200920389 九、發明說明： 4 【發明所屬之技術領域】 本發明係關於活化動物體内免疫細胞活性的組成物 及其製備方法。 【先前技術】 近年來有ό午多研究發現菇蕈菌絲體或子實體中含有 相當多的生理活性成分，包含多醣體、三帖類 (tnterpencnds)、蛋白質、核酸、類固醇等，上述成分 具有抗腫瘤、增強免疫力、降低高血壓、降血糖、抑制血 小板凝集、抗病毒等功效。 已知多醣體具有抗腫瘤的功效，其生理活性可依多釀 體之分子量大致區分為三類：⑴分子量S 3，_〜5_ 左右者，具有降低血糖之功能，⑻分子量在 1〇,〇〇〇〜1〇〇，_之間者，具消炎的作用，（〇分子^ 30, 000以上者，則呈右杆陆 里在 愈佳。 L有抗腫瘤作用，且分子量愈大效用 以往兹輩多膽體大多萃取自子實體，但是子實 有限’近來研究已知約冑5"兹蕈類 產、 生胞外多醣體，所以可取苴谇養 反心k養中產 於子實體培養，液態培養較;㈣純化;相較 萃取==蕈:Γ物之產品以，特別是_蕈 卞取物之健康…因此，增加菇蕈早 方、菇蕈萃取物之新應用或新產品等相關需7:之配 要性。 自需求’仍有其必 U0560 5 200920389 ‘在健康食品的領域中，益生菌亦是研究重點之—。人 .體的腸道内戶斤分布的菌相，據估計有超過i〇m重以上的细 ，存在’其中由30〜40種細菌構成99%的菌量，且厭气 氧菌占優勢’約多—百到—萬倍。消化道不同區： 存在者不同的菌種及不同的數量分佈。這些腸内菌群彼此 升:成共生 '拮抗的關係，而在健康的個體中維持 :=ν·:部分細菌對人體有害，例如:產㈣ Γ 2 (心wper如她）及金黃葡萄球菌 咖），會因宿主免疫力下降而祠 大:成疾病，然而，棲息於健康人體腸道的細菌絶 二」人體有盈，-般稱為益菌（beneficlal teri/)，其中以乳酸菌群（Lactic acid bacter 或雙叉乳桿g群W /id⑽咖⑺·⑽）最為人所知。 之功处有=研九報告心出’盈菌具有多種促進人體健康 ^⑴維持腸道正常微生物相：藉由益菌代 、值，的醋酸和乳酸’使腸道有效維持在較低的邱 ::使她及腐敗性細菌不易生長；（2)活化免疫系 性.二囷具有刺激免疫系統、增加抗體或巨嘆細胞的活 壁含有某些抗原成分，可誘發宿主免疫系統活 膜ΓΓ瘤，或使宿主產生抗體以抑制病原菌吸附在腸 （3)提高營養價值：以雙叉桿菌為例，其 物==用及蛋白質水解活性，可改變食物中含氮化合 高蛋白質消化率，並可促料及其他營養之吸 進而預防骨質疏鬆症，甚至，某些菌株可合成多種維 110560 6 200920389 ▲素與葉酸，（4 )促進乳酸并他· ^ κ、 L — 孔齔代5射，（5)改善腹瀉便秘： .ϋ囷產生的有機酸能增進腊洁φ令舌^ ^ θ s道柘動，提高糞便中的水分含 置，以降低腸阻滯；（6 )降桐A A缺门 ^ ~低血中膽固醇：雖然此功能 仍具爭議，但由部分體外續矜姓 π3Κ, t χ 卜°式鲂結果侍知，某些益菌能使膽 固醇與去結合型膽鹽共沈殿而隋 _ 此我而蚣糞便排出人體，亦有動物 5式'1¾結果頒不，攝食穿丨祕Μ -r r?女 攝"L酉夂囷可降低血中膽固醇的濃度;⑺ 抑制南血屢：益菌的羋也并％ 4 /、二代6射物此抑制血管收縮素轉化而 使血壓降低。 科學豕研究盈菌的作用嫂連,丨& n α’作用機制而發現益菌會生產益菌 助生貝(prebi〇tics)，對人财辟由 好的作用。 子人粗健康的促進或維護有最良 纟^月依上述概念並因應市場需求’開發出具 保健功效的新穎組合物。 【發明内容】 活性2:::目的在提供-種活化動物體内免疫細胞 為=述及其他目的，本發明的紅成物係包括：益蕈 成刀、涵·菌成分，及賦形劑。 根據本發明，兹蕈成分包含兹葦菌絲體 物之菇蕈成分白人w 4 4 月、‘且成 成刀匕3姑覃多醣體，特別是葡 明的菇蕈成分降了釭笛夕Α Ρ匍來椐本發 铷η 除 覃多畴體之外’亦可包含兹蕈代謝產 物’例如：茶白暂 座 Α白貝夕肽類、三帖類、多種胺基酸、壬私 驗、酯類和有機酸望抓所 物 …/ 質。本發明之組成物，可包含單- 早成分，或多種菇蕈成分之混合物。 H0560 7 200920389 根據本發明，所述兹蕈係選自裂褶菌（Sci] izophy 7 J⑽ Λ co/nfflue)、巴西蘑菇（Agarics )、冬蟲夏草 (Cordyceps sinensis')、I lt(Ganoderim lucidwn)、雲 文（Coriolus versi’ccJojr)、樟芝（ camphor ate )、桑黃(Phellinus linteus )、珊姑月 | 去 (Pleurotus citrinopileatus )、香益 (Lentinula edodes)' Agrocybe aegeri ta) - m m Hericium eri naceus )、杏鮑益(Pleurotus eryngi ig^) ^ 花瓣茸 (Sparrasis crispa)、木茸（ji/Wcdaria auricda)、 金針菇（Flammulina velutipes)及上述兹蕈之組合。 根據本發明’益菌成分包含益菌菌體，且該益菌係選 自嗜酸性乳酸桿菌（LactohaciHws acidopWks)、雙 叉乳桿菌（5i/idc^acieriu/n7o叹⑽）、副乾酪乳酸桿菌 paracaseO、腸球菌（如ter〇c〇ccus /aeciu/n)及其組合。 ( 根據本發明，益菌成分包含益菌菌種於培養過程中所 產生的益菌助生質。 根據本發明，益菌成分係益菌菌體、益菌助生質或其 混合物。 根據本發明，賦形劑係，舉例但非限制’薏仁、乳糖、 白糖、D -甘露醇、殺粉、玉米殿粉、纖維素晶體，或無水 矽酸·’於較佳實施例中，賦形劑為乳糖；於另一較佳實施 例中，賦形劑為薏仁。 本發明之組成物進一步包含添加物，該添加物可為藥 110560 8 200920389 學上、食品上、或美容上可接受之添加物。 於本發明之實施例中，添加物為食品；於較佳實施例 中’添加物係選自米類、麥類、黍類、豆類、芝麻、蓖麻 及其混合物；於更佳實施例中，添加物為鍵米、玉米，或 兩者之混合物。 本發明之另一構想係提供包含一種或多種菇蕈成 分、一種或多種益菌成分及賦形劑的組成物。於較佳實施 例中’該兹蕈係選自裂褶菌co腳i/e)、巴 / : 西蘑菇（dearies biaze )、冬蟲夏草（Cordyceps sijiensjs)、靈芝（Ga/ioder/na !uciduffl)、雲芝（CorioJus vers!co!or)、樟芝（Znaodia ca/np/]〇rate)、桑黃 (Phel 1inus 11nteus )、珊蝴兹(P1eurotus cjtrj'Tiopj'ieaius)、香兹（Le/7ii/7i/_/a edodes)、柳松兹 (Agrocybeaegerita)、^S^^(Hericiuinerinaceus)、 杏飽菇（W euro tus 发）、花瓣茸（Spar r as is crispa )、木茸（如rict/iaWa awricuia )、金針兹 l... (Flammul ina velutipes )及其組合；該益菌係選自嗜酸 性乳酸桿菌（Lacto6acj_nzis acidop/jj’Ji/s)、雙叉乳桿 菌 (Bifidobacterium longum )、畐1J摩乞路乳酸桿菌 〔Lactobaci 1 lus paracasei )、腸球菌（Enterococcus /aeciuffi)及其組合；且該賦形劑為乳糖或薏仁。 根據本發明，該組成物可供直接使用，或可視需要混 合載體、藥物、食品、美容產品，或其他成分以供後續使 用。 9 110560 200920389 " 本發明之另一構想係提供包含一種或多種菇蕈成 分、一種或多種益菌成分及添加物的組成物。於較佳實施 例中，該兹蕈係選自裂褶菌co丽i/e)、巴 西蘑H ( paries bhze )、冬蟲夏草（Cordyceps sinensis)、靈芝（Ganoderma lucidum)、雲芝（Coriolus versi col or)、禅芝（Anthodi a camphorate )、桑黃 (Phe 11 inus 1 inteus )、珊蝴益（_P_/eL/rotL/s ciiriwopUeatus)、香菇edoefes)、柳松菇 (Agrocybe aegeri ta ) ^ ^ ^ ( Hericium erinaceus) ' 杏鮑菇（尸Jeurotus ejT/3《ij·^·) ' 花瓣茸（Sparrasis crispa )、木茸（iiirici/iaria auricula ) ' 金針菇 (Flammul ina velutipes )及其組合；該益菌係選自嗜酸 性乳酸桿菌（Lactohacinus acj’dop/n’Jtys)、雙叉乳桿 菌 （Bifidobacterium longum )、畐丨J乾酿乳酸桿菌 (Lactobaci 1 lus paracasei)、月昜球菌(Enterococcus /aeciL//H)及其組合；該添加物係選自米類、麥類、黍類、 V*. 