A uric acid-lowering composition containing gardenia and preparation method thereof

Abstract

The invention proposes a uric acid lowering composition containing gardenia and a preparation method thereof, and belongs to the field of medical technology. After mulberry leaves, chicory, gardenia and kudzu root are fermented by Lactobacillus rhamnosus and Bifidobacterium longum, probiotics and fermentation broth are separated, immobilized ionic liquid is added to the fermentation broth, carbon dioxide is introduced for adsorption, filtration is performed, solids are added to ethanol, heating for desorption, and concentration is performed to obtain extract 1, and the filtrate is ethanol precipitated to obtain extract 2, which is embedded with probiotics together with sodium alginate to obtain a uric acid lowering embedding material, and evenly mixed with extract 1, vitamin C, and tea polyphenols to obtain a uric acid lowering composition containing gardenia. The uric acid lowering composition containing gardenia prepared by the invention can promote uric acid excretion, inhibit sodium urate crystallization, improve antioxidant effect, repair intestinal barrier, reduce inflammation, inhibit xanthine oxidase (XOD) activity and expression, reduce the level of blood uric acid, has a good uric acid lowering effect, and has broad application prospects.

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to a uric acid-lowering composition containing gardenia and a preparation method thereof. Background Art

[0002] Hyperuricemia refers to blood uric acid levels higher than 420 μmol / L in men and higher than 360 μmol / L in women. Gout refers to the deposition of monosodium urate crystals in joints, kidneys and other parts of the body, which can cause gouty nephropathy, gouty arthritis and other diseases. Gout is directly related to hyperuricemia.

[0003] At present, lowering the concentration of uric acid in the blood is the main method for treating gout. There are two main types of drugs: one is the inhibitor of xanthine oxidase, a key enzyme in the process of uric acid synthesis, and the other is the drug that promotes the excretion of uric acid in the human body. There are several commonly used drugs in clinical practice to inhibit the activity of xanthine oxidase. However, studies have also found that most of these drugs have adverse effects on normal metabolism and are prone to cause serious side effects, such as liver and kidney damage and allergic reactions, which brings great limitations to clinical treatment.

[0004] Traditional Chinese medicine believes that gout is a rheumatic arthralgia syndrome, which is mainly caused by exogenous wind, cold and dampness. Long-term illness obstructs the meridians and the circulation of qi, blood and body fluids. It is mostly caused by improper diet, excessive consumption of fatty and sweet food, internal dampness and turbidity, and heat over time. The pathogenesis of gout is mainly related to dampness, phlegm and blood stasis, and heat toxicity. Western medicine is currently used as the main treatment for gout. Commonly used ones include colchicine, non-steroidal anti-inflammatory analgesics, adrenal glucocorticoids and probenecid. Although these Western medicines are effective quickly, most of them only treat the symptoms and not the root cause. After stopping the medication, the pain is easy to relapse. Moreover, these drugs often have large toxic side effects. In addition to causing gastrointestinal reactions, long-term use may also lead to side effects such as bone marrow suppression, liver cell damage, mental depression, ascending paralysis, respiratory depression, etc. Summary of the invention

[0005] The purpose of the present invention is to provide a uric acid-lowering composition containing Gardenia jasminoides and a preparation method thereof, which can promote uric acid excretion, inhibit sodium urate crystallization, improve antioxidant effect, repair intestinal barrier, reduce inflammation, inhibit xanthine oxidase (XOD) activity and expression, and reduce blood uric acid levels. It has a good uric acid-lowering effect and has broad application prospects.

[0006] The technical solution of the present invention is achieved in this way:

[0007] The invention provides a preparation method of a uric acid lowering composition containing gardenia. The method comprises the following steps: fermenting mulberry leaf, chicory, gardenia and kudzu root with Lactobacillus rhamnosus and Bifidobacterium longum, separating probiotics and fermentation liquid, adding immobilized ionic liquid into the fermentation liquid, introducing carbon dioxide for adsorption, filtering, adding solid into ethanol, heating for desorption, concentrating to obtain extract 1, ethanol precipitation of the filtrate to obtain extract 2, embedding probiotics with sodium alginate to obtain uric acid lowering embedding material, and uniformly mixing with extract 1, vitamin C and tea polyphenols to obtain the uric acid lowering composition containing gardenia.

[0008] As a further improvement of the present invention, the following steps are included:

[0009] S1. Wash, dry and crush mulberry leaves, chicory, gardenia and kudzu root to prepare a mixed powder;

[0010] S2. activating Lactobacillus rhamnosus and Bifidobacterium longum to obtain a bacterial seed solution;

[0011] S3. The mixed powder is added to water, sterilized, Lactobacillus rhamnosus and Bifidobacterium longum seed solution is added, fermented, filtered, washed with solids, the culture solution is collected, freeze-dried, and probiotics are obtained, and the filtrate is retained;

[0012] S4. The immobilized ionic liquid was added to the filtrate, carbon dioxide was adsorbed, filtered, the filtrate was retained, the solid was added to ethanol, heated for desorption, filtered, the solid was washed, dried, and reused, and the filtrate was decompressed to remove the solvent to obtain extract 1;

[0013] S5. The filtrate was precipitated by adding ethanol, washed, and dried to obtain extract 2;

[0014] S6. The extract 2 and sodium alginate were dissolved in water, probiotics and lecithin were added, the mixture was stirred and evenly mixed, and the mixture was added dropwise to the fish oil, emulsified, and calcium chloride solution was added dropwise, solidified at room temperature, filtered, washed, and dried to obtain a uric acid-lowering embedding material;

[0015] S7. The extract 1, the uric acid-lowering embedding material, vitamin C and tea polyphenols are mixed evenly to prepare a uric acid-lowering composition containing gardenia.

[0016] As a further improvement of the present invention, the mass ratio of mulberry leaf, chicory, gardenia and kudzu root in step S1 is 7-10:12-15:14-20:4-7; the activation conditions in step S2 are 28-33°C, 100-150r / min, under anaerobic conditions, the activation treatment is 18-24h, and the bacterial content of the bacterial seed liquid is 10 8 -10 9 cfu / mL.

[0017] As a further improvement of the present invention, the solid-liquid ratio of the mixed powder and water in step S3 is 1:7-10 g / mL, the inoculation amounts of the rhamnosus Lactobacillus and longum Bifidobacterium seed liquids are 1-3 v / v% and 2-4 v / v%, respectively, and the fermentation culture conditions are 35-37°C, 100-150 r / min, and anaerobic conditions for fermentation culture for 36-48 h.

[0018] As a further improvement of the present invention, the preparation method of the immobilized ionic liquid in step S4 is as follows:

[0019] T1. Adding silica gel to hydrochloric acid, heating to reflux, filtering, washing, and drying to obtain activated silica gel;

[0020] T2. Add activated silica gel to water, add tannic acid and a catalyst, heat and stir to react, filter, wash, and dry to obtain modified silica gel;

[0021] T3. Add the modified silica gel to water, add NHS and EDC, stir to activate, add 1-butyl-3-methylimidazole L-proline, stir to react, filter, wash, and dry to obtain an amino acid imidazole immobilized material;

[0022] T4. Mix the amino acid imidazole immobilized material and salt and add them into acetonitrile, stir for reaction, filter, wash and dry to obtain an immobilized ionic liquid.

[0023] As a further improvement of the present invention, the concentration of hydrochloric acid in step T1 is 1-2 mol / L, and the heating reflux time is 3-4 h; the mass ratio of activated silica gel, tannic acid and catalyst in step T2 is 10:3-5:0.2-0.3, the temperature of the heating and stirring reaction is 40-50°C, the time is 2-4 h, and the catalyst is a Tris-HCl solution with a pH of 8.5-9.5; the mass ratio of modified silica gel, NHS, EDC, and 1-butyl-3-methylimidazole L-proline in step T3 is 10:2-3:1-3:3-4, the stirring activation time is 20-30 min, and the stirring reaction time is 7-10 h; the mass ratio of amino acid imidazole immobilization material and salt in step T4 is 1:2-3, the salt is NaBF4 or KPF6, and the stirring reaction time is 20-24 h.

[0024] As a further improvement of the present invention, in step S4, the mass ratio of the filtrate to the immobilized ionic liquid is 10:2-3, the ventilation rate of the carbon dioxide is 3-5 mL / min, the adsorption time is 30-50 min, and the heating desorption temperature is 40-50°C.

[0025] As a further improvement of the present invention, in step S5, ethanol is added until the ethanol content of the system is 70-80wt%, and precipitation is carried out for 1-2h; in step S7, the mass ratio of extract 1, uric acid-lowering embedding material, vitamin C, and tea polyphenols is 10-12:15-20:2-3:1-3.

[0026] As a further improvement of the present invention, the mass ratio of the extract 2, sodium alginate, probiotics and lecithin in step S6 is 7-10:8-12:10-15:1-2, and the room temperature curing time is 20-30 minutes.

[0027] The present invention further protects a uric acid lowering composition containing gardenia prepared by the above preparation method.

[0028] The present invention has the following beneficial effects:

[0029] Uric acid is the final product of purine metabolism. The intestine degrades about 1 / 3 of urate every day, and has great potential for uric acid excretion. The intestinal microecology is the key to the intestinal degradation of uric acid. It is composed of intestinal microbial flora, intestinal metabolites, intestinal barriers and intestinal uric acid transporters. Intestinal microorganisms participate in purine metabolism and uric acid decomposition to reduce uric acid levels. The intestinal flora metabolizes uric acid into short-chain fatty acids (SCFAs) such as acetate, butyrate and propionate, succinate, xanthine and lactate, participates in energy metabolism, can provide energy for intestinal epithelial cells, and promotes uric acid excretion. The rhamnosus lactobacillus of the present invention promotes the decomposition and excretion of uric acid in the intestine by producing SCFAs. At the same time, Bifidobacterium longum can regulate the expression of intestinal epithelial uric acid transporter glucose transporter 9 and ATP binding cassette subfamily G member 2 to affect uric acid excretion. The byproduct butyrate can nourish the intestinal mucosa and intestinal villi, stimulate the growth of beneficial bacteria and inhibit the growth of harmful bacteria to support intestinal metabolic function. Bifidobacterium longum and its secretions can repair the intestinal barrier and reduce inflammation. Therefore, the two bacterial communities have a synergistic effect.

[0030] Mulberry leaf, chicory, gardenia and kudzu root are rich in polysaccharide substances, such as inulin, mulberry leaf polysaccharide, gardenia polysaccharide, kudzu root polysaccharide, etc., wherein inulin is the highest active ingredient in chicory extract, and is a water-soluble dietary fiber that regulates sugar and lipid metabolism and improves intestinal microenvironment, and can enhance the abundance of bifidobacteria and maintain intestinal health. These polysaccharide structures can be obtained by the method of alcohol precipitation, therefore, the extract 2 prepared by the present invention contains abundant polysaccharide substances, and with this as shell material, mixed with sodium alginate to embed probiotics, on the one hand, can protect probiotics, improve its stability and acid resistance, on the other hand, the extract 2 is rich in carboxylic acid group substances, which have obvious inhibitory effect on sodium urate crystallization, and at the same time, can trigger a cascade reaction so that probiotics can quickly degrade urea and accelerate intestinal uric acid excretion.

[0031] In addition, mulberry leaves, chicory, gardenia and kudzu root also contain rich active substances, such as resveratrol can increase the abundance of lactobacillus and host-associated lactobacillus, enhance cecal uric acid degradation, polyphenol extract can upregulate the relative abundance of intestinal beneficial bacteria Firmicutes and Lactobacillaceae, ferulic acid upregulates ABCG2 expression, reduces SLC2A9 to promote uric acid intestinal excretion, etc., these substances are extracted to a large extent in the raw materials through the probiotic fermentation process and enter the fermentation liquid, but the fermentation liquid has complex components and contains a large amount of other substances, resulting in a low concentration of these substances, and it is impossible to play a good effect of reducing uric acid. Therefore, the present invention prepares a kind of immobilized ionic liquid, retains the respective properties of ionic liquid and solid material, has good stability, strong extraction selectivity, high recovery rate and other characteristics, and can also solve the problem of ionic liquid residue well.

[0032] The immobilized ionic liquid prepared by the present invention activates silica gel through acid, increases the number of silanol groups on the surface of silica gel and eliminates metal oxides and nitrogen-containing impurities, and then the surface is modified by tannic acid, which is a type of highly active tannic acid and can be coupled with 1-butyl-3-methylimidazole L-proline, so that the imidazole group is fixed on the silica gel, and then the immobilized ionic liquid is prepared by salt exchange. Under the condition of introducing carbon dioxide, the polarity of the immobilized ionic liquid is improved, thereby greatly improving its adsorption capacity for active substances. After heating and desorption in ethanol, the solvent is removed under reduced pressure to obtain a high-purity active substance extract 1, which has good effects of promoting uric acid excretion, inhibiting the activity of xanthine oxidase, having anti-inflammatory effects, and can also inhibit uric acid crystallization, reduce the occurrence of gout, and reduce the symptoms of joint swelling and pain during gout attacks.

[0033] Vitamin C can lower blood uric acid levels by promoting uric acid excretion and inhibiting uric acid production. Tea polyphenols can inhibit the activity and expression of xanthine oxidase (XOD), reduce the expression of urate transporter 1 (URAT1) in the kidney, and enhance the expression of organic anion transporters (OAT) 1 and OAT3, thereby increasing uric acid excretion and further lowering blood uric acid levels. Both can also reduce inflammation caused by peroxidation through antioxidant effects, thereby promoting uric acid excretion and playing a good synergistic role.

[0034] The uric acid lowering composition containing gardenia prepared by the present invention can promote uric acid excretion, inhibit sodium urate crystallization, improve antioxidant effect, repair intestinal barrier, reduce inflammation, inhibit xanthine oxidase (XOD) activity and expression, and reduce the level of blood uric acid. It has a good uric acid lowering effect and has broad application prospects. DETAILED DESCRIPTION

[0035] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0036] Lactobacillus rhamnosus, 10 billion cfu / g, and Bifidobacterium longum, 10 billion cfu / g, were purchased from Weikang Probiotics (Suzhou) Co., Ltd.

[0037] Preparation Example 1 Preparation of immobilized ionic liquid

[0038] Here’s how:

[0039] T1. Add 10 g of silica gel to 200 mL of 1 mol / L hydrochloric acid, heat under reflux for 3 h, filter, wash, and dry to obtain activated silica gel;

[0040] T2. Add 10g activated silica gel to 200mL water, add 3g tannic acid and 0.2g catalyst, heat to 40℃, stir and react for 2h, filter, wash and dry to obtain modified silica gel;

[0041] The catalyst is a Tris-HCl solution with a pH of 8.5;

[0042] T3. 10 g of modified silica gel was added to 200 mL of water, 2 g of NHS and 1 g of EDC were added, and the mixture was stirred for activation for 20 min. 3 g of 1-butyl-3-methylimidazole L-proline was added, and the reaction was stirred for 7 h. The mixture was filtered, washed, and dried to obtain an amino acid imidazole immobilized material.

[0043] T4. 10 g of amino acid imidazole immobilized material and 20 g of NaBF4 were mixed into 200 mL of acetonitrile, stirred for 20 h, filtered, washed, and dried to obtain an immobilized ionic liquid.

[0044] Preparation Example 2 Preparation of immobilized ionic liquid

[0045] Here’s how:

[0046] T1. Add 10 g of silica gel to 200 mL of 2 mol / L hydrochloric acid, heat under reflux for 4 h, filter, wash, and dry to obtain activated silica gel;

[0047] T2. Add 10g activated silica gel into 200mL water, add 5g tannic acid and 0.3g catalyst, heat to 50℃, stir and react for 4h, filter, wash and dry to obtain modified silica gel;

[0048] The catalyst is a Tris-HCl solution with a pH of 9.5;

[0049] T3. 10 g of modified silica gel was added to 200 mL of water, 3 g of NHS and 3 g of EDC were added, and the mixture was stirred for activation for 30 min, 4 g of 1-butyl-3-methylimidazole L-proline was added, and the reaction was stirred for 10 h, filtered, washed, and dried to obtain an amino acid imidazole immobilized material;

[0050] T4. 10 g of amino acid imidazole immobilized material and 30 g of NaBF4 were mixed and added into 200 mL of acetonitrile, stirred and reacted for 24 h, filtered, washed, and dried to obtain an immobilized ionic liquid.

[0051] Preparation Example 3 Preparation of immobilized ionic liquid

[0052] Here’s how:

[0053] T1. Add 10 g of silica gel to 200 mL of 1.5 mol / L hydrochloric acid, heat under reflux for 3.5 h, filter, wash, and dry to obtain activated silica gel;

[0054] T2. Add 10g activated silica gel to 200mL water, add 4g tannic acid and 0.25g catalyst, heat to 45°C, stir and react for 3h, filter, wash and dry to obtain modified silica gel;

[0055] The catalyst is a Tris-HCl solution with a pH of 9;

[0056] T3. 10 g of modified silica gel was added to 200 mL of water, 2.5 g of NHS and 2 g of EDC were added, and the mixture was stirred for activation for 25 min. 3.5 g of 1-butyl-3-methylimidazole L-proline was added, and the reaction was stirred for 8 h. The mixture was filtered, washed, and dried to obtain an amino acid imidazole immobilized material.

[0057] T4. 10 g of amino acid imidazole immobilized material and 25 g of NaBF4 were mixed into 200 mL of acetonitrile, stirred for reaction for 22 h, filtered, washed, and dried to obtain an immobilized ionic liquid.

[0058] Comparative Preparation Example 1

[0059] Compared with Preparation Example 3, the difference is that step T1 is not performed.

[0060] The details are as follows:

[0061] T1. Add 10g silica gel to 200mL water, add 4g tannic acid and 0.25g catalyst, heat to 45°C, stir and react for 3h, filter, wash and dry to obtain modified silica gel;

[0062] The catalyst is a Tris-HCl solution with a pH of 9;

[0063] T2. 10 g of modified silica gel was added to 200 mL of water, 2.5 g of NHS and 2 g of EDC were added, and the mixture was stirred for activation for 25 min, 3.5 g of 1-butyl-3-methylimidazole L-proline was added, and the reaction was stirred for 8 h, filtered, washed, and dried to obtain an amino acid imidazole immobilized material;

[0064] T3. 10 g of amino acid imidazole immobilized material and 25 g of NaBF4 were mixed and added into 200 mL of acetonitrile. The mixture was stirred for 22 h, filtered, washed and dried to obtain an immobilized ionic liquid.

[0065] Comparative Preparation Example 2

[0066] Compared with Preparation Example 3, the difference is that step T2 is not performed.

[0067] The details are as follows:

[0068] T1. Add 10 g of silica gel to 200 mL of 1.5 mol / L hydrochloric acid, heat under reflux for 3.5 h, filter, wash, and dry to obtain activated silica gel;

[0069] T2. 10 g of activated silica gel was added to 200 mL of water, 2.5 g of NHS and 2 g of EDC were added, and the mixture was stirred for 25 min. 3.5 g of 1-butyl-3-methylimidazole L-proline was added, and the reaction was stirred for 8 h. The mixture was filtered, washed, and dried to obtain an amino acid imidazole immobilized material.

[0070] T3. 10 g of amino acid imidazole immobilized material and 25 g of NaBF4 were mixed and added into 200 mL of acetonitrile. The mixture was stirred for 22 h, filtered, washed and dried to obtain an immobilized ionic liquid.

[0071] Example 1

[0072] This embodiment provides a method for preparing a uric acid-lowering composition containing Gardenia jasminoides, comprising the following steps:

[0073] S1 7g mulberry leaves, 12g chicory, 14g gardenia and 4g Pueraria root were washed, dried and crushed to obtain a mixed powder;

[0074] S2. Lactobacillus rhamnosus and Bifidobacterium longum were activated for 18-24 hours at 28°C, 100 r / min, and anaerobic conditions to obtain a bacterial count of 10 8 -10 9 cfu / mL bacterial seed solution;

[0075] S3. Add 20 g of the mixed powder to 140 mL of water, sterilize, add Lactobacillus rhamnosus and Bifidobacterium longum seed solution, the inoculation amount is 1 v / v% and 2 v / v%, respectively, at 35°C, 100 r / min, under anaerobic conditions, ferment and culture for 36 h, filter, wash the solid, collect the bacterial solution, freeze-dry, prepare probiotics, and retain the filtrate;

[0076] S4. Add 20 g of the immobilized ionic liquid prepared in Preparation Example 1 to 100 g of the filtrate, introduce carbon dioxide for adsorption, the ventilation rate is 3 mL / min, the adsorption time is 30 min, filter, retain the filtrate, add ethanol to the solid, heat to 40 ° C, desorb, filter, wash the solid, dry, reuse, and remove the solvent from the filtrate under reduced pressure to obtain Extract 1;

[0077] S5. The filtrate was adjusted to a system ethanol content of 70 wt%, precipitated for 1 h, washed, and dried to obtain an extract 2;

[0078] S6. 7 g of extract 2, 8 g of sodium alginate was dissolved in 300 mL of water, 10 g of probiotics and 1 g of lecithin were added, the mixture was stirred for 15 min, and then added dropwise to 500 mL of fish oil, emulsified at 8000 r / min for 15 min, 20 mL of 2 wt% calcium chloride solution was added dropwise, and the mixture was cured at room temperature for 20 min, filtered, washed, and dried to obtain a uric acid-lowering embedding material;

[0079] S7. 10 g of extract 1, 15 g of uric acid-lowering embedding material, 2 g of vitamin C, and 1 g of tea polyphenols were mixed for 15 minutes to prepare a uric acid-lowering composition containing gardenia.

[0080] Example 2

[0081] This embodiment provides a method for preparing a uric acid-lowering composition containing Gardenia jasminoides, comprising the following steps:

[0082] S1. Wash 10g mulberry leaves, 15g chicory, 20g gardenia and 7g Pueraria root, dry and crush to obtain a mixed powder;

[0083] S2. Lactobacillus rhamnosus and Bifidobacterium longum were activated at 33°C, 150 r / min, and anaerobic conditions for 18-24 hours to obtain a bacterial content of 10 8 -10 9 cfu / mL bacterial seed solution;

[0084] S3. Add 20 g of the mixed powder to 200 mL of water, sterilize, add Lactobacillus rhamnosus and Bifidobacterium longum seed solution, the inoculation amount is 3 v / v% and 4 v / v%, respectively, at 37 ° C, 150 r / min, under anaerobic conditions, ferment and culture for 48 hours, filter, wash the solid, collect the bacterial solution, freeze-dry, prepare probiotics, and retain the filtrate;

[0085] S4. Add 30 g of the immobilized ionic liquid prepared in Preparation Example 2 to 100 g of the filtrate, introduce carbon dioxide for adsorption, the ventilation rate is 5 mL / min, the adsorption time is 50 min, filter, retain the filtrate, add ethanol to the solid, heat to 50 ° C, desorb, filter, wash the solid, dry, reuse, and remove the solvent from the filtrate under reduced pressure to obtain Extract 1;

[0086] S5. The filtrate was adjusted to an ethanol content of 80 wt % and precipitated for 2 h, washed, and dried to obtain an extract 2;

[0087] S6. 10 g of extract 2 and 12 g of sodium alginate were dissolved in 300 mL of water, 15 g of probiotics and 2 g of lecithin were added, the mixture was stirred for 15 min, and then added dropwise to 500 mL of fish oil. The mixture was emulsified at 8000 r / min for 15 min, and 20 mL of 3 wt% calcium chloride solution was added dropwise. The mixture was cured at room temperature for 30 min, filtered, washed, and dried to obtain a uric acid-lowering embedding material.

[0088] S7. 12 g of the extract 1, 20 g of the uric acid-lowering embedding material, 3 g of vitamin C, and 3 g of tea polyphenols were mixed for 15 minutes to prepare a uric acid-lowering composition containing gardenia.

[0089] Example 3

[0090] This embodiment provides a method for preparing a uric acid-lowering composition containing Gardenia jasminoides, comprising the following steps:

[0091] S1 8g mulberry leaves, 13g chicory, 17g gardenia and 5g Pueraria root were washed, dried and crushed to obtain a mixed powder;

[0092] S2. Lactobacillus rhamnosus and Bifidobacterium longum were activated at 30°C, 120 r / min, and anaerobic conditions for 18-24 hours to obtain a bacterial content of 10 8 -10 9 cfu / mL bacterial seed solution;

[0093] S3. Add 20 g of the mixed powder to 170 mL of water, sterilize, add Lactobacillus rhamnosus and Bifidobacterium longum seed solution, the inoculation amount is 2 v / v% and 3 v / v%, respectively, at 36 ° C, 120 r / min, under anaerobic conditions, ferment and culture for 42 hours, filter, wash the solid, collect the bacterial solution, freeze-dry, prepare probiotics, and retain the filtrate;

[0094] S4. Add 25 g of the immobilized ionic liquid prepared in Preparation Example 3 to 100 g of the filtrate, introduce carbon dioxide for adsorption, the ventilation rate is 4 mL / min, the adsorption time is 40 min, filter, retain the filtrate, add ethanol to the solid, heat to 45 ° C, desorb, filter, wash the solid, dry, reuse, and remove the solvent from the filtrate under reduced pressure to obtain Extract 1;

[0095] S5. The filtrate was adjusted to an ethanol content of 75 wt % and precipitated for 1.5 h, washed, and dried to obtain an extract 2;

[0096] S6. 8 g of extract 2, 10 g of sodium alginate were dissolved in 300 mL of water, 12 g of probiotics and 1.5 g of lecithin were added, stirred and mixed for 15 min, added dropwise to 500 mL of fish oil, emulsified at 8000 r / min for 15 min, 20 mL of 2.5 wt% calcium chloride solution was added dropwise, cured at room temperature for 25 min, filtered, washed, and dried to obtain a uric acid-lowering embedding material;

[0097] S7. 11 g of the extract 1, 17 g of the uric acid-lowering embedding material, 2.5 g of vitamin C, and 2 g of tea polyphenols were mixed for 15 minutes to prepare a uric acid-lowering composition containing gardenia.

[0098] Comparative Example 1

[0099] Compared with Example 3, the difference is that the immobilized ionic liquid is prepared by Comparative Preparation Example 1.

[0100] Comparative Example 2

[0101] Compared with Example 3, the difference is that the immobilized ionic liquid is prepared by Comparative Preparation Example 2.

[0102] Comparative Example 3

[0103] Compared with Example 3, the difference is that the Lactobacillus rhamnosus seed liquid is not inoculated in step S3.

[0104] The details are as follows:

[0105] S3. Add 20g of the mixed powder into 170mL of water, sterilize, add Bifidobacterium longum seed solution with an inoculation rate of 5v / v%, culture at 36℃, 120r / min under anaerobic conditions, ferment for 42h, filter, wash the solid, collect the bacterial solution, freeze-dry, obtain probiotics, and retain the filtrate.

[0106] Comparative Example 4

[0107] Compared with Example 3, the difference is that the seed liquid of Bifidobacterium longum is not inoculated in step S3.

[0108] The details are as follows:

[0109] S3. Add 20g of the mixed powder into 170mL of water, sterilize it, add Lactobacillus rhamnosus seed solution with an inoculation rate of 5v / v%, culture it at 36℃, 120r / min under anaerobic conditions for 42h, filter it, wash the solid, collect the bacterial solution, freeze-dry it to obtain probiotics, and retain the filtrate.

[0110] Comparative Example 5

[0111] Compared with Example 3, the difference is that fermentation is not performed in step S3, and heating and boiling extraction is adopted.

[0112] The details are as follows:

[0113] S3. Add 20g of the mixed powder into 170mL of water, heat and boil for 2h, filter, and keep the filtrate.

[0114] Comparative Example 6

[0115] Compared with Example 3, the difference is that no carbon dioxide is introduced in step S4.

[0116] The details are as follows:

[0117] S4. Add 25 g of the immobilized ionic liquid prepared in Preparation Example 3 to 100 g of the filtrate for adsorption. The adsorption time is 40 min. Filter and retain the filtrate. Add the solid to ethanol and heat to 45°C for desorption. Filter and wash the solid, dry and reuse. Remove the solvent from the filtrate under reduced pressure to obtain Extract 1.

[0118] Comparative Example 7

[0119] Compared with Example 3, the difference is that dichloromethane is added for extraction in step S4.

[0120] The details are as follows:

[0121] S4. To 100 g of the filtrate was added 500 mL of dichloromethane for extraction, and the mixture was filtered. The solvent was removed from the filtrate under reduced pressure to obtain an extract 1.

[0122] Comparative Example 8

[0123] Compared with Example 3, the difference is that the extract 2 is not added in step S6.

[0124] The details are as follows:

[0125] S6. Dissolve 18g sodium alginate in 300mL water, add 12g probiotics and 1.5g lecithin, stir and mix for 15min, add dropwise into 500mL fish oil, emulsify at 8000r / min for 15min, add dropwise 20mL 2.5wt% calcium chloride solution, solidify at room temperature for 25min, filter, wash and dry to obtain uric acid-lowering embedding material.

[0126] Comparative Example 9

[0127] Compared with Example 3, the difference is that the extract 1 is not added in step S7.

[0128] The details are as follows:

[0129] S7. 28 g of the uric acid-lowering embedding material, 2.5 g of vitamin C, and 2 g of tea polyphenols were mixed for 15 minutes to prepare a uric acid-lowering composition containing gardenia.

[0130] Comparative Example 10

[0131] Compared with Example 3, the difference is that no uric acid-lowering embedding material is added in step S7.

[0132] The details are as follows:

[0133] S7. 28 g of the extract 1, 2.5 g of vitamin C, and 2 g of tea polyphenols were mixed for 15 minutes to prepare a uric acid-lowering composition containing gardenia.

[0134] Test Example 1: Sustained-release test

[0135] 1g of the uric acid-lowering embedding material prepared in Examples 1-3 of the present invention and Comparative Example 8 was added to 9mL of artificial simulated gastric juice and 9mL of artificial simulated intestinal juice, respectively, and reacted at 37°C and 100r / min for 2h and 3h, respectively. In addition, an equal amount of the uric acid-lowering embedding material was added to 9mL of artificial simulated gastric juice, first placed in a shaker, reacted at 37°C and 100r / min for 2h, centrifuged, and then 9mL of artificial simulated intestinal juice was added to continue reacting at 37°C and 100r / min for 3h. After the reaction was completed, the probiotic cell count was performed. The results are shown in Table 1. The survival rate was calculated according to the following formula:

[0136] Survival rate (%) = N t / N0×100%In the formula, N t is the concentration of probiotics that survived after a certain period of incubation in vitro (cfu / g), and N0 is the original concentration of probiotics added (cfu / g).

[0137] The release rate was calculated according to the following formula:

[0138] Release rate (%) = (W t -W0) / W0×100%Wherein, Wt is the initial weight of the sample; W0 is the weight of the sample after a certain period of incubation in vitro.

[0139] Table 1

[0140]

[0141] It can be seen from the above table that the uric acid-lowering embedding materials prepared in Examples 1-3 of the present invention can maintain good integrity in artificial simulated gastric fluid and have good acid resistance. After being transferred to artificial simulated intestinal fