CN115364161A - Tea oil glycoprotein saponin nano intestinal probiotic accelerant and preparation method and application thereof - Google Patents
Tea oil glycoprotein saponin nano intestinal probiotic accelerant and preparation method and application thereof Download PDFInfo
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Abstract
The invention discloses a tea oil glycoprotein saponin nano intestinal probiotic accelerant and a preparation method and application thereof. The preparation method comprises the following steps: extracting camellia seeds by buffer solution, passing through macroporous resin, obtaining glycoprotein by membrane dialysis and freeze drying of the solution, eluting the adsorption solution by ethanol water solution, drying in vacuum to obtain camellia saponin, adding lipase into camellia oil for hydrolysis reaction, then adding glycoprotein for reaction to obtain a camellia oil glycoprotein conjugate, respectively preparing the camellia oil glycoprotein conjugate and camellia saponin into the ethanol water solution and the buffer solution, stirring at high speed, and passing through a microporous filter membrane to obtain the camellia oil saponin-coated nanoemulsion of the camellia oil glycoprotein conjugate. The product can inhibit harmful bacteria, and promote proliferation of intestinal lactobacillus and Bacillus bifidus, so as to prevent diseases caused by intestinal harmful bacteria.
Description
Technical Field
The invention belongs to the field of medicines, and particularly relates to a tea oil glycoprotein saponin nano intestinal probiotic accelerant as well as a preparation method and application thereof.
Background
The intestinal flora is a normal microorganism in the intestinal tract of a host and is of various types. Generally, in healthy adults, the intestinal tract contains about 10 microorganisms 14 About 10 times of human body cell number, and can form a complex and unique system in vivo to regulate the normal physiological functions of hostThe composition and metabolism of (a) and (b) can also be influenced by the environment surrounding the host. The intestinal probiotics comprise lactobacillus, bifidobacterium and the like, are indispensable elements for human health, can synthesize various vitamins, participate in the digestion of food, promote the intestinal peristalsis, inhibit the growth of pathogenic flora, decompose harmful and toxic substances and the like. By regulating intestinal flora and increasing probiotics, intestinal diseases caused by various pathogenic bacteria can be inhibited, and metabolism and immunity of human body can be regulated.
The tea polysaccharide has the function of regulating intestinal flora, CN106188323A discloses a method for preparing Fuzhuan tea polysaccharide with the intestinal probiotic function, the method comprises the steps of decoloring Fuzhuan tea by adopting 80% ethanol, drying, crushing, extracting with hot water, precipitating with ethanol, redissolving, decoloring DEAE Sepharose fast flow resin, dialyzing, concentrating, and freeze-drying to obtain the Fuzhuan tea polysaccharide, and an in vitro anaerobic fermentation experiment shows that the Fuzhuan tea polysaccharide has the intestinal probiotic effect. Tea oil also has the function of regulating intestinal flora, and CN111329928A discloses a composition containing camellia oil, which has the functions of reducing blood sugar level, reducing insulin resistance and regulating intestinal flora. However, the single polysaccharide or tea oil can not provide various nutrients for the probiotics and can not inhibit the propagation of harmful bacteria such as escherichia coli and the like.
The camellia seeds contain various nutritional ingredients such as tea saponin and glycoprotein besides polysaccharide and tea oil, and after oil pressing, the cake dregs cannot be effectively utilized, so that resource waste is caused. Although there are reports of extracting theasaponin and glycoprotein from the raw materials, no product development is carried out on intestinal flora.
Disclosure of Invention
In order to overcome the defects of the prior art, the invention mainly aims to provide the tea oil glycoprotein saponin nano intestinal probiotic accelerant.
The invention also aims to provide a preparation method of the tea oil glycoprotein saponin nano intestinal probiotic accelerant.
The invention also aims to provide application of the tea oil glycoprotein saponin nano intestinal probiotic accelerant in preparation of intestinal flora regulating medicines.
