bacteria, transfer to a fresh medium and cultivate at constant temperature, and wait for the bacteria to grow vigorously; (2) inoculate the bacteria to kill In the space bag of the fibrous material of the bacteria (such as wheat kernels or wood chips), continue to culture at a constant temperature; (3) When the mycelium in the Taizhi bag is full of hyphae, remove the uppermost piece of mycelium , And then you can inoculate them into the space pack in large quantities. The proliferation step before the mycelium culture stage, or fermentation in liquid culture to mass culture bacteria, and then inoculated into the space bag. The formula of the fermentation broth is 1-3% fructose, 0. 0 1-0. 1% magnesium sulfate, 0 J J% yeast extract 0. 05-0.5. /. Potassium dihydrogen phosphate. Particularly preferred is a composition of 2% fructose, 0.05% osmium sulfate, 0.5% yeast extract, and 0.1% potassium dihydrogen phosphate. After oblique interview tube culture, 'inoculate a large amount of fermented bacteria in 5 liters of fermentation broth' at 2 4-2 6 ° C, rotate and shake at 240 rpm for about 4 days, and continue shaking at 90 rpm Cultivate for about 14 days, and inoculate it into 20 liters of liquid fermentation broth and stir for 14 days. The space bag referred to here is a plastic bag containing fibrous matter such as stems, stalks, fruits or sawdust of any mushroom or plant (particularly the stems, stalks, fruits or Fiber sawdust is better), 10-j 0% "Yi Fen Yuan (especially for horses), and 彳 J, 1)% of grains (especially rice bran is better), 1-10% Carbohydrates (v V especially in Portuguese

U: \ 71 \ 71526.D0C \ WCK

565430 V. Description of the invention: Glucose is preferred) '0.5. 2% phosphate (especially potassium dihydrogen phosphate as one) and 0.1-1/0 sulfate (especially magnesium sulfate is preferred). Its history is about 60-90%, 7 "r > 7 is preferably 80%, and its pH value is adjusted to be close to neutral. The culture stage of buttoned mycelium in this article refers to the cultivation of bacteria. The space temperature of the space bag is about 60 days. The growth temperature of the mycelium bone bean of Bazhong is between 5 and 32 ° C, especially the optimal growth temperature of 2 8 C. The humidity of the space bag is preferably 60-80%. / ° is better. As for its air condition, it is better to use 0.2-1% carbon dioxide. After the mycelium culture is completed, the fruiting body cultivation stage is performed, that is, the space bag is removed in about 61-90 days. , So that it is completely exposed to the air. During the growth of the daughter body, there is a shortage of g ^ ^ The daily battle & difference must be strictly controlled, and the daytime temperature is generally between 20 and 30 ^, and 25n; rc is particularly preferred; at night The temperature is preferably 1 1 C and 9 1 C; the temperature difference between day and night is preferably 5 r. Fruiting body cultivation P "and 2 air humidity should be controlled between 9 0-95%; in addition fruit seed t # material Cultivate under air circulation, in which the carbon dioxide content cannot exceed: 1%. Generally, the solid culture can be harvested within 90-120 days. 〃According to the present invention, the strains deposited in Taiwan Food Industry Development Research Institute No. CCRC 3 5 3 9 8 were used as the source. Straight length is about 1 〇_ 3 0 male knife 'interval is about 1 355 cm and weight is about 0.2-〇. Kg. The solids of Antrodia camphorata obtained by the above-mentioned cultivation method can be learned through any method

O: \ 71 \ 71520.DOC \ WCK -9- 56543〇

7 The processing methods familiar to the Institute of Technology, such as the active ingredients of solid cultures, continue to extract the two (vacuum concentrati 00) method with known alcohol or alcohol, such as true more concentrated cocn.entration) to prepare concentrated products. —7 Chen Zhi (freeze or drying treatment, such as, etc., its + Λ helmet + office, ground coffee tile drying, freeze drying among them ... heat, dry, can get <dry powder. Especially cold Cambodia drying method It is better to remove the moisture of the solid matter between normal temperature and normal temperature, and the solid nutrients are not affected by oxygen during the drying period when the temperature is lower than <4 to ^ κ. Properties: Or make the spray-dried micro-powder particles or micro particles prepared by other drying methods to increase the agglomeration and particle size of the powder through granulation (agg10merat10n). The structure of the granulated product is porous. It has good wettability. The high-performance liquid chromatography (ΗPLC) spectrum data comparison of the solid culture of Antrodia cinnamomea and the Antrodia camphorata obtained from the conventional liquid culture method and wild Antrodia camphorata, as shown in Figure 2 .Analyzed by PLC, the daughter body obtained by the solid culture method of the present invention has a composition very similar to that of natural wild Antrodia camphorata. When it has the same activity and use as natural wild Antrodia camphorata, including calming, timidity, and fasting blood circulation. Wenzhong Xiaoji, detoxification and swelling, and sedative pain, and has the ability to inhibit bacteria, viruses and tumor cells' and the effects of strengthening heart, immune regulation, anti-sympathetic effect, anti-serotonin activity. It can be used to treat gastrointestinal pain , Diarrhea, vomiting, diabetes, gout, kidney-10-

0; \ 7] \ 71526.D〇C \ WCK This paper size applies to China National Standard (CNS) A4 (210X297 mm)

