JPS6382362A - Blood coagulation promoting agent - Google Patents

Abstract

PURPOSE:To achieve a better stability after coagulation, by using a blood coagulation promoting agent containing a specified organic compound, an anti- fibrinolysis agent and the like to coagulate blood quickly regardless of the content of heparin. CONSTITUTION:This is a blood coagulation promoting agent which contains a cyclic organic compound as given by the formula while adjacent carbonyl groups therein exist on virtually the same plane in a solid manner, an organic compound having amino salt or quaternary nitrogen and an anti-ray solvent or anti-plasmin. The compound of the formula may be any of homocyclic, heterocyclic, monocyclic and polycyclic ones each of which composed of two adjacent carbonyl groups and a residual group A and preferably, the ring containing two carbonyl carbons is 6-member ring or 5-member ring. The organic compound having an amine salt or quaternary nitrogen is hexadecyl dimethyl amine chlorate, dodecyl trimethyl ammonium chloride or the like. The anti-fibrinolysis agent or the anti-plasmin agent employs aprotinin, soy bean trypsin inhibiter, epsilon-aminocaproic acid and the like in a single or a properly combined form.

Description

Detailed Description of the Invention (Industrial Field of Application) The present invention relates to blood Py, a coagulation promoter, and particularly a blood coagulation promoter capable of promoting the coagulation of a blood specimen of a test subject receiving heparin administration. .

(Prior Art) With the remarkable progress in testing technology, blood tests such as serum biochemical tests, serum immunological tests, and blood cell tests have become widely used, and are useful for disease prevention and early diagnosis. Most blood tests are serum tests, and the serum required for these tests is usually
After coagulating the blood collected in a blood test container, it is separated from blood clots (gel-like lumps of fibrin and blood cells) with different specific gravities by centrifugation, and then using a pipette,
Alternatively, it is collected by decanting.

Blood taken from a subject requires a relatively long time to coagulate. For example, even if a glass test container, which has a relatively short blood coagulation time, is used, it takes 40 to 60 minutes for the blood to coagulate, while a synthetic resin test container can be left standing for over 4 hours. Is required. Therefore, it has the disadvantage that serum necessary for testing cannot be quickly secured. This becomes a problem especially when there is a need to carry out an inspection urgently.

According to the conventional serum fractionation method, in addition to the problem that it takes a long time for blood coagulation, there is also the problem that it is difficult to properly separate serum and blood clots when coagulated whole blood is separated by centrifugation. There is also. If the separation conditions are poor, contamination with red blood cells is inevitable when pipetting and/or decanting the serum portion, even with the greatest care. As a result, clinical test results may be adversely affected or centrifugation may be required again.

Another problem arises when blood samples from patients undergoing dialysis or thrombosis patients are exchanged. In such patients, heparin is administered to prevent blood clots, so there is 1 to 20 units of heparin per 10 sl of blood. This heparin binds to antithrombin ■ in the blood and significantly inhibits the action of thrombin. moreover,
It is said to also inhibit the effects of blood coagulation factors such as factor Ⅰ. Therefore, conversion of fibrinogen to fibrin does not occur, and as a result, blood does not coagulate. Therefore, it becomes difficult to separate serum.

In order to resolve these rlRlIll, the inventor (
a) a cyclic organic compound represented by the following general formula (I), in which adjacent carbonyl groups exist substantially on the same plane; and (b) an amine salt and/or a quaternary proposed a blood coagulation promoter containing an organic compound containing nitrogen (Japanese Patent Application Laid-Open No. 60-27858?@).

(C=0 (Here, A represents a residue of a cyclic compound.) Examples of the compound represented by formula (I) include gallic acid alkyl ester oxide, elladic acid oxide, etc.Amine Alkylamine hydrochlorides and the like are used as organic compounds containing salt and/or quaternary nitrogen.When blood coagulation promoters containing these compounds are used, the amine salts and the like contained therein adsorb and absorb heparin. Neutralized and inactivated, the compound represented by formula (I) activates blood coagulation factors in blood and can coagulate blood in a short time.

However, as time passes after coagulation, it is unavoidable that fibrin clots begin to dissolve due to the action of degrading enzymes present in plasma, and therefore, problems remain regarding stability over time after coagulation. It had been.

(Problems to be Solved by the Invention) The inventors have further studied the above-mentioned blood coagulation promoter and found that it is possible to rapidly coagulate not only blood that does not contain heparin but also blood that contains heparin, and that We attempted to develop a blood coagulation promoter that can also improve blood stability. An object of the present invention is to provide a blood coagulation promoter that can rapidly coagulate blood regardless of whether it contains heparin, has good stability after coagulation, and has good serum separation properties.