豆類、芝麻、蓖麻及其混合物。於更佳實施例中，該添加 物為链米、玉米’或兩者之混合物。 根據本發明’該組成物可供直接使用，或可視需要混 合載體、藥物、食品、美容產品，或其他成分以供後續使 用。 ' 本發明之另一目的在提供一種製備活化動物體内免 疫細胞活性的組成物之方法’係包括下列步驟：取菇蕈菌 絲體進行液態培養，以獲得菇蕈成分；以適當培養基培養 10 110560 200920389 盈囷’以獲付盈囷成分；及混合該兹蕈成分、益菌成分與 賦形劑，以獲得該組成物。 根據本發明，該兹蕈係選自裂褶菌（ScAizop/jyn⑽ CO/»腳e)、巴西蘑菇（知arics Maze )、冬蟲夏草 (Cordyceps sinensis')、I 芝（Ganoderma lucidum)、雲 定（Coriolus versicolor)、槔芝（Anthodia camphorate)、桑黃(Phellinus linteus )、珊蝴益 (Pleurotus citrinopileatus )、香兹 (Lentinula f ·. edodes) ' Agrocybe aegeri ta)- m ^ tiericium eri/7acei/s)、杏飽菇（PJeurotiys 、花瓣茸 (Sparrasis crispa)、木茸（di/j-icdaria auricLda)、 金針兹(F lammul ina velut ipes )及其組合。 根據本發明，液癌培養所得之兹蕈成分包含兹蕈菌絲 體，及/或菇蕈多醣體。 根據本發明’該益菌係選自嗜酸性乳酸桿菌 (Lactobacillus acidophilus ) 、雙叉 桿菌 (Bifidobacterium longum )、畐ij 章乞路 ^ 酉复桿菌 (Lactobacillus paracasei)、腸球镜（Enter〇c〇ccus faeci⑽）及其組合。 根據本發明’經適當培養基所獲得之益菌成分包含益 菌菌體、及/或益菌助生質。 根據本發明’該賦形劑係薏仁 '乳糖、白糖、D_甘露 醇、澱粉、玉米澱粉、纖維素晶體或無水矽酸；於較佳實 施例中’ §玄赋形劑為乳糖，於另一較佳實施例中，該賦形 110560 11 200920389 劑為薏仁。200920389 IX. INSTRUCTIONS: 4 TECHNICAL FIELD OF THE INVENTION The present invention relates to a composition for activating immune cell activity in an animal and a preparation method thereof. [Prior Art] In recent years, many studies have found that the mycelium or fruiting body of the mushroom has a considerable number of physiologically active components, including polysaccharides, tnterpencnds, proteins, nucleic acids, steroids, etc. Anti-tumor, enhance immunity, reduce high blood pressure, lower blood sugar, inhibit platelet aggregation, anti-virus and other effects. Polysaccharides are known to have anti-tumor effects, and their physiological activities can be roughly classified into three categories according to the molecular weight of the multi-breast: (1) the molecular weight S 3, _~5_ or so, has the function of lowering blood sugar, and (8) the molecular weight is 1 〇, 〇 〇〇~1〇〇, _ between, has anti-inflammatory effect, (〇 〇 ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ Most of the genus of the genus is extracted from the fruiting body, but the genus is limited. Recently, it has been known that 胄5" 蕈 蕈 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 Compared with the extraction ==蕈: the product of the sputum, especially the health of the sputum... Therefore, the new application of the mushroom, the new application of the mushroom extract, or the new product is required. 7: The need for sex. Since the demand 'still has its U0560 5 200920389 'Probiotics are also the focus of research in the field of health foods. - The body's intestinal flora is distributed in the gut phase, it is estimated that there are More than i〇m heavy, there exists 'where 30 to 40 kinds of bacteria It is 99% of the amount of bacteria, and the anaerobic bacteria predominates 'about more than one hundred to ten thousand times. Different areas of the digestive tract: different species of bacteria and different quantity distribution. These intestinal flora rises each other: Symbiosis 'antagonistic relationship, and maintained in healthy individuals: = ν ·: Some bacteria are harmful to the human body, for example: production (four) Γ 2 (heart wper like her) and golden staphylococcus), due to decreased host immunity祠大:成病, however, the bacteria that inhabit the intestines of healthy humans are the second most abundant, the human body is full, commonly known as beneficlal teri/, which is a group of lactic acid bacteria (Lactic acid bacter or double-forked rods) W / id (10) coffee (7) · (10)) is best known. There are = research report 9 out of the heart of the 'small bacteria have a variety of promoting human health ^ (1) maintain the normal microbial phase of the intestine: by the beneficial bacteria, the value of acetic acid and lactic acid 'to keep the intestinal tract effectively in the lower Qiu :: Makes her and spoilage bacteria difficult to grow; (2) activates