201125636 六、發明說明： 【發明所屬之技術領域】 [議1] 本發明係有關於一種萃取方法及其萃取成份，特別是關 於一種從雙子葉植物胎座中萃取出植物生物素及其植 物性機能成份之方法。 【先前技術·】 [0002] 近年來，由於睡蓮在食用上的開發利用，在台南縣六甲 與白河地區出現食用睡蓮的栽培，且因社區都市高度發 展，亦出現盆花的栽培，主要在田尾與台南市安南區。 目前全省栽培面積以嘉義縣的水上地區居多，近5公頃 ，其次是台南縣的六曱與白河地區。睡蓮在台灣一年四 季皆可不斷開花，以香水睡蓮而言，每年每株的開花量 約在280至340朵之間。 [0003] 然目前對於睡蓮等雙子葉植物的經濟應用，除觀賞外， 係將其花朵乾燥後作為泡茶飲品。高雄醫學大學藥物學 研究所將蓮花茶等睡蓮抽出物進行急性及慢性毒性分析 後，結果證明蓮花茶等睡蓮抽出物無毒性，食用安全性 高。 [0004] 因此，如何自蓮花及睡蓮等雙子葉植物中，抽取出穩定 結構之植物機能性成份，藉以有效提升此等雙子葉植物 農產品之經濟附加價值，符合農業休閒產業利用性，實 為目前業界亟待解決之課題。 【發明内容】 [0005] 為了解決上述課題，本發明提供了 一種從雙子葉植物胎 座中萃取出植物生物素及其植物性機能成份之方法，係 099101624 表單編號A0101 第4頁/共22頁 0992003176-0 201125636 0 [0006] [0007] Ο [0008] 包含以下步驟：⑷以機械方式破碎雙子葉植物胎座， 得出雙子葉植物胎座破碎物；（b)以逆境方式處理由步 驟（a)所得之雙子葉植物胎座破碎物，藉此，以取得雙 子葉植物胎座前萃取物；（c)將步驟（b)所得之雙子 葉植物胎座前萃取物進行生物轉換方式處理，藉此，以 取得植物胎座粗萃取物；（d)將步驟（c)所得之雙子 葉植物胎座粗萃取物施以震動過濾或是離心方式處理後 ，取得渡液，以及（e)將由步驟（d)所得之該遽液進 行一乾燥處理，藉此以取得該植物性機能成份。 因此，本發明之主要目的係提供一種從雙乎葉植物胎座 中萃取出植物生物素及其植物性機能成份之方法，係利 用逆境處理及生物轉換之技術，即可提取出含有多醣及 多酚之植物性機能成份，可有效降低成冬，並處理縮短 時間。 本發明之次要目的係提供一種從雙子葉植参胎座中萃取 出植物生物素及其植物性機能成份之方法，由於避免使 用化學藥劑，故具有高安全性、耳啤減低能源浪費及縮 短加工製程之時效，且取得具有穩定結構之產物。 【實施方式】 由於本發明係揭露一種雙子葉植物胎座中萃取出植物生 物素及其植物性機能成份之方法，其中所利用之萃取基 礎技術及相關萃取原理，已為相關技術領域具有通常知 識者所能明瞭，故以下文中之說明，不再作完整描述。 同時’以下，文中所對照之圖式，係表達與本發明特徵有 關之結構示意，無需依據實際尺寸完整繪製，合先敘明 099101624 表單編號A0101 第5頁/共22頁 0992003176-0 201125636 [〇〇〇9]首先請參考第1圖，係本發明提出之第一較佳實施例從 雙子葉植物胎座中萃取出植物生物素及其植物性機能成 伤之方法步驟流程圖。雙子葉植物胎座中萃取出植物 生物素及其植物性機能成份之方法係包含以下步驟。 [0010]步驟（a): [0011] 將雙子葉植物胎座（如第6A圖及第6B圖所示，第6A圖為 完整雙子葉植物胎座，第6B圖為雙子葉植物胎座經洗淨 剖切後之樣態）置入均質機、超音波震盪機、果汁機、 . ： 截切蔬菜機或食物調理機等，以機械方式進行破碎，藉 此得出雙子葉植物胎座破碎物^雙子葉植物以蓮屬（Ne_ lurabo)或睡蓮屬（Nymphaea)植物為隹， [0012] 又，使用前述的機械方式以破碎雙子葉植物胎座，係將 雙子葉植物胎座置入均質機、超音波震盪機、果汁機、 截切蔬菜機或食物調理機中，施加以瞬間（transien1： )攪動或攪拌2-3秒，甚至可以用手工切碎的方式加以取 代。其目的在於，將雙子葉植物胎座裁切為小塊，並使 裁切後的小塊保有雙子葉植物胎座的細胞；亦即，雙子 葉植物胎座經此機械方式破碎後，並非使其所有細胞完 全破碎。所以經過步驟（a)處理後而得的雙子葉植物胎 座破碎物，除由破碎組織或細胞流出的組織液及細胞液 外’仍可保有部份完整之細胞或組織團塊。 [0013] 步驟（b): [0014] 099101624 將步驟（a)得到的雙子葉植物胎座破碎物進行逆境處理 表單編號A0101 第6頁/共22頁 0992003176-0 201125636 [0015] [0016] Ο [0017] Ο [0018] ，亦即將將步驟（a)得到的雙子葉植物破碎物添加氯化 鈉（最終濃度範圍為重量百分比〇. 〇2%到2〇%)，並於 40 °C至100 °C之溫度範圍進行加熱，並加熱至多9〇 分鐘。藉此，以取得雙子葉植物胎座前萃取物。 步驟（c): 將步驟（b)㈣之雙子葉植物胎座冑萃取物進行生物轉 換方式處理，亦即將將步驟（b)所得之雙子葉植物胎座 前萃取物，於無菌環境中以天然發酵方式培養“至“小 時，誘導其中的植物胎座麵胞進行轉化，藉此，以取得 雙子葉植物胎座粗萃取物，且此時所取得之雙子葉植物 胎座粗萃取物呈現凝膠狀態。 上述以天然發酵方式進行培養係指將雙子考植物胎座前 萃取物，於無菌環境中室溫下直接進行培養，無須額外 加入其他微生物或化學物質，即可進行發酵程序。此外 ’經過機械破碎後取得的雙子禁植物胎座破碎物，仍保 有雙子葉植物胎座細胞或組織團塊，因此，在上述的培 養環境中，並誘使雙子葉植物胎座細胞或組織困塊進行 轉化’使其大量地分泌出包含有植物生物素及其植物性 機能成份的凝膠狀雙子葉植物胎座粗萃取物。 又，如第6C圖所示’其為雙子葉植物胎座直接經過手工 擠壓（未經破碎處理）後之情形。如圖所示，雙子葉植 物胎座經過手工擠壓後流出透明凝膠狀之粗萃取物，其 内即含有植物生物素及其植物性機能成份。然僅經手 工擠壓’並無法自雙子葉植物胎座中大量取得之凝膠狀 099101624 表單編號A0101 第7頁/共22頁 0992003176-0 201125636 之含有植物生物素及其植物性機能成份，故若需大旦 取得含有植物生物素及其植物性機能成份的凝膠狀雙 子葉植物胎座粗萃取物，仍需經由步驟（b)處理。 19] 001 e κ(ν 步 [0020] 將由步驟（d)所得之濾液進行熱風乾燥 '冷風乾燥、嘴 霧乾燥、冷束乾燥或減壓乾燥之乾燥處理，而乾燥後即 可取得如第7B圖所示之無色無味的植物生物素及其植物 性機能成份。 [0021] 前述乾燥後的植物生物素及其植物性機能成份係包含有 植物多醣及多酚（polyphenol)，其中植物多酚如可水 解之單寧（hydrolyzable tannins)及類苯丙醇（ phenylpropanoid，例如黃酹素（Flav0noid)、異黃201125636 VI. Description of the invention: [Technical field to which the invention pertains] [1] The present invention relates to an extraction method and an extract thereof, and more particularly to an extraction of plant biotin from a placenta of a dicotyledon and its phytochemical The method of functional ingredients. [Prior Art·] [0002] In recent years, due to the development and utilization of water lily in edible food, the cultivation of water lily has appeared in the Liujia and Baihe areas of Tainan County, and due to the high development of the community, there is also the cultivation of potted flowers, mainly in Tianwei and Annan District, Tainan City. At present, the cultivated area of the province is mostly in the water area of Chiayi County, nearly 5 hectares, followed by the Liuyi and Baihe areas of Tainan County. Water lilies can be continuously flowered in Taiwan every year and four seasons. In the case of perfume water lily, the flowering amount per plant is between 280 and 340 per year. [0003] At present, for the economic application of dicotyledonous plants such as water