JPS6390505A - Novel polysaccharide - Google Patents
Novel polysaccharideInfo
- Publication number
- JPS6390505A JPS6390505A JP23583686A JP23583686A JPS6390505A JP S6390505 A JPS6390505 A JP S6390505A JP 23583686 A JP23583686 A JP 23583686A JP 23583686 A JP23583686 A JP 23583686A JP S6390505 A JPS6390505 A JP S6390505A
- Authority
- JP
- Japan
- Prior art keywords
- polysaccharide
- galactose
- freeze
- filtration chromatography
- gel
- 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.)
- Granted
Links
- 150000004676 glycans Chemical class 0.000 title claims abstract description 27
- 229920001282 polysaccharide Polymers 0.000 title claims abstract description 27
- 239000005017 polysaccharide Substances 0.000 title claims abstract description 27
- 229930182830 galactose Natural products 0.000 claims abstract description 11
- 239000000126 substance Substances 0.000 claims abstract description 8
- IAJILQKETJEXLJ-UHFFFAOYSA-N Galacturonsaeure Natural products O=CC(O)C(O)C(O)C(O)C(O)=O IAJILQKETJEXLJ-UHFFFAOYSA-N 0.000 claims abstract description 7
- 235000000346 sugar Nutrition 0.000 claims abstract description 7
- 239000000470 constituent Substances 0.000 claims abstract description 6
- 238000001641 gel filtration chromatography Methods 0.000 claims abstract description 6
- SHZGCJCMOBCMKK-UHFFFAOYSA-N D-mannomethylose Natural products CC1OC(O)C(O)C(O)C1O SHZGCJCMOBCMKK-UHFFFAOYSA-N 0.000 claims abstract description 5
- SHZGCJCMOBCMKK-JFNONXLTSA-N L-rhamnopyranose Chemical compound C[C@@H]1OC(O)[C@H](O)[C@H](O)[C@H]1O SHZGCJCMOBCMKK-JFNONXLTSA-N 0.000 claims abstract description 5
- PNNNRSAQSRJVSB-UHFFFAOYSA-N L-rhamnose Natural products CC(O)C(O)C(O)C(O)C=O PNNNRSAQSRJVSB-UHFFFAOYSA-N 0.000 claims abstract description 5
- 241000208173 Apiaceae Species 0.000 claims abstract description 4
- WQZGKKKJIJFFOK-PHYPRBDBSA-N alpha-D-galactose Chemical compound OC[C@H]1O[C@H](O)[C@H](O)[C@@H](O)[C@H]1O WQZGKKKJIJFFOK-PHYPRBDBSA-N 0.000 claims abstract description 3
- PYMYPHUHKUWMLA-WDCZJNDASA-N arabinose Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)C=O PYMYPHUHKUWMLA-WDCZJNDASA-N 0.000 claims abstract description 3
- PYMYPHUHKUWMLA-UHFFFAOYSA-N arabinose Natural products OCC(O)C(O)C(O)C=O PYMYPHUHKUWMLA-UHFFFAOYSA-N 0.000 claims abstract description 3
- SRBFZHDQGSBBOR-UHFFFAOYSA-N beta-D-Pyranose-Lyxose Natural products OC1COC(O)C(O)C1O SRBFZHDQGSBBOR-UHFFFAOYSA-N 0.000 claims abstract description 3
- 238000001962 electrophoresis Methods 0.000 claims abstract description 3
- 238000004128 high performance liquid chromatography Methods 0.000 claims abstract description 3
- 150000008163 sugars Chemical class 0.000 claims abstract description 3
- AEMOLEFTQBMNLQ-YMDCURPLSA-N D-galactopyranuronic acid Chemical compound OC1O[C@H](C(O)=O)[C@H](O)[C@H](O)[C@H]1O AEMOLEFTQBMNLQ-YMDCURPLSA-N 0.000 claims abstract 4
- HMFHBZSHGGEWLO-HWQSCIPKSA-N L-arabinofuranose Chemical compound OC[C@@H]1OC(O)[C@H](O)[C@H]1O HMFHBZSHGGEWLO-HWQSCIPKSA-N 0.000 claims description 4
- 230000003287 optical effect Effects 0.000 claims 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 abstract description 20
- 239000008213 purified water Substances 0.000 abstract description 14
- 239000002244 precipitate Substances 0.000 abstract description 8
- 208000023275 Autoimmune disease Diseases 0.000 abstract description 4
- 238000004108 freeze drying Methods 0.000 abstract description 4
- 239000006228 supernatant Substances 0.000 abstract description 4
- 206010018367 Glomerulonephritis chronic Diseases 0.000 abstract description 3
- 239000003125 aqueous solvent Substances 0.000 abstract description 3
- 239000000706 filtrate Substances 0.000 abstract description 3
- 150000002632 lipids Chemical class 0.000 abstract description 2
- 239000007788 liquid Substances 0.000 abstract description 2
- 239000003960 organic solvent Substances 0.000 abstract description 2
- 206010043778 thyroiditis Diseases 0.000 abstract description 2
