JP2007290982A - Artificial blood - Google Patents

Abstract

【Task】
An object is to provide a new artificial blood.
[Solution]
Artificial blood containing iron corficene represented by the following formula (1) combined with globin.

[Selection figure] None

Description

  The present invention relates to artificial blood.

  During blood transfusion, blood type and the like must be examined, but there is a global demand for artificial components that can be transfused in place of real blood because there is a risk that undesirable components will be mixed in during blood transfusion. If the introduction of artificial blood is successful, the risk of spreading other diseases such as AIDS or jaundice during surgery (due to blood transfusion) may be fundamentally reduced. In addition, since synthetic blood can be transfused regardless of blood type, it is considered that transfusion that affects life and death can be performed in the field or in an ambulance immediately after an accident occurs.

  As techniques related to conventional artificial blood, techniques using hemoglobin or perfluoro hydrocarbon are described in Patent Documents 1 to 3 below.

JP 2000-51322 A Special table 2003-522094 gazette Special Table 2002-507185

  However, the technique using hemoglobin has a problem that an allosteric effector such as an organic phosphate is decomposed and gradually loses a normal oxygen carrying capacity over time. Further, since the oxygen affinity of hemoglobin varies depending on pH, there is a problem that it is difficult to set solution conditions for use as artificial blood.

  In addition, perfluorohydrocarbons that can dissolve oxygen at a high concentration as described above are used for the purpose of supplying oxygen to the body, but even in the technology using perfluorohydrocarbons, overexpanded non-collapsible lung syndrome is manifested. It is difficult to control the residence time in the body after administration, and it is not easy to extract a single isomeric form of perfluorohydrocarbon, which is most suitable for oxygen transport, and perfluorohydrocarbon is used. In some cases, it may be necessary to wear a ventilator, and there is a problem that a decrease in QOL of a patient who breathes with perfluoro hydrocarbon is suggested. Perfluorohydrocarbons are also a global warming gas and their use is being restricted from the standpoint of protecting the global environment.

  In view of the above problems, an object of the present invention is to provide a novel artificial blood.

That is, the artificial blood according to one means for solving the above problem includes iron corficene represented by the following formula (1) combined with globin.
However, the above formula ₍₁₎ R 1 ~R ₈ is a hydrogen atom or an optionally substituted alkyl group, an aryl group, an ester group, an ether group, a carbonyl group or carboxyl group. R _{1 to} R ₈ may be the same or different from each other.

The artificial blood according to another means of the present invention contains iron corficene represented by the following formula (2) combined with globin.

The artificial blood according to another means of the present invention contains iron corficene represented by the following formula (3) combined with globin.

  Further, the artificial blood according to the present invention is not limited, but it is desirable that the pH is in the range of 7 or more and 8 or less. Moreover, although not necessarily limited, it is desirable that artificial myoglobin is contained within a range of 1 mM to 50 mM.

  According to the present invention, a novel artificial blood can be provided.

  Embodiments of the present invention will be described below with reference to the drawings.

The artificial blood according to the present embodiment (hereinafter referred to as “the artificial blood”) includes iron corficene represented by the following formula (1) coordinated with globin. As shown by the following formula (1), iron corphycene has a structure in which four 5-membered rings called pyrrole are arranged in a trapezoidal shape, and is an iron complex in which an iron atom is arranged in the vicinity of the center thereof. The iron colliferin described in the following formula (1) is bound to the globin in the hydrophobic pocket of globin. In addition, although iron corficene shown by following (1) is not necessarily limited, it can obtain by synthesis | combination, for example.
However, the above formula ₍₁₎ R 1 ~R ₈ is a hydrogen atom or an optionally substituted alkyl group, an aryl group, an ester group, an ether group, a carbonyl group or carboxyl group. R _{1 to} R ₈ may be the same or different from each other. The number of carbons of R _{1 to} R ₈ is not limited, but is preferably in the range of 1 or more and 20 or less.

  The globin can be extracted from meat and animal organs without being limited as long as it is derived from mammals. For example, myoglobin or hemoglobin in an animal body can be obtained by a known method (for example, Biochim. Biophys, Acta, 1959, 35, 543), and heme can be removed therefrom. The artificial blood can be obtained by binding with globin.

  In addition, the artificial blood is not limited, but the iron colficene bound to the globin is preferably contained in a solution such as Ringer's solution or physiological saline, and the solution is pharmaceutically acceptable. It is also preferable to include other substances (pH adjusters, inorganic salts, amino acids, vitamins, reducing agents, etc.) that do not inhibit the oxygen carrying function as far as possible. Although it does not necessarily limit as a density | concentration of iron corficene, it is desirable to exist in the range of 1 mM or more and 50 mM or less. Furthermore, although this artificial blood is not necessarily limited, it is preferable that pH is in the range of 7 or more and 8 or less.

  In the artificial blood, it is also preferable that iron corphycene combined with a plurality of the globins is contained in a lipid membrane such as phospholipid.

In this artificial blood, the oxygen affinity of myoglobin, which is considered to be unusable as blood by using corficene described in the above formula (1), can be used for the human body. It can be suitably adjusted to the extent that appropriate oxygen carrying capacity can be imparted. In addition, this artificial blood has little variation in oxygen affinity due to factors such as pH, and is very easy to adjust for conventional artificial blood using hemoglobin, perfluorohydrocarbon, or the like. In addition, by adjusting R _{1 to} R ₈ in the above formula (1) as appropriate, finer adjustment with oxygen affinity becomes possible.

  As described above, a novel artificial blood can be provided.

Example 1
(Synthesis of iron corficene)
The compound of the following formula (I) was dissolved in a solution obtained by mixing 25 ml of ethanol and 25 ml of water in which 2.5 g of sodium bicarbonate was dissolved. And the temperature of this solution was 45 degreeC, and the solution which melt | dissolved 1.1 g of iodine and 2.0 g of potassium iodide in 20 ml of water was dripped at this solution over 15 minutes. Then, it stirred at 45 degreeC for 90 minutes. The precipitate was collected, washed with a large amount of water and dried. As a result, (II) was obtained.

Next, 5.0 g of the compound of (II) was dissolved in 100 ml of tetrahydrofuran to which 0.70 g of 10% palladium carbon activated by 10 drops of triethylamine and carbon was dropped, and the reaction was performed overnight in the presence of hydrogen. And after filtering and evaporating a solvent, recrystallization of the product was performed using the mixed solution of water and ethanol. This operation yielded (III).

Then, 10.00 g of the compound of (III) was dissolved in 100 ml of N, N-dimethylformamide, the phosphorous oxide was brought to a temperature of 5 ° C. or less, and added in the presence of nitrogen. The mixed solution was allowed to react overnight at room temperature while taking care to prevent moisture from entering, and the reaction solution was added to 500 ml of water. Thereafter, sodium hydroxide was added until the amine smelled, the product was washed with water, and recrystallized with a mixed solution of water and ethanol. By this operation, (IV) was obtained.

Then, to a solution of 10.00 g of compound (IV) in 90 ml of acetic acid, 4.90 g of 3,4-dimethylpyrrole and 30% hydrobromic acid in 20 ml of acetic acid was added at a temperature of 60 ° C. It was. Thereafter, the reaction was performed while heating for 1 hour, and the mixture was cooled. Then, it cooled on the ice bath for 1 hour, the crystal | crystallization was deposited, the deposit was collected, and (V) was obtained by recrystallizing using the mixed solution of water and ethanol.

2.70 g of the compound (V) and 5.70 g of ferric chloride dihydrate were dissolved in 230 ml of dimethylformamide, and the mixture was heated to reflux for 1 hour with stirring. Thereafter, the cooled solution was mixed with 500 ml of chloroform, washed with 500 ml of water four times, and then evaporated to dryness. Then, column chromatography using silica gel was performed using chloroform as a solvent. A black-green band was collected and evaporated. Further, the compound obtained after evaporation was dissolved in 100 ml of a mixed solution of 50 ml of sulfuric acid and 50 ml of ethanol, stirred overnight, and dissolved in 300 ml of chloroform. The mixed solution was washed twice with 400 ml of water, twice with 400 ml of 0.5 M sodium hydroxide solution, and then once with 400 ml of water. Thereafter, the solution was evaporated, and column chromatography using silica gel was performed using chloroform. By this operation, corphycene of the following formula (V) was obtained, and iron was further coordinated to obtain iron corphycene of the following (2).
(Production and evaluation of artificial blood)
Natural heme was extracted from the whale myoglobin and replaced with a corphycene iron complex represented by the following formula (3). With respect to this artificial myoglobin, the P ₅₀ value, which is a measure that quantitatively represents the oxygen transport ability, was measured and found to be 37.0 mmHg. Conventional natural myoglobin has been reported to be 1.0 mmHg (Chem. Rev., 1994, Vol. 94, 699), confirming that the oxygen motility of this artificial myoglobin has been significantly improved. Considering the oxygen partial pressure difference between the lungs and peripheral tissues in the human body, it is considered that this artificial myoglobin can improve the oxygen carrying capacity by about 21 times. The oxygen carrying capacity of this myoglobin is as thin as about 80% of that of hemoglobin (reference: Strayer “Biochemistry”, 4th edition, Chapter 7).

(Example 2)
In Example 1, iron corficene shown in the following (3) was obtained by substantially the same method except that the following (I ′) was used instead of (I).

First, natural heme was extracted from the whale myoglobin and replaced with a corphycene iron complex represented by the above formula (2). With respect to this artificial myoglobin, the P ₅₀ value, which is a measure that quantitatively represents the oxygen transport ability, was measured and found to be 7.0 mmHg. Conventional natural myoglobin has been reported to be 1.0 mmHg (Chem. Rev., 1994, Vol. 94, 699), confirming that the oxygen motility of this artificial myoglobin has been significantly improved. Considering the oxygen partial pressure difference between the lungs and peripheral tissues in the human body, this artificial myoglobin can improve the oxygen carrying capacity by about 8 times.

  From the results of the above examples, it can be seen that the oxygen carrying capacity of myoglobin can be controlled by adjusting the position and substituent of the corphycene molecule. In other words, it is extremely important for the molecular design of corficene. As described above, according to the above examples, it was confirmed that myoglobin reconstituted with a corphycene iron complex becomes artificial blood that can be more easily adjusted and functions as an excellent artificial blood material.

The present invention can be used as artificial blood and has industrial applicability.

Claims (5)

Artificial blood containing iron corficene represented by the following formula (1) combined with globin.
(However, R < ₁ > -R < ₈ > is a hydrogen atom or the alkyl group, ester group, or aryl group which may have a substituent. R < ₁ > -R < ₈ > may be same or different, respectively. ) Artificial blood containing iron corficene represented by the following formula (2) combined with globin.
Artificial blood containing iron corficene represented by the following formula (3) combined with globin.
  The artificial blood according to any one of claims 1 to 3, wherein the pH is in the range of 7 to 8. The artificial blood according to any one of claims 1 to 3, comprising the iron corphycene within a range of 1 mM to 50 mM.