A preparation method of a tea oil glycoprotein saponin nano intestinal probiotic accelerant comprises the following steps:
(1) Adding buffer solution with the mass of 8-30 times of that of the degreased tea seed cake and the pH value of 5-10 at 50-70 ℃ into the degreased tea seed cake, extracting for 1-3 h, filtering, and adsorbing the filtrate for 8-24 h by using macroporous resin;
(2) Dialyzing the unadsorbed solution obtained in the step (1) for 24-48 h by using a membrane with the molecular weight cutoff of 50-200 kDa, and freeze-drying the trapped solution for 24-48 h to obtain camellia oleifera glycoprotein; eluting the macroporous resin after adsorption by using ethanol water solution which is 1 to 3 times of the mass of the macroporous resin and has the volume fraction of 50 to 70 percent, and drying the eluent at the temperature of between 50 and 70 ℃ and under the pressure of between 0.01 and 0.1MPa for 3 to 5 hours to obtain the sasanquasaponin;
(3) Adding lipase accounting for 5-10% of the weight of the tea oil and pH 5-10 buffer solution accounting for 8-30 times of the weight of the tea oil at 50-70 ℃ for reacting for 1-3 h, adding organic solvent accounting for 5-10 times of the weight of the tea oil and camellia glycoprotein obtained in the step (2) accounting for 0.5-1.5 times of the weight of the tea oil, continuously reacting for 1-3 h, standing for 8-24 h, and concentrating and drying supernatant at 50-70 ℃ and 0.01-0.1 MPa for 3-5 h to obtain a tea oil glycoprotein conjugate;
(4) And (3) preparing the camellia saponin obtained in the step (2) and the camellia oil glycoprotein conjugate obtained in the step (3) with 5-20 times of the mass of the camellia saponin into 5-10% solution by using an ethanol water solution with the volume fraction of 60-80% and a buffer solution with the pH value of 5-10 respectively, adding the camellia saponin solution into the camellia oil glycoprotein conjugate solution under the stirring of 5000-20000 revolutions per minute, stirring for 10-60 minutes, and then passing through a 0.1-0.5 mu m microporous filter membrane to obtain the camellia oil glycoprotein saponin nano intestinal probiotic accelerant.
Preferably, the defatted tea seed cake in the step (1) is prepared by the following method: crushing the tea seed cake, sieving with a 20-30 mesh sieve, adding n-hexane, stirring, centrifuging at the rotating speed of 3000-4000 rpm for 20-30 min, and obtaining solid matter, namely the degreased tea seed cake.
Preferably, the macroporous resin in the step (1) is weak polar macroporous resin, including but not limited to AB-8, HPD100, HPD400, D101 and YKDH-2, and the dosage of the macroporous resin is 1-3 times of the mass of the defatted tea seed meal.
Preferably, the organic solvent in step (3) is one or more of ethyl acetate, tert-amyl alcohol and cyclohexane.
Preferably, the buffer solution in the steps (1), (3) and (4) is any one of acetate buffer solution, phosphate buffer solution and Tris-HCl buffer solution.
A tea oil glycoprotein saponin nano intestinal probiotic accelerant is prepared by the preparation method. The probiotic accelerant is a nano preparation which is composed of a camellia oil glycoprotein conjugate serving as a shell and camellia saponin serving as a core.
The tea oil glycoprotein saponin nano intestinal probiotic accelerant is applied to preparation of intestinal flora regulating medicines.
Preferably, the intestinal flora modulation comprises promoting proliferation of lactobacillus and/or bifidobacterium and inhibiting proliferation of escherichia coli. The probiotic accelerant can inhibit harmful bacteria after oral administration to promote the obvious proliferation of intestinal lactobacillus and bifidobacteria, thereby preventing diseases caused by the intestinal harmful bacteria.