lily, in addition to viewing, the flowers are dried and used as a tea drink. The Institute of Pharmacology at Kaohsiung Medical University analyzed the acute and chronic toxicity of lotus root tea and other water lily extracts. The results showed that lotus root tea and other water lily extracts were non-toxic and safe to eat. [0004] Therefore, how to extract plant functional components of stable structure from dicotyledonous plants such as lotus and water lily, thereby effectively increasing the economic added value of these dicotyledonous agricultural products, in line with the utilization of agricultural leisure industry, The problem that the industry needs to solve urgently. SUMMARY OF THE INVENTION [0005] In order to solve the above problems, the present invention provides a method for extracting plant biotin and its phytofunctional components from a dicotyledonous placenta, 099101624 Form No. A0101 Page 4 of 22 0992003176-0 201125636 0 [0006] [0007] [0008] The following steps are included: (4) mechanically breaking the dicotyledonous placenta, resulting in a dicotyledonous placenta fragment; (b) treating the step by adversity (step) a) the resulting dicotyledonous placenta fragment, whereby the dicotyledonous pre-fetal extract is obtained; (c) the dicotyledonous pre-fetal extract obtained in step (b) is subjected to a biotransformation treatment, Thereby, obtaining a crude extract of the plant placenta; (d) subjecting the crude extract of the dicotyledon of the dicotyledon obtained in the step (c) to vibration filtration or centrifugation, obtaining the liquid, and (e) The mash obtained in the step (d) is subjected to a drying treatment to obtain the phytofunctional component. Therefore, the main object of the present invention is to provide a method for extracting plant biotin and its phytofunctional component from a bile leaf plant placenta, which can extract polysaccharides and polysaccharides by using the techniques of stress treatment and biotransformation. The botanical functional ingredient of phenol can effectively reduce wintering and reduce the time. A secondary object of the present invention is to provide a method for extracting plant biotin and its phytofunctional components from a dicotyledonous gynecological placenta, which has high safety, avoids energy waste and shortens by avoiding the use of chemical agents. The aging of the processing process, and the product with a stable structure. [Embodiment] The present invention discloses a method for extracting plant biotin and its phytofunctional components from a dicotyledonous placenta, wherein the extraction basic technology and related extraction principles utilized have common knowledge in the related art. The person can understand, so the description below will not be fully described. At the same time, the following figures are used to express the structure related to the features of the present invention, and need not be completely drawn according to the actual size. The first description is 099101624 Form No. A0101 Page 5 / Total 22 Pages 0992003176-0 201125636 [〇 〇〇9] First, referring to Fig. 1, a flow chart of a method for extracting plant biotin and its vegetative function from a dicotyledonous placenta of the first preferred embodiment of the present invention is shown. The method of extracting plant biotin and its phytofunctional components from the dicotyledonous placenta includes the following steps. [0010] Step (a): [0011] A dicotyledonous placenta (as shown in Figures 6A and 6B, Figure 6A is a complete dicot plant placenta, and Figure 6B is a dicot plant placenta After washing and cutting, put it into a homogenizer, ultrasonic oscillating machine, juice machine, . : cutting vegetable machine or food conditioning machine, etc., mechanically crushing, thereby producing a dicotyledonous placenta fragmentation The dicotyledonous plant is a nematode (Ne_lurabo) or a Nymphaea plant, [0012] Further, using the aforementioned mechanical