CN103012544B - A kind of method extracting saponin and polysaccharide from tea seed grouts - Google Patents

Abstract

The invention provides a method for extracting saponin and polysaccharide from defatted camellia oleifera seed cake by using water as a solvent. The method comprises the following process steps: (1) grinding the camellia seed cake into powder, and degreasing with petroleum ether; (2) extracting saponin and polysaccharide in the defatted camellia seed cake powder with water; (3) concentrating the extract and adjusting the pH , using macroporous resin to adsorb saponin, and ethanol to elute and purify saponin; (4) the effluent is then adsorbed with macroporous resin to obtain polysaccharide. The invention is extracted with water, which is green and pollution-free; the ethanol solution is used as the eluent, the boiling point is low, easy to recycle and reuse, and the production cost is low; the production process is simple, the operation is convenient, it is suitable for industrial production, and high-purity saponin can be obtained. and polysaccharides.

Description

A method for extracting saponin and polysaccharide from oil tea seed cake

technical field

The invention relates to a method for extracting saponin and polysaccharide from camellia seed cake, and belongs to the technical field of extracting active components of camellia seed cake.

Background technique

Camellia oleifera ( Camellia oleifera Abel. ) is a perennial woody oil crop. It is one of the four woody edible oil source tree species that are as famous as oil palm, olive and coconut. It has unique resource advantages in China and is mainly distributed in Guangxi, Jiangxi and Hunan. and so on. Camellia oleifera fruit is mainly used to extract camellia oil, and the residue after extracting camellia oil is camellia seed cake, also known as camellia oleifera, tea bran, etc. Camellia oleifera seed cake contains a large amount of dietary fiber, as well as various active ingredients such as saponin, polysaccharide and polyphenol.

Saponin is the general term for saponin compounds extracted from Camellia genus Camellia, so it is also called saponin. Its structure is complex and consists of three parts: ligand, sugar body and organic acid. The content of saponin in Camellia oleifera seed cake is about 15%, which is the main product of deep processing of Camellia oleifera fruit. Saponin not only has various surface activities such as emulsification, decontamination, and foaming, but also has significant anti-inflammatory, analgesic, bactericidal, insecticidal, and plant growth-promoting effects. It is widely used in daily chemicals, medicine, food and other fields. At present, there are many studies on the extraction and purification of saponin in Camellia oleifera seed cake, such as ultrafiltration membrane separation technology, organic solvent extraction, flocculant method, recrystallization method, silica gel column chromatography, etc. insufficient.

Plant polysaccharides are formed by the dehydration of monosaccharides in plants, a general term for a class of physiologically active high molecular polymers that are non-starch and non-cellulose. Plant polysaccharides have anti-tumor, hypoglycemic, anti-coagulant, anti-thrombosis, anti-oxidation, anti-aging, anti-virus and other pharmacological effects and various health functions, and are currently one of the hot research areas in the field of food and medicine. At present, the technology for extracting polysaccharides in Camellia oleifera cake is not mature enough and difficult to scale up.

At present, in the method of deproteinization, toxic chemical reagents such as Sevag reagent (a mixture of chloroform and n-butanol), trifluoroacetic acid, etc. are commonly used to cause protein denaturation to achieve the purpose of deproteinization, and deproteinization often takes 5 to 6 times Above, the reagents used are expensive, consume a large amount, are toxic, and are difficult to recycle. They are difficult to be used for industrial scale-up and easily cause problems such as solvent residues.

Contents of the invention

The purpose of the present invention is to provide a method for extracting saponin and polysaccharide from camellia oleifera seed cake with mild production conditions, safety and non-toxicity, simple operation and water as solvent.

The purpose of the present invention is achieved through the following technical solutions.

A method for extracting saponin and polysaccharide from camellia seed cake can be realized through the following two ways.

1. A method for extracting saponin and polysaccharide from camellia seed cake, comprising the following steps:

(1) Add petroleum ether to the camellia seed cake, so that the mass volume ratio of the camellia seed cake to petroleum ether is 1: (6-12) kg/L, extract at 65-90°C, separate the petroleum ether to obtain degreasing Camellia seed cake;

(2) Add water to the defatted camellia seed cake, so that the mass volume ratio of the defatted camellia seed cake to water is 1: (8-16) kg/L, extract at 60-90°C, and filter to obtain the filter residue and Filtrate and filter residue are dried to obtain dietary fiber;

