JP3161882B2 - Ginseng-derived polysaccharide, its production method and use - Google Patents
Ginseng-derived polysaccharide, its production method and useInfo
- Publication number
- JP3161882B2 JP3161882B2 JP19927593A JP19927593A JP3161882B2 JP 3161882 B2 JP3161882 B2 JP 3161882B2 JP 19927593 A JP19927593 A JP 19927593A JP 19927593 A JP19927593 A JP 19927593A JP 3161882 B2 JP3161882 B2 JP 3161882B2
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- ginseng
- polysaccharide
- extracted
- water
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- Medicines Containing Plant Substances (AREA)
- Polysaccharides And Polysaccharide Derivatives (AREA)
Description
【発明の詳細な説明】DETAILED DESCRIPTION OF THE INVENTION
【0001】[0001]
【産業上の利用分野】本発明は、老人はもとより近年若
年層で増加傾向にある腎臓疾患、とりわけ慢性腎不全に
至る前段階であるネフローゼ症候群と診断された患者に
対し、あるいはウイルス性及び薬物性肝炎等の肝障害の
症状を有する患者に対し、通院可能で安全な経口及び筋
肉内投与治療剤を提供する発明に関する。The present invention is intended for patients diagnosed with nephrotic syndrome, which is an early stage leading to chronic renal failure, especially kidney disease, which is increasing in young people as well as elderly people, or for viral and drug use. The present invention relates to an invention that provides a safe and therapeutic agent for oral and intramuscular administration that can be visited in a patient with symptoms of liver disorder such as hereditary hepatitis.
【0002】[0002]
【従来の技術】丹参は中国では、血液循環,血行停止の
改善に良いとされ伝統的に使用されている薬草で、近
年、血管拡張作用(大浦彦吉:和漢医薬学会誌.,5:
227−237.,1988.)、降圧作用(中山医学
院論:医歯薬出版,東京.,257−258.,198
0.)、抗凝血作用(羅厚蔚ら:Acta Pharm
aceutica Sinica.,23(11):8
30−834.,1988.)、細胞内コレステロール
の合成抑制作用(Sun Xi−ming;中草薬.,
22(1):20−23.,1991.)、肝細胞保護
(戚心広ら:中西医結合雑誌.,11(2):102−
104.,1991.)及びラジカル消去作用(胡 天
喜:上海中医薬雑誌,9:28−30.,1988.)
などの多彩な生理活性作用があること、また腎機能の改
善(大浦彦吉:第二回和漢薬の医学薬学的研究に関する
日中シンポジウムpp.148−157,198
8.)、慢性腎疾患及び***の患者に対して有効(張
鏡人:上海中医薬雑誌.,1.17−18.,198
1.)であることが内外において報告されているが、そ
の本質については何等言及されていないか、言及されて
いても本願発明の物質とは全く異なる低分子物質であ
る。BACKGROUND OF THE INVENTION Dansin is a herb that has been traditionally used in China for improving blood circulation and blood circulation arrest, and has recently been shown to have a vasodilator effect (Hikichi Ohura: Journal of the Wakan Pharmaceutical Society., 5:
227-237. 1988. ), Antihypertensive effect (Nakayama Medical School theory: Medical and Dental Publishing, Tokyo., 257-258., 198)
0. ), Anticoagulant effect (Ra Thao Ul et al .: Acta Pharm)
aceutica Sinica. , 23 (11): 8
30-834. 1988. ), Inhibition of intracellular cholesterol synthesis (Sun Xi-ming; herbal medicine.,
22 (1): 20-23. 1991. ), Protection of hepatocytes (Rinshin Hiroshi: Nakanishi Medical Combined Magazine., 11 (2): 102-)
104. 1991. ) And radical scavenging action (Tianki Hu: Shanghai China Pharmaceutical Magazine, 9: 28-30, 1988.)
And various other physiologically active activities, and improvement of renal function (Oura Hikichi: 2nd Japan-China Symposium on Medical and Pharmaceutical Research on Wakan Yakuhin pp. 148-157, 198)
8. ), Effective for patients with chronic kidney disease and uremic disease (Zhang Jingjin: Shanghai China Pharmaceutical Magazine, 1.17-18., 198)
1. ) Is reported internally and externally, but the substance is not mentioned at all, or even if it is mentioned, it is a low molecular substance completely different from the substance of the present invention.
【0003】微小変化型ネフローゼの内科的な治療薬と
して、合成薬であるステロイド剤或いは抗血小板薬であ
るジピリダモールが用いられているが、長期間使用しな
くてはならず、若年層に投薬する際には特に満月様顔
貌,月経不順,めまい,頭痛,悪心,嘔吐で、重篤な場
合には感染症,消化管出血,代謝異常,骨粗症,血栓
症,副腎不全,精神障害などの副作用が懸念されてい
る。[0003] Steroid drugs as synthetic drugs or dipyridamole as antiplatelet drugs have been used as medical treatments for minimal change nephrosis, but they must be used for a long time and are administered to young people. In some cases, especially full moon-like face, irregular menstruation, dizziness, headache, nausea, vomiting, and in severe cases, such as infection, gastrointestinal bleeding, metabolic abnormalities, osteoporosis, thrombosis, adrenal insufficiency, mental disorders, etc. Side effects are a concern.
【0004】本発明と同じ原料からの腎疾患への治療薬
の研究は盛んに行われているが、低分子のフェナントレ
ンキノン類であるタンシノン類似物並びにカフェイン酸
4量体であるマグネシウムリトスペルミン酸B等が単離
されているが(T.Yoozawa,H.Y.Chun
g,H.Oura,G.Nonaka,and I.N
ishioka,Chem.Pharm.Bull.,
36,316(1988))、何れも有機溶媒含有下で
抽出される脂溶性の高い物質だけであり、まだ臨床応用
には至っていない。[0004] Although active research has been conducted on remedies for renal diseases from the same raw materials as in the present invention, tanshinone analogs, which are low-molecular phenanthrenequinones, and magnesium lithospermine, which is a tetramer of caffeic acid Acid B and the like have been isolated (T. Yozawa, HY.
g, H .; Oura, G .; Nonaka, and I .; N
isioka, Chem. Pharm. Bull. ,
36, 316 (1988)), all of which are only fat-soluble substances extracted in the presence of an organic solvent, and have not yet reached clinical application.
【0005】また、肝炎にはウイルス性、薬物性のもの
もあるがウイルス性のものが大半を占めており、それら
の治療薬としてはインターフェロン、インターロイキン
2のようなBiological response
modifier(BRM)、グリチルリチンの靜注用
注射剤であるネオミノファーゲンCなどが用いられてい
る。しかしながらBRMには発熱等の副作用が見られる
ことにより長期投与には問題がある。またグリチルリチ
ンは基本的には抗炎症剤であり、長期に使用する場合は
血圧、電解質濃度の管理が必要となる。この他にも生薬
成分であるサイコサポニン、サポニン、ゴミシンなどが
知られているが、いずれも臨床応用されるには至ってい
ない。[0005] Hepatitis includes viral and drug-related hepatitis, but most of them are viral. Therapeutic agents include biologic response such as interferon and interleukin-2.
