JPS6339574B2 - - Google Patents

Description

[Detailed description of the invention] Trauma (injury) in a state as close to the fetal environment as possible
What is believed to be needed is a product that gives rise to a zone and thus allows individual cells within this zone to have an optimal opportunity for regeneration. muscles, nerves,
Regardless of the skin, the rapidly dividing cells of the embryo or fetus develop from a single fertilized cell and then divide into specific tissues, so the inventors have developed We sought a readily available and inexpensive protein source material obtained from.
The inventor has discovered that freshly slaughtered immature poultry feet are a suitable product. Since poultry by-products are readily available, this would be an ideal source.

The greatest obstacle to regenerative wound treatment is antigenicity. It is possible to simulate the fetal state near the wound, thus stimulating the cell's memory that the cell will probably give rise to other cells, and the biological defenses in such immature cells to advance the foreign protein. The immaturity of the protein source is the most important factor in order to produce a product that, due to its non-rejection, allows for a more regenerative recovery or at least a more rapid therapeutic rate of recovery. For example, in dogs, the body's ability to develop antibodies does not occur until 8-9 weeks after birth. In cats it is 12-13 weeks and in humans it is much later.

The protein hydrolysis products described below are believed to have the properties described above.

Briefly, the medical protein hydrolysis product of the present invention comprises soluble polypeptides and soluble amino acids and inorganic contents of an immature animal protein, such as baby bird's foot, and the product dilutes the animal protein. It is obtained by hydrolysis with an acid, preferably a weak organic acid such as acetic acid, and the resulting solution is then dried or concentrated.

The method involves first cleaning the raw material, such as 8 or 9 week old broilers, and then subdividing the raw material and treating it with dilute acid to controlled PH and temperature conditions. The resulting solution is decanted, strained or filtered, and then spray dried or otherwise dehydrated or concentrated to a dry powder or dry gel.

In use, this powder is applied topically to a moist, open wound, or reconstituted with water and injected with a syringe into the area to be treated. Gels are used to treat relatively dry areas and can be reconstituted from dry powders or partially dry gels.

Preferably, freshly sacrificed immature broiler paws, or other protein sources from freshly sacrificed immature animals, are used as a raw material from which polypeptides and amino acids lacking antigenicity are extracted.

In doing this, the raw material is washed and subdivided, and the protein is extracted from the raw material using a dilute organic weak acid such as acetic or lactic acid as a hydrolyzing agent while controlling the extraction temperature to minimize biological degradation. selectively dissolves. I tried using hydrochloric acid and phosphoric acid, but the results were not very good. Much better is to use acetic acid. The solution thus formed is a liquid product, which is then dried by spray drying or dehydration into a dry product, which can be reconstituted into a gel.

In detail, the young "broiler"
Feet from young "broiler" chickens are used because they have sufficient immature protein and are cheap and plentiful. The paws are thoroughly washed to remove dirt, debris, and blood, and the paws are ground with a 1/4-inch plate in a Hobart grinder. Although 1/8-1/2 inch plates may be used, it is believed that 1/4 inch plates may be broken to the most effective size for rapid and uniform extraction of soluble protein.

In a separate container, an aqueous glacial acetic acid solution is prepared by adding glacial acetic acid to water to form a first dilute acetic acid solution containing about 1 to 11/2 percent by weight glacial acetic acid. The feet are placed in this dilute acetic acid solution on a 1:1 weight basis at room temperature, ie, about 70° to about 90°, to allow the acid to destroy or hydrolyze blood and other deposits. The minced leg and dilute acetic acid were left to work overnight, ie, 8-12 hours, with periodic stirring to ensure thorough mixing. The mixture is then washed or rinsed with fresh water until the minced foot mix is free of debris. This initial treatment or pretreatment with acetic acid solution is performed to remove blood and remove fat and serum.

Then adjust another glacial acetic acid and water mixture to PH
Make the second solution of 4.6. PH is PH6.5 if desired.
It should be PH3.6. However, using these extreme ranges, the resulting protein material was found to be less effective. To this second solution was then added the washed minced foot mix in a 1:1 weight ratio, and the resulting mix was heated to 130-140°C for about 30 min.
-Stir constantly for 40 minutes. This mix can be mixed and heated together for a shorter period of time, but the yield will be reduced.

The solubilized mixture is then decanted or drained and placed in a centrifuge to remove most of the fat and particulate matter. The defatted and centrifuged solution is then passed through a filter. So far, we have found that the best filter to use is "Speed Flow" diatomaceous earth, but we believe that other filters could be used that are equally effective. It will be done.

The dissolved protein solution is then allowed to solidify. Then skim off any remaining fat. To dissolve this gelled protein again, heat it again to 100-120 degrees so that it can be pumped. This mixture (solution) of amino acids and polypeptides generally has a solid content of approximately 4%.
-9%. When preparing the gel, the coagulum (gelling solution) is dried by low temperature air drying, and when necessary, solubilized water is added again at 100%
- Recompose it by making it up to 130〓.

If desired, a further extraction or second extraction can be carried out using the extracted crushed feet and an acetic acid solution (PH 4.6). After extraction for 30 minutes at 140 - 155 degrees and going through the same defatting and cleaning steps as the first extraction, the mixture is combined with the filtered and cleaned protein solution of the first extract. It was found that this second product was not as good as the first extract, as the proteins appeared to be denatured too much in this second product.

To produce a dry powder, the resulting solution is spray dried. The powder has an average particle size of about 1 micron to about 20 microns.

The same procedure was performed using lactic acid diluted to a pH of approximately 4.6. The resulting powder was effective, but inferior to that obtained using acetic acid. Other weak organic acids can be used as is conventional, with similar results. The same thing happens when using hydrochloric acid or phosphoric acid.

The invention may be better understood by reference to the following examples.

Example 8 years old raised and processed in Maryland, USA
- 100 pounds of frozen poultry legs obtained from a 9-week-old freshly slaughtered market broiler were washed with water to remove coarse debris, dirt, and blood. Next, use a Hobart grinder with a 1/4″ plate to crush the legs.
The subdivided material was then immersed in 100 pounds of a 1% aqueous glacial acetic acid solution for 10 hours at room temperature. The mixture was stirred at least once every hour to maximize the zone exposed to dilute acetic acid.

The solution was then drained from the paws by pouring the minced paws onto a 1/32'' mesh screen.
The material on the screen was then rinsed with water until the wash water was clear. This resulted in 87 pounds of minced or shredded broiler or poultry feet after washing.

A first pour solution was made by mixing glacial acetic acid and water to prepare a solution containing approximately 1% by weight glacial acetic acid.

To 87 pounds of the first extraction solution was added 87 pounds of washed and minced poultry feet. The pH of this mixture was then adjusted to PH4.6 by addition of glacial acetic acid. The mixture is then stirred constantly and brought to a temperature of about 130 〓 and about 130 〓
- maintained at about 140°C for about 1/2 hour to dissolve soluble proteins from the crushed material.

This hot solubilized mixture was then subjected to a milk centrifuge to extract the fat from the solution and remove any solid or insolubilized material.

Next, the centrifuged solution was mixed with diatomaceous earth [Speed Flow - a product of Dow Chemical (Grefco)].
by passing it through. At this stage, the solution was a clear solution containing approximately 5.7 pounds of polypeptide and amino acid mix.

Approximately 100-120 degrees of the clear solution was passed through a spray dryer while it was in liquid form. The inlet temperature of the air was about 250㎓ and the outlet temperature was about 120〓. The resulting hydroscopic non-antigenic protein hydrolyzate powder was then immediately placed in an airtight, moisture-tight container to avoid caking.

The resulting powder was analyzed as follows.