(Means and effects for solving the problems) The blood coagulation promoter of the present invention is represented by the general formula (wherein, human represents a residue of a cyclic compound),
and (b) an amine salt or an organic compound having a quaternary nitrogen, and (c) an anti-fibrinosolvent. Alternatively, it contains an anti-plasmin agent, thereby achieving the above object.

The above-mentioned cyclic organic compound used as a component of the blood coagulation promoter of the present invention may be either a homocyclic or a heterocyclic compound formed by two adjacent carbonyl groups and residue A. In addition, such cyclic compounds may be monocyclic or polycyclic compounds, but according to the present invention, the ring containing at least two carbonyl carbons is a 6-membered ring or a 5-membered ring.
Cyclic compounds that are membered rings are preferably used.

In particular, the 6-membered cyclic compound preferably used in the present invention is represented by the following formula I (where R1 to R4 represent hydrogen, a hydrocarbon group, a polar substituent, or a residue in a polycyclic compound).
- A compound having a quinone ring, the hydrocarbon group in the above formula is not particularly limited, but is preferably an alkyl group, and the polar substituent is also not particularly limited, but Examples include a carboxyl group, a carboxylic acid ester group, a hydroxyl group, an amino group, and a mercapto group. Therefore, as preferred specific examples of compounds having an O-quinone ring, for example, 〇-quinone, a gallic acid alkyl ester oxide represented by the following general formula (wherein R5 represents an alkyl group), and a gallic acid alkyl ester oxide represented by the following formula, respectively. Niradinic acid partial oxide represented by 1,4-di(3,4-dihydroxyphenyl) 2,
Examples include 3-dimethylbutane partial oxide and complete oxide.

Further, as a preferable specific example of a homocyclic compound in which the ring containing two carbonyl carbon atoms is a 5-membered ring, 1,2,3-)liketohydroindene represented by the following formula 0 can be mentioned.

Further, one of the heterocyclic compounds that can be preferably used in the present invention has the following general formula (wherein R6 is hydrogen,
Represents a hydrocarbon group or a residue in a polycyclic compound, R7
and R8 represents hydrogen, a hydrocarbon group, a polar substituent, or a residue in a polycyclic compound. ), where the above hydrocarbon group and polar substituent are the same as above.

A preferred specific example of such a compound is isatin represented by the following formula.

All of these groups of compounds have two adjacent carbonyl groups on the same or approximately the same plane within the molecule, and although the detailed mechanism of action is unknown, they are effective in blood coagulation. Shows specific effects on factors.

The second component constituting the blood coagulation promoter of the present invention may be an amine salt or an organic compound containing quaternary nitrogen.

Specific examples include hydrohalic acids such as hydrochloric acid, inorganic acid salts such as sulfuric acid and sulfite, and organic acid salts such as formic acid and acetic acid. The organic residue of the amine salt is usually an alkyl group, but this alkyl group may also be an alkyl group containing a different element such as an imino group or an ether group, and the amine salt shall include an inner salt. .

Therefore, specific examples of preferable amine salts include hexadecyldimethylamine hydrochloride represented by the following formula % and tetradecyldi(aminoethyl)glycine represented by the following formula %.

Further, among organic compounds having quaternary nitrogen, the organic monomer is preferably tetraalkylammonium, but it may partially have an aryl group, and as described above, an alkyl The group may be an alkyl group containing a different element such as an imino group or an ether group. For example, a preferred specific example is dodecyltrimethylammonium chloride represented by the following formula %[).

The polymer among the organic compounds having quaternary nitrogen preferably has the following general formula (where R9 is hydrogen or an alkyl group, X is a halogen atom or an acid radical, and Y is an alkylene group or -alkylene group -5o
2- is shown. ) It is a polycation having a repeating unit represented by:

In particular, a polycation having a repeating unit represented by the following formula % is preferably used.

On the other hand, antifibrinolytic agents or antiplasmin agents include aprotinin, (soybean trypsin inhibitor, C-aminocarb decanoic acid, P-
Aminomethylbenzoic acid, aminomethylcyclohexanecarboxylic acid, etc. may be used alone or in appropriate combinations.

In the present invention, a cyclic organic compound, an amine salt, a monomer or polymer having quaternary nitrogen, and an antifibrinolytic agent or an antiplasmin agent as described above are present in the blood as blood coagulation promoters. These blood coagulation promoters may be added to blood as they are, or dissolved or dispersed in an appropriate solvent, or these blood coagulation promoters may be supported on a carrier with a large specific surface area, and this may be placed in a blood test container. It may also be added to blood.

