CN104839018A - Preparation method for selenium-rich hair weed product - Google Patents

Abstract

The invention belongs to the technical field of food bio-enrichment, and in particular relates to a method for preparing a selenium-enriched hair vegetable product. In the present invention, the Nostocella cells convert the inorganic selenium sodium selenite in the culture solution into organic selenium through their own transformation, which reduces the harm to the human body and improves the absorption and utilization of selenium by humans and animals. The growth rate of the Nostoc sativa cells in the culture process is 30 times that of the wild Nostoc sativa, which realizes the rapid growth and reproduction of the Nostoc sativa. The hair vegetable cells produced by the invention can be used as natural biological raw materials for selenium supplementation, and can be widely used in various health foods, beverages and new resource foods to meet the demand of human body for selenium. In the present invention, the culture solution after removing the Nostocella cells contains abundant Nostocella exopolysaccharides, which have antiviral activity, and can replace Nostocs as raw materials for producing functional food. Moreover, the selenium-enriched cultivation method of Nostocs edulis of the present invention is easy to operate, does not pollute the environment, has low cost, and is suitable for industrial production.

Description

A kind of preparation method of selenium-enriched hair vegetable product

technical field

The invention belongs to the technical field of food bio-enrichment, and in particular relates to a method for preparing a selenium-enriched Nostocella product.

Background technique

Selenium is honored as "the fire of life" by the medical and nutrition circles at home and abroad, and enjoys the reputation of "longevity element", "king of anti-cancer", "patron saint of the heart", "natural antidote" and so on. The content of selenium in human tissue is 1/10000000, but it determines the existence of life, and its great effect on human health is irreplaceable by other substances. Selenium deficiency will directly lead to the decline of human immunity. Clinical medicine has proved that more than 40 kinds of diseases that threaten human health and life are related to human selenium deficiency, such as cancer, cardiovascular disease, liver disease, cataract, pancreatic disease, diabetes, reproductive systemic diseases, etc. Whether it is animal experiments or clinical practice, it has been shown that sufficient selenium should be continuously obtained from the diet, and if it cannot be supplemented in time, the ability to cure diseases will be reduced.

Facai, also known as Nostoc hairiformis, is named for its black and slender color, like human hair, and Facai has a homonym with "facai", which has been loved by people for thousands of years. Nostoc is rich in nutritional value. Every 100g of natural Nostoc contains 22.8g of protein, which is 1.5 times that of eggs. It is low in fat and rich in calcium, iron, iodine, magnesium, zinc, selenium, manganese and other trace elements. In recent years, excessive harvesting of Nostocs has caused great harm to the ecological environment. At present, the liquid suspension culture of Nostocella cells isolated from Nostocella algae has gradually attracted people's attention. The Nostocella cell culture technology is to separate and purify Nostocella cells with vigorous division ability from the algal body of Nostocypris spp., to multiply the cells of Nostocys sativa in large quantities through liquid suspension culture, and to realize the industrial production of Nostocys sativa cells through reprocessing. During the cultivation process, the growth rate of the Nostoc sativa cells is 30 times that of the wild Nostoc sativa, which realizes the rapid growth and reproduction of the Nostoc sativa resources.

There are two forms of selenium in the cells of Nostocs: one is the ionic form of selenite, and the other is the form of organic selenium combined with organic matter. Ionized selenium can be dissolved in water and absorbed into the cells, a part of which is combined with protein and further converted into organic selenium, which is the premise for the enrichment of organic selenium in Nostocs cells. However, ionic selenium dissolved in water is particularly easy to lose during the production process, resulting in low utilization rate and insignificant economic benefits. If you ingest high levels of ionic selenium for a long time, it will not only cause waste, but also cause selenium poisoning. On the contrary, organic selenium has low toxicity, high biological activity, and is easily absorbed and utilized by humans and animals. It is a safe and efficient selenium preparation. Cells enriched with organic selenium have the characteristics of high production utilization and food safety. Organic selenium hair vegetable cells can be used for various diseases caused by selenium deficiency. Therefore, it is currently a promising way to use Nostocati cells as carriers for bioaccumulation and conversion of inorganic selenium into organic selenium.

