Preparation method of ganoderma lucidum protein peptide microelement chelate

Abstract

The preparation method of the ganoderma lucidum protein peptide microelement chelate comprises the following steps: carrying out enzymolysis on ganoderma lucidum protein by adopting compound protease to prepare a ganoderma lucidum protein peptide compound solution; carrying out liquid chelation on the trace elements in the inorganic compound containing the trace elements and the ganoderma lucidum protein peptide in the ganoderma lucidum protein polypeptide compound solution by adopting a liquid chelation method, so as to prepare a ganoderma lucidum protein peptide-trace element chelation solution; and centrifuging, filtering and freeze-drying the ganoderma lucidum protein peptide-microelement chelate liquid to obtain the ganoderma lucidum protein peptide microelement chelate. According to the invention, the ganoderma lucidum protein from ganoderma lucidum is taken as a raw material, the extraction process of the ganoderma lucidum protein is optimized by an alkali-solution and acid-isolation method, and the activity of metal chelated calcium and metal chelated selenium is efficiently realized by controlling enzymolysis of compound protease and chelating active peptides by using calcium chloride and sodium selenite. The invention provides a new thought for the application of the ganoderma lucidum protein.

Description

Technical Field

[0001] The invention belongs to the technical field of functional polypeptide preparation, and in particular relates to a method for preparing a ganoderma lucidum protein peptide capable of chelating with trace element ions. Background Art

[0002] Mineral elements are an indispensable substance for maintaining the functions of organisms, including growth, development, and reproduction. They have obvious effects on the regulation of physiological functions and nutritional supplementation of organisms. Although the human body has very little demand for mineral elements, they are inseparable from the physiological health of organisms and the prevention of related physiological diseases. Phenomena such as selenium deficiency, calcium deficiency, and iron deficiency have become common phenomena in today's world, so the development of mineral element supplements has become another major topic. The chelate of polypeptides and mineral elements is composed of the mutual coordination between mineral element ions and the N-terminal amino group, amino acid side chain, C-terminal carboxyl group, and carbonyl and imino groups in the peptide chain. Compared with the chelate formed by the interaction of simple amino acids and mineral elements, it can be found that the chelate of polypeptides and mineral elements has better stability and coordination rate, and also has the advantages of high biological benefits, excellent absorption effect, excellent nutritional properties, etc., and can combine the complementary functional activities of polypeptides and mineral elements such as anti-oxidation, anti-inflammatory, antibacterial, immune enhancement, blood pressure reduction, lipid reduction and blood sugar reduction. Therefore, the chelate of polypeptides and mineral elements will have excellent development and utilization value and broad research and development prospects, and will surely become a hot topic for experts and scholars from all walks of life. Due to the high proportion of plant-based foods in my country's dietary structure and relatively low intake of dairy products, calcium deficiency often occurs, especially in some children and the elderly. Chelated calcium is calcium embedded in amino acids or polypeptides. It is not affected by plant acids, oxalic acid in food and the alkaline environment in the intestines, avoiding the formation of calcium ions. Precipitation increases the solubility and stability of calcium ions, so that it can be directly absorbed through the intestinal mucosa, improving its absorption and utilization rate. Chelated calcium has high bioavailability and fast absorption, has no effect on the absorption of other nutrients, and can enhance the nutritional value of the food itself. Domestic and foreign scholars have isolated polypeptides with metal ion chelating activity from active protease hydrolysates from different sources.

[0003] Peptides are a class of compounds with a molecular structure between amino acids and proteins, which gives proteins certain physiological functions. In human life activities, peptides are better digested and absorbed in the body than free amino acids, and have an in vivo transport system that is different from amino acids. While providing nutrients necessary for human growth and development, certain short peptides can also prevent and treat diseases and regulate human functions. These biologically active peptides are called bioactive peptides. Studies have found that many peptides from different sources have antioxidant capacity, such as hydrolyzed casein, soy protein, bovine serum albumin, ovalbumin, oilseed protein, wheat alcohol-soluble protein and corn protein, all of which have certain antioxidant properties.