565430 The invention says ϋ inflammation, arthritis, inflammation, allergies, high urine protein, uremia, cirrhosis, liver cancer and influenza. It even has the ability to rebuild the skin, and can also be used as a material for human skin or callus, and can be used as a patch for bedsores and skin trauma. According to the present invention, the solid culture of the present invention is further concretely demonstrated to have anti-free radical and anti-lipid peroxidation capabilities, protect liver function, and inhibit cancer cell efficacy. It is apparent that the solid culture of the present invention can be used as a healthy food for survival, or can be further developed into various drugs. The following examples are provided to further illustrate the present invention, and are not intended to limit the present invention. Modifications and applications achieved by anyone skilled in the art in accordance with the teachings of the description of the present invention are all examples of the present invention. EXAMPLES A strain 'from the food industry development institute registration number CCRC 35398 was cultured and fermented in liquid to ferment a large number of strains, and then inoculated into a space bag. The formula of the fermentation broth is 2% fructose, 0.05% magnesium sulfate, 0.5% yeast extract and 0.1% diammonium diammonium bell. Then the mycelium culture was carried out in a space bag method. The space bag composition was 65% of the stems, stalks, fruits, and sawdust fibrous matter of the herbaceous plants, 20% of yam potato, 10% of rice bran, and 35% of the total. Glucose, 1% potassium dihydrogen phosphate and 0.5% magnesium sulfate were adjusted to have a clarity of 80% and a pH value of 7. Its clarity is about 60-90%, and S0% is better, and its pH value is adjusted to be near neutral. The term “silk culture stage” referred to in this article means that the bacteria are planted into the space pack for about 60 days U: \ 7l \ 71526.DOC \ WCK-1 1 ·

This paper size is in accordance with Chinese National Standard (CNS) A4 specification (210X297 mm) 565430 A7 B7 V. Description of the invention () 9, in which the mycelium growth temperature is between 5 and 3 2 ° C, especially 2 8 ° C is the optimum growth temperature. The humidity of the space bag is 80%, and the air condition is 0.2-1% carbon dioxide. After the mycelium culture is completed, the fruiting body cultivation stage is performed, that is, the space bag is removed in about 6 1 -90 days, and it is completely exposed to the air. The temperature difference between day and night during the growth of the fruiting body must be strictly controlled. The daytime temperature is generally between 20 and 30 ° C; the nighttime temperature is 11 ° C soil J ° C; the day and night temperature difference is maintained at 15 ° C. The air humidity of the fruiting body in the cultivation stage should be controlled between 90 and 95%. In addition, the fruiting body should be cultivated under air circulation, and the carbon dioxide content should not exceed 1%. Within 90-120 days, the solid culture can be harvested when the conical fruiting body grows to a diameter of 15 cm and a height of 25 cm. Its weight is about 0.4 kg, as shown in Figure 1. The high-performance liquid chromatography (ΗPLC) analysis of triterpenoids, its main active ingredient, is shown in Figure 2 (1). The values are shown in Table 1 below. -12-

O: \ 71 \ 71520.DOC \ WCK The paper size is applicable to the Chinese National Standard (CNS) A4 specification (210 X 297 mm) 565430 Α7 Β7 V. Description of the invention (10 Table 1. Camphor zhi obtained from the cultivation method of the present invention Physical high performance liquid chromatogram data of the physical solution tbpm curve product · phase »《degree" concentration "degree 缜 缜 w (uv« =) atst mice 96 name s47e32317137373373sso7343 $ o97 $ 3606783904323279390lo131390s767z3lsoso57LOo} 32r303773o03 .0.56el 4.s.9.3.l7.0n.l9.9.34.82.32.19998131ll998 $ 8632s96s91945079573619ss32sss &quot; 丨 10.10.11.12.12.14.17.18.18.19.20.22.23.23.25., :: 30.33.33.37.:::42. ::: 50:32. -10111213141ss1? 1bi9202122232425s27s29303132333435s3738ss414243 "454647 5 6 7 8 9 22 ^^ 2333345566s 2795β870 6320387 6333234 13521140 393321X 4614312 2519874 2084 2 312 2589 2962 2264 2264 2264 2264 2264 2264 1000.279 6.5362 0.0000 2.1028 999.303 6.5299 0.0000 2.1070 999.158 β.5289 0.0000 4.49Θ4 998.886 6.5271 0.0000 1.3007 3 28.263 2.1450 0.0000 1.5352 303.141 1.9B08 0.0000 0.8394 192.129 1 0.0000 0.693S 144.477 0.9441 0.0000 1.4192 149.458 0.9766 0.0000 2.5239 460.320 3.0079 0.0000 0.55β1 71.272 0.4657 0.0000 0.2469 54.184 0.3341 0.0000 0.420? 62.749 0.4X00 0.0000 0.4322 0.4002 58.580 0.00. 0.2815 0.0000 1.9504 317.837 2.0769 0.0000 0.3103 29.915 0.19S5 0.0000 0.4172 34.691 0.2267 0.0000 0.1790 22.B99 0.1496 0.0000 0.0347 12.337 0.0806 0.0000 0.4104 29.716 0.1942 0.0000 0.2090 19.25β 0.1258 0.0000 77 07000330307 7 3 B792124 9 8800091 ^ 4 ^ 6925804947927 58885550001 88889990001 88889990001 I2590SS 0.418 »37.37β 0.2442 0.0000 2982S81 0.9923 90.403 0.5254 0.0000 2321967 0.8390 68.342 0.4466 0.0000 627346 0.2097 19.934 0.1303 0.0000 117339 0.0390 4.690 0.0306 0.0000 663514 0.2207 20.199 0.1320 0,0000 2932057 0.9755 32.760 0.344β 0.0000 252149 0.0839 3.193 0.0636 0.00630 0.00630 0.00630 0.00630 0.00630 0.004191130832 0.3752 31.023 0.2027 0.0000 4135840 1.37β0 78.025 0.5098 0.0000 1070268 0.2561 22.420 0.1465 0.0000 2127664 0.7079 44.217 0.2889 0.0000 13997700 S.3224 304.306 1.998S 0.0000 5256445 1.74ββ »2.3« 0, € 036 ο.οοοο 462920 0.1540 9.40S 0.05 4416830 1.4695 79.089 0.S168 0.0000 261669 0.0871 5.522 0.0361 0.0000 4411945 1.4 € 7B 73.32 «0.4791 0.0000 2821209 0.3386 50.536 0.3329 0.οοοο 3ββΟβ 0.0122 1.071 0.0070 0.οοοο 10087950 3.3562 134.950 0.8813 0.οοοο 291 0 1 4294 0.149 4.5109 «0.37« 0.3943 0.οοοο 357 0.0414 2.047 0.0134 0.0000 938 1.6391 340.488 2.2249 0.οοοο 754 2.4336 «25.513 4.0974 0.οοοο 732 1.5260 463.198 3.0267 0.0000 .58β 1.2308 189.756 1.2399 0.οοοο 1056724 0.3516 118.120 0.7718 0.00 .2Ι8 0.8770 0.0000 1992234 0.S393 137.325 1.0293 0.οοοο 144S816 ο.Αβια 124.837 0.8137 0.0000 313869 «1.0442 171.327 1.1195 0.οοοο 1098725 0.3β5 5 β9.7 »4 0.5ββ7 0.0000 1481909 0.4930 102.S06 0.6698 0.0000 340« 597 1.1334 2Z2.Α22 1.3223 ύ.ΰΰΰο 16450Θ4 0.5473 104.449 0.6825 0.οοοο 10387770 3.5225 757.281 4. »484 0.οοοο 1 £ 2« 563 0.5412 117.086 0.7β31 0.οοοο 856049 0.2848 79.162 0.5173 0.οοοο 1490212 0.4984 ββ.3S4 0.5773 0.0000 735Θ936 2.4483 S38.464 3.5182 0.οοοο 10076ΒΘ 0.3353 76.731 0.5014 0.0000 αΛ