- 241000202722 Bupleurum falcatum Species 0.000 abstract 2
- 230000003171 anti-complementary effect Effects 0.000 abstract 1
- 238000001914 filtration Methods 0.000 abstract 1
- 239000000463 material Substances 0.000 abstract 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 9
- 230000000694 effects Effects 0.000 description 8
- 230000002391 anti-complement effect Effects 0.000 description 7
- 108010008730 anticomplement Proteins 0.000 description 7
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 6
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 6
- 238000002360 preparation method Methods 0.000 description 5
- 239000002775 capsule Substances 0.000 description 4
- 238000005119 centrifugation Methods 0.000 description 4
- 238000002156 mixing Methods 0.000 description 4
- 239000000243 solution Substances 0.000 description 4
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 3
- DNIAPMSPPWPWGF-UHFFFAOYSA-N Propylene glycol Chemical compound CC(O)CO DNIAPMSPPWPWGF-UHFFFAOYSA-N 0.000 description 3
- IAJILQKETJEXLJ-RSJOWCBRSA-N aldehydo-D-galacturonic acid Chemical compound O=C[C@H](O)[C@@H](O)[C@@H](O)[C@H](O)C(O)=O IAJILQKETJEXLJ-RSJOWCBRSA-N 0.000 description 3
- 238000006243 chemical reaction Methods 0.000 description 3
- 238000000502 dialysis Methods 0.000 description 3
- 238000000605 extraction Methods 0.000 description 3
- 239000000499 gel Substances 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 239000002504 physiological saline solution Substances 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- 239000011780 sodium chloride Substances 0.000 description 3
- FTOAOBMCPZCFFF-UHFFFAOYSA-N 5,5-diethylbarbituric acid Chemical compound CCC1(CC)C(=O)NC(=O)NC1=O FTOAOBMCPZCFFF-UHFFFAOYSA-N 0.000 description 2
- LZZYPRNAOMGNLH-UHFFFAOYSA-M Cetrimonium bromide Chemical compound [Br-].CCCCCCCCCCCCCCCC[N+](C)(C)C LZZYPRNAOMGNLH-UHFFFAOYSA-M 0.000 description 2
- 229920002271 DEAE-Sepharose Polymers 0.000 description 2
- 229920002684 Sepharose Polymers 0.000 description 2
- VZTDIZULWFCMLS-UHFFFAOYSA-N ammonium formate Chemical compound [NH4+].[O-]C=O VZTDIZULWFCMLS-UHFFFAOYSA-N 0.000 description 2
- 238000004440 column chromatography Methods 0.000 description 2
- 230000004154 complement system Effects 0.000 description 2
- 238000007796 conventional method Methods 0.000 description 2
- 239000003085 diluting agent Substances 0.000 description 2
- 239000007903 gelatin capsule Substances 0.000 description 2
- 238000002523 gelfiltration Methods 0.000 description 2
- 239000008187 granular material Substances 0.000 description 2
- 239000007924 injection Substances 0.000 description 2
- 238000002347 injection Methods 0.000 description 2
- 238000004255 ion exchange chromatography Methods 0.000 description 2
- HQKMJHAJHXVSDF-UHFFFAOYSA-L magnesium stearate Chemical compound [Mg+2].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O HQKMJHAJHXVSDF-UHFFFAOYSA-L 0.000 description 2
- 239000003755 preservative agent Substances 0.000 description 2
- 210000002966 serum Anatomy 0.000 description 2
- 239000003826 tablet Substances 0.000 description 2
- GUBGYTABKSRVRQ-XLOQQCSPSA-N Alpha-Lactose Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@H](O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 description 1
- 206010008909 Chronic Hepatitis Diseases 0.000 description 1
- 239000004278 EU approved seasoning Substances 0.000 description 1
- 108010010803 Gelatin Proteins 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Chemical compound OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- 229920000084 Gum arabic Polymers 0.000 description 1
- 206010018910 Haemolysis Diseases 0.000 description 1
- 206010020751 Hypersensitivity Diseases 0.000 description 1
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 241001494479 Pecora Species 0.000 description 1