(3) Concentrate the filtrate obtained in step (2) to 1/3-1/2 of the original volume, adjust the pH value to 2-6 to precipitate the protein, centrifuge to obtain the aqueous phase and precipitate, and dry the precipitate to obtain the protein;

(4) Adsorb the aqueous phase obtained in step (3) with AB-8 macroporous resin for 2 to 5 hours, collect the effluent I, and sequentially add AB-8 macroporous resin to water, 0.1 to 0.2% NaOH aqueous solution, and 10 to 30% ethanol Solution elution, and finally use 70-95% ethanol solution to analyze, collect the analysis solution, concentrate and dry to obtain saponin;

(5) Adsorb the effluent I in step (4) with D3520 macroporous resin for 2-4 hours, collect the effluent II, and carry out alcohol precipitation or drying on the effluent II to obtain camellia oleifera polysaccharide.

2. A method for extracting saponin and polysaccharide from camellia seed cake, comprising the following steps:

(1) Add petroleum ether to the camellia seed cake, so that the mass volume ratio of the camellia seed cake to petroleum ether is 1: (6-12) kg/L, extract at 65-90°C, separate the petroleum ether to obtain degreasing Camellia seed cake;

(2) Add water to the defatted camellia seed cake, so that the mass volume ratio of the defatted camellia seed cake to water is 1: (8-16) kg/L, extract at 60-90°C, and filter to obtain the filter residue and Filtrate and filter residue are dried to obtain dietary fiber;

(3) Concentrate the filtrate obtained in step (2) to 1/3-1/2 of the original volume, adjust the pH value to 2-6 to precipitate the protein, centrifuge to obtain the aqueous phase and precipitate, and dry the precipitate to obtain the protein;

(4) Adsorb the aqueous phase obtained in step (3) with D3520 macroporous resin for 2-4 hours, collect the effluent A, elute the D3520 macroporous resin with water, 10-20% ethanol solution in sequence, and finally use 60-80% ethanol solution Analyze the ethanol solution, collect the analysis solution, concentrate and dry to obtain Camellia oleifera polyphenols;

(5) Adsorb the effluent A in step (4) with AB-8 macroporous resin for 2-5 hours, collect the effluent B, and carry out alcohol precipitation or drying on the effluent B to obtain camellia oleifera polysaccharide;

(6) The AB-8 macroporous resin is eluted with water, 0.1-0.2% NaOH, and 10-30% ethanol solution in sequence, and finally analyzed with 70-95% ethanol solution, and the analytic solution is collected, concentrated and dried to obtain saponin.

In the above step (1), extract at 65-90°C for 1-3 times, each time for 1-4 hours.

In the above step (2), extract at 60-90° C. for 2-4 hours.

Hydrochloric acid or sulfuric acid is used to adjust the pH value in the above step (3).

Advantages and positive effects of the present invention: (1) Using water as a solvent to extract saponin and polysaccharide avoids the disadvantages of high cost and poor safety caused by organic solvent extraction. (2) Macroporous resin has the advantages of large processing capacity and reusability. Using macroporous resin adsorption to effectively separate and purify saponin and polysaccharides improves production efficiency and reduces energy consumption. (3) The present invention first uses isoelectric point sedimentation to remove part of the protein, and then uses the macroporous resin to remove the rest of the protein and decolorize. The macroporous resin can be reused, which not only solves the traditional method's difficulty in enlarging, toxic solvent residues, and affecting products. Bottleneck problems such as biological activity also simplify the process and reduce production costs. (4) The production conditions are mild, safe and non-toxic, and the operation is simple, which is suitable for industrial production.

Description of drawings

Fig. 1 is process flow chart of the present invention.

detailed description

The present invention will be further elaborated below in conjunction with specific examples.

In the invention, water is used as an extraction solvent, and macroporous adsorption resin is used to extract, separate and purify saponin and polysaccharide from camellia oleifera seed cake. During the experiment, LSA-700B, AB-8, D101, D941 and D3520 macroporous resins were selected to investigate their adsorption on proteins and polyphenols. The results show that: D3520 macroporous resin has good adsorption and analysis ability to protein and polyphenol, but weak adsorption ability to saponin and polysaccharide, it can be used to adsorb protein and enrich polyphenol in water extract, so D3520 was used Macroporous resin removes protein in polysaccharides (protein is the main factor affecting the purity of polysaccharides) and achieves the purpose of enriching polyphenols. AB-8 macroporous resin has very good selective adsorption to saponin, and has a large adsorption capacity and strong analytical ability, so AB-8 is selected to purify saponin.

