JP3547835B2 - Prophylactic / therapeutic agent for throat inflammation and hemolytic toxin and oral composition containing the same - Google Patents
Prophylactic / therapeutic agent for throat inflammation and hemolytic toxin and oral composition containing the same Download PDFInfo
- Publication number
- JP3547835B2 JP3547835B2 JP06794395A JP6794395A JP3547835B2 JP 3547835 B2 JP3547835 B2 JP 3547835B2 JP 06794395 A JP06794395 A JP 06794395A JP 6794395 A JP6794395 A JP 6794395A JP 3547835 B2 JP3547835 B2 JP 3547835B2
- Authority
- JP
- Japan
- Prior art keywords
- present
- eucalyptus
- compound
- hemolytic
- prophylactic
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Lifetime
Links
- QSDZEUSKRROZOO-MRTOFGJQSA-N CC(C1)Oc2c(C=O)c(O)c(C=O)c(O)c2[C@@H]1[C@H](CC1)[C@](C)(CC[C@H](C2)C(C)(C)O)[C@@H]2[C@]1(C)O Chemical compound CC(C1)Oc2c(C=O)c(O)c(C=O)c(O)c2[C@@H]1[C@H](CC1)[C@](C)(CC[C@H](C2)C(C)(C)O)[C@@H]2[C@]1(C)O QSDZEUSKRROZOO-MRTOFGJQSA-N 0.000 description 1
Landscapes
- Cosmetics (AREA)
- Medicines Containing Plant Substances (AREA)
- Acyclic And Carbocyclic Compounds In Medicinal Compositions (AREA)
Description
【０００１】
【産業上の利用分野】
本発明は、ユーカリ属植物由来の喉炎症・溶血毒の予防・治療剤およびこれを含有する口腔用組成物に関する。
【０００２】
【従来の技術】
溶血性連鎖球菌（Ｓｔｒｅｐｔｏｃｏｃｃｕｓ ｐｙｏｇｅｎｅｓ：ストレプトコッカス・ピオゲネス）は、ヒトの口腔内や咽頭などに分布する通性嫌気性グラム陽性球菌である。本菌は連鎖球菌の中で最も病原性が強く、粘膜では咽頭炎、扁桃炎、中耳炎等を起こし、皮膚では種々の化膿性炎症をを引き起こすことが知られており、いずれも咽頭もしくは扁桃腺において増殖する。
【０００３】
本菌の病原性因子として、溶血毒（ストレプトリジンＯ）、ディック毒素、ストレプトキナーゼ、ＤＮＡ分解酵素、ヒアルロニダーゼ等が知られており、咽頭炎、扁桃炎、気管支炎等の呼吸器系疾患の病因的役割を果たしている。
【０００４】
これらの呼吸器系疾患を予防および治療するためには、病巣において本菌類の生育を阻害することおよび本菌類の病原性因子を抑制することが重要である。そして、菌の抑制という観点では、本菌類に対して有効な抗菌性物質の応用が効果的である。
【０００５】
従来、本菌類に対する抗菌性物質としてペニシリン系、セフェム系、セファロスポリン系の抗生物質が用いられていたが、これらは作用が強力である反面、耐性菌の出現や副作用の発現等から日常的な利用に適しているとはいえない。
【０００６】
本発明者等は、非抗生物質系で、本菌に対する抗菌活性および本菌の産生する溶血毒阻害活性を有する物質を得んとして、各種天然物の抽出物につきスクリーニングを行った結果、ユーカリの精油（ユーカリ油）以外の抽出物に着目した。ユーカリは、ハーブティーや香料成分として用いられているものであり、その安全性についての問題はなく、抗生物質を応用する場合のような副作用の問題は生じない。
【０００７】
現在までに、ユーカリ抽出物の効果としては、ユーカリ油およびその主要成分であるシネオール（ユーカリプトール）に防腐剤としての効果や、抗菌活性物質の作用を増強することが知られている（特開昭６２−２８９５１１号公報、抗細菌性口腔用組成物）。しかしながら、このユーカリ油やシネオール自体の溶血性連鎖球菌に対する抗菌活性は弱いのみならず、その強い独特の臭気により、例えば口腔内で使用するに当たり用途や添加量においてかなりの制約を受ける欠点がある。
【０００８】
また、特開昭５８−３９６１５号公報（歯磨き組成物）は、ユーカリ抽出物とアルキル硫酸ナトリウムとの併用が歯周病原因菌の一つであるフゾバクテリウム・ヌクレイタムに対する抗菌活性を示すことを開示している。このユーカリ抽出物は、アルキル硫酸ナトリウムとの併用をもって初めて抗菌活性を発現するもので、それ自体には抗菌活性がないか、例えあったとしても非常に微弱なことが示唆される。しかも、この組成物には溶血性連鎖球菌に対する効果は示されていない。
【０００９】
さらに、特開平５−３０６２５２号公報（新規マクロカルパール類及びその製造法）は、ユーカリ属植物の溶媒抽出物より採取したマクロカルパール類がアルドースリダクターゼ阻害活性および抗菌活性を示すことを開示しているが、その抗菌活性はいくつかのグラム陽性菌に対して作用することが明らかにされているに過ぎず、溶血性連鎖球菌に対する抗菌活性は示されていない。
【００１０】
