200815601 九、發明說明·· 【發明所屬之技術領域】 本發明係關於一種利用微生物培養所產生之可塑性高分子在 固型模下製作之生物膜及其方法與固型模，尤指一種生物膜之方 法、生物膜以及固型模具。 【先前技術】200815601 IX. INSTRUCTION DESCRIPTION OF THE INVENTION The present invention relates to a biofilm produced by using a plastic polymer produced by microbial culture in a solid mold, a method thereof and a solid mold, especially a biofilm. Methods, biofilms, and solid molds. [Prior Art]
基於人體肌雜養中之各種需求，例如深層清潔、刺激、收 緊、緩和、滋養、爽膚、濕潤等。市售之面膜產品種類繁多。其 中主要係以「不織布」為基材’而添加各類保養品及藥妝品成分 而此種*織布」為基材之面膜，由於該「不織布」不是 用線編織成的布狀物’是—種球(婦）或合錢_料且未經 織布過程而製成的纺織品，透氣性、吸難、耐久性、耐藥品性 、絕緣性料性質。而由於透氣，因此導致其氣密性不佳，且因 所添加的各種滩雙向絲在外造餅解轉發過快，以 至於其功效未能全然發揮。 等，有各種型態之「生物面膜」、「生物纖維面膜_ 皮膚㈣且具有「類離子導人」的魏，因此可以加5 去。且生物面理的保養品及藥妝品成分深入到真皮層， 問題。、可鱗衫的營養物質，幾料會5丨起皮膚獅 然而’目前一般所謂+「，w 天然植物纖維）為載!/，、 面膜」’主要係採用纖維成份 _以纖維大小約50奈米，並塗佈精華液 200815601 即藉由宣稱之天然草本成份配合奈米科技所製成。 如我國專利申請第094210129號「複合式保養膜組合包」之 專利案，其雖然揭露一種涉及生物性保養膜之技術，即「複合式 保養膜組合包」專利案，#内容主要包括一保養膜，具有效成分 ，該肴效成分係以冷凍乾燥之形式附著於該保養膜上；以及一濃 縮精華液。其有效成分係選自膠原蛋白、維他命c、生長因子等其 它易氧化變性之美容保養物質及其組合之群組。以及該濃縮精華 # 係選自玻尿酸、甘草精、金縷梅萃取液、其它溶於水中且不易氧 化變性之美各保養物質及其組合之群組。又，該保養膜之材質包 含不織布、棉布、軟凍材質、紙張或生物科技纖維。 然而，此種藉由有效成分以冷凍乾燥之形式附著於該保養膜 上，而所謂之「生物性」保養膜，大多只是將載體以生物性之原 料製成，配合所謂有效成分之附著，對於使用者之吸收度仍然限 於所谓有效成分，並未能對於該面膜之生物性精華有全面性之吸 • ㈣果： 且前述各種面膜（包括生物面膜），成品貼覆與吸收的過程 ，一般為了避免使用者被敷之部位處於完全遮蔽狀態以及燠熱感 等不適，且避免臉部之眼、鼻、口等器官皆被遮蔽而無法視、嗅 以及飲食、說話。因此基於人體需求，不論為傳統面膜或生物面 膜、生物纖維面膜等皆預設有預留孔，以滿足使用者前述之透孔 需求。而’該預留孔或透孔之製作，必須藉由刀膜進行开克成裁切 。此種必須以一次加工之刀模裁製該預留孔，不但施工繁瑣且耗 費成本。 6 200815601 【發明内容】 本發明主要之目的在於設計一種利用微生物培養所產生之可 塑性高分子在固型模下製作之生物膜及其方法與固型模，主要之 目的在於利用於微生物種菌於已滅菌洋菜固體培養基上著床培養 ，而達成一種令面膜成為含豐富生物菌種之生物膜，而令其對於 人體之吸收度可以較佳，該方法主要包括以下步驟： 1、 由以下選取可產生高分子之微生物種菌： (A)、Gluconacetobacter xylinus subsp· Xylinus ATCC 10821 或 ATCC700178，Gluconacetobacter hansenii ATCC 23769，以及由自然界所分離到屬於醋酸菌類 Acetobacter spp等可產生高分子多糖體類之微生物。 (B )、Xathomonas Campeastris，以及由自然界類似 Xathomonas spp等可產生高分子多糖體類之微生物。 (C )、Bacillus subtil is var Natto，以及由自然界類 似Bacillus spp枯草桿菌等可產生聚麩胺酸鹽 (r -Polyglutamate)高分子之微生物。 (D)、利用遺傳工程技術將動物蛋白膠(如：絲蛋白、彈力 蛋白、膠原蛋白等）、植物蛋白膠(如：大豆蛋白、植 物性膠原蛋白）等蛋白基因，轉植到細菌類或酵母菌類 後’而具備了生產該生物蛋白之基因轉殖菌種。 2、 製備已滅菌洋菜固體培養基，將以上之種菌斜面培養於試管 中0 3、 種菌繁殖；該種菌之繁殖藉由各種營養成分以令其繁殖。 7 200815601 4、生物膜的生產；可選自： (1) 於液態培養液中靜置，培養生產多糖體高分子生物膜; (2) 攪拌槽或發酵槽培養生產多糖體高分子生物膜或 (3) 攪拌槽或發酵槽培養生產蛋白質類高分子膜。 本發明次一目的在於發明一種利用微生物培養所產生之可塑 性高分子在固型模下製作之生物膜，該生物膜之成分主要包括已 滅菌洋菜固體培養基，並以可產生高分子之微生物種菌繁殖而形 • 成之生物膜。藉由此種生物膜，可以將基體包括成分均以生物培 養製成’而非僅為成分簡易附著於載體上，可以全面提升面膜之 生物性精華之吸收效果。· 本發明其他目的一在於設計一種生物膜之固型模，該固型模 適合於生物性之培養皿或生物纖維之培養皿。主要係為一皿體， 該皿體之底板内侧至少設有一生物膜預設孔之肋體，藉由該肋體 所占底板之局部面積，令培養形成之生物膜生成時，可以產生生 _ 物膜對應之預留孔，並藉由該預留孔，免除生物膜製成時必須以 一次加工之刀模裁製該預留孔之施工繁瑣以及節省製造成本。 本發明其他目的二在於實施前述之生物膜之固型模，其中該 肋體了為任思方向之直條狀，藉以令生物膜生成時可以產生對 應該直條狀之預留孔，以提供肌膚有一接觸外界之透氣性。 本發明其他目的三在於實施前述之生物膜之固型模，其中該 肋體可以選擇或另外複合—對應人體贿框圍之形狀，且該形狀 开>/成缺口 ’生物膜可以自框圍内之部分由該對應該缺口所形成 之連接η卩翻折’對應人體目哺之形狀，藉以令生物膜生成 8 200815601 時可以產生對應該肋體之預留孔，以提供目艮睛有一可視穿之視孔。 本發明其他目的四在於實施前述之生物膜之固型模，其中該 肋體可以選擇或糾複合—對應人體鼻部_之形狀，且該形^ 形成-缺口，生物膜可以自框圍内之部分由該對應該缺口所形成 之連接部翻折，_成對應人體鼻部之形狀，藉以令生物膜生成 時了以產生對應該肋體之預留孔’以提供鼻部有一可哞吸之透孔。 本發明其他目的五在於實施前述之生物膜之固型模，其中該 φ 肋體可以選擇或另外複合一對應人體嘴巴框圍，而概呈横橢之形 狀，藉以令生物膜生成時可以產生對應該肋體之預留孔，以提供 嘴巴有一說話或飲食之透孔。 此外’該肋體係由底部之外侧面向内侧凹陷，令内侧面形成 相對浮凸，可以節省材質。 【實施方式】 以下說明本發明之内容、特點以及實施例，俾使貴審查人 ® 員對於本發明有更進一步之了解。 本發明是利用微生物培養所產生之可塑性高分子在固型的模 型下製作一種生物膜之平台技術。該平台技術包含以下幾個步驟 首先係利用微生物培養產生高分子，而可產生高分子之微生 物很多類，包括： A - Gluconacetobacter xylinus subsp. Xylinus ATCC 10821 或 ATCC700178，Gluconacetobacter hansenii ATCC 23769， 9 200815601 以及由自然界所分離到屬於醋酸菌類Acet〇bacter spp等可 產生高分子多糖體類之微生物。 B、 Xathomonas Campeastris，以及由自然界類似 xathomonas spp等可產生高分子多糖體類之微生物。 C、 BacillussubtilisvarNatto，以及由自然界類似 Bacillus spp枯草桿菌等可產生聚麩胺酸鹽（7 -P〇lyglutaraate)高分 子之微生物。 φ D、利用这傳工程技術將動物蛋白膠(如：絲蛋白、彈力蛋白、膠 原蛋白等）、植物蛋白膠(如：大豆蛋白、植物性膠原蛋白）等 蛋白基因，轉植到細菌類或酵母菌類後，而具備了生產該生 物蛋白之基因轉殖菌種。 其次有關菌種的培養及保存 如配方一、二、三、四配方所製備之已滅菌洋菜固體培養基， 將以上之種_斜面培養於試管中，Gluconacetobacter spp及 _ Xathomonas spp於30°C生長2〜7天後冷藏保存於8°C中。納豆菌 類及基因轉植類之大腸桿菌則培養於3rt生長丨〜3天後冷藏保存 於8°C中。基因轉植類之酵母菌類則培養於3〇ΐ生長2〜7天後冷 藏保存於8 C中。在長期之菌種保存則以冷象乾燥保存法進行保存 之。 再則關於種菌的繁殖： 適量將斜面培養之種菌卜5個白金耳量接種至將已滅菌之液 體培養基配方（如下配方五、六、七、八之培養基）中， Gluconacetobacter spp培養於配方五之液體培養基配方及 200815601Based on various needs in human muscle nourishment, such as deep cleansing, irritation, tightening, easing, nourishing, toning, moistening, etc. There are a wide variety of commercially available mask products. Among them, the "non-woven fabric" is used as the base material for the "non-woven fabric" as the base material, and the "non-woven fabric" is not the fabric woven by the thread. It is a kind of textile made from a ball (women) or a combination of money and without a weaving process, which is breathable, sucking, durable, chemical resistant, and insulating. However, due to the venting, the airtightness is not good, and the various types of beach skein added are too fast to be transferred, so that the