[0004] Lingzhi enjoys the reputation of "immortal grass" among the people because of its nourishing, longevity, qi-replenishing and tranquilizing effects. The active ingredients contained in Lingzhi are complex. Current research focuses on the biological activity of its polysaccharides, and there is less research on Lingzhi protein. Biotechnology is used to deeply process the protein resources of Lingzhi, screen and extract and separate bioactive peptides, and then chelate them with inorganic calcium and inorganic selenium to prepare polypeptide-selenium and polypeptide-calcium chelates. This Lingzhi bioactive polypeptide-trace element chelate can significantly improve the absorption rate and utilization rate of trace elements, and also has antioxidant, antibacterial, immunomodulatory, lipid-lowering and blood sugar-lowering activities. It has high development value and application prospects, and has opened up new ways for the utilization of edible and medicinal fungi resources. Summary of the invention

[0005] The purpose of the present invention is to provide a method for preparing a ganoderma lucidum protein peptide trace element chelate.

[0006] To achieve the above-mentioned purpose and other related purposes, the technical scheme provided by the present invention is: a method for preparing a Ganoderma protein peptide trace element chelate, comprising enzymolysis of Ganoderma protein by a composite protease to prepare a Ganoderma protein polypeptide complex solution; liquid chelation of trace elements in an inorganic compound containing trace elements with Ganoderma protein peptides in the Ganoderma protein polypeptide complex solution by a liquid chelation method to prepare a Ganoderma protein peptide-trace element chelate liquid; and centrifugation, filtration and freeze-drying of the Ganoderma protein peptide-trace element chelate liquid to obtain a Ganoderma protein peptide trace element chelate.

[0007] The preferred technical scheme is: the method for extracting Ganoderma protein comprises: drying Ganoderma fruiting bodies and then crushing them, mixing the crushed Ganoderma debris with water at a w / v ratio of 1:30-50, adjusting the pH value to 8.9-9.1, ultrasonically extracting for 40-60 minutes or heating and extracting for 80-120 minutes at a temperature of 50-70°C; filtering the extract to remove Ganoderma residue; adjusting the pH value of the obtained filtrate to 2.0-3.0 for protein precipitation, standing, centrifuging to obtain precipitated protein, washing with water to neutrality, centrifuging again, and freeze-drying the precipitate to obtain Ganoderma protein powder.

[0008] The preferred technical solution is: the centrifugal speed is 4000-5000r / min, and the centrifugal time is 10-30min.

[0009] The preferred technical scheme is: the enzymatic hydrolysis method comprises: dissolving the ganoderma lucidum protein in water, adjusting the pH value to 6.5-8.5, heating to 40-60°C, then adding the composite protease, stirring evenly, enzymolyzing for 30-90 minutes, then taking it out and killing the enzyme in a boiling water bath, cooling to room temperature; centrifuging under the premise of impurities, the centrifugal condition is 4000-5000r / 10min, taking the supernatant after centrifugation, and storing it in a refrigerator at 4°C, so as to obtain the ganoderma lucidum protein polypeptide complex solution The preferred technical solution is: the activity of the composite protease used is 120u / mg, the action temperature is 40-60°C, and the action pH value is 6.5-8.5.

[0010] The preferred technical scheme is: using a liquid chelation method to carry out liquid chelation of trace elements in an inorganic compound containing trace elements with Ganoderma protein peptides in a Ganoderma protein polypeptide complex solution to prepare a Ganoderma protein peptide-trace element chelate liquid, including: using a liquid chelation method to carry out liquid chelation of calcium in calcium chloride with Ganoderma protein peptides in a Ganoderma protein polypeptide complex solution to prepare a Ganoderma protein peptide-calcium chelate liquid, and using a liquid chelation method to carry out liquid chelation of selenium in sodium selenite with Ganoderma protein peptides in a Ganoderma protein polypeptide complex solution to prepare a Ganoderma protein-selenium chelate liquid.

[0011] The preferred technical scheme is: when conducting trace element ion liquid chelation, Ganoderma lucidum protein polypeptide complex solution: inorganic trace element ion solution = 2:1 volume ratio is mixed and added, stirred evenly, adjusted to pH 7.0, reacted at 60°C for 1 hour, cooled after the reaction, centrifuged to remove impurities, added 3-5 times the volume of 95wt% ethanol, precipitated and stood for more than 12 hours, centrifuged to remove the supernatant, obtained Ganoderma lucidum protein peptide-trace element chelate liquid, and freeze-dried to obtain Ganoderma lucidum protein calcium chelate powder.