Contents of the invention

The purpose of the present invention is to provide a preparation method of a selenium-enriched Nostoc sativa product, which can not only significantly increase the growth rate of Nostoc sativa, but also promote the absorption of selenium by the human body.

The technical scheme adopted by the present invention is: a preparation method of a selenium-enriched Nostocchi sativa product, including the step of separating and purifying the Nostocella sativa; the step of cultivating the cells of the Nostoc sativa obtained by separation and purification; and the step of treating the selenium-enriched Nostoc sativa cells ; In the step of separating and purifying Nostocchi, the cleaned and sterilized Nostoc is soaked in the basic medium for 8-11 hours, then homogenized with a glass homogenizer to obtain Nostoc cell slurry, and the Nostoc cell slurry is inoculated Purifying and culturing on the basic medium to obtain the Nostocella cells;

In the step of culturing the Nostocella cells obtained by separation and purification: under aseptic conditions, inoculate the obtained Nostocchi cells on the selenium-enriched liquid medium in the culture bottle, and then culture the Nostocia cells connected with them The bottle is placed in a constant temperature biochemical incubator with a temperature of 25°C and an average light intensity of 2000 Lux for 15 to 20 days. During the cultivation process, light and darkness are alternated every 12 hours, and then the selenium-rich hair is obtained by filtering and separating the lettuce cells Vegetable cells;

In the step of treating the selenium-enriched hair vegetable cells: washing the selenium-enriched hair vegetable cells with distilled water to remove the extracellular polysaccharides and inorganic selenium on the cell surface, and then placing the selenium-enriched hair vegetable cells at a temperature of -50 Freeze-drying in an environment of ~-55°C can produce selenium-enriched hair vegetables;

The preparation method of the selenium-enriched liquid medium is to sterilize the Na2SeO3 aqueous solution with a mass concentration of _{1 mg} /mL and add it to the basic medium until the mass concentration _of Na2SeO3 is ₂ to 40 mg/L. The selenium-enriched liquid medium is prepared when the pH value reaches 7.5.

The basic medium consists of NaNO ₃ , MgSO ₄ 7H ₂ O, CaCl ₂ 2H ₂ O, H ₃ BO ₄ , MnCl ₂ 4H ₂ O, ZnSO ₄ 7H ₂ O, CuSO ₄ 5H ₂ O, NaMoO ₄ 2H ₂ O, CoCl ₂ , EDTA-Na ₂ , FeSO ₄ 7H ₂ O, citric acid, Na ₂ SiO ₃ 9H ₂ O and K ₂ HPO ₄ , and the parts by weight of each component are respectively 250 parts of NaNO ₃ , 75 parts of MgSO ₄ 7H ₂ O, 36 parts of CaCl ₂ 2H ₂ O, 2.86 parts of H ₃ BO ₄ , 1.8 parts of MnCl ₂ 4H ₂ O, 0.2 parts of ZnSO ₄ 7H ₂ O, CuSO 0.8 parts of ₄ 5H ₂ O, 0.4 parts of NaMoO ₄ 2H ₂ O, 0.01 parts of CoCl ₂ , 1 part of EDTA-Na ₂ , 4.8 parts of FeSO ₄ 7H ₂ O, 0.6 parts of citric acid, Na ₂ SiO ₃ 9H ₂ O 58 parts, K ₂ HPO ₄ 40 parts.

Beneficial effect

1. In the present invention, the cells of Nostocella sativa convert the inorganic selenium sodium selenite in the culture medium into organic selenium through their own transformation, which reduces the harm to the human body and improves the absorption and utilization of selenium by humans and animals.

2. The growth rate of the Nostoc sativa cells in the culture process is 30 times that of the wild Nostoc sativa, which realizes the rapid growth and reproduction of the Nostoc sativa.

3. The hair vegetable cells produced by the present invention can be used as natural biological raw materials for selenium supplementation, and are widely used in various health foods, beverages and new resource foods to meet the human body's demand for selenium.

4. In the present invention, the culture medium after removing the Nostocella cells contains rich Nostocchi exopolysaccharides which have antiviral activity and can replace Nostocs as a raw material for the production of functional foods.

