JPS6382361A - Blood test container - 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 regardless of the content of heparin. CONSTITUTION:A blood coagulation promoting agent which contains a cyclic organic compound given by the formula where adjacent carbonyl groups in the formula exist solidly and on virtually the same plane, an organic compound having an amine salt or quaternary nitrogen and an anti-fibrinolysis agent or an anti-plasmin agent is made to exist in a test container. The compound of the formula may be any of homocyclic, heterocyclic, monocyclic and polycyclic ones and preferably, the ring containing two carbonyl carbons is 6-member ring or 5-member ring. The organic compound having amine salt or quaternary nitrogen acts as heparin neutralizing agent to inactivate heparin by adsorbing and neutralizing. 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 combined form.

Description

DETAILED DESCRIPTION OF THE INVENTION (Field of Industrial Application) The present invention relates to a blood test container, and particularly to a blood test container capable of promoting coagulation of a blood sample obtained from a patient receiving heparin administration.

(Conventional technology) With the remarkable progress in testing technology, blood tests such as serum biochemistry tests, serum immunology tests, and hematology tests have become widespread, and many of the oral blood tests that are useful for disease prevention and early diagnosis are based on serum It is a test, and the serum required for the test 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 blood to coagulate. If a test container made of Ij oil is used, it will actually be necessary to leave it for more than 4 hours. Therefore, it has the disadvantage that it is not possible to quickly secure serum necessary for testing. 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 blood coagulation takes a long time, the coagulated whole blood is centrifuged for 11 minutes. There is also the problem that it is difficult to separate serum and blood clots well when they are separated over a long period of time. If the separation is poor, pipette the serum part.

Even with the greatest care, contamination with red blood cells is inevitable when aspirating and/or decanting. As a result, clinical test results may be adversely affected or centrifugation may be required again.

Another problem arises when dealing with blood samples from patients undergoing dialysis or thrombosis patients. In such patients, heparin is administered to prevent blood clots, so there is 1 to 20 units of heparin per 10 m of blood. Ren heparin binds to antitropin m in the blood and significantly inhibits the action of thrombin. Furthermore, No.
It is said that it also inhibits the action of blood coagulation factors such as factor II. 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 solve these problems, the inventors have developed a cyclic organic compound represented by the general formula TII below, in which adjacent carbonyl groups exist substantially on the same plane, and (b) proposed a blood coagulation promoter containing a 7E salt or an organic compound containing quaternary nitrogen (Japanese Patent Application Laid-Open No. 1983-1999).
27858).

(Here, person indicates the residue of a cyclic compound).

Examples of the compound represented by the +I1 formula include gallic acid alkyl ester oxide and elladic acid oxide. As the amine salt and/or the organic compound having quaternary nitrogen, an alkylamine hydrochloride or the like is used. When a blood coagulation promoter containing these compounds is used, the above-mentioned amine salts and the like contained therein absorb and neutralize heparin to inactivate it, and the compound represented by formula (I) promotes blood coagulation in the blood. Blood can be coagulated in a short time by activating factor XII.

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 problems and found that not only blood that does not contain heparin but also blood that contains heparin can be rapidly coagulated, and that We attempted to develop a blood test container that can also improve stability.

An object of the present invention is to provide a blood test container that can rapidly coagulate blood regardless of whether it contains heparin, has good stability after coagulation, and has good serum separation.

(Means and effects for solving the problems) The blood test container of the present invention is (a) represented by the following general formula, and in which adjacent carbonyl groups exist substantially on the same plane. A cyclic organic compound containing (BL amine salt or an organic compound having quaternary nitrogen) and a blood coagulation promoter containing a +61 antifibrinolytic agent or an antiplasmin agent is present inside, This achieves the above objective.

(Here, person indicates the residue of a cyclic compound).

The compound represented by the above formula (I1) contained in the blood coagulation promoter used in the present invention may be a homocyclic compound or a heterocyclic compound, or may be a monocyclic compound. In such a cyclic compound, the ring containing the two carbonyl carbons is preferably a 6-membered ring or a 5-membered ring.

