JPH11192086A - Tempeh enzyme - Google Patents

Abstract

PROBLEM TO BE SOLVED: To readily obtain a super oxide dismutase enzyme agent, which can not be separated from common grains themselves, from a tempeh. SOLUTION: This tempeh enzyme having super oxide dismutase enzyme activity is obtained by extraction from the tempeh by using water or a diluted salt solution. The prepared substance is purified directly as it is or by combination of one or more operations of a proper polar organic solvent treatment, adsorption chromatography, ion exchange chromatography, gel filtration or the like.

Description

【発明の詳細な説明】DETAILED DESCRIPTION OF THE INVENTION

【０００１】[0001]

【発明の属する技術分野】本発明はテンペより水、ある
いは希薄塩溶液を用いて抽出することを特徴とするスー
パーオキシドジスムターゼ活性を有するテンペ酵素に関
する。今日、癌や各種炎症性疾患、その他の成人病予防
目的に、食品由来の本酵素は極めて強力な活性を持つた
め安価に調整でき、食して安全性が高く、また熱に対す
る安定性もよく、利用価値が高いと考えられる。TECHNICAL FIELD The present invention relates to a tempe enzyme having superoxide dismutase activity, which is extracted from tempe using water or a dilute salt solution. Today, for the purpose of preventing cancer, various inflammatory diseases, and other adult diseases, this enzyme derived from food has extremely strong activity and can be adjusted at low cost, is safe to eat, and has good stability to heat, It is considered that the utility value is high.

【０００２】[0002]

【従来の技術】各種疾患に対してそれを治療、あるいは
予防するためのスーパーオキシドジスムターゼ酵素剤と
して、これまで動物及び植物由来のものが報告されてい
るが、発酵によりいくらでも大量生産が可能で、安価、
且つ食べたり飲んで安全性の保障されたものは少なかっ
た。特に、リゾープスを用いた発酵食品であるテンペ由
来の強力な本酵素は、これまで全く報告がなかった。2. Description of the Related Art As a superoxide dismutase enzyme preparation for treating or preventing various diseases, those derived from animals and plants have been reported so far, but any amount can be mass-produced by fermentation. Cheap,
Few foods or drinks were guaranteed safe. In particular, a strong present enzyme derived from tempeh, a fermented food using resorps, has never been reported.

【０００３】[0003]

【発明が解決しようとする課題】一般の穀類そのものか
らは分離しえない強力なスーパーオキシドジスムターゼ
酵素をテンペから容易に取得する。このものは安価で、
食べたり飲んだりしても安全であるがゆえに、薬品とし
てのみならず、特に成人病予防及び予後目的のドリンク
剤あるいは機能性食品素材としての応用が期待できる。SUMMARY OF THE INVENTION A powerful superoxide dismutase enzyme which cannot be separated from general cereals itself can be easily obtained from Tempe. This one is cheap,
Since it is safe to eat and drink, it can be expected to be used not only as a drug but also as a drink or a functional food material especially for the prevention and prognosis of adult diseases.

【０００４】[0004]

【課題を解決するための手段】我々は、食品を中心に天
然素材中に種々の生理活性物質を検索することに鋭意努
力し、特に日本の伝統的発酵食品である納豆やインドネ
シア納豆とも呼ばれるテンペ中に血栓溶解酵素であるナ
ットウキナーゼ、あるいはテンペキナーゼを発見し、そ
の取得方法及び酵素学的性質について報告してきた（須
見ら、機能性食品素材、食品由来の生理活性物質におけ
る研究と開発、ｐ．８８、工業技術会、１９８９；日本
生化学会誌、６１：８３４、１９８９；月刊フードケミ
カル、１２：７２、１９９０；食品機能学への招待、
ｐ．３６、三共出版、１９９６）。このように対象とし
て発酵食品由来のものを主に選んだのは、長年摂取され
ても安全性に問題が少ないからである。本発明は、そう
した研究の過程で発見するに至った極めて強力なテンペ
由来のスーパーオキシドジスムターゼ活性を有するテン
ペ酵素に関するものである。[Means for Solving the Problems] We have worked diligently to search for various physiologically active substances in natural materials, especially foods, and in particular, tempeh, which is also called Japanese traditional fermented food, natto or Indonesian natto. Nattokinase or tempekinase, which is a thrombolytic enzyme, has been discovered in them, and their acquisition method and enzymatic properties have been reported (Sumi et al., Research and Development on Functional Food Materials, Food-Derived Physiologically Active Substances, p. 88, Industrial Technology Association, 1989; Journal of Biochemical Society of Japan, 61: 834, 1989; Monthly Food Chemical, 12:72, 1990;
p. 36, Sankyo Publishing, 1996). The reason why fermented food-derived foods are mainly selected in this way is that there is little problem in safety even if ingested for many years. The present invention relates to a tempe enzyme having superoxide dismutase activity derived from tempe, which was discovered during the course of such research.