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• 14-

O: \ 71 \ 71520.DOC \ WCK The paper scale is applicable to the Chinese National Standard (CNS) A4 (210 X 297 mm) 565430 A7 B7 V. Description of the invention () 12 In order to compare the solids of Antrodia camphorata with other cultures The differences between Antrodia cinnamomea obtained in this way and Antrodia cinnamomea and wild Antrodia spp. Obtained by liquid culture were performed by HPLC. The spectra are shown in Figures 2 (2) and (3), and the data are shown in Tables 2 and 3. It can be seen from the map that the fruit body of Antrodia camphorata cultivated by the present invention has the same bar shape as that of wild Antrodia camphorata, and it also indicates that the child bodies cultivated by the method of the present invention should have the same activity and use as wild Antrodia camphorata. -15-

O: \ 71 \ 71526.DOCWVCK The paper size is applicable to Chinese National Standard (CNS) A4 (210 X 297 mm) 565430 A7 B7 V. Description of the invention (13 Table II. High-performance liquid phase of Antrodia camphorata obtained by liquid cultivation method Chromatogram data m Retention area area * Relative height * Concentration concentration baseline component number time (uv * sec) Height unit series g 〇1 1.620 262998 1.2134 38.745 2.2045 0.0000 2 1.857 8294342 38.4242 660.535 37.5912 0.0000 3 2.520 5417475 25.0969 726.393 41.3301 0.0000 4 2.820 1692634 7.8413 99.406 5.6559 0.0000 5 3.790 731157 3.3S71 23.209 1.3205 0.0000 6 4.123 271974 1.2599 11.713 0.6664 0.0000 7 4.623 154068 0.7137 7.810 0.4444 0.0000 β 5.023 154230 0.7145 4.580 Q.2S06 0.0000 9 5.897 29822 0.1382 1.652 0.0940 0.0000 10 7.860 150572 0.6 5.210 0.2964 0.0000 11 S.210 123419 0.5717 3.314 0.2227 0.0000 12 11.930 130807 0.6060 3.563 0.2027 0.0000 13 12.740 37712 0.1747 1.232 0.0701 0.0000 14 15.103 504463 2.3370 11.048 0.6286 0.0000 15 16.890 143477 0.6647 3.512 0.1998 0.0000 16 19.350 56 said € 0 0.2634 1.827 0.1039 0.0000 17 23.510 806878 3.7379 11.274 0.6415 0.0000 18 27.167 92042 0.4264 1.224 0.024 0.00696 19 30.633 144285 0.6684 2.243 0.1276 0.0000 20 33.747 299568 1.3414 3.94 * 7 0.2245 0.0000 21 38.727 250488 1.1604 3.231 0.1839 0.0000 22 41.340 61644 0.2856 0.7 0.0438 0.0000 23 43.727 29928 0.1386 0.4 70 0.0267 0.0000 24 57.047 112074 0.5192 9.096 0.5175 0.0000 25 57.433 175711 0.8140 10.275 0.5840 0.0000 26 57.907 107598 0.4985 4.508 0.2565 0.0000 27 53.137 25810 0.1196 2.279 0.1296 0.0000 28 61.380 23245 0.1077 2.024 0.1152 0.0000 29 62.743 196612 1.2385 0.0000 30 64.223 30411 0.1409 2.094 0.1192 0.0000 31 64.717 441196 2.0439 41.620 2.36Θ1 0.0000 32 64.997 48524 0.2248 4.870 0.2771 0.0000 33 65.420 40354 0.1869 3.529 0.2008 0.0000 34 65.833 67766 0.3139 3.823 0.2175 0.0000 35 66.950 21293 0.0987 1.691 0.0962 0.0000 36 67.773 32 0.0972 0.0000 37 79.013 29414 0.1363 0.920 0.0523 0.0000 38 97.297 21884 0.1014 0.456 0.0260 0.0000 39 100.053 38474 0.1782 0.738 0.0420 0.0000 40 104.457 126686 0.5869 2.358 0.1342 0.0000 41 107.010 57252 0.2652 2.113 0.1202 0.0000 42 111.397 35854 0.1661 4.236 0.2410 0.0000 43 111.313 28337 0.1313 2.512 0.1429 0.0000 44 113.790 25408 0.1177 1.310 0.1030 0.0000 45 1.808.883 43 0.2167 0.0000 46 119.590 25451 0.1179 1.656 0.0942 0.0000