Among the homocyclic compounds, preferable 6-membered cyclic compounds include compounds having an O-quinone ring represented by the following general formula.

R1 R6 (where R, , R, , R, and R4 represent hydrogen, a hydrocarbon group, a polar substituent, or a residue in a polycyclic compound).

In the above formula, the hydrocarbon group is not particularly limited, but an alkyl group, particularly an alkyl group having 1 to 18 carbon atoms, is preferable. The polar substituent is also not particularly limited.

Examples include carboxyl groups, carboxylic acid ester groups, hydroxyl groups, amino acids, and mercapto groups. Examples of compounds having an O-quinone ring include O-quinone and compounds represented by the following formulas (2): (Left below) Gallic acid furkyl ester oxide (where RS represents an alkyl group) ) Elazinic acid partial oxide Niladic acid complete oxide 1,4-di(3,4-dihydroxyphenyl)2,3-
Dimethylbutane partial oxide 1,4-di(3,4-dihydroxyphenyl) (capital) Among allocyclic compounds, preferred specific examples of 5-membered cyclic compounds include 1, 2, and 3-trikehydroindene is mentioned.

Examples of the sm cyclic compound include compounds represented by the following general formula (5).

(Here, R6 represents hydrogen, a hydrocarbon group, or a residue in a polycyclic compound, and R1 and R. represent hydrogen, a hydrocarbon group, a polar substituent, or a residue in a polycyclic compound.

Regarding the hydrocarbon group and the polar substituent (same as in formula II1), a preferable specific example of the compound represented by formula (3) is isatin represented by the following formula.

The amine salt or quaternary nitrogen-containing organic compound contained in the blood coagulation promoter used in the present invention acts as a heparin neutralizer that adsorbs and neutralizes heparin to inactivate it. The amine constituting the amine salt may be any of primary, secondary, and tertiary amines, and the acid constituting the amine salt may be either an inorganic acid or an organic acid.

Inorganic acids include hydrohalic acids such as hydrochloric acid, sulfuric acid,
Examples include sulfurous acid, and organic acids include formic acid and acetic acid. The organic residue of the amine salt is usually a 7'alkyl group, but it may also be a hydrocarbon group containing a different element such as an imino group or an ether group.The 7' salt may be an inner salt.

Specific examples of preferred amine salts include (XI
) Hexadecyldimethylamine hydrochloride represented by the formula,
There is tetradecyl di(aminoethyl)glycine represented by the formula (XII).

+ Cu”u-NH(CHl)z・CI (X
I ) + C, 4H,, NHCH, CH, NHCH, CH, NH,
CH, COO (XII) Organic compounds having quaternary nitrogen include, for example, tetraalkylammonium. A compound having a 7-aryl group instead of an alkyl group, or a compound having a hydrocarbon group containing a different element such as an imino group or an ether group may be used. As a preferable specific example, dodecyltrimethylammonium chloride represented by the formula (XI I I) is OC,,H! SN (CH,), -CI- (XIII)
In addition to such relatively low molecular weight compounds, organic polymers containing quaternary nitrogen may also be utilized.

Examples of such polymers include polycations having repeating units represented by general formula (XIV).

(Here, R, -wRl is ice water or an alkyl group,
X is a halogen group or an acid group, Y is a fullylene group or -
It represents an alkylene group -8O-, and the number of repeats of the above unit is 5 to 2000. ) Among the compounds represented by (XIV), polycations having a repeating unit represented by (XV) or (XVI) are particularly preferred.

On the other hand, as antifibrinolytic agents or antiplasmin agents, aprotinin, soybean trypsin inhibitor, ε-aminocaproic acid, P-7 minomethylbenzoic acid, aminomethylcyclohexanecarboxylic acid, etc., which have been conventionally used clinically, can be used alone or in appropriate combinations. They can be used together.

The blood coagulation promoter in the present invention is I/ml of blood.
It is preferable to use it in a proportion of X 1 G-'' to 1X10'y. If it is too small, the effect of promoting blood coagulation cannot be obtained. Even if it is in excess, the effect corresponding to the amount used cannot be obtained.