the immune system. Dioxins have a living wall that stimulates the immune system, increases antibodies or stun cells, contains certain antigenic components, and induces the immune system of the host immune system. Or causing the host to produce antibodies to inhibit the adsorption of pathogenic bacteria in the intestine (3) to improve the nutritional value: for example, the use of bifidobacteria, its use = and protein hydrolyzing activity, can change the digestibility of high-protein containing nitrogen in food, and Promote and other nutrient absorption to prevent osteoporosis, and even some strains can synthesize multiple dimensions 110560 6 200920389 ▲ 素 and folic acid, (4) promote lactic acid and he · ^ κ, L - 龀 龀 代 5, (5 Improve diarrhea constipation: . The organic acid produced by ϋ囷 can enhance the 洁 φ 令 ^ ^ ^ ^ θ s 柘 柘 , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , ~ low blood cholesterol: although this It can still be controversial, but some of the in vitro 矜3π, t χ ° 鲂 鲂 侍 侍 侍 侍 侍 侍 侍 侍 侍 侍 侍 侍 侍 侍 侍 侍 侍 侍 侍 侍 侍 侍 侍 侍 某些 某些 某些 某些 某些 某些 某些 某些 某些 某些 某些 某些 某些 某些 某些There are also animal type 5 '13⁄4 results are not awarded, eating through the secrets -rr? Female photo " L酉夂囷 can reduce the concentration of blood cholesterol; (7) inhibit the South blood repeatedly: the bacteria of the beneficial bacteria also% 4 / The second generation of 6 shots inhibits the conversion of angiotensin and lowers blood pressure. The scientific research on the role of bacteria, Qilian, 丨 & n α' mechanism of action and discovery of beneficial bacteria will produce probiotics (prebi〇tics), good for people. The promotion or maintenance of the sub-health is the best. 依^月 Based on the above concepts and in response to market demand, we have developed a novel composition with health benefits. SUMMARY OF THE INVENTION Activity 2::: The purpose of providing immune cells in an activated animal is as described for other purposes. The red-yielding system of the present invention includes: Yisui Chengdao, han·bacteria components, and excipients. . According to the present invention, the scorpion ingredient comprises the white scorpion scorpion component of the scorpion scorpion body w 4 April, and the scorpion scorpion polysaccharide of the scorpion scorpion Ρ匍 椐 椐 铷 铷 覃 覃 覃 覃 覃 覃 覃 覃 覃 覃 覃 覃 覃 覃 覃 覃 覃 ' ' ' ' ' ' ' ' ' ' ' 蕈 蕈 蕈 蕈 蕈 蕈 蕈 蕈 蕈 蕈 蕈 蕈 蕈 蕈 蕈Esters and organic acids are expected to catch.../quality. The composition of the present invention may comprise a single-early ingredient, or a mixture of various mushroom ingredients. H0560 7 200920389 According to the present invention, the scorpion is selected from the group consisting of Sci] izophy 7 J(10) Λ co/nfflue), Brazilian mushroom (Agarics), Cordyceps sinensis, lt (Ganoderim lucidwn), cloud (Coriolus versi'ccJojr), camphor ate, Phellinus linteus, Pleurotus citrinopileatus, Lentinula edodes 'Agrocybe aegeri ta) - mm Hericium eri naceus ), Pleurotus eryngi ig^ ^Sparrasis crispa, ji/Wcdaria auricda, Flammulina velutipes and combinations of the above. According to the present invention, the beneficial bacteria component comprises a beneficial bacterial cell selected from the group consisting of Lactohaci Hws acidop Wks, Lactobacillus bifidum (5i/idc^acieriu/n7o sigh (10)), and Lactobacillus paracasei paracaseO. Enterococcus (such as ter〇c〇ccus / aeciu/n) and combinations thereof. (According to the present invention, the beneficial bacteria component comprises the beneficial bacteria helper produced by the beneficial bacteria in the culture process. According to the present invention, the beneficial fungi component is a beneficial bacterial cell, a beneficial bacterial helper or a mixture thereof. According to the present invention, Excipients are, but are not limited to, 'barley, lactose, white sugar, D-mannitol, powdered powder, corn