PVVVVVVVVVBBVVWWPPPPPPV-VPPPPPPPSPPPPPPPPPPPPPVVVWWVPPVVVVVVVVVVVPPPPPPPPPPPPVVVPVPPVPP Class and 21586254 1757.553 0.0000 -16-

O: \ 7l \ 71526.DOC \ WCK This paper size is applicable to Chinese National Standard (CNS) A4 specification (210X297 mm) 565430 A7 B7 V. Description of the invention (14 Table III. High performance liquid chromatogram of wild Antrodia Data compilation © Remaining school rents Relative height% Moss S Degree detected component number time (uv ^ sec) Island unit brick name 1 1.550 1332009 0.5787 162.045 1.1488 0.0000 2 1.913 12708000 5.5210 797.430 5.6535 0.0000 3 2.607 7990504 3.4715 374.149 2.6526 0.0000 4 2.997 3336181 1.4494 151.464 1.0738 0.0000 5 3.480 827969 0.3597 90.536 0.6419 0.0000 5 3.687 2100942 0.9127 89.231 0.S326 0.0000 7 4.110 1665378 0.7235 80.468 0.5705 0.0000 Θ 4.590 986857 0.4287 61.543 0.4363 0.0000 9 4.867 1300649 0.5651 59 6.445 0.4214 37.086 0.2629 0.0000 11 5.623 712514 0.3095 37.666 0.2670 0.0000 12 6.020 517412 0.2682 32.787 0.2324 0.0000 13 6.347 380678 0.1654 24.939 0.1768 0-0000 14 6.660 622333 0.2704 24.053 0.1705 0.0000 15 7.060 557302 0.2421 18.364 0.1302 0.0000 16 7 .710 585765 0.2545 27.106 0.1922 0.0000 17 8.353 276221 0.1200 10.957 0.0777 0.0000 18 8.867 379887 0.1650 19.904 0.1411 0.0000 19 S.327 316234 0.1374 21.157 0.1500 0.0000 20 9.523 72843 0.031S 4,914 0.0348 0.0000 21 10.593 120581 0.0524 6.584 0.0467 0.0000 22 11.260 57846 0.0251 2.283 0.0162 0.0000 23 11.747 32476 0.0141 2.199 0.0156 0.0000 24 12.263 209326 0.090 $ 11.437 0.0811 0.0000 25 12.837 131315 0.0570 6.603 0.0468 0.0000 25 13.733 270732 0.1176 7.438 0.0527 0.0000 27 15.300 1563,80 0.0679 6.542 0.0464 0.0000 28 17.513 346774 0.1507 10.369 0.0735 0.0000 29 1S. 270 1011777 0.4396 25.904 0.1837 0.0000 30 19.863 298337 0.1296 7.175 0.0509 0.0000 31 21.567 237547 0.1032 6.715 0.0476 0.0000 32 22.657 261555 0.1136 7.129 0.0505 0.0000 33 23.960 419821 0.1824 8.165 0.0579 0.0000 34 26.780 1014237 0.4405 20.361 0.1444 0.0000 35 23.320 4482109 0.7900 0.0000 103. 3S 32.017 6999710 3.0410 156.204 1.1074 0.0000 37 33.343 399151 0.1734 8.733 0.0619 0.0000 3Θ 35.053 97125 0.0422 2.428 0.0172 0.0000 39 37.007 3062612 1.3305 4S.150 0.3272 0.0000 40 39.587 832484 0.3617 15.926 0.1129 0.0000 41 40.897 2411303 1.0476 30.354 0.2152 0.0000 42 43.603 94412 0.0410 2.102 0.0149 0.0000 43 46.093 950329 0.4129 12.948 0.0918 0.00918 0.00918 0.00 0.697 0.0049 0.0000 45 51.273 22016 0.0096 0.505 0.0036 0.0000 45 53.253 391511 0.1701 6.542 0.0464 0.0000 47 S5.177 418679 0.1819 6.082 0.0431 0.0000 48 57.243 39 Θ8680 1.7329 240.883 1.7078 0.0000 49 57.500 4155318 1.8053 345.611 2.4503 0.0000 50 57.787 3545656 1.0404 0.00404 257 0.5244 175.620 1.2451 0.0000 52 58.257 2986313 1.2974 222.955 1.5807 0.0000 53 58.400 5093407 2.2128 431.601. 3.0599 0.0000 54 58.673 2540255 1.1036 286.541 2.0315 0.0000 55 58.943 5651787 2.4597 471.025 3.3394 0.0000 56 59.233 2400643 1.0430 197.347 1.3991 0.005.00 0.0059 57222 233.006 0.9684 16 4.165 1.1639 0.0000 53 60.060 2928869 1.2290 335.781 2.3806 0.0000 60 60.410 17547630 7.6235 994.818 7.0529 0.0000 U: \ 71 \ 71526.DOC \ WCK -17- Packing 297 mm) 565430 A7 B7 V. Description of invention (15 61 60.743 13833800 6.0101 994.628 7.0516 0.0000 62 60.920 9930270 4.3142 941.167 6.6726 0.0000 63 61.343 3601150 1.5645 265.761 1.8842 0.0000 64 61.723 604S523 2.6278 525.238 3.7238 0.0000 65 62.127 2249845 6 62.357 2849680 1.2380 152.131 1.0786 0.0000 67 63.207 11335680 4.924Θ 9Θ6.220 6.2 said 30 0.0000 68 63.460 3758168 1.6327 214.761 1.5226 0.0000 69 64.087 17403430 7.5609 992.676 7.0377 0.0000 70 04.807 6379895 2.7717 332.965 2.3606 0.0000 71 65.487 10.5522 1105522. 