JPH0546502B2 - - Google Patents
Info
- Publication number
- JPH0546502B2 JPH0546502B2 JP59031794A JP3179484A JPH0546502B2 JP H0546502 B2 JPH0546502 B2 JP H0546502B2 JP 59031794 A JP59031794 A JP 59031794A JP 3179484 A JP3179484 A JP 3179484A JP H0546502 B2 JPH0546502 B2 JP H0546502B2
- Authority
- JP
- Japan
- Prior art keywords
- blood
- blood coagulation
- compound
- residue
- present
- 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
- 230000023555 blood coagulation Effects 0.000 claims description 32
- 125000002915 carbonyl group Chemical group [*:2]C([*:1])=O 0.000 claims description 11
- 150000001923 cyclic compounds Chemical class 0.000 claims description 10
- -1 cyclic organic compound Chemical class 0.000 claims description 9
- 108010039209 Blood Coagulation Factors Proteins 0.000 claims description 7
- 102000015081 Blood Coagulation Factors Human genes 0.000 claims description 7
- 239000003114 blood coagulation factor Substances 0.000 claims description 7
- 229940019700 blood coagulation factors Drugs 0.000 claims description 7
- 102000004190 Enzymes Human genes 0.000 claims description 6
- 108090000790 Enzymes Proteins 0.000 claims description 6
- 239000012190 activator Substances 0.000 claims description 6
- 230000002255 enzymatic effect Effects 0.000 claims description 4
- 125000003275 alpha amino acid group Chemical group 0.000 claims description 3
- 125000000539 amino acid group Chemical group 0.000 claims description 3
- 230000003301 hydrolyzing effect Effects 0.000 claims 1
- 210000004369 blood Anatomy 0.000 description 21
- 239000008280 blood Substances 0.000 description 21
- 210000002966 serum Anatomy 0.000 description 14
- 238000009534 blood test Methods 0.000 description 11
- 238000012360 testing method Methods 0.000 description 10
- 150000001875 compounds Chemical class 0.000 description 8
- 108091005804 Peptidases Proteins 0.000 description 6
- 239000004365 Protease Substances 0.000 description 6
- 208000007536 Thrombosis Diseases 0.000 description 5
- 238000000926 separation method Methods 0.000 description 5
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 description 4
- 230000000694 effects Effects 0.000 description 4
- 229940088598 enzyme Drugs 0.000 description 4
- 125000004435 hydrogen atom Chemical group [H]* 0.000 description 4
- 125000001424 substituent group Chemical group 0.000 description 4
- WOAHJDHKFWSLKE-UHFFFAOYSA-N 1,2-benzoquinone Chemical group O=C1C=CC=CC1=O WOAHJDHKFWSLKE-UHFFFAOYSA-N 0.000 description 3
- AFSDNFLWKVMVRB-UHFFFAOYSA-N Ellagic acid Chemical compound OC1=C(O)C(OC2=O)=C3C4=C2C=C(O)C(O)=C4OC(=O)C3=C1 AFSDNFLWKVMVRB-UHFFFAOYSA-N 0.000 description 3
- ATJXMQHAMYVHRX-CPCISQLKSA-N Ellagic acid Natural products OC1=C(O)[C@H]2OC(=O)c3cc(O)c(O)c4OC(=O)C(=C1)[C@H]2c34 ATJXMQHAMYVHRX-CPCISQLKSA-N 0.000 description 3
- 229920002079 Ellagic acid Polymers 0.000 description 3
- 108090000190 Thrombin Proteins 0.000 description 3
- 108090000631 Trypsin Proteins 0.000 description 3
- 102000004142 Trypsin Human genes 0.000 description 3
- 230000015271 coagulation Effects 0.000 description 3
- 238000005345 coagulation Methods 0.000 description 3
- 239000006185 dispersion Substances 0.000 description 3
- 235000004132 ellagic acid Nutrition 0.000 description 3
- 229960002852 ellagic acid Drugs 0.000 description 3
- 239000011521 glass Substances 0.000 description 3
- 125000001183 hydrocarbyl group Chemical group 0.000 description 3