powder, cellulose crystals, or anhydrous citric acid.' In a preferred embodiment, the excipient is lactose In another preferred embodiment, the excipient is coix seed. The composition of the present invention further comprises an additive which may be a pharmaceutical, food, or cosmetically acceptable addition to the drug 110560 8 200920389. In an embodiment of the invention, the additive is a food product; in a preferred embodiment the 'additive is selected from the group consisting of rice, wheat, medlar, beans, sesame, ramie, and mixtures thereof; in a more preferred embodiment The additive is a key rice, corn, or a mixture of the two. Another idea of the present invention is to provide a composition comprising one or more mushroom trout ingredients, one or more beneficial fungi ingredients, and excipients. 'The 蕈 蕈 is selected from Pleurotus co-foot i/e), Bar/: dearies biaze, Cordyceps sijiensjs, Ganoderma lucidum (Ga/ioder/na!uciduffl), Coriolis (Ceo s vers! co!or), Antrodia Znaodia ca/np/]〇rate), Phel 1inus 11nteus, P1eurotus cjtrj'Tiopj'ieaius, Lez 7i/7i/_/a edodes, Agrocybeaegerita ), ^S^^(Hericiuinerinaceus), Apricot mushroom (W euro tus hair), Spar r as is crispa, Wood velvet (such as rict/iaWa awricuia), Golden needles l... (Flammul ina velutipes And a combination thereof; the beneficial bacteria are selected from the group consisting of Lactobacillus acidophilus (Lacto6acj_nzis acidop/jj'Ji/s), Bifidobacterium longum, Lactobaci 1 lus paracasei, intestine Enterococcus / aeciuffi and combinations thereof; and the excipient is lactose or coix seed. According to the present invention, the composition can be used as it is, or a carrier, a drug, a food, a cosmetic product, or other ingredients can be mixed as needed for subsequent use. 9 110560 200920389 " Another idea of the present invention is to provide a composition comprising one or more mushroom components, one or more beneficial components and additives. In a preferred embodiment, the strain is selected from the group consisting of Schizophyllum coli i/e), Paria bhze, Cordyceps sinensis, Ganoderma lucidum, Coriolus versi col or ), Anthodi a camphorate, Phe 11 inus 1 inteus, _P_/eL/rotL/s ciiriwopUeatus, edoefes, Agrocybe aegeri ta ^ ^ ^ ( Hericium erinaceus) 'Pleurotus eryngii (Jeurotus ejT/3 "ij·^·) 'Sparrasis crispa, iiirici/iaria auricula 'Flammul ina velutipes' and combinations thereof; It is selected from Lactohacinus acj'dop/n'Jtys, Bifidobacterium longum, Lactobaci 1 lus paracasei, and Enterococcus /aeciL//H And combinations thereof; the additive is selected from the group consisting of rice, wheat, medlar, V*. beans, sesame, ramie, and mixtures thereof. In a more preferred embodiment, the additive is a chain of rice, corn' or a mixture of both. According to the present invention, the composition can be used as it is, or a carrier, a drug, a food, a cosmetic product, or other ingredients can be mixed as needed for subsequent use. 'Another object of the present invention is to provide a method for preparing a composition for activating immune cell activity in an animal' comprising the steps of: cultivating a mycelium of the mushroom to carry out liquid culture to obtain a mushroom meal component; and cultivating the medium in an appropriate medium 10 110560 200920389 Ying Ying 'to obtain the profit component; and to mix the ingredients, beneficial components and excipients to obtain the composition. According to the present invention, the mites are selected from the group consisting of Schizophyllum (ScAizop/jyn (10) CO/» foot e), Brazilian mushroom (known arcics Maze), Cordyceps sinensis, Ganoderma lucidum, Coriolus Versicolor), Anthodia camphorate, Phellinus linteus, Pleurotus citrinopileatus, Lentinula f. edodes 'Agrocybe aegeri ta)- m ^ tiericium eri/7acei/s), PJeurotiys, Sparrasis crispa, di/j-icdaria auric Lda, F