# US 4,394,374 A · Thymus gland extracts

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- Number: US 4,394,374 A
- Document ID: US4394374A
- Status: Granted
- Kind: A
- Application number: 06/273,581
- Filed: 1981-06-15
- Granted: 1983-07-19
- Earliest claimed: 1981-06-15
- Inventors: Richard N. Ushijima
- Web page: https://patentgrep.com/patent/US4394374A

## CPC

- A61K 35/26: Lymph; Lymph nodes; Thymus; Spleen; Splenocytes; Thymocytes
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)

## Abstract

A process for the manufacture of a new thymus gland extract is disclosed. Comminuted thymus glands are mixed with hot distilled water which dissolves target protein substances. After a period of time, solids and liquid fats are separated from the aqueous solution. The solution is mixed with a large volume of a sulfate salt to form a pricipitate containing a high yield of the target substances.

## Claims

### Claim 1

A process for producing a thymus extract in the substantial absence of added acids and organic solvents comprising:
comminuting defatted thymus glands;
mixing the comminuted glands with water at a ratio of not more than 300 g of comminuted glands per 400 ml of water;
maintaining the resulting mixture at an elevated temperature and for a time sufficient that large proteins and proteolytic enyzymes are at least partially denatured;
mechanically separating the mixture, by centrifugation or equivalent means, into oils, undissolved solids, and an aqueous solution including protein substances;
decanting to isolate the aqueous solution;
after the aforesaid steps, forming a precipitate including the protein substances by adding a source of sulfate ions to the aqueous solution in an amount sufficient to produce a precipitate yield at least as great as a yield obtained when from 400 to 600 ml. of distilled water are mixed with each 300 g. of comminuted glands to produce the aqueous solution and from 9.5 to eleven parts to saturated ammonium sulfate solution are combined with each seven parts of the aqueous solution at room temperature;
separating the precipitate from supernatant liquid by centrifugation or equivalent means;
combining the precipitate with water so that a portion of the precipitate dissolves to form an extract solution;
dializing the extract solution to remove dissolved, low molecular weight substances from the extract solution; and
purifying the extract solution by removing any undissolved precipitate from the extract solution by centrifugation or equivalent means.

### Claim 2 (depends on 1)

The process of claim 1 wherein between about eight and ten volumes of water are combined with each volume of precipitate.

### Claim 3 (depends on 1)

The process of claim 1 wherein the low molecular weight substances are recycled by mixing with a new volume of the aqueous solution to form additional amounts of the precipitate including the protein substances.

### Claim 4 (depends on 1)

The process of claim 1 wherein:
about 500 ml. of water is mixed with each 300 grams of comminuted glands; and
the precipitate is formed by adding about ten parts of saturated ammonium sulfate solution to each seven parts of the aqueous solution.

### Claim 5 (depends on 1)

The process of claim 1 wherein:
from 400 to 600 ml. of water is mixed with each 300 grams of comminuted glands; and
the precipitate is formed by adding from 240 to 280 grams of sodium sulfate to each liter of the aqueous solution.

### Claim 6 (depends on 5)

The process of claim 5 wherein:
about 500 ml. of water is mixed with each 300 grams of comminuted glands; and
between about 240 and 250 grams of sodium sulfate is added to each liter of the aqueous solution.

### Claim 7 (depends on 1)

The process of claim 1 wherein the temperature is maintained at less than 100.degree. C.

### Claim 8 (depends on 7)

The process of claim 7 wherein the temperature is maintained between 65.degree. C. and 90.degree. C.

### Claim 9 (depends on 1)

A thymus extract manufactured according to the process of claim 1.

### Claim 10 (depends on 8)

The process of claim 8 wherein the precipitate is formed by adding Na.sub.2 SO.sub.4 in an amount sufficient to produce a solution containing from 1.75 to 1.98 moles per liter of Na.sub.2 SO.sub.4.