The above-mentioned carrier is not limited to any particular type as long as it does not have a harmful effect on blood tests and has a large specific surface area, and various carriers can be used, but for example,
Nonwoven fabrics, woven fabrics, tM resin, etc. can be suitably used. In order to support blood Fi and coagulation promoters on such a carrier, it is necessary to control the solution and the amount! No. 2 can be applied or dipped, dried, and attached to a 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 above compound in the blood is at least lXl0-''P per 1 d of blood, and if it is less than this, the effect of promoting blood coagulation is poor.However, if it is present in too large a quantity, Since it causes various problems in blood tests, it is preferable to set it to 1×10”y or less.

Further, in the blood coagulation promoter of the present invention, the amine salt or quaternary salt is added to 10,021 t parts of the cyclic organic compound.
It is preferable to contain an organic compound having nitrogen at a ratio of 5 to 10,000 parts from the viewpoint of promoting blood coagulation.
Further, it is preferable to use the antifibrinolytic agent or the antiplasmin agent in a clinically recommended amount. For example, aprotinin is used at a dosage of about 100 to 600 KIU (units) per 1 liter of blood, and soybean trypsin inhibitor is used at a dosage of about 500 to 4 KIU per ld of blood.
．． In the usage amount of OOOF U (unit), also ε-aminocaproic acid, P-aminomethy,...? alzy;z
For 7-y-R-carboxylic acid, it is preferred to use about 10-'' to 1?l' per blood wIld.

According to the present invention, the above-described amine salt or monomer or polymer having quaternary nitrogen adsorbs and neutralizes heparin when it comes into contact with blood containing heparin, and precipitates together with it, In addition to quickly eliminating the action of heparin and thus restoring normal blood coagulation function, the above-mentioned cyclic compounds have specific effects on blood coagulation factors, so glue and blood test containers are used. The blood contained therein coagulates within a short time, and therefore serum can be obtained with good separation by centrifugation.

To explain this point in more detail, in normal blood, when the blood comes into contact with the inner wall surface of the blood container, factor #1 of the coagulation factors immediately becomes activated, which acts as a starting point to cause a chain reaction in the blood. Coagulation progresses, and finally, thrombin generated by activation of prothrombin acts on fibrinogen to form an insoluble fibrin network and complete coagulation.

On the other hand, it is said that in the blood of a patient receiving heparin administration, there is usually 1 unit to 10 units of heparin per 10 (!e), and thus, in the case of blood containing heparin, Heparin acts cooperatively with antithrombin present in the blood and significantly inhibits the action of thrombin.In this way, heparin not only inhibits the action of thrombin, but also inhibits antithrombin and other factors. It is said that it also inhibits the action of coagulation factors, and therefore, normally, in blood containing heparin, the conversion of fibrinogen to fibrin does not occur, and as a result,
Serum cannot be separated because the blood does not clot.

However, according to the present invention, as described above, since heparin is adsorbed to an amine salt or a monomer or polymer having quaternary nitrogen and removed from the blood, blood coagulation factors including thrombin and other blood coagulation factors are removed. may restore normal blood coagulation, and the specific action of the cyclic organic compound may significantly promote blood coagulation.

Furthermore, antifibrinolytic agents or antiplasmin agents inhibit the fibrin-degrading action of plasmin, which is produced competitively during the blood coagulation reaction process, to ensure coagulation, and to stabilize the coagulation state even after coagulation. It can be kept at

As described above, when the blood coagulation promoter of the present invention is present in blood, heparin is removed from the blood, and blood coagulation factors are rapidly activated, which facilitates coagulation after blood is collected in a container. The time required is significantly shortened, and the separation of serum and blood clots is significantly facilitated, thereby eliminating the problem of blood clot components being mixed in the separated and collected serum.

Furthermore, according to the present invention, as a result of sufficient contraction of blood clot components, the yield of serum is significantly increased.Furthermore, the coagulation state can be maintained stably even after coagulation, and the blood clot components can be kept stable over time. This can solve many stability problems.

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 Elazic acid oxide represented by the above formula (ma) and the above formula (
A polyester nonwoven fabric was impregnated with an aqueous dispersion of the polycation represented by 1) and aprotinin, and thoroughly dried. The amounts of each of the above components per unit area of the nonwoven fabric were 4 x 10-', 4 x 10 y, and 500 KIU, respectively.

Add 1.0% heparin to a commercially available 5D polyethylene spittoon.
2 ml of fresh cut blood containing a concentration of I U/ml was injected, and then 15# of nonwoven fabric supporting the above components was added, stirred gently, and left at 20°C. Serum was collected 1 hour and 30 hours after release, and fibrinogen and fibrin degradation products (hereinafter abbreviated as FDP) were measured.