一方、特開昭６２−５９２１５号公報（外用鎮咳、去啖、鎮痛、鎮静剤）では、ｄｌ−カンフル、１−メントール、ニクズク油およびウイキョウ油配合クリームに、さらにユーカリ油を配合することにより、外用時における鎮咳、去啖効果を増強することを開示している。この公報では、溶血性連鎖球菌に対する抗菌活性が微弱なユーカリ油を有効成分としているのみならず、外用という投与形態により揮発性成分の作用を期待するものであり、口腔内において非揮発性で使用することが求められる溶血性連鎖球菌に対する抗菌活性剤とは基本的に異なるものである。
【００１１】
このようにユーカリ属抽出物は各種開示されているが、いずれも喉炎症・溶血毒の予防・治療には不十分で汎用性に乏しいものである。
【００１２】
【発明が解決しようとする課題】
そこで本発明者等はユーカリ属植物から喉炎症・溶血毒両者の予防、治療に有効な成分を抽出せんとして鋭意研究を行った結果、ユーカリ属植物から精油成分（ユーカリ油）を除去した残渣を極性溶媒で抽出した抽出物およびこれに含まれるフロログルシノール誘導体が喉炎症起因菌（溶血性連鎖球菌）に対して非常に強い抗菌活性を有し、さらに本菌種の産生する溶血毒の阻害活性をも併せ持つことを見出し、本発明を完成させるに至った。
【００１３】
なお、ユーカリ属植物植物に含まれるフロログルシノール誘導体については抗ウイルス作用［Ｃｈｅｍ．Ｐｈａｒｍ．Ｂｕｌｌ．，３８（５）１４４４〜１４４６（１９９０）］、ＨＩＶ逆転写酵素阻害作用［Ｔｅｔｒａｈｅｄｒｏｎ Ｌｅｔｔ．，３３（２１）２９８３〜２９８６（１９９２）］などが報告されているが、本発明における喉炎症・溶血毒の予防・治療効果については知られておらず、本発明によって初めて明らかにされたものである。
【００１４】
【課題を解決するための手段】
本発明はユーカリ属植物から精油成分（ユーカリ油）を除去した残渣を極性溶媒で処理した抽出物およびこの抽出物から活性本体として得られるフロログルシノール誘導体を有効成分とする喉炎症・溶血毒の予防・治療剤並びにこれらを用いた口腔用組成物である。
【００１５】
以下に、本発明製剤の抽出製造例を挙げて説明する。
【００１６】
先ず、ユーカリ属植物、例えばユーカリ葉を粉砕機等の適当な粉砕手段で粉砕し、好ましくは通常精油抽出で使用する水蒸気蒸留装置を用いて精油成分を除去する。この精油成分の除去はｎ−ヘキサン、石油エーテル等の低極性溶媒を用いて常温で抽出することもできる。
【００１７】
このようにして得られた精油抽出残渣に酢酸エチル、アセトン、エタノール、メタノール、水等の極性溶媒の少なくとも１つの溶媒を適用して抽出を行い、得られた抽出液を減圧濃縮することにより抽出物の形態で本発明製剤を得る。
【００１８】
このようにして得られた製剤を、さらにカラムクロマトグラフィ等の適当な分離精製手段を用いて分離精製することにより化合物の形態の本発明製剤であるフロログルシノール誘導体を得る。
【００１９】
このフロログルシノール誘導体は、一般式
【化１０】
【００２０】
本発明製剤として使用するフロログルシノール誘導体の具体例としては、次の構造式のものが挙げられる。
【００２１】
【化１１】
【化１２】
【化１３】
【化１４】
【化１５】
【化１６】
【化１７】
【化１８】
【化１９】
なお、フロログルシノール誘導体の分離精製に使用するカラムクロマトグラフィとしては、吸着クロマトグラフィ、分配クロマトグラフィ、高速液体クロマトグラフィ、薄層クロマトグラフィ等が挙げられる。
【００３１】
本発明製剤の用途については種々考えられるが、香り、呈味性に優れ、安全性が高いことからチューインガム、キャンディ、錠菓、含そう剤等に配合する等の日常的な利用が可能である。本発明製剤を食品等に配合する場合、０．００１〜１０重量％の割合になるように添加するのが好適である。
【００３２】
以下に実施例、試験例、応用例を挙げて説明するが、これらの例は本発明の範囲を制限するものではない。
【００３３】
【実施例】
実施例１
ユーカリ葉粉末１ｋｇから水蒸気蒸留装置を用いて精油成分を除去した後、エタノール５リットルを加え、３時間加熱還流抽出を行った。得られた濾液を減圧下濃縮することにより本発明製剤である抽出物２１４ｇを得た。
【００３４】
実施例２
ユーカリ葉粉末１ｋｇから水蒸気蒸留装置を用いて精油成分を除去した後、エタノール−水（１：１）混液５リットルを加え、３時間加熱還流抽出を行った。得られた濾液を減圧下濃縮することにより本発明製剤である抽出物２３４ｇを得た。
【００３５】
実施例３
実施例２で得られた抽出物１００ｇに水を加えて分散させた後、酢酸エチルで分配し、酢酸エチル可溶画分（２９．５ｇ）を得た。この酢酸エチル可溶画分をシリカゲル（ワコーゲルＣ−２００、和光純薬）１ｋｇを充填したカラムに吸着させた。
【００３６】
このカラムをジクロロメタン−メタノール（５０：１〜２：１）混液にて溶出させ、分画を行った。ＴＬＣプレート（６０Ｆ２５４、メルク）を用いてジクロロメタン−メタノール（３：１）混合溶液で展開した時、Ｒｆ値０．２〜０．４付近にＵＶ吸収があり、１０％硫酸により紫色、灰色もしくは茶色のスポットを呈する画分を集めた。この画分を高速液体クロマトグラフィ（ＨＰＬＣ）用のオクタデシルシラノール型（ＯＤＳ）分取カラム（センシュー科学）を用いて、メタノール−水（９０：１０〜１００：０）で溶出させ、２８５ナノメータのＵＶ吸収をモニターしながら各ピークを分取した。
【００３７】
それぞれのピークは、さらにＨＰＬＣ用シリカゲル分取カラム（センシュー科学）を用いてジクロロメタン−メタノール（９８：２〜８５：１５）で、またＨＰＬＣ用ＯＤＳ分取カラムを用いて、さらにアセトニトリル−水（９０：１０〜１００：０）およびメタノール−水（９０：１０〜１００：０）で繰り返し精製して本発明製剤である前記化合物１〜９を得た。
【００３８】
因みに化合物５〜９の物理化学的性質を以下に記載する。
（化合物５）
１）性状：微黄色粉末
２）融点：非結晶のため測定せず。
３）分子式：Ｃ28Ｈ38Ｏ7
４）ＦＡＢ−マススペクトル：４８７［Ｍ＋１］
【数１】
【化２０】
（化合物６）
１）性状：微黄色粉末
２）融点：非結晶のため測定せず。
３）分子式：Ｃ28Ｈ40Ｏ6
４）ＥＩ−マススペクトル：４７２［Ｍ］,454,251,195
【数３】
【化２１】
（化合物７）
１）性状：微黄色粉末
