LU507172B1 - A METHOD FOR THE PRODUCTION AND USE OF VITAMIN D2 FROM EDIBLE MUSHROOMS TO PROMOTE CALCIUM ABSORPTION, PREVENTION OF RICKETS AND STRENGTHENING THE IMMUNE SYSTEM - Google Patents

Abstract

This invention relates to the field of vitamin production technology, and specifically to a method for producing and applying vitamin D2 from edible mushrooms, which promotes calcium absorption, prevents rickets, and strengthens the immune system. It includes the following steps: preparation of raw material, selection of edible mushrooms with a high content of ergosterol as raw material; cultivation and optimization, genetic modification of the fermented and cultivated edible mushrooms; light conversion, irradiation of the cultures with ultraviolet light. The beneficial effect is that: by constructing a gene expression vector, transforming fungal cells, and selecting and identifying transformants with high synthesis capacity, the gene expression associated with the synthesis of ergosterol and conversion to vitamin D2 is increased. This leads to the breeding of edible mushroom varieties with higher ergosterol contents to increase the production and efficiency of vitamin D2.

Description

A process for the production and use of vitamin D2 from edible mushrooms for-V507172

Promote calcium absorption, prevent rickets and strengthen the

immune system

technical area

The present invention relates to the field of vitamin production, in particular a

Process for the production and use of vitamin D2 from edible mushrooms to promote

Calcium absorption, to prevent rickets and to strengthen the immune system.

technology in the background

Vitamin D2 appears as white to almost white flaky or needle-shaped crystals and is unstable when exposed to light or air. Vitamin D2 is mainly used for the prevention and

Treatment of vitamin D deficiency; at the same time, vitamin D2 can also be used for

Treatment of chronic hypocalcemia, hypophosphatemia, rickets and osteomalacia in

Association with chronic ~~ renal insufficiency, familial hypophosphatemia,

Hypoparathyroidism (postoperative, idiopathic or pseudohypoparathyroidism); vitamin D2 is also effective in acute, chronic and potential postoperative tetany and idiopathic tetany.

In existing technology, the production processes for vitamin D2 are more efficient and environmentally friendly. Researchers will continue to explore new extraction and refining methods to improve the purity and activity of vitamin D2 while

to reduce energy consumption and emissions during the production process.

For example, advanced technologies such as microwave-assisted

Extraction and supercritical fluid extraction allow for more efficient extraction of vitamin D2 from edible mushrooms. However, the optimization of the vitamin D2 source - the edible mushrooms - has been neglected, i.e., the optimization of the edible mushrooms themselves to increase the yield of extracted vitamin D2 is lacking.

Therefore, a process for the production and application of vitamin D2 from edible mushrooms is needed, which solves the problem of insufficient raw material optimization in the existing vitamin

D2 extraction; this could optimize the genetic properties of the raw materials to

to increase the yield of extracted vitamin D2.

Content of the invention

The aim of this invention is to provide a process for the preparation and application of

To provide vitamin D2 from edible mushrooms to promote calcium absorption, prevent rickets and strengthen the immune system, in order to overcome the problems addressed in the existing technology of the lack of optimization of the raw materials for vitamin D2 production.

extraction; it can optimize the genetic characteristics of the raw materials to

to increase the yield of extracted vitamin D2.

To achieve this goal, the present invention offers the following technical solution:

A process for producing vitamin D2 from edible mushrooms which promotes calcium absorption, prevents rickets and strengthens the immune system, comprising the following steps:

Raw material preparation, selection of edible mushrooms rich in ergosterol;

Cultivation and optimization, genetic modification of fermented and cultivated edible mushrooms;

Light conversion, irradiation of the culture medium with ultraviolet light;

Extraction and purification, using nanotechnology to purify vitamin D2.

Preferably, the specific steps of raw material preparation include:

Selection of edible mushrooms rich in ergosterol as raw material for the manufacture of 507172

Vitamin D2.