Modifier (BRM), neominophagen C, which is an intravenous injection of glycyrrhizin, and the like are used. However, there is a problem with long-term administration of BRM due to side effects such as fever. Glycyrrhizin is basically an anti-inflammatory agent, and when used for a long period of time, it is necessary to control blood pressure and electrolyte concentration. In addition, saikosaponin, saponin, gomisin and the like, which are crude drug components, are known, but none of them has been applied to clinical applications.
【0006】[0006]
【発明が解決しようとする課題】腎疾患は大別して、糸
球体由来の原発性とそれ以外の疾患に由来する続発性に
分類され、前者が約80%以上を占め、低年齢層ほど多
い(約90%)15) 。病理組織所見からは微小変化群,
単状糸球体硬化症,膜性腎症,メサンギゥム増殖性腎症
(IgA腎症,非IgA腎症),膜性増殖糸球体腎炎,
半月体形成糸球体腎炎などに分類される。蛋白尿は臨床
所見の特徴として、いずれの組織学的病型にも共通して
認められ、重篤な場合にはネフローゼ症候群として診断
されている16) 。また近年腎臓疾患、特に腎臓の器質的
な衰えにより誘発される疾患は老人にとどまらず若年層
に増加しつつあるため、器質の改善から疾患を治療する
副作用発現の少ない生薬,漢方薬成分で水溶性のある経
口或いは筋肉内に投薬し治療する薬剤の開発が望まれて
いる。[0006] Kidney diseases are roughly classified into primary glomerular-derived diseases and secondary diseases derived from other diseases, with the former accounting for about 80% or more, and are more common in lower age groups ( About 90%) 15) . From the histopathological findings, a group of small changes,
Monoglomerular sclerosis, membranous nephropathy, mesangial proliferative nephropathy (IgA nephropathy, non-IgA nephropathy), membranous proliferative glomerulonephritis,
It is classified as crescent-forming glomerulonephritis. Proteinuria is a common clinical feature of all histological types, and is diagnosed as nephrotic syndrome in severe cases16 ) . In recent years, renal diseases, especially those induced by the organic deterioration of the kidney, are increasing not only in the elderly but also in younger people. There is a need for the development of a drug that can be administered orally or intramuscularly.
【0007】また、肝炎は大別して、ウイルス性のもの
とアルコール等の薬物によるものがあるが、B型及びC
型肝炎ウイルスによるものが圧倒的に多い。近年治療薬
として特にインターフェロンがC型肝炎に対して有効で
あることが報告されており、使用頻度も高まってきてい
る(飯野司郎ら:基礎と臨床.26:339,199
2;鈴木宏ら:肝・胆・膵.23:1065,199
1)。しかしながら、短期投与で発熱等のインフルエン
ザ様症状が表れ、長期投与でも体重減少、下痢、嘔吐、
不整脈、脱毛、自己免疫異常等の副作用が見られる。ま
たこれらの薬剤はすべて注射剤であることからより簡便
に使用できる新たな治療薬あるいはインターフェロン等
との併用補助薬として副作用発現の少ない生薬、漢方薬
成分で水溶性のある経口或いは筋肉内に投与し治療する
薬剤の開発が望まれている。[0007] Hepatitis can be roughly classified into two types: viral ones and drugs such as alcohol.
Mostly due to hepatitis virus. In recent years, interferon has been reported to be particularly effective as a therapeutic agent against hepatitis C, and its use has been increasing (Shiro Iino et al .: Basic and Clinical. 26: 339, 199).
2: Hiroshi Suzuki et al .: Liver, gall, and pancreas. 23: 1065, 199
1). However, short-term administration showed flu-like symptoms such as fever, and long-term administration resulted in weight loss, diarrhea, vomiting,
Side effects such as arrhythmia, hair loss, and autoimmune abnormalities are observed. In addition, since all of these drugs are injections, they can be used as a new therapeutic agent that can be used more easily or as a supplementary drug combined with interferon, etc. There is a need for the development of therapeutic drugs.
【0008】[0008]
【課題を解決するための手段】本発明者らは、広く繁用
されている蛋白尿***を顕著に起こし、さらに腎臓の器
質変化を示すことで良く知られるピューロマイシンで惹
起したラットのネフローゼ症候群モデルおよび肝障害モ
デルを利用して鋭意検討した結果、丹参の水抽出物より
上記モデル対して有効な多糖類を見いだした。DISCLOSURE OF THE INVENTION The present inventors have developed a nephrotic syndrome in rats induced by puromycin, which is prominent in the widely used proteinuria excretion and is also well known to show renal organic changes. As a result of intensive studies using a model and a liver injury model, a polysaccharide effective for the above model was found from a water extract of Danshen.
【0009】本発明はこの知見に基づくもので、 (I)丹参から水または水性溶媒で抽出されうる下記の
特性を有する多糖類 A.糖含量:60〜100% (1)糖組成:ウロン酸 40〜80% 中性糖 10〜30% (2)中性糖組成:ラムノース 0〜15% グルコース 0〜15% ガラクトース 25〜55% アラビノース 30〜60% マンノース 0〜15% B.分子量 150,000〜300,000 (II)丹参を水または水性溶媒で抽出し、抽出液から残
渣を除去した液を無極性多孔性ポリマー樹脂を通過さ
せ、限外濾過で濃縮し、ついで限外濾過クロマトグラフ
ィーに付することを特徴とする上記の多糖類の製造法お
よび (III)上記の多糖類よりなるネフローゼ症候群の寛解剤
である。The present invention is based on this finding, and (I) a polysaccharide having the following properties, which can be extracted from water ginseng with water or an aqueous solvent. Sugar content: 60-100% (1) Sugar composition: uronic acid 40-80% Neutral sugar 10-30% (2) Neutral sugar composition: rhamnose 0-15% glucose 0-15% galactose 25-55% arabinose 30-60% Mannose 0-15% B. Molecular weight 150,000-300,000 (II) Dansin is extracted with water or an aqueous solvent, and the liquid obtained by removing the residue from the extract is passed through a non-polar porous polymer resin, concentrated by ultrafiltration, and then concentrated by ultrafiltration. A method for producing the above polysaccharide, which is characterized by subjecting the polysaccharide to filtration chromatography, and (III) an agent for ameliorating nephrotic syndrome comprising the above polysaccharide.
【0010】丹参は植物丹参(Salvia milt
iorrhiza Bunge)の根および根状の茎で
あるが、同属植物の南丹参(S.bowleyana
Dunn)、甘粛丹参(S.przewalskii
Maxim)、雲南丹参(S.yunnanensis
C.H.Wright)等の根も用いられ、本発明で
はこれらを総称して丹参という。The ginseng is a plant ginseng (Salvia milt)
iorrhiza Bunge's roots and root-like stems, but a congener plant, S. bowleyana.
Dunn), Gansu Dansin (S.przewalskii)
Maxim), Yunnan Dansin (S. yunnanensis)
C. H. The roots such as Wright are also used. In the present invention, these are collectively referred to as Dansin.
【0011】本発明で用いられる丹参の産地は特に限定
されるものではないが、好ましくは中国,四川地区産の
丹参が本発明の多糖類を効率よく抽出精製することがで
きる。The place of production of ginseng used in the present invention is not particularly limited, but preferably ginseng from Sichuan, China can efficiently extract and purify the polysaccharide of the present invention.
