CN1067589C

CN1067589C - Protein granules suitable for intravenous injection

Info

Publication number: CN1067589C
Application number: CN95117945A
Authority: CN
Inventors: 徐家祥
Original assignee: Individual
Current assignee: Individual
Priority date: 1995-11-06
Filing date: 1995-11-06
Publication date: 2001-06-27
Anticipated expiration: 2015-11-06
Also published as: CN1150047A

Abstract

The present invention relates to a protein particle which can reduce excessive bleeding in operations and medical treatment and increase the cure speed of the medical treatment and the operations, particularly to a protein particle suitable for intravenous injection. The protein particle suitable for intravenous injection of the present invention is made from human albumin in the following modes: firstly, making a suspending liquid of the protein particle, wherein dissolving protein powder in a buffer solution with proper osmotic pressure; adding a surface active agent into the protein solution; then, adding a crosslinking agent into the mixed solution of protein and the surface active agent; finally, adding an alcoholic solution into the protein, the surface active agent and the crosslinking agent. Thus, the protein particle suitable for intravenous injection of the present invention is made.

Description

A kind of intravenous protein granule and production method and purposes of being fit to

The present invention relates to reduce bleeding profusely in operation and medical treatment, and can quicken the protein granule of medical treatment and operation recovery from illness speed, refer to a kind of intravenous protein granule and production method and purposes of being fit to especially.

As everyone knows, the blood of human body abundance is healthy basis, many operations and medical treatment, and for example cancer operation all can cause and lose blood.Yet blood transfusion is again a very complicated process, may cause acquired immune deficiency syndrome (AIDS) or introduce hepatitis virus; In addition, especially for the patient of aleukia, then danger is very big especially for severe loss of blood.Thereby, be badly in need of clinically a kind of the helpful material of blood coagulation being solved above-mentioned these problems.

Because the effect of this type of material is as platelet, can reduce and operate on and the patient's of surgical patient and aleukia blood loss, thereby after it is injected into human body, to its character certain requirement will be arranged: one: because the diameter of human microvascular is 7 microns, this type of corpuscle diameter must be less than 7 microns so; Its two: itself should not condense this corpuscle in blood flow, but can condense in the wound, promptly is that this corpuscle can not cause that hemocyte condenses in blood vessel, but can participates in or promote hemocyte to condense in the wound.

Because this type of corpuscle helps blood pool, these microgranules must have the surface molecular that can allow their participation blood coagulations, for example Fibrinogen so.

In the prior art, as people such as K.J.Widder in " development of pharmacology and Chemo-Therapy therapy ", Vol.16, " the active microsphere of the magnetic in the spike of antitumor and anticancer agent biophysics and other carriers " literary composition in the 213-271 page or leaf is related, with protein aqueous solution and the oily emulsion synthetic protein microsphere that obtains by heating or polymerization.Yet make the half-life of this microgranule very short owing to protein is very easily rotten in this process.Thereby in actual therapeutic, need it can in blood vessel, preserve the particle that undergoes no deterioration more than at least one day.

People such as Oppenhein are in U.S. Pat 4,107,288, another kind of method is proposed in " effective atomic particle and production method thereof in the injectable composition ", wherein make protein solution reach the limit of desolvating by the concentration of using a kind of opposite solubilizing agent to regulate the reagent that desolvates.The problem that this method exists is that condition is extremely unstable when the reaction beginning, is accompanied by a large amount of polymerization that causes spheroid to generate in the middle of course of reaction, and follows the polymerism above the blood clotting degree to occur.The blood clotting sphere diameter has surpassed blood vessel diameter greater than 7 microns, and this will cause angiemphraxis and cause danger.Thereby in actual therapeutic, need it can not condense and its diameter than 7 microns much smaller microgranules.

The object of the present invention is to provide a kind of intravenous protein granule and production method and purposes of being fit to, this particle does not condense in suspension, and its diameter is littler than 1 micron, can play good blood coagulation effect when being connected with Fibrinogen on its surface.

So being fit to intravenous albumen plasmid, make through following mode this kind of the present invention by human albumin:

1, at first make the protein granule suspension, comprising:

A, in the suitable buffer of osmotic pressure, the solubilising protein powder;

B, in the protein solution described in a, add surfactant;

C, in protein described in the b and surfactant mixed liquor, add cross-linking agent;

D, in the protein described in the c, surfactant and cross-linking agent, add alcoholic solution.

At once, the protein globules liquid suspension of a muddiness occurs, this suspension is suitable for intravenous injection and can occluding vascular.

Among the present invention, described protein solution is a human albumin solution.

Among the present invention, described surfactant is a detergent.

Among the present invention, described surfactant can be selected from a kind of among Texapon K 14 (Sodiumtetradecylsulfate), Tween-80 or the TritonX-151;

Among the present invention, described cross-linker molecules is the covalency micromolecule that comprises glutaraldehyde (glutaraldahyde).

Among the present invention, described ph value of buffer solution scope is 4-8.