5. The selenium-enriched cultivation method of Nostocs edulis involved in the present invention is easy to operate, does not pollute the environment, has low cost, and is suitable for industrial production.

Detailed ways

A method for preparing a selenium-enriched Nostocchia product, comprising the steps of separating and purifying Nostocchis; culturing the cells obtained from the separation and purification; treating the selenium-enriched Nostoma cells; In the steps: soak the cleaned and sterilized Nostocchi in the basic medium for 8-11 hours, then homogenize with a glass homogenizer to obtain the Nostoc cell slurry, and inoculate the Nostoc cell slurry on the basic medium for purification and cultivation. Fat lettuce cells;

In the step of culturing the Nostocella cells obtained by separation and purification: under aseptic conditions, inoculate the obtained Nostocchi cells on the selenium-enriched liquid medium in the culture bottle, and then culture the Nostocia cells connected with them The bottle is placed in a constant temperature biochemical incubator with a temperature of 25°C and an average light intensity of 2000 Lux for 15 to 20 days. During the cultivation process, light and darkness are alternated every 12 hours, and then the selenium-rich hair is obtained by filtering and separating the lettuce cells Vegetable cells;

In the step of treating the selenium-enriched hair vegetable cells: washing the selenium-enriched hair vegetable cells with distilled water to remove the extracellular polysaccharides and inorganic selenium on the cell surface, and then placing the selenium-enriched hair vegetable cells at a temperature of -50 Freeze-drying in an environment of ~-55°C can produce selenium-enriched hair vegetables;

The preparation method of the selenium-enriched liquid medium is to sterilize the Na2SeO3 aqueous solution with a mass concentration of _{1 mg} /mL and add it to the basic medium until the mass concentration _of Na2SeO3 is ₂ to 40 mg/L. The selenium-enriched liquid medium is prepared when the pH value reaches 7.5.

The basic medium consists of NaNO ₃ , MgSO ₄ 7H ₂ O, CaCl ₂ 2H ₂ O, H ₃ BO ₄ , MnCl ₂ 4H ₂ O, ZnSO ₄ 7H ₂ O, CuSO ₄ 5H ₂ O, NaMoO ₄ 2H ₂ O, CoCl ₂ , EDTA-Na ₂ , FeSO ₄ 7H ₂ O, citric acid, Na ₂ SiO ₃ 9H ₂ O and K ₂ HPO ₄ , and the parts by weight of each component are respectively 250 parts of NaNO ₃ , 75 parts of MgSO ₄ 7H ₂ O, 36 parts of CaCl ₂ 2H ₂ O, 2.86 parts of H ₃ BO ₄ , 1.8 parts of MnCl ₂ 4H ₂ O, 0.2 parts of ZnSO ₄ 7H ₂ O, CuSO 0.8 parts of ₄ 5H ₂ O, 0.4 parts of NaMoO ₄ 2H ₂ O, 0.01 parts of CoCl ₂ , 1 part of EDTA-Na ₂ , 4.8 parts of FeSO ₄ 7H ₂ O, 0.6 parts of citric acid, Na ₂ SiO ₃ 9H ₂ O 58 parts, K ₂ HPO ₄ 40 parts.

Example 1

A method for preparing a selenium-enriched Nostocchia product, comprising the steps of separating and purifying Nostocchis; culturing the cells obtained from the separation and purification; treating the selenium-enriched Nostoma cells; In the steps: soak the cleaned and sterilized Nostocchi in the basic medium for 8 hours, then homogenize it with a glass homogenizer to obtain the cell slurry of Nostoc, inoculate the cell slurry of Nostoc on the basic medium for purification and culture to obtain the cell slurry of Nostoc. Vegetable cells;

In the step of culturing the Nostocella cells obtained by separation and purification: under aseptic conditions, inoculate the obtained Nostocchi cells on the selenium-enriched liquid medium in the culture bottle, and then culture the Nostocia cells connected with them The bottle was placed in a constant temperature biochemical incubator with a temperature of 25°C and an average light intensity of 2000 Lux for 15 days. During the cultivation process, light and darkness were alternated every 12 hours, and then filtered and separated. ;