0.4613 41.562 0.2947 0.0000 73 66.490 1903467 0.8270 65.469 0.4642 0.0000 74 67.563 1016818 0.4418 30.915 0.2192 0.0000 75 68.387 1511594 0.6567 37.104 0.2631 0.0000 76 6 9.390 10743430 4.6675 526.207 3.7306 0.0000 77 70.583 358833 0.1559 13.087 0.0928 0.0000 78 71.257 2732845 1.1873 127.212 0.9019 0.0000 79 72.317 237862 0.1033 7.948 0.0563 0.0000 60 72.730 245290 0.1066 6.784 0.0481 0.0000 81 73.783 87523 0.0380 3.200 0.0227 0.000 0.0000 0.00 74 74.250 136. 86115 0.0374 2.980 0.0211 0.0000 84 / ^ 65 75.973 28738 0.0125 1.165 0.0083 0.0000 77.503 92275 0.0401 1.898 0.0135 0.0000 86 80.060 508744 0.2210 11-408 0.0809 0.0000 87 82.200 28158 0.0122 0.871 0.0062 0.0000 88 84.293 79909 0.0347 1.534 0.0109 0.0000 89 87.500 1092447 0.4746 24.011 0.0000 90 87.630 300046 0.1304 24.553 0.1741 0.0000 91 87.893 382295 0.1661 25.603 0.1815 0.0000 92 88.107 224523 0.0975 21.631 0.1534 0.0000 93 88.397 717983 0.3119 37.094 0.2630 0.0000 94 89.010 518852 0.2254 22.242 0.1577 0.0000 95 89.237 307090 0.1334 21.551 0.1158 0.00358 0.0000 96 89.315 97 90.007 4050733 1.7598 326.533 2.3150 0.000 0 98 91.020 376589 0.1636 16.743 0.1187 0.0000 99 91.427 152969 0.0665 11.423 0.0810 0.0000 100 91.S43 205346 0.0892 11.033 0.0782 0.0000 101 92.013 142333 0.0618 11.613 0.0823 0.0000 102 92.157 268901 0.1158 11.479 0.0814 0.0000 103 92.633 124915 0.0543 8.073 0.0572 0.0000 104 93.083.938 0.6266 0.0000 105 93.690 210904 0.0916 15.423 0.1093 0.0000 106 94.053 323059 0.1404 22.173 0.1572 0.0000 107 94.547 63428 0.0276 3.102 0.0220 0.0000 108 95.123 29222 0.0127 2.059 0.0146 0.0000 109 95.963 45135 0.0196 2.967 0.0210 0.0000 110 98.950 58309 0.0253 3.644 0.0258 0.0000 111 100.273 0.0025 0.00 0.0000 112 105.563 38299 0.0166 1.804 0.0128 0.0000 113 112.263 55710 0.0242 1.468 0.0104 0.0000 114 126.417 44605 0.0194 0.851 0.0061 0.0000 vvvvvvvvvvvvvvvvwvvvvwvvwvvvvvvvvvvvvvvvvvpg; £ aa 3VVVVVVVVVVVVVVVVVVVVVVVVVVVVVVVVVVVV suppression offerings a £ g; g; sum 230177S56 14105.03 0.0000 O: \ 71 \ 71526.DOC \ WCK ~ 1 8-This paper size applies to Chinese National Standards (CNS) A4 specification (210X297 mm) 565430 A7 B7 V. Description of the invention (16 The following composition analysis test and pharmacological test describe the composition and efficacy of the concentrated product of Antrodia camphorata fruit solids obtained according to the cultivation method of the present invention. After analysis, the main components include triterpenoids, water-soluble polysaccharides, and chitin. The water-soluble polysaccharides were tested by the Japanese Food Analysis Center and found that the solid body of Antrodia camphorata fruit obtained by the culture method of the present invention has a high amount of water-soluble polysaccharides, and each 100 grams contains about 1.2 to 6.5 grams of water-soluble polysaccharides. body. Heavy metal content According to the atomic spectrometer, the heavy metal analysis of Antrodia cinnamomea solids cultivated according to the method of the present invention was performed according to the atomic spectrometer in the Micro-analysis Laboratory of the Department of Atomic Science, Tsinghua University.