- 241000978776 Senegalia senegal Species 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 239000000205 acacia gum Substances 0.000 description 1
- 235000010489 acacia gum Nutrition 0.000 description 1
- 239000002671 adjuvant Substances 0.000 description 1
- 230000000844 anti-bacterial effect Effects 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 239000003899 bactericide agent Substances 0.000 description 1
- 229960002319 barbital Drugs 0.000 description 1
- 239000003086 colorant Substances 0.000 description 1
- 230000000295 complement effect Effects 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 239000012059 conventional drug carrier Substances 0.000 description 1
- 239000013078 crystal Substances 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 239000002552 dosage form Substances 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 210000003743 erythrocyte Anatomy 0.000 description 1
- 238000011049 filling Methods 0.000 description 1
- 235000011194 food seasoning agent Nutrition 0.000 description 1
- 235000003599 food sweetener Nutrition 0.000 description 1
- 239000003205 fragrance Substances 0.000 description 1
- 239000008273 gelatin Substances 0.000 description 1
- 229920000159 gelatin Polymers 0.000 description 1
- 235000019322 gelatine Nutrition 0.000 description 1
- 235000011852 gelatine desserts Nutrition 0.000 description 1
- JEGUKCSWCFPDGT-UHFFFAOYSA-N h2o hydrate Chemical compound O.O JEGUKCSWCFPDGT-UHFFFAOYSA-N 0.000 description 1
- 208000006454 hepatitis Diseases 0.000 description 1
- 241000411851 herbal medicine Species 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- 239000011777 magnesium Substances 0.000 description 1
- 235000019359 magnesium stearate Nutrition 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 235000010270 methyl p-hydroxybenzoate Nutrition 0.000 description 1
- 239000000546 pharmaceutical excipient Substances 0.000 description 1
- 229940124531 pharmaceutical excipient Drugs 0.000 description 1
- 230000002335 preservative effect Effects 0.000 description 1
- 235000010232 propyl p-hydroxybenzoate Nutrition 0.000 description 1
- 125000001453 quaternary ammonium group Chemical group 0.000 description 1
- 238000010992 reflux Methods 0.000 description 1
- 206010039073 rheumatoid arthritis Diseases 0.000 description 1
- 238000007493 shaping process Methods 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 239000003381 stabilizer Substances 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 239000000829 suppository Substances 0.000 description 1
- 239000003765 sweetening agent Substances 0.000 description 1
- 235000020357 syrup Nutrition 0.000 description 1
- 239000006188 syrup Substances 0.000 description 1
- 239000000454 talc Substances 0.000 description 1
- 229910052623 talc Inorganic materials 0.000 description 1
- 235000012222 talc Nutrition 0.000 description 1
Landscapes
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Polysaccharides And Polysaccharide Derivatives (AREA)
Abstract
Description
【発明の詳細な説明】
［産業上の利用分野］
本発明は抗補体活性を有し、関節リウマチ、甲状腺炎、
慢性肝炎、慢性腎炎等の自己免疫疾患の治療に有効な新
規な多糖に関するものである。[Detailed Description of the Invention] [Industrial Application Field] The present invention has anti-complement activity and is effective against rheumatoid arthritis, thyroiditis,
This invention relates to a new polysaccharide that is effective in treating autoimmune diseases such as chronic hepatitis and chronic nephritis.
［従来の技術および問題点］
血清中に存在する補体系は、アレルギー反応にも関与す
ることが知られており、この補体系の活性を抑制する物
質、すなわち抗補体活性を有する物質が、自己免疫疾患
の治療に有効であることか示唆されている。[Prior Art and Problems] It is known that the complement system present in serum is also involved in allergic reactions, and substances that suppress the activity of the complement system, that is, substances that have anti-complement activity, It has been suggested that it may be effective in treating autoimmune diseases.
従って、抗補体活性を有する薬剤の開発が望まれていた
。Therefore, it has been desired to develop a drug having anti-complement activity.