[0012] Due to the application of the above technical solution, the present invention has the following advantages compared with the prior art: The invention is based on the Ganoderma lucidum protein from Ganoderma lucidum as raw material, optimizes the extraction process of Ganoderma lucidum protein by alkali dissolution and acid precipitation, and then controls the enzymatic hydrolysis by composite protease, uses calcium chloride and sodium selenite to chelate active peptides, so that the metal chelated calcium and metal chelated selenium activities can be efficiently realized. The invention provides a new idea for the application of Ganoderma lucidum protein. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] Figure 1 It is the antioxidant capacity (free radical scavenging rate) of Ganoderma lucidum crude protein, Ganoderma lucidum chelated protein, and Ganoderma lucidum protein extract before and after enzymatic hydrolysis. DETAILED DESCRIPTION

[0014] The following is a description of the implementation of the present invention by means of specific embodiments. Those skilled in the art can easily understand other advantages and effects of the present invention from the contents disclosed in the embodiments.

[0015] See also Figure 1 . It should be noted that the structures, proportions, sizes, etc. illustrated in the drawings of this specification are only used to match the contents disclosed in the specification for people familiar with this technology to understand and read, and are not used to limit the limiting conditions under which the present invention can be implemented. Therefore, they have no substantive technical significance, and any structural modification, change in proportional relationship or adjustment of size. The following examples are provided to better understand the present invention, but are not intended to limit the present invention. The experimental methods in the following examples are all conventional methods unless otherwise specified. The experimental materials used in the following examples are all purchased from conventional biochemical reagent stores unless otherwise specified.

[0016] Example 1: A method for preparing a Ganoderma lucidum protein peptide trace element chelate Step 1: Crush 1g of Ganoderma lucidum, mix the crushed Ganoderma lucidum with ultrapure water at a w / v ratio of 1:40, adjust the pH to 9.0 with NaOH, extract in a water bath for 100 minutes at 60°C, filter with a Buchner funnel after extraction, the filtrate is an alkali-extracted Ganoderma protein solution, adjust the pH of the alkali-extracted Ganoderma protein solution to 3.0 with hydrochloric acid for protein precipitation, centrifuge at 4000r / min for 20 minutes to obtain precipitated protein, wash with water until neutral, and centrifuge again. After freeze-drying the precipitate, Ganoderma protein is obtained.

[0017] Step 2: Dissolve the Ganoderma protein in water, adjust the pH value of the Ganoderma protein solution to about 9.0 with NaOH solution, wait for the temperature to rise to 50°C, immediately add the composite protease, stir evenly, and after the enzymatic hydrolysis time is about 60 minutes, take it out and put it in a boiling water bath to inactivate the enzyme for 10 minutes. After rapid cooling to room temperature, if there are impurities, centrifuge it at 4500r / 10min. After centrifugation, take the supernatant and store it in a 4°C refrigerator to obtain the Ganoderma protein polypeptide complex solution.

[0018] Step 3: Take 4 ml of the stored Ganoderma lucidum protein peptide complex solution, add 2 ml of 0.5M calcium chloride solution, stir evenly, adjust the pH to 7.0, and continue stirring at 60°C for 1 hour. After the reaction, cool in ice water, centrifuge to remove impurities, add 3-5 times the volume of 95wt% ethanol, let the precipitation stand for 12 hours, centrifuge to remove the supernatant, and freeze-dry for 24 hours to obtain Ganoderma lucidum protein peptide-calcium chelate.

[0019] Step 4: Measure the selenium content in Ganoderma lucidum protein peptide-selenium chelate. (1) Prepare a selenium standard curve. 1. Take 0, 2, 4, 6, 8, and 10 ml of 5ug / ml selenium standard solution (toluene is used as a blank) 2. Place in a separatory funnel, dilute to 40 ml with water, adjust the pH to 2-3, add 2 ml of 5% EDTA-2NA solution 3. Add 2 ml of 0.5% 3,3-diaminobenzidine solution, place in the dark to react, shake for 50 minutes, and the solution turns yellow at this time) 4. Adjust the pH to neutral, add 10 ml of toluene and shake for 2 minutes, let it stand to separate, and measure the absorbance of the toluene layer at 420 nm. Selenium standard solution: Accurately weigh 2.194 g of dry sodium selenite, dissolve in ultrapure water, and dilute to 1 L to prepare a standard solution containing 1 g / L of selenium. Dilute to a selenium standard solution with a concentration of 5 μg / mL when used. (2) Take 0.2g of freeze-dried Ganoderma lucidum protein selenium chelate powder, add ultrapure water, and sonicate to fully dissolve. (3) Use 1M HCl to adjust the pH to 2-3. (4) Add 2ml of 5% EDTA-2NA solution. (5) Add 2ml of 0.5% DBA (3,3-diaminobenzidine) (6) Place the solution in a 60℃ shaker for 50 minutes and protect from light. (7) After adjusting the pH to 7.0-7.5, accurately add 10ml of toluene, shake well, and let stand to separate. (8) Measure the absorbance of the toluene layer at 420nm. (Toluene is used as a blank) (9) Calculation formula: The calculation formula for selenium content is: Selenium content in the sample (ug / g) = pv / mN Please make sure the formula is correct p: Standard curve query is equivalent to the standard concentration of selenium v: Sample volume obtained by toluene extraction, ml m: sample mass in g N: sample volume used for determination as a percentage of the total volume of the sample after constant volume.