means to break the dicotyledonous placenta, the dicot plant placenta is placed in homogenization In a machine, ultrasonic oscillating machine, juicer, cutting vegetable machine or food conditioner, apply a momentary (transien1:) agitation or agitation for 2-3 seconds, or even replace it by hand chopping. The purpose is to cut the dicotyledonous placenta into small pieces and to keep the cut pieces of cells of the dicotyledonous placenta; that is, the dicotyledonous placenta is broken by this mechanical means. All of its cells are completely broken. Therefore, the dicotyledonous placental fragment obtained after the treatment of the step (a) can retain a part of the intact cell or tissue mass except for the tissue fluid and the cell fluid flowing out of the broken tissue or cells. [0013] Step (b): [0014] 099101624 The dicotyledonous placenta fragment obtained in step (a) is subjected to stress treatment. Form No. A0101 Page 6 of 22 0992003176-0 201125636 [0016] Ο [0018] 双 [0018], will also be added to the dicotyledonous material crushed product obtained in step (a) sodium chloride (final concentration range of weight% 〇. 〇 2% to 2%), and at 40 ° C to Heat at a temperature of 100 °C and heat for up to 9 minutes. Thereby, the dicotyledonous pre-fetal extract is obtained. Step (c): biologically converting the dicotyledonous placenta scorpion extract of step (b) (iv), and also preparing the dicotyledonous pre-fetal extract obtained in step (b) in a sterile environment. Fermentation method cultures "to" hours, induces the transformation of the plant placenta cells, thereby obtaining a crude extract of the dicotyledon of the dicot, and the crude extract of the dicotyledon of the dicot is presented as a gel at this time. status. The above-mentioned culture by natural fermentation means that the extract of the Gemini test plant placenta is directly cultured in a sterile environment at room temperature, and the fermentation process can be carried out without adding additional microorganisms or chemicals. In addition, the "double-forbidden plant placenta broken pieces obtained after mechanical crushing still retain the dicotyledonous placental cells or tissue mass, and therefore, in the above-mentioned culture environment, and induce the dicotyledonous placental cells or tissues The trapped block is transformed to 'substantially secrete a crude extract of the gelatinous dicotyledonous placenta containing plant biotin and its vegetal functional ingredients. Further, as shown in Fig. 6C, it is a case where the dicotyledonous placenta is directly subjected to manual extrusion (unbroken treatment). As shown in the figure, the dicotyledonous plant placenta is hand-squeezed and then flows out of a transparent gel-like crude extract containing plant biotin and its botanical functional ingredients. However, it has only been manually squeezed 'and can not be obtained from the dicotyledonous placenta in a large amount of gelatinous 099101624 Form No. A0101 Page 7 / Total 22 Page 0992003176-0 201125636 Contains plant biotin and its phytofunctional ingredients, so If it is necessary to obtain a crude extract of the gelatinous dicotyledonous placenta containing plant biotin and its phytofunctional components, it is still necessary to be treated by the step (b). 19] 001 e κ (ν step [0020] The filtrate obtained in the step (d) is subjected to hot air drying, cold air drying, mist drying, cold beam drying or vacuum drying to dry, and after drying, it can be obtained as in 7B. The colorless and odorless plant biotin and its phytofunctional component are shown in the figure. [0021] The above-mentioned dried plant biotin and its phytofunctional component comprise plant polysaccharides and polyphenols, wherein plant polyphenols are Hydrolyzable tannins and phenylpropanoids such as flavonoids (Flav0noid)