efficacy is not fully exerted. In addition, there are various types of "bio-mask", "bio-fiber mask _ skin (four) and "ion-like lead" Wei, so you can add 5 to. And the bio-care products and cosmeceutical ingredients go deep into the dermis, the problem. The nutrients of the scalloped shirts will be picked up by the skin lions. However, the current general so-called + ", w natural plant fiber" is contained! /,, the mask "mainly uses fiber components _ with a fiber size of about 50 奈Rice, and coated serum 200815601 is made by claiming the natural herbal ingredients in combination with nanotechnology. For example, the patent application No. 094210129 of the "Plastic Maintenance Film Combination Package" discloses a technology related to a biological maintenance film, that is, a "composite maintenance film combination package" patent case, and the content mainly includes a maintenance film. An effective ingredient, the medicinal ingredient is attached to the maintenance film in a freeze-dried form; and a concentrated essence. The active ingredient is selected from the group consisting of collagen, vitamin C, growth factors and other oxidatively degradable cosmetic care substances and combinations thereof. And the concentrated essence # is selected from the group consisting of hyaluronic acid, liquorice extract, witch hazel extract, other water-soluble and non-oxidative denatured beauty maintenance materials and combinations thereof. Moreover, the material of the care film comprises non-woven fabric, cotton cloth, soft frozen material, paper or biotechnology fiber. However, such an active ingredient is attached to the care film in a freeze-dried form, and the so-called "biological" maintenance film is usually made of a biological material and is blended with a so-called active ingredient. The absorption of the user is still limited to the so-called active ingredients, and has not been comprehensively absorbed for the biological essence of the mask. (4) Fruit: and the above various masks (including bio-masks), the process of product attachment and absorption, generally for Avoid discomfort such as the user's being covered area and the feeling of heat, and avoid the eyes, nose, mouth and other organs of the face being covered and unable to see, smell, eat, and talk. Therefore, based on the needs of the human body, conventional masks or biofilms, bio-fiber masks, and the like are pre-set with reserved holes to meet the user's aforementioned through-hole requirements. The production of the reserved hole or the through hole must be cut into a cut by a knife film. This type of hole must be cut in a single-machined die, which is cumbersome and costly to construct. 6 200815601 SUMMARY OF THE INVENTION The main object of the present invention is to design a biofilm produced by using a plastic polymer produced by microbial culture in a solid mold, a method thereof and a solid mold, the main purpose of which is to utilize microbial inoculum. The sterilized agarwood solid medium is cultured on the bed, and a mask is formed into a biofilm rich in biological species, so that the absorption to the human body can be better. The method mainly comprises the following steps: 1. Select from the following: A microbial inoculant producing a polymer: (A), Gluconacetobacter xylinus subsp. Xylinus ATCC 10821 or ATCC700178, Gluconacetobacter hansenii ATCC 23769, and microorganisms which are derived from the natural world and which are capable of producing a polymeric polysaccharide such as Acetobacter spp. (B), Xathomonas Campeastris, and microorganisms that produce high molecular weight polysaccharides such as Xathomonas spp in nature. (C), Bacillus subtil is var Natto, and a microorganism capable of producing a polymer of poly-glutamate (r-Polyglutamate), such as Bacillus sp. (D) using genetic engineering techniques to transfer protein proteins such as animal protein glue (eg silk protein, elastin, collagen, etc.), vegetable protein glue (eg, soy protein, plant collagen) to bacteria or After the yeast, it has the gene transfer strain that produces the biological protein. 2. Preparing a solid medium for sterilized agar, incubating the above-mentioned seed slant in a test tube, and breeding the inoculum; the breeding of the bacterium is propagated by various nutrients. 