[0020] Step 5: Measure the calcium content in the Ganoderma lucidum protein peptide-calcium chelate. (1) Dissolve the Ganoderma lucidum protein-calcium chelate peptide in 1 ml ultrapure water. (2) After centrifugation, take the supernatant for use. (3) Prepare Ca colorimetric solution: Take 10 ml Ca Assay buffer reagent and 10 ml MTB colorimetric solution, mix them in equal amounts, and use them immediately. Store at 4 ° C in the dark and do not store for a long time. (4) Ca addition: Three dry test tubes are used as blank tubes, standard tubes, and assay tubes respectively. Add 0.025 ml ultrapure water and 2 ml Ca colorimetric working solution to the blank tube in sequence. Add 0.025 ml of 5 mmol / L calcium standard solution and 2 ml Ca colorimetric working solution to the standard tube in sequence. Add 0.025 ml of the sample to be tested and 2 ml Ca colorimetric working solution to the assay tube in sequence. (5) Ca determination: Mix well, let stand at room temperature for 5 min, adjust to zero with a blank tube, and use a spectrophotometer to measure the absorbance of the standard tube and the measurement tube at 610 nm (i.e., A standard, A measurement). (6) Calculation formula: Ca content (mmol / L) = (A measurement / A standard) × 5.

[0021] Application Effect 1. Size: determined by SDS-PAGE protein electrophoresis analysis before and after Ganoderma lucidum enzymatic hydrolysis; The size of Ganoderma lucidum protein before enzymatic hydrolysis was about 43kDa; after enzymatic hydrolysis with composite protease, a 25kda band appeared.

[0022] 2. Antioxidant activity of Ganoderma lucidum protein peptides - free radical scavenging ability (ABTS method) Principle: The ABTS method can be used to determine the antioxidant capacity of hydrophilic and lipophilic substances. Antioxidant ingredients can react with ABTS free radicals to fade the reaction system and reduce the absorbance at 405 nm. Within a certain range, the change in absorbance is proportional to the degree of free radical removal.

[0023] The free radical scavenging rate of the sample was calculated by the formula: ABTS free radical scavenging rate D% = [A blank - (A test - A control)] ÷ A blank × 100%.

[0024] The results show that the chelated Ganoderma lucidum protein peptide has stronger antioxidant activity than the original Ganoderma lucidum protein. Generally speaking, the antioxidant capacity will be enhanced after protease hydrolysis.

[0025] Example 2: A method for preparing a Ganoderma lucidum protein peptide trace element chelate An optimized process for extracting ganoderma lucidum protein and a method for preparing optimal chelated trace elements (such as calcium and selenium).

[0026] The method for preparing the ganoderma lucidum protein chelated trace elements (such as calcium and selenium) of the present invention comprises the following steps: (1) Extraction of Ganoderma lucidum protein: extract the protein from Ganoderma lucidum by alkali dissolution and acid precipitation. (2) Enzymatic hydrolysis: hydrolyze the Ganoderma lucidum protein obtained in step 1 by using a composite protease to inactivate the enzyme and prepare a Ganoderma lucidum protein polypeptide complex solution; (3) Preparation of polypeptide-calcium chelate and selenium chelate: liquid chelation method is used to make the calcium in inorganic calcium chloride and Ganoderma lucidum protein peptide undergo liquid chelation to prepare Ganoderma lucidum protein peptide-calcium chelate liquid; liquid chelation method is used to make the selenium in sodium selenite and Ganoderma lucidum protein peptide undergo chelation to prepare Ganoderma lucidum protein-selenium chelate. (4) The Ganoderma lucidum protein peptide-trace element chelate obtained in step 3 is centrifuged, filtered, and freeze-dried to obtain the final desired chelate. (5) Measure the content of trace elements in the prepared chelate.