lammul ina velut ipes, and combinations thereof. According to the present invention, the liquid component obtained by liquid cancer culture contains According to the present invention, the probiotic strain is selected from the group consisting of Lactobacillus acidophilus, Bifidobacterium longum, and 畐ij 乞路^酉酉(Lactobacillus paracasei), enterococci (Enter〇c〇ccus faeci (10)), and combinations thereof. According to the present invention, the beneficial bacteria component obtained by the appropriate medium comprises the beneficial bacteria, and/or the beneficial bacteria. The present invention is an excipient which is a hazelnut, lactose, white sugar, D-mannitol, starch, corn starch, cellulose crystals or anhydrous citric acid; in a preferred embodiment, the § 玄 excipient is lactose, in another In a preferred embodiment, the formulation 110560 11 200920389 is a coix seed.
I 本發明之方法進一步包含加入添加物以形成該組成 物’其中該添加物係選自米類 '麥類、黍類、豆類、芝麻、 蓖麻，及其混合物。 【實施方式】 以下係藉由特定的具體實施例說明本發明之實施方 式’熟習此技蟄之人士可由本說明書所揭示之内容瞭解本 發明之其他優點與功效。 1.菇蕈菌種培養 本發明可使用的兹蕈菌株如裂褶菌（Sc/jizophynuffi co丽ue)、巴西磨兹（)、冬蟲夏草 (Cordyceps sinensis')、靈芝（Ganoderma lucidum)、雲 芝(Coriolus versicolor)、掉芝(Anthodia camphorate)、桑黃（Phellinus linteus)、珊瑚兹 (Pleurotus citrinopileatus )、香兹(Lentinula edodes)' m M ( Agrocybe sp. ) ^ ^ ^ ^ ( Agrocybe ae^erita )、猴頭益(Hericium erinaceus)、杏，鮑益 (Pleurotus eryngiig) ' 花瓣茸(Sparrasis crispa)、 木茸（hriciiJaria ai/ricivh)、金針菇（Flammulina velutipes )等，但不限於上述菌株。選擇其一進行培養。 將菌種接種於YM瓊脂培養基（YM瓊脂培養基包含 0. 3%(w/w)酵母菌萃取物、0. 3%麥芽萃取物、〇. 5%消化蛋 白（peptone)、1 · 0%右旋糖（dextrose)、1. 5%洋菜（agar) 12 110560 200920389 等）上，置於28°C培養，並中片 ^ 八中依囷種不同，菌絲體長滿 u版培養基所需培養時問 约W… 20天，·例如：裂褶菌 相養傻2 —3天可見菌絲體，約5-7天可長滿固體培養基。 將上述固態培養菌鮮 、土 真 口，、糸肢切—小塊植入YM培養液50 笔升（m 1 )，於80 丁的τ荆知仏/ h 0 T型動物細胞培養盒中，在28°C 口養一週，作為液態前置醱酵的菌種。 ::口血清瓶中配製_毫升的⑽培養液並放入 在Τ型塑膠培養盒培養完成的兹輩菌液I The method of the present invention further comprises adding an additive to form the composition' wherein the additive is selected from the group consisting of rice, wheat, alfalfa, legumes, sesame, ramie, and mixtures thereof. [Embodiment] The following describes the embodiments of the present invention by way of specific embodiments. Those skilled in the art can understand other advantages and advantages of the present invention from the disclosure of the present disclosure. 1. Mushroom cultivar culture The sputum strains which can be used in the present invention, such as Schizophyllum sp. (Sc/jizophynuffi coliue), Brassica sinensis, Cordyceps sinensis, Ganoderma lucidum, Yunzhi ( Coriolus versicolor), Anthodia camphorate, Phellinus linteus, Pleurotus citrinopileatus, Lentinula edodes ' m M ( Agrocybe sp. ) ^ ^ ^ ^ ( Agrocybe ae^erita ), Hericium erinaceus, apricot, Pleurotus eryngiig 'Sparrasis crispa, wood stalk (hriciiJaria ai/ricivh), Flammulina velutipes, etc., but not limited to the above strains. Choose one to cultivate. The strain was inoculated on a YM agar medium (YM agar medium containing 0.3% (w/w) yeast extract, 0.3% malt extract, 〇. 5% digested protein (peptone), 1 · 0% Dextrose (dextrose), 1.5% acacia (agar) 12 110560 200920389, etc.), placed at 28 ° C culture, and medium tablets ^ eight in the different species, mycelium is overgrown with u-plate medium When it is necessary to raise the culture, it takes about 20 days. For example, the mycelium can be seen in 2 to 3 days, and the solid medium can be filled in about 5-7 days. The above-mentioned solid-state culture bacteria, fresh soil, and sputum limbs were cut into small pieces and implanted into YM culture solution for 50 liters (m 1 ) in 80 butyl τ 仏 仏 / h 0 T type animal cell culture box, It is incubated at 28 ° C for one week as a liquid pre-fermented strain. :: Prepare _ml of (10) medium in the serum bottle and place it in the sputum plastic culture box.