Na 48.7 ppm Mg: 767 ppm A1 41.6 ppm Ca: 605 ppm

Mn: 10.9 ppm Cd: ND

Fe: 100 ppm Cr: ND

Zn: 16.4 ppm As: ND K: 0.560% Hg ·· ND Total antioxidant activity (T ota 1 antioxidant activity) According to the concentrated powder of Antrodia camphorata solids and wild Antrodia camphorata powder and Ganoderma lucidum fruit body concentrated powder For comparison, check the content of each gram of powder, its -19-

CJ: V7! \ 71526.DOC \ WCK This paper size applies to Chinese National Standard (CNS) A4 (210 X 297 mm) 565430 A7 B7 5. Description of the invention () 17 is equivalent to the milligrams of vitamin C. Content per gram of powder / mg equivalent to vitamin C. Wild Antrodia powder 633 ug ascorbic acid / g Ganoderma lucidum fruit body concentrated powder 854 · ug ascorbic acid / g Concentration of solids of Antrodia Powder 1180ug ascorbicacid / g This result shows that the anti-oxidative power of the solid substance of Antrodia camphorata in the present invention is much higher than that of Ganoderma lucidum fruit body and wild Antrodia camphorata. Toxicity test The Antrodia camphorata according to the present invention uses rats to perform an acute oral toxicity test LD50. The rats in the experimental group take 2,000 mg / Kg orally a single time according to the present invention. The control group is given purified water as a vehicle control. The experimental results showed that there were no abnormal or dead mice in the experimental group. Therefore, the L D 50 value of the acute oral toxicity test is 2,000 m g / K g or more. Salmonella inverse mutation test (Ames test) The Antrodia camphorata line according to the present invention uses the flat-plate hybrid test method to perform Salmonella inverse mutation measurement (A m e s t e s t). The test strains included five strains of Salmonella typhimurium TA 9 7, TA 9 8, TA 1 0 0, TA 1 02, and TA 1 5 3 5. Five concentrations were analyzed, which were 3 1 2. 5, 6 2 5, 1 2 50, 25, 0, and 5 0 0 ug / plate, including both the negative control group and the strain-specific positive control group, each group is O: \ 71 \ 71526.DOCWVCK ~ 20 ~ This paper size applies China National Standard (CNS) A4 specification (210 X 297 mm) 565430 A7 B7 V. Description of the invention () 18 Repeatedly. The test results showed that the background of the reverse mutant colonies caused by the negative control group all fell within an acceptable range, while the positive control group also caused a significant increase in the number of reverse mutant colonies. All the tested concentrations of Antrodia camphorata according to the present invention did not significantly increase the number of reverse mutant colonies of the test strains, regardless of whether they contained the rat liver enzyme (S 9) metabolism system, as shown in Table 4. Table 4. Salmonella reverse mutation test subject number of experimental cases ingestion • Periodic experiment results Salmonella (Salmonella ty phimurium) 5 strains of TA97 TA98 TA100 TA102 TA1535 each require Histidine mutant test concentration of histidine 31g, 312.5, 625, 1250, 2500, 5000 plate mixed f-type test was repeated three times at 37 ± 1 ° C constant temperature culture for 48 ~ 72 hours. The number of mutant colonies was calculated. All test concentrations were processed with or without rat liver enzyme metabolism system (S9). In the following, no significant increase in the number of reverse mutant colonies of the test strain was caused. The test result was negative. Test for anti-lipid peroxidation ability The lipid peroxidation reaction of rat brain homogenate was used to detect the anti-free radical and anti-oxidation ability of the solid culture concentrate of Antrodia camphorata according to the present invention. Figure 3 shows that the stimulation of the rat brain homogenate with ferrous ions induced lipid peroxidation of free radicals and caused an increase in lipid peroxidation components (TBARS). Three-fold and eight-fold concentrated liquids of solid culture of Antrodia camphorata according to the present invention As its concentration increases, the results show that it can significantly inhibit the peroxidation reaction and have the ability to resist free radicals. O: \ 7l \ 71526.DOC \ WCK-21 _ This paper size is applicable to Chinese National Standard (CNS) A4 (210 X 297 mm) 565430 A7 B7 V. Description of the invention (19) Liver protection experiment established with tetrachloride The experimental model of chemical carbon-induced chronic liver injury in mice was explored to investigate the effects of Antrodia camphorata at different concentration multiples on chronic liver injury in mice. Twenty-four mice without disease (ICR strain) were divided into four groups, as shown in Table 5. Table V. Antrodia cinnamomea test dose and animal group distribution group Carbon tetrachloride CCU dose dose level A 0 (control group) 0 (control group) 6 B 40% CCU / olive oil (0.3 ml / 100 g BW) 0 (control group) 6 C as above 50 mg (triple) / kg 6 D as above 50 mg (eight times) / kg 6 A and B to D mice were injected subcutaneously with olive oil (0.1 m 1/10 g BW) and 40% carbon tetrachloride CC 14 / olive oil (0.1 m 1/10 g BW) twice a week. Mice in groups A and B were administered with gastric tubes. In salt water, in the C and D groups, two concentrated doses of Antrodia camphorata cultivated in accordance with the method of the present invention were orally administered separately for five weeks. After the fifth week, all mice were tested for blood-related biochemical values by orbital blood collection. The mice were sacrificed and their livers were taken and fixed to the largest right lobe liver group. O: \ 71 \ 71520.DOC \ WCK ~ 22 ~ Paper size applies Chinese National Standard (CNS) A4 specification (210 X 297 mm) 565430 A7 B7 V. Description of the invention (20 Weaving for histopathological observation, such as liver cell damage, steatosis, necrosis and fibrosis, etc. The degree of biochemical function includes the blood surface: S Ai-pyramid grape transaminase (GPTamic) and Glutamic oxaloacetic transaminase (G〇T) ) Enzyme activity, GPT and GOT detection principles are based on international federal clinical chemistry standard methods. Male test results showed 7F, GPT and GOT values in mouse plasma under chronic liver injury with carbon tetrachloride increased significantly after 5 weeks of experiment The GpT value is about S50 units, which is 20-22 times the normal value; and the got value is about 1700 units, which is 40-42 times the normal value (Figures 4 and 5). , Feeding three times and eight times the concentrated Antrodia camphorata The increase in GPT and G0T values caused by carbon tetrachloride. Especially when the GpT and GOT values reach their peaks, the use of three times the concentrated dose can inhibit the increase of about 37% GPT and 65% GOT values. It shows that compared with normal mice liver tissues, after six weeks of treatment with four gaseous carbon, the liver tissues were significantly enlarged and the surface was gray and rough; however, silver food mice with different concentrations of Antrodia camphorata The liver did not show swelling or grayish rough surface. In addition, significant accumulation of fat particles was found under staining. Hepatocytes near the central veins were significantly enlarged. Giant heart cells containing iron metabolites and their erythrocytes were also inflammatory. Only in the liver cells' and part of the cell necrosis can be seen in nodular transformation transf () rmatl () n). Different silver food -23

O: \ 7l \ 71520.DOC \ WCK This paper size is applicable to the Chinese National Standard (CNS) A4 specification (21 × x297 mm) 565430, .A7 V. Description of the invention 1 ~ &quot; 21 from Toshiba, '' And, ', A &lt; liver tissues did not find the above pathological changes (see Figure 7). From the outside, Table 6 is an example of mouse liver pathological tissue analysis (N table normal group) Table D, carbon tetrachloride injury group, table X3 fed with barley citrate three-times concentrated group and table X8 table fed antrodia cinnamomea eight-fold concentrated group), of which, it can be confirmed that Antrodia cinnamomea condensate significantly improved pathological changes. Table 6. Semi-quantitative analysis of liver pathological tissues in mice. Necrosis of bile duct hypercalcification cells Necrosis Connection necrosis tubercle spine j N 0 0 0-1 0 0 0 D 2-3 2-3 2 2 0-1 1 X3 1 1 -2 0-1 1-2 0-1 0-1 X8 0-1 — 0-1 1 0 0-1 m ^; js w system inflammation and degeneration of liver 0 0-1 0-1 0-1, food industry development The institute accepts the entrustment of the Antrodia camphorata solid culture concentrate for cervical cancer (H e L a), wr Λ &quot; J ^ ^ (AGS), milk brigade (MCF-7), liver cancer (HepG2), and etiquette redundant

uotisn) and other cancer cell inhibition tests, the rate of inhibition of fat growth is shown in Figure 尽, and the inhibitory power of the solid culture concentrate of Antrodia camphorata (75%-87%) is invented, where the milk # ^ ^ / 、 Strong Dingzha cells (MCF_7) -24-