[0027] In step (1), the specific operation of the alkaline dissolution and acid precipitation method is as follows: the ganoderma lucidum fruiting body is dried and crushed, the crushed ganoderma lucidum debris is mixed with ultrapure water at a w / v ratio of 1:40, the pH is adjusted to 9.0 using 1M NaOH, ultrasonic extraction is performed for 50 minutes, water bath heating extraction is performed for 100 minutes, and the water bath temperature is 60°C. After extraction, the ganoderma lucidum liquid is filtered with a Buchner funnel to remove the ganoderma lucidum residue. The obtained ganoderma lucidum liquid is adjusted with 1M hydrochloric acid so that the solution pH is between 2.0 for protein precipitation, and after standing overnight for 12 hours, the precipitated protein is obtained by centrifugation, washed with water until neutral, centrifuged again, and the precipitate is freeze-dried to obtain ganoderma lucidum protein powder.

[0028] The protein content was determined using the bca protein detection kit, and the optimized conditions for the alkali dissolution and acid precipitation method were obtained.

[0029] The optimized process conditions for extracting Ganoderma lucidum protein are as follows: solid-liquid ratio of 1:40, extraction water bath temperature of 60° C., extraction time of 100 min, and alkaline solution pH of 9.0. The amount of Ganoderma lucidum protein obtained from 1 g of Ganoderma lucidum is 0.9 mg.

[0030] The centrifugal sedimentation treatment conditions are: centrifugal speed is 4500r / min, and the centrifugal time is 20min.

[0031] The specific operation steps of enzymatic hydrolysis in step (2) are as follows: dissolve the prepared Ganoderma lucidum protein in ultrapure water, adjust the pH of the Ganoderma lucidum protein solution to 6.5 with NaOH solution, heat the water bath and wait for the temperature to rise to 40°C, immediately add the composite protease, stir evenly, and take out the enzyme for about 30 minutes, then take it out and put it in a boiling water bath to inactivate the enzyme for 10 minutes, quickly cool it to room temperature, centrifuge it if there are impurities, the centrifugation condition is 4500r / 10min, take the supernatant after centrifugation and store it in a refrigerator at 4°C, and obtain the Ganoderma lucidum protein polypeptide complex solution. The activity of the composite protease used is 120u / mg, the action temperature is 50°C, and the action pH value is 7.5.

[0032] The specific operation of step (3) is as follows: when conducting trace element ion liquid chelation, a volume ratio of Ganoderma lucidum protein polypeptide complex solution: inorganic trace element ion solution = 2:1 is mixed and added, stirred evenly, the pH is adjusted to 7.0, and the reaction is continued at 60° C. for 1 hour. After the reaction, the mixture is cooled and centrifuged to remove impurities. Three times the volume of 95wt% ethanol is added, and the precipitate is allowed to stand for more than 12 hours. The supernatant is removed by centrifugation to obtain a Ganoderma lucidum protein peptide-trace element chelate liquid, and freeze-dried to obtain a Ganoderma lucidum protein calcium chelate powder.

[0033] Step (5) Measure the selenium content: Principle: Under acidic conditions, selenium reacts with 3,3′-diaminobenzidine (DAB) to form a stable yellow Se-DAB complex. Under neutral and alkaline conditions, the maximum wavelength of the organic layer system complex is 420nm through toluene extraction. The optimized extraction of Ganoderma lucidum protein peptide-selenium chelate and enzymatic hydrolysis process was carried out, with selenium content as the index. The optimized enzymatic hydrolysis conditions of chelated selenium were obtained as follows: enzymatic hydrolysis time 90min, enzymatic hydrolysis temperature 50 degrees, enzymatic hydrolysis pH 6.5, and enzyme substrate ratio 0.05%.

[0034] Step (5) Measuring calcium content: The methyl muscol blue colorimetric method utilizes the ability of calcium ions in the solution to combine with MTB (methyl muscol blue) in an alkaline solution to generate a navy / blue complex. A magnesium ion chelator is added to remove background interference from magnesium ions. The absorbance at 610 nm is detected by a spectrophotometer, and the calcium content is calculated according to the formula.