接種於血清瓶内，接種比 A 例為母10耄升新鮮培養液接種 笔i液’而後將血清瓶置於磁石攪拌機（轉速期 rpni)上，於宮、丁與拙 72 主'皿下攪拌，以進行第一次前置醱酵，培養 96小時後，胞外多醣體濃度達最高。 以相同步驟’於10公升血清瓶中配製約8公升i立養 液丨公升培養液中含10克消化蛋白、丨。克牛肉萃升取:養 克酵母菌萃取物、2G克右旋糖、i克Wn⑽、 檬酸錄、5古Γΐί rnrm 兄可丁 克 CH3C_a、〇. 1 克 MgSCUC . 7仏0、〇. 05 克 nSCUC 10與2克KzHPO4;並將酸鹼值調成pH 6 2〜6 5 )， 接種=次前置輯完成之菌液，進㈣二次前置_。 當然，得視需要進行多次的前置醱酵。接著， 酸酵完成之酸酵液進行大量酸酵。 ;20 a升聚碳酸樹脂（p〇iycarb〇nate，ρ〇典養容哭 中製13公升最適化的生產培養基（包含4㈣萄糖； 旧一 Y酵母菌萃取物）’攪拌均勻後，蓋上矽膠塞，以高 '皿网壓滅菌’其中滅菌條件為121〇C、15分鐘，放涼至室 110560 13 200920389 f。而後，將2—3公升上述第二次前置_完成之菌液接 .重入聚碳酸樹脂培養容器中，放置於搖擺機上，設定轉速 立為⑽聊，於常溫（約25。〇下進行_至生長曲線的 静止期（sut ionary phase)，此時期的菌絲體細胞數盥多 酶體濃度係維持衡定，即停止醱酵。所得培養液體包含輩 益多醣體、蛋白質、多肽類、三祐類、多種胺基酸、生物 鹼、酯類和有機酸等物質。 2·益菌培養及益菌代謝物製備 進行盈菌菌種培養，宜中贫κ益你$心 。脣” Y 3皿囷係為嗜酸性乳酸桿菌 L=ClUus acidophllus )、雙又乳样菌 (ifidobactenum longum )、副乾酪乳酸桿菌Inoculate in a serum bottle, inoculate a pentoid liquid in a sample of 10 liters of fresh culture solution in the case of A, and then place the serum bottle on a magnet mixer (rotation period rpni), and stir under the main dish of the palace, butyl and sorghum 72 For the first pre-fermentation, the concentration of extracellular polysaccharides was highest after 96 hours of culture. In the same procedure, about 8 liters of the literate liter culture solution containing 10 grams of digested protein and mash was prepared in a 10 liter serum bottle. Gram extract: yoke yeast extract, 2G gram dextrose, i gram Wn (10), citric acid record, 5 Γΐ ί rnrm brother can Dingke CH3C_a, 〇. 1 gram MgSCUC. 7仏0, 〇. 05 grams nSCUC 10 and 2 grams of KzHPO4; and adjust the pH value to pH 6 2~6 5), inoculate = the pre-set of the complete bacterial liquid, into the (four) secondary pre-_. Of course, it is necessary to carry out multiple pre-fermentation. Then, the sourdough of the acid yeast is used to carry out a large amount of acid fermentation. 20 a liter of polycarbonate resin (p〇iycarb〇nate, 〇 〇 养 养 哭 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 13 The silicone plug is sterilized by high 'steam pressure'. The sterilization condition is 121〇C, 15 minutes, and it is allowed to cool to room 110560 13 200920389 f. Then, 2-3 liters of the above second pre-completed bacterium is connected. Re-enter the polycarbonate resin culture vessel, place it on the swing machine, set the rotation speed to (10), and at room temperature (about 25. 〇 进行 to the sut ionary phase of the growth curve, the mycelium of this period The cell number and the concentration of the enzyme are maintained, that is, the fermentation is stopped. The obtained culture liquid contains polysaccharides, proteins, peptides, Sanyou, various amino acids, alkaloids, esters and organic acids. 2. Beneficial culture and probiotics metabolite preparation for the cultivation of bacteria, it is appropriate to be poor and you can benefit from the heart. The lip "Y 3 dish is Lactobacillus acidophilus L = ClUus acidophllus", double and milky Bacteria (ifidobactenum longum), Lactobacillus paracasei
Uactobaci】iUSparacasei) ’ 或腸球菌（加⑽c〇ccus faecium)。 將菌種接種於平面固體培養基（1公升培養液中含ι〇 克消化蛋白、ίο克牛肉萃取物、5克酵母菌萃取物、2〇 克右旋糖、1克Tween 8〇、2克檸檬酸銨、5克cH3C_a、 〇. 1 克 Mgs〇4c .瓜〇、〇· 05 克 MnS〇4C · h2〇、2 克— 與15克洋菜，並將酸鹼值調成ρΗ 6 2_6 5)，置於π =培養。依菌種不同，、約i天〜3天可於培養基上長出 洛0 挑單-菌落進行小量液態培養，接著進行大量 :料同的習知生長培養基成分可獲得不同的益菌。代謝 4共血囷助以。培養液可直接用於後續組成物製備，亦 Π0560 14 200920389 而僅以菌體供後續使 可將菌體與培養液以離心方式分開 用。 3.本發明組成物之製備 (實例一） 、方法所付的姑覃醱酵 後，利用高速均質19 囷培養液混合 )貝機以12,〇〇〇1*卵處理1 焓液。而後加入乳糖賤形“而侍均質 例為1:1，混人均七β 八中均貝浴液與賦形劑之比 心口均勻後’經冷凍後， (iy〇phiiization)處理，其中 -，皿…王乾燥 -4〇°C、0. 3 Mpa，获、，脸、_ '里，、二乾燥的操作條件為 藉以將混合物濃縮至开j 5 %的粉末。 、 ^成$水率低於 視冩要將乳 例如：薏仁、白 晶體’或無水石夕 曰状句丹他賦形 糖、D-甘露醇、澱粉、玉米澱粉、 酸。 (實例二） 合，液與益菌培養液混 溶液=::: = =處理1◦分鐘而得均質 為2: 1)，均勻混合後 』=液將比例 的混合物纟10〇C下利用言，處理。將冷;東乾燥後 有菇蕈成分盥^ 门1句貝機磨成粉末，以獲得含 、^囷成y刀的組成物粉末。 可視需要將輪来替換為其他穀類’例如：米類'麥類、 110560 15 200920389 暴類、豆類、芝麻、蓖麻，及其混合物。 « 4.本發明組成物用於活化免疫細胞活性之效果 動物模式： 選用五週齡的雄性Wi star大白鼠（來源：台灣大學 動物中心）’重量為380〜450克。實驗前將大白鼠分別單 獨飼養，室溫控制於21±2。(：、濕度65±5%，光暗循環時 間為12D : 12L，待適應兩星期後開始進行實驗。 f-·. 以本發明組成物餵食大白鼠三天，每天0. 2毫升。