### Claim 11 (depends on 1)

The process of claim 1 further comprising forming a second precipitate by adding an additional amount of a sulfate salt to the separated supernatent liquid.

### Claim 12 (depends on 1)

The process of claim 1 wherein, to form the precipitate, at least 9.5 parts of saturated, aqueous ammonium sulfate solution are added per each seven parts of the aqueous solution that includes the protein substances.

### Claim 13 (depends on 1)

The process of claim 1 wherein, to form the precipitate, at least 250 grams of solid sodium sulfate are added per liter of the aqueous solution that includes the protein substances.

### Claim 14

A process for producing a thymus extract in the substantial absence of added acids and organic solvents comprising:
comminuted defatted thymus glands;
mixing from 400 to 600 ml. of a an aqueous solvent with each 300 grams of comminuted glands;
maintaining the resulting mixture at an elevated temperature sufficiently to at least partially denature large protein fractions and proteolytic enzymes;
mechanically separating the mixture, by centrifugation or equivalent means, into oils, undissolved solids, and a solution including protein substances;
decanting to isolate the the solution,
after the aforesaid steps forming a precipitate including the protein substances by adding a source of sulfate ions to the solution in an amount sufficient to produce a precipitate yield at least as great as a yield obtained when from 9.5 to 11 parts of saturated ammonium sulfate solution are combined with seven parts of the solvent solution at room temperature;
separating the precipitate from supernatent liquid by centrifugation or equivalent means;
combining the precipitate with water so that a portion of the precipitate dissolves to form an extract solution;
dializing the extract solution to remove dissolved, low molecular weight substances from the extract solution; and
purifying the extract solution by removing any undissolved precipitate from the extract solution by centrifugation or equivalent means.

### Claim 15 (depends on 14)

The process of claim 14 wherein:
the defatted thymus glands are comminuted by grinding to a pulp;
the aqueous solvent is hot water;
the maintaining at an elevated temperature is accomplished in a hot water bath from 65.degree. C. to 90.degree. C. for ten to fifteen minutes;
the mechanically separating is accomplished by centrifuging;
the oils are removed by suction;
the solution including protein substances is separated from the undissolved solids by pouring off the liquid;
the sulfate salt is selected from the group consisting of (NH.sub.4).sub.2 SO.sub.4, Na.sub.2 SO.sub.4, and mixtures thereof;
the precipitate is separated from supernatent liquid by centrifugation; and
the precipitate is combined with about ten volumes of water for a sufficient time that a portion of the precipitate is dissolved to form an extract solution.

### Claim 16

A process for producing two distinct thymus extracts in the substantial absence of added acids and organic solvents comprising:
comminuted defatted thymus glands;
mixing the comminuted glands with water at a ratio of not more than 300 g of comminuted glands per 400 ml of water;
maintaining the resulting mixture at an elevated temperature and for a time sufficient that large proteins and proteolytic enyzymes are at least partially denatured;
mechanically separating the mixture, by centrifugation or equivalent means, into oils, undissolved solids, and an aqueous solution including protein substances;
decanting to isolate the aqueous solution;
after the aforesaid steps, forming a first precipitate including protein substances by adding source of sulfate ions to the aqueous solution in an amount sufficient to produce a precipitate yield not greater than a yield obtained when from 400 to 600 ml of distilled water are mixed with each 300 grams of comminuted glands to produce the aqueous solution and seven parts of saturated ammonium sulfate solution are combined with each seven parts of the aqueous solution at room temperature;
separating the first precipitate from supernatant liquid by centrifugation or equivalent means;
forming a second precipitate including protein substances by adding a source of sulfate ions to the supernatant liquid;
separately purifying each of the precipitates by combining each precipitate with water so that portions of the precipitates are dissolved to form extract solutions;
separately dialyzing the extract solutions to remove dissolved, low molecular weight substances from the extract solutions; and
separately purifying the extract solutions by removing any undissolved precipitates from the extract solutions.