_素（isoflavonoid)等成分，具有抗氧化能力及抑制 路胺酸酶（tyrosinase )之效果，可應用於化妝品 、營養品、保健食品品及健it食讓等甩途以下第一至 第四實驗例，係就本發明第一較佳實施例所製得之植物 性機能成份測試其抗氧化能力、和卩善j路胺酸酶生成能 \J 力、總醣含量，以及總多酚含量。 [0022] 第一實驗例： [0023] 將第一較佳實施例中所提供之方法，自不同花色的蓮花 胎座中取得含有植物生物素及其植物性機能成份之提 取液’並利用Arnao等人所提出之方法，測定各花色蓮花 胎座中所取得含有植物性機能成份之提取液的總抗氧化 能力。Arnao等人研究指出抗氧化能力之測定可利用 099101624 表單編號Α0101 第8頁/共22頁 0992003176-0 201125636 ❹ [0024] [0025] Ο [0026] 2, 2’-連氮基-雙- (3 -乙基苯并二氫。塞〇坐琳__6 -續酸 )二銨鹽（ 2, 2’-azino-bis(3-ethylbenzthiazoline-6-sulph onic acid) ’ ABTS)與過氧化酶（preoxidase)進 行氧化反應’藉以形成安定藍綠色之陽離子自由基；而 待測樣本的抗氧化能力則是以其清除經由上述反應所生 成的安定藍綠色之陽離子自由基之多寡為表示。由於此 陽離子自由基在734 nm波長下有強烈吸收波峰。因此利 用公式： 清除率（％)= [1 —(樣品734 nm吸光值/控制組734 nm 吸光值）]X 100% :厂 可得到計算出清除效應百分率。亦即’當吸光值越低， 表示樣品清除ABTS自由基之清除能力越強。此外，本實 驗例中另以不同濃度的trolox抗氧化劑作為清除ABTS + 自由基之陽性對照組’並建立標準曲線。trolox為水溶 性維他命E的類似物，故trolox可作為抗氧化劑，且具有 預防與保護皮膚免受紫外線輻射傷害的作用。 試驗結果如第2圖所示’不同花色蓮花胎座植物生物素之 提取液，對清除ABTS+自由基之清除能力皆接近9〇%，且 經標準曲線換算，0. 36 mM的trolox其抗氧化能力為 95%。因此，經由第一較佳實施例中所提供之方法，自不 同花色的蓮花胎座中取得含有植物生物素及其植物性 機能成份’具有有效清除ABTS+自由基之清除能力及良 好之抗氧化效果。 099101624 表單編號A0101 第9頁/共22頁 0992003176-0 201125636 [0027] 第二實驗例： [0〇28] 本第一實驗例係利用不同，j洛落aa η卞扪用不间化色4化提取液對酪胺酸酶 (trosinase)抑制能力，乃透過多巴色素 (d〇PaChr〇me)來測定，其係為從赂鞍酸（tyr〇sine) 生合成黑色素（melanin)過程中的—個中間產物以評 估對抑制黑色素合成的效果。將第_較佳實施例中所提 供之方法’自不同花色的蓮花胎座中取得含有植物生物 素及其才皇物性機能成份之提取液，並利用|等人於1996 年所提出之方法，測定各花色蓮花胎座中所取得含有 植物生物素及其植物性機能成份之提取液的酪胺酸酶 抑制能力；由於酪胺酸酶在黑色素合成中，扮演著速 率限制酵素角色，故利用分解反應，可藉由測定於475 nm波長下的吸光值來計算酪胺酸酶抑制能力。當吸光 值愈大’表示黑色素的形成被抑制程度愈大。本實驗係 依總吸光度方法進行測定。 .::....:. r 1. ：'. ...;丨：\ [0029] 首先，準備4支樣品瓶分別為At、Ai、Ab及AQ，（ 1 )在4 支樣品分別加入0.9 ml的緩衝溶興”（2)在4支樣品分 別加入1 ml酷胺酸水溶液；（3)在人+及41加入1 ml受 測檢品，（4)在Ab及A〇加入1 ml去離子水’（5)置於 37 °C水浴槽中作用10 min後’ （6)在人丨及八^加入1 ml缓衝溶液’（7)在At及Ab加入0.1 m1’濃度為350 units/ml的酪胺酸酶水溶液’將上述溶液混合均勻。（ 8)於37。(：水浴槽中反應25 min後，（9)利用光譜儀 測定波長475 nm的吸光值。受測檢品是由第一較佳實施 例中所提供之方法自不同花色的蓮花胎座中取得含有植 099101624 表單編號A0101 第10頁/共22頁 0992003176-0 201125636 物生物素及其植物性機能成份之提取液、以同體積之市 售麵酸（k〇jic acid，1 mg/ml)及熊果苷（arbutin，1 mg/ml)溶液，其中市售麴酸及熊果苷溶液係作為酪胺酸 酶抑制劑之指標化合物（陽性對照組）。 _]㈣酸酶抑制率的計算，係利用下列公式計算， [_ 抑制率u)= {[(Ab_A〇HArV] / % χ1〇〇 % [0032] 上述代號說明如下： 圆At :待測樣本總吸光度，代表黑色素合成中加入受測檢 品後多巴色素吸收度 闕A1 :無路胺酸酶空白對照組之吸光度’代表受測檢品的 吸收度 _ Ab :無待測樣本空白對照組織吸光度，代表黑色素生合 成中多巴色素吸收度 〇 圆A〇:總空白對照組，吸光度，代表溶媒本身的吸收光产 [0〇37]測定結果如第3圖所示，其中黃花對路胺酸酶抑 最佳，甚至較陽性對照組熊果苷（1 mg/nll)抑此2 力更佳’其次是粉紅、紫紅及藍花。故經由第〜較制月b 施例中所提供之方法，自不同花色的蓮花胎座中取得 1 實 有植物生物素及其植物性機能成份，具有有欵抑'含 胺酸酶生成之效果β [0038] 第三實驗例： 099101624 表單編號A0101 第11頁/共22頁 0992003176-0 201125636 [0039] [0040] 由第—較佳實施例中所提供之方法，自不同花色的蓮花 胎座中取得含有植物生物素及其植物性機能成份之提 取'液’ 並利用 phenol-sulforic 法（Dubois et al·， 1 956)進行分析。phen〇i_suijforic法係由於五碳糖和 碳糖在酸性、高溫下皆會因脫水分解而形成喃甲醛， 而嗔甲链會進—步和苯酚反應而呈現橘色，因此本第三 實驗例係利用此原理，以測定自不同花色的蓮花胎座中 所取彳卞含有植物生物素及其植物性機能成份之提取液 的總糖含量，並以葡萄糖溶液製作標準曲線（y = 0. 291x 一 ^ 1724 ’ r2 = 〇. 99)，將測定值換算成相對應的濃度， 即可求得樣品溶液之多醣濃度。 本實驗之測定結果如第4圖所示，可發現自不同花色的蓮 化*月台座中所取得含有植物生物素及其植物性機能成份 之提取液的總醣含量’其中自黃花蓮花胎座中所取得含 有植物生物素及其植物性機能成份之提取液的總醣含 量相當於5.6 mg/ml葡萄糖，自藍花蓮花胎座中所取得 含有植物生物素及其植物性機能成份之提取液的總醣 含量相當於3.81 mg/ml葡萄糖，自粉紅花蓮花胎座中所 取侍含有植物生物素及其植物性機能成份之提取液的 總酶含量相當於3.92 mg/ml葡萄糖，自紫紅花蓮花胎座 t所取得Μ _生物素及其植物性滅成份之提取 液的總聽含量相當於4. 35 mg/mi葡萄糖。就結果而言， 係以黃花蓮花胎座中所取得含有植物生物素及其植物 性機能成份之提取液的總醣含量為最多，而理論上不同 花色之蓮花胎座中所取得含有植物生物素及其植物性 099101624 表單編號A0101 第丨2頁/共22頁 0992003176-0 201125636 機能成份之提取液的總醣含量亦有所差異。因此，可得 知經由第一較佳實施例中所提供之方法，自不同花色的 連花船座中取得含有M物生物素及其矛直物性機能成份 ，具有多醣成份。 [0041]第四實驗例： _]湘第—較佳實施射所提供之方法，自不同花色的蓮 花胎座中取得含有@物生物素及其植物性機能成份之 提取液，並利用Singleton等人於1 965年所提出之方法 ◎ ’藉由比色分析法，以酚類指示劑(Folin and_ elemental (isoflavonoid) and other ingredients, with antioxidant capacity and inhibition of tyrosinase (tyrosinase) effect, can be applied to cosmetics, nutritional products, health food products and health and food, etc. For example, the phytofunctional components prepared according to the first preferred embodiment of the present invention were tested for their antioxidant capacity, 卩 j 路 胺 生成 \ 总 总 , , , , , , , , , , , , , , , , , , , , , [0022] First Experimental Example: [0023] Taking the method provided in the first preferred embodiment, an extract containing plant biotin and its phytofunctional components was obtained from lotus seat of different colors and using Arnao The method proposed by et al. measures the total antioxidant capacity of the extract containing the phytofunctional component obtained in