- FAARLWTXUUQFSN-UHFFFAOYSA-N methylellagic acid Natural products O1C(=O)C2=CC(O)=C(O)C3=C2C2=C1C(OC)=C(O)C=C2C(=O)O3 FAARLWTXUUQFSN-UHFFFAOYSA-N 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 239000003998 snake venom Substances 0.000 description 3
- 229960004072 thrombin Drugs 0.000 description 3
- 239000012588 trypsin Substances 0.000 description 3
- 206010053567 Coagulopathies Diseases 0.000 description 2
- 102000004157 Hydrolases Human genes 0.000 description 2
- 108090000604 Hydrolases Proteins 0.000 description 2
- 102000035195 Peptidases Human genes 0.000 description 2
- 108010022999 Serine Proteases Proteins 0.000 description 2
- 102000012479 Serine Proteases Human genes 0.000 description 2
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 2
- 125000000217 alkyl group Chemical group 0.000 description 2
- 230000035602 clotting Effects 0.000 description 2
- 230000001112 coagulating effect Effects 0.000 description 2
- 230000000052 comparative effect Effects 0.000 description 2
- 150000002391 heterocyclic compounds Chemical class 0.000 description 2
- 229910052739 hydrogen Inorganic materials 0.000 description 2
- 239000001257 hydrogen Substances 0.000 description 2
- JXDYKVIHCLTXOP-UHFFFAOYSA-N isatin Chemical compound C1=CC=C2C(=O)C(=O)NC2=C1 JXDYKVIHCLTXOP-UHFFFAOYSA-N 0.000 description 2
- 230000001737 promoting effect Effects 0.000 description 2
- 230000002345 thrombinlike Effects 0.000 description 2
- 239000005995 Aluminium silicate Substances 0.000 description 1
- 101000772006 Bombus ignitus Venom serine protease Bi-VSP Proteins 0.000 description 1
- 239000004215 Carbon black (E152) Substances 0.000 description 1
- 108090000712 Cathepsin B Proteins 0.000 description 1
- 102000004225 Cathepsin B Human genes 0.000 description 1
- 102000009123 Fibrin Human genes 0.000 description 1
- 108010073385 Fibrin Proteins 0.000 description 1
- BWGVNKXGVNDBDI-UHFFFAOYSA-N Fibrin monomer Chemical compound CNC(=O)CNC(=O)CN BWGVNKXGVNDBDI-UHFFFAOYSA-N 0.000 description 1
- 229920000084 Gum arabic Polymers 0.000 description 1
- 241000519695 Ilex integra Species 0.000 description 1
- 102000005741 Metalloproteases Human genes 0.000 description 1
- 108010006035 Metalloproteases Proteins 0.000 description 1
- 239000004698 Polyethylene Substances 0.000 description 1
- 241000978776 Senegalia senegal Species 0.000 description 1
- 101710097834 Thiol protease Proteins 0.000 description 1
- 206010052428 Wound Diseases 0.000 description 1
- 208000027418 Wounds and injury Diseases 0.000 description 1
- 235000010489 acacia gum Nutrition 0.000 description 1
- 239000000205 acacia gum Substances 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 125000002723 alicyclic group Chemical group 0.000 description 1
- 235000012211 aluminium silicate Nutrition 0.000 description 1
- 125000003277 amino group Chemical group 0.000 description 1
- LNTHITQWFMADLM-UHFFFAOYSA-N anhydrous gallic acid Natural products OC(=O)C1=CC(O)=C(O)C(O)=C1 LNTHITQWFMADLM-UHFFFAOYSA-N 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 239000012752 auxiliary agent Substances 0.000 description 1
- 239000000440 bentonite Substances 0.000 description 1
- 229910000278 bentonite Inorganic materials 0.000 description 1
- SVPXDRXYRYOSEX-UHFFFAOYSA-N bentoquatam Chemical compound O.O=[Si]=O.O=[Al]O[Al]=O SVPXDRXYRYOSEX-UHFFFAOYSA-N 0.000 description 1
- 230000000740 bleeding effect Effects 0.000 description 1
- 210000000601 blood cell Anatomy 0.000 description 1
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 1
- 125000003262 carboxylic acid ester group Chemical group [H]C([H])([*:2])OC(=O)C([H])([H])[*:1] 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000011109 contamination Methods 0.000 description 1
