RU2732928C9 - Biopolymer cross-linking method - Google Patents

Abstract

FIELD: chemistry.SUBSTANCE: invention relates to a method of biopolymers crosslinking wherein biopolymers are brought into contact with sea-buckthorn extracts. Sea-buckthorn extracts contain 10–95 wt. % polyphenols in terms of dry weight in the extract. Polyphenols are derived from a group of tanning substances, flavonoids and catechines. Biopolymer cross-linking takes place in an aqueous medium.EFFECT: invention relates to application of sea-buckthorn extracts for biopolymers cross-linking in food products, fodders for animals, cosmetics and pharmaceutics, in medical products or for tanning of hides and skins for fabric finishing.12 cl, 1 dwg, 1 tbl, 8 ex

Description

The present invention relates to a method for crosslinking biopolymers with sea buckthorn extracts and to the preparation and use of sea buckthorn extracts as non-toxic agents for crosslinking biopolymers such as proteins, peptides or polysaccharides. Such biopolymers are used in particular in food, animal feed, pharmaceuticals or medicine or in other industrial applications.

Crosslinked biopolymers such as proteins or polysaccharides are common in nature and are widely used in food, animal feed, enzymes, packaging materials, or biomedical applications as well as industrial applications. In addition, crosslinking of biopolymers is used in tanning hides and skins and in finishing textiles.

Various methods are used to immobilize or crosslink biopolymers, using, for example, aldehydes or dialdehydes such as formaldehyde or glutaric dialdehyde or acetic acid, or enzymes or crosslinking with metal salt complexes, as in tanning hides. The use of iridoids and secoiridoids of olive leaves is specifically described as the use of natural ingredients for crosslinking biopolymers (EP 1 489 135 A1).

EP 1 489 135 A1 describes in detail the disadvantages of the prior art. In particular, the toxicological properties of substances such as formaldehyde or glutaric dialdehyde limit their use in food, animal feed, cosmetics or medicine. The use of heavy metals in leather tanning has significant drawbacks from the point of view of humans and the environment and toxicology and involves complex wastewater treatment or waste disposal.

There is a growing need for natural, in particular herbal, alternatives for crosslinking proteins that can be obtained in sufficient quantities and at a reasonable cost.

Sea buckthorn grows over 2 million hectares worldwide. The use of the fruit is well described. The harvest produces a significant amount of wood, leaves and bark. These raw materials have so far only been used commercially on a limited basis, but also had the potential to add value to farmers.

The aim of the invention is to provide a method for crosslinking biopolymers using plant extracts. Another object of the invention is to provide natural sea buckthorn extracts for crosslinking biopolymers.

These goals are achieved by creating a method for crosslinking biopolymers, in which the biopolymers are brought into contact with sea buckthorn extracts to crosslink these biopolymers.

Extracts of sea buckthorn aerial components or sea buckthorn fruit processing residues are used as sea buckthorn extracts. Waste from processing of sea buckthorn fruits are components of sea buckthorn residues or unused plant components obtained during processing of sea buckthorn fruits. Sea buckthorn berries are reserved for fruit processing. Low proportions of berries when extracting the aboveground components of the sea buckthorn plant do not harm the cross-linking properties of the extract.

In one embodiment, the sea buckthorn residue components or unused plant components obtained during processing of the sea buckthorn fruit are wood, leaves, bark and / or shoots.

In one embodiment, the sea buckthorn extracts are selected from the Ascola, Hergo, Leikora, Pollmix 1 and / or Pollmix 4 sea buckthorn varieties, preferably from the Hergo sea buckthorn variety.

Sea buckthorn extracts contain polyphenols with a content of 10-95 wt.% (Based on dry weight in the extract), and the polyphenols come from the group of tannins, flavonoids and catechins. Polyphenols can be determined using a modified Folin-Ciocalteu (Singleton and Rossi (1965)) colorimetry method for total phenols with phospho-molybdenum-phosphotungstic acid reagents. Am J Enol Vitic. 16: 144-158).

In a preferred embodiment, the sea buckthorn extracts contain polyphenols in a content of 25 to 50 wt% (based on the dry weight of the sea buckthorn extract).

Polyphenols, in particular hydrolysable and condensed tannins, predominantly lead to crosslinking of biopolymers.

In one embodiment, the sea buckthorn extracts are composed exclusively of natural ingredients, with "natural ingredients" being understood as meaning unmodified plant substances that occur naturally.

Sea buckthorn extracts for crosslinking biopolymers are obtained by extracting sea buckthorn plant material in dry or fresh form using water, an organic solvent or a mixture of water and an organic solvent as an extractant and / or solvent.