［問題点を解決するための手段］
本発明者等は、抗補体活性を有する物質を見いだすべく
鋭意研究を重ねた結果、セリ科（Ｉｌｍｂｅｌｌｉｆｅ
ｒａｅ）のミシマサイコ（Ｂｕｐｌｅｕｍｆａｌｃａｔ
ｕｎ＋ Ｌ、）の根である生薬、すなわち柴胡（Ｂｕｐ
ｌｅｕｒｉ Ｒａｄｉｘ）に含有される本発明の多糖体
を単離し、本発明を完成した。本発明の多糖体の理化学
的性質を示すと次の如くである。[Means for Solving the Problems] As a result of intensive research to find a substance having anti-complement activity, the present inventors discovered that the Ilmbelliferae (Umbelliferae)
rae)'s Mishima Psycho (Bupleumfalcat)
The herbal medicine that is the root of un + L, ) is Chaihu (Bup
The present invention was completed by isolating the polysaccharide of the present invention contained in P. leuri Radix. The physicochemical properties of the polysaccharide of the present invention are as follows.
■性状 ：白色繊維状
■分子量 ：３６，０００（ゲル濾過クロマトグラフィ
ー）、
■比旋光度： ［α］晶０＝＋１２０、■構成糖 ：ラ
ムノース、アラビノース、ガラクトース、ガラクツロン
酸、
■単一性 ：電気泳動、ゲル濾過クロマトグラフィー、
高速液体クロマトグラフィーにより単一性を示す、
■構成糖結合様式：
１５結合アラビノフラノース １．４％末端ガラクト
ース ２．７％末端ガラクツロン酸
２．７％３−５分岐アラビノフラノース ２．
８％２−４分岐ラムノース ２．２％ｉ４結
合ガラクツロン酸 ８０．０％！−６結合ガラクト
ース １．９％ｉ４結合ガラクトース
４．２％３−６分岐ガラクトース ０．７％
本発明の多糖体は上記に示した如くの糖構成、結合様式
を持つものであり、このような多糖体に関する発表は全
くなされていないことから、本発明の多糖体は新規な多
糖体であることが認められる。■Properties: white fibrous ■Molecular weight: 36,000 (gel filtration chromatography), ■Specific rotation: [α] crystal 0 = +120, ■Constituent sugars: rhamnose, arabinose, galactose, galacturonic acid, ■Singarity: electrophoresis, gel filtration chromatography,
Uniformity shown by high performance liquid chromatography ■ Constituent sugar binding mode: 15-linked arabinofuranose 1.4% terminal galactose 2.7% terminal galacturonic acid
2.7% 3-5 branched arabinofuranose 2.
8% 2-4 branched rhamnose 2.2% i4-linked galacturonic acid 80.0%! -6-linked galactose 1.9% i4-linked galactose
4.2% 3-6 branched galactose 0.7%
The polysaccharide of the present invention has the sugar composition and binding mode as shown above, and since there has been no publication regarding such a polysaccharide, the polysaccharide of the present invention is a novel polysaccharide. It is recognized that
本発明の多糖体は例えば次のようにして得ることができ
る。The polysaccharide of the present invention can be obtained, for example, as follows.
柴胡を粉砕し、１０倍量程度の水性溶剤にて抽出し、抽
出液を濾過して得た濾液を遠心分離し、上清を凍結乾燥
して乾燥物を得る。この乾燥物をアセトン、メタノール
、エタノール等の有機溶媒に溶解し、場合によっては還
流した後、遠心分離して脂質成分を除いた沈澱物を精製
水に溶解し、更に透析を行い、非透析物を凍結乾燥して
粗分画を得る。The saiko is ground, extracted with about 10 times the volume of an aqueous solvent, the extract is filtered, the resulting filtrate is centrifuged, and the supernatant is freeze-dried to obtain a dried product. This dried product is dissolved in an organic solvent such as acetone, methanol, or ethanol, and if necessary, refluxed, centrifuged to remove lipid components, the precipitate is dissolved in purified water, and further dialysis is performed. Lyophilize to obtain a crude fraction.