Using these two resins to separate and purify saponin and polysaccharides can be achieved by the following two methods:

Method 1: first extract saponin, polysaccharide, protein, polyphenol and other components with water to obtain water extract, concentrate the water extract to 1/3~1/2 of the original volume, adjust pH=2~6 Most of the protein precipitated and was centrifuged to obtain an aqueous phase and a pellet. Use AB-8 macroporous resin to enrich and adsorb saponin in the water phase, remove pigment and other impurities during elution to obtain high-purity saponin; then use D3520 macroporous resin to adsorb proteins, polyphenols, etc., after this process The effluent absorbed by the two macroporous resins mainly contains camellia oleifera polysaccharide, which can be obtained through alcohol precipitation or spray drying.

Method 2: First extract saponin, polysaccharide, protein, polyphenol and other components with water to obtain a water extract, concentrate the water extract to 1/3~1/2 of the original volume, adjust the pH=2~6 to make most Protein precipitation, centrifugation to obtain the aqueous phase and precipitate. Use D3520 macroporous resin to adsorb and enrich proteins and polyphenols in the water phase, and obtain tea polyphenols during elution; then use AB-8 macroporous resin to enrich and adsorb saponin, and remove pigments during elution After waiting for impurities, high-purity saponin is obtained. The effluent absorbed by these two macroporous resins mainly contains camellia oleifera polysaccharide, and the camellia oleifera polysaccharide is obtained after alcohol precipitation or spray drying.

Example 1

Add petroleum ether to 50g of Camellia oleifera seed cake powder, so that the mass volume ratio (g/mL) of Camellia oleifera seed cake powder to petroleum ether is 1:7, extract twice at 75°C for 1 hour each time, filter, and combine the extracts , concentrating and recovering petroleum ether to obtain defatted camellia seed cake. Add water to the defatted camellia oleifera seed cake, so that the mass volume ratio (g/mL) of defatted camellia oleifera seed cake to water is 1:16, extract at 70°C for 2 hours, filter, and dry the residue to obtain dietary fiber and water The extract was concentrated to 1/3 of the original volume, 5% hydrochloric acid was added to adjust the pH=3.5, and the precipitated protein was removed by centrifugation. Take the water phase part, absorb it with AB-8 macroporous resin for 3h, and collect the effluent Ⅰ. The AB-8 macroporous resin was eluted sequentially with water, 0.15% NaOH, and 20% ethanol solution to remove impurities, then analyzed with 80% ethanol solution, collected the analysis solution, concentrated and dried to obtain refined saponin with a yield of 8.6 %, the purity is 97%. After being adsorbed by AB-8 macroporous resin, the effluent I was adsorbed with D3520 macroporous resin for 3 hours, the effluent II was collected, and 4 times the volume of absolute ethanol was added to the effluent II, stirred, left standing, and dried to obtain camellia oleifera Polysaccharide, the yield is 1.04%, and its glucose equivalent is 0.3763. Dietary fiber can be obtained by drying the residue after water extraction.

Example 2:

Add petroleum ether to 500g of Camellia oleifera seed cake powder, so that the mass volume ratio (g/mL) of Camellia oleifera seed cake powder to petroleum ether is 1:9, extract twice at 85°C for 2 hours each time, filter, and combine the extracts , concentrating and recovering petroleum ether to obtain defatted camellia seed cake. Add water to the defatted camellia oleifera seed cake, so that the mass volume ratio (g/mL) of the defatted camellia oleifera seed cake to water is 1:12, extract at 85°C for 3 hours, filter, and dry the residue to obtain dietary fiber and water The extract was concentrated to half of its original volume, 5% hydrochloric acid was added to adjust the pH to 4, and the precipitated protein was removed by centrifugation. Take part of the water phase, absorb it with AB-8 macroporous resin for 3.5h, and collect the effluent Ⅰ. The AB-8 macroporous resin was eluted sequentially with water, 0.15% NaOH, and 20% ethanol solution to remove impurities, then analyzed with 80% ethanol solution, collected the analysis solution, concentrated and dried to obtain refined saponin with a yield of 8.3 %, purity 96.3%. After being adsorbed by AB-8 macroporous resin, the effluent I was adsorbed with D3520 macroporous resin for 3.5 hours, the effluent II was collected, and 4 times the volume of absolute ethanol was added to the effluent II, stirred, allowed to stand, and dried to obtain Camellia oleifera polysaccharide, the yield is 0.98%, and its glucose equivalent is 0.3321. Dietary fiber can be obtained by drying the residue after water extraction.