- 230000008602 contraction Effects 0.000 description 1
- 125000004122 cyclic group Chemical group 0.000 description 1
- OILAIQUEIWYQPH-UHFFFAOYSA-N cyclohexane-1,2-dione Chemical compound O=C1CCCCC1=O OILAIQUEIWYQPH-UHFFFAOYSA-N 0.000 description 1
- 230000006806 disease prevention Effects 0.000 description 1
- 238000013399 early diagnosis Methods 0.000 description 1
- 229950003499 fibrin Drugs 0.000 description 1
- 238000004108 freeze drying Methods 0.000 description 1
- 235000004515 gallic acid Nutrition 0.000 description 1
- 229940074391 gallic acid Drugs 0.000 description 1
- 230000005484 gravity Effects 0.000 description 1
- 230000009931 harmful effect Effects 0.000 description 1
- 230000023597 hemostasis Effects 0.000 description 1
- 229930195733 hydrocarbon Natural products 0.000 description 1
- 150000002430 hydrocarbons Chemical class 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- NLYAJNPCOHFWQQ-UHFFFAOYSA-N kaolin Chemical compound O.O.O=[Al]O[Si](=O)O[Si](=O)O[Al]=O NLYAJNPCOHFWQQ-UHFFFAOYSA-N 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000010534 mechanism of action Effects 0.000 description 1
- 239000004745 nonwoven fabric Substances 0.000 description 1
- HCZKYJDFEPMADG-UHFFFAOYSA-N nordihydroguaiaretic acid Chemical compound C=1C=C(O)C(O)=CC=1CC(C)C(C)CC1=CC=C(O)C(O)=C1 HCZKYJDFEPMADG-UHFFFAOYSA-N 0.000 description 1
- 239000013618 particulate matter Substances 0.000 description 1
- 229920000573 polyethylene Polymers 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 235000004252 protein component Nutrition 0.000 description 1
- 235000018102 proteins Nutrition 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 229920005989 resin Polymers 0.000 description 1
- 239000011347 resin Substances 0.000 description 1
- 239000012508 resin bead Substances 0.000 description 1
- 239000000377 silicon dioxide Substances 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 229920003002 synthetic resin Polymers 0.000 description 1
- 239000000057 synthetic resin Substances 0.000 description 1
- 125000003396 thiol group Chemical group [H]S* 0.000 description 1
- 239000002759 woven fabric Substances 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/86—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving blood coagulating time or factors, or their receptors
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Hematology (AREA)
- Engineering & Computer Science (AREA)
- Molecular Biology (AREA)
- Biomedical Technology (AREA)
- Chemical & Material Sciences (AREA)
- Immunology (AREA)
- Urology & Nephrology (AREA)
- Biotechnology (AREA)
- Biochemistry (AREA)
- Cell Biology (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- Microbiology (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Description
（技術分野）
本発明は、血液凝固促進剤に関する。
（従来技術）
近年、検査技術の目覚しい進歩と相俟つて、血
清生化学検査、血清免疫学検査、血球検査等の血
液検査が広く普及し、病気予防や早期診断に大き
く貢献するに至つている。なかでも血清検査は血
液検査の主体をなしており、この検査において必
要な血清は、通常、血液を血液検査用容器に採取
し、これを凝固させた後、遠心分離によつて比重
の異なる血餅、即ち、フイブリンと血球が混合し
たゲル様塊状物を分離させ、血清部分をピペツト
で吸い上げたり、或いはデカンテーシヨンして採
取している。
しかしながら、一般に血液は凝固するまでにか
なりの時間を要し、従来、迅速に検査を実施する
ことが困難である。最も血液凝固時間が短いとさ
れているガラス製血液検査用容器でさえ、血液を
注入した後、凝固に至るまでに40分乃至60分を必
要とし、合成樹脂製血液検査用容器に至つては、
血液凝固までに４時間以上の放置を必要とする。
このため、血液凝固第因子活性化能を有
し、血液凝固を促進するガラス、カオリン、ベン
トナイト、シリカ、エラジン酸等が血清検査にお
ける血液凝固促進剤としてのほか、血液の凝固機
能検査の一つである活性化部分トロンボプラスミ
ン時間の測定試薬の一成分として実用に供されて
いるが、その純度や組成等によつてその活性化能
が安定しない問題がある。
更に、これらの血液凝固促進剤による場合、一
般に血液凝固後に血清から血餅を遠心分離し、血
清を採取する際に血清の分離性がよくなく、血清