【０００５】[0005]

【発明の実施の形態】次に本発明を実施例にて詳細に説
明する。Next, the present invention will be described in detail with reference to examples.

【０００６】第１例 １ｋｇの蒸煮ハトムギをオートクレーブで１２０℃、３
０分間滅菌処理済みの綿栓付きの１０ｌのステンレス容
器に入れ、ｐＨ４．０になるまで乳酸を添加混合した
後、２ｇのテンペ菌（秋田今野商店、秋田）を加え、よ
く撹拌した後、３７℃の恒温槽内で静置培養した。３日
後の発酵に生理的食塩水を加えて２時間撹拌後、東洋濾
紙Ｎｏ．５０で濾過し、得られた瀘液をエバポレーター
で濃縮した。薄黄色の本物質のスーパーオキシドジスム
ターゼ（ＳＯＤ）活性をＥｌｓｔｎｅｒの亜硝酸法（Ａ
ｎａｌ．Ｂｉｏｃｈｅｍ．，７０：６１６，１９７６）
で調べた結果、テンペの乾燥品ｇ当たりに換算して２９
０単位（ｃｕ）のＳＯＤ活性が確認できた（表１）。こ
の活性はＳＯＤ市販製品（日本薬品開発（株）のグリー
ンマグマ、プリセプト（株）の百種百源）と比べても、
そのままの状態で見劣りしない強いものであった。また
このようなＳＯＤ活性は、抽出濃縮されたものを１００
℃で３０分間加熱処理してももとの９０％以上の活性を
持ち、ヒト血球ＳＯＤ剤のように失活することはなかっ
た。First Example 1 kg of steamed barley is autoclaved at 120.degree.
After placing in a 10-liter stainless steel container with a cotton stopper sterilized for 0 minutes, adding and mixing lactic acid until the pH becomes 4.0, 2 g of Tempeh bacteria (Akita Konno Shoten, Akita) was added, and the mixture was stirred well. The culture was allowed to stand still in a constant temperature bath at ℃. To the fermentation after 3 days, physiological saline was added and stirred for 2 hours. The mixture was filtered at 50, and the obtained filtrate was concentrated by an evaporator. The light yellow superoxide dismutase (SOD) activity of the substance was determined by the Elstner nitrite method (A
nal. Biochem. , 70: 616, 1976).
As a result of the above, it was calculated as 29 per g of dried tempeh.
SOD activity of 0 units (cu) was confirmed (Table 1). This activity is comparable to that of commercially available SOD products (green magma of Nippon Pharmaceutical Development Co., Ltd., 100 kinds and 100 sources of Precept Co., Ltd.)
It was strong as it was. In addition, such SOD activity is determined by extracting and concentrating the SOD activity by 100%.
Even after heat treatment at 30 ° C. for 30 minutes, it had an activity of 90% or more of the original activity, and did not inactivate unlike human blood cell SOD agents.

【表１】 [Table 1]

【０００７】第２例 １００ｇの市販オカラ（ニシナ（株）、岡山市）をオー
トクレーブで１２０℃、３０分間滅菌処理したものに少
量の乳酸（ｐＨ４．０とする）及びテンペ菌（秋田今野
商店、３０ｍｇ）を添加し、よく撹拌した後、３７℃の
恒温槽内で２日間保温し、オカラテンペを作製した。な
お、容器は乾熱滅菌した５００ｍｌの三角フラスコを使
用した。テンペを凍結乾燥し、ミキサーにかけて約１３
ｇの粉末を得た。このものに５００ｍｌの水を加えて１
時間撹拌抽出後、東洋濾紙Ｎｏ．５０で濾過したものを
凍結乾燥した。このものに含まれる活性を第１例と同じ
亜硝酸法で測定した結果、乾燥ｇ当たり１１５ＣＵのＳ
ＯＤ活性を示した。なお、オカラに重量当り５％量の大
豆蒸煮で得られた煮汁を加えると、それから得られたＳ
ＯＤ活性は約１８％高まった。Second Example 100 g of commercially available okara (Nishina Co., Okayama City) was sterilized in an autoclave at 120 ° C. for 30 minutes, and a small amount of lactic acid (pH 4.0) and tempeh bacteria (Akita Konno Shoten, After stirring well, the mixture was kept warm in a thermostat at 37 ° C. for 2 days to produce okara tempe. The container used was a dry-heat sterilized 500 ml Erlenmeyer flask. Lyophilize Tempe and mix in a mixer for about 13
g of powder was obtained. Add 500 ml of water to this and add 1
After stirring and extracting for hours, Toyo Filter Paper No. Those filtered at 50 were lyophilized. The activity contained therein was measured by the same nitrous acid method as in the first example.
OD activity was shown. In addition, when the broth obtained by soybean steaming of an amount of 5% by weight is added to okara, S obtained from it is added.
OD activity increased by about 18%.

【０００８】第３例 １ｋｇの蒸煮した大豆、及び黒豆の各々を１２０℃、３
０分間滅菌処理した後、少量のクエン酸（ｐＨ４．０と
する）を加え撹拌後、５００ｍｇのテンペ菌（秋田今野
商店、秋田）を添加し、再度よく撹拌し、４０℃の恒温
槽内で５日間静置培養したものを５０℃で減圧乾燥し、
各々４１の蒸留水で抽出した。これらの抽出液をメンブ
ランフィルター（０．２μ）に通した後、ＤＡＥＡ−セ
ルロース及びＣＭ−セルロースカラムクロマトグラフィ
ーで精製後、その溶出液をエバポレーターで濃縮したも
のには第１例と同じ亜硝酸法でｇ当たり、各々２８００
ＣＵ、及び２３４０ＣＵという強力なＳＯＤ活性が確認
できた。Third Example 1 kg of steamed soybean and black bean were each heated at 120 ° C. for 3 hours.
After sterilizing for 0 minutes, add a small amount of citric acid (pH 4.0), stir, add 500 mg of Tempeh bacteria (Akita Konno Shoten, Akita), stir well again, and in a 40 ° C constant temperature bath. The stationary culture for 5 days was dried under reduced pressure at 50 ° C.
Each was extracted with 41 distilled water. These extracts were passed through a membrane filter (0.2 µ), purified by DAEA-cellulose and CM-cellulose column chromatography, and the eluate was concentrated by an evaporator. 2g each per g
Strong SOD activity of CU and 2340 CU was confirmed.

【０００９】第４例 第１例に順じてハトムギテンペから水で抽出し、凍結乾
燥したＳＯＤ剤（Ｌｏｔ．Ｎｏ．２２５７５）の３０ｇ
を１回分として、６人の健常成人に毎日朝１０時に経口
摂取させ、それを１カ月続けて経時的に採血し、血液中
の線溶活性及び血圧の変化を調べてみた。その結果、表
２に示すように線溶亢進を示す有意なＥＬＴ（ｅｕｇｌ
ｏｂｕｌｉｎ ｌｙｓｉｓ ｔｉｍｅ）の延長（ｐ
〈０．０５）と血圧降下傾向が確認された。Fourth Example According to the first example, 30 g of a SOD agent (Lot. No. 22575) extracted with water from adlay tempe and freeze-dried.
Was taken orally in 6 healthy adults every day at 10 o'clock in the morning, and blood was collected over time for one month continuously to examine changes in blood fibrinolytic activity and blood pressure. As a result, as shown in Table 2, significant ELT (eugl
Obulin lysis time) (p
<0.05), indicating a tendency to lower blood pressure.

【表２】 [Table 2]

【００１０】[0010]

【発明の効果】本発明によれば、大量生産が容易で、安
価、且つ食べたり飲んだりしても安全なスーパーオキシ
ドジスムターゼ活性を有するテンペ酵素が提供できる。According to the present invention, it is possible to provide a tempe enzyme having a superoxide dismutase activity which can be easily mass-produced, is inexpensive, and is safe to eat and drink.

Claims (2)

【特許請求の範囲】[Claims] 【請求項１】 テンペより水、あるいは希薄塩溶液を用
いて抽出することを特徴とするスーパーオキシドジスム
ターゼ活性を有するテンペ酵素。1. A tempe enzyme having superoxide dismutase activity, which is extracted from tempeh with water or a dilute salt solution. 【請求項２】 得られた物質をそのまま、または適当な
極性有機溶媒処理、限外濾過処理、吸着クロマトグラフ
ィー、イオン交換クロマトグラフィー、ゲル濾過などの
操作を一種類以上組み合わせて精製する請求項１に記載
の取得法。2. The obtained substance is purified as it is or by a combination of one or more operations such as appropriate polar organic solvent treatment, ultrafiltration treatment, adsorption chromatography, ion exchange chromatography, and gel filtration. Acquisition method described in.