２）融点：非結晶のため測定せず。
３）分子式：Ｃ28Ｈ42Ｏ7
４）ＥＩ−マススペクトル：４７２［Ｍ−Ｈ2Ｏ］,454,251,195
【数５】
【化２２】
（化合物８）
１）性状：微黄色粉末
２）融点：非結晶のため測定せず。
３）分子式：Ｃ28Ｈ42Ｏ7
４）ＥＩ−マススペクトル：４７２［Ｍ−Ｈ2Ｏ］,454,251,195
【数７】
【化２３】
（化合物９）
１）性状：微黄色粉末
２）融点：非結晶のため測定せず。
３）分子式：Ｃ28Ｈ38Ｏ7
４）ＥＩ−マススペクトル：４６８［Ｍ−Ｈ2Ｏ］,427,251,217,195
【数９】
【化２４】
試験例１
実施例１〜３で調製した本発明製剤の溶血性連鎖球菌および腸内細菌に対する抗菌活性を、以下の方法により試験した。
【００４４】
試料として本発明製剤、ユーカリ油および殺菌剤として知られるチモールを用いて本発明製剤との抗菌活性の比較を行った。
【００４５】
（試験方法）
１）供試菌株、使用培地および培養条件
溶血性連鎖球菌２株および腸内細菌４株をそれぞれの培地および培養条件で２４〜４８時間培養し、菌を充分に生育させる。９６穴平底マイクロプレートを用い、各ウエル中で試料溶液（１００μｌ）の２倍希釈系列を調製する。２倍濃度で調製した培地に各々の菌を接種し、その１００μｌを各ウエルに添加する。
【００４６】
２４〜４８時間培養した後、マイクロプレートリーダーを用いて各ウエルの濁度（５５０ｎｍ）を測定し、菌の生育を判定し、その最小発育阻止濃度（ＭＩＣ）を求める。
【００４７】
（試験結果）
試験結果を表１に示す。本発明製剤は溶血性連鎖球菌に対して強い抗菌活性を示した。また、チモール、ユーカリ油は本発明製剤の抗菌活性と比較すると、その作用は著しく劣るものであった。さらに、本発明製剤の腸内細菌に対する抗菌活性は溶血性連鎖球菌に対する活性と比較してかなり弱いものであり、本発明製剤による腸内細菌に対する影響は少ないものと思われた。
【００４８】
【表１】
試験例２
実施例１〜３で調製した本発明製剤の溶血毒（ストレプトリジンＯ）阻害活性を以下の方法により試験した。
【００５０】
（試験方法）
ウサギ脱繊維血液２５ｍｌを緩衝食塩水（ｐＨ６．５）で３回洗浄（３０００ｒｐｍ×１０分間）し、赤血球を集める。緩衝食塩水で２００ｍｌに希釈し、ウサギ赤血球浮遊液とする。試料規定量をジメチルスルフォキシド（ＤＭＳＯ）に溶解してから、さらに緩衝食塩水に溶解し（ＤＭＳＯ４％）試料溶液とする。
【００５１】
試料溶液６００μｌとストレプトリジンＯ（栄研）水溶液（１結合力価）３００μｌを１３ｍｍ径の試験管中で混合し、３７℃で３０分間反応させる。ウサギ赤血球浮遊液３００μｌを添加し、３７℃で４５分間反応させる。反応終了後、１５００ｒｐｍで１分間遠心分離し、上清の溶血度合を肉眼で判定する。
【００５２】
判定基準 −：溶血のみられないもの、＋：不完全溶血、＋＋：完全溶血とする。
【００５３】
（試験結果）
試験結果を表２に示す。本発明製剤の殆どは１０００μｇ／ｍｌで完全にストレプトリジンＯによる溶血を阻害し、５００μｇ／ｍｌにおいても阻害活性を示した。
【００５４】
【表２】
応用例１
実施例１〜２の方法により調製した本発明製剤を用いて、次の処方によりチューインガム、キャンディ、錠菓、含そう剤を製造した。
【００５６】
１．チューインガムの処方
ガムベース ２０．０％
砂糖 ５５．０
グルコース １５．０
水飴 ９．３
香料 ０．５
本発明製剤（実施例１） ０．２
１００．０％
【００５７】
２．キャンディの処方
砂糖 ５０．０％
水飴 ３４．０
クエン酸 １．０
本発明製剤（実施例１） ０．２
水 １４．８
１００．０％
【００５８】
３．錠菓の処方
砂糖 ７６．４％
グルコース １９．０
ショ糖脂肪酸エステル ０．２
本発明製剤（実施例２） ０．２
水 ４．２
１００．０％
【００５９】
４．含そう剤の処方
エタノール ３０．０％
香料 １．０
銅クロロフィリンナトリウム ０．１
サッカリン ０．０５
塩酸クロルヘキシジン ０．０１
本発明製剤（実施例２） ０．２
水 ６８．６４
１００．０％
【００６０】
応用例２
応用例１の１の処方で、実施例１の本発明製剤の代わりに実施例３で得られた化合物１を０．００５％、砂糖を５５．１９５％含有する他は同様にしてチューインガムを製造した。
【００６１】
応用例３
応用例１の２の処方で、実施例１の本発明製剤の代わりに実施例３で得られた化合物２を８．２％、砂糖を４２％含有する他は同様にしてキャンディを製造した。
【００６２】
応用例４
応用例２の処方で、化合物１の代わりに実施例３で得られた化合物３を使用する他は同様にしてチューインガムを製造した。
【００６３】
応用例５
応用例３の処方で、化合物２の代わりに実施例３で得られた化合物４を使用する他は同様にしてキャンディを製造した。
【００６４】
応用例６
応用例２の処方で、化合物１の代わりに実施例３で得られた化合物５を使用する他は同様にしてチューインガムを製造した。
【００６５】
応用例７
応用例２の処方で、化合物１の代わりに実施例３で得られた化合物６を使用する他は同様にしてチューインガムを製造した。
【００６６】
応用例８
応用例３の処方で、化合物２の代わりに実施例３で得られた化合物７を使用する他は同様にしてキャンディを製造した。
【００６７】
応用例９
応用例２の処方で、化合物１の代わりに実施例３で得られた化合物８を使用する他は同様にしてチューインガムを製造した。
【００６８】
応用例１０
応用例２の処方で、化合物１の代わりに実施例３で得られた化合物９を使用する他は同様にしてチューインガムを製造した。
【００６９】
【発明の効果】
本発明製剤は、喉炎症起因菌である溶血性連鎖球菌に対する強い成育阻害効果を有するばかりでなく、本菌種の産生する溶血毒に対する阻害効果を併せ持つものであり、喉の炎症および化膿性疾患をはじめ種々の感染症に対して有効に作用する口腔用組成物として有用である。
【００７０】
本発明製剤の原料となるユーカリはハーブティや香料成分として用いられているものであり、その安全性については問題なく、抗生物質を使用した場合にみられる副作用等の恐れがない。[0001]