In one embodiment, a method for producing sea buckthorn extracts for crosslinking biopolymers is carried out by extracting the sea buckthorn plant material in dried or fresh form using water, an organic solvent, or a mixture of water and an organic solvent as an extractant and / or solvent. In an additional embodiment, the method of obtaining sea buckthorn extracts for crosslinking biopolymers is carried out by extracting the sea buckthorn plant material in frozen form using water, an organic solvent or a mixture of water and an organic solvent as an extractant and / or solvent.

Extracts are obtained from aerial plant parts such as leaves, wood and bark or shoots of sea buckthorn or from processing residues from sea buckthorn fruits. In this case, the material can be processed fresh, preferably dried. Waste from processing of sea buckthorn fruits are components of sea buckthorn residues or unused plant components obtained during processing of sea buckthorn fruits. Sea buckthorn berries are reserved for fruit processing. Low proportions of berries when extracting the aboveground components of the sea buckthorn plant do not harm the cross-linking properties of the extract.

As extractants, you can use water, polar organic solvents or mixtures of polar organic solvents with water. Polar organic solvents in the context of the present invention are alcohols or ketones according to one embodiment of the invention. Particularly preferred are alcohols with 1-4 carbon atoms or ketones with 1-4 carbon atoms. Examples of organic solvents are methanol, ethanol, 1-propanol, isopropanol, 1-butanol, 2-butanol, ethylene glycol, acetone, butanone, or any desired mixtures of these solvents.

Preferably, the extractant or solvent is a mixture of an aqueous solution and at least one polar organic solvent. The polar organic solvent in the mixture is usually 10 to 95% by volume. Preferably, the proportion of the polar organic solvent is 10 to 90 vol%, More preferably 20 to 80 vol%, Most preferably 30 to 70 vol%. The preferred polar organic solvents in the mixture correspond to the preferred polar organic solvents listed above. Particularly preferred is a water / alcohol or water / ketone mixture with a solvent content of 20 to 80% by volume.

Auxiliaries can be added to the extractant to adjust pH or reduce surface tension. Extraction takes place at a temperature of 20 to 95 ° C and a pH of 2 to 11.

According to one embodiment of the invention, additives, in particular acids or alkalis and / or anionic, nonionic or amphoteric surfactants, can be added to the extractant, in particular to water.

Extraction can be carried out by conventional means, such as maceration, percolation, or cleavage, in a continuous, discontinuous, one-stage or multi-stage method under normal or elevated pressure.

Treatment of the crude extracts to enrich the polyphenols can be carried out by membrane methods or adsorption. In addition, processing of the crude extracts to enrich polyphenols can be carried out by liquid-liquid extraction.

In one embodiment, the enrichment of the polyphenols is carried out with an adsorbent resin.

After the implementation of the method, the extracts are subjected to enzymatic or acid-catalytic treatment.

Acid catalytic or enzymatic degradation of bound monosaccharides results in improved activation of extracts.

In one embodiment of the method, the cleavage of bound monosaccharides occurs by acid catalytic or enzymatic treatment of the extracts, resulting in aglycones. "Aglycones" mean organic compounds that are a non-monosaccharide component in a glycoside. For example, as an aglycone, isohamnetin can be formed from isorchamethin-3-O-glycoside and / or quercetin from quercetin-3-O-rutinoside.

After the implementation of the method, enzymatic treatment is carried out using glucosidases. By "glucosidases" is meant a group of enzymes that catalyze the hydrolysis of glycosidically linked glucose.

In another embodiment of the method, the extracts are fermented using microorganisms. In one embodiment, fermentation is performed using microorganisms that generate glucosidases.

The extracts can be used in solution or in powder form. Drying of the extracts can be carried out by known drying methods.

Biopolymers in the context of the invention are polymers that are synthesized in the cell of a living being. In one embodiment, the biopolymers are selected from proteins, peptides and polysaccharides, with the biopolymers preferably being proteins.

In one embodiment, the proteins can be selected from milk proteins, preferably casein or β-lactoglobulin; and / or cereal proteins, preferably gliadins, glutenins, albumin or globulins; collagen or gelatin.

Advantageously, in the process according to the invention, it is not necessary to modify the biopolymers prior to crosslinking by bringing the biopolymers into contact with sea buckthorn extracts.

In one embodiment of the method, the crosslinking of biopolymers occurs in an aqueous medium. By "environment" is meant the chemical environment in which the biopolymer is located and the biopolymer is crosslinked. In one embodiment of the method, the aqueous medium comprises water or an aqueous buffer solution.