【0012】丹参はできるだけ小片にして抽出するのが
望ましい。抽出溶媒として水または水性溶媒が用いら
れ、水性溶媒としてはリン酸緩衝液、酢酸緩衝液等の緩
衝液または食塩等の塩類水溶液を用いるのが好ましい。It is desirable that the ginseng be extracted as small pieces as possible. As the extraction solvent, water or an aqueous solvent is used, and as the aqueous solvent, a buffer such as a phosphate buffer or an acetate buffer or an aqueous salt solution such as a salt is preferably used.
【0013】抽出溶媒のpHは約4〜約9に調整するの
が好ましい。抽出は約50〜約100℃で行うのが好ま
しく、さらに好ましくは70〜90℃である。たとえ
ば、丹参を予め50〜100℃好ましくは70〜90℃
にした温水或いはpH4〜9の範囲にある緩衝液または
塩類溶液に入れるか、入れたあと温浴中などで所定の温
度になるまで加温して抽出する。[0013] Preferably, the pH of the extraction solvent is adjusted to about 4 to about 9. The extraction is preferably performed at about 50 to about 100C, more preferably at 70 to 90C. For example, ginseng is pre-heated to 50 to 100 ° C, preferably 70 to 90 ° C.
It is added to warm water or a buffer solution or a salt solution having a pH in the range of 4 to 9, or after it is added, it is heated to a predetermined temperature in a warm bath or the like and extracted.
【0014】かくして、1〜8時間、好ましくは3〜5
時間抽出すれば粗出液が得られる。抽出液中に混在する
丹参の残渣は目の粗い濾布やヌッチェなどで濾去するこ
とができる。Thus, 1 to 8 hours, preferably 3 to 5 hours
A crude extract can be obtained by extraction over time. Dansin residues mixed in the extract can be filtered off with a coarse filter cloth or Nutsche.
【0015】残渣を除去した抽出液は水で平衡化した無
極性の多孔性ポリマー樹脂を充填材とするカラムクロマ
トグラフィーによりさらに精製する。このポリマー樹脂
としては、たとえばHP−20およびMCI−ゲル(三
菱化成社製)やアンバーライトXAO−2(オルガノ社
製)などが挙げられる。The extract from which the residue has been removed is further purified by column chromatography using a nonpolar porous polymer resin equilibrated with water as a filler. Examples of the polymer resin include HP-20 and MCI-gel (manufactured by Mitsubishi Kasei Co., Ltd.) and Amberlite XAO-2 (manufactured by Organo Corporation).
【0016】丹参の活性成分についてMCI−ゲルの吸
着分画で、50%メタノール溶液で溶出される低分子物
質であるマグネシウムリトスペルミン酸Bが報告されて
おり、発明者らも、同様の分画に蛋白尿***を抑制する
活性のあることを確認した。しかし、非吸着部分を含め
溶出された全画分について、詳細に蛋白尿***を抑制す
る作用を試験した結果、当初の予想に反して、素通り画
分にも上記作用を示す成分のあることが判明し、本発明
の物質の発見の端緒となった。この時点で、この処理か
らの素通り画分は、極性が高いことが予想され、又、蛋
白質に固有の紫外部吸収スペクトルを示さないから蛋白
質以外の成分であろうと思われた。It has been reported that magnesium lithospermic acid B, which is a low molecular weight substance eluted with a 50% methanol solution in the MCI-gel adsorption fraction of the active ingredient of Dansin, was reported by the present inventors. Was confirmed to have an activity to suppress proteinuria excretion. However, as a result of a detailed test of the effect of suppressing proteinuria excretion of all the eluted fractions including the non-adsorbed part, contrary to the initial expectation, there is a component that has the above effect even in the pass-through fraction. It turned out to be the starting point of the discovery of the substance of the present invention. At this point, the flow-through fraction from this treatment was expected to be highly polar and would not be a protein-specific UV absorption spectrum, indicating a non-protein component.
【0017】素通り画分の一部を、次いで2枚の限外ろ
過膜(カットオフ,分子量300,000及び50,0
00の)システムに通し、濃縮且つ精製ステップの検討
を行った。実際には分子量300,000以上、50,
000〜300,000及び50,000以下の各物質
群に分け、各成分についてピューロマインシン惹起ラッ
トモデルを用いて、蛋白尿***を抑制する活性を試験し
た。その結果、強い活性が分子量50,000〜30
0,000に範囲にある分画に見られ、50,000以
下にも弱い活性が見られた。A portion of the flow-through fraction was then separated by two ultrafiltration membranes (cut-off, molecular weight 300,000 and 50,000
A review of the concentration and purification steps was performed throughout the system. In practice, the molecular weight is more than 300,000, 50,
Each substance group was divided into 000 to 300,000 and 50,000 or less, and the activity of suppressing proteinuria excretion was tested for each component using a purominsin-induced rat model. As a result, a strong activity has a molecular weight of 50,000-30.
It was found in fractions in the range of 0000, with weak activity below 50,000.
【0018】そこで、先の素通り画分の濃縮に分子量3
000のカットオフ,限外ろ過膜を用いた。この工程で
の限外ろ過膜はカットオフのサイズが同じであれば何れ
の種類のものでも本発明の物質の濃縮に用いることが可
能である。Accordingly, the concentration of the molecular weight of 3
A 000 cut-off, ultrafiltration membrane was used. Any type of ultrafiltration membrane in this step can be used for concentrating the substance of the present invention, as long as it has the same cutoff size.
【0019】次に濃縮液中の活性成分をより狭い分子量
範囲に限定するために、ゲルろ過カラムクロマトグラフ
ィーによる精製を行った。一般に蛋白質などの場合に
は、ゲルろ過クロマトグラフィーの担体としては、セフ
ァデックス及びセファロース類(ファルマシア社製),
セルロファイン(生化学工業社製),バイオゲル(バイ
オラッド社製)などが使用されており、本発明の物質の
クロマトグラフィーにも上記のものが使用できるが、好
ましくは合成手段によって調製された材質によるクロマ
ト材、例えばバイオゲルが本発明の物質の精製に適して
いる。クロマト材には編目サイズ、即ち排除限界により
各種のものがあるが、分画範囲として10000〜30
0,000のものであればいずれでも構わない。発明者
らは、好んでバイオゲルP−30,−60,−100、
中でもバイオゲルP−60を使用した。そこで、濃縮液
を水で平衡化したバイオゲルP−60を充填したカラム
に通した。モニターは暫定的に280nmの吸収で行
い、吸収のある分画と吸収のない分画をカラム容量の2
倍量まで数分画に分け、次いでピューロマイシン惹起ラ
ットモデルでの蛋白尿***の抑制作用を試験したところ
高分子領域に活性を有する多糖類が溶出されていること
がわかった。Next, in order to limit the active ingredient in the concentrated solution to a narrower molecular weight range, purification was performed by gel filtration column chromatography. Generally, in the case of proteins and the like, as carriers for gel filtration chromatography, Sephadex and Sepharose (manufactured by Pharmacia),
Cellulofine (manufactured by Seikagaku Corporation), biogel (manufactured by Bio-Rad) and the like can be used, and the above-mentioned substances can also be used for the chromatography of the substance of the present invention. Chromatographic materials such as biogels are suitable for the purification of the substances according to the invention. There are various types of chromatographic materials depending on the stitch size, that is, the exclusion limit.