Further, in the blood coagulation promoter used in the present invention, the amine salt or the organic compound having quaternary nitrogen is contained in a proportion of 5 to 10.000 parts by weight per 100 parts by weight of the cyclic organic compound. It is preferable to use anti-fibrinolytic agents or anti-plasmin agents in a clinically recommended amount to promote blood coagulation. For example, 7-protinin is used at a dosage of about 100-600 KIU (units) per d of blood, soybean trypsin inhibitor is used at a dosage of about SOO-4000 FU (units) per liter of blood, and ε
-7minocaproic acid, P-7minomethylbenzoic acid and 7minomethylcyclohexanecarboxylic acid, blood 1
It is preferable to use an amount of about 10'' to 1012 per d.10 The blood test container of the present invention may be made of glass or resin.

In addition, in order to have a blood coagulation accelerator present inside a blood test container, it is necessary to store it in powder form, or dissolve or disperse it in an appropriate solvent, or apply it to the inner wall of the container. good.

In addition, in order to prevent a highly concentrated blood coagulation promoter from coming into contact with blood and denaturing protein components, it is also possible to carry the blood coagulation promoter on a carrier with a large specific surface area and store this in a container. .

Such a 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. For example, nonwoven fabrics, woven fabrics, resin beads, etc. are preferably used. In order to support the blood coagulation promoter on such a carrier, for example, a solution or dispersion thereof is applied to the carrier, or the carrier is immersed in the solution or dispersion to be impregnated, and then dried. It is also possible to prepare an aqueous dispersion of a blood coagulation promoter containing an appropriate auxiliary agent such as gum arabic, and then quickly freeze-dry this to obtain a blood coagulation promoter-supported particulate material.

When blood containing heparin is collected into the blood test container of the present invention, the heparin in the blood is adsorbed and neutralized by a neutralizing agent such as an amine salt and precipitates, so that heparin inhibits thrombin and factor XII. It disappears. Therefore,
Blood regains normal clotting function. Furthermore, the cyclic organic compound contained in the blood coagulation promoter has the ability to act on and activate factor XII in the blood. Chapter XI
Activation of factor I causes blood coagulation to proceed in a chain reaction in a short time, and finally, thrombin generated by activation of prothrombin acts on fibrinogen to form an insoluble fibrin network, leading to blood coagulation. is completed.

The time required for blood coagulation varies depending on the type of cyclic organic compound and neutralizing agent in the blood coagulation promoter, the amount of blood coagulation promoter, the material of the container used, the amount of heparin in the blood, etc.
When a synthetic resin container is used, it usually takes 20 to 40 minutes.

Furthermore, antifibrinolytic agents or antiplasmin agents inhibit the fibrin-degrading action of plasmin, which plays an antagonistic role in the blood coagulation reaction process, ensuring coagulation, and also maintain the coagulation state even after coagulation. can be kept stable.

In this way, not only normal blood but also blood containing heparin can be coagulated in a short time, and the coagulated state can be maintained stably even after coagulation. Furthermore, since serum and blood clots can be easily separated, the problem of blood clot components being mixed in the separated and collected serum is also solved. Since the degree of contraction of blood clot components is sufficient, the serum yield is also high.

The blood test container of the present invention can also be used as a vacuum blood collection tube. A vacuum blood collection tube is prepared by evacuating the inside of a container in which a blood coagulation promoter is present, and then sealing the container with a stopper having excellent sealing properties, such as a stopper made of butyl rubber.

(Example) The invention will be explained below with reference to examples.

Example 1 A preester-based nonwoven fabric was impregnated with an aqueous dispersion containing elladic acid oxide (compound represented by formula (M)), polycation (compound represented by formula (XVI)), and 7-brotenin, and thoroughly dried. I let it happen.