[Industrial applications]
TECHNICAL FIELD The present invention relates to a prophylactic / therapeutic agent for throat inflammation and hemolytic toxin derived from a plant of the genus Eucalyptus, and an oral composition containing the same.
[0002]
[Prior art]
Streptococcus pyogenes (Streptococcus pyogenes) is a facultative anaerobic gram-positive cocci distributed in the human oral cavity, pharynx, and the like. This bacterium is the most pathogenic among streptococci, causing pharyngitis, tonsillitis, otitis media, etc. on mucous membranes, and causing various purulent inflammations on skin, all of which are pharyngeal or tonsils. Proliferate in
[0003]
Hemolytic toxin (streptolysin O), Dick toxin, streptokinase, DNA degrading enzyme, hyaluronidase, etc. are known as virulence factors of this bacterium, and the etiology of respiratory diseases such as pharyngitis, tonsillitis, bronchitis, etc. Play a role.
[0004]
In order to prevent and treat these respiratory diseases, it is important to inhibit the growth of the fungus in the lesion and to suppress the pathogenic factor of the fungus. From the viewpoint of controlling bacteria, it is effective to apply an antibacterial substance effective against the fungus.
[0005]
Conventionally, penicillin, cephem, and cephalosporin antibiotics have been used as antibacterial substances against this fungus.However, these are powerful, but they are routinely used due to the emergence of resistant bacteria and the appearance of side effects. It cannot be said that it is suitable for various uses.
[0006]
The present inventors obtained a non-antibiotic substance having an antibacterial activity against the present bacterium and a hemolytic toxin inhibitory activity produced by the present bacterium, and performed screening for extracts of various natural products. We focused on extracts other than the essential oil (eucalyptus oil). Eucalyptus is used as a herbal tea or a fragrance ingredient, and there is no problem with its safety, and there is no problem of side effects as in the case of applying antibiotics.
[0007]
Until now, the effects of eucalyptus extract have been known to enhance the effect of eucalyptus oil and its main component, cineol (eucalyptol), as an antiseptic and to enhance the action of antibacterial active substances. JP-A 62-289511, antibacterial oral composition). However, the antibacterial activity of eucalyptus oil and cineole itself against hemolytic streptococci is not only weak, but also has a drawback that, due to its strong and unique odor, for example, when used in the oral cavity, the use and the amount of addition are considerably restricted.