The principle of the invention is as follows: the defatted tea seed cake is extracted by buffer solution to be mixed solution of tea-oil glycoprotein and saponin, the mixed solution is absorbed and separated by macroporous resin to obtain tea-oil saponin, the unadsorbed solution is dialyzed and separated to obtain tea-oil glycoprotein, the tea oil is hydrolyzed by lipase to obtain fatty acid, and the fatty acid and the tea-oil glycoprotein are esterified and connected in a two-phase system to form a tea-oil glycoprotein conjugate. After the camellia saponin and the camellia oil glycoprotein conjugate is subjected to nano emulsification, the camellia oil glycoprotein conjugate has amphipathy, hydrophilic glycoprotein is outside, and hydrophobic fatty acid group is connected with the camellia saponin internally, so that a nano structure with the camellia oil glycoprotein as a shell and the camellia saponin as a core is formed. The camellia oleifera glycoprotein is acidic glycoprotein, is stable in gastric juice, is hydrolyzed in intestinal tracts to release polysaccharide, protein, fatty acid and saponin, not only provides multiple nutrients for intestinal beneficial bacteria, but also inhibits harmful bacteria by the saponin, so that the proliferation of the intestinal beneficial bacteria can be promoted, and the health of a human body can be kept.
Compared with the prior art, the invention has the following advantages and effects:
(1) The invention provides a camellia oleifera glycoprotein saponin nano preparation, which can keep the stability of camellia oleifera saponin in gastric juice.
(2) The tea oil glycoprotein saponin nano preparation provided by the invention overcomes the defects that tea polysaccharide or tea oil can only provide single nutrition and cannot inhibit harmful bacteria, and can realize the proliferation of effective bacteria in intestinal tracts and the inhibition of the harmful bacteria.
(3) The method provided by the invention has mild reaction conditions and is easy for industrial production.
Detailed Description
The present invention will be described in further detail with reference to examples, but the embodiments of the present invention are not limited thereto.
The degreased tea seed cake used in the following examples is obtained by pulverizing tea seed cake, sieving with 20 mesh sieve, adding n-hexane reagent, stirring, and centrifuging at 3000 rpm for 20 min.
Example 1
(1) Adding 8kg of 50 deg.C pH5 acetic acid buffer solution into 1kg of defatted tea cake, extracting for 3h, filtering, and adsorbing the filtrate with AB-8 macroporous resin 1kg for 8h;
(2) Dialyzing the unadsorbed solution for 24h by using a membrane with the molecular weight cutoff of 50kDa, and freeze-drying the cutoff solution for 24h to obtain 170g of camellia oleifera glycoprotein; eluting the adsorbed macroporous resin with 1kg of 50% ethanol water solution, and drying the eluate at 50 deg.C under 0.01MPa for 3 hr to obtain Camellia saponin 90g;
(3) Adding 5g of lipase into 100g of tea oil and 800g of 50-DEG C pH5 acetic acid buffer solution for reacting for 3h, adding 500g of ethyl acetate and 50g of camellia glycoprotein, continuing to react for 1h, standing for 8h, concentrating and drying the supernatant for 3h at 50 ℃ and 0.01MPa to obtain 110g of tea oil glycoprotein conjugate;
(4) Respectively preparing a 5% solution from 10g of camellia saponin and 50g of a camellia oil glycoprotein conjugate by using a 70% ethanol aqueous solution and a pH5 acetic acid buffer solution in volume fraction, adding the camellia saponin solution into the camellia oil glycoprotein conjugate solution under stirring at 5000 r/min, stirring for 10min, and then filtering through a 0.1 mu m microporous filter membrane to obtain the camellia oil glycoprotein saponin nano intestinal probiotic accelerant.
Example 2
(1) Adding 30kg of 70 ℃ pH10 phosphate buffer solution into 1kg of degreased tea meal, extracting for 1h, filtering, and adsorbing the filtrate for 24h by using 3kg of HPD100 macroporous resin;
(2) Dialyzing the unadsorbed solution for 48h by using a membrane with the molecular weight cutoff of 200kDa, and freeze-drying the cutoff solution for 48h to obtain 240g of camellia oleifera glycoprotein; eluting the adsorbed macroporous resin with 9kg 70% ethanol water solution, and drying the eluate at 70 deg.C under 0.1MPa for 5 hr to obtain sasanquasaponin 125g;
(3) Adding 10g of lipase into 100g of tea oil, reacting with 3000g of 70 ℃ and pH10 phosphate buffer solution for 3h, adding 1000g of tert-amyl alcohol and 150g of camellia oleifera glycoprotein, continuing to react for 3h, standing for 24h, and concentrating and drying the supernatant at 70 ℃ under 0.1MPa for 5h to obtain 180g of tea oil glycoprotein conjugate;
(4) Respectively preparing 10% solution from 10g of sasanquasaponin and 200g of tea oil glycoprotein conjugate by using 80% ethanol water solution in volume fraction and pH10 phosphate buffer solution, adding the sasanquasaponin solution into the tea oil glycoprotein conjugate solution under the stirring condition of 20000 revolutions per minute, stirring for 60min, and then filtering through a 0.5 mu m microporous filter membrane to obtain the tea oil glycoprotein saponin nano intestinal probiotic accelerant.
