DE2248804A1 - METHOD FOR MANUFACTURING LYSOZYME - Google Patents

Description

Ϊ ¹ . 2620 / P. - (SPA 051)

8 MUNICH 86, DEN

MDHLSTRASSE 22, CALL NUMBER 98 3921/22

SPA - Societa Prodotti Antibiotic! Spa
8 Via Biella
Milan / Italy

Process for the production of lysozyme

The invention relates to a new method of manufacture
of lysozyme from protein by means of ultrafiltration.

The term "ultrafiltration" is understood to mean the separation of compounds with low molecular weights contained in the same solution from compounds with higher molecular weights, the ability of the former
Exploits connections for passage through semipermeable membranes with suitable pore size. By applying more appropriate
Compounds with low molecular weight can thus be

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separate weights from the original solution. The foregoing Process is already on a large scale for concentrating sugar solutions, cleaning sewers and concentrating for example protein and enzyme solutions have been used.

Egg white contains proteins with very different molecular weights: For example, the molecular weight of lysozyme is about 14-500, of ovalbumin 4-5,000, of gonalbumin about 76,000, from Ovomucoid about 28,000 and by Avidin et v / a 66,000.

The object of the present invention is to provide a simple and effective process for the production of lysozyme from protein to provide. This object is achieved by the invention.

The subject of the invention is .somit a process for the production

of lysozyme from protein, which is characterized in that one the

/ E ~ Lweiss subjected to ultrafiltration and that enriched with lysozyme Permeate wins.

The present invention is based on the position that one can separate not only water, but also lysozyme directly from protein using large-pored membranes, because of that Molecular weight is considerably lower than the molecular weights of most of the other proteins present in egg white.

In the process according to the invention, of course, the salts contained in the egg and all compounds in general also occur low molecular weight such as glucose, the semipermeable membrane.

An interesting point of view is that in the invention Process after ultrafiltration, in which a lysozyme containing

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tendes permeate is obtained, the volume of the remaining protein is reduced; is because some of the water has been separated. When drying the lysozyme-free protein, a
considerable amount of heat saved.

From the lysozyme fraction separated off together with part of the water contained in the protein, the enzyme can be obtained in a simple manner by means of ion exchangers (cf. eg; GB-PS 1 110 466). Furthermore, one can zur'Gewinnung the lysozyme
the permeate of a second ultrafiltration using
, subjecting less permeable membranes than before, the.
Water, and the low molecular weight compounds

be separated and a concentrated lysozyme solution back-;

remain. The lysozyme can be obtained in a simple manner from this solution by adding salts. _:

When using the method according to the invention, a qualitatively high-quality lysozyme is obtained. Also occur, based on
is turned on only very small losses of ^proteins: c.as dried protein that comparable mainly for animal feed.

In the process according to the invention, membranes consisting of organic polymers, for example vinyl and acrylic polymers, are particularly preferred as semipermeable membranes *. It can
however, numerous other membranes can be used provided they have the appropriate pore size.

The example illustrates the invention. ,

3 0 9 8 1 5./Q S 8 iß

ORIGINAL INSPECTED example

10 kg of protein with a lysozyme content of 0.3 to 0.5 percent by weight are filtered through a permeator with large-pore membranes made of organic polymers at a pressure of 2 to JO kp / cm ". About 5 liters of solution are obtained which most of the inorganic ions (sodium and potassium ions),
Glucose and depending on the pore size of the membrane used and the number of circulations varying amounts of lysozyme of up to

Contains 1 to 2 g / liter and more. It should also be noted that the pressure and temperature of the ultrafiltration of.der
Depending on the type of membrane used.

The permeate is adjusted to pH 6 to 7 and passed through about 1 liter of a weakly acidic ion exchanger ("Ämberlite" IRC-50) equilibrated to pH 6.5. The exchanger
is washed first with water and then with a 0.2 M sodium phosphate buffer with a pH of 6.5 until no more protein-containing material is eluted with a ß weight percent ammonium sulfate solution. The eluate is then with further
Ammonium sulfate is added up to a concentration of about 40 percent by weight, the lysozyme precipitating out.

The lysozyme-containing permeate can also be subjected to a second ultrafiltration using small-pore membranes in order to reduce the lysozyme concentration to 0.5 to

2 percent increase. The enzyme is then precipitated by adding a salt and purified in a known manner.

The lysozyme prepared according to the invention can be used if desired in salts with non-toxic and physiologically compatible

Inorganic or organic acids are converted.

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INSPECTED

Claims (3)

-. Patent claims 1. Process for the production of lysozyme from protein,
through this. ge-ken ni ^ z-ve ich η et that the protein is subjected to ultrafiltration and that with lysozyme
enriched permeate wins. 2. The method according to claim 1, characterized; that one the permeate enriched with lysozyme from a second ultrafiltration using a smaller-pore membrane than in the first ultrafiltration. 3. The method according to opposition 1 and 2, characterized in that that one consists of a vinyl or acrylic polymer Membrane sets in. 309815/0886