O: \ 71 \ 71520.DOC \ WCK This paper size is applicable to China National Standard (CNS) A4 (210 X 297 mm) 5 ^ 430 A7 B7

Strongest inhibition Table VII. Results of tumor cell test results of Antrodia camphorata solid culture concentrated solution Cell strains ~ 1 ------ Mean (%) ---------- Soil H e 1 a 8 3 Shigs 84 soil HepG2 75 soil MCF-7 8 7 The final concentration of the soil test sample is one tenth of the original sample concentration

SD 2.7 2.4 1.4

In addition, five kinds of powder samples (10 mg / ml) (Newl, New2, New3, New4, and New5) and eleven liquid samples (20L, New2, New3, 5 2 0, 521, 5 2 2, 5) were used. 2 3, 5 2 4, 5 2 5, 5 2 6 and 5 2 7) and their 10-fold diluted samples were detected by microculture tetrazoliUrn assay for colon cancer cells (C 0 L〇3 2 0 HSR) inhibition test. The results are shown in Fig. 8. It shows that at the final concentration of 1/10 the stock solution has at least 30% inhibitory power, and the sample 20 L has the strongest inhibitory power, which can inhibit the intestinal cancer cell ability up to 90%. In vitro chromosome structural variation I

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565430 A7 B7 V. Description of the invention (23) Further analysis of chromosomal structure in vitro using Antrodia camphorata according to the present invention, the method is to treat Chinese hamster ovary cells with Antrodia camphorata according to the present invention, and then analyze . Metaphase cells, observe and calculate how often they cause abnormal chromosomes. This chromosome structural variation analysis method can be used to evaluate the ability of Antrodia camphorata to cause damage to cell chromosomes. The analysis was performed in the following three treatments. The first was treated with Antrodia camphorata for 3 hours without rat liver activating enzyme (S 9) system; the second was treated with S 9 and treated with cells 3 Hours; the third is to continuously treat the cells for 20 hours without adding ς9. The test concentrations and experimental results are shown in Table 8. The test concentrations of Antrodia camphorata according to the present invention under the three treatment methods did not significantly increase the frequency of chromosome mutation. Table 8. Analysis of in vitro chromosome structural variation. Experimental subjects. Number of experimental cases. Intake and period of experiment. Chinese hamster ovary cells (ATCC, repository No. CCL-61 CH〇-Kl). 60 per plate according to test concentration. mm cell culture dishes with 2 concentrations of each, divided into 3 groups according to different processing conditions. The first and second groups are tested at concentrations of 0,312.5, 625, 1250, 2500 and 5000 pg / ml. The third group is tested at 0, 30. , 100, 300, 1000, and 3000 pg / ml cells with 18 to 21 chromosomes and uniform spread after culture according to individual conditions are selected for observation. The results of the three groups show that: Antrodia camphorata according to the present invention is treated before and after metabolic activation and at different times. There was no significant increase in the frequency of chromosomal abnormalities induced by Chinese hamster ovary cells. Therefore, the result of this experiment is negative. -26-

O: \ 71 \ 71526.DOC \ WCK This paper size is applicable to the Chinese National Standard (CNS) A4 specification (210 X 297 mm) 565430 A7 B7 V. Description of the invention (24 Small nuclear analysis in animals according to the invention Antrodia In the case of small nucleated red blood cells causing small nuclei, to determine whether Antrodia camphorata according to the present invention has the ability of chromosomal mutation (clastogenicity) in animals, and further provide whether Antrodia camphorata according to the present invention is capable of transmitting to humans. Risk assessment of toxicity. 25 mice were divided into five groups for testing, and the test methods and results are shown in Table 9. The micronucleus analysis of the mice showed that Antrodia camphorata according to the present invention has a significant effect on the peripheral blood of mice. The results of small nuclear analysis were negative. Table IX. The number of small nuclear analysis subjects in animals. Experimental results • ICR mice scores 1. Solvent control group 2. High-dose group 3. Medium-dose group 4. Low-dose group 5. Positive control group: 5 male rats in each group, 1-4 groups were orally administered with doses of 0, 500, 1000, and 2000 mg / Kg; positive control group received 1.0 mg / Kg MMC. Light microscope Observation of reticular red blood cells 1. There was no significant difference in body weight between the dose group and the solvent control group 2. Analysis of the proportion of reticular red blood cells: proved that no bone marrow toxicity was caused in mice 3. There was no dose-response trend in the proportion of small nucleus, and the comparison of control There is no significant increase. Therefore, the micronucleus analysis results of Antrodia camphorata on the peripheral blood of mice are negative. O: \ 71 \ 71526.DOC \ WCK -27- This paper size applies the Chinese National Standard (CNS) A4 specification ( 210 X 297 mm) 565430 A7 B7 V. Description of the invention (25 Based on the above pharmacological data, it can support that the Antrodia camphorata solids according to the present invention not only have components similar to wild forms, such as polysaccharides and triterpenoids, and Anti-lipid peroxidation, anti-free radical or anti-oxidant effects are also found. In addition, the solid substance of Antrodia camphorata according to the present invention has liver protection and anti-cancer effects. -28-

O: \ 71 \ 71526.DOC \ WCK This paper size applies to China National Standard (CNS) A4 (210 X 297 mm)

Claims (1)