に血餅成分が混入するのを避けられない。
（発明の目的）
本発明は上記した問題を解決するためにされた
ものであつて、因子を活性化させ、血液凝固
に要する時間を大幅に短縮させると共に、その血
液凝固の促進効果が極めて安定しており、更に、
血清の分離性にもすぐれた血液凝固促進剤を提供
することを目的とするものである。
（発明の構成）
本発明の血液凝固促進剤は、血液凝固第因
子の非酵素的活性化剤と、アミノ酸配列において
Arg又はLys残基と任意のアミノ酸残基との間の
結合の加水分解酵素とを含有することを特徴とす
るものである。
本発明による血液凝固促進剤における一成分で
ある非酵素的活性化剤としては、
一般式
（但し、Ａは環式化合物の残基を示す。）
で表わされ、且つ、上記二つの相隣るカルボニル
基が立体的に実質的に同一の平面上にある環式有
機化合物が用いられる。
上記環式有機化合物は、二つの相隣るカルボニ
ル基と残基Ａとが形成する同素環式又は異節環式
化合物のいずれであつてもよく、また、このよう
な環式化合物は単環式であつても、多環式化合物
であつてもよいが、少なくとも二つのカルボニル
炭素を含む環が６員環又は５員環である環式化合
物である。
特に、好ましい６員環式化合物は次式
（但し、R1，R2，R3及びR4は水素基、炭化水
素、極性置換基又は多環式化合物における残基を
示す。）
で表わされるｏ−キノン環を有する化合物であ
る。上記式において炭化水素基は特に制限される
ものではないが、好ましくはアルキル基であり、
また、上記極性置換基も特に制限されるものでは
ないが、例えば、カルボキシル基、カルボン酸エ
ステル基、水酸基、アミノ基、メルカプト基等で
ある。従つて、ｏ−キノン環を有する化合物の好
ましい具体例として、例えば、ｏ−キノン、次の
一般式
（但し、R5はアルキル基を示す。）
で表わされる没食子酸アルキルエステル酸化物、
次式
でそれぞれ表わされるエラジン酸部分酸化物及び
完全酸化物、次式
で表わされる１，４−ジ（３，４−ジヒドロキシ
フエニル）２，３−ジメチルブタン部分酸化物及
び完全酸化物等を挙げることができる。
また、二つのカルボニル炭素を含む環が５員環
である同素環式化合物の好ましい具体例として、
次式
で表わされる１，２，３−トリケトヒドロインデ
ンを挙げることができる。
更に、上記環式化合物として好ましく用いるこ
とができる異節環式化合物の一つは、次の一般式
（但し、R6は水素、炭化水素基又は多環式化合
物における残基を示し、R7及びR8は水素、炭化
水素基、極性置換基又は多環式化合物における残
基を示す。）
で表わされる、ここに、上記炭化存水素基及び極
性置換基については前記と同じである。
このような化合物の好ましい具体例として、例
えば、次式で表わされるイサチンを挙げることが
できる。
これら一群の化合物は、いずれも分子内に立体
的に同一の乃至は近似的に同一の平面上に二つの
相隣るカルボニル基をもち、詳細な作用機序は不
明であるが、血液凝固因子に対して特異的な効果
を示す。一方、同じく分子内に相隣るカルボニル
基をもちながら、それらが同一平面上にないため
に血液凝固促進剤機能をもたない化合物として
は、例えば次式で表わされる１，２−ジケトシク
ロヘキサンがある。
この場合、二つの相隣るカルボニル基をつなぐ
脂環はかなりのフレキシビリテイーを有し、その
ためにこれらカルボニル基同士は立体反発によつ
て相互にゴーシユの位置にあり、同一平面上にな
い。そして、この化合物は血液凝固因子に対して
何ら特異的な効果を示さない。これらの事実はお
そらく立体的な特別な位置にある二つの相隣るカ
ルボニル基がタンパク質である血液凝固因子のあ
る特定の部位と立体構造的に特殊な関係をもつた
めであろうと推測される。
次に、本発明による血液凝固促進剤の他の成分
である加水分解酵素は、アミノ酸配列において
Arg又はLys残基と任意のアミノ酸残基との間の
結合の加水分解酵素であり、このような加水分解
酵素としては、特にプロテアーゼが好ましく用い
られる。このプロテアーゼの具体例として、例え
ば、トリプシン、トロンビン、ヘビ毒トロンビン
様酵素等のセリンプロテアーゼ、カテプシンＢ、
フイシン等のチオールプロテアーゼ、キニナーゼ
のような金属プロテアーゼ等を挙げることがで
きるが、セリンプロテアーゼが入手容易でもある
ので、使用するのに好適である。これらプロテア
ーゼは、単独でも血液凝固を活性化し得るが、本
発明に従つて、前記した非酵素的活性化剤、特
に、前記環式化合物からなる因子活性化剤と
併用することによつて、血液凝固の活性化能が飛
躍的に向上するのである。
本発明の血液凝固促進剤の使用においては、例
えば、血液を血液検査用容器に採取し、これを凝
固させる際に促進剤を血液中に存在させる。この
場合、上記血液凝固促進剤は、これをそのままの
粉末状で血液中に存在させてもよいが、好ましく
は、血液凝固促進剤を適宜の溶剤に溶解若しくは
分散させて、血液中に添加する。上記血液検査用
容器は特に制限されず、従来より通常に用いられ
ているガラス製又は樹脂製の容器が適宜に用いら
れる。
尚、血液が血液検査用容器内において瞬間的
に、又は部分的に高濃度のこれら血液凝固促進剤
と接触し、血液中のタンパク質成分が変質するお
それがあるときは、比表面積の大きい担体に血液
凝固促進剤を担持させ、これを血液検査用容器中
の血液に添加してもよい。上記担体としては、血
液検査に有害な影響を与えず、大きい比表面積を
有するものであれば、特に制限されることなく、
種々のものを用いることができるが、例えば、不
織布、織布、樹脂ビーズ等を好適に用いることが
できる。このような担体に血液凝固促進剤を担持
させるには、例えば、その溶液や分散液を塗布
し、又はこれに浸漬した後、乾燥して、担体に付
着させればよい。また、アラビアゴム等の適宜の
助剤と混合して水分散液とし、これを急速凍結乾
燥する等の方法により、血液凝固促進剤を担持し
た粒子状物を得ることもできる。
血液凝固促進剤の血液中における存在量は、血
液１mlについて少なくとも１×10-10ｇであり、
これよりも少ないときは、血液凝固の促進効果が
乏しい。しかし、余り多量に存在させるときは、
却つて血液検査に種々の支障を来すおそれがある
ので、10-1ｇ以下とするのが好ましい。
（発明の効果）
本発明の血液凝固促進剤によれば、これを血液
中に存在させるとき、因子が迅速に活性化さ
れ、容器に血液を採取後の凝固に要する時間が著
しく短縮されると共に、血餅成分の収縮が十分に
行なわれる結果、血清と血餅との分離性にすぐ
れ、分離採取した血清に血餅成分が混入すること
がなく、更に、血清の収量も著しく増大する。従
つ、本発明の血液凝固促進剤は、臨床検査分野に
おいて広く用いることができるほか、出血創の止
血等にも使用することができる。
以下に実施例を挙げて本発明を説明するが、本
発明はこれら実施例により何ら限定されるもので
はない。
（実施例）
実施例 １
血液凝固第因子活性化剤としての前記環式
化合物として、エラグ酸酸化物、１，２，３−ト
リケトヒドロインデンを、また、プロテアーゼと
してトリプシン、トロンビン及び蛇毒トロンビン
様酵素をそれぞれ用いて、本発明による血液凝固
促進剤水溶液を調製した。尚、各血液凝固促進剤
における各成分の含有量は、環式化合物が0.5重
量％、プロテアーゼは、トリプシン、トロンビン
及び蛇毒トロンビン様酵素がそれぞれについて
0.05重量％、500単位／ml及び0.005重量％とし
た。
市販のポリエチレンプレーンスピツツを用い
て、本発明による血液凝固促進剤30μを人新鮮
血３mlに加え、血液が流動性を失なうまでに要し
た時間を凝固時間として測定し、また、凝固後、
3000回転／分で５分間遠心分離して、分離状態を
観察した。
結果を第１表に示す。
比較例 １
比較のために、実施例１で用いた環式化合物及
びプロテアーゼをそれぞれ単独で用いた場合の凝
固時間及び分離状態を第２表に示す。
比較例 ２
血液凝固促進剤を用いないほかは、実施例１と
同様に血液処理したときの凝固時間及び分離状態
を第３表に示す。