[0008]
JP-A-58-39615 (toothpaste composition) discloses that the combined use of a eucalyptus extract and sodium alkylsulfate exhibits antibacterial activity against Fusobacterium nucleitam, one of the periodontal disease-causing bacteria. ing. This eucalyptus extract expresses antibacterial activity only when used in combination with sodium alkylsulfate, suggesting that the eucalyptus extract itself has no antibacterial activity or is very weak, if at all. Moreover, this composition does not show any effect on hemolytic streptococci.
[0009]
Further, Japanese Patent Application Laid-Open No. 5-306252 (New macrocarpals and a method for producing the same) discloses that macrocarpals collected from a solvent extract of Eucalyptus plants exhibit aldose reductase inhibitory activity and antibacterial activity. However, its antibacterial activity has only been shown to act against some Gram-positive bacteria, and no antibacterial activity against hemolytic streptococci has been shown.
[0010]
On the other hand, in Japanese Patent Application Laid-Open No. 62-59215 (external coughing, exudation, analgesia, sedative), eucalyptus oil is further added to a cream containing dl-camphor, 1-menthol, nutmeg oil and fennel oil. It discloses that it enhances the antitussive and bodily effects when used externally. In this gazette, not only eucalyptus oil, which has a weak antibacterial activity against hemolytic streptococci, is used as an active ingredient, but it is also expected that the action of volatile components is expected by a dosage form of external use, and it is used in a non-volatile manner in the oral cavity. It is fundamentally different from the antibacterial active agent against hemolytic streptococci that is required to be used.
[0011]
As described above, various Eucalyptus extracts are disclosed, but all of them are insufficient for prevention and treatment of throat inflammation and hemolytic toxin and have poor versatility.
[0012]
[Problems to be solved by the invention]
Accordingly, the present inventors have conducted intensive research on extracting effective components for prevention and treatment of both throat inflammation and hemolytic toxin from Eucalyptus plants. As a result, the residue obtained by removing the essential oil component (eucalyptus oil) from the Eucalyptus plants was removed. The extract extracted with a polar solvent and the phloroglucinol derivative contained in it have a very strong antibacterial activity against throat inflammation-causing bacteria (hemolytic streptococci), and also inhibit the hemolytic toxins produced by this species The present inventors have also found that they also have an activity, and have completed the present invention.
[0013]
The phloroglucinol derivative contained in the plant of the genus Eucalyptus has an antiviral effect [Chem. Pharm. Bull. , 38 (5) 1444-1446 (1990)], HIV reverse transcriptase inhibitory activity [Tetrahedron Lett. , 33 (21) 2983-2986 (1992)], but the preventive and therapeutic effects of throat inflammation and hemolytic toxin in the present invention are not known, and those first revealed by the present invention. It is.
[0014]
[Means for Solving the Problems]
The present invention relates to an extract obtained by treating a residue obtained by removing an essential oil component (eucalyptus oil) from a plant of the genus Eucalyptus with a polar solvent and a throat inflammation / hemolytic toxin containing as an active ingredient a phloroglucinol derivative obtained as an active substance from the extract. It is a prophylactic / therapeutic agent and an oral composition using the same.
[0015]
Hereinafter, an example of extraction and production of the preparation of the present invention will be described.
[0016]
First, a eucalyptus plant, for example, eucalyptus leaves, is pulverized by a suitable pulverizing means such as a pulverizer, and the essential oil component is removed preferably using a steam distillation apparatus usually used for essential oil extraction. The removal of the essential oil component can be carried out at room temperature using a low-polarity solvent such as n-hexane or petroleum ether.
[0017]
Extraction is performed by applying at least one of polar solvents such as ethyl acetate, acetone, ethanol, methanol and water to the thus obtained essential oil extraction residue, and concentrating the obtained extract under reduced pressure. The preparation of the present invention is obtained in the form of a product.
[0018]
The preparation thus obtained is further separated and purified using a suitable separation and purification means such as column chromatography to obtain a phloroglucinol derivative which is the preparation of the present invention in the form of a compound.
[0019]
This phloroglucinol derivative has the general formula:
[0020]
Specific examples of the phloroglucinol derivative used as the preparation of the present invention include those having the following structural formula.
[0021]
Embedded image
Embedded image
Embedded image
Embedded image
Embedded image
Embedded image
Embedded image
Embedded image
Embedded image
In addition, examples of column chromatography used for separation and purification of phloroglucinol derivatives include adsorption chromatography, partition chromatography, high-performance liquid chromatography, and thin-layer chromatography.
[0031]
Although various uses of the preparation of the present invention are conceivable, it can be used daily for chewing gum, candy, confectionery, mouthwash, etc. because of its excellent aroma and taste and high safety. . When the preparation of the present invention is incorporated into foods and the like, it is preferable to add the preparation so as to have a ratio of 0.001 to 10% by weight.
[0032]
Hereinafter, the present invention will be described with reference to examples, test examples, and application examples, but these examples do not limit the scope of the present invention.
[0033]
【Example】
Example 1
After removing essential oil components from 1 kg of eucalyptus leaf powder using a steam distillation apparatus, 5 liters of ethanol was added, and the mixture was subjected to heating and reflux extraction for 3 hours. The obtained filtrate was concentrated under reduced pressure to obtain 214 g of an extract of the present invention.