Any one of 0000 is acceptable. The inventors prefer biogels P-30, -60, -100,
Among them, Biogel P-60 was used. Therefore, the concentrate was passed through a column filled with Biogel P-60 equilibrated with water. The monitor was tentatively performed at 280 nm absorption, and the fraction with absorption and the fraction without absorption were measured at the column capacity of 2 columns.
The fraction was divided into several fractions up to twice the amount, and the inhibitory effect on proteinuria excretion in a puromycin-induced rat model was examined. As a result, it was found that a polysaccharide having activity in the high molecular weight region was eluted.
【0020】この多糖類は過ヨウ素酸酸化あるいは還元
処理すると活性を失う。また本物質は血小板凝集抑制作
用,赤血球膜保護作用を有することがin vitro
の実験結果より明らかになった。さらにピューロマイシ
ン惹起ネフローゼモデルラットまたは四塩化炭素誘発肝
障害モデルラットにおいてはラジカルが関与することが
知られているが、本物質には直接ラジカルを消去する作
用はないことがESRを用いた実験により示された。This polysaccharide loses its activity when subjected to periodate oxidation or reduction treatment. In addition, this substance has an inhibitory effect on platelet aggregation and a protective effect on erythrocyte membrane in vitro.
It became clear from the experimental result. Further, it is known that radicals are involved in puromycin-induced nephrotic model rats or carbon tetrachloride-induced liver injury model rats. However, it has been confirmed by experiments using ESR that this substance has no action of directly scavenging radicals. Indicated.
【0021】試験例および実施例においては、丹参の抽
出液を無極性ポーラスポリマー樹脂及びゲルろ過クロマ
トグラフィーで精製(精製画分)した多糖類の特性を下
記のように試験した。In Test Examples and Examples, the properties of polysaccharide obtained by purifying (purified fraction) the extract of Dansin using a nonpolar porous polymer resin and gel filtration chromatography were tested as follows.
【0022】(1)全糖含量:フェノール硫酸法(Ho
dge,J.E.and Hofreiter,B.
T.(1962)Methods in Carboh
ydrate Chemistry(Academic
Press).Vol.1,p.338) 試験溶液0.5mlを試験管にとり5%(w/v)フェ
ノール液0.5mlを加え、濃硫酸2.5mlを速やか
に液面に直接に加えよくまぜ、室温に20分保った後、
480nmの吸光度を測定する。標準溶液にはグルコー
ス(10〜90μg/ml)水溶液を用いた。(1) Total sugar content: phenol-sulfuric acid method (Ho
dge, J.M. E. FIG. and Hoofreiter, B.A.
T. (1962) Methods in Carboh
ydrate Chemistry (Academic
Press). Vol. 1, p. 338) Take 0.5 ml of the test solution into a test tube, add 0.5 ml of 5% (w / v) phenol solution, immediately add 2.5 ml of concentrated sulfuric acid directly to the liquid surface, mix well, and keep at room temperature for 20 minutes. ,
The absorbance at 480 nm is measured. An aqueous solution of glucose (10 to 90 μg / ml) was used as a standard solution.
【0023】(2)ウロン酸含量:メタヒドロキシジフ
ェニル法(Blumenkrantz,N.and A
sboe Hansen,G.(1973)Anal.
Biochem.54,484−489.) 試料溶液0.2mlに0.0125Mホウ酸ナトリウム
の濃硫酸溶液1.2mlを添加し、氷冷後攪はんする。
100℃で5分間処理した後氷冷し、0.15%メタヒ
ドロキシジフェニルの0.5%水酸化ナトリウム溶液2
0μlを加え、520nmの吸光度を測定する。標準溶
液にはガラクツロン酸溶液を用いた。(2) Uronic acid content: metahydroxydiphenyl method (Blumenkrantz, N. and A.
sboe Hansen, G .; (1973) Anal.
Biochem. 54, 484-489. ) Add 0.2 ml of a 0.0125 M sodium borate concentrated sulfuric acid solution to 0.2 ml of the sample solution, and stir after cooling on ice.
After treatment at 100 ° C. for 5 minutes, the mixture was cooled on ice, and 0.5% sodium hydroxide solution of 0.15% metahydroxydiphenyl 2
Add 0 μl and measure the absorbance at 520 nm. A galacturonic acid solution was used as a standard solution.
【0024】(3)中性糖含量(アルジトールアセテー
ト−GC−MS法)(Borchardt,L.G.a
nd Piper,C.V.(1970)Tappi.
53,257−260.)(3) Neutral sugar content (Alditol acetate-GC-MS method) (Borchardt, LGa)
nd Piper, C.I. V. (1970) Tappi.
53, 257-260. )
【0025】丹参の精製画分50mgを72%(v/
v)硫酸3mlに溶かし、30℃,1時間処理したもの
に蒸留水84mlを加え、120℃で1時間オートクレ
ープする。内部標準としてイノシトール4mgを加え、
その30mlを水酸化バリウムを加えてpH5.5に調
整した後、遠心分離して上清を得る。その25mlに水
素化ホウ素ナトリウム80mgを添加し、室温で2時間
処理後、酢酸で反応をとめる。濃縮乾燥後、蒸留水に溶
かしDEAEセファデックスカラムにかけ素通り画分を
集める。濃縮乾燥後、無水酢酸とピリジンによりアセチ
ル化した後、ジクロルメタンと1N塩酸の混液で抽出す
る。有機層を蒸留水で洗った後、濃縮し、アセトンに溶
かしてガスクロマトグラフィーマススペクトロメトリー
(GC−MS)にて定量した(カラム:Supelco
糖分析用キャピラリーカラムを使用)。また標品は上記
の還元操作により処理して同様に分析した。The purified fraction of Dansin 50 mg was added to 72% (v / v
v) After dissolving in 3 ml of sulfuric acid and treating at 30 ° C. for 1 hour, add 84 ml of distilled water and autoclave at 120 ° C. for 1 hour. Inositol 4mg was added as an internal standard,
30 ml thereof is adjusted to pH 5.5 by adding barium hydroxide, and then centrifuged to obtain a supernatant. 80 mg of sodium borohydride is added to 25 ml of the mixture, and the mixture is treated at room temperature for 2 hours, and the reaction is stopped with acetic acid. After concentration and drying, the residue is dissolved in distilled water and passed through a DEAE Sephadex column to collect a flow-through fraction. After concentration and drying, the mixture is acetylated with acetic anhydride and pyridine, and then extracted with a mixture of dichloromethane and 1N hydrochloric acid. The organic layer was washed with distilled water, concentrated, dissolved in acetone, and quantified by gas chromatography mass spectrometry (GC-MS) (column: Supelco).