この粗分画を精製水に溶解し、臭化セチル−トリメチル
−アンモニウム等の４級アンモニウムを加えて混和し、
放置した後遠心分離する。その沈澱物を５〜３０％程度
の塩化ナトリウム水溶液に溶解し、エタノールを加え、
遠心分離して得た沈澱物を更に精製水に溶解し、透析を
行い、非透析分画を凍結乾燥して多糖含有分画を得る。This crude fraction was dissolved in purified water, and quaternary ammonium such as cetyl-trimethyl-ammonium bromide was added and mixed.
After standing, centrifuge. Dissolve the precipitate in a 5-30% aqueous sodium chloride solution, add ethanol,
The precipitate obtained by centrifugation is further dissolved in purified water, dialyzed, and the non-dialyzed fraction is freeze-dried to obtain a polysaccharide-containing fraction.
この多糖含有分画を、通常用いられるゲル濾過カラムク
ロマトグラフィーおよび／またはイオン交換クロマトグ
ラフィーに付すことにより本発明の多糖体を得る。The polysaccharide of the present invention is obtained by subjecting this polysaccharide-containing fraction to commonly used gel filtration column chromatography and/or ion exchange chromatography.
抽出する水性溶剤としては水、特に精製水が好ましく、
抽出にあたっては熱時抽出が望ましい。Water, especially purified water, is preferred as the aqueous solvent for extraction.
For extraction, hot extraction is preferable.
遠心分離の条件としては６０００〜８０００ ｒｐｍｓ
２０〜４０分程度であれば十分である。透析に際しては
、流水または精製水に対し２〜５日間程度で行うことが
できる。Centrifugation conditions are 6000-8000 rpms
About 20 to 40 minutes is sufficient. Dialysis can be performed against running water or purified water for about 2 to 5 days.
ゲル濾過カラムクロマトグラフィーのゲルの具体例とし
てはセファロース（５ｅｐｈａｒｏｓｅ） ＣＬ−６Ｂ
等が挙げられ、イオン交換クロマトグラフィーのゲルの
具体例としてはＤＥＡＥ−セファロース（５ｅｐｈａｒ
ｏｓｅ） ＣＬ−６Ｂ等が挙げられる。A specific example of gel for gel filtration column chromatography is Sepharose CL-6B.
A specific example of a gel for ion exchange chromatography is DEAE-Sepharose (5ephar).
ose) CL-6B and the like.
次に本発明の多糖体の製造の具体例を示す。Next, a specific example of the production of the polysaccharide of the present invention will be shown.
具体例
柴胡１．ＯｋｇにＩＯＣの精製水を加え、液量が半量に
なるまで加熱抽出した。抽出液を濾過し、濾液を７５０
Ｏｒｐｍ、３０分間遠心分離し、上清を得た。沈澱物
に上記と同様の操作を繰り返し、上滑を合併し、凍結乾
燥して得た乾燥物１３６．３ｇをメタノールに溶解し、
還流、遠心分離、沈澱物を精製水に溶解した後、流水に
対し３日間、精製水に対し２日間透析した。非透析分画
を凍結乾燥し、粗分画３５．３９を得た。Specific example: Saiko 1. IOC purified water was added to Okg, and the mixture was heated and extracted until the liquid volume was reduced to half. Filter the extract and reduce the filtrate to 750
Orpm, centrifugation was performed for 30 minutes to obtain a supernatant. The same operation as above was repeated on the precipitate, the upper layer was combined, and 136.3 g of the dried product obtained by freeze-drying was dissolved in methanol,
After refluxing, centrifugation, and dissolving the precipitate in purified water, it was dialyzed against running water for 3 days and purified water for 2 days. The non-dialyzed fraction was lyophilized to obtain crude fraction 35.39.
この粗分画のうちＩ５．８９を精製水に溶解し、同量の
８％臭化セチル−トリメチル−アンモニウムを加え、混
和して２０℃で４時間放置した。この溶液を遠心分離し
、沈澱物をｌＯ％塩化ナトリウム水溶液に溶解し、更に
エタノールを加えて遠心分離した。得られた沈澱物を精
製水に溶解後、流水に対して３日間、精製水に対して２
日間透析し、得られた非透析分画を凍結乾燥して多糖含
有分画３．５４９を得た。Of this crude fraction, I5.89 was dissolved in purified water, the same amount of 8% cetyl-trimethyl-ammonium bromide was added, mixed and left at 20°C for 4 hours. This solution was centrifuged, and the precipitate was dissolved in 10% sodium chloride aqueous solution, and ethanol was added and centrifuged. After dissolving the obtained precipitate in purified water, dissolve it in running water for 3 days and in purified water for 2 days.
Dialysis was carried out for several days, and the resulting non-dialyzed fraction was freeze-dried to obtain a polysaccharide-containing fraction of 3.549.
次に、多糖含有分画５００■を精製水に溶解し、精製水
で平衡化したＤＥＡＥ−セファロースＣＬ−６Ｂカラム
に吸着させ、ギ酸アンモニウムを溶媒として、順次濃度
を変えて溶出させた。０．３Ｍギ酸アンモニウムで°溶
出した溶出液を濃縮し、更に０．２Ｍ塩化ナトリウムで
平衡化したセファロースＣＬ−６Ｂカラムでゲル濾過を
行い白色繊維状の物質を得た。Next, 500 μl of the polysaccharide-containing fraction was dissolved in purified water, adsorbed on a DEAE-Sepharose CL-6B column equilibrated with purified water, and eluted using ammonium formate as a solvent at successively different concentrations. The eluate eluted with 0.3M ammonium formate was concentrated and gel filtrated using a Sepharose CL-6B column equilibrated with 0.2M sodium chloride to obtain a white fibrous substance.
次に本発明の多糖体が、抗補体活性を有し、慢性腎炎等
の自己免疫疾患の治療に有効であることを実験例を挙げ
て説明する。Next, it will be explained using experimental examples that the polysaccharide of the present invention has anti-complement activity and is effective in treating autoimmune diseases such as chronic nephritis.
実験例
具体例で得た多糖体１〜ｌＯ■を生理食塩水に溶解し、
そのうち５０／ｉｌを、５０−のＭｇ”および１５ （
ＩＭのＣａ”を含むゼラチン添加ベロナール緩衝液（ｐ
Ｈ７，４，ＧＶＢ”）で希釈し、更に正常ヒト血清を添
加して、３７°Ｃで３０分間インキュベートした。Experimental Example Dissolve 1 to 1 O of the polysaccharide obtained in the specific example in physiological saline,
Of this, 50/il was added to 50-Mg" and 15 (
Gelatin-added veronal buffer (p
H7,4,GVB"), normal human serum was added, and the mixture was incubated at 37°C for 30 minutes.
この溶液を２倍連続希釈し、試験管に分注し、一定量の
ＩｇＭ−ヘモリノン感作ヒツジ赤血球を加え、更に３７
℃で６０分間、混和しながら反応させた。This solution was serially diluted 2 times, dispensed into test tubes, added with a certain amount of IgM-hemolinone sensitized sheep red blood cells, and further
The reaction was carried out at ℃ for 60 minutes with mixing.
反応終了後、直ちにリン酸−生理食塩水緩衝液（ＰＢＳ
−）を加え、２５００ ｒｐｍで８分間遠心分離し、溶
血反応に基づく波長４１２ｎｍでの上清の吸光度より、
総残存補体価（Ｔ ＣＨｓ。）を測定した。Immediately after the reaction is complete, add phosphate-physiological saline buffer (PBS).
-) and centrifuged at 2500 rpm for 8 minutes. From the absorbance of the supernatant at a wavelength of 412 nm based on the hemolytic reaction,
Total residual complement values (TCHs.) were measured.
具体例で得た多糖体に加えない以外は、上記と同様に反
応させた場合のＴ ＣＨ５ｏを１００として、本発明の
多糖体によるＴＣＨ５゜の減少率をもって抗補体活性と
した。The anti-complement activity was defined as the rate of decrease in TCH5° by the polysaccharide of the present invention, assuming that TCH5o was 100 when the reaction was carried out in the same manner as above, except that it was not added to the polysaccharide obtained in the specific example.
その結果、本発明の多糖体の抗補体活性は、８０−１０
０％であった。As a result, the anti-complement activity of the polysaccharide of the present invention was 80-10
It was 0%.
本発明の多糖体はそのまま、あるいは慣用の製剤担体と
共に動物および人に投与することができる。投与形態と
しては、特に限定かなく、必要に応じ適宜選択して使用
され、錠剤、カプセル剤、顆粒剤等の経口剤、注射剤、
坐剤等の非経口剤が挙げられる。The polysaccharide of the present invention can be administered to animals and humans as it is or together with a conventional pharmaceutical carrier. The dosage form is not particularly limited and may be selected and used as required, including oral preparations such as tablets, capsules, and granules, injections,
Examples include parenteral preparations such as suppositories.