each flower lotus placenta. Arnao et al. pointed out that the determination of antioxidant capacity can be used 099101624 Form No. 1010101 Page 8 / Total 22 Pages 0992003176-0 201125636 ❹ [0024] [0025] Ο [0026] 2, 2'-N-N-D-( 3-ethylbenzodihydrogen. Sesame __6 - continued acid) diammonium salt (2, 2'-azino-bis (3-ethylbenzthiazoline-6-sulph onic acid) 'ABTS) and peroxidase ( The preoxidase is subjected to an oxidation reaction to form a stable blue-green cation radical; and the antioxidant capacity of the sample to be tested is represented by the amount of the cationic blue-green cation free radical generated by the above reaction. Since this cationic radical has a strong absorption peak at a wavelength of 734 nm. Therefore, the formula is used: Clearance (%) = [1 - (sample 734 nm absorbance / control group 734 nm absorbance)] X 100% : The factory can calculate the percentage of clearing effect. That is, the lower the absorbance value, the stronger the ability to remove ABTS free radicals from the sample. In addition, in this example, different concentrations of trolox antioxidants were used as a positive control group for removing ABTS + free radicals' and a standard curve was established. Trolox is an analogue of water-soluble vitamin E, so trolox acts as an antioxidant and has the effect of preventing and protecting the skin from UV radiation. The test results are shown in Figure 2, 'The extracts of different flower color lotus placenta plant biotin, the scavenging ability to remove ABTS+ free radicals are close to 9〇%, and the anti-oxidation of 0.36 mM trolox is converted by the standard curve. The ability is 95%. Therefore, through the method provided in the first preferred embodiment, the plant-containing biotin and its phytofunctional component are obtained from the lotus seat of different colors, which has the ability to effectively remove ABTS+free radicals and has good antioxidant effect. . 099101624 Form No. A0101 Page 9 / Total 22 Pages 0992003176-0 201125636 [0027] Second Experimental Example: [0〇28] This first experimental example uses different, j Luo Luo aa η 卞扪 with no color 4 The inhibitory effect of the extract on tyrosinase is determined by dopa pigment (d〇PaChr〇me), which is derived from the synthesis of melanin from tyr〇sine. An intermediate product to evaluate the effect on inhibition of melanin synthesis. Taking the method provided in the first preferred embodiment, an extract containing plant biotin and its functional substance is obtained from lotus seat of different colors, and using the method proposed by | et al. The tyrosinase inhibitory ability of the extract containing plant biotin and its phytofunctional component obtained in each flower lotus placenta is determined; since tyrosinase plays a role of rate-limiting enzyme in melanin synthesis, decomposition is utilized. For the reaction, the tyrosinase inhibition ability can be calculated by measuring the absorbance at a wavelength of 475 nm. When the absorbance value is larger, 'the formation of melanin is suppressed to a greater extent. This experiment was performed according to the total absorbance method. .::....:. r 1. :'. ...;丨:\ [0029] First, prepare 4 vials for At, Ai, Ab and AQ, respectively (1) in 4 samples Add 0.9 ml of buffer to dissolve" (2) Add 1 ml of urethane aqueous solution to each of the 4 samples; (3) Add 1 ml of the test sample to human + and 41, (4) Add 1 to Ab and A? Ml deionized water '(5) was placed in a 37 °C water bath for 10 min after '(6) Add 1 ml buffer solution to human 丨 and 八^(7) at At and Ab to add 0.1 m1' concentration 350 units / ml of tyrosinase aqueous solution 'mix the above solution evenly. (8) at 37. (: 25 minutes after reaction in a water bath, (9) use a spectrometer to measure the absorbance at a wavelength of 475 nm. It is obtained from the lotus placenta of different colors by the method provided in the first preferred embodiment. The plant contains 099101624 Form No. A0101 Page 10 / Total 