Use a capillary column for sugar analysis). The sample was processed by the above-mentioned reduction operation and analyzed in the same manner.
【0026】(4)分子量の測定 トーソーHPLCシステム(トーソー社製)にアサヒパ
ックGS−520(旭化成社製)カラム(7.6mmI
Dx500mm)を接続し、ミリQ水を用いて、本願物
質試料溶液10μlをカラムに付加して分子量を測定
(温度:室温、流速:0.5ml/分、検出:RID)
した。分子量マーカーにはアミロースキット(中埜酢店
社製):分子量10200,30100,75200,
111400,364200を用い、これらの溶出パタ
ーンから得た検量線より、本願物質の分子量を算出し
た。(4) Measurement of molecular weight Asahi Pack GS-520 (Asahi Kasei) column (7.6 mmI) was applied to a Tosoh HPLC system (Tosoh).
Dx500 mm), and using Milli-Q water, add 10 μl of the sample solution of the substance of the present invention to the column and measure the molecular weight (temperature: room temperature, flow rate: 0.5 ml / min, detection: RID)
did. Amylose kit (manufactured by Nakano Vinegar Co., Ltd.): molecular weight 10200, 30100, 75200,
Using 111400 and 364200, the molecular weight of the substance of the present invention was calculated from a calibration curve obtained from these elution patterns.
【0027】(5)ピューロマイシン惹起ネフローゼ症
候群モデルラットによる評価 1)ネフローゼ症候群モデルラットの作製:ウィスター
系雌性ラット(約9週令、体重約150g)にピューロ
マイシンアミノヌクレオシド(:以下ピューロマイシ
ン、シグマ社製)60mg/kgを1回、頸静脈内投与
してネフローゼ症候群モデルラットを作製した(東條静
夫ら:ネフローゼ症候群 京極方久 編,ソフトサイエ
ンス社出版,479−488,1984)。(5) Evaluation using a puromycin-induced nephrotic syndrome model rat 1) Preparation of a nephrotic syndrome model rat: Puromycin aminonucleoside (hereinafter referred to as puromycin, sigma) was administered to a Wistar female rat (about 9 weeks old, weighing about 150 g). A nephrotic syndrome model rat was prepared by injecting 60 mg / kg once into the jugular vein (Shizuo Tojo et al .: Nephrotic Syndrome Hikaru Kyogoku, Soft Science Publishing, 479-488, 1984).
【0028】2)試料溶液の投与:ピューロマイシン投
与の開始の日から1日1回各試料溶液を胃カニューレを
通じて21日間経口或いは筋肉内投与した。陰性対照群
には注射用水を、陽性対照群にはジピリダモール溶液を
用いた。2) Administration of sample solutions: Each sample solution was orally or intramuscularly administered for 21 days through a gastric cannula once a day from the start of puromycin administration. Water for injection was used for the negative control group, and dipyridamole solution was used for the positive control group.
【0029】3)尿蛋白質量の測定:尿蛋白質量は、ピ
ューロマイシン投与終了後5日目から、2〜3日おきに
24時間尿をラット代謝ケージを用いて採尿し、尿量を
計測した。次いで遠心分離(3000rpmx10分)
して得た上清1mlにスルホサリチル酸3mlを加え、
室温で10分間放置した後、波長660nmの吸収を測
定し、検量線より尿中のタンパク質濃度を求め、更に尿
容量を乗じて尿中への総蛋白質***量を算出した。検量
線作成に当たっては牛血清アルブミン(シグマ社製)を
標準タンパク質に用いた。3) Measurement of urine protein content: From the 5th day after the end of puromycin administration, urine was collected for 24 hours every 2 to 3 days using a rat metabolic cage, and the amount of urine protein was measured. . Then centrifugation (3000 rpm x 10 minutes)
3 ml of sulfosalicylic acid was added to 1 ml of the resulting supernatant,
After standing at room temperature for 10 minutes, the absorption at a wavelength of 660 nm was measured, the protein concentration in the urine was determined from a calibration curve, and the total protein excretion in the urine was calculated by multiplying the urine volume. In preparing a calibration curve, bovine serum albumin (Sigma) was used as a standard protein.
【0030】4)血清コレステロール,血清アルブミ
ン,血清中の総蛋白質量及び血清過酸化脂質の測定:ピ
ューロマイシン投与終了後7日目,14日目,21日目
に各群のラットをエーテル麻酔を施した後、静脈より採
血し、血清中のコレステロール1アルブミン,総蛋白質
量及び過酸化脂質量を測定した(いずれも和光純薬工業
社製)。4) Measurement of serum cholesterol, serum albumin, total protein content in serum and serum lipid peroxide: Rats of each group were anesthetized with ether on days 7, 14 and 21 after completion of puromycin administration. After the administration, blood was collected from a vein, and cholesterol 1 albumin, total protein and lipid peroxide in serum were measured (all manufactured by Wako Pure Chemical Industries, Ltd.).
【0031】(6)四塩化炭素誘発肝障害モデルによる
評価 1)肝障害モデルの作成:Wistar系雌性ラット
(9−11週令、体重160g前後)に対し、オリーブ
油に等量の割合で混和した四塩化炭素(和光純薬製)溶
液1.5ml/kgを腹空内に1回投与することにより
作成した。(炎症学書:炎症炎症動物実験法、実験的肝
繊維症.医学書院出版,253−277)(6) Evaluation using a carbon tetrachloride-induced liver injury model 1) Preparation of a liver injury model: Female Wistar rats (9-11 weeks old, weighing around 160 g) were mixed with olive oil in an equal amount. It was prepared by injecting 1.5 ml / kg of carbon tetrachloride (manufactured by Wako Pure Chemical Industries) solution intraperitoneally once. (Inflammation book: Inflammation inflammation animal experiment method, experimental liver fibrosis. Medical Shoin Publishing, 253-277)
【0032】2)試料溶液の投与:四塩化炭素投与3日
前から投与前日まで試料を1日1回筋肉内投与あるいは
7日前から投与前日まで経口投与を行った。陰性対照群
には注射用水を投与した。2) Administration of the sample solution: The sample was administered intramuscularly once a day from 3 days before the administration of carbon tetrachloride to the day before the administration, or orally from 7 days before the administration to the day before the administration. Water for injection was administered to the negative control group.
【0033】3)血清生化学的検査:四塩化炭素投与2
4時間後にエーテル麻酔下で開胸、心臓より2mlの血
液を採血し、常温に1時間放置後、遠心分離して血清を
得た。血清トランスアミナーゼ(GOT:glutam
ic oxaloacetic transamina
se,GPT:glutamic pyruvictr
ansaminase)活性の測定は和光純薬製キット
を用いて行った。3) Serum biochemical test: administration of carbon tetrachloride 2
Four hours later, the chest was opened under ether anesthesia, 2 ml of blood was collected from the heart, left at room temperature for 1 hour, and centrifuged to obtain serum. Serum transaminase (GOT: glutam)
ic oxaloacetic transamino
se, GPT: glutamic pyruvictr
An activity of the enzyme was measured using a kit manufactured by Wako Pure Chemical Industries.