The results are shown in Table 1, and F after 1 hour and 30 hours.
There was no difference in the DP measurement values, indicating that the blood clot decomposition reaction was well suppressed. The results of Examples 2 to 7 and Comparative Example are also shown in Table 1.

Example 2 Gallic acid n-propyl oxide, tetradecyl di(aminoethyl)glycine t, and aprotinin were each mixed at 0.0%.
5% by weight, 0.5% by weight and 10, OOOKIU/
A saline dispersion containing a concentration of 1 was prepared.

Add 1 ml of heparin to a commercially available 5 ml polyethylene Spitz.
．． Inject 2 d of fresh cut blood containing a concentration of 0 IU/d, then 50 μl of the above dispersion! After the addition, the same treatment and evaluation as in Example 1 were conducted.

Example 3 Isatin 12, hexadecyldimethylamine hydrochloride 0.
4f, 50 μm of aminomethylcyclohexanecarboxylic acid and 1 polystyrene bead carrier with an average particle size of 1.5 tm.
kg was thoroughly mixed with glue using a small amount of ethanol as a dispersion aid, and then dried.

Add 1.5 mL of heparin to a commercially available 5 ml polyethylene Spitz.
Inject 2 d of cut fresh blood containing OIU/waste/,
Next, after adding the blood coagulation promoter 12, Example 1
It was processed and evaluated in the same manner as above.

Example 4 O-quinone, polycation (compound represented by formula (1)) instead of n-propyl gallic oxide, tetradecyl di(aminoethyl)glycine, and aprotinin
and ε-aminocaproic acid each at 0.51! Quantity, 0
．． The same procedure as in Example 2 was carried out except that 5% by weight and 0.1% by weight were used.

Example 5 In place of gallic acid n-propyl oxide, tetradecyl di(aminoethyl)glycine and aprotinin,
2,3-triketohydroindene, polycation ((1
1) The same procedure as in Example 2 was carried out except that the compound represented by the formula (2) and ε-aminocaproic acid were used.

Example 6 Same as Example 2 except that dodecyltrimethylammonium chloride was used instead of tetradecyldi(aminoethyl)glycine. Example 7 Gallic acid n-propyl oxide $ and replacement of tetradecyldi(aminoethyl)glycine. ni 1・4-di (3・4-
The procedure of Example 2 was repeated except that 2,3-dihydroxyphenyl (dihydroxyphenyl) 2,3-dimethylbutane oxide and a polycation (a compound represented by the formula <XV>) were used.

Comparative Example 1 Gallic acid n-propyl oxide and tetradecyl di(aminoethyl)glycine were added at 0.5 wt ffi% and 0.5%, respectively.
! A physiological saline dispersion containing the following concentrations was prepared.

1. Heparin was added to a commercially available 5D polyethylene spittoon. O
After injecting 2 ml of fresh blood at a concentration of IU/Ml and then adding 50 μl of the above-mentioned dispersion, it was treated and evaluated in the same manner as in Example 1.

Example 8 The preparation of the coagulation promoter described in each of Examples 1 to 7, the preparation of a spitz for blood collection, and the injection of heparin-containing fresh blood into the spitz were repeated, and after gentle stirring, the mixture was left at 20°C. The time required for whole blood to stop flowing completely,
That is, when the blood coagulation time was measured, the blood coagulated in 30 to 35 minutes in all Examples.

Also, immediately after blood coagulation, 3. Centrifugation was performed for 5 minutes at a rotation speed of OOO revolutions/minute, and the serum separation layer was observed and the situation of serum collection with a pipette was examined. In all examples, good results were obtained for serum separation and serum yield. Indicated.

(The following is a blank space) (Effects of the Invention) According to the present invention, it is possible to provide a blood coagulation promoter that rapidly coagulates not only normal blood samples but also blood containing heparin. Furthermore, when the coagulation accelerator of the present invention is used, the coagulation state after coagulation can be maintained stably,
Furthermore, the yield of serum when serum and blood clots are separated is also increased.

Such a blood coagulation promoter is suitably used for collecting serum for blood tests of artificial dialysis patients or thrombosis patients receiving heparin administration.

Patent applicant Sekisui Chemical Co., Ltd. Representative Hiroshi 1) Kaoru

Claims (1)

[Claims] (a) A compound represented by the general formula ▲ includes mathematical formulas, chemical formulas, tables, etc. ▼ (I) (where A represents a residue of a cyclic compound), and which has the above two phases. A cyclic organic compound in which adjacent carbonyl groups are sterically substantially on the same plane; (b) an amine salt or an organic compound having a quaternary nitrogen; and (c) an antifibrinolytic agent or an antiplasmin agent. A blood coagulation promoter.