22 Pages 0992003176-0 201125636 Biotin and its botanical functional ingredients are extracted. a solution of commercially available facial acid (k〇jic acid, 1 mg/ml) and arbutin (1 mg/ml) in the same volume, wherein commercially available citric acid and arbutin solutions are used as tyrosine Enzyme inhibitor Standard compound (positive control group). _] (4) Calculation of acidase inhibition rate, calculated by the following formula, [_ inhibition rate u) = {[(Ab_A〇HArV] / % χ1〇〇% [0032] Description of the above code As follows: Circle At: Total absorbance of the sample to be tested, representing the dopa pigment absorbance after adding the test sample to melanin synthesis 阙A1: Absorbance of the no-lumenase blank control group' represents the absorbance of the test article _ Ab : No sample test blank control tissue absorbance, representing dopa pigment absorption in melanin synthesis 〇 round A 〇: total blank control group, absorbance, representing the absorption light of the solvent itself [0〇37] determination results as shown in Figure 3. As shown, yellow flower is optimal for lysinase, even better than arbutin (1 mg/nll) in the positive control group, followed by pink, purple and blue flowers. The method provided in the monthly b example, obtained from the different flower color lotus placenta 1 real plant biotin and its phyto-functional components, has the effect of suppressing 'aminase production β [0038] Experimental example: 099101624 Form number A0101 Page 11 of 22 0992003176-0 201125636 [0040] [0040] The method of the first preferred embodiment obtains an extract 'liquid' containing plant biotin and its phytofunctional component from a lotus seat of different colors and utilizes phenol The -sulforic method (Dubois et al., 1 956) was performed. The phen〇i_suijforic method is formed by the dehydration and decomposition of the five carbon sugars and carbon sugars under acidic and high temperature, and the indole chain will react with the phenol to give an orange color. Therefore, the third experimental example is Using this principle, the total sugar content of the extract containing plant biotin and its phytofunctional components taken from the lotus placenta of different colors is determined, and a standard curve is prepared with glucose solution (y = 0. 291x ^ 1724 ' r2 = 〇. 99), the measured value is converted to the corresponding concentration, and the polysaccharide concentration of the sample solution can be obtained. The results of the experiment are shown in Fig. 4. It can be found that the total sugar content of the extract containing plant biotin and its phytofunctional ingredients obtained from the lotus buds of different colors is 'from the yellow lotus The extract containing plant biotin and its phytofunctional ingredients has a total sugar content equivalent to 5.6 mg/ml glucose, which is derived from the extract of plant biotin and its phyto-functional components from the blue flower placenta. The total sugar content of the liquid is equivalent to 3.81 mg/ml glucose. The total enzyme content of the extract containing plant biotin and its plant functional ingredients from the pink flower placenta is equivalent to 3.92 mg/ml glucose, from purple The total content of the extract of the extract of Μ _ biotin and its phytosanitary component is equivalent to 4. 35 mg / mi glucose. As a result, the total sugar content of the extract containing plant biotin and its phytofunctional components obtained from the placenta of the yellow lotus flower is the most, and the plant organisms obtained in the lotus seat of the different colors are theoretically obtained. Prime and its phytochemical 099101624 Form No. A0101 Page 2 of 22 0992003176-0 201125636 The total sugar content of the extracts of functional ingredients also varies. Therefore, it can be understood that, by the method provided in the first preferred embodiment, the M-containing biotin and its spear-like functional component are obtained from the flower hulls of different colors, and have a polysaccharide component. [0041] The fourth experimental example: _] Xiangdi - preferred implementation of the method provided by the lotus roots of different colors to obtain the extract containing @物生物素 and its phytofunctional ingredients, and the use of Singleton, etc. Method proposed by the person in 1965 ◎ 'by colorimetric analysis, with phenolic indicator (Folin and