【0034】7)統計解析:それぞれの測定データはす
べて平均±S.Eで示し、Student’sのt検定
により統計解析した。7) Statistical analysis: All measured data are mean ± SEM This was indicated by E and statistically analyzed by Student's t-test.
【0035】下記の実施例に示されるように、本発明の
多糖類は動物実験でネフローゼ症候群の寛解や肝障害症
状の改善に有効であり、ネフローゼ症候群の寛解剤また
は肝障害症状改善剤として、通常成人1人当たり1日
量、経口的に約10〜100mg、筋肉内注射として約
0.5〜10mg投与することができる。As shown in the following examples, the polysaccharides of the present invention are effective in ameliorating nephrotic syndrome and ameliorating hepatic impairment symptoms in animal experiments. Usually, a daily dose per adult is about 10 to 100 mg orally, and about 0.5 to 10 mg can be administered as an intramuscular injection.
【0036】[0036]
【実施例】以下に実施例および試験例を挙げて本発明を
さらに説明する。 実施例1 中国,四川地区産の丹参の根部の小片10kgに水80
Lを加え、投げ込みヒーターを投じ80℃にした後、3
時間攪はんしながら抽出した。抽出液中の根部残渣はヌ
ッチェでろ過して取り除き、回収した根部残渣に更に水
80Lを加えて1回目と同様に処理して再抽出液を得
た。1回目と2回目の抽出液(130L)を合わせ、減
圧下、40℃でエバポレーターを用いて濃縮し濃縮液
(10L)を得た。The present invention will be further described below with reference to examples and test examples. Example 1 Water was added to 10 kg of small pieces of ginseng root from Sichuan, China.
L, add a throwing heater and bring to 80 ° C.
Extracted with time stirring. The root residue in the extract was removed by filtration with Nutsche, and the collected root residue was further treated with 80 L of water and treated in the same manner as the first time to obtain a re-extract. The first and second extractions (130 L) were combined, and concentrated under reduced pressure at 40 ° C. using an evaporator to obtain a concentrated liquid (10 L).
【0037】次に、濃縮液をHP−20(4L,三菱化
成社製)を充填したカラム(11×45cm)に付加し
た。平衡化液及び展開液には水を用い、室温下でクロマ
トグラフィーを行い、素通り分画及び水洗浄分画(非吸
着)液を回収した。カラムは50%メタノール及びアセ
トニトリルで洗浄した後、最後に水に完全に置換して再
使用した。上記非吸着画分は繰り返し3回程同様に処理
して完全に非吸着分画の液(55L)を得た。この全非
吸着液は分子量カットオフ3000の限外ろ過膜を装備
した限外ろ過装置SEP−1013(旭化成)を用いて
限外ろ過濃縮し、通過液(分子量 約3000以下,S
EP−OUT)と非通過液(分子量 約3000以上,
SEP−IN)を得た。濃縮した非通過液(SEP−I
N)を水で平衡化したバイオゲルP−60を充填したカ
ラム(1.5×100cm)に付加し、水を展開液とし
て用いクロマトグラフィーを行った。モニターは暫定的
に波長280nmで吸収を測定し、横軸に分画番号を、
縦軸に各分画液の吸収を記載したグラフを作成し、吸収
を示さない分画を中心に全糖の測定を行い、その測定結
果を同じグラフに記載した(図1)。Next, the concentrate was added to a column (11 × 45 cm) packed with HP-20 (4 L, manufactured by Mitsubishi Kasei). Chromatography was performed at room temperature using water as the equilibrating solution and the developing solution, and a flow-through fraction and a water-washed fraction (non-adsorbed) solution were collected. The column was washed with 50% methanol and acetonitrile, and finally was completely replaced with water and reused. The non-adsorbed fraction was repeatedly treated in the same manner about three times to obtain a completely non-adsorbed fraction (55 L). The total non-adsorbed liquid was subjected to ultrafiltration and concentration using an ultrafiltration apparatus SEP-1013 (Asahi Kasei) equipped with an ultrafiltration membrane having a molecular weight cutoff of 3000, and the passed liquid (molecular weight of about 3000 or less, S
EP-OUT) and non-passing liquid (molecular weight about 3000 or more,
(SEP-IN) was obtained. Concentrated non-passage liquid (SEP-I
N) was added to a column (1.5 × 100 cm) packed with Biogel P-60 equilibrated with water, and chromatography was performed using water as a developing solution. The monitor tentatively measures the absorption at a wavelength of 280 nm, and displays the fraction number on the horizontal axis,
A graph in which the vertical axis represents the absorption of each fraction was prepared, the total sugar was measured mainly on the fractions showing no absorption, and the measurement results were described in the same graph (FIG. 1).
【0038】目的の多糖類成分は分画番号11から22
の分画液(AF)に得られ、ついで凍結乾燥して粉末
(30g,ロットA)を得た。ここで得た精製多糖類の
特性を前述した試験法により試験した結果、以下の持性
を得た。The objective polysaccharide components are fractions 11 to 22
And then freeze-dried to obtain a powder (30 g, lot A). The properties of the purified polysaccharide obtained here were tested by the test method described above, and as a result, the following durability was obtained.
【0039】1)糖含量(フェノール硫酸法):79% (1)糖組成:ウロン酸含量(62%、メタヒドロキシ
ジフェニル法) 中性糖含量(17%、アルジトールアセテート−GC−
MS法) (2)中性糖組成(アルジトールアセテート−GC−M
S法) グルコース含量(5.5%) ガラクトース含量(39.7%) アラビノース含量(44.6%) マンノース含量(5.5%) ラムノース含量(4.7%) (3)その他の糖:アミノ糖、アルドヘキソース、2−
デオキシ糖は含まない。1) Sugar content (phenol-sulfuric acid method): 79% (1) Sugar composition: uronic acid content (62%, metahydroxydiphenyl method) Neutral sugar content (17%, alditol acetate-GC-)
MS method) (2) Neutral sugar composition (Alditol acetate-GC-M)
Method S) Glucose content (5.5%) Galactose content (39.7%) Arabinose content (44.6%) Mannose content (5.5%) Rhamnose content (4.7%) (3) Other sugars: Amino sugar, aldohexose, 2-
Does not contain deoxy sugar.
【0040】2)分子量:259000(標準多糖:ア
ミラーゼキット、分子量100〜360K)2) Molecular weight: 259000 (standard polysaccharide: amylase kit, molecular weight 100-360K)
【0041】3)ネフローゼ症候群モデルラットによる
評価 精製試料10mg/kg,40mg/kgを経口投与、
0.5mg/kg,2.5mg/kgを筋肉内投与した
ところ、図2,図3に示したようにいずれの場合も尿蛋
白***が有意に抑制された。また、表1,表2に示した
ように血清コレステロール(Cholesterol)
値,血清アルブミン(Albumin)値及びA/G
比,血清過酸化脂質(MDA)値においても有意な改善
効果が認められた。3) Evaluation by Nephrotic Syndrome Model Rat Orally administered purified samples 10 mg / kg and 40 mg / kg,
When 0.5 mg / kg and 2.5 mg / kg were intramuscularly administered, urine protein excretion was significantly suppressed in each case as shown in FIGS. In addition, as shown in Tables 1 and 2, serum cholesterol (Cholesterol)
Value, serum albumin (Albumin) value and A / G
A significant improvement was also observed in the ratio and serum lipid peroxide (MDA) value.