In the step of treating the selenium-enriched hair vegetable cells: washing the selenium-enriched hair vegetable cells with distilled water to remove the extracellular polysaccharides and inorganic selenium on the cell surface, and then placing the selenium-enriched hair vegetable cells at a temperature of -50 Freeze-drying in the environment of ℃ can make selenium-enriched hair vegetables products;

The preparation method of the selenium _- enriched liquid medium is to sterilize the _Na2SeO3 aqueous solution with a mass concentration of 1mg/mL and add it to the basic medium until the mass concentration _of Na2SeO3 is _2mg /L, and adjust the pH value To 7.5 promptly make the selenium-enriched liquid medium.

The basic medium consists of NaNO ₃ , MgSO ₄ 7H ₂ O, CaCl ₂ 2H ₂ O, H ₃ BO ₄ , MnCl ₂ 4H ₂ O, ZnSO ₄ 7H ₂ O, CuSO ₄ 5H ₂ O, NaMoO ₄ 2H ₂ O, CoCl ₂ , EDTA-Na ₂ , FeSO ₄ 7H ₂ O, citric acid, Na ₂ SiO ₃ 9H ₂ O and K ₂ HPO ₄ , and the parts by weight of each component are respectively 250 parts of NaNO ₃ , 75 parts of MgSO ₄ 7H ₂ O, 36 parts of CaCl ₂ 2H ₂ O, 2.86 parts of H ₃ BO ₄ , 1.8 parts of MnCl ₂ 4H ₂ O, 0.2 parts of ZnSO ₄ 7H ₂ O, CuSO 0.8 parts of ₄ 5H ₂ O, 0.4 parts of NaMoO ₄ 2H ₂ O, 0.01 parts of CoCl ₂ , 1 part of EDTA-Na ₂ , 4.8 parts of FeSO ₄ 7H ₂ O, 0.6 parts of citric acid, Na ₂ SiO ₃ 9H ₂ O 58 parts, K ₂ HPO ₄ 40 parts.

Example 2

A method for preparing a selenium-enriched Nostocchia product, comprising the steps of separating and purifying Nostocchis; culturing the cells obtained from the separation and purification; treating the selenium-enriched Nostoma cells; In the steps: soak the cleaned and sterilized Nostocchi in the basic medium for 11 hours, then homogenize it with a glass homogenizer to obtain the cell slurry of Nostoc, inoculate the cell slurry of Nostoc on the basic medium for purification and culture to obtain the cell slurry of Nostoc. Vegetable cells;

In the step of culturing the Nostocella cells obtained by separation and purification: under aseptic conditions, inoculate the obtained Nostocchi cells on the selenium-enriched liquid medium in the culture bottle, and then culture the Nostocia cells connected with them The bottle was placed in a constant temperature biochemical incubator with a temperature of 25°C and an average light intensity of 2000 Lux for 20 days. During the cultivation process, light and darkness were alternated every 12 hours, and then filtered and separated. ;

In the step of treating the selenium-enriched hair vegetable cells: washing the selenium-enriched hair vegetable cells with distilled water to remove the extracellular polysaccharides and inorganic selenium on the cell surface, and then placing the selenium-enriched hair vegetable cells at a temperature of -50 Freeze-drying in an environment of ~-55°C can produce selenium-enriched hair vegetables;

_The preparation method of the selenium _- enriched liquid medium is to sterilize the _Na2SeO3 aqueous solution with a mass concentration _of 1mg/mL and add it to the basic medium until the mass concentration of Na2SeO3 is 40mg/L, and adjust the pH value To 7.5 promptly make the selenium-enriched liquid medium.