[0035] The optimized extraction of Ganoderma lucidum protein peptide-calcium chelate and enzymatic hydrolysis process is based on calcium content. The higher the concentration of calcium in the supernatant, the lower the chelation rate of calcium ions in the chelate. Therefore, the optimal enzymatic hydrolysis conditions are: enzymatic hydrolysis pH 7.5, enzymatic hydrolysis time 90 minutes, enzymatic hydrolysis temperature 40 degrees, enzyme substrate ratio 0.025% The ganoderma lucidum protein peptide of the present invention is used for chelating trace element ions, such as calcium and selenium, especially in the fields of food and medicine.

[0036] The invention also provides a calcium and selenium ion chelate product, which is prepared by chelating calcium ions and selenium ions with the ganoderma lucidum protein peptide prepared by the invention.

[0037] The invention optimizes the extraction of the enzymatic hydrolysis process of ganoderma lucidum protein peptide-calcium chelate and selenium chelate, taking the calcium content and selenium content as indicators.

[0038] Example 3: A method for preparing a Ganoderma lucidum protein peptide trace element chelate A method for preparing a ganoderma protein peptide trace element chelate comprises the following steps: enzymolysis of ganoderma protein by a composite protease to prepare a ganoderma protein polypeptide complex solution; liquid chelation of trace elements in an inorganic compound containing trace elements with ganoderma protein peptides in the ganoderma protein polypeptide complex solution by a liquid chelation method to prepare a ganoderma protein peptide-trace element chelate liquid; and centrifugation, filtration and freeze-drying of the ganoderma protein peptide-trace element chelate liquid to obtain the ganoderma protein peptide trace element chelate.

[0039] A preferred embodiment is: the method for extracting Ganoderma protein comprises: drying and crushing Ganoderma fruiting bodies, mixing the crushed Ganoderma debris with water at a w / v ratio of 1:50, adjusting the pH value to 9.1, ultrasonically extracting for 60 minutes or heating and extracting at a temperature of 70°C for 120 minutes; filtering the extract to remove Ganoderma residue; adjusting the pH value of the obtained filtrate to 3.0 for protein precipitation, standing, centrifuging to obtain precipitated protein, washing with water to neutrality, centrifuging again, and freeze-drying the precipitate to obtain Ganoderma protein powder.

[0040] The preferred embodiment is as follows: the centrifugal speed is 5000 r / min and the centrifugal time is 10 min.

[0041] The preferred embodiment is: the enzymatic hydrolysis method comprises: dissolving the ganoderma lucidum protein in water, adjusting the pH value to 8.5, heating to 60°C, then adding the composite protease, stirring evenly, enzymolyzing for 30 minutes, then taking it out and killing the enzyme in a boiling water bath, cooling to room temperature; centrifuging under the premise of impurities, the centrifugal condition is 5000r / 10min, taking the supernatant after centrifugation, and storing it in a refrigerator at 4°C, so as to obtain the ganoderma lucidum protein polypeptide complex solution The preferred embodiment is as follows: the activity of the composite protease used is 120u / mg, the action temperature is 60°C, and the action pH value is 6.5-8.5.

[0042] The preferred embodiment is: using a liquid chelation method to carry out liquid chelation of trace elements in an inorganic compound containing trace elements with Ganoderma protein peptides in a Ganoderma protein polypeptide complex solution to prepare a Ganoderma protein peptide-trace element chelate liquid, including: using a liquid chelation method to carry out liquid chelation of calcium in calcium chloride with Ganoderma protein peptides in a Ganoderma protein polypeptide complex solution to prepare a Ganoderma protein peptide-calcium chelate liquid, and using a liquid chelation method to carry out liquid chelation of selenium in sodium selenite with Ganoderma protein peptides in a Ganoderma protein polypeptide complex solution to prepare a Ganoderma protein-selenium chelate liquid.

[0043] A preferred implementation is: when conducting trace element ionic liquid chelation, Ganoderma lucidum protein polypeptide complex solution: inorganic trace element ion solution = 2:1 volume ratio is mixed and added, stirred evenly, the pH is adjusted to 7.0, the reaction is continued at 60°C for 1 hour, cooled after the reaction, centrifuged to remove impurities, 5 times the volume of 95wt% ethanol is added, the precipitation is allowed to stand for more than 12 hours, the supernatant is removed by centrifugation to obtain Ganoderma lucidum protein peptide-trace element chelate liquid, and freeze-dried to obtain Ganoderma lucidum protein calcium chelate powder.