[0034]
Example 2
After removing essential oil components from 1 kg of eucalyptus leaf powder using a steam distillation apparatus, 5 liters of a mixed solution of ethanol-water (1: 1) was added, and the mixture was heated and refluxed for 3 hours. The obtained filtrate was concentrated under reduced pressure to obtain 234 g of an extract of the present invention.
[0035]
Example 3
After 100 g of the extract obtained in Example 2 was dispersed by adding water, the mixture was partitioned with ethyl acetate to obtain an ethyl acetate-soluble fraction (29.5 g). This ethyl acetate-soluble fraction was adsorbed on a column packed with 1 kg of silica gel (Wako Gel C-200, Wako Pure Chemical Industries, Ltd.).
[0036]
The column was eluted with a dichloromethane-methanol (50: 1 to 2: 1) mixture to fractionate. When developed with a dichloromethane-methanol (3: 1) mixed solution using a TLC plate (60F254, Merck), UV absorption is observed at an Rf value of about 0.2 to 0.4, and purple, gray or brown with 10% sulfuric acid. Fractions showing spots were collected. This fraction was eluted with methanol-water (90:10 to 100: 0) using an octadecylsilanol type (ODS) preparative column (HPLC) for high performance liquid chromatography (HPLC), and the UV absorption at 285 nm was obtained. Each peak was collected while monitoring the temperature.
[0037]
The respective peaks were further analyzed using dichloromethane-methanol (98: 2 to 85:15) using a silica gel preparative column for HPLC (senshu science), and further analyzed using acetonitrile-water (90%) using an ODS preparative column for HPLC. : 10 to 100: 0) and methanol-water (90:10 to 100: 0) to obtain Compounds 1 to 9 of the present invention.
[0038]
Incidentally, the physicochemical properties of Compounds 5 to 9 are described below.
(Compound 5)
1) Property: slightly yellow powder 2) Melting point: not measured because it is non-crystalline.
3) Molecular formula: C 28 H 38 O 7
4) FAB-mass spectrum: 487 [M + 1]
(Equation 1)
Embedded image
(Compound 6)
1) Property: slightly yellow powder 2) Melting point: not measured because it is non-crystalline.
3) Molecular formula: C 28 H 40 O 6
4) EI-mass spectrum: 472 [M], 454, 251, 195
[Equation 3]
Embedded image
(Compound 7)
1) Property: slightly yellow powder 2) Melting point: not measured because it is non-crystalline.
3) Molecular formula: C 28 H 42 O 7
4) EI- Mass spectrum: 472 [M-H 2 O ], 454,251,195
(Equation 5)
Embedded image
(Compound 8)
1) Property: slightly yellow powder 2) Melting point: not measured because it is non-crystalline.
3) Molecular formula: C 28 H 42 O 7
4) EI- Mass spectrum: 472 [M-H 2 O ], 454,251,195
(Equation 7)
Embedded image
(Compound 9)
1) Property: slightly yellow powder 2) Melting point: not measured because it is non-crystalline.
3) Molecular formula: C 28 H 38 O 7
4) EI-mass spectrum: 468 [MH 2 O], 427, 251, 217, 195
(Equation 9)
Test example 1
The antibacterial activities of the preparations of the present invention prepared in Examples 1 to 3 against hemolytic streptococci and intestinal bacteria were tested by the following methods.
[0044]
The antimicrobial activity of the preparation of the present invention, eucalyptus oil and thymol known as a bactericide were compared with the preparation of the present invention.
[0045]
(Test method)
1) Test strain, culture medium and culture conditions
[0046]
After culturing for 24-48 hours, the turbidity (550 nm) of each well is measured using a microplate reader to determine the growth of the bacterium, and its minimum inhibitory concentration (MIC) is determined.
[0047]
(Test results)
Table 1 shows the test results. The preparation of the present invention showed strong antibacterial activity against hemolytic streptococci. Thymol and eucalyptus oils were significantly inferior to the antimicrobial activity of the preparation of the present invention. Furthermore, the antimicrobial activity of the preparation of the present invention against enteric bacteria was considerably weaker than that of hemolytic streptococci, and it was considered that the preparation of the present invention had little effect on enteric bacteria.
[0048]
[Table 1]
Test example 2
The hemolytic toxin (streptolysin O) inhibitory activity of the preparations of the present invention prepared in Examples 1 to 3 was tested by the following method.
[0050]
(Test method)
Rabbit defibrinated blood (25 ml) is washed three times with buffered saline (pH 6.5) (3000 rpm × 10 minutes) to collect red blood cells. Dilute to 200 ml with a buffered saline to obtain a rabbit erythrocyte suspension. A prescribed amount of the sample is dissolved in dimethyl sulfoxide (DMSO), and then further dissolved in buffered saline (DMSO 4%) to obtain a sample solution.
[0051]
600 μl of the sample solution and 300 μl of an aqueous solution of streptolysin O (Eiken) (1 binding titer) are mixed in a 13 mm diameter test tube, and reacted at 37 ° C. for 30 minutes. Rabbit erythrocyte suspension (300 μl) is added and reacted at 37 ° C. for 45 minutes. After the completion of the reaction, the mixture is centrifuged at 1500 rpm for 1 minute, and the degree of hemolysis of the supernatant is visually determined.
[0052]
Judgment criteria-: Only hemolysis is possible, +: Incomplete hemolysis, ++: Complete hemolysis.
[0053]
(Test results)
Table 2 shows the test results. Most of the preparations of the present invention completely inhibited hemolysis by streptolysin O at 1000 μg / ml, and exhibited inhibitory activity even at 500 μg / ml.