Ciocalteu s phenol 檢測樣品中酚類化 σ物的3量。若待測樣品中含有盼類化合物，則會與紛 類才曰不劑反應呈色’其呈色反應之程度可利用光譜儀測 量其於波長760 rnn之吸光值；並以配置不同已知浪度之 沒食子酸（gallic acid)與酚類指示劑反應作為標準 品並製作檢量線，檢測待測樣本與酚類指示劑反應後之 吸光值，對應檢量蟓則可測得其酚類化合物的含量。 〇 國0此’本實驗利用上述原理測定自不同花色的蓮花胎座 中取彳于含有植物生物素及其植物性機能成份之提取液 中的總多酚含量，並以含沒食子酸溶液製作標準曲線（y =〇.2705X _ 0.4349，R2 = 0.99)，即可算出含沒食 子酸之相對量。 [0044]本實驗之結果如第5圖所示。由圖中可發現，黃花蓮花胎 座中取得含有植物生物素及其植物性機能成份之提取 液中的總多酚含量相當於0.67 mg/ml沒食子酸，藍花 0992003176-0 099101624 表單編號A0101 第13頁/共22頁 201125636 蓮花胎座中取得含有植物生物素及其植物性機能成份 之提取液中的總多酚含量相當於1.45 mg/ml沒食子酸 ，粉紅花蓮花胎座中取得含有植物生物素及其植物性 機能成份之提取液中的總多紛含量相當於2.95 mg/ml 沒食子酸，紫紅花蓮花胎座中取得含有植物生物素及其 植物性機能成份之提取液中的總多酚含量相當於0. 81 m g / m 1沒食子酸。就結果而言，紫紅花蓮花胎座中取得含 有植物生物素及其植物性機能成份之提取液中的總多 紛含量較其它花色為高，而花色之差異亦會影響盼類化 合物含量。因此，可得知經由第一較佳實施例中所提供 之方法，自不同花色的蓮花胎座中取得含有植物生物素 及其植物性機能成份，具有多酚成份。 第四實驗例： [0045] 本實驗例係採取不同花色（黃色、粉紅、粉紫及藍色） 蓮花各取20朵，測其花朵重量（花朵總重量/數量）、胎 座重量（胎座總重量/數量），且經由第一較佳實施例所 述之方法以取得含有植物生物素及其植物性機能成份之 後，測定其植物生物素之pH值，並計算植物生物素萃取 率（植物生物素總重量/胎座總重量xlOO%)。結果如表 一所示： [0046] 表一、探討不同花色之植物生物素之萃取率 099101624 表箪編號A0101 第14頁/共22頁 0992003176-0 201125636 [0047] 4匕巴 __名稱 數量 花朵重量 27.82±4.76 公克 黃色 胎座重量 5.91±1.21 公克 萃取率· 16.93 % DH佶 4.91 花朵總重量 21.85±2.8 公克 粉紅 胎座重量 3·85±1·42 公克 萃取率 25.97 % __ρΗΛ_ 4.99 花朵總重量 21.85±5.16 公克 粉紫 胎座重量 3.85±1.42 公克 萃取率 17.47% ΡΗ 值 4.95 藍色 胎座重量 萃取率· pmCiocalteu s phenol detects 3 amounts of phenolated σ in the sample. If the sample to be tested contains a compound of interest, it will react with a variety of dyes to form a color. The degree of color reaction can be measured by a spectrometer at a wavelength of 760 rnn; and configured with different known waves. The gallic acid reacts with the phenolic indicator as a standard and prepares a calibration curve to detect the absorbance of the sample to be tested and the phenolic indicator, and the corresponding phenol can be measured. The content of the compound. 〇国0This 'This experiment uses the above principle to determine the total polyphenol content in the lotus roots of different colors from the extract containing plant biotin and its phytofunctional ingredients, and contains gallic acid solution By making a standard curve (y = 〇.2705X _ 0.4349, R2 = 0.99), the relative amount of gallic acid can be calculated. [0044] The results of this experiment are shown in Figure 5. It can be seen from the figure that the total polyphenol content in the extract containing plant biotin and its phytofunctional components is equivalent to 0.67 mg/ml gallic acid, blue flower 0992003176-0 099101624 No. A0101 Page 13 of 22 201125636 The total polyphenol content in the extract containing plant biotin and its phytofunctional ingredients in the lotus placenta is equivalent to 1.45 mg/ml gallic acid, pink flower lotus placenta The total content of the extract containing plant biotin and its phytofunctional components is equivalent to 2.95 mg/ml gallic acid, and the plant buds and plant functions are obtained in the sternal lotus placenta. The total polyphenol content in the extract is equivalent to 0.81 mg / m 1 gallic acid. As a result, the total amount of extracts containing plant biotin and its phyto-functional components in the purple primordial placenta is higher than that of other colors, and the difference in flower color also affects the content of the desired compound. Thus, it can be seen that, via the method provided in the first preferred embodiment, plant biotin and its phytofunctional ingredients are obtained from lotus seat of different colors, having a polyphenol component. The fourth experimental example: [0045] This experiment is to take different colors (yellow, pink, pink and