Preferably, the specific steps of cultivation and optimization include:

Inoculation of the fungi into a culture medium containing carbon sources, nitrogen sources and

Contains minerals and trace elements, controls the pH of the culture medium to neutral or slightly acidic, places the culture vessel in a constant temperature shaker or a fermenter, sets the appropriate temperature at 25-30° C;

Construction of gene expression carriers, transformation of fungal cells and selection and

Identification of transformants with efficient synthetic ability, increasing gene expression associated with the synthesis of ergosterol and its conversion to vitamin D2.

Preferably, the specific steps of light conversion include:

By crushing the mushrooms, the cell walls are destroyed, allowing vitamin D2 to enter the

culture medium; ethanol is used for extraction.

The use of nanofiltration membranes or nano-adsorbents enables the

Filtration and adsorption of the extract to remove impurities and ensure the purity of

to increase vitamin D2.

The vitamin D2 solution obtained after extraction and purification is

subjected to a crystallization process, the crystals are separated from the mother liquor by centrifugation and the moisture is removed by drying processes.

Preferably, the process also includes:

The dried vitamin D2 is ground and filled into well-sealed packaging containers to ensure the hygiene and safety of the product.

The use of vitamin D2 produced from edible mushrooms using this method to promote calcium absorption.

The use of vitamin D2 produced from edible mushrooms by this method for the prevention of rickets.

The use of vitamin D2 produced from edible mushrooms using this method to strengthen the immune system.

The use of vitamin D2 from edible mushrooms produced by this method to simultaneously promote calcium absorption, prevent rickets and strengthen the

immune system.

Compared to existing technology, the invention offers the following advantages:

The process proposed by the invention for the production and application of

Vitamin D2 from edible mushrooms to promote calcium absorption, prevent rickets and strengthen the immune system made possible by the construction of gene expression carriers that

Transformation of fungal cells and the selection and identification of transformants with high synthesis capacity lead to an increase in gene expression associated with the synthesis of ergosterol and the conversion to vitamin D2. Mushroom varieties with higher

ergosterol content to improve the yield and efficiency of vitamin D2; this further solves the problem of insufficient optimization of raw materials in the existing

Vitamin D2 extraction; genetic properties of the raw materials can be optimized to increase the yield of extracted vitamin D2.

Detailed description

In order to clearly and fully describe the objects, technical solutions and advantages of this invention, it is to be understood that the specific embodiments described herein represent only a portion of the embodiments of the invention and not all507172

They are intended only to explain the embodiments of the invention and not to limit the embodiments of the invention. All other embodiments that can be achieved by persons with ordinary skill in the art without creative effort are within the scope of this invention.

Example One

This invention offers a technical solution: A process for the production of vitamin

D2 from edible mushrooms, which promotes calcium absorption, prevents rickets and

The procedure includes the following steps: 1. Raw material preparation: selection of edible mushrooms rich in ergosterol as

Raw material. First, edible mushrooms rich in ergosterol must be selected as raw material for the production of vitamin D2. Ergosterol is a precursor substance of vitamin D2, which can be converted into vitamin D2 through a specific conversion process. Among the various edible mushrooms, mushrooms are often selected as a high-quality raw material for the production of vitamin D2 due to their high content, easy availability and processing. The selected mushrooms must be thoroughly cleaned to remove surface dirt and contaminants. This step is crucial for the hygiene and quality of the subsequent manufacturing process. During cleaning, running water or a special food cleaning agent is used to clean the surface of the mushrooms. After cleaning, the mushrooms must be dried to remove excess water. Drying is done either by natural

Air-drying or using a food dryer. Dried mushrooms are easier to store and transport and also facilitate subsequent chopping and

Extraction processes. The dried mushrooms must be ground to a fine powder to enable efficient extraction. The grinding is done mechanically or manually to ensure that the mushrooms are finely ground enough to

To increase extraction efficiency. The crushed mushroom powder must be carefully stored to avoid moisture, contamination and spoilage. The storage conditions should be dry, cool and well ventilated, and avoid direct sunlight. At the same time, it is important to regularly check and stir the stored mushroom powder to ensure its good condition. 2. Second, cultivation and optimization: genetic modification of fermented and cultivated edible mushrooms; to support the growth and metabolism of the mushrooms, a nutrient medium rich in carbon sources, nitrogen sources, minerals and trace elements is prepared. Carbon sources such as glucose or sucrose provide energy for the

Fungi; nitrogen sources such as yeast extract or peptone support cell growth; minerals and trace elements ensure the normal life activities of the fungi. At the same time, the pH value of the medium is adjusted to neutral or slightly acidic to maintain the natural

The selected mushrooms are placed in the prepared

Medium, whereby the inoculation process must be carried out sterilely to avoid contamination by foreign organisms. After inoculation, the culture vessel is placed in a constant temperature shaker or fermenter, with the appropriate temperature normally set between 25-30° C to ensure good growth and a good&/507172

To ensure the fungal growth. In addition, depending on the

Growth characteristics of the fungi, the shaking speed or the ventilation of the

Fermenter must be adjusted to meet the oxygen requirements of the fungi. To increase the ability of

To further improve the ability of fungi to synthesize vitamin D2, genetic engineering

Techniques are used. First, a gene expression carrier is constructed and the genes that are

Key enzymes encoding the synthesis of ergosterol and its conversion to vitamin D2 are cloned into the carrier. Then these carriers are transformed into the

fungal cells so that the fungi are able to express these genes. The transformed fungal cells must be selected by screening and identification by comparing the synthesis capabilities of different transformants in order to obtain strains with high

synthesis performance. To further increase the production of vitamin D2, the culture conditions must also be optimized. This includes adjusting the

Composition of the medium, pH value, temperature, humidity and

Light conditions to enable optimal growth and metabolism of the fungi.

The cultivated and optimized mushroom voices must be carefully stored in order to

To ensure the stability of their genetic characteristics and their sustainability. Methods such as

Freeze preservation, dry preservation or storage in liquid nitrogen can be used to store the strains for a long time. At the same time, the strains can be

Passaged and resuscitated as needed to ensure that they are continuously available for

production of vitamin D2. 3. Third, light conversion: irradiation of the culture medium with ultraviolet light;

Preparation of a professional UV light source and appropriate lighting equipment.

The selection of the UV light source is crucial because it must be ensured that the emitted UV wavelength is suitable for the conversion of ergosterol to vitamin D2 in the fungal cells. At the same time, the lighting equipment should provide stable output and adjustable light intensity to meet the needs of different fungal strains or different growth phases. Before light conversion, the fungi are cultivated to a suitable growth phase. This is usually done during the exponential growth phase or the stable growth phase of the fungi to ensure that the cells contain sufficient ergosterol as a precursor substance. At the same time, the

Concentration and homogeneity of the fungal culture medium must be controlled to ensure the consistency of the reaction during the light conversion. The key parameters for the

Light conversion includes light intensity and irradiation time. In this example, the

Light intensity is set to 10 watts per square meter, an optimized parameter that ensures efficient conversion without excessive damage to the fungal cells. The

Irradiation time depends on the characteristics of the fungal strain and the experimental

The irradiation time is adjusted to the requirements and is usually between six and fourteen hours. Too long an irradiation time may cause cell damage or degradation of the product, while too short an irradiation time may not be enough to achieve complete conversion. The prepared mushroom culture medium is placed in the lighting facility to ensure that the culture medium is evenly exposed to UV light. It is also important to control the temperature and humidity of the lighting environment to simulate the natural growth environment of the mushrooms. After the lighting has ended, the mushroom cultures must be further processed. This includes stopping the lighting,

Collection of the culture medium, extraction and purification of vitamin D2. During a64/507172

Extraction and purification process, care must be taken to ensure that the light does not damage the product to ensure the quality and purity of the final product. 4. Fourth, extraction and purification, using nanotechnology to purify 5 vitamin D2; In order to release vitamin D2 from the fungal cells, the cell wall of the

Fungi are destroyed. This is usually achieved by physical or chemical methods, such as high pressure homogenization, ultrasonic grinding or enzyme hydrolysis. It is crucial to select a grinding method suitable for the properties of the fungi to avoid the destruction of the vitamin D2 molecules. The crushed fungal cells release

Vitamin D2 and other cell contents are released into the culture medium, which facilitates the subsequent

Extraction is facilitated. A suitable solvent is then selected to extract vitamin D2 from the culture medium. Ethanol is used due to its good solubility for vitamin

D2 and its relatively low cost. During the extraction process, the appropriate temperature and duration are controlled to ensure that vitamin D2 is completely dissolved in ethanol while avoiding other unnecessary contaminants.