[0054]
[Table 2]
Application example 1
Using the preparations of the present invention prepared by the methods of Examples 1 and 2, chewing gum, candy, tablet confectionery and mouthwash were prepared according to the following formulation.
[0056]
1. Chewing gum prescription gum base 20.0%
Sugar 55.0
Glucose 15.0
Candy 9.3
Fragrance 0.5
Formulation of the Present Invention (Example 1) 0.2
100.0%
[0057]
2. Candy prescription sugar 50.0%
Syrup 34.0
Citric acid 1.0
Formulation of the Present Invention (Example 1) 0.2
Water 14.8
100.0%
[0058]
3. Prescription sugar for tablet confections 76.4%
Glucose 19.0
Sucrose fatty acid ester 0.2
Formulation of the Present Invention (Example 2) 0.2
Water 4.2
100.0%
[0059]
4. Prescription of mouthwash Ethanol 30.0%
Spice 1.0
Copper chlorophyllin sodium 0.1
Saccharin 0.05
Chlorhexidine hydrochloride 0.01
Formulation of the Present Invention (Example 2) 0.2
Water 68.64
100.0%
[0060]
Application example 2
A chewing gum is produced in the same manner as in Application Example 1 except that the compound 1 obtained in Example 3 is replaced by 0.005% and sugar is 55.195% in place of the preparation of the present invention. did.
[0061]
Application example 3
A candy was produced in the same manner as in Application Example 1 except that the composition of the present invention of Example 1 was replaced with 8.2% of compound 2 and 42% of sugar instead of the preparation of the present invention.
[0062]
Application example 4
A chewing gum was produced in the same manner as in Application Example 2, except that Compound 3 obtained in Example 3 was used instead of Compound 1.
[0063]
Application example 5
A candy was produced in the same manner as in Application Example 3, except that Compound 4 obtained in Example 3 was used instead of Compound 2.
[0064]
Application example 6
A chewing gum was produced in the same manner as in Application Example 2, except that Compound 5 obtained in Example 3 was used instead of Compound 1.
[0065]
Application example 7
A chewing gum was produced in the same manner as in Application Example 2, except that Compound 6 obtained in Example 3 was used instead of Compound 1.
[0066]
Application example 8
A candy was produced in the same manner as in Application Example 3, except that Compound 7 obtained in Example 3 was used instead of Compound 2.
[0067]
Application example 9
A chewing gum was produced in the same manner as in Application Example 2, except that Compound 8 obtained in Example 3 was used instead of Compound 1.
[0068]
Application example 10
A chewing gum was produced in the same manner as in Application Example 2, except that Compound 9 obtained in Example 3 was used instead of Compound 1.
[0069]
【The invention's effect】
The preparation of the present invention not only has a strong growth inhibitory effect against hemolytic streptococci, which is a throat inflammation-causing bacterium, but also has an inhibitory effect on the hemolytic toxin produced by this strain of the present invention. And is useful as an oral composition that effectively acts against various infectious diseases.
[0070]
Eucalyptus, which is a raw material of the preparation of the present invention, is used as a herbal tea or a fragrance ingredient, and there is no problem in its safety, and there is no fear of side effects or the like seen when an antibiotic is used.
Claims (2)
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP06794395A JP3547835B2 (en) | 1995-03-27 | 1995-03-27 | Prophylactic / therapeutic agent for throat inflammation and hemolytic toxin and oral composition containing the same |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP06794395A JP3547835B2 (en) | 1995-03-27 | 1995-03-27 | Prophylactic / therapeutic agent for throat inflammation and hemolytic toxin and oral composition containing the same |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPH08259452A JPH08259452A (en) | 1996-10-08 |
| JP3547835B2 true JP3547835B2 (en) | 2004-07-28 |
Family
ID=13359530
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP06794395A Expired - Lifetime JP3547835B2 (en) | 1995-03-27 | 1995-03-27 | Prophylactic / therapeutic agent for throat inflammation and hemolytic toxin and oral composition containing the same |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JP3547835B2 (en) |
Families Citing this family (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1998013055A1 (en) * | 1996-09-27 | 1998-04-02 | Takeshi Karita | Antioxidizing composition for scavenging free radicals, pharmaceutical composition comprising the same, and process for preparing the same |
| US6159473A (en) * | 1998-06-24 | 2000-12-12 | Botanical Laboratories, Inc. | Sore throat spray |
| DE60139795D1 (en) * | 2000-10-18 | 2009-10-15 | Auris Ehf | DISPENSING SYSTEM AND METHOD FOR TREATING OR PREVENTING MEDIUM EARLY IGNITION |
| ITMI20032287A1 (en) * | 2003-11-24 | 2005-05-25 | Indena Spa | COMPOSITIONS FOR THE TREATMENT OF THE ORAL CABLE AFFECTIONS AND THE FIRST RESPIRATORY ROUTES |
| AU2006331925A1 (en) * | 2005-12-16 | 2007-07-05 | Bakto Natural Preservatives, Llc | Recovery of residual plant components after distillation of essential oils |
| JP2009209065A (en) * | 2008-03-03 | 2009-09-17 | Lotte Co Ltd | Chewing gum blended with eucalyptus extract |
| JP5602346B2 (en) * | 2008-06-17 | 2014-10-08 | 株式会社ロッテ | Preparation method of eucalyptus extract |