Example 3
(1) Adding 20kg of 60 ℃ Tris-HCl buffer solution into 1kg of degreased tea meal, extracting for 2h, filtering, and adsorbing the filtrate for 16h by using 2kg of HPD400 macroporous resin;
(2) Dialyzing the unadsorbed solution for 36h by using a membrane with the molecular weight cutoff of 100kDa, and freeze-drying the cutoff solution for 36h to obtain 210g of camellia oleifera glycoprotein; eluting the adsorbed macroporous resin with 5kg of 60% ethanol water solution, and drying the eluate at 60 deg.C under 0.05MPa for 4 hr to obtain sasanquasaponin 110g;
(3) Adding 8g of lipase and 2000g of Tris-HCl buffer solution with the pH value of 8 at 60 ℃ into 100g of tea oil, reacting for 2 hours, adding 800g of cyclohexane and 120g of camellia oleifera glycoprotein, continuing to react for 2 hours, standing for 16 hours, and concentrating and drying supernatant at 60 ℃ and 0.05MPa for 4 hours to obtain 140g of tea oil glycoprotein conjugate;
(4) Preparing a 8% solution from 10g of sasanquasaponin and 100g of tea oil glycoprotein conjugate by using a 75% ethanol aqueous solution with volume fraction and a Tris-HCl buffer solution with pH of 8 respectively, adding the sasanquasaponin solution into the tea oil glycoprotein conjugate solution under stirring at 10000 r/min, stirring for 40min, and then filtering through a 0.3 mu m microporous filter membrane to obtain the tea oil glycoprotein saponin nano intestinal probiotic promoter.
Example 4
(1) Adding 15kg of acetic acid buffer solution with the temperature of 55 ℃ and the pH value of 6 into 1kg of degreased tea meal, extracting for 1.5h, filtering, and adsorbing the filtrate for 12h by using 1.5kg of D101 macroporous resin;
(2) Dialyzing the unadsorbed solution for 30h by using a membrane with the molecular weight cutoff of 75kDa, and freeze-drying the cutoff solution for 30h to obtain 186g of camellia oleifera glycoprotein; eluting the macroporous resin after adsorption by using 3kg of 55% ethanol aqueous solution with volume fraction, and drying the eluent at 65 ℃ and 0.1MPa for 3.5h to obtain 96g of camellia saponin;
(3) Adding 7g of lipase into 100g of tea oil and 1000g of acetic acid buffer solution with the temperature of 55 ℃ and the pH value of 5.5, reacting for 1.5h, adding 600g of tert-amyl alcohol and 70g of tea-oil glycoprotein, continuing to react for 1.5h, standing for 12h, concentrating and drying the supernatant for 3.5h at the temperature of 65 ℃ and the pressure of 0.05MPa, and obtaining 135g of tea-oil glycoprotein conjugate;
(4) Preparing 6% solution of 10g of sasanquasaponin and 80g of tea oil glycoprotein conjugate by respectively using 60% ethanol water solution and pH5.5 acetic acid buffer solution in volume fraction, adding the sasanquasaponin solution into the tea oil glycoprotein conjugate solution under stirring at 6000 rpm, stirring for 30min, and filtering through a 0.2 mu m microporous filter membrane to obtain the tea oil glycoprotein saponin nano intestinal probiotic accelerant.