The basic medium consists of NaNO ₃ , MgSO ₄ 7H ₂ O, CaCl ₂ 2H ₂ O, H ₃ BO ₄ , MnCl ₂ 4H ₂ O, ZnSO ₄ 7H ₂ O, CuSO ₄ 5H ₂ O, NaMoO ₄ 2H ₂ O, CoCl ₂ , EDTA-Na ₂ , FeSO ₄ 7H ₂ O, citric acid, Na ₂ SiO ₃ 9H ₂ O and K ₂ HPO ₄ , and the parts by weight of each component are respectively 250 parts of NaNO ₃ , 75 parts of MgSO ₄ 7H ₂ O, 36 parts of CaCl ₂ 2H ₂ O, 2.86 parts of H ₃ BO ₄ , 1.8 parts of MnCl ₂ 4H ₂ O, 0.2 parts of ZnSO ₄ 7H ₂ O, CuSO 0.8 parts of ₄ 5H ₂ O, 0.4 parts of NaMoO ₄ 2H ₂ O, 0.01 parts of CoCl ₂ , 1 part of EDTA-Na ₂ , 4.8 parts of FeSO ₄ 7H ₂ O, 0.6 parts of citric acid, Na ₂ SiO ₃ 9H ₂ O 58 parts, K ₂ HPO ₄ 40 parts.

Example 3

A kind of preparation method of selenium-enriched hair vegetables product specifically comprises the following steps:

Step 1: Preparation of basic medium

1. Liquid A: Weigh 1.36g of CaCl ₂ and 3.75g of MgSO ₄ 7H ₂ O respectively, dissolve in distilled water, dilute to 500mL, and store at room temperature for later use;

2. Solution B: weigh NaMoO ₄ 2H ₂ O 20mg, ZnSO ₄ 7H ₂ O 10mg, H ₃ BO ₄ 143mg, MnCl ₂ 4H ₂ O 90mg, EDTA-Na ₂ 50mg, FeSO ₄ 7H ₂ O 240mg, CuSO ₄ 5H ₂ O 40mg, CoCl ₂ 6H ₂ O 0.5mg, citric acid 30mg, dissolve in distilled water, dilute to 500mL, refrigerate at 4°C for later use;

3. P solution: Weigh 2g of K ₂ HPO ₄ 3H ₂ O, dissolve in distilled water, dilute to 500mL, store at room temperature for later use;

4. Si solution: Weigh 2.9g of Na ₂ SiO ₃ 9H ₂ O, dissolve in distilled water, dilute to 500mL, store at room temperature for later use;

5. Accurately draw 20mL of solution A and 2mL of solution B with a pipette, add distilled water to make up to 1000mL, divide into ten 250mL Erlenmeyer flasks, each 100mL, wrap and sterilize. Moist heat sterilization: temperature 121°C, time 20min.

6. Accurately draw 20mL of P solution and 20mL of Si solution with a pipette, add 2.5g of NaNO3, add distilled water to 1000mL, divide into ten 250mL Erlenmeyer flasks, each 100mL, wrap and sterilize. Moist heat sterilization: temperature 121°C, time 20min.

7. Cool the sterilized 5 and 6 to room temperature under aseptic conditions on the ultra-clean workbench, then mix them into one bottle, shake them well, and then divide them into two bottles. The basic medium is ready.

Step 2: preparation of selenium-enriched medium

Weigh 50 mg of Na ₂ SeO ₃ , dissolve it in distilled water, dilute to 50 mL, and prepare a 1 mg/mL mother solution, and sterilize the prepared solution by filtration. Add the prepared mother liquor of sodium selenite to the basic medium, so that the concentration of sodium selenite in the medium reaches 2-40 mg/L, which is the selenium-enriched medium of Nostocchia selenium, and adjust the pH value of the medium to 7.5 .

Step 3: Separation and purification of Nostococci cell strains.

Dried hair vegetables should be washed with distilled water for 3 to 4 times before use, and then sterilized with 75% alcohol for 0.5 to 1.0 min after drying, rinsed with distilled water and soaked in basic medium for 8 to 11 hours, and then homogenized with a glass homogenizer , A large number of algal filaments and free single cells were observed under a microscope. Streak inoculation was carried out on the basic medium, and Nostocchis cell culture was obtained through repeated purification and culture. Pick the purified single algae colony on the solid medium, and put it in the basic medium to expand the culture with aeration. The cells cultured to the logarithmic growth phase were harvested and used for inoculation in this experiment.