(Technical Field) The present invention relates to a blood coagulation promoter. (Prior art) In recent years, with the remarkable progress in testing technology, blood tests such as serum biochemistry tests, serum immunology tests, and hematology tests have become widespread, and have greatly contributed to disease prevention and early diagnosis. . Among these, serum tests are the main body of blood tests, and the serum required for this test is usually obtained by collecting blood into a blood test container, coagulating it, and then centrifuging it to collect blood with different specific gravities. The mochi, that is, a gel-like mass containing a mixture of fibrin and blood cells, is separated, and the serum portion is collected by pipetting or decanting. However, blood generally takes a considerable amount of time to coagulate, making it difficult to conduct tests quickly. Even with glass blood test containers, which are said to have the shortest blood coagulation time, it takes 40 to 60 minutes for blood to coagulate after blood is injected, and synthetic resin blood test containers do not. ,
It takes 4 hours or more for blood to coagulate. For this reason, glass, kaolin, bentonite, silica, ellagic acid, etc., which have the ability to activate blood coagulation factor and promote blood coagulation, are used as blood coagulation promoters in serum tests, as well as in blood coagulation function tests. Although it has been put to practical use as a component of a reagent for measuring the activated partial thromboplasmin time, there is a problem that its activation ability is unstable depending on its purity, composition, etc. Furthermore, when these blood coagulation promoters are used, blood clots are generally centrifuged from serum after blood coagulation, and serum separation is not good when serum is collected, making it difficult to avoid contamination of blood clot components with serum. do not have. (Objective of the Invention) The present invention has been made to solve the above-mentioned problems, and it activates factors, significantly shortens the time required for blood coagulation, and has an extremely stable blood coagulation promoting effect. In addition,
The object of the present invention is to provide a blood coagulation promoter that also has excellent serum separation properties. (Structure of the Invention) The blood coagulation promoter of the present invention comprises a non-enzymatic activator of blood coagulation factor and an amino acid sequence of
It is characterized by containing an enzyme that hydrolyzes the bond between Arg or Lys residue and any amino acid residue. The non-enzymatic activator, which is one component of the blood coagulation promoter according to the present invention, has the general formula (However, A represents a residue of a cyclic compound.) A cyclic organic compound is used in which the above two adjacent carbonyl groups are sterically located on substantially the same plane. . The above-mentioned cyclic organic compound may be either a homocyclic or a heterocyclic compound formed by two adjacent carbonyl groups and the residue A, and such a cyclic compound may be a monocyclic compound. Although it may be a cyclic or polycyclic compound, it is a cyclic compound in which the ring containing at least two carbonyl carbons is a 6-membered ring or a 5-membered ring. Particularly preferred six-membered cyclic compounds are of the following formula: (However, R 1 , R 2 , R 3 and R 4 represent a hydrogen group, a hydrocarbon, a polar substituent, or a residue in a polycyclic compound.) It is a compound having an o-quinone ring represented by: In the above formula, the hydrocarbon group is not particularly limited, but is preferably an alkyl group,
