RU2279885C2 - Method for simultaneous production of lecithin, cholesterol, kephalin, and biological solvent cake - Google Patents

Abstract

FIELD: biotechnology, in particular method for production of lecithin, cholesterol, kephalin, and biological solvent cake, being value feed supplement for domestic animals.

SUBSTANCE: claimed method includes raw material pretreatment, lipids extraction, phospholipid fraction separation, lecithin extraction and kephalin precipitation; cholesterol precipitation from neutral lipid fraction. As raw materials waste from meet processing industry, namely frozen brain tissue of domestic animals are used. Tissue is defrosted, ground and homogenized in ethanol, filtered, obtained mass is extracted with ethanol at 15-75°C in one or more steps in biomass/extractant ratio of 1:2-1:100 for 25-120 min in each step to produce brain solvent cake and lipids. Lipids are redissolved in any polar organic solvent having polarity less than acetone in ratio of 1:1-1:10 respectively, and phospholipid fraction is precipitated by addition of 2-10-aliquot acetone volume intosolution. Precipitate is collected on filter, phospholipids are extracted with ethanol in ratio of 1:1-1:50 at 15-75°C in one or more steps, kephalin fraction is collected on filter, filtrate is cooled to 5-10°C, then ethanol is distilled to obtain lecithin. Neutral lipid fraction is redissolved in ethanol in ratio of 1:1-1:50 and conditioned for 30-60 min to produce cholesterol suspension in ethanol. Cholesterol is collected on filter and dried.

EFFECT: method for production of biologically active substances without application of high toxic solvents.

2 tbl, 3 ex

Description

The invention relates to the field of biotechnology and for the production of lecithin, cholesterol, mullet, as well as valuable protein feed additives for animals - bio-meal.

Currently, lecithin is widely used in the production of medicines, cosmetics and food. A number of foreign companies based on lecithin organized the production of highly effective drugs (Lipostabil, Germany, Essential, Essential Fort, Slovenia, etc.) intended for liver treatment. Lecithin both in our country and abroad is widely used to obtain liposomal forms of medical and cosmetic preparations. In recent decades, drugs with a high cholinlecithin content have been increasingly used in medicine: as emulsifiers to obtain an emulsion of fats, as medicines for diseases of the circulatory system, diseases of the liver and gall bladder, and also as a choline donor for diseases of the central nervous system. In our country, lecithin is most widely used for the production of perfumes and in the food industry.

Cholesterol is currently used to obtain the substance of vitamin D ₃ , as well as in the perfume and cosmetics industries.

Vitamins of group D are antirachitic vitamins related to fat-soluble vitamins. Insufficient insolation or malabsorption of vitamin D ₃ in the intestine leads to a violation of calcium-phosphorus metabolism - rickets. Rickets is found in all countries, but especially often where there is a lack of sunlight. Children born in autumn or winter suffer from rickets more often and more severely. With insufficient insolation due to climatic features (frequent fogs, cloudiness, smoke of the atmospheric air) or living conditions, the intensity of the synthesis of vitamin D ₃ decreases. In this case, an additional introduction of Group D vitamins into the children's diet is necessary. In recent years, the frequency of rickets in Russia among young children ranges from 54% to 66%.

For commercial use, only those sources of lecithin that have high concentrations of phospholipids are of interest. In the animal kingdom, eggs are the best source, but this source is very expensive. Plant sources are more economical. Sunflower seed, corn, peanuts and other oilseeds are somewhat less important sources of phospholipids. The most popular of all lecithins is soya lecithin.

A known method of producing lecithin from egg yolks by precipitation with acetone, extraction with ethanol, precipitation with cadmium chloride, reprecipitation with ethanol - Patent RF №2058787.

This method has a number of significant drawbacks: the high cost and shortage of raw materials (eggs), large volumes of organic solvents, the inability to use the method in an industrial environment, in addition, it is proposed at one stage of the process to use chloroform - a solvent that is highly toxic and is prohibited for use in the food industry.

A known method of producing soya lecithin - RF Patent No. 1231658, including degreasing a phosphatide concentrate with acetone, extraction with ethanol, purification on alumina, followed by isolation of the target product.

This method also has a number of significant drawbacks, the main ones of which are: the use of soybean processing intermediate as a phospholipid concentrate as a raw material, a significant amount of soybeans currently grown is genetically modified, and the safety of long-term use of components of genetically engineered strains as biologically active additives plants are still not finally confirmed; the use of column chromatography at one stage of the process is a very time-consuming and expensive stage, the use of which in industrial conditions is fraught with considerable difficulties, in particular with the regeneration and utilization of an adsorbent - aluminum oxide, which makes this method difficult to achieve on an industrial scale.

Closest to the proposed is a method of obtaining valuable cerebroside - nerve, which, like lecithin, is attributed to the polar fractions of the biolipid complex of animal brain tissue. This method includes the stage of extraction of animal raw materials - the brain of pigs. The method is described in RF Patent No. 94012010. However, in this method it is recommended to obtain only one target biologically active product - nerve cerebroside, a number of highly toxic solvents are used - methanol, chloroform, sulfur ether, pyridine, which are significant disadvantages, since it increases the cost of the target product, makes it impossible to use technology waste, it is difficult feasible in an industrial environment.

The objective of the invention is to develop a cost-effective process for obtaining a gamut of biologically active compounds: lecithin, cholesterol, mullet and bio-meal without the use of highly toxic solvents, with the possibility of using the inventive method in semi-industrial and industrial conditions.

The solution to this problem is achieved as follows:

- the use of raw materials with a significant content in its composition of target components with a minimum cost;

- selection of extractants suitable for the process;

- maintaining a special technological process;

As raw materials for production, waste from the meat processing industry is used - the brain of farm animals: cattle, pigs, or rams.

Organic solvents of domestic production are used as extractants in the technological process, which are approved for use in the food industry both in our country, and in Europe and the USA.

The proposed technology does not use organochlorine (chloroform), and other solvents prohibited in the food industry (methanol, pyridine, sulfur ether).

The process under development includes the following main stages: preparation of the extraction material, extraction of lipids with ethyl alcohol, separation of the fraction of water-soluble substances and fractionation of phospholipids to obtain neutral lipids, cephalin and lecithin itself. Cholesterol is obtained from neutral lipids by crystallization in an organic solvent.

Due to the use of ethyl alcohol as an extractant, the bioshrot obtained does not lose its nutritional properties. Moreover, the cholesterol content in raw materials during its processing is reduced to almost zero, which improves the nutritional properties of bio-meal, which can be used for food or feed purposes.

Due to the high content of protein substances it is possible to use bio-meal as a valuable component of feed for various animals, and in the future even for the production of therapeutic and prophylactic food products and medical preparations. The bio-meal obtained from the brain of farm animals contains a wide range of amino acids, including essential amino acids. Amino acid and mineral composition of the biosheath of the brain of cattle are presented in tables 1 and 2.

Table 1
Amino acid composition of bio-meal of brain tissue of farm animals lysine
histidine
arginine
aspartic acids
threonine
serine
glutamic acid
proline
glycine
alanine
cystine
valine
methionine
isoleucine
leucine
tyrosine
phenylalanine 3.90
1.88
3.80
4.10
2.36
2.58
7.10
1.38
2.22
2.72
1.05
2.50
1.70
1.86
4.28
2.42
2.70 Table 2
The content of mineral components in the brain tissue of farm animals. (mg / kg) TO
Na
Sa
Mg
Fe
Cu
Zn
Mn 4390
3390
3405
695
110
8.3
67.2
1.7

The proposed method is as follows.

The frozen brain of farm animals GOST 16677-71, thawed, crushed and homogenized in ethanol, filtered, the resulting biomass is extracted with ethanol at a temperature of 15-75 ° C in one or several stages with a ratio of biomass: extractant equal to 1: 2-1 : 100, for 25-120 minutes at each step, brain bio-meal and lipids are obtained, lipids are redissolved in any organic solvent less polar than acetone at a ratio of 1: 1-1: 10, respectively, and the phospholipid fraction is precipitated by adding lysine to the solution of 2-10 times the volume of acetone, the precipitate of phospholipids is collected on the filter, the phospholipids are extracted with ethyl alcohol at a ratio of 1: 1-1: 50, at a temperature of 15-75 ° C, in one or more steps, the mullet fraction is collected on the filter, the filtrate is cooled to a temperature of 5-10 ° C and re-filtered, then ethanol is distilled off and lecithin is obtained; the neutral lipid fraction is redissolved in ethanol at a ratio of 1: 1-1: 50, maintained for 30-60 minutes and a suspension of cholesterol in ethanol is obtained, the cholesterol is collected on a filter and dried.

The invention is illustrated by the following examples, which illustrate but do not limit the application of the claimed method.

Example 1

Frozen tissue of the brain of a cow in an amount of 1000 g is thawed at a temperature of 20-25 ° C for 2 hours and ground in a meat grinder to obtain a homogeneous forcemeat. The resulting biomass is homogenized with 1000 g of ethanol. The homogenate is extracted with 1000 g of ethanol (biomass: extractant ratio 1: 2) for 120 minutes in three steps at a temperature of 15 ° C. The result is a bioshrot of the brain of a cow and a lipid extract. After drying under vacuum at 60 ° C for 3 hours, 108.0 g of bio-meal are obtained, the composition of which is shown in Tables 1 and 2. The lipid extract is evaporated and lipids are obtained in the amount of 90.0 g, which are evaporated under vacuum and dissolved in 90.0 g of petroleum ether (1: 1 ratio). 180.0 g of acetone (2 volumes) are poured into the resulting solution, the solution is incubated for 15 minutes at room temperature. The precipitate of phospholipids is collected on a filter and dried to obtain 59.0 g of phospholipids. 59 g of ethanol are added to the precipitate of phospholipids (1: 1 ratio) and extraction is carried out at a temperature of 75 ° C in one step. The cephalic fraction was collected on a filter and dried, yielding 42.0 g of cephalin. The filtrate is cooled to a temperature of 5 ° C and re-filtered. The filtrate obtained in turn is evaporated on a rotary-film evaporator under vacuum and 17.0 g of lecithin are obtained. The neutral lipids in a solution of acetone and petroleum ether are evaporated and 31.0 g of neutral lipids are obtained, which are redissolved in 31.0 g of ethanol (1: 1 ratio), the solution is kept for 30 minutes and filtered. Cholesterol is collected from the filter, which is dried in a vacuum oven at a temperature of 60 ° C for 3 hours and 15.0 g of cholesterol is obtained. The solution of neutral lipids is evaporated and sent for further processing to obtain fatty acids.

Example 2

Frozen ram brain tissue in an amount of 100.0 g is thawed at a temperature of 30-35 ° C for 3 hours and minced in a meat grinder to obtain a homogeneous forcemeat. The resulting biomass is homogenized with 1000.0 g of ethanol. The homogenate is extracted with 9000.0 g of ethanol (biomass: extractant ratio 1: 100) for 25 minutes at a temperature of 75 ° C, in one step. The result is a bioshrot of the ram’s brain and a lipid extract. After drying under vacuum at a temperature of 80 ° C for 2 hours, 104.0 g of bio-meal are obtained. The lipid extract was evaporated to give 85.0 g of lipids, which were evaporated in vacuo and dissolved in 850.0 g of ethyl acetate (1:10 ratio). 8500.0 g of acetone (10 volumes) are poured into the resulting solution, the solution is incubated for 10 minutes at room temperature. The precipitate of phospholipids is collected on a filter and dried, to obtain 53.0 g of phospholipids. 2650.0 g of ethanol (ratio 1:50) are added to the precipitate of phospholipids and extraction is carried out at a temperature of 15 ° C in two stages using the same amount of solvent on each. The cephalic fraction was collected on a filter and dried to obtain 34.0 g of cephalin. The filtrate is cooled to a temperature of 10 ° C and re-filtered. The filtrate obtained in turn is evaporated on a rotary-film evaporator under vacuum and 19.0 g of lecithin are obtained. Neutral lipids in a solution of acetone and ethyl acetate are evaporated and 34.0 g of neutral lipids are obtained, which are redissolved in 1700.0 g of ethanol (ratio 1:50), the solution is kept for 60 minutes and filtered. Cholesterol is collected from the filter, which is dried in a vacuum oven at a temperature of 80 ° C for 2 hours and 11.0 g of cholesterol is obtained. The solution of neutral lipids is evaporated and sent for further processing to obtain fatty acids.

Example 3

Frozen tissue of the brain of a pig in the amount of 1000.0 g is thawed at a temperature of 20 ° C for 2 hours and crushed in a meat grinder to obtain a homogeneous forcemeat. The resulting biomass is homogenized with 5000.0 g of ethanol. The homogenate is extracted with 5000.0 g of ethanol (biomass: extractant ratio 1:10), for 90 minutes at 55 ° C in two stages, with the same amount of extractant used at each stage. As a result, a pig brain bio-meal and a lipid extract are obtained. After drying under vacuum at a temperature of 60 ° C for 3 hours, 111.0 g of bio-meal are obtained. The lipid extract is evaporated and lipids are obtained in an amount of 79.0 g, which are evaporated under vacuum and dissolved in 395.0 g of refined vegetable oil (1: 5 ratio). 1975.0 g of acetone (5 volumes) are poured into the resulting solution, the solution is kept for 15 minutes at room temperature. The precipitate of phospholipids is collected on a filter and dried, to obtain 50.0 g of phospholipids. 200.0 g of ethanol (ratio 1: 4) are added to the precipitate of phospholipids and extraction is carried out at a temperature of 55 ° C in three stages using the same amount of solvent on each. The cephalic fraction was collected on a filter and dried, yielding 38.0 g of cephalin. The filtrate is cooled to a temperature of 7 ° C and re-filtered. The filtrate obtained in turn is evaporated on a rotary-film evaporator under vacuum and 16.0 g of lecithin are obtained. Neutral lipids in a solution of acetone and vegetable oil are evaporated to remove acetone impurities from the solution. Then get 415.8 g of neutral lipids in a solution of vegetable oil, which is re-dissolved in 4158.0 g of ethanol (ratio 1:10), the solution is incubated for 45 minutes and filtered. Cholesterol is collected from the filter, which is dried in a vacuum oven at a temperature of 70 ° C for 2.5 hours to obtain 9.0 g of cholesterol. A solution of neutral lipids dissolved in refined vegetable oil is separated from ethanol by distillation of the latter and 406.8 g of neutral lipids in vegetable oil are obtained (which corresponds to a 5.0% solution of neutral brain lipids in vegetable oil), which are used for the preparation of perfumes and cosmetic compositions.

Information sources

1. RF patent No. 2058787 C1. A 61 K 31/685 // A 61 K 35/54, 1996.

2. RF patent No. 1231658 S. A 61 K 35/78, 1994.

3. RF patent No. 94012010 A1. A 61 K 35/30, 1996.

Claims (1)

A method for the simultaneous production of lecithin, cholesterol, mullet and khephalin bio-meal, which consists in the fact that the frozen brain tissue of farm animals is thawed, crushed and homogenized in ethanol, filtered, the resulting biomass is extracted with ethanol at a temperature of 15-75 ° C in one or several steps with a ratio of biomass: extractant equal to 1: 2-1: 100, for 25-120 min at each stage, biosmeat of the brain and lipids are obtained, lipids are redissolved in any organ polar less than acetone ohm solvent at a ratio of 1: 1-1: 10, respectively, and the phospholipid fraction is precipitated by adding 2-10-fold volume of acetone to the solution, the precipitate of phospholipids is collected on a filter, phospholipids are extracted with ethyl alcohol at a ratio of 1: 1-1: 50 at a temperature of 15 -75 ° C in one or more steps, the mullet fraction is collected on the filter, the filtrate is cooled to a temperature of 5-10 ° C and filtered again, then ethanol is distilled off and lecithin is obtained; the fraction of neutral lipids is redissolved in ethanol at a ratio of 1: 1-1: 50, maintained for 30-60 minutes and a suspension of cholesterol in ethanol is obtained, the cholesterol is collected on a filter and dried.