Among the present invention, the osmotic pressure scope of described buffer is 14-680mOsm.

2, the spheroid suspension to above-mentioned human albumin is processed into powder again, comprise dilute, filtration, dialysis, refilter, centrifugalize, electrophoretic separation separate with chromatographic column, again by the powder reconstructization of vacuum freezing after dry and freezing.Wherein need to add buffer or biofacies solutions such as change buffer and adding antiseptic in this process, the kind of these reagent and consumption are well-known to those skilled in the art, so do not repeat.

Like this, just make the intravenous protein granule that is fit to of the present invention.

Its purposes of protein granule of the present invention is: if after the muddy appearance of human albumin spheroid suspension, surface protein is added in the spheroid in the suspension, like this, unreacted crosslink part will be linked on the human albumin spherome surface surface protein is former by covalence key on the spheroid; It is characterized in that: the surface protein among the present invention is former can be Fibrinogen.This protein granule suspension composition is suitable for intravenous injection so, its mean particle dia of this compositions is less than 7 microns, and this does not condense this microgranule in blood vessel, but can condense in the wound, this microgranule can not cause that hemocyte condenses in blood vessel, but can participate in or promote hemocyte to condense in the wound.

The present invention is further described below by embodiment, although following examples are to use human albumin, other protein of human body equally also can use and can form spheroid as immune protein, Myoglobin, collagen and gelatin.

Embodiment 1

Do not have Fibrinogen to form the method for microgranule:

The human albumin powder put into 0.9% sodium chloride solution, and adding concentration is at the surfactant of 0.05mg/ml-15mg/ml, a kind of as Texapon K 14, tween-80 or TritonX-151, the concentration that forms protein solution is at 5-20% (percentage by weight); In the serial test tube that contains the above-mentioned protein solution of 2ml of the surfactant that contains variable concentrations, adding 2ml diluted with sodium chloride solution, its concentration range is the glutaraldehyde of 0.01-3.0% then; After 10 minutes, add the ethanol (being diluted with water to 80% (volume ratio)) of 10ml again, at this moment upright this of solution generates cloudy suspensions; After 10 minutes, dilute with the sodium chloride solution of 5 times of volumes and to observe that this suspension is whether clear to confuse, the result is for negating.After this with 1000 times this suspension of microscopic examination, the diameter of finding particle in this suspension is less than 1 micron.

Also finding simultaneously, is that the mixture effect made of the TritonX-151 of 1-2.5mg/ml and albumin that concentration is 100-200mg/ml is best with concentration.Also find with concentration to be the coefficient of variation minimum of its mean particle dia of mixture of making of the Texapon K 14 of 1-12mg/ml in addition.

Claims

1, a kind of suitable intravenous protein granule, it is characterized in that: this particle prepares in the following manner:

A, in the suitable buffer of osmotic pressure, the solubilising protein powder is made protein solution,

B, in the protein solution described in a, add surfactant,

C, in protein described in the b and surfactant mixed liquor, add cross-linking agent,

D, in the protein described in the c, surfactant and cross-linking agent solution, add alcoholic solution, make suspension,

E, through separation, drying and reconstruct processing, make the protein granule powder;

Described protein solution is a human albumin solution; The weight percent concentration of this protein solution is 5-20%; This surfactant concentrations is 0.05-15mg/ml; The concentration of this cross-linking agent is 0.01-3.0%; The consumption of this alcoholic solution is 10ml.

2, according to the described protein granule of claim 1, it is characterized in that: described surfactant is a detergent.

3, according to the described protein granule of claim 1, it is characterized in that: described surfactant is selected from a kind of among Texapon K 14, Triton X-151 or the Tween 80.

4, according to the described protein granule of claim 1, it is characterized in that: described cross-linking agent is the covalency micromolecule that comprises glutaraldehyde.

5, according to the described protein granule of claim 1, it is characterized in that: described buffer is PH=4～8.

6, according to the described protein granule of claim 1, it is characterized in that: the osmotic pressure scope of described buffer is 14-680mOsm.

7, according to the described protein granule of claim 1, it is characterized in that: described alcoholic solution is an alcoholic solution.

8, a kind of production method that is fit to intravenous protein granule, it is characterized in that: this method may further comprise the steps:

A, in the suitable buffer of osmotic pressure the solubilising protein powder, make protein solution, described protein solution is a human albumin solution, the weight percent concentration of this protein solution is 5-20%;

B, in the protein solution described in a, add surfactant, this surfactant concentrations is 0.05-15mg/ml;

C, in protein described in the b and surfactant mixed liquor, add cross-linking agent, the concentration of this cross-linking agent is 0.01-3.0%;

D, add alcoholic solution in the protein described in the c, surfactant and cross-linking agent solution, make suspension, the consumption of this alcoholic solution is 10ml;

E, through separation, drying and reconstruct processing, make the protein granule powder.

9, the application of the described protein granule of claim 1 in the preparation intravenous formulations.