565430 Patent Application No. 090118468 A8 Chinese Application for Patent Scope Replacement (1992 g) C8 VI. Application for Patent Scope 1 · A method for cultivating solid Antrodia camphorata, which includes the registration number CCRC 3 5 3 98 is the source of the bacteria in the space bag, the temperature is 5 to 32 ° C, and the mycelium is cultured in the air containing 0.2-1% carbon dioxide, and then the space bag is removed and exposed Under the condition that the carbon dioxide content is less than 1% and the air humidity is between 90 and 95%, the fruiting body is cultivated for a period of time until the cone-shaped fruiting body is formed; 60-70% fibrous material selected from mushrooms, plant stems, stalks, fruits and sawdust, 20-30% yam potato, 5-15% rice bran, 1-10% glucose, 0.2 % Dihydrogen phosphate bell and 0 · 1-1% magnesium sulfate, its humidity is between 6 〇 § 0%, and its p Η value is adjusted to neutral; and the daytime temperature during the cultivation of fruiting bodies is medium Between 20 and 30. (: Between 'and the night temperature is between 8 _ 丨 4, and the temperature difference between day and night is controlled between 12 and 16 ° C. 2 · According to the method in the scope of the patent application, the space pack The culture medium consists of 65% herb stems, stalks, fruits or sawdust, 20% yam potato, 10% rice bran, 3.5% glucose, 1% dihydrogen dihydrogen and 0.5% magnesium sulfate. 3. The method according to item i of the scope of patent application, wherein the temperature of mycelium culture is 28 ° C. 4. The method according to the first scope of patent application, wherein the air in the space bag O: \ 71 \ 71526-920820.DOC \ 5-This paper size applies to Chinese National Standard (CNS) A4 (210 x 297 mm) 565430 Αδ B8 C8 The humidity is 80%. 5. According to Article 1 of the scope of the patent application, the daytime temperature of the fruiting body when it is cultivated is 2 4-2 6 t, and the temperature is 10-12. (:. 6. According to the scope of the patent application No. 1 ^ negative negative method, in which the temperature difference between day and night is 15 ° c. 7. According to the scope of the patent application No. 1 million method, the mycelium culture is carried out 50 ° 80 days, and the fruiting body culture must be 20-50 days. 8. According to the application of the scope of the patent application, i, Wanfa, before the mycelium culture stage, it can be treated with a large number of bacteria tomb, ruler ^ The cultivation step includes (1) taking the activated bacteria from the liquid nitrogen preservation, taking one mountain, one, one, and one 廿 — a small piece of mycelium mycelium, transferred to a fresh medium and cultivated at a constant temperature; (?, You_ # t, 2) When the maggot grows to a vigorous level, inoculate the bacterium to a sterilized fibrous material (such as wheat kernels or wood chips)… in a bag and continue to grow at a constant temperature; When the seed is full of mycelium, remove the uppermost old hypha piece, and then inoculate it into the space bag in large quantities. 9. According to the method of item 丨 in the scope of the article, it contains a Large number of bacteria culture steps, including liquid culture fermentation to mass culture bacteria, oblique interview tube culture After that, 5 liters of fermented culture medium inoculated with a large number of propagated bacteria was cultured at 24 rpm for 24 days at 24 rpm with rotary shaking and shaking, and then continued at 90 rpm for about 14 days, and then inoculated to 20 liters. Stir culture in liquid fermentation broth for 14 days; the formula of the fermentation broth is fructose, -2- binding O: \ 71 \ 71526-920820.DOC 5 Winter paper scale suitable for China National Takazaki Standard (CNS) A4 Specification (210X 297 (Mm) 565430 A8 B8 C8 D8 6. Application scope of patents 0 · 0 0 0 1% magnesium sulfate, 0 · ii% yeast extract 0005 _ 05% potassium dihydrogen phosphate 10. According to the scope of patent application The method of 9 items, wherein the formula of the fermentation broth is 2% fructose, 0.05% magnesium sulfate, 0.5% yeast extract, and 0.1% potassium dihydrogen phosphate. The solid body of stick zhizhi fruit cultivated by the method according to the item is characterized in that the tangzhi fruit body has the physiological activity of triterpenoids similar to that of wild-type cinnamomum cinnamomea, and its south performance liquid chromatography conditions are: A Mobile phase: 1) 30% acetonitrile / 70% 1% acetic acid-for 50 minutes 2) 7 0% acetic acid cream / 30% 1% acetic acid-for 50 minutes 3) 9 9% acetonitrile / 1% acetic acid-for 30 minutes B · Stationary phase: C 1 8-chromatographic column (Merck No. 938071) 0 · 5 cm diameter X 2 5 cm length C · Temperature: 42 ° CD · Flow rate: 1 ml / min E. Detector: UV (2 54nm), with a retention time of about 46.6, 69.6 4 specific wave peaks of 80.7 and 87.8 minutes, and its shape is conical, with a diameter of 10-20 cm and a height of 20_ O: \ 71 \ 71526-920820.DOC 5 -3- The size of clothing and paper is common in China Standard (CNS) Α4 size (210 X 297 mm) Η 565430 6. Scope of Patent Application 30 cm and weight 0.3-0.5 kg. 12. —A product of Antrodia camphorata fruit solids, characterized in that the Antrodia camphorata fruit solids according to item 11 of the patent application is pulverized, and extracted with ^ or water and concentrated to a viscous state to obtain a concentrated liquid. . 13. The product according to item 12 of the scope of patent application, which can be further dried into a dry powder. 14. The product according to item 13 of the scope of patent application, the dried powder of which can be conditioned by water and then dried to obtain granulated products. 15. The product according to item 12 of the scope of patent application, which is a food composition. 16. The solid body of Antrodia camphorata according to item 11 of the scope of patent application, which has a liver-protecting function. -4- O: \ 71 \ 71526-920820.DOC 5 This paper is suitable for Chinese National Standard (CNS) A4 size (210 X 297 mm)