Example 5
(1) Extracting 1kg of defatted tea cake with 25kg of 65 deg.C phosphate buffer solution with pH of 7.5 for 2.5 hr, filtering, and adsorbing the filtrate with YKDH-2 macroporous resin 2.5kg for 20 hr;
(2) Dialyzing the unadsorbed solution for 40h by using a membrane with the molecular weight cutoff of 150kDa, and freeze-drying the cutoff solution for 40h to obtain 213g of camellia oleifera glycoprotein; eluting the macroporous resin after adsorption by using 5kg of ethanol water solution with the volume fraction of 65%, and drying the eluent at 65 ℃ under 0.06MPa for 4.5h to obtain 107g of camellia saponin;
(3) Adding 6g of lipase into 100g of tea oil and 1500g of phosphate buffer solution with the pH value of 7.5 at 65 ℃ for reacting for 2.5h, adding 800g of cyclohexane and 100g of tea-oil glycoprotein, continuing to react for 2.5h, standing for 20h, concentrating and drying supernatant at 65 ℃ and 0.06MPa for 4.5h to obtain 153g of tea oil glycoprotein conjugate;
(4) Respectively preparing a 8% solution from 10g of camellia saponin and 160g of tea oil glycoprotein conjugate by using a 65% ethanol water solution and a pH7.5 buffer solution in volume fraction, adding the camellia saponin solution into the tea oil glycoprotein conjugate solution under the stirring of 12000 r/min, stirring for 25min, and then filtering through a 0.4 mu m microporous filter membrane to obtain the tea oil glycoprotein saponin nano intestinal probiotic accelerant.
Comparative example 1
The preparation method was performed as in example 1, except that no camellia glycoprotein was added in step (3), and the prepared product was control 1.
Comparative example 2
The procedure of example 1 was followed except that the tea oil glycoprotein conjugate was replaced with lecithin in step (4), and the product thus obtained was control 2.
Test 1:
compositional analysis of tea oil glycoprotein conjugates prepared in examples 1 to 5
The method comprises the following steps: the tea oil glycoprotein conjugates prepared in the embodiments 1 to 5 are taken, a proper amount of sugar content in the samples is measured according to the agricultural industry standard of the people's republic of China-the measurement of the content of crude polysaccharide in edible fungi (NY/T1676-2008), the protein content in the samples is measured according to the export and import inspection and quarantine industry standard of the people's republic of China-the Coomassie brilliant blue method (SN/T3926-2006) for measuring the protein content in milk, egg and bean foods, and the fatty acid content in the samples is measured according to the national food safety standard-the first method for measuring fatty acid in foods (GB 5009.168-2016).
As a result: the contents of sugar, protein and fatty acid in the tea oil glycoprotein conjugates prepared in examples 1 to 5 are shown in table 1, and each 100g of the product contains 8.4 to 11.6 percent of sugar, 10.5 to 17.4 percent of protein and 67.9 to 78.7 percent of fatty acid, which indicates that the conjugates simultaneously contain three nutritional components of sugar, protein and fatty acid.
TABLE 1 content composition of tea oil glycoprotein conjugates
| Sample source | Sugar content% | Protein content% | Content of fatty acid% |
| Example 1 | 9.7 | 11.8 | 76.4 |
| Example 2 | 10.3 | 15.3 | 73.2 |
| Example 3 | 8.4 | 10.5 | 78.7 |
| Example 4 | 9.2 | 12.6 | 71.5 |
| Example 5 | 11.6 | 17.4 | 67.9 |
And (3) testing 2:
determination of the nanoparticle size and the sasanquasaponin drug-loading rate in the tea oil glycoprotein saponin nano intestinal probiotic accelerant prepared in examples 1 to 5
The method comprises the following steps: the tea oil glycoprotein saponin nano preparation prepared in the embodiment 1-5, the comparison 1 and the comparison 2 are measured for particle size by a Malvern nano particle size analyzer, a proper amount of free saponin is removed by a micro-column centrifugation method, the content of the tea saponin is measured according to the determination (SN/T1852-2006) of the saponin content in tea saponin of the export and export inspection and quarantine industry standard of the people's republic of China, and the drug loading amount is calculated.
As a result: the grain size and the content of the camellia saponin nano intestinal probiotic accelerant prepared in the embodiments 1-5 are shown in table 2, the grain size is 219-284 nm, the drug loading is 3.7-12.6%, and the product is a tea saponin-loaded nano preparation. Control 1, no drug loading, shows that the tea oil fatty acid has no entrapment capability; control 2 has a small drug loading, indicating that the liposomes can entrap camellia saponin, but the drug loading is low.
TABLE 2 particle size and tea saponin loading of tea oil glycoprotein saponin nano intestinal probiotic accelerant
Test 3 determination of stability of tea oil glycoprotein saponin nano intestinal probiotic accelerator prepared in examples 1 to 5 in simulated gastrointestinal fluids
The method comprises the following steps: the stability of the sweet almond oil glycoprotein phenolic acid nano preparation is evaluated by adopting simulated artificial gastric juice (containing 1% pepsin and pH = 1.2) and simulated artificial intestinal juice (containing 1% trypsin and pH = 6.8). Adding 1mL of the tea oil glycoprotein saponin nano preparation prepared in examples 1-5 and the control 2 into 4mL of simulated artificial gastric juice or intestinal juice, incubating at 37 ℃ under oscillation at 100rpm, sampling 200 μ L at intervals of 2h, removing exuded free saponin by a micro-column centrifugation method, transferring the obtained filtrate into a 5 mL volumetric flask, diluting with methanol to a constant volume, and performing ultrasonic demulsification. The content of saponin is determined by adopting the determination (SN/T1852-2006) of saponin content in tea saponin of the export of the entry and exit inspection and quarantine industry standard of the people's republic of China, and the percentage of the residual saponin at each time point of the preparation is calculated by taking the drug concentration at 0 time point as 100 percent.
As a result: the tea oil glycoprotein saponin nano intestinal probiotic accelerant prepared in the examples 1-5 still has the saponin more than 80% retained in simulated gastric fluid within 4h, but degrades in intestinal fluid faster, and the results are shown in table 3. The product has the function of improving the gastric juice stability, and can quickly release saponin in the intestinal tract. Control 2 released all the drugs rapidly in the gastrointestinal fluid, indicating that the liposomes were unstable in the gastrointestinal fluid.
Table 3 retention of saponin (%)% in simulated gastrointestinal fluids by tea oil glycoprotein saponin nano-intestinal probiotic accelerator
Test 4
Effect of tea oil glycoprotein saponin nano intestinal probiotic promoter prepared in example 1
The method comprises the following steps: 40 male SD rats (250 + -25 g) were randomly divided into 8 groups of 5 rats each. Normal rats were fed with diet in group 1, high-fat diet (87.6% standard diet, 2% cholesterol, 0.2% sodium cholate, 10% lard) was fed to group 2, and high-fat diet was fed to groups 3 to 8, and then tea oil, the camellia glycoprotein prepared in example 1, tea saponin, a tea oil glycoprotein conjugate, a tea oil glycoprotein saponin nano intestinal probiotic promoter, and control 2 were separately infused with stomach tea oil, twice a day at a dose of 1g/100g body weight. In 15 days and 30 days, respectively, rat feces were taken, subjected to 16sRNA metagenomic sequencing of intestinal flora and probiotics (such as bifidobacterium and lactobacillus), and abundance analysis (relative percentage) of various probiotics and escherichia coli.
As a result: the results are shown in table 4, which shows that the group 2 model rats have significantly reduced probiotics but increased escherichia coli compared with the normal group, and the groups 3-5 and 8 are improved compared with the second group, but have little difference with the normal group; the group 6 is remarkably increased, and the group 7 is optimal, which shows that the tea oil glycoprotein conjugate can provide more comprehensive nutrition for the intestinal probiotics and promote the proliferation of the intestinal probiotics, and the tea oil glycoprotein saponin nano intestinal probiotics promoter can not only provide comprehensive nutrition, but also better promote the proliferation of the intestinal probiotics by inhibiting harmful flora escherichia coli through tea saponin.
Table 4 relative abundance of probiotics in rat feces (%)
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.
Claims (8)
1. A preparation method of a tea oil glycoprotein saponin nano intestinal probiotic accelerant is characterized by comprising the following steps: the method comprises the following steps:
(1) Adding buffer solution with the mass of 8-30 times of that of the degreased tea seed cake and the pH value of 5-10 at 50-70 ℃ into the degreased tea seed cake, extracting for 1-3 h, filtering, and adsorbing the filtrate for 8-24 h by using macroporous resin;
(2) Dialyzing the unadsorbed solution obtained in the step (1) for 24-48 h by using a membrane with the molecular weight cutoff of 50-200 kDa, and freeze-drying the trapped solution for 24-48 h to obtain camellia oleifera glycoprotein; eluting the adsorbed macroporous resin with ethanol water solution which is 1 to 3 times of the mass of the macroporous resin and has the volume fraction of 50 to 70 percent, and drying the eluent at the temperature of between 50 and 70 ℃ and under the pressure of between 0.01 and 0.1MPa for 3 to 5 hours to obtain the sasanquasaponin;
(3) Adding lipase with the mass of 5-10% of the tea oil and 8-30 times of buffer solution with the pH value of 5-10 at 50-70 ℃ for reacting for 1-3 h, adding organic solvent with the mass of 5-10 times of the tea oil and 0.5-1.5 times of the tea oil glycoprotein obtained in the step (2), continuing to react for 1-3 h, standing for 8-24 h, and concentrating and drying the supernatant for 3-5 h at 50-70 ℃ and 0.01-0.1 MPa to obtain a tea oil glycoprotein conjugate;
(4) And (3) preparing the camellia saponin obtained in the step (2) and the camellia oil glycoprotein conjugate obtained in the step (3) with 5-20 times of the mass of the camellia saponin into 5-10% solution by using an ethanol water solution with the volume fraction of 60-80% and a buffer solution with the pH value of 5-10 respectively, adding the camellia saponin solution into the camellia oil glycoprotein conjugate solution under the stirring of 5000-20000 revolutions per minute, stirring for 10-60 minutes, and then passing through a 0.1-0.5 mu m microporous filter membrane to obtain the camellia oil glycoprotein saponin nano intestinal probiotic accelerant.
2. The method for preparing the tea oil glycoprotein saponin nano intestinal probiotic accelerant as claimed in claim 1, wherein the method comprises the following steps:
the macroporous resin in the step (1) is AB-8, HPD100, HPD400, D101 or YKDH-2, and the dosage of the macroporous resin is 1 to 3 times of the mass of the degreased tea seed meal.
3. The method for preparing the tea oil glycoprotein saponin nano intestinal probiotic accelerant as claimed in claim 1, wherein the method comprises the following steps:
the organic solvent in the step (3) is one or more of ethyl acetate, tertiary amyl alcohol and cyclohexane.
4. The method for preparing the tea oil glycoprotein saponin nano intestinal probiotic accelerant as claimed in claim 1, wherein the method comprises the following steps:
the buffer solution in the steps (1), (3) and (4) is any one of acetic acid buffer solution, phosphate buffer solution and Tris-HCl buffer solution.
5. The preparation method of the tea oil glycoprotein saponin nano intestinal probiotic accelerant as claimed in claim 1, is characterized in that:
the defatted tea seed cake in the step (1) is prepared by the following method: crushing the tea seed cake, sieving with a 20-30 mesh sieve, adding n-hexane, stirring, centrifuging at the rotating speed of 3000-4000 rpm for 20-30 min, and obtaining solid matter, namely the degreased tea seed cake.
6. A tea oil glycoprotein saponin nano intestinal probiotic accelerant is characterized in that: prepared by the preparation method of any one of claims 1 to 5.
7. The use of a tea oil glycoprotein saponin nano-intestinal probiotic promoter as claimed in claim 6 in the manufacture of a medicament for the modulation of intestinal flora.
8. The tea oil glycoprotein saponin nano intestinal probiotic promoter according to claim 7, wherein the application in intestinal flora regulation is characterized in that:
the intestinal flora regulation comprises promoting proliferation of lactobacillus and/or bifidobacterium and inhibiting proliferation of escherichia coli.
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