錠剤、カプセル剤、顆粒剤等の経口剤は常法に従って製
造される。錠剤は本発明の多糖体をゼラチン、でん粉、
乳糖、ステアリン酸マグネシウム、滑石、アラビアゴム
等の製剤学的賦形剤と混合し賦形することによりつくら
れ、カプセル剤は、上記化合物を不活性の製剤充填剤、
もしくは希釈剤と混合し、硬質ゼラチンカプセル、軟質
ゼラチンカプセル等に充填することによりつくられる。Oral preparations such as tablets, capsules, and granules are manufactured according to conventional methods. The tablet contains the polysaccharide of the present invention, gelatin, starch,
Capsules are made by mixing and shaping the above compounds with pharmaceutical excipients such as lactose, magnesium stearate, talc, gum arabic, etc.
Alternatively, it can be prepared by mixing it with a diluent and filling it into hard gelatin capsules, soft gelatin capsules, etc.
シロップ剤、エリキシル剤は、本発明の多糖体をショ糖
等の甘味剤、メチルパラベンおよびプロピルパラベン類
等の防腐剤、着色剤、調味剤、芳香剤、補助剤と混合し
て製造される。Syrups and elixirs are produced by mixing the polysaccharide of the present invention with sweeteners such as sucrose, preservatives such as methylparabens and propylparabens, colorants, seasonings, fragrances, and adjuvants.
非経口剤は常法に従って製造され、希釈剤として一般に
注射用蒸留水、生理食塩水、デキストロース水溶液、プ
ロピレングリコール等を用いることができる。さらに必
要に応じて、殺菌剤、防腐剤、安定剤を加えてもよい。Parenteral preparations are manufactured according to conventional methods, and distilled water for injection, physiological saline, aqueous dextrose solution, propylene glycol, etc. can generally be used as diluents. Furthermore, a bactericide, a preservative, and a stabilizer may be added as necessary.
また、この非経口剤は安定性の点から、カプセル等に充
填後冷凍し、通常の凍結乾燥技術により水分を除去し、
使用直前に凍結乾燥物から液剤を再調製することもてき
る。In addition, from the viewpoint of stability, this parenteral preparation is frozen after being filled into capsules, etc., and the water is removed using normal freeze-drying technology.
It is also possible to reconstitute the solution from the lyophilizate immediately before use.
特許出願人 北里研究所（社団法人）代表者 水之江
公英Patent applicant: Kitasato Research Institute (Incorporated Association) Representative: Kimihide Mizunoe
Claims (1)
性質 ［１］性状：白色繊維状 ［２］分子量：３６，０００（ゲル濾過クロマトグラフ
ィー）、 ［３］比旋光度：［α］＾２＾０＿Ｄ＝＋１２０、［４
］構成糖：ラムノース、アラビノース、ガラクトース、
ガラクツロン酸、 ［５］単一性：電気泳動、ゲル濾過クロマトグラフィー
、高速液体クロマトグラフィーにより単一性を示す、 ［６］構成糖結合様式： １−５結合アラビノフラノース １．４％ 末端ガラクトース ２．７％ 末端ガラクツロン酸 ２．７％ ３−５分岐アラビノフラノース ２．８％ ２−４分岐ラムノース ２．２％ １−４結合ガラクツロン酸 ８０．０％ １−６結合ガラクトース １．９％ １−４結合ガラクトース ４．２％ ３−６分岐ガラクトース ０．７％ を有する多糖体。[Claims] Obtained from the root of Apiaceae, it has the following physical and chemical properties: [1] Properties: white fibrous [2] Molecular weight: 36,000 (gel filtration chromatography), [3] Specific optical rotation : [α]^2^0_D=+120, [4
] Constituent sugars: rhamnose, arabinose, galactose,
Galacturonic acid, [5] Unity: Shows unity by electrophoresis, gel filtration chromatography, high performance liquid chromatography, [6] Constituent sugar binding mode: 1-5 linked arabinofuranose 1.4% terminal galactose 2.7% Terminal galacturonic acid 2.7% 3-5 branched arabinofuranose 2.8% 2-4 branched rhamnose 2.2% 1-4 linked galacturonic acid 80.0% 1-6 linked galactose 1.9% Polysaccharide having 4.2% 1-4-linked galactose and 0.7% 3-6-branched galactose.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP23583686A JPH0753762B2 (en) | 1986-10-03 | 1986-10-03 | Novel polysaccharide |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP23583686A JPH0753762B2 (en) | 1986-10-03 | 1986-10-03 | Novel polysaccharide |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPS6390505A true JPS6390505A (en) | 1988-04-21 |
| JPH0753762B2 JPH0753762B2 (en) | 1995-06-07 |
Family
ID=16991981
