CH641963A5 - Process for the production of a porous collagen skin dressing material - Google Patents

Description

The invention now relates to a method for producing a porous collagen skin dressing material, which is characterized in that a collagen gel is formed, the collagen gel is extruded, a collagen sheet is formed from the extruded gel, the extruded collagen is crosslinked and the crosslinked sheet is subjected to freeze-drying.

Solubilized collagen has many NH2 and COOH groups in its framework. Chemical modifications of the molecule can be done well; e.g. all or some of the amino groups can be acylated by reaction with a mixture of acetic anhydride and acetic acid. Similarly, succinic anhydride reacts with collagen, replacing the amino groups with carboxyl groups. The carboxyl groups contained in the molecule are esterified after the standard reaction with acidified alcohol, e.g. Reaction with anhydrous methanol, which is acidified with HCl, accessible. In the above reactions, the pure isoelectric point of collagen can be controlled and set to be either negative or positive or completely neutralized.

All known types of collagen and chemically modified collagen can be used in the practice of this invention, e.g. native collagen; denatured collagen (neutral isoelectric point); esterified collagen (alkaline isoelectric point); and in the form of modified amino groups, e.g. as anhydride derivatives (acidic isoelectric point). In the manufacture of gel type dressings of this invention, the use of collagen anhydride derivatives (acidic isoelectric point), e.g. Succinic anhydride derivatives, preferred.

When producing the collagen gel, the skin or fur is solubilized in an enzyme solution at an acidic pH. The gel obtained is a viscous material which can be filtered, e.g. through a cheesecloth and / or a Millipor filter. The viscous solution is made alkaline by adding alkali to a pH of about 10. At this stage the material is left to inactivate residual enzyme. The material is then neutralized, the collagen collected via a centrifuge and washed with water. A second cleaning step follows, namely redissolving in aqueous acid (pH 2.0-5.0), reprecipitation by neutralization to a pH of 6 to 7 and cleaning to remove acid by dialysis against water. The neutral gel is recovered, and at this stage antibiotics or bactericides, or both, can be added before storing the gel material.

The collagen material used in the preparation of the gel is preferably not a multimer; therefore this material is not subjected to tanning during its manufacture.

The porous collagen sheet is preferably made as follows:

Solubilized collagen gel (pH 2.0-3.5, collagen concentration 1-10%) was extruded from a tubular die into a coagulation bath (saturated NaCl solution). The coagulated cylindrical collagen was cut lengthwise into sheets and tanned for 0.5 to 3.0 hours with 1-5% glutaraldehyde in saturated NaCl containing 0.05 M Na2HP04. The tanned collagen sheet was washed repeatedly with water, then freeze-dried on a methyl methacrylate plate. To produce a semi-porous, sheet-like sheet in which the top surface of the sheet is more concentrated in collagen (resulting in an upper sheet-like surface) and the lower surface of the sheet less concentrated in collagen (ie, more porous), the sheet is freeze-dried subjected to partial air drying. The collagen sheet was sterilized by ethylene oxide gas and soaked in a typical stock solution containing one or more bactericidal agents, such as silver nitrate (0.5 g / 100 ml) or silver lactate (0.5 g / 100 ml) or lactated Ringer's solution 25 mg / ml gentamicin, 25 mg / ml lincomycin, 25 mg / ml colistimethate, 25 mg / ml kanamycin and 5 mg / ml amphotericin B, or contained lactated Ringer's solution containing 25 mg / ml linomy-cin, 5 mg / ml Amphotericin B and 25 mg / ml Gentamicin.

An effective skin dressing material should have the following properties:

1. good adhesion to wounded surfaces

2. Avoidance of protein, fluid and electrolyte losses

3. Avoid infection

4. Pain relief

5. no stimulation of local tissue irritation and the like.

The collagen skin dressings described here satisfy the above properties, are easy to use, and are less expensive. Porous sheet and semi-porous film type

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Dressing materials adhere firmly to the wound, provide effective protection against infections and enable the wounds to heal well. Extensive cell growth was observed in the porous collagen sheet.