Example 3:

Weigh 10kg of camellia oleifera seed cake powder, add petroleum ether, the mass ratio of camellia oleifera seed cake to petroleum ether (kg/L) is 1:6, extract twice at 80°C for 3 hours each time, filter, and combine the extracts , concentrating and recovering petroleum ether to obtain defatted camellia seed cake. Add water to the defatted camellia oleifera seed cake, so that the mass volume ratio (kg/L) of the defatted camellia oleifera seed cake to water is 1:8, extract at 90°C for 4 hours, filter, and dry the residue to obtain dietary fiber. Concentrate the water extract to half of its original volume, add 5% sulfuric acid to adjust the pH=4.5, and centrifuge to remove the precipitated protein. Take the water phase part, absorb it with AB-8 macroporous resin for 4h, and collect the effluent Ⅰ. The AB-8 macroporous resin was eluted sequentially with water, 0.15% NaOH, and 20% ethanol solution to remove impurities, then analyzed with 80% ethanol solution, collected the analysis solution, concentrated and dried to obtain refined saponin with a yield of 8.1 %, purity 95.1%. After being adsorbed by AB-8 macroporous resin, the effluent I was adsorbed with D3520 macroporous resin for 3.5 hours, the effluent II was collected, and 4 times the volume of absolute ethanol was added to the effluent II, stirred, allowed to stand, and dried to obtain Camellia oleifera polysaccharide, the yield is 0.7%, and its glucose equivalent is 0.3221. Dietary fiber can be obtained by drying the residue after water extraction.

Example 4:

Similar to Example 1, the difference is that D3520 macroporous resin is first used for adsorption and then AB-8 macroporous resin is used for adsorption.

Weigh 50g of camellia oleifera seed cake powder, add petroleum ether, make the mass volume ratio (g/mL) of camellia oleifera seed cake powder to petroleum ether 1:7, extract twice at 75°C for 1 hour each time, filter, Combine the extracts, concentrate and recover petroleum ether to obtain defatted camellia oleifera seed cake. Add water to the defatted camellia oleifera seed cake, so that the mass volume ratio (g/mL) of defatted camellia oleifera seed cake to water is 1:16, extract at 70°C for 2 hours, filter, and dry the residue to obtain dietary fiber. The water extract was concentrated to 1/3 of the original volume, 5% hydrochloric acid was added to adjust the pH=3.5, and the precipitated protein was removed by centrifugation. Take part of the water phase, absorb it with D3520 macroporous resin for 4 hours, and collect the effluent (A). The D3520 macroporous resin was eluted successively with water and 15% ethanol solution to remove impurities, then analyzed with 70% ethanol solution, collected the analysis solution, concentrated and dried to obtain Camellia oleifera polyphenols, the yield was 0.64%, and its gallic acid equivalent is 0.08572. The effluent (A) after being adsorbed by D3520 macroporous resin was adsorbed with AB-8 macroporous resin for 3 hours, the effluent (B) was collected, and the effluent (B) was concentrated and dried to obtain Camellia oleifera polysaccharide, with a yield of 0.66%. The glucose equivalent is 0.6651; the AB-8 macroporous resin is eluted with water, 0.15% NaOH, and 20% ethanol solution to remove impurities, and then analyzed with 80% ethanol solution, and the analysis solution is collected, concentrated, and dried to obtain refined saponin , the yield was 7.6%, and the purity was 98.5%. Dietary fiber can be obtained by drying the residue after water extraction.

Compared with Example 1, the saponin obtained in Example 4 has a whiter color and higher purity.

Polyphenols are the main reason for the darkening of saponin, and how to remove polyphenols in the process of purifying saponin is the main factor to be considered. The method described in Example 4 can be used to obtain Camellia oleifera polyphenols while eluting D3520.

Method 1 of the present invention uses AB-8 macroporous resin earlier, then uses D-3520 macroporous resin to purify, because AB-8 macroporous resin has strong selective adsorption to saponin, but it also adsorbs polyphenols simultaneously, in In the process of purifying saponin, 0.15% NaOH can be used to remove polyphenols on the AB-8 macroporous resin. Since alkali elution is used, polyphenols cannot be recovered at this time. After elution with 0.15% NaOH, there is still a small amount of polyphenols remaining on the AB-8 macroporous resin, so when the saponin is decomposed with 80% ethanol, the small amount of polyphenols remaining on the resin is also eluted together, resulting in the opposite color of the saponin. Poor, relatively small purity.