【0042】[0042]
【表1】 (表中、PAはピューロマイシンアミノヌクレオシド、
AFは本発明の多糖類を表す。以下の表においてもおな
じ)[Table 1] (In the table, PA is puromycin aminonucleoside,
AF represents a polysaccharide of the invention. The same applies to the table below)
【0043】[0043]
【表2】 [Table 2]
【0044】4)四塩化炭素誘発肝障害モデルによる評
価 精製試料(AF)7mg/kg,35mg/kgを経口
投与、0.3mg/kg,1.5mg/kgを筋肉内投
与したところ、表4に示したようにいずれの場合もAF
前投与群では血清GOT,GPT値の上昇が用量依存的
に抑制された。4) Evaluation by carbon tetrachloride-induced liver injury model Purified samples (AF) were orally administered at 7 mg / kg and 35 mg / kg and intramuscularly administered at 0.3 mg / kg and 1.5 mg / kg. As shown in FIG.
In the pre-administration group, increases in serum GOT and GPT levels were suppressed in a dose-dependent manner.
【0045】実施例2 実施例1に記載の方法によりロットの異なる丹参原料よ
り各々精製を行い、1に記載した以外に計3ロット(ロ
ットB,C,D)の精製品を取得し、同様の特性評価を
行った。Example 2 According to the method described in Example 1, purification was carried out from raw materials of different lots of Dansan, and purified lots of 3 lots (lots B, C, D) were obtained in addition to those described in 1 above. Was evaluated.
【0046】1)糖含量(フェノール硫酸):ロットB
(65%),ロットC(80%),ロットD(93%) (1)糖組成1) Sugar content (phenol sulfate): Lot B
(65%), Lot C (80%), Lot D (93%) (1) Sugar composition
【0047】[0047]
【表4】 [Table 4]
【0048】(2)中性糖組成(アルジトールアセテー
ト−GC−MS法)(2) Neutral sugar composition (alditol acetate-GC-MS method)
【0049】[0049]
【表5】 [Table 5]
【0050】(3)その他の糖:ロットB,C,Dのい
ずれもアミノ糖、アルドヘキソース、2−デオキシ糖は
含まない。(3) Other sugars: None of lots B, C and D contain amino sugars, aldohexoses and 2-deoxy sugars.
【0051】2)分子量:(標準多糖:アミロースキャ
ット、分子量100〜360K) ロットB(150,000),ロットC(280,00
0),ロットD(250,000)2) Molecular weight: (Standard polysaccharide: amylose cat, molecular weight 100-360K) Lot B (150,000), Lot C (280,00)
0), Lot D (250,000)
【0052】3)ネフローゼ症候群モデルラットによる
評価 薬効評価についてはいずれの精製品も実施例1で得られ
たものとほぼ同等の効果が得られた。3) Evaluation by Nephrotic Syndrome Model Rat In the evaluation of drug efficacy, all purified products obtained almost the same effects as those obtained in Example 1.
【0053】実施例3 活性の本体が糖であること及びウロン酸の薬効に対する
関与について調べる目的で本精製品を以下の操作によっ
て酸化及び還元し、それらの効果について調べた。投与
量はネフローゼ症候群ラットにはそれぞれ40mg/k
gを経口投与し、肝障害ラットにはそれぞれ1.5mg
/kgを筋肉内投与した。Example 3 The purified product was oxidized and reduced by the following operation to examine the fact that the main ingredient of the activity was sugar and the contribution of uronic acid to the medicinal effect, and the effects thereof were examined. The dose was 40 mg / k for nephrotic syndrome rats.
g was orally administered to the liver-impaired rats.
/ Kg was administered intramuscularly.
【0054】(1)酸化(Noble,D.W.and
Sturgeon,R.J.(1970)Carbo
hyd,Res.12,448による)精製試料600
mgを蒸留水150mlに溶かし、0.2M過ヨウ素酸
ナトリウム溶液150mlを加えて37℃で240時間
反応させる。エチレングリコール150mlを加えて反
応を止めた後、水素化ホウ素ナトリウム500mgを加
えて室温24時間処理する。酢酸で反応を止め、水に透
析した後、凍結乾燥する。(1) Oxidation (Noble, DW and
Sturgeon, R .; J. (1970) Carbo
hyd, Res. 12,448) purified sample 600
mg is dissolved in 150 ml of distilled water, 150 ml of a 0.2 M sodium periodate solution is added, and the mixture is reacted at 37 ° C. for 240 hours. After stopping the reaction by adding 150 ml of ethylene glycol, 500 mg of sodium borohydride is added and the mixture is treated at room temperature for 24 hours. The reaction is stopped with acetic acid, dialyzed against water, and lyophilized.
【0055】(2)還元(Taylor,R.L.an
d Conrad,H.E.(1972)Bioche
mistry.11,1383−1388による)精製
試料50mgを蒸留水に溶かし、EDC(1−ethy
l−3−(dimethyl−aminopropy
l)−carbodiimide)249mgを加えた
後、0.1N塩酸でpH4.75に保ちながら19時間
反応させる。その後、2M水素化ホウ素ナトリウム溶液
10mlを1時間かけて滴下し、その間pH7に保つ。
さらに1時間攪はんした後、水に透析し凍結乾燥する。(2) Reduction (Taylor, RL ann)
d Conrad, H .; E. FIG. (1972) Bioche
mistry. Dissolve 50 mg of purified sample (according to 11,1383-1388) in distilled water and add EDC (1-ethyl
l-3- (dimethyl-aminopropy)
After adding 249 mg of l) -carbodiimide), the mixture is reacted for 19 hours while maintaining pH 4.75 with 0.1N hydrochloric acid. Thereafter, 10 ml of a 2M sodium borohydride solution are added dropwise over 1 hour, during which the pH is kept at 7.
After stirring for an additional hour, the mixture is dialyzed against water and freeze-dried.
【0056】結果を図4ならびに表3、表4および表5
に示したが、酸化及び還元した試料については全く効果
が認められなかった。このことは精製試料中の活性物質
は確かに多糖類であり、中でも特にウロン酸が薬効に大
きく寄与していることが示された。また還元前後の中性
糖含量の比較から本試料に含まれるウロン酸のほとんど
はガラクツロン酸であることがわかった。The results are shown in FIG. 4 and Tables 3, 4 and 5.
No effect was observed for the oxidized and reduced samples. This indicates that the active substance in the purified sample is indeed a polysaccharide, and that uronic acid in particular greatly contributes to the drug efficacy. Comparison of the neutral sugar content before and after the reduction indicated that most of the uronic acid contained in this sample was galacturonic acid.