The amounts of each of the above components per 1 d of nonwoven fabric were 4 x 10'p, 4 x 10'y, and 500 KIU, respectively.◎ The nonwoven fabric 1- was housed in a commercially available 5-polyethylene spitz to prepare. Add heparin to the Spitz
Inject human fresh, a2m/ containing at a concentration of IU/IR1,
After gentle stirring, the mixture was left at 20°C. Hou Ii! Serum was collected 1 hour and 30 hours later, and fibrinogen and fibrin degradation products (hereinafter abbreviated as FDP) were measured. The results are shown in Table 1. After 1 hour, 3
There was no difference in the FDP measurement values after 0 minutes, 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 and tetradecyl di(aminoethyl)glycine and aprotinin were each combined with α5
A saline dispersion was prepared containing % by weight, α5% and a concentration of 10000 KIU/d.

door! 5 ol of the dispersion was placed in a commercially available 5-polyethylene Spitz and dried to prepare a dispersion. Two liters of fresh cut blood containing heparin at a concentration of LOIU/j was injected into the spirane, treated in the same manner as in Example 1, and evaluated in the same manner.

Example 3 Isatin 12, hexadecyldimethylamine 23M acid salt a
, 4y, aminomethylcyclohexanecarboxylic acid 50q
and polystyrene bead carrier IKI with an average particle size of L5■
F was thoroughly mixed with a small amount of ethanol as a dispersion aid, and then dried.

A commercially available 5D polyethylene spittoon containing the beads was prepared. LQ IU of heparin to the spirane
Example 1
It was processed and evaluated in the same manner as above.

Example 4 O-quinone, polycation (compound represented by formula (XV)) instead of n-propyl gallic oxide, tetradecyl di(7-minoethyl)glycine, and aprotinin
and ε-aminocaproic acid, α5% by weight and α5M, respectively.
The same procedure as in Example 2 was carried out except that the amount was used at a ratio of 1% by weight and α1% by weight.

Example 5 In place of gallic acid n-propyl oxide, tetradecyl di(aminoethyl)glycine and aprotinin, l.
2,3-triketohydroindene, polycation ((X
The procedure of Example 2 was repeated except that the compound represented by the formula VI) and ε-aminocaproic acid were used.

Example 6 The same procedure as Example 2 was carried out except that dodecyltrimethylammonium chloride was used instead of tetradecyldi(aminoethyl)glycine.

Example 7 1,4-di(3,4-
Comparative Example 1 The same procedure as Example 2 was carried out except that dihydroxyphenyl) 2,3-dimethylbutane oxide and polycation (compound represented by formula (XV)) were used. Physiological saline dispersions containing (7minoether)glycine at concentrations of α5% by weight and α5% by weight, respectively, were prepared. Commercially available 5
dLoad heparin in a polyethylene spittoon. OIV/d
After injecting 2 d of fresh human blood containing a concentration of , and then adding 50 μl of the above dispersion, the same treatment and evaluation as in Example 1 were performed.

Example 8 The heparin-containing fresh blood was repeatedly injected into the blood test container in each of Examples 1 to 7, and after gentle stirring, it was left at 20°C until the whole blood had completely flowed. When the time required for the blood to disappear, that is, the blood coagulation time, was measured, the blood coagulated in 30 to 35 minutes in all Examples.

In addition, after blood coagulation, the blood was immediately centrifuged at a rotational speed of 3000 rpm for 5 minutes, and the state of serum separation was observed, as well as the state of serum collection with a pipette. The yield showed good results.

(The following is a blank space) (Effects of the invention) According to the present invention, a container for blood tests can be obtained that can quickly coagulate not only normal blood samples but also blood containing heparin, and furthermore, can be used after coagulation. The stability of the coagulation state is also good. In addition, the state of separation between serum and blood clots is also good. Such blood test containers are suitably used for fractionating serum during blood tests of artificial dialysis patients or thrombosis patients receiving heparin administration.

that's all

Claims (1)

[Claims] 1. (a) a cyclic organic compound represented by the following general formula (I), in which adjacent carbonyl groups exist substantially on the same plane; (b) an amine salt; or an organic compound having quaternary nitrogen;
and (c) a blood test container characterized in that a blood coagulation promoter containing an antifibrinolytic agent or an antiplasmin agent is present therein. ▲There are mathematical formulas, chemical formulas, tables, etc.▼(I) (Here, A indicates the residue of a cyclic compound).