Preferred embodiments of the invention are explained using the following examples.

example 1

Fresh calfskin (about 5 kg) was depilated, cleaned with scissors and cut into small pieces. The fur was washed repeatedly with 10% NaCl containing 0.2% sodium azide bactericide and sterilized with water. The fur was dissolved in 10 liters of water (pH 2.5, HCl), which contained 30 mg / ml gentamicin, by adding 1 g of pepsin (approximate ratio of enzyme to collagen was 1/400) at 20 ° C. for 4 days with interim Stirring solubilized. The viscous solubilized collagen obtained was filtered through a cheesecloth, the pH was raised to 10 by NaOH and left to stand at 20 ° C. for 24 hours in order to inactivate the pepsin. The pH of the collagen was then adjusted to 7-8 (HCl), the collagen precipitate was collected by centrifugation and washed with sterilized water. The washed precipitate was redissolved in acidic solution and precipitated again at pH 7-8 for further purification.

The collagen was dissolved in dilute HCl solution (final pH 2.5, collagen concentration 3%) and degassed under vacuum. Acidic collagen gel was extruded into a coagulation bath (saturated NaCl) through a suitable nozzle. The coagulated "tube" material was isolated and cut lengthways into sheets and tanned for 1 hour with 3% glutaraldehyde in saturated NaCl containing 0.05 M Na2HP04. After repeated washing with water, the collagen sheet was freeze-dried on a plate made of methyl methacrylate. The freeze-dried leaves (10 cm x 10 cm) were sterilized by treatment with ethylene oxide gas and preserved by soaking in 0.5% silver nitrate solution. The final thickness of the sheet was 3 mm. This skin dressing material was excellent in terms of adhesion to the wound, protection against fluid loss and infection, and wound healing.

Example 2

A collagen sheet was made by extrusion, tanning and washing using the io method described in Example 1, except that 5% acidic collagen gel was used. The washed collagen sheet was then partially (partially) air dried on a methyl methacrylate plate until the sheet thickness reached half the original thickness. This partial drying reduced the porosity (higher collagen concentration) of the upper surface of the leaf. It was then freeze-dried to make the lower surface porous (collagen concentration lower) and sterilized with ethylene oxide gas. The sterilized sheet was conserved in sterile 0.5% silver lactate solution (pH 7.4). The final thickness of the sheet was 2 mm. This skin dressing material had a finer porosity and greater strength than the sheet of Example 1. It was excellent in protection against protein, fluid and electrolyte loss, protection against infection and wound healing.

Example 3

A sterile freeze-dried collagen sheet was made according to the method described in Example 1 with the exception that the collagen concentration was 5%. The leaf was preserved by soaking in sterile lactated Ringer's solution (pH adjusted to 7.4) containing the following antibiotics: 25 mg / ml gentamicin, 25 mg / ml lincomycin, 25 mg / ml colistimethate, 25 mg / ml Kanamycin 35 and 5 mg / ml amphotericin B. The final thickness of the leaf was 4 mm. This sheet also showed excellent skin dressing properties.

Claims (4)

641 963 1. A method for producing a porous collagen skin dressing material, characterized in that a collagen gel is formed, the collagen gel is extruded, a collagen sheet is formed from the extruded gel, the extruded collagen is crosslinked and the crosslinked sheet is subjected to freeze-drying. 2. The method according to claim 1, characterized in that the crosslinking is carried out by tanning with glutaraldehyde. 2nd PATENT CLAIMS 3. The method according to claim 1, characterized in that at least one antibiotic and / or bactericide is added to the extruded gel or the collagen sheet. 4. The method according to claim 1, characterized in that the cross-linked sheet is partially air-dried before freeze-drying. The National Fire Protection Association reported in 1962 that approximately 1,800,000 people suffer fires each year and use over 11,000 hospital beds a day. There is therefore a great need for a readily available, easy-to-store and temporary replacement for human skin for the effective treatment of heat burns or other forms of skin loss . Such treatments protect against infection as well as protein, body fluid and electrolyte loss from exposed tissue. However, these treatments have the following disadvantages: grafts are difficult to obtain and store for long periods, and are also very expensive. These difficulties could be alleviated by developing artificial skin dressing materials that are less expensive and quickly available for use. Collagen is a major protein of the connective tissue such as the skin, cornea and the like and can be treated by treatment with proteolytic enzymes (other than collagenase) e.g. Proctase, pepsin, trypsin and pronase can be solubilized, separated and purified. Solubilized collagen is low in telopeptides, relatively cheap and ideally suited as a material for processing into a skin wound dressing material.