【0057】[0057]
【表3】 [Table 3]
【0058】[0058]
【表4】[Table 4]
【0059】[0059]
【表5】[Table 5]
【0060】[0060]
【発明の効果】本発明によれば、丹参から抽出される新
規な多糖類が提供され、それはネフローゼ症候群の寛解
や肝障害症状の改善に用いることができる。According to the present invention, there is provided a novel polysaccharide extracted from ginseng, which can be used for remission of nephrotic syndrome and improvement of liver disorder symptoms.
【0061】[0061]
図1は実施例1におけるゲルろ過クロマトグラフィーに
より展開された分画の波長280nm(A280 )および
480nm(A480 )の吸光度、図2および図3は実施
例1においてネフローゼ症候群モデルラットに本発明の
多糖類(AF)を種々の投与量で経口投与した場合の尿
蛋白***量、図4はAFを酸化または還元して同様に投
与した場合の尿蛋白***量、をそれぞれ示すグラフであ
る。FIG. 1 shows the absorbance at 280 nm (A 280 ) and 480 nm (A 480 ) of the fraction developed by gel filtration chromatography in Example 1, and FIGS. 2 and 3 show the present invention in a nephrotic syndrome model rat in Example 1. FIG. 4 is a graph showing the amount of urinary protein excreted when the polysaccharide (AF) is orally administered at various doses, and FIG. 4 is a graph showing the amount of urinary protein excreted when the same is administered by oxidizing or reducing AF.
───────────────────────────────────────────────────── フロントページの続き (72)発明者 阿部 博子 大阪府堺市向陵西町四丁目10−8 サン ライズガーデン三国ケ丘406号 (58)調査した分野(Int.Cl.7,DB名) C08B 37/00 A61K 35/78 CA(STN)────────────────────────────────────────────────── ─── Continuation of front page (72) Inventor Hiroko Abe 4-10-8 Kouri Nishimachi, Sakai City, Osaka Sunrise Garden Mikunigaoka 406 (58) Field surveyed (Int. Cl. 7 , DB name) C08B 37/00 A61K 35/78 CA (STN)
Claims (8)
る下記の特性を有する多糖類 A.糖含量:60〜100% (1)糖組成:ウロン酸 40〜80% 中性糖 10〜30% (2)中性糖組成:ラムノース 0〜15% グルコース 0〜15% ガラクトース 25〜55% アラビノース 30〜60% マンノース 0〜15% B.分子量 150,000〜300,0001. A polysaccharide having the following properties, which can be extracted from ginseng with water or an aqueous solvent. Sugar content: 60-100% (1) Sugar composition: uronic acid 40-80% Neutral sugar 10-30% (2) Neutral sugar composition: rhamnose 0-15% glucose 0-15% galactose 25-55% arabinose 30-60% Mannose 0-15% B. Molecular weight 150,000-300,000
液から残渣を除去した液を無極性多孔性ポリマー樹脂を
通過させ、限外濾過で濃縮し、ついで限外濾過クロマト
グラフィーに付することを特徴とする請求項1記載の多
糖類の製造法。2. Ginseng is extracted with water or an aqueous solvent, and the liquid obtained by removing the residue from the extract is passed through a non-polar porous polymer resin, concentrated by ultrafiltration, and then subjected to ultrafiltration chromatography. The method for producing a polysaccharide according to claim 1, wherein
項2記載の製造法。3. The method according to claim 2, wherein the ginseng is extracted at a pH of about 4 to about 8.
る請求項2記載の製造法。4. The method according to claim 2, wherein the aqueous solvent is a buffer or an aqueous salt solution.
請求項2または3記載の製造法。5. The method according to claim 2, wherein the ginseng is extracted at about 40 to about 100 ° C.
ゼ症候群の寛解剤。6. An agent for ameliorating nephrotic syndrome comprising the polysaccharide according to claim 1.
状の改善剤。7. An agent for ameliorating hepatic disorder comprising the polysaccharide according to claim 1.
5記載の寛解剤。8. The ameliorating agent according to claim 5, which is administered intramuscularly or orally.
Priority Applications (11)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP19927593A JP3161882B2 (en) | 1993-06-03 | 1993-07-15 | Ginseng-derived polysaccharide, its production method and use |
FI943242A FI943242A (en) | 1993-07-15 | 1994-07-07 | A remedy for kidney and liver diseases |
US08/271,795 US5547945A (en) | 1993-07-15 | 1994-07-07 | Remitting agent for nephrotic syndrome and hepatopathy symptoms |
CA002127934A CA2127934A1 (en) | 1993-07-15 | 1994-07-13 | Remitting agent for nephrotic syndrome and hepatopathy symptoms |
KR1019940016859A KR960013374A (en) | 1993-07-15 | 1994-07-13 | Nephrotic syndrome and liver disease symptoms |
RU94026096A RU2119341C1 (en) | 1993-07-15 | 1994-07-14 | Agent attenuating a sickness course at nephrotic syndrome and hepatopathy symptoms, water-soluble polysaccharide, method of polysaccharide preparing |
AT94305146T ATE171462T1 (en) | 1993-07-15 | 1994-07-14 | POLYGLUCURONIC ACID AS A REMITTING AGENT FOR NEPHROTIC SYNDROME AND SYMPTOMS OF HEPATOPATHY |
AU67470/94A AU686161B2 (en) | 1993-07-15 | 1994-07-14 | Remitting agent for nephrotic syndrome and hepatopathy symptoms |
EP94305146A EP0635519B1 (en) | 1993-07-15 | 1994-07-14 | Polyglucuronic acid as remitting agent for nephrotic syndrome and hepatopathy symptoms |
DE69413467T DE69413467T2 (en) | 1993-07-15 | 1994-07-14 | Polyglucuronic acid as a remitting agent for nephrotic syndrome and symptoms of hepatopathy |
ZA945205A ZA945205B (en) | 1993-07-15 | 1994-07-15 | Remitting agent for nephrotic syndrome and hepatopathy symptoms |
Applications Claiming Priority (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP16037393 | 1993-06-03 | ||
JP5-160373 | 1993-06-03 | ||
JP19927593A JP3161882B2 (en) | 1993-06-03 | 1993-07-15 | Ginseng-derived polysaccharide, its production method and use |
Publications (2)
Publication Number | Publication Date |
---|---|
JPH0748403A JPH0748403A (en) | 1995-02-21 |
JP3161882B2 true JP3161882B2 (en) | 2001-04-25 |
Family
ID=26486913
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JP19927593A Expired - Fee Related JP3161882B2 (en) | 1993-06-03 | 1993-07-15 | Ginseng-derived polysaccharide, its production method and use |
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JP (1) | JP3161882B2 (en) |
Families Citing this family (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6149915A (en) * | 1999-09-29 | 2000-11-21 | Shiva Biomedical, Llc | Treatment of diabetic nephropathy and microalbuminuria |
CN1785284B (en) | 2004-12-10 | 2012-04-18 | 天津天士力制药股份有限公司 | Medicinal composition containing flowery knotwood root |
JP2010233497A (en) * | 2009-03-31 | 2010-10-21 | Institute Of National Colleges Of Technology Japan | Extract from soybean curd refuse |
-
1993
- 1993-07-15 JP JP19927593A patent/JP3161882B2/en not_active Expired - Fee Related
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JPH0748403A (en) | 1995-02-21 |
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