blue). Take 20 lotus flowers, measure the weight of the flowers (total weight/number of flowers), the weight of the placenta (the placenta) Total weight/quantity), and after obtaining the plant biotin and its phytofunctional components by the method described in the first preferred embodiment, determining the pH of the plant biotin and calculating the plant biotin extraction rate (plant) Total biotin weight / total carcass weight x100%). The results are shown in Table 1: [0046] Table 1. Exploring the extraction rate of plant biotin of different colors 099101624 Table No. A0101 Page 14 of 22 Page 0992003176-0 201125636 [0047] 4匕巴__Name number of flowers Weight 27.82±4.76 g Yellow seat weight 5.91±1.21 g extraction rate · 16.93 % DH佶4.91 total flower weight 21.85±2.8 g pink seat weight 3.85±1·42 g extraction rate 25.97 % __ρΗΛ_ 4.99 total flower weight 21.85 ±5.16 gram pink purple placenta weight 3.85±1.42 gram extraction rate 17.47% ΡΗ value 4.95 blue placen weight extraction rate · pm
5.86 ±1.5 公克 17.08% a on [0048] 以上所述僅為本發明之較佳實施例及實驗例，並非用以 限疋本發明之申請專利權利；同時以上的描述，對於熟 知本技術領域之專門人士應可明瞭及實施，因此其他未 脫離本發明所揭示之精神下所完成的等效改變或修飾， 均應包含在申請專利範圍中。5.86 ± 1.5 gram 17.08% a on [0048] The above description is only the preferred embodiments and experimental examples of the present invention, and is not intended to limit the patent application rights of the present invention; and the above description is well known in the art. It should be understood and implemented by those skilled in the art that other equivalent changes or modifications may be made without departing from the spirit of the invention.
【圖式簡單說明】 [0049] 第1圖為一流程圖，係根據本發明提供之第一較佳實施例 ，為從雙子葉植物胎座中萃取出植物生物素及其植物 性機能成份之方法。 [0050] 第2圖為根據本發明第一實驗例中所提取之不同花色蓮花 胎座植物生物素及其植物性機能成份之提取液之總抗 氧化能力。 [0051] 第3圖為根據本發明第二實驗例中所提取之不同花色蓮花 胎座植物生物素及其植物性機能成份之提取液之抑制 099101624 表單編號A0101 第15頁/共22頁 099: 201125636 酪胺酸酶能力》 [0052] [0053] [0054] [0055] [0056] [0057] 第4圖為根據本發明第三實驗例中所提取之不同花色蓮花 胎座植物生物素及其植物性機能成份 之提取液之總醣 含量。 第5圖為根據本發明第四實驗例中所提取之不同花色蓮 化胎座植物生物素及其植物性機能成份之提取液之總 多齡含量。 第6A、6B、6C圖所示為—雙子葉植物胎座。 第7A圖為根據本發0$提供之第—較佳實施例經步驟⑷ 處理後所得之濾液，呈現凝膠狀乂 第7B圖為根據本發明提供之第—較佳實施例，經步驟（e) 處理後所得乾燥之無色無味的植物生物素及其植物性機 能成份。 【主要元件符號說明】 步驟（a)、（b)、（c)、（時、（θ) 099101624 表單煸號A0101 第丨6頁/共22頁 0992003176-0BRIEF DESCRIPTION OF THE DRAWINGS [0049] FIG. 1 is a flow chart showing the extraction of plant biotin and its phytofunctional components from a dicotyledonous placenta according to a first preferred embodiment of the present invention. method. 2 is a graph showing the total antioxidant capacity of extracts of different color lotus flower placental plant biotin and its plant functional ingredients extracted according to the first experimental example of the present invention. Figure 3 is a diagram showing the inhibition of extracts of different flower color lotus placenta plant biotin and its plant functional ingredients extracted according to the second experimental example of the present invention 099101624 Form No. A0101 Page 15 of 22 099: 201125636 tyrosinase ability [0057] [0057] [0057] FIG. 4 is a diagram showing the different flower color lotus placenta plant biotin extracted according to the third experimental example of the present invention and The total sugar content of the extract of plant functional ingredients. Fig. 5 is a graph showing the total age of the extracts of the different flower coloring placental plant biotin and its plant functional ingredients extracted according to the fourth experimental example of the present invention. Figures 6A, 6B, and 6C show a dicotyledonous placenta. Figure 7A is a view of the filtrate obtained after the step (4) according to the first preferred embodiment of the present invention, which exhibits a gelatinous state. Figure 7B shows a first preferred embodiment according to the present invention. e) dried, colorless and odorless plant biotin and its phytofunctional ingredients. [Description of main component symbols] Steps (a), (b), (c), (hour, (θ) 099101624 Form nickname A0101 Page 6 of 22 0992003176-0