The extract contains vitamin D2, but may also contain some impurities. To remove these and increase the purity of vitamin D2, nanofiltration membranes or

Nano-adsorbents are used for filtration and adsorption. Nanofiltration membranes have a very high filtration accuracy and can effectively remove particles larger than their pore size; nano-adsorbents can selectively adsorb and remove certain impurities. These steps can significantly increase the purity of vitamin D2 and provide high-quality raw materials for the subsequent crystallization process. The extracted and purified vitamin D2 solution needs to be crystallized. By controlling the appropriate temperature, concentration and crystallization conditions, vitamin D2 is precipitated in crystallized form. After crystallization, the crystal is separated from the mother liquor by centrifugation. During centrifugation, the high

Rotation speed that the crystals are deposited at the bottom of the centrifuge tube, separating them from the mother liquor. Finally, the separated

Vitamin D2 crystal dried. This is usually done by vacuum drying or spray drying to remove moisture from the crystals and produce a dry

Vitamin D2 product.

Example Two

Based on Example One, the following method is proposed:

production of vitamin D2

First step: raw material preparation

Fresh mushrooms are used as raw material for the production of vitamin D2.

The mushrooms should be cleaned, dried and chopped to ensure an even

to obtain mushroom powder.

Second step: mushroom cultivation

Preparation of the nutrient medium: glucose, yeast powder, inorganic salts and

Trace elements are mixed in a specific ratio to produce a liquid nutrient medium. The pH of the nutrient medium is adjusted to 6.5.

Inoculation and cultivation: Streptomyces is inoculated into the nutrient medium and placed on a constant temperature shaker, with the temperature set to 28° C and the speed at

150 rpm. The cultivation time is 48 hours to ensure that ak 4507172

Fungi grow and metabolize sufficiently.

Third step: genetic modification

Vector construction: Genes encoding key enzymes for the synthesis of ergosterol and

conversion to vitamin D2 are cloned into an expression vector.

Transformation and screening: The constructed expression vector is transformed into Streptomyces and positive transformants are identified by resistance screening.

Expression validation: The positive transformants are cultured and their production of ergosterol and vitamin D2 is measured to select strains with high synthesis capacity.

Fourth step: light conversion

The cultured fungal cell suspensions are evenly distributed on culture dishes and then exposed to light conversion under a UV lamp. The light intensity is set to 5W/m ?, the irradiation time to 8 hours. The light stimulation

Ergosterol is converted into vitamin D2 in the fungal cells.

Fifth step: extraction and purification

Extraction: The light-converted fungal cells are crushed and treated with ethanol as

solvent. The extraction temperature is set to 40 ° C, the

Extraction time was 2 hours and the solvent to biomass ratio was 3:1.

Filtration and concentration: The extract is filtered to remove impurities and then concentrated using a rotary evaporator to obtain a crude vitamin D2 extract.

Nanotechnology purification: Using a nanofiltration membrane to filter the

crude extract to remove further impurities and increase the purity of vitamin D2.

Sixth step: crystallization and drying

The purified vitamin D2 solution is crystallized. By checking the

Crystallization conditions precipitate vitamin D2 in crystallized form. The crystals are then dried in a vacuum dryer to remove moisture and

vitamin D2 product.

Seventh step: packaging and storage

The dried vitamin D2 product is packaged using well-sealable

Packaging materials are used to ensure the hygiene and safety of the product. The packaged vitamin D2 is stored in a cool, dry and light-protected

environment to avoid moisture and light influence.

Eighth step: quality control

The vitamin D2 products manufactured are tested for quality, including purity,

Content and activity. Comparison with standard products ensures that the

Product quality meets the relevant standards and requirements.

Example Three

Application of vitamin D2 produced by a process for producing vitamin D2 from edible mushrooms, which promotes calcium absorption, prevents rickets and strengthens the immune system. The produced vitamin D2 is used to promote calcium absorption.

Example Four

Use of vitamin D2 obtained by a process for the production of vitamin D2 from

; edible mushrooms, which promotes calcium absorption, prevents rickets and strengthens the immune system507172. The vitamin D2 produced is used to prevent rickets.

Example Five

Application of vitamin D2 produced by a process for producing vitamin D2 from edible mushrooms, which promotes calcium absorption, prevents rickets and strengthens the immune system. The produced vitamin D2 is used to strengthen the immune system.

Example Six

Application of vitamin D2 produced by a process for producing vitamin D2 from edible mushrooms, which promotes calcium absorption, prevents rickets and strengthens the immune system. The produced vitamin D2 is simultaneously used to promote

Calcium absorption, prevention of rickets and strengthening of the immune system.

Comparison Example One

Enzymatic process for the production of vitamin D2, comprising the following steps:

First step: raw material preparation

Using yeast rich in ergosterol as a raw material. The yeast is cultivated, collected and dried to prepare it for subsequent use.

Second step: selection and preparation of the enzyme

Selection of an enzyme with high efficiency for the conversion of ergosterol to vitamin

D2, such as the vitamin D2 synthase enzyme. This enzyme can be genetically engineered into

Microorganisms are expressed and obtained by purification and concentration.

Third step: Enzymatic conversion

The prepared yeast mixture is mixed with an appropriate amount of enzyme solution. The reaction temperature, pH and reaction time are controlled to enable the enzymatic conversion of ergosterol to vitamin D2. The specific

Reaction conditions can be optimized depending on the properties of the enzyme and the nature of the raw material.

Fourth step: extraction and purification

After enzymatic conversion, vitamin D2 is converted into

Extraction solvents and conditions are used to extract vitamin D2 from the reaction system. Methods such as chromatography and crystallization are then used to remove impurities and increase the purity of vitamin D2.

Fifth step: drying and packaging

The purified vitamin D2 is dried to remove moisture. It is then packed in suitable packaging materials to ensure the hygiene and safety of the product. It is stored in a cool, dry environment to

Avoid exposure to moisture and light.

Sixth step: quality control

The vitamin D2 products manufactured are tested for quality, including purity,

Content and activity. Comparisons with standard products ensure that the

Product quality meets the relevant standards and requirements.

Comparison Example Two

Proposal for a photobiological production process for vitamin D2, which has the following

Steps include:

First step: raw material preparation

Selection of plant leaves or microorganisms rich in ergosterol, at4/507172

Raw material. The raw materials are cleaned, crushed and pretreated to prepare them for subsequent photobiological synthesis.

Second step: optimizing the lighting conditions

By adjusting the light intensity, wavelength and lighting duration, the

Conditions for photobiological synthesis optimized. Selection of suitable light sources and

Lighting devices to ensure the stability and uniformity of the light source.

Third step: Photobiological synthesis reaction

The prepared raw materials are placed under light conditions, whereby ergosterol is converted into vitamin D2 under the influence of light. By controlling the

Efficient photobiological synthesis is achieved by selecting the right light conditions and reaction time.

Fourth step: extraction and purification

After the photobiological synthesis reaction, vitamin D2 is extracted using suitable

Extraction solvents and methods are used to extract vitamin D2 from the raw material. Then, chromatographic separation, crystallization and other purification techniques are used to remove impurities and increase the purity of vitamin D2.

Fifth step: drying and packaging

The purified vitamin D2 is dried to remove moisture and then packaged in well-sealable packaging materials to ensure the hygiene and safety of the

product. It is stored in a cool, dry environment to protect it from

to be protected from moisture and light.

Sixth step: quality control

Quality tests are carried out on the vitamin D2 products manufactured, including purity, content and activity. Comparison with standard products ensures that the product quality meets the relevant standards and requirements.

It was found that embodiment two provides the best result by using a

Gene expression vector constructed, fungal cells transformed and transformants with high

synthesis ability. This increases gene expression associated with the

synthesis of ergosterol and its conversion to vitamin D2, and cultivates edible mushroom varieties with higher ergosterol contents to increase the production and efficiency of vitamin

D2. This further solves the problem of insufficient raw material optimization in existing vitamin D2 extraction; it enables the optimization of genetic

Properties of raw materials to increase the vitamin D2 extraction rate.

Although examples and descriptions of this invention have been presented,

It will be understood by those skilled in the art that various changes, modifications,

Substitutions and variations are possible without departing from the principles and spirit of the invention as defined by the appended claims and their equivalents.

Claims (10)

claims LU507172 1. A process for producing vitamin D2 from edible mushrooms, which promotes calcium absorption, prevents rickets and strengthens the immune system, characterized by the following steps: raw material preparation, selection of edible mushrooms with high ergosterol content as raw material; cultivation and optimization, genetic modification of the fermented and cultured edible mushrooms; light conversion, irradiation of the cultures with ultraviolet light; extraction and purification, use of nanotechnology to purify vitamin D2. 2. Process according to claim 1 for the production of vitamin D2 from edible mushrooms, which promotes calcium absorption, prevents rickets and strengthens the immune system, characterized by the specific implementation of the raw material preparation: selection of edible mushrooms with a high ergosterol content as raw material for the production of vitamin D2. 3. Process according to claim 1 for the production of vitamin D2 from edible mushrooms, which promotes calcium absorption, prevents rickets and strengthens the immune system, characterized by the specific implementation of the cultivation and optimization: inoculation of mushrooms in a nutrient medium containing carbon source, nitrogen source as well as minerals and trace elements, control of the pH of the medium to neutral or slightly acidic, placement of the culture vessel on a constant temperature shaker or in a fermenter, setting the appropriate temperature to 25-30° C; construction of a gene expression vector, transformation of the fungal cells and selection and identification of transformants with high synthesis performance, increase of gene expression associated with the synthesis of ergosterol and the conversion to vitamin D2. 4. Process according to claim 1 for the production of vitamin D2 from edible mushrooms, which promotes calcium absorption, prevents rickets and strengthens the immune system, characterized by the specific implementation of the light conversion: During the cultivation process of the mushrooms, these are exposed to ultraviolet irradiation, with a light intensity of 10 watts per square meter and an irradiation time of six to fourteen hours. 5. Process according to claim 1 for the production of vitamin D2 from edible mushrooms, which promotes calcium absorption, prevents rickets and strengthens the immune system, characterized by specifically carrying out the extraction and purification: by crushing the cell walls of the mushrooms are broken down to release vitamin D2, which is then extracted with ethanol; using nanofiltration membranes or nanoadsorbents to filter and adsorb the extract in order to remove impurities and increase the purity of vitamin D2; after extraction and purification, the vitamin D2 solution is crystallized, the crystals are separated from the mother liquor by centrifugation and dehydrated by drying. 6. Process according to claim 1 for the production of vitamin D2 from edible mushrooms, which promotes calcium absorption, prevents rickets and strengthens the immune system, characterized by: The dried vitamin D2 is ground and stored in well-sealable Packaging containers to ensure the hygiene and safety of the product LUS071 72 7. The use of vitamin D2 produced by a process according to any one of claims 1-6, which promotes calcium absorption, prevents rickets and strengthens the immune system, characterized in that the vitamin D2 produced is used to promote calcium absorption. 8. The use of vitamin D2 produced by a process according to any one of claims 1-6, which promotes calcium absorption, prevents rickets and strengthens the immune system, characterized in that the vitamin D2 produced is used for the prevention of rickets. 9. The use of vitamin D2, produced by a process according to any one of claims 1-6, which promotes calcium absorption, prevents rickets and strengthens the immune system, characterized in that the vitamin D2 produced is used to strengthen the immune system. 10. The use of vitamin D2, produced by a process according to any one of claims 1-6, which promotes calcium absorption, prevents rickets and strengthens the immune system, characterized in that the vitamin D2 produced is used simultaneously to promote calcium absorption, prevent rickets and strengthen the immune system.