7 200815601 4. Production of biofilm; can be selected from: (1) standing in liquid culture solution to culture and produce polysaccharide polymer biofilm; (2) cultivating polysaccharide polymer biofilm or stirring tank or fermentation tank (3) A protein-based polymer membrane is produced by stirring in a stirred tank or a fermentation tank. The second object of the present invention is to invent a biofilm produced by using a plastic polymer produced by microbial culture in a solid mold, the biofilm component mainly comprising a solid medium for sterilized agar and a microbial inoculum capable of producing a polymer. Reproduction and shape • Biofilm. With this biofilm, the substrate and the components can be made by biological cultivation, rather than simply attaching the component to the carrier, which can comprehensively enhance the absorption effect of the biological extract of the mask. Another object of the present invention is to design a solid mold for a biofilm which is suitable for a biological petri dish or a petri dish for biofibers. The main body is a dish body, and the inner side of the bottom plate of the dish body is provided with at least one rib of a biofilm preset hole, and the local area of the bottom plate occupied by the rib body can generate a biofilm when the cultured biofilm is formed. The reserved hole corresponding to the object film, and by the reserved hole, the construction of the reserved hole which is necessary to cut the reserved hole in the one-time processing of the biofilm is eliminated, and the manufacturing cost is saved. Another object of the present invention is to implement the above-mentioned solid film mold of the biofilm, wherein the rib is in the shape of a straight strip of any direction, so that the biofilm can be formed with a reserved hole corresponding to the straight strip to provide The skin has a breathability that is in contact with the outside world. Another object of the present invention is to implement the above-mentioned solid film mold of the biofilm, wherein the rib body can be selected or otherwise composited - corresponding to the shape of the human bribe frame, and the shape is opened and the biofilm can be self-framed. The inner part is formed by the connection η卩 formed by the corresponding gap, which corresponds to the shape of the human body, so that the biofilm formation 8 200815601 can produce a reserved hole corresponding to the rib to provide a visual Wear the sight hole. Another object of the present invention is to implement the above-mentioned solid film mold of the biofilm, wherein the rib can be selected or entangled-corresponding to the shape of the human nose _, and the shape forms a gap, and the biofilm can be self-framed. Partially folded by the connecting portion formed by the corresponding notch, _ is formed into a shape corresponding to the nose of the human body, so that the biofilm is formed to generate a reserved hole corresponding to the rib to provide a snuffing of the nose. Through hole. Another object of the present invention is to implement the above-mentioned solid film mold of the biofilm, wherein the φ rib body can be selected or otherwise combined with a corresponding body mouth frame, and has a shape of a horizontal ellipse, so that the biofilm can be generated when it is generated. The holes should be reserved for the ribs to provide a through hole for the mouth or diet. In addition, the rib system is recessed from the outer side of the bottom side to the inner side to form a relatively embossed inner side surface, which can save material. [Embodiment] The contents, features, and embodiments of the present invention are described below, so that the examiner has a better understanding of the present invention. The present invention is a platform technology for producing a biofilm by using a plastic polymer produced by microbial culture under a solid mold. The platform technology comprises the following steps: firstly, microbial culture is used to produce a polymer, and a plurality of microorganisms can be produced, including: A-Gluconacetobacter xylinus subsp. Xylinus ATCC 10821 or ATCC700178, Gluconacetobacter hansenii ATCC 23769, 9 200815601 and Naturally, microorganisms which can produce high molecular weight polysaccharides such as Acet〇bacter spp, which are acetic acid bacteria, are isolated. B, Xathomonas Campeastris, and microorganisms that produce high molecular weight polysaccharides such as xathomonas spp in nature. C, Bacillus subtilisvar Natto, and microorganisms such as Bacillus spp Bacillus subtilis which can produce high molecular weight polyglutamate (7-P〇lyglutaraate). φ D, using this transfer engineering technology to transfer animal protein glue (such as: silk protein, elastin, collagen, etc.), vegetable protein glue (such as: soy protein, plant collagen) and other protein genes to bacteria or After the yeast, the gene-transforming