Furthermore, the polar substituents mentioned above are not particularly limited, but include, for example, a carboxyl group, a carboxylic acid ester group, a hydroxyl group, an amino group, a mercapto group, and the like. Therefore, as a preferable specific example of a compound having an o-quinone ring, for example, o-quinone, the compound having the following general formula (However, R 5 represents an alkyl group.) Gallic acid alkyl ester oxide represented by
The following formula ellagic acid partial oxide and complete oxide, respectively represented by the following formula: Examples include 1,4-di(3,4-dihydroxyphenyl)2,3-dimethylbutane partial oxide and complete oxide represented by: Further, as a preferable specific example of a homocyclic compound in which the ring containing two carbonyl carbons is a 5-membered ring,
The following formula 1,2,3-triketohydroindene represented by: Furthermore, one of the heterocyclic compounds that can be preferably used as the above-mentioned cyclic compound has the following general formula: (However, R 6 represents hydrogen, a hydrocarbon group, or a residue in a polycyclic compound, and R 7 and R 8 represent hydrogen, a hydrocarbon group, a polar substituent, or a residue in a polycyclic compound.) Here, the hydrocarbon-existing hydrogen group and polar substituent are the same as described above. A preferred specific example of such a compound is isatin represented by the following formula. All of these group of compounds have two adjacent carbonyl groups on the same or approximately the same steric plane within the molecule, and although the detailed mechanism of action is unknown, blood coagulation factors It shows a specific effect on. On the other hand, examples of compounds that have carbonyl groups adjacent to each other in the molecule but do not have a blood coagulation promoter function because they are not on the same plane include 1,2-diketocyclohexane represented by the following formula. There is. In this case, the alicyclic ring connecting two adjacent carbonyl groups has considerable flexibility, and therefore these carbonyl groups are in mutually gauche positions due to steric repulsion and are not on the same plane. This compound does not exhibit any specific effect on blood coagulation factors. It is speculated that these facts are probably due to the fact that two adjacent carbonyl groups in special steric positions have a special steric structure with a specific site of a blood coagulation factor, which is a protein. Next, the hydrolase, which is another component of the blood coagulation promoter according to the present invention, has an amino acid sequence of
It is an enzyme that hydrolyzes the bond between an Arg or Lys residue and any amino acid residue, and protease is particularly preferably used as such a hydrolase. Specific examples of this protease include trypsin, thrombin, serine proteases such as snake venom thrombin-like enzymes, cathepsin B,