In an additional embodiment of the method, biopolymers are crosslinked by adding sea buckthorn extract to an aqueous medium.

In a further embodiment of the method, the biopolymers are crosslinked by adding from 0.1 to 100 wt% sea buckthorn extract based on biopolymer, preferably from 0.5 to 50 wt% sea buckthorn extract based on biopolymer.

In an additional embodiment of the method, the crosslinking of the biopolymers occurs over a period of 1 to 72 hours.

The invention also relates to the use of the sea buckthorn extracts according to the invention, alone or in mixture, for the cross-linking of biopolymers in food, animal feed, cosmetics and pharmaceuticals, as well as in medical products or for industrial use, for tanning hides and skins, and for textile finishing.

In one embodiment, sea buckthorn extracts are used to crosslink biopolymers in food, animal feed, cosmetics and pharmaceuticals, as well as in medical products or industrial applications, for tanning hides and skins, and for finishing textiles.

In one embodiment, extracts of the above-ground components of the sea buckthorn plant or sea buckthorn fruit processing residues are used as sea buckthorn extracts.

In one embodiment, sea buckthorn extracts are used to crosslink biopolymers by extracting dried or fresh sea buckthorn plant material using water, an organic solvent, or a mixture of water and an organic solvent as an extractant and / or solvent.

In one embodiment, sea buckthorn extracts are used to crosslink biopolymers, where the mixture of water and organic solvent used as extractant contains 10 to 95 vol% organic solvent.

In one embodiment, sea buckthorn extracts are used to crosslink biopolymers, wherein additives, in particular acids or alkalis and / or anionic, nonionic or amphoteric surfactants are added to the extractant.

In one embodiment, sea buckthorn extracts are used to crosslink biopolymers, the extracts being subjected to enzymatic or acid catalytic treatment.

The invention will be explained in more detail with reference to the following practical examples.

Determination of polyphenols:

Polyphenols are determined using a modified Folin-Ciocalteu method (Singleton und Rossi (1965)) for colorimetry of total phenolic compounds with phosphorus-molybdenum-phosphotungstic acid reagents. Am J Enol Vitic. 16: 144-158).

Determination of flavonoids:

Flavonoids are measured by HPLC on an RP 18 column. The content is calculated using external standards (quercetin-3-O-rutinoside, isorchamethin-3-O-rutinoside, isorchamethin-3-O-glucoside, quercetin and isorchamethin).

The content of protein, sugar, fat and ash is determined in accordance with the methods of the German Food and Feed Code (Lebensmittel-, Bedarfsgegenstände- und Futtermittelgesetzbuch - LFGB 2013).

Example 1

Dried and crushed sea buckthorn leaves are thoroughly extracted with water at a temperature of 40 ° C. The extract is filtered, concentrated and dried. The content of polyphenols in the obtained extract is 20 wt% (based on dry weight in the extract).

In an alternative embodiment of example 1, sea buckthorn leaves of the Hergo variety are used, and the ratio of the mass of dried and crushed sea buckthorn leaves to the mass of the extractant is 1:10. The content of polyphenols in the obtained extract is 15 wt% (based on dry weight in the extract), and the extract yield is 17%.

The extract yield is understood as the dry weight of the extract based on the weight of the dried and crushed sea buckthorn leaves.

Example 2

Dried and crushed sea buckthorn leaves are thoroughly extracted with water at a temperature of 40 ° C. After filtration of the extract, the pH is adjusted to 7, followed by the dosage of β-glucosidase (from almonds). The incubation is carried out at 25 ° C for 3 hours. Control over the decomposition of glycosidically bound polyphenols is carried out using isohamnetin-3-O-glucoside molecules into isorchamethin and quercetin-3-O-rutinoside into quercetin using HPLC. More than 80% of the glycosides have been degraded. The extract is filtered, concentrated and dried.

Example 3

Dried and crushed leaves of the Hergo sea buckthorn are thoroughly extracted at 40 ° C with an ethanol / water mixture, where the mass fraction of ethanol is 60%. The ratio of the mass of dried and crushed sea buckthorn leaves to the mass of the extractant is 1:10. The extract is filtered, concentrated and dried. The content of polyphenols in the obtained extract is 20 wt% (based on dry weight in the extract). The extract yield is 20%.

Table 1. Content of polyphenols, flavonoids, proteins, total sugar and ash in sea buckthorn extracts according to examples 1 and 3: The values are based on the dry weight of the extract.

polyphenols flavonoids protein total sugar ash An extract according to an alternative embodiment Example 1 15 wt% 2.9 wt% 7 wt% 21 wt% 12 wt% Extract according to Example 3 20 wt% 3.8 wt% 5 wt% 16 wt% 9 wt%

Example 4

The defatted natural yoghurt is mixed with 15 g / L of the dry extract from Example 1. After a 24 hour incubation period at 11 ° C., the viscosity was checked using a Bostwick consistometer. The result of the experiment showed that the addition of the sea buckthorn extract according to the invention leads to an increase in viscosity.

Example 5

To demonstrate the crosslinking properties of the sea buckthorn extract from Example 1, a casein solution was used. A solution of 10 g / l of casein in sodium phosphate buffer (pH 7) is mixed with 10 g / l of sea buckthorn extract according to example 1. The incubation is carried out at room temperature for 45 minutes. Based on the turbidity that occurs in the solution, crosslinking of the casein solution is established.

Example 6

To demonstrate the cross-linking properties of the sea buckthorn extract according to example 3, a 2.5 wt% aqueous gelatin solution was prepared. The dried extract of sea buckthorn in accordance with example 3 is prepared in a concentration of 2.5 wt.% In an aqueous solution and filtered to transparency through a 0.45 μm filter. Both clear solutions are mixed in a 1: 1 ratio at room temperature. Crosslinking of gelatin by adding a sea buckthorn extract solution is visualized by the immediate turbidity of the solution.

Example 7

In silage from grass-based animal feed, sea buckthorn extract is added to the silage according to example 1. By adding 5 wt% sea buckthorn extract based on the weight of the silage, the proteins in the silage can be protected from hydrolysis and thus its feed value can be improved. The addition of sea buckthorn extract reduces the degree of protein hydrolysis in animal feed by 18%. The degree of protein hydrolysis is determined using the OPA method (Frister H, Meisel H, Schlimme E (1990) Einsatzmöglichkeiten der modifizierten OPA-Methode in Der Proteinanalytik). Ernährungs-Umschau 37: 442-445).

Example 8

Sea buckthorn extract in accordance with example 2 is used for tanning and retanning sheep skins. Sea buckthorn extract is used in the amount of 25% of the sheep skin mass. It can be shown that tanning of sheep skins is possible by cross-linking protein structures. The attained shrinkage temperatures were from 68.

Claims (12)

1. A method of crosslinking biopolymers, characterized in that for crosslinking biopolymers they are brought into contact with sea buckthorn extracts, while the sea buckthorn extracts contain 10-95 wt.% Polyphenols in terms of dry weight in the extract, and the polyphenols come from the group of tannins, flavonoids and catechins, and the crosslinking of biopolymers occurs in the aquatic environment. 2. The method according to claim 1, characterized in that extracts of ground components of sea buckthorn or the remains of processing sea buckthorn fruits are used as sea buckthorn extracts. 3. A method according to claim 1 or 2, characterized in that sea buckthorn extracts for crosslinking biopolymers are obtained by extracting sea buckthorn plant material in dry or fresh form using water, an organic solvent or a mixture of water and an organic solvent as an extractant and / or solvent. 4. The method according to claim 3, characterized in that the mixture of water and an organic solvent used as an extractant contains from 10 to 95 vol.% Of an organic solvent. 5. A method according to claim 3 or 4, characterized in that additives, in particular acids or alkalis, and / or anionic, nonionic, or amphoteric surfactants are added to the extractant. 6. A method according to any one of claims 3 to 5, characterized in that the extracts are subjected to enzymatic or acid catalytic treatment. 7. Application of sea buckthorn extracts for crosslinking biopolymers in food, animal feed, cosmetics and pharmaceuticals, as well as in medical products or for tanning hides and skins and for finishing fabrics. 8. Use according to claim 7, characterized in that extracts of the above-ground components of the sea buckthorn plant or the remains of processing sea buckthorn fruits are used as sea buckthorn extracts. 9. Use according to claim 7 or 8, characterized in that sea buckthorn extracts for crosslinking biopolymers are obtained by extraction of sea buckthorn plant material in dry or fresh form using water, an organic solvent or a mixture of water and an organic solvent as an extractant and / or solvent. 10. Use according to claim 9, characterized in that the mixture of water and organic solvent used as extractant contains from 10 to 95 vol.% Of organic solvent. 11. Use according to claim 9 or 10, characterized in that additives, in particular acids or alkalis, and / or anionic, nonionic, or amphoteric surfactants are added to the extractant. 12. Use according to any one of claims 9 to 11, characterized in that the extracts are subjected to enzymatic or acid catalytic treatment.