strain for producing the biological protein is provided. Secondly, the culture and preservation of the strains are as follows. The solid medium of sterilized agar vegetables prepared in the formulas 1, 2, 3 and 4 is cultured in the test tube, and the Gluconacetobacter spp and _Xathomonas spp are grown at 30 ° C. After 2 to 7 days, it was stored refrigerated at 8 °C. The natto bacteria and the gene-transplanted Escherichia coli were cultured at 3 rt for 3 days and then stored at 8 ° C in a refrigerated state. The yeasts of the gene transfer type were cultured in 3〇ΐ and grown for 2 to 7 days, and then stored in 8 C in a cold storage. Preservation in long-term strains is preserved by cold-image drying. In addition, the breeding of the inoculum: Inoculate 5 kinds of white gold ear of the slant culture to the sterilized liquid medium formula (the following formula 5, 6, 7, and 8 medium), and Gluconacetobacter spp is cultured in formula 5 Liquid medium formula and 200815601
Xathomonas spp培養於配方六之液體培養基中，於3〇它振盪培養 生長1 2天以2〜30倍培養液進行再次放大培養，此放大培養可 以在發酵槽或振盈培養箱進行，生長卜2天後即可為量產規模之 種菌苗。基因轉植類之大腸桿_培養於配方七之紐培養基、 納豆菌類則培養於配方八之液體培養基，3rc生長卜3天後，以 2〜30倍培魏進行再奴放大鱗，放大培射式可以在發酵槽 或振盪培養箱進行，生長天後即可為量產規模之種菌苗。 φ 有關生物膜的生產： 靜置培養生產多糖體高分子生物膜之程序 A、 生產容器之製作 將種菌約1%〜50%量與配方九之培養液相混後置入已經設計之 模型谷器中（如第一圖所示）中，於定溫下培養至生物膜生成 至適當之厚度即完成生產階段。 B、 生物膜之蒸煮及漂洗 . • C、生物膜之力π工 本發明亦可利用挽拌槽或發酵槽培養生產多糖體高分子生物 膜’其程序為： A、 多糖體高分子之生產 將種菌約1%〜50%量與配方九之培養液相混後置入已經設計之 攪拌槽或發酵槽培養中，於定溫下培養至細菌高分子生成至 適當之濃度(0· 1%〜20%)即完成生產階段。 B、 高分子之蒸煮及粹取 C、 固型高分子膜之製備 11 200815601 D、生物膜之加工 本發明亦可利用授拌槽或發酵槽培養生產蛋白質類高分子膜 ，其製作程序為： A、將大腸桿菌（如BL21(DE3)類)培養於發酵槽或振盪培養箱， 37 C生長至〇D6〇〇達〇· 2〜0. 8後，加入IPTG誘導劑，即可以 產生該蛋白。 Β、蛋白質之萃取 • C、固型高分子膜之製備 D、生物膜之加工 以下就各種固體培養基與液體培養基之配方提出說明： 配方一、Gluconacetobacter spp之固體培養基配方 可利用醣類物質作為碳原來源，包括：單醣、雙醣、多醣以 上的醣類，或是具有carb〇xy group的物質，例如·· Giyc〇1的物 質其-人營養條件也須包含氮源，包括：Yeast、pept〇ne ⑩ 、頁豆粉、可溶性澱粉、Gelatin的物質等，以及輔助因子：如氯 化銨、硫化銨及填酸鉀、磷、硫、鎂等礦物質，與成長因子維生 素的添加等，其主要配方包括： (1) 1%〜30% Glucose (2) 0.1%-3% Peptone (3) 0·1%〜3% Yeast extract (4) 0· 05%〜2% Citric acid (5) 0·〇5%〜3% di-Natriumhydrogen-phosphat wasserfrei (6) 1%〜3% Agar 12 200815601 本發明關於配方之實施例更可為： 2% Glucose、1% Peptone、1% Yeast extract、0· 1% Citric acid、 0· 3% di-Natriumhydrogen-phosphat wasserfrei、1% Agar。 配方二、Xathomonas之固體培養基配方 可利用醣類物質作為碳原來源，包括：單醣、雙醣、多醣以 上等各種的醣類，或是具有carboxy group的物質，例如：Glycol 肇 的物質。其次營養條件也須包含氣源，包括：Yeast extract、 Peptone、黃豆粉、可溶性澱粉、Gelatin的物質等，以及辅助因 子：如氯化銨、硫化鏔及磷酸卸、構、硫、鎂等礦物質，與成長 因子維生素的添加等，其主要配方包括： (1) 1%〜30% Glucose (2) 1%〜30% Fructose (3) 0· 1 %〜3% Peptone (4) 0· 1%〜3% Yeast extract (5) 0· 05%〜2% Citric acid (6) 0.5%〜2% Potassium Dihydrogenphosphate (7) 1%〜3% Agar 本發明關於配方之實施例更可為： 4% Glucose、2% Fructose、2% Peptone、2% Yeast extract、0· 2% Citric acid、0·2% % Potassium Dihydrogenphosphate 1% Agar ° 配方三、大腸桿菌（E· coli)之固體培養基配方 13 200815601 通常可選取下列之胰蛋白大豆瓊脂培養基（Trypt〇ne s〇y agar ’ TSA)或培養皿計數培養基（piate courrt agar，pCA)與 最常用的Luria broth來培養細菌。 依下列配方配製培養基，或使用市售商品化的培養基均可。 A、 騰蛋白大旦壤脂培養基 (1) 0.01%-2% Tryptone (2) 〇·〇〇1%〜1% Peptone (3) 0.001%〜1%氯化鈉 (4) 1%〜3% Agar B、 培養皿計數培養基 (1 )0·01%〜2% Glucose (2) 0. 01%-1% Peptone (3) 0· 01%〜2% Yeast extract (4) 1°/〇-3°/〇 Agar C、 Luri a broth 培養基 (1) 1%〜5% Luria broth (2) 1%〜3% Agar 本發明關於配方之實施例更可為： 2% Luria broth、1% Agar 配方四、酵母菌類之固體培養基配方 使用沙氏葡萄糖培養基（Sabouraud Dextrose Agar，SDA) 或馬鈴薯葡萄糖培養基（Potato dextrose agar，PDA)來培養酵 200815601 母菌。 A、 沙氏匍萄糖培養基 (1) 2%〜10% Dextrose (2) 1%〜3% Peptone (3) 1%〜3% Agar B、 馬铃薯葡萄糖培養基 (1) 0· 1%〜5% Glucose (2) 0.01%〜1% Peptone (3) 0.01%〜1% Yeast extract (4) 1%〜3% Agar 本發明關於配方之實施例更可為： 2% Glucose、1% Peptone、1% Yeast extract、1% Agar 配方五、Gluconacetobacter spp之固體培養基配方 可利用醣類物質作為碳原來源，包括··單醣、雙醣、多醣以 上等各醣類，或是具有carboxy group的物質，例如：Glyc〇1的 物質。其次營養條件也須包含氮源，包括：Yeast extract 'Xathomonas spp is cultured in liquid medium of formula 6. It is shaken and cultured for 3 days at 3 〇, and then re-amplified by 2~30 times of culture medium. This amplification culture can be carried out in fermentation tank or vibrating incubator, growth growth 2 After the day, it can be a seedling of the mass production scale. Gene-transplanted large intestine rod _ cultured in the formula seven of the New Medium, natto bacteria are cultured in the liquid medium of the formula eight, 3rc growth b 3 days later, 2 to 30 times Pei Wei to re-slave the scales, zoom in and shoot The type can be carried out in a fermentation tank or a shaking incubator, and the seedlings of the mass production scale can be produced after the growth day. φ Production of biofilm: Procedure for static culture of polysaccharide polymer biofilm A. Production of production container The amount of inoculum is about 1% to 50% mixed with the culture liquid of formula IX and placed in the designed model valley. In the