Examples include thiol proteases such as fuicin and metalloproteases such as kininase, but serine proteases are easily available and are therefore suitable for use. Although these proteases can activate blood coagulation alone, according to the present invention, they can activate blood coagulation by using them in combination with the above-mentioned non-enzymatic activator, especially the factor activator consisting of the cyclic compound. The ability to activate coagulation is dramatically improved. In using the blood coagulation promoter of the present invention, for example, blood is collected into a blood test container and the promoter is present in the blood when coagulating the blood. In this case, the blood coagulation promoter may be present in the blood as it is in powder form, but preferably, the blood coagulation promoter is dissolved or dispersed in an appropriate solvent and then added to the blood. . The blood test container is not particularly limited, and conventional glass or resin containers may be used as appropriate. In addition, if blood comes into contact with these blood coagulation promoters momentarily or partially in a blood test container and there is a risk that protein components in the blood may be denatured, use a carrier with a large specific surface area. A blood coagulation promoter may be supported and added to the blood in the blood test container. The carrier is not particularly limited as long as it does not have a harmful effect on blood tests and has a large specific surface area.
Although various materials can be used, for example, nonwoven fabrics, woven fabrics, resin beads, etc. can be suitably used. In order to support a blood coagulation promoter on such a carrier, for example, the carrier may be coated with a solution or dispersion thereof, or immersed therein, and then dried and attached to the carrier. Further, particulate matter carrying a blood coagulation promoter can also be obtained by mixing with an appropriate auxiliary agent such as gum arabic to form an aqueous dispersion, and then rapidly freeze-drying the resulting aqueous dispersion. The amount of the blood coagulation promoter present in the blood is at least 1 x 10 -10 g per ml of blood;
When the amount is less than this, the effect of promoting blood coagulation is poor. However, when present in too large a quantity,
On the contrary, it may cause various problems in blood tests, so it is preferable to limit the amount to 10 -1 g or less. (Effects of the Invention) According to the blood coagulation promoter of the present invention, when it is present in blood, factors are rapidly activated, and the time required for coagulation after blood is collected into a container is significantly shortened. As a result of sufficient contraction of blood clot components, the separability of serum and blood clot is excellent, and the blood clot components are not mixed into the separated and collected serum, and furthermore, the yield of serum is significantly increased. Therefore, the blood coagulation promoter of the present invention can be widely used in the field of clinical testing, and can also be used for hemostasis of bleeding wounds. The present invention will be explained below with reference to Examples, but the present invention is not limited to these Examples in any way. (Example) Example 1 Ellagic acid oxide and 1,2,3-triketohydroindene were used as the cyclic compound as a blood coagulation factor activator, and trypsin, thrombin and snake venom thrombin-like were used as proteases. A blood coagulation promoter aqueous solution according to the present invention was prepared using each enzyme. The content of each component in each blood coagulation promoter is 0.5% by weight for the cyclic compound, and 0.5% by weight for the protease, trypsin, thrombin, and snake venom thrombin-like enzyme.