Step 4: Cultivation of selenium-enriched hair vegetable cells

Under sterile conditions, inoculate the isolated and purified Nostocchis cells in selenium-enriched liquid medium, and then put the culture bottle connected with Nostocs cells into a constant temperature biochemical incubator at 25°C, and the culture condition is that the light cycle is 12h Light for 12 hours and dark, light intensity 2000Lux, culture for 15-20 days, let the culture medium stand still, filter and separate the lettuce cells for later use;

Step 5: Preparation of selenium-enriched hair vegetable cells

Washing the isolated Nostocchis cells with distilled water for two to three times to remove the Nostocia officinalis exopolysaccharide on the cell surface and the inorganic selenium in the culture solution. The washed Nostocchi cells are poured into a plate, and freeze-dried to obtain selenium-enriched Nostoc cells.

The results show that: the growth rate of the Nostoc sativa cells in the culture process is 30 times that of the wild Nostoc sativa, and the rapid growth and reproduction of the Nostoc sativa is realized.

Claims (2)

1. A preparation method for a selenium-enriched Nostocella product, comprising the step of separating and purifying Nostocchis; the step of cultivating the cells of Nostocchis obtained by separation and purification; the step of processing the cells of Nostocys selenium; it is characterized in that: In the step of separating and purifying Nostocchi sativa, soak the cleaned and sterilized Nostoc sativa in the basic medium for 8-11 hours, then homogenize it with a glass homogenizer to obtain the cell slurry of Nostoc sativa, and inoculate the cell slurry of Nostoc sativa on the Carry out purification and culture on the basic medium to obtain Nostocchi cells; In the step of culturing the Nostocella cells obtained by separation and purification: under aseptic conditions, inoculate the obtained Nostocchi cells on the selenium-enriched liquid medium in the culture bottle, and then culture the Nostocia cells connected with them The bottle is placed in a constant temperature biochemical incubator with a temperature of 25°C and an average light intensity of 2000 Lux for 15 to 20 days. During the cultivation process, light and darkness are alternated every 12 hours, and then the selenium-rich hair is obtained by filtering and separating the lettuce cells Vegetable cells; In the step of treating the selenium-enriched hair vegetable cells: washing the selenium-enriched hair vegetable cells with distilled water to remove the extracellular polysaccharides and inorganic selenium on the cell surface, and then placing the selenium-enriched hair vegetable cells at a temperature of -50 Freeze-drying in an environment of ~-55°C can produce selenium-enriched hair vegetables; The preparation method of the selenium-enriched liquid medium is to sterilize the Na2SeO3 aqueous solution with a mass concentration of _{1 mg} /mL and add it to the basic medium until the mass concentration _of Na2SeO3 is ₂ to 40 mg/L. The selenium-enriched liquid medium is prepared when the pH value reaches 7.5. 2. The preparation method of a selenium-enriched hair vegetable product according to claim 1, characterized in that: the basic medium is composed of NaNO ₃ , MgSO ₄ ·7H ₂ O, CaCl ₂ ·2H ₂ O, H ₃ BO ₄ , MnCl ₂ 4H ₂ O, ZnSO ₄ 7H ₂ O, CuSO ₄ 5H ₂ O, NaMoO ₄ 2H ₂ O, CoCl ₂ , EDTA-Na ₂ , FeSO ₄ 7H ₂ O, citric acid, Na ₂ SiO ₃ 9H ₂ O and K ₂ HPO ₄ , and the parts by weight of each component are 250 parts of NaNO ₃ , 75 parts of MgSO ₄ 7H ₂ O, 36 parts of CaCl ₂ 2H ₂ O, H ₃ BO ₄ 2.86 parts, MnCl ₂ 4H ₂ O 1.8 parts, ZnSO ₄ 7H ₂ O 0.2 parts, CuSO ₄ 5H ₂ O 0.8 parts, NaMoO ₄ 2H ₂ O 0.4 parts, CoCl ₂ 0.01 parts, EDTA-Na ₂ 1 part, FeSO ₄ ·7H ₂ O 4.8 parts, citric acid 0.6 parts, Na ₂ SiO ₃ ·9H ₂ O 58 parts, K ₂ HPO ₄ 40 parts.