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP23583686A Expired - Lifetime JPH0753762B2 (en) | 1986-10-03 | 1986-10-03 | Novel polysaccharide |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH0753762B2 (en) |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5166196A (en) * | 1989-05-12 | 1992-11-24 | Kao Corporation | Method for removing immunocomplexes from blood |
| EP0635519A1 (en) * | 1993-07-15 | 1995-01-25 | JCR PHARMACEUTICALS Co., LTD. | Polyglucuronic acid as remitting agent for nephrotic syndrome and hepatopathy symptoms |
| CN101700342A (en) * | 2009-10-30 | 2010-05-05 | 杨雄志 | Pharmaceutical composition for treating hepatitis B and preparation method thereof |
| CN103739733A (en) * | 2014-01-09 | 2014-04-23 | 南京农业大学 | Anti-duck virus hepatitis (DVH) subprostrate sophora polysaccharide and molecular modifier thereof |
| CN110498865A (en) * | 2018-05-17 | 2019-11-26 | 复旦大学 | Big fruit Chinese juniper polysaccharide and preparation method thereof and preparing the purposes in anticomplement medicament |
| CN110498864A (en) * | 2018-05-17 | 2019-11-26 | 复旦大学 | Arborvitae polysaccharide and preparation method thereof and preparing the purposes in anticomplement medicament |
| CN110498863A (en) * | 2018-05-17 | 2019-11-26 | 复旦大学 | Climbing fig leaf polyose and preparation method thereof and preparing the purposes in anticomplement medicament |
-
1986
- 1986-10-03 JP JP23583686A patent/JPH0753762B2/en not_active Expired - Lifetime
Cited By (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5166196A (en) * | 1989-05-12 | 1992-11-24 | Kao Corporation | Method for removing immunocomplexes from blood |
| EP0635519A1 (en) * | 1993-07-15 | 1995-01-25 | JCR PHARMACEUTICALS Co., LTD. | Polyglucuronic acid as remitting agent for nephrotic syndrome and hepatopathy symptoms |
| AU686161B2 (en) * | 1993-07-15 | 1998-02-05 | Jcr Pharmaceuticals Co., Ltd. | Remitting agent for nephrotic syndrome and hepatopathy symptoms |
| CN101700342A (en) * | 2009-10-30 | 2010-05-05 | 杨雄志 | Pharmaceutical composition for treating hepatitis B and preparation method thereof |
| CN103739733A (en) * | 2014-01-09 | 2014-04-23 | 南京农业大学 | Anti-duck virus hepatitis (DVH) subprostrate sophora polysaccharide and molecular modifier thereof |
| CN110498865A (en) * | 2018-05-17 | 2019-11-26 | 复旦大学 | Big fruit Chinese juniper polysaccharide and preparation method thereof and preparing the purposes in anticomplement medicament |
| CN110498864A (en) * | 2018-05-17 | 2019-11-26 | 复旦大学 | Arborvitae polysaccharide and preparation method thereof and preparing the purposes in anticomplement medicament |
| CN110498863A (en) * | 2018-05-17 | 2019-11-26 | 复旦大学 | Climbing fig leaf polyose and preparation method thereof and preparing the purposes in anticomplement medicament |
| CN110498865B (en) * | 2018-05-17 | 2021-09-07 | 复旦大学 | Selaginella maxima polysaccharide, preparation method thereof and application thereof in preparation of anticomplement medicines |
| CN110498863B (en) * | 2018-05-17 | 2021-09-07 | 复旦大学 | Ficus pumila leaf polysaccharide, preparation method thereof and application thereof in preparation of anticomplement medicines |
| CN110498864B (en) * | 2018-05-17 | 2021-09-07 | 复旦大学 | Selaginella polysaccharide, preparation method thereof and application thereof in preparation of anticomplement medicines |
Also Published As
| Publication number | Publication date |
|---|---|
| JPH0753762B2 (en) | 1995-06-07 |
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