0.05% by weight, 500 units/ml and 0.005% by weight. Using a commercially available polyethylene plain spit, 30μ of the blood coagulation promoter according to the present invention was added to 3ml of fresh human blood, and the time required for the blood to lose its fluidity was measured as the coagulation time. ,
The mixture was centrifuged at 3000 rpm for 5 minutes and the state of separation was observed. The results are shown in Table 1. Comparative Example 1 For comparison, Table 2 shows the clotting time and separation state when the cyclic compound and protease used in Example 1 were used alone. Comparative Example 2 Table 3 shows the clotting time and separation state when blood was treated in the same manner as in Example 1, except that no blood coagulation promoter was used.
【表】【table】
【表】【table】
Claims (1)
基が立体的に実質的に同一の平面上にある環式有
機化合物からなり、該二つの相隣るカルボニル基
を含む環が６員環又は５員環である、血液凝固第
因子の非酵素的活性化剤と、アミノ酸配列に
おいてArg又はLys残基と任意のアミノ酸残基と
の間の結合の加水分解酵素とを含有することを特
徴とする血液凝固促進剤。[Claims] 1. General formula (However, A represents a residue of a cyclic compound.) and consists of a cyclic organic compound in which the two adjacent carbonyl groups are sterically substantially on the same plane, A non-enzymatic activator of blood coagulation factor, in which the ring containing two adjacent carbonyl groups is a 6-membered ring or a 5-membered ring, and an Arg or Lys residue and any amino acid residue in the amino acid sequence. A blood coagulation promoter characterized by containing a hydrolyzing enzyme and a bond-hydrolyzing enzyme.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP59031794A JPS60174952A (en) | 1984-02-21 | 1984-02-21 | Blood coagulation accelerator |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP59031794A JPS60174952A (en) | 1984-02-21 | 1984-02-21 | Blood coagulation accelerator |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPS60174952A JPS60174952A (en) | 1985-09-09 |
| JPH0546502B2 true JPH0546502B2 (en) | 1993-07-14 |
Family
ID=12340972
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP59031794A Granted JPS60174952A (en) | 1984-02-21 | 1984-02-21 | Blood coagulation accelerator |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPS60174952A (en) |
Families Citing this family (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH0643342B2 (en) * | 1986-04-11 | 1994-06-08 | 積水化学工業株式会社 | Blood coagulation promoter |
| KR950006614B1 (en) * | 1986-04-11 | 1995-06-19 | 세끼스이 가가꾸 고교 가부시기가이샤 | Method for acceleration of blood coagulation |
| JPH05506309A (en) * | 1990-04-17 | 1993-09-16 | アナリティカル・コントロール・システムズ・インコーポレーテッド | Coagulation assays and reagents |
| AU674265B2 (en) * | 1993-11-04 | 1996-12-12 | Dade Behring Inc. | Tetrahydroxyquinone as an activator component for activated partial thromboplastine time test of blood coagulation and as a detector of blood coagulation disorders |
| JP2000187032A (en) * | 1998-12-21 | 2000-07-04 | Nagase & Co Ltd | Method and device for separating blood serum |
-
1984
- 1984-02-21 JP JP59031794A patent/JPS60174952A/en active Granted
Also Published As
| Publication number | Publication date |
|---|---|
| JPS60174952A (en) | 1985-09-09 |
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