[0044] The above description is only used to explain the preferred embodiments of the present invention, and is not intended to limit the present invention in any form. Therefore, any modifications or changes made to the present invention under the same inventive spirit should still be included in the scope of protection intended by the present invention.

Claims

1. A method for preparing a Ganoderma lucidum protein peptide trace element chelate, characterized in that: The method comprises the following steps: enzymolysis of ganoderma lucidum protein by composite protease to prepare a ganoderma lucidum protein polypeptide complex solution; liquid chelation of trace elements in an inorganic compound containing trace elements with ganoderma lucidum protein peptides in the ganoderma lucidum protein polypeptide complex solution by liquid chelation to prepare a ganoderma lucidum protein peptide-trace element chelate liquid; and centrifugation, filtration and freeze-drying of the ganoderma lucidum protein peptide-trace element chelate liquid to obtain a ganoderma lucidum protein peptide trace element chelate.

2. The method for preparing the Ganoderma lucidum protein peptide trace element chelate according to claim 1, characterized in that: The method for extracting ganoderma protein comprises: drying and crushing ganoderma fruiting bodies, mixing the crushed ganoderma scraps with water at a w / v ratio of 1:30-50, adjusting the pH value to 8.9-9.1, ultrasonically extracting for 40-60 minutes or heating and extracting for 80-120 minutes at a temperature of 50-70°C; filtering the extract to remove ganoderma residue; adjusting the pH value of the obtained filtrate to 2.0-3.0 for protein precipitation, standing, centrifuging to obtain precipitated protein, washing with water to neutrality, centrifuging again, and freeze-drying the precipitate to obtain ganoderma protein powder.

3. The method for preparing the Ganoderma lucidum protein peptide trace element chelate according to claim 2, characterized in that: The centrifugal speed is 4000-5000r / min, and the centrifugal time is 10-30min.

4. The method for preparing the Ganoderma lucidum protein peptide trace element chelate according to claim 1, characterized in that: The enzymatic hydrolysis method comprises: dissolving the ganoderma protein in water, adjusting the pH value to 6.5-8.5, heating to 40-60°C, then adding the composite protease, stirring evenly, enzymolyzing for 30-90 minutes, then taking it out and putting it into a boiling water bath to inactivate the enzyme, and cooling it to room temperature; centrifuging under the premise of impurities, the centrifugal condition is 4000-5000r / 10min, taking the supernatant after centrifugation, and storing it in a refrigerator at 4°C to obtain the ganoderma protein polypeptide complex solution.

5. The method for preparing the Ganoderma lucidum protein peptide trace element chelate according to claim 4, characterized in that: The activity of the composite protease used is 120u / mg, the action temperature is 40-60°C, and the action pH value is 6.5-8.

5.

6. The method for preparing the Ganoderma lucidum protein peptide trace element chelate according to claim 1, characterized in that: The invention adopts a liquid chelation method to make the trace elements in the inorganic compound containing trace elements and the ganoderma protein peptide in the ganoderma protein polypeptide complex solution to carry out liquid chelation to prepare the ganoderma protein peptide-trace element chelate liquid, which comprises: adopting a liquid chelation method to make the calcium in the calcium chloride and the ganoderma protein peptide in the ganoderma protein polypeptide complex solution to carry out liquid chelation to prepare the ganoderma protein peptide-calcium chelate liquid, and adopting a liquid chelation method to make the selenium in the sodium selenite and the ganoderma protein peptide in the ganoderma protein polypeptide complex solution to carry out liquid chelation to prepare the ganoderma protein-selenium chelate liquid.

7. The method for preparing the Ganoderma lucidum protein peptide trace element chelate according to claim 1, characterized in that: When conducting trace element ionic liquid chelation, Ganoderma lucidum protein polypeptide complex solution: inorganic trace element ion solution = 2:1 volume ratio is mixed and added, stirred evenly, adjusted to pH 7.0, reacted at 60°C for 1 hour, cooled after the reaction, centrifuged to remove impurities, added 3-5 times the volume of 95wt% ethanol, precipitated and stood for more than 12 hours, centrifuged to remove the supernatant, obtained Ganoderma lucidum protein peptide-trace element chelate liquid, and freeze-dried to obtain Ganoderma lucidum protein calcium chelate powder.

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