PH 4.6 Water 7.9 Total N 15.73 Ash 6.10 Amino acid distribution Aspartic acid 5.51 Glutamic acid 10.24 Histidine 0.81 Lysine 3.80 Argenine 8.01 Hydroxylysine 1.19 Hydroxyproline 10.14 Threonine 2.26 Serine 2.86 Tyrosine 0.73 Glycine 23.41 1/2 Cystine 0.32 Proline 11.83 Alanine 9.14 Valine 2.43 Methionine 1.24 Leucine 3.00 Isoleucine 1.76 Phenylalanine 2.23 Example The procedure of the Example was repeated, but instead of spray drying, the post-filter solution was dried with dry air at a temperature of 65-75 to contain approximately 5% water by weight. .

The weight of the resulting protein hydrolysis product material is 6.1
lb and placed in an airtight, moisture-proof container.

The material is then mixed with water to produce approximately 15-30% by weight
Various gel samples were prepared containing the following materials. approx. water
It was preheated to 120㎓ - about 130㎓ and the ingredients were added to this hot water. The gel was kept under refrigeration.

In the Examples and Examples, a Sharpless centrifuge or cream separator with fixtures was used. A Bowen spray dryer was used in the example.

In the protein hydrolysis product procedure described above by extracting polypeptides and amino acids using acetic acid as a solvent for the extracted proteins, proteins are broken down from their complex protein structures to simpler structures. For best results, it is important to retain in the resulting product the solids or minerals that precipitate when the filtered hydrolyzed protein is dehydrated and cooled. Although such mineral-free protein hydrolysis products provide excellent wound healing, they also produce concomitant tissue overgrowth, which must be re-debrided to bring the area into conformity with adjacent tissue. It is something. After that, the wound will continue to slowly heal.

In other words, it is undesirable to remove precipitated minerals that precipitate as the extracted (solubilized) protein loses water, and it may be undesirable to filter the product after it is in concentrated form. found. When using the non-precipitating mixtures of Examples and Examples, tissue abnormal growth of the regenerating tissue disappeared and significant regrowth of muscle tissue and skin and nerve tissue regenerated back to its normal position, shape and function. , it was shown that memory returned to the tissue.
Treatment using the medical protein hydrolysis product of the present invention.

The protein hydrolysis products of the present invention are particularly useful in the treatment of burns, especially third degree burns. A dry powder is sprinkled over the wound area, which combines with water in the tissue to produce protective cells that resemble a capsule.
The gel is preferably used for first and second degree burns that include relatively dry zones.
The pain usually stops 3-5 minutes after applying the powder to the wound area. It has been found preferable to apply a sprinkling layer of dry powder or gel to the burn zone approximately every three days.

In the case of third-degree burns, this powder allows fiberblasts or regenerating cells to continue to grow from the center and edges of the wound, substantially reducing or completely expelling the skin graft. obtain. Furthermore, it appears that new skin is formed from normal cells and that hair actually grows normally from the new skin.

In most cases of animal wounds treated with the material of the invention in spray-dried or gel form, bacterial infection was generally eliminated after the first three days, and in all cases by the end of the seventh day (second At the end of application) the infection appeared to be eliminated. This was manifested in gas gangrene (clinical), an extensive disintegrating scar of tissue resulting from autologous and other traumatic breakdown of the skin and underlying muscle, nerve and ductal tissues.

When the material of the present invention in spray-dried powder or gel form is topically applied, the regenerative tissue abnormal growth disappears and the significant regrowth of muscle tissue, skin and nerve tissue regenerates and returns to its normal position and shape and function. returns, indicating that memory has been restored to the tissue.

Bedsores that had been present for some time healed very well after treatment with the material of the invention. In fact, even cells that were thought to be dead around the wound recovered and became normal.