| JP2010254603A (en) * | 2009-04-23 | 2010-11-11 | Osaka Univ | Oral cavity composition |
| CN104645058A (en) * | 2015-03-19 | 2015-05-27 | 许兰兰 | Medicament for treating children acute suppurative tonsillitis in paediatric internal medicine center |
-
1995
- 1995-03-27 JP JP06794395A patent/JP3547835B2/en not_active Expired - Lifetime
Also Published As
| Publication number | Publication date |
|---|---|
| JPH08259452A (en) | 1996-10-08 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Policegoudra et al. | Antimicrobial, antioxidant, cytotoxicity and platelet aggregation inhibitory activity of a novel molecule isolated and characterized from mango ginger (Curcuma amada Roxb.) rhizome | |
| US7214392B2 (en) | Process and composition for inhibiting growth of microorganisms | |
| CN104114176A (en) | Maillard reaction inhibitor | |
| KR101676043B1 (en) | Process for preparing a eucalyptus extract | |
| JP2009501708A (en) | Korean thistle extract, its use and formulations containing it | |
| KR102026407B1 (en) | A composition for antioxidating comprising extracts of fermented tenebrio molitor | |
| US20110059193A1 (en) | Method for obtaining an extract of cranberry marc that can be used in particular in the prevention and treatment of conditions such as caries, gingivitis or sore throats | |
| JP3547835B2 (en) | Prophylactic / therapeutic agent for throat inflammation and hemolytic toxin and oral composition containing the same | |
| JP2804232B2 (en) | Anti-caries, periodontal agent and oral composition containing it | |
| KR20180065799A (en) | A composition for anti-bacterial effect and anti-inflammation comprising unripe apple extracts and baicalin | |
| JP3749743B2 (en) | Antibacterial activity against throat inflammation-causing bacteria and hemolytic toxin inhibitor, and oral composition and food and drink containing the same | |
| JP4873805B2 (en) | Anti-periodontal agent | |
| WO2007020830A1 (en) | Oral composition | |
| Sharma et al. | Role of plant extract in the inhibition of dental caries | |
| EP4403180A1 (en) | Antibacterial and antioxidant composition containing grape callus culture medium as active ingredient | |
| JP4457035B2 (en) | Oral composition for antibacterial against throat inflammation-causing bacteria (hemolytic streptococci) and for inhibiting hemolytic toxins produced by this species | |
| JP5600067B2 (en) | Anti-Helicobacter pylori activator | |
| Alemeye et al. | Antimicrobial activities and formulations of the extracts of chewing sticks commonly used in Ethiopia for oral cleansing | |
| KR20070047743A (en) | Antimicrobial composition which comprising the hibiscus ethanol extract | |
| KR100967541B1 (en) | Antibacterial pharmaceutical composition comprising Aceriphyllum rossii extract and active compounds isolated from them | |
| US20100179227A1 (en) | Production method of hop preparation, hop preparation, anti-inflammatory agent, food product and beverage, and oral product | |
| JP2772552B2 (en) | Preventive agent for dental caries and periodontal disease | |
| JP7577351B2 (en) | Novel ellagitannins and oral preparations | |
| JP4515732B2 (en) | Glucosyltransferase inhibitors, plaque formation inhibitors, antibacterial agents, oral preparations and foods for preventing touch | |
| KR102274808B1 (en) | Composition for eradicating Korean helicobacter pylori comprising compounds from fruits of Cudrania tricuspidata |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| A521 | Written amendment |
Free format text: JAPANESE INTERMEDIATE CODE: A523 Effective date: 20031224 |
|
| A521 | Written amendment |
Free format text: JAPANESE INTERMEDIATE CODE: A523 Effective date: 20031225 |
|
| A131 | Notification of reasons for refusal |
Free format text: JAPANESE INTERMEDIATE CODE: A131 Effective date: 20040129 |
|
| A521 | Written amendment |
Free format text: JAPANESE INTERMEDIATE CODE: A523 Effective date: 20040309 |
|
| TRDD | Decision of grant or rejection written | ||
| A01 | Written decision to grant a patent or to grant a registration (utility model) |
Free format text: JAPANESE INTERMEDIATE CODE: A01 Effective date: 20040401 |
|
| A61 | First payment of annual fees (during grant procedure) |
Free format text: JAPANESE INTERMEDIATE CODE: A61 Effective date: 20040415 |
|
| R150 | Certificate of patent or registration of utility model |
Free format text: JAPANESE INTERMEDIATE CODE: R150 |
|
| R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
| FPAY | Renewal fee payment (event date is renewal date of database) |
Free format text: PAYMENT UNTIL: 20100423 Year of fee payment: 6 |
|
| S111 | Request for change of ownership or part of ownership |
Free format text: JAPANESE INTERMEDIATE CODE: R313111 |
|
| FPAY | Renewal fee payment (event date is renewal date of database) |
Free format text: PAYMENT UNTIL: 20100423 Year of fee payment: 6 |
|
| R350 | Written notification of registration of transfer |
Free format text: JAPANESE INTERMEDIATE CODE: R350 |
|
| FPAY | Renewal fee payment (event date is renewal date of database) |
Free format text: PAYMENT UNTIL: 20130423 Year of fee payment: 9 |
|
| FPAY | Renewal fee payment (event date is renewal date of database) |
Free format text: PAYMENT UNTIL: 20130423 Year of fee payment: 9 |
|
| FPAY | Renewal fee payment (event date is renewal date of database) |
Free format text: PAYMENT UNTIL: 20150423 Year of fee payment: 11 |
|
| EXPY | Cancellation because of completion of term |