apparatus (as shown in the first figure), the production phase is completed by culturing at a constant temperature until the biofilm is formed to an appropriate thickness. B. Biofilm cooking and rinsing. • C. Biofilm force π work The invention can also be used to produce polysaccharide polymer biofilm by using a mixing tank or a fermentation tank. The procedure is as follows: A. Production of polysaccharide polymer will be The inoculum is mixed with the culture liquid of the formula 9 and placed in the culture tank of the designed formula or the fermentation tank, and cultured at a constant temperature until the bacterial polymer is formed to an appropriate concentration (0.1%~ 20%) completes the production phase. B. Preparation of polymer and extraction of C and solid polymer membranes 11 200815601 D. Processing of biofilms The present invention can also be used to produce protein-based polymer membranes by using a mixing tank or a fermentation tank. The preparation procedure is as follows: A. Escherichia coli (such as BL21 (DE3)) is cultured in a fermentation tank or a shaking incubator, 37 C is grown to 〇D6〇〇达〇·2~0. 8 and the IPTG inducer is added to produce the protein. . Extraction of hydrazine and protein • Preparation of solid polymer membrane D. Processing of biofilm The following formulas describe the formulation of various solid and liquid media: Formulation 1. The solid medium formulation of Gluconacetobacter spp can use carbohydrate as carbon. Original sources, including: monosaccharides, disaccharides, sugars above polysaccharides, or substances with carb〇xy group, such as · · Giyc〇1 substances - human nutritional conditions must also contain nitrogen sources, including: Yeast, Pept〇ne 10, page bean powder, soluble starch, Gelatin substances, and cofactors: such as ammonium chloride, ammonium sulfide and potassium, phosphorus, sulfur, magnesium and other minerals, and the addition of growth factor vitamins, etc. The main formulas include: (1) 1%~30% Glucose (2) 0.1%-3% Peptone (3) 0·1%~3% Yeast extract (4) 0· 05%~2% Citric acid (5) 0·〇5%~3% di-Natriumhydrogen-phosphat wasserfrei (6) 1%~3% Agar 12 200815601 The embodiment of the present invention relating to the formulation may further be: 2% Glucose, 1% Peptone, 1% Yeast extract, 0 · 1% Citric acid, 0·3% di-Natriumhydrogen-phosphat wasserfr Ei, 1% Agar. Formulation 2, Xathomonas Solid Medium Formulations Sugar substances can be used as a carbon source, including: monosaccharides, disaccharides, polysaccharides, various kinds of sugars, or substances having a carboxy group, such as Glycol®. Secondly, the nutrient conditions must also include gas sources, including: Yeast extract, Peptone, soy flour, soluble starch, Gelatin, etc., and cofactors such as ammonium chloride, barium sulfide and phosphate decommissioning, sulphur, magnesium and other minerals. The main formulas include: (1) 1%~30% Glucose (2) 1%~30% Fructose (3) 0· 1 %~3% Peptone (4) 0· 1% ~3% Yeast extract (5) 0·05%~2% Citric acid (6) 0.5%~2% Potassium Dihydrogenphosphate (7) 1%~3% Agar The embodiment of the present invention relating to the formulation may be: 4% Glucose 2% Fructose, 2% Peptone, 2% Yeast extract, 0.2% Citric acid, 0.2%% Potassium Dihydrogenphosphate 1% Agar ° Formula 3, E. coli solid medium formula 13 200815601 Usually available The following trypsin soy agar medium (Trypt〇ne s〇y agar ' TSA) or culture dish counting medium (piate courrt agar, pCA) was used to culture the bacteria with the most commonly used Luria broth. The medium may be prepared according to the following formula, or a commercially available medium may be used. A, Teng protein large dendritic lipid medium (1) 0.01% -2% Tryptone (2) 〇 · 〇〇 1% ~ 1% Peptone (3) 0.001% ~ 1% sodium chloride (4) 1% ~ 3% Agar B, culture dish counting medium (1) 0·01%~2% Glucose (2) 0. 01%-1% Peptone (3) 0· 01%~2% Yeast extract (4) 1°/〇-3 °/〇Agar C, Luri a broth medium (1) 1%~5% Luria broth (2) 1%~3% Agar The embodiment of the invention relating to the formulation may further be: 2% Luria broth, 1% Agar Formula 4 The solid medium formulation of the yeast type was cultured using Sabouraud Dextrose Agar (SDA) or Potato dextrose agar (PDA) to culture the yeast 200815601. A, Sabouraud sugar medium (1) 2% ~ 10% Dextrose (2) 1% ~ 3% Peptone (3) 1% ~ 3% Agar B, potato glucose medium (1) 0 · 1% ~ 5% Glucose (2) 0.01%~1% Peptone (3) 0.01%~1% Yeast extract (4) 1%~3% Agar The embodiment of the present invention relating to the formulation may further be: 2% Glucose, 1% Peptone, 1% Yeast extract, 1% Agar Formula 5, Gluconacetobacter spp solid medium formula can use carbohydrates as a carbon source, including monosaccharides, disaccharides, polysaccharides and other sugars, or substances with carboxy group For example, the substance of Glyc〇1. Secondly, the nutritional conditions must also include nitrogen sources, including: Yeast extract '