And method 2 first uses D3520 macroporous resin, and then uses the purification of AB-8 macroporous resin. Since the D3520 macroporous resin has a strong adsorption effect on polyphenols and proteins (it also has a certain adsorption effect on saponin), after the D3520 macroporous resin is put into the water extract, most of the polyphenols in the extract are absorbed by the D3520 macroporous resin. Resin adsorption, when eluting D3520 macroporous resin, polyphenols can be collected because alcohol is used instead of alkali for elution. In addition, because the D3520 macroporous resin is used first, it basically absorbs the polyphenols in the water extract, and then when the AB-8 macroporous resin is used to adsorb saponin, the influence of polyphenols on the purification of saponin has been minimized. , so that the color and purity of saponin are correspondingly improved. But at this time, saponin has undergone two-step adsorption, so that the loss becomes unavoidable, and the yield decreases accordingly.

Claims (3)

1. a method for extracting saponin and polysaccharides from oil tea seed cake, is characterized in that comprising the following steps: (1) Add petroleum ether to the camellia seed cake, so that the mass volume ratio of the camellia seed cake to petroleum ether is 1: (6-12) kg/L, extract 1-3 times at 65-90°C, 1 to 4 hours each time, separate the petroleum ether to obtain the degreased camellia seed cake; (2) Add water to the defatted camellia seed cake, so that the mass volume ratio of the defatted camellia seed cake to water is 1: (8-16) kg/L, extract at 60-90°C, and filter to obtain the filter residue and Filtrate and filter residue are dried to obtain dietary fiber; (3) Concentrate the filtrate obtained in step (2) to 1/3-1/2 of the original volume, use hydrochloric acid or sulfuric acid to adjust the pH value to 2-6 to precipitate the protein, centrifuge to obtain the aqueous phase and precipitate, and dry the precipitate to obtain the protein ; (4) Adsorb the aqueous phase obtained in step (3) with AB-8 macroporous resin for 2 to 5 hours, collect the effluent I, and sequentially add AB-8 macroporous resin to water, 0.1 to 0.2% NaOH aqueous solution, and 10 to 30% ethanol Solution elution, and finally use 70-95% ethanol solution to analyze, collect the analysis solution, concentrate and dry to obtain saponin; (5) Adsorb the effluent I in step (4) with D3520 macroporous resin for 2-4 hours, collect the effluent II, and carry out alcohol precipitation or drying on the effluent II to obtain camellia oleifera polysaccharide. 2. A method for extracting saponin and polysaccharides from Camellia oleifera cake, is characterized in that comprising the following steps: (1) Add petroleum ether to the camellia seed cake, so that the mass volume ratio of the camellia seed cake to petroleum ether is 1: (6-12) kg/L, extract 1-3 times at 65-90°C, 1 to 4 hours each time, separate the petroleum ether to obtain the degreased camellia seed cake; (2) Add water to the defatted camellia seed cake, so that the mass volume ratio of the defatted camellia seed cake to water is 1: (8-16) kg/L, extract at 60-90°C, and filter to obtain the filter residue and Filtrate and filter residue are dried to obtain dietary fiber; (3) Concentrate the filtrate obtained in step (2) to 1/3-1/2 of the original volume, use hydrochloric acid or sulfuric acid to adjust the pH value to 2-6 to precipitate the protein, centrifuge to obtain the aqueous phase and precipitate, and dry the precipitate to obtain the protein ; (4) Adsorb the aqueous phase obtained in step (3) with D3520 macroporous resin for 2-4 hours, collect the effluent A, elute the D3520 macroporous resin with water, 10-20% ethanol solution in sequence, and finally use 60-80% ethanol solution Analyze the ethanol solution, collect the analysis solution, concentrate and dry to obtain Camellia oleifera polyphenols; (5) Adsorb the effluent A in step (4) with AB-8 macroporous resin for 2-5 hours, collect the effluent B, and carry out alcohol precipitation or drying on the effluent B to obtain camellia oleifera polysaccharide; (6) The AB-8 macroporous resin is eluted with water, 0.1-0.2% NaOH, and 10-30% ethanol solution in sequence, and finally analyzed with 70-95% ethanol solution, and the analytic solution is collected, concentrated and dried to obtain saponin. 3. The method for extracting saponin and polysaccharides from Camellia oleifera seed cake according to claim 1 or 2, characterized in that: in step (2), extract at 60-90°C for 2-4 hours.