Comparative Tests The following tests were conducted to demonstrate the importance of using immature animal tissue, particularly immature poultry feet. Four cats of the same weight, maturity and health condition were operated on and three muscles of the upper right thigh (Bicepts femoris, Vastus lateralis,
Semitendinosus) was severed. Each cat was treated with the therapeutic agent applied topically and the wounds were covered with the therapeutic agent twice a week until all wounds were healed.

The first cat was treated with a protein hydrolyzate prepared according to the example above. The wound healed in 21 days without complications. This cat walked without running over in seven days.

A second cat was treated with a protein hydrolyzate similarly prepared from the paws of approximately one year old baby birds. It took 33 days to heal, and abnormal tissue growth appeared on the 8th day. Normal gait was observed on the 15th day.

A third cat was treated with a prior art protein hydrolyzate, hydrolyzed casein, Borden product. Tissue overgrowth occurred on day 7, and this tissue overgrowth and secondary infection persisted until final resolution at 33 days. The cat was harassed until it finally subsided.

The fourth cat tested the broad spectrum antibiotics 5-nitro-2-fureldehyde and semicarbazone (Eaton).
Laboratories product "FURACIN"). It took 33 days to recover. Mild tissue abnormalities were observed and the cat walked with a limp for the entire period.

In the third and fourth cats, the muscles posterior to the incision gradually atrophied until the thigh pulsed.

This test demonstrates the effectiveness and advantages of the protein hydrolyzate prepared according to the present invention, in particular the rapid recovery rate, absence of infection, absence of tissue abnormal growth and complete recovery of muscle function. Show that.

Thus, it can be seen that the therapeutic agent of the present invention, ie, the medical protein hydrolysis product, is a hydroscopic mixture of polypeptides and amino acids. The therapeutic agents of the present invention appear to have the remarkable ability to create conditions that allow tissue regeneration around the wound zone, without the production of any antibodies, even in the most critically infected wounds.

Preferably, the wound is heavily coated with the powder or gel of the invention. When using powder, the wound area must be moist so that the powder can stick.
No malignant tissue reactions are observed in all treatments with the protein hydrolysis products of the present invention.

It is advisable to cover the wound with a non-adhesive dressing (Telfa) to keep the powder/gel in contact with the wound.
Wound treatment with the material of the invention should be repeated every 3 or 4 days.

Claims (1)

[Claims] 1. Mixing unripe poultry feet with dilute acid at elevated temperature up to 155°C at a pH of 6.5-3.6 to remove fat components and solubilizing protein at elevated temperature. A therapeutic agent for trauma comprising a non-antigenic protein hydrolysis product containing a soluble polypeptide, a soluble amino acid and a dissolved inorganic substance obtained by collecting a hydrolyzate and a dissolved inorganic substance. 2. The wound treatment agent according to claim 1, wherein the elevated temperature is 130-140. 3. The wound treatment agent according to claim 1, wherein the poultry foot is obtained from a baby bird that is not more than 9 weeks old. 4. The wound treatment agent according to claim 1, wherein the acid is acetic acid. 5. A patent in which the poultry feet are crushed and pretreated with an organic mild acid solution containing 1 to 11/2% by weight of acid at 70 to 90 degrees for 8 to 12 hours, and after this pretreatment, the poultry feet are washed with water. The wound treatment agent according to claim 1. 6. The wound treatment agent according to claim 1, wherein the step of removing fat components includes cooling the solution and separating the fat layer. 7. The therapeutic agent of claim 6, wherein the therapeutic agent is a dehydrated gel and is obtained by reheating a cooled solution and air drying the reheated solution to form the dehydrated gel. Trauma treatment agent. 8. A therapeutic agent for wounds according to claim 1, wherein the therapeutic agent is a powder and is obtained by spray drying the solution resulting after removal of fatty substances. 9. the precipitate removal step comprises decanting;
A wound treatment agent according to claim 1. 10. The wound treatment agent according to claim 1, wherein the step of removing the precipitate comprises diatomaceous earth filtration.