Peptone、頁豆粉、可溶性殿粉、Geiatin的物質等，以及辅助因 子·如氯化銨、硫化銨及鱗酸卸、填、硫、鎂等礦物質，與成長 因子維生素的添加等，其主要配方包括： (1) 1%〜30% Glucose (2) 0.1%-3% Peptone (3) 0.1%-3°/〇 Yeast extract 15 200815601 (4) 0.05%〜2% Citric acid (5) 0.05%^3% di-Natriumhydrogen-phosphat wasserfrei (6) 1%〜3% Agar 本發明關於配方之實施例更可為： 2% Glucose、1% Peptone、1% Yeast extract、0· 1% Citric acid、 0·3% di-Natriumhydrogen-phosphat wasserfrei、1% Agar o 配方六、Xathomonas之固體培養基配方 可利用醣類物質作為碳原來源，包括：單醣、雙醣、多醣以 上等各醣類，或是具有carboxy group的物質，例如：Glycol的 物質。其次營養條件也須包含氮源，包括：Yeast extract、 Peptone、黃豆粉、可溶性澱粉、Gelatin的物質等，以及辅助因 子：如氯化銨、硫化銨及磷酸斜、磷、硫、鎂等礦物質，與成長 因子維生素的添加等，其主要配方包括： (1) 1%〜30% Glucose (2) 1%〜30% Fructose (3) 0· 1%〜3% Peptone (4) 0·1%〜3% Yeast extract (5) 0.05%〜2% Citric acid (6) 0. 5%-2% Potassium Dihydrogenphosphate (Ό 1%〜3% Agar 本發明關於配方之實施例更可為： 4% Glucose、2% Fructose、2% Peptone、2% Yeast extract、0· 2% 200815601Peptone, page bean powder, soluble temple powder, Geiatin substances, and auxiliary factors such as ammonium chloride, ammonium sulfide and sulphate unloading, filling, sulfur, magnesium and other minerals, and growth factor vitamins, etc. Formulations include: (1) 1%~30% Glucose (2) 0.1%-3% Peptone (3) 0.1%-3°/〇Yeast extract 15 200815601 (4) 0.05%~2% Citric acid (5) 0.05% ^3% di-Natriumhydrogen-phosphat wasserfrei (6) 1% to 3% Agar The embodiment of the present invention relating to the formulation may further be: 2% Glucose, 1% Peptone, 1% Yeast extract, 0.1% Citric acid, 0 ·3% di-Natriumhydrogen-phosphat wasserfrei, 1% Agar o Formulation 6. The solid medium formulation of Xathomonas can use carbohydrates as a carbon source, including: monosaccharides, disaccharides, polysaccharides and other sugars, or A substance of a carboxy group, such as a substance of Glycol. Secondly, the nutrient conditions must also include nitrogen sources, including: Yeast extract, Peptone, soy flour, soluble starch, Gelatin, etc., and cofactors such as ammonium chloride, ammonium sulfide and phosphate, phosphorus, sulfur, magnesium and other minerals. The main formulas include: (1) 1%~30% Glucose (2) 1%~30% Fructose (3) 0·1%~3% Peptone (4) 0·1% ~3% Yeast extract (5) 0.05%~2% Citric acid (6) 0. 5%-2% Potassium Dihydrogenphosphate (Ό 1%~3% Agar The embodiment of the invention for the formulation may be: 4% Glucose, 2% Fructose, 2% Peptone, 2% Yeast extract, 0·2% 200815601
Citric acid、0· 2% Potassium Dihydrogenphosphate 1% Agar 〇 配方七、大腸桿菌（E_ coli)之固體培養基配方 通苇可選取下列之騰蛋白大旦瓊脂培養基（Tryptone soy agar ’ TSA)或培養皿計數培養基（piate count agar，PCA)與 最常用的Luria broth來培養細菌。 依下列配方配製培養基，或使用市售商品化的培養基均可。 _ A、胰蛋白大豆瓊脂培養基 (1) 0. 01%-2% Tryptone (2) 0.001%〜1% Peptone (3) 0.001%〜1%氯化鈉 (4) l%-3% Agar B、 培養皿計數培養基 (1) 0· 01%〜2% Glucose • (2) 0· 01%〜1% Peptone (3) 0.01%〜2% Yeast extract (4) 1%〜3% Agar C、 Luria broth 培養基 (1) 1%〜5% Luria broth (2) 1%〜30/〇 Agar 本發明關於配方之實施例更可為： 2% Luria broth、1% Agar 200815601 • 配方八、酵母菌類之固體培養基配方 使用沙氏葡萄糖培養基（Sabouraud Dextrose Agar ’ SI)A) 或馬鈐薯葡萄糖培養基（potato dextrose a§ar ’ PDA)來培養酵 母菌。 A、 沙氏葡萄糖培養基 (1) 2%〜10% Dextrose (2) 1%〜3% Peptone φ (3) 1%〜3% Agar B、 馬鈴薯葡萄糖培養基 (1) 0.1%-5% Glucose (2) 0· 01%〜1% Peptone (3) 0.01%〜1% Yeast extract (4) 1%〜3% Agar 本發明關於配方之實施例更可為： φ 2% Glucose > 1% Peptone > 1% Yeast extract ^ 1% Agar 配方九、生產生物膜之培養液配方之培養液配方 實驗研究上常用到-些複雜的培養基，其主要成分則由複杂 的養分（包括植物或動物）戶斤提供的一些不詳萃取物。但培養細 菌(yeast)其培養基必須提供下列幾種基本的營養， 同作為能源⑵水分（3)氮源⑷鱗（5)硫⑻； 源（=Γ 鐵和鎂。切翻和酵賴能在只有一翻 源（如糖類物質：赚）與簡單的氮源（如氣化銨、硫纖 200815601 磷、硫等礦物質）的簡單培養基中生長，酵母菌則還需要幾種成 長因子維生素。液態培養基主要配方包括·· (1) 1%〜30% Glucose (2) 0· 1%〜3% Peptone (3) 0.1%-3% Yeast extract (4) 0· 05%〜3% Calcium Carbonate (5) 0. 05%-3% Magnesium Sulfate Heptahydrate φ 本發明關於配方之實施例更可為： 2% Glucose、1% Peptone、1% Yeast extract、0· 5% Calcium Carbonate、0· 1% Magnesium Sulfate Heptahydrate 從而可以製作出一種利用微生物培養所產生之可塑性高分子 在固型模下製作之生物膜，該生物膜之成分主要包括已滅菌洋菜 固體培養基，並以可產生高分子之微生物種菌繁殖而形成之生物 膜。藉由此種生物膜，可以將基體包括成分均以生物培養製成， φ 而非僅為成分簡易附著於載體上，可以全面提升面膜之生物性精 華之吸收效果。 請參閱第二圖所示，有關於本發明中對於生物膜固型用之固 型模，該固型模適合以生物方法對於培養物（2 〇)進行培養， 其概可以而形成生物膜，甚至亦可使用於以不織布為載體之傳統 面膜生成之用。又，本發明所製成之「生物膜」，不僅為人體「臉 部表面」使用，亦可作為眼膜、胸膜、唇膜。 請參閱第一圖所示，主要係為一皿體（工），該皿體（丄）之 底板（10)内側至少設有一面膜預設孔之肋體（丄i )，藉由該 19 200815601 肋體（11)所占底板之局部面積，令培養形成之生物膜（2) 生成時，可以產生如第三圖所示之生物膜（2 )對應之預留孔（2 1)。並藉由該預留孔（2 1)，免除生物膜（2 )製成時必須以 二次加工之刀模裁製該預留孔（21)之施工繁瑣以及節省製造 成本。 至於該肋體之實施例，該肋體（i丄A)可為一任意方向之 直條狀，而當如第三圖所示之生物膜（2)生成時，可以產生對 ⑩應該直條狀肋體（1 1A)之預留孔（2 1A)，以提供肌膚有一 接觸外界之透氣性。 ❿該肋體（1 1B)亦可以選擇或另外複合一對應人體眼睛 框圍之形狀’且該形狀形成―缺口（i 2 B )，而#如第三圖所示 之生物膜（2 )生成時，可以自框圍内之部分由該對應該缺口（工 2 B)所形成之連接部（2 2 B )贿，而形成對應人體眼睛之 形狀，藉以令生物膜⑵生成時可以產生對應該肋體（工工b ) _ 之預留孔（2 1 B )，以提供眼睛有-可視穿之視孔。 該麵（1 1 C)亦可以選擇或另外複合一對應人體鼻部框 圍之形狀，且該形狀形成一缺口（工2 C ) ’而當如第三圖所示之 生物膜（2)生成時，生物膜（2)可以自框圍内之部分由該對 應該缺口（ ；L 2 C )所形成之連接部（2 2 c )翻折，而形成對 ^體鼻部之形狀’藉以令生物膜⑵生成時可以產生對應該 -(1 1 C)之預留孔（2 ! c)，以提供鼻部有—可呼吸之透 孔。 該肋體（11D)亦可以選擇或另外複合一對應人體嘴巴框 20 200815601 圍，而概呈橫橢之形狀，而當如第三圖所示之生物膜（2)生成 k ’產生對應該肋體（1 1 D)之預留孔（2 1 d )，以提供嘴巴 有一說話或飲食之透孔。 此外，前述之各個肋體（i i A)〜α i D)均可以由底 部之外侧面向内侧凹陷，令内侧面形成相對浮凸，可以節省材質 〇 又，本發明之固型模材質可以由Pvc (聚氯乙烯）、pp (聚丙 φ 稀）或PE (聚乙烯）等材質所製成，或者由抗氧化之金屬製成， 或者，本發明亦可以於材質之表面塗佈(COATING)PEP (防銹膜） 該PEP (p猶膜）為尚分子防錄钱膜，可以包覆阻隔方式防止腐 飿生氣體牙透，在腐姓性氣體穿透防錄膜時將中和，以及將被包 覆在内潛藏的顧性氣體反應將其巾和，使包覆物_空氣不再 具有腐紐，它不會在被保護的物體上殘留有任何化學物質。並 且，、有抑制細菌’防止黴菌細菌等對物質所造成的損害。對於生 • 物膜之培養，可以有效減少其他細菌之干擾。 綜上所述’本發明確實符合產業利用性，且未於申請前見於 刊物或公開使用，縣為絲所知悉，且具有非顯而易知性，符 合可專利之要件，爰依法提出專利申請。 惟上述所陳’為本創作產業上一較佳實施例，舉凡依本創作 申睛專利範圍所作之均等變化，皆屬本案訴求獅之範蜂。 21 200815601 【圖式簡單說明】 第一圖係本發明之立體圖 第二圖係本發明培養狀態之立體圖 第三圖係本發明之生物膜之平面圖 第四圖係本發明之生物膜成孔狀態之平面圖 【主要元件符號說明】 (1):皿體Citric acid, 0.2% Potassium Dihydrogenphosphate 1% Agar 〇 Formula VII, E. coli solid medium formula The following selection of Tryptone soy agar ' TSA or culture dish counting medium can be selected overnight. (piate count agar, PCA) and the most commonly used Luria broth to culture bacteria. The medium may be prepared according to the following formula, or a commercially available medium may be used. _ A, tryptic soy agar medium (1) 0. 01% - 2% Tryptone (2) 0.001% ~ 1% Peptone (3) 0.001% ~ 1% sodium chloride (4) l% -3% Agar B, Petri dish counting medium (1) 0· 01%~2% Glucose • (2) 0· 01%~1% Peptone (3) 0.01%~2% Yeast extract (4) 1%~3% Agar C, Luria broth Medium (1) 1%~5% Luria broth (2) 1%~30/〇Agar The embodiment of the present invention relating to the formulation may further be: 2% Luria broth, 1% Agar 200815601 • Formulation 8, solid medium of yeast The formulation was cultured using Sabouraud Dextrose Agar 'SI A) or potato dextrose a§ar 'PDA. A, Sabouraud glucose medium (1) 2%~10% Dextrose (2) 1%~3% Peptone φ (3) 1%~3% Agar B, potato glucose medium (1) 0.1%-5% Glucose (2 0·01%~1% Peptone (3) 0.01%~1% Yeast extract (4) 1%~3% Agar The embodiment of the present invention relating to the formulation may further be: φ 2% Glucose > 1% Peptone > 1% Yeast extract ^ 1% Agar Formula IX. Formulation of culture solution for the production of biofilm culture solution The experimental study used to use some complex media, the main components of which are complex nutrients (including plants or animals). Some unknown extracts are provided. However, the culture medium (yeast) must provide the following basic nutrients, as energy (2) moisture (3) nitrogen source (4) scale (5) sulfur (8); source (= strontium iron and magnesium. Only one source (such as sugar: earned) and a simple nitrogen source (such as ammonium sulfate, sulfur fiber 200815601 phosphorus, sulfur and other minerals) in simple medium growth, yeast also needs several growth factors vitamins. The main formulation of the medium includes: (1) 1%~30% Glucose (2) 0·1%~3% Peptone (3) 0.1%-3% Yeast extract (4) 0· 05%~3% Calcium Carbonate (5 0. 05%-3% Magnesium Sulfate Heptahydrate φ The embodiment of the present invention relating to the formulation may further be: 2% Glucose, 1% Peptone, 1% Yeast extract, 0.5% Calcium Carbonate, 0.1% Magnesium Sulfate Heptahydrate Therefore, a biofilm produced by using a plastic polymer produced by microbial culture in a solid mold can be produced, and the composition of the biofilm mainly includes a solid medium of sterilized agar and is formed by breeding microorganisms capable of producing a polymer. Biofilm. With this organism Membrane, the substrate and the components can be made by biological culture, and φ, instead of merely simply attaching the component to the carrier, can comprehensively enhance the absorption effect of the biological extract of the mask. Please refer to the second figure, relating to the present invention. In the solid mold for biofilm solidification, the solid mold is suitable for biologically cultivating the culture (2 〇), which can form a biofilm, and can even be used for a conventional mask with a non-woven fabric as a carrier. In addition, the "biofilm" produced by the present invention is not only used for the "face surface" of the human body, but also as an eye mask, a pleura, or a lip membrane. Referring to the first figure, the main one is a body (work), at least one of the inner side of the bottom plate (10) of the dish body (10) is provided with a rib body (丄i) of a predetermined hole of the film, and the partial area of the bottom plate occupied by the rib (11) of the 19 200815601 is When the biofilm (2) formed by the culture is formed, a reserved hole (2 1) corresponding to the biofilm (2) as shown in the third figure can be produced, and the biological hole is exempted by the reserved hole (2 1). When the film (2) is made, it must be cut by a secondary processing knife. The construction of the reserved hole (21) is cumbersome and saves manufacturing costs. As for the embodiment of the rib, the rib (i丄A) can be a straight strip in any direction, and as shown in the third figure. When the biofilm (2) is formed, a reserved hole (2 1A) for the straight rib (1 1A) of 10 should be produced to provide the skin with a breathability to the outside. The rib (1 1B) may also be selected or otherwise combined with a shape corresponding to the frame of the human eye and the shape forms a notch (i 2 B ), and the biofilm (2) generated as shown in the third figure is generated. At the time, the portion of the frame can be bribed by the connection portion (2 2 B) formed by the corresponding gap (Work 2 B) to form a shape corresponding to the human eye, so that the biofilm (2) can be generated when it is generated. The rib (worker b) _ is reserved for the hole (2 1 B) to provide the eye with a visible hole. The surface (1 1 C) may also be selected or otherwise combined with a shape corresponding to the contour of the human nose frame, and the shape forms a gap (Work 2 C ) ' and when the biofilm (2) is generated as shown in the third figure When the biofilm (2) can be folded from the portion of the frame by the corresponding portion (2 2 c ) formed by the corresponding gap ( ; L 2 C ), the shape of the nose portion is formed. When the biofilm (2) is generated, a reserved hole (2! c) corresponding to -(1 1 C) can be generated to provide a breathable through hole in the nose. The rib (11D) may also be selected or otherwise combined with a corresponding human mouth frame 20 200815601, and has a shape of a horizontal ellipse, and when the biofilm (2) as shown in the third figure generates k 'produces a corresponding rib A reserved hole (2 1 d) of the body (1 1 D) to provide a through hole for the mouth or diet. In addition, each of the aforementioned ribs (ii A) 〜α i D) may be recessed from the outer side of the bottom side to the inner side to form a relatively embossed inner side surface, which can save material 〇, and the solid mold material of the present invention can be made of Pvc. (Polyvinyl chloride), pp (polypropylene φ thin) or PE (polyethylene) or other materials, or made of anti-oxidation metal, or the invention can also be coated on the surface of the material (COATING) PEP ( Anti-rust film) The PEP (p jujube) is a molecular anti-money film, which can be covered to prevent the corrosion of gas generated by the rot, and will be neutralized when the rot gas penetrates the anti-recording film, and will be The gas that coats the inner hidden gas reacts with the towel, so that the wrap_air no longer has a ruin, and it does not leave any chemicals on the object to be protected. Moreover, there are inhibitors against bacteria that prevent damage to substances caused by mold bacteria. For the culture of the biofilm, it can effectively reduce the interference of other bacteria. In summary, the present invention is indeed in line with industrial applicability, and has not been found in publications or publicly used before the application, the county is known to the wire, and has non-obvious knowledge, conforms to the patentable requirements, and filed a patent application according to law. However, the above-mentioned Chen's is a preferred embodiment of the creative industry. The average change made by the scope of the patent application of this creation belongs to the actor of the lion. 21 is a perspective view of the present invention. The second diagram is a perspective view of the culture state of the present invention. The third diagram is a plan view of the biofilm of the present invention. The fourth diagram is the state of the biofilm of the present invention. Plan view [main component symbol description] (1): dish
(1 0 )··底板 (1 1 ):肋體 (1 1 A): 肋體 (1 1 B): 肋體 (1 1 C)： 肋體 (1 1 D)： 肋體 (1 2 B)： 缺口 (1 2 C)： 缺口 (2 ): :生物膜 (2 0 ):培養物 (2 1 ):預留孔 (2 1 A): 預留孔 (2 1 B): 預留孔 (2 1 C)： 預留孔 (2 1 D)： 預留孔 (2 2 B): 連接部 22 200815601 (2 2 C ):連接部(1 0 )··Bottom plate (1 1 ): rib (1 1 A): rib (1 1 B): rib (1 1 C): rib (1 1 D): rib (1 2 B ): Notch (1 2 C): Notch (2): : Biofilm (2 0 ): Culture (2 1 ): Reserved hole (2 1 A): Reserved hole (2 1 B): Reserved hole (2 1 C): Reserved hole (2 1 D): Reserved hole (2 2 B): Connection part 22 200815601 (2 2 C ): Connection part