三 天後使用 LIVE/DEAD Cell-Mediated Cytotoxicity 套經 (Molecular Probes)測定血液中自然殺手細胞活性，實驗 步驟如下： 將YAC-1細胞濃度調成ΐχΐ〇6細胞/毫升。依比例加 入DIOCu ( 10//LDIOCu/毫升YAC-1細胞懸浮液）並混和 均勻。於37°C培養20分鐘。將細胞濃度調成lxl06cell/ 毫升。Uactobaci] iUSparacasei) ' or Enterococcus (plus (10) c〇ccus faecium). Inoculate the strain in a flat solid medium (1 liter of culture medium containing ι grams of digested protein, ίο克 beef extract, 5 grams of yeast extract, 2 grams of dextrose, 1 gram of Tween 8 〇, 2 grams of lemon Ammonium acid, 5 g cH3C_a, 〇. 1 g Mgs〇4c. Melon, 〇· 05 g MnS〇4C · h2〇, 2 g - with 15 g of amaranth, and adjust the pH to ρΗ 6 2_6 5), Placed in π = culture. Depending on the strain, about 1 day to 3 days, you can grow a small amount of liquid culture on the medium, and then carry out a small amount of liquid culture. Then, a large amount of the same growth medium composition can be used to obtain different beneficial bacteria. Metabolism 4 total blood help. The culture solution can be directly used for the preparation of the subsequent composition, and is also used for the subsequent separation of the cells and the culture solution by centrifugation only by using the cells for subsequent use. 3. Preparation of the composition of the present invention (Example 1) After the agaric fermentation of the method, the high-quality homogenized 19 囷 culture medium was mixed with a shell machine to treat 1 mash with 12, 〇〇〇 1* eggs. Then add lactose 贱 shape and the average temperament is 1:1, mixed with the average of seven β 八 中 贝 浴 浴 与 与 赋形剂 赋形剂 赋形剂 赋形剂 赋形剂 ' ' ' ' ' ' ' ' ' ' ' ' ' ' ' 经 经 经 经 经 经 经 经 经 经 经 经...wang dry-4〇°C, 0. 3 Mpa, get, face, _ 'Li, 2, dry conditions are used to concentrate the mixture to j 5% powder. ^, the water rate is lower than It is necessary to treat milk such as: coix seed, white crystal 'or waterless stone, D-mannitol, starch, corn starch, acid. (Example 2), liquid mixed with beneficial bacteria culture solution Solution =::: = = 1 min to be treated to obtain a homogenization of 2: 1), after mixing uniformly, the mixture is treated at a ratio of 10 ° C to 10 ° C. It will be treated. It will be cold;盥^ The door is ground into a powder to obtain a powder containing the composition of the y knife. The wheel can be replaced with other cereals as needed. For example: rice type 'milk, 110560 15 200920389 violent class, beans , sesame, ramie, and mixtures thereof. « 4. Effect of the composition of the present invention on activation of immune cell activity Animal mode: selection Five-week-old male Wi star rats (source: National Taiwan Animal Center) have a weight of 380 to 450 g. The rats were individually housed before the experiment, and the room temperature was controlled at 21 ± 2. (:, humidity 65 ± 5%) The light-dark cycle time was 12D: 12L, and the experiment was started two weeks after the adaptation. f-·. The rats of the present invention were fed the rats for three days, 0.2 ml per day. Three days later, LIVE/DEAD Cell-Mediated was used. Cytotoxicity kits (Molecular Probes) were used to determine the activity of natural killer cells in the blood. The experimental procedure was as follows: The concentration of YAC-1 cells was adjusted to 6 cells/ml. DIOCu (10//LDIOCu/ml YAC-1 cell suspension was added in proportion. Liquid) and mix well. Incubate for 20 minutes at 37 ° C. Adjust the cell concentration to lxl06cell / ml.
V 以RPMI 1 640培養基（Hyc 1 one )將淋巴球細胞調成 細胞濃度2xl06cell/毫升。將染色後的YAc—丨細胞2〇微 升加入的脾臟淋巴球懸浮液2〇〇微升中，並加入220微升 的PI溶液（40微克/毫升）染色，以1 〇〇〇Xg離心3〇秒 後’再於37°C培養2小時。以流式細胞儀測量淋巴球活 性。 結果如第2圖所顯示，與對照組相比較，本發明組成 物確實可提升自然殺手細胞之活性。 16 110560 200920389 •本4明的組成物除可活化免疫細胞活性外，更且有以 .下所述之優點與功效： … 本發明之組成物復具有其他保健功效，如抑制有宏細 菌生長、補充營養、吸附有毒物質、改善膚質等，故;增 進動物體或人體之健康、減少疾病損害。 …本發明之組成物所包含的益菌助生質包含有機酸（如 如 子，可抑制對動物體或人體有害細 菌之生長。 σ、 f - 1 本發明之組成物可提供幽含>* U、八+ ,、 促1A丑田言養成分，主要為菇蕈及 显_在培養過程中戶斤& tl·、 / ! » 柱甲所生成’例如：菇蕈多醣體、蛋白質、 多肽類、三帖類、多藉脫装缺 、 種胺基齩、生物鹼、酯類、有機酸、 維生素B群及酵素等，可、、天 ”，了从、加至食品或飼料，供動物體或 人肢所萬之營養，做為添加用。 另外，本發明之組成物可中和與吸附毒素，故低 人體對硝胺及苯酚類的叨你；政& t + - 喊自0及收而降低致癌風險。本發明之組 ί成物亦可降低膽固醇吸收率、％几。。德、 4又早強化早雙醣之應用並降低j： 體内吸收率’可應用於糖 …、 丁思尿病患者之飲食控管與營養補 充。 本毛明之組成物可溶於水，取其水溶液塗抹於皮膚， I迅速被皮膚吸收，並活化皮膚組織中的免疫細胞（如吞 °巫細胞及製造膠原蛋白的纖維母細胞），可改善膚質並預 防皮膚病。 上述實施例僅例示性說明本發明之組成物與製備方 法，而非用於限制本發明。任何熟習此項技藝之人士均可 Π0560 17 200920389 夸不違背本發明之精神及範疇下，對上述實施例進行修飾 •與改變。因此，本發明之權利保護範圍如後述申請專利範 圍所載。 【圖式簡單說明】 弟1圖為本發明之料與益菌組成物製備流程。 第2圖係長條圖，其係根據本發明之實施例，顯示本 、、舌勺始早與证菌組成物對於大白鼠體内自然殺手細胞 菌響；其中一為對照組4與B為饒食益生 围組、C與D為饒食本發明組成物組。 18 110560V Lymphocytes were adjusted to a cell concentration of 2 x 106 cells/ml in RPMI 1 640 medium (Hyc 1 one ). The stained YAc-丨 cells were added to the spleen lymphocyte suspension in 2 μl of microliters, and stained with 220 μl of PI solution (40 μg/ml) and centrifuged at 1 〇〇〇Xg. After leap seconds, it was incubated at 37 ° C for 2 hours. Lymphocyte activity was measured by flow cytometry. As a result, as shown in Fig. 2, the composition of the present invention did enhance the activity of natural killer cells as compared with the control group. 16 110560 200920389 • In addition to the activation of immune cell activity, the composition of the present invention has the advantages and effects described below: The composition of the present invention has other health effects, such as inhibiting the growth of macrobacteria, Supplementing nutrition, adsorbing toxic substances, improving skin quality, etc., so as to improve the health of animals or human bodies and reduce disease damage. The probiotic-promoting substance contained in the composition of the present invention contains an organic acid (for example, can inhibit the growth of harmful bacteria to the animal or the human body. σ, f - 1 The composition of the present invention can provide a cryptic >* U, eight +, , and promote the 1A ugly field, mainly for mushroom sputum and _ in the process of cultivation, jin·amp; tl·, / ! » column agglutination 'for example: mushroom 蕈 polysaccharide, protein, peptide , three posts, more borrowing and deficiencies, alkaloid hydrazine, alkaloids, esters, organic acids, vitamin B groups and enzymes, etc., can,, and, from, to food or feed, for animal body Or the nutrients of the human body, for added. In addition, the composition of the present invention can neutralize and adsorb toxins, so the body is low on the nitramine and phenols; political & t + - shouted from 0 and In order to reduce the risk of carcinogenesis, the group of the present invention can also reduce the rate of cholesterol absorption, %. De, 4 and early use of early disaccharide application and reduce j: in vivo absorption rate can be applied to sugar ..., Dietary control and nutritional supplements for patients with Dingsi's disease. The composition of this hair is soluble in water. The aqueous solution is applied to the skin, I is quickly absorbed by the skin, and activates immune cells in the skin tissue (such as swallowing witch cells and collagen-producing fibroblasts) to improve skin texture and prevent skin diseases. The above examples are merely illustrative. The present invention is not intended to limit the invention, and any person skilled in the art can modify the above embodiments without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention is as set forth in the following patent application. [Simple Description of the Drawings] Figure 1 is a flow chart for preparing the composition of the present invention and the beneficial bacteria composition. Fig. 2 is a bar graph based on In the embodiment of the present invention, the natural killer cell bacteria of the body, the early start of the tongue and the syndrome composition for the body of the rat are displayed; one of them is the control group 4 and the B is the food and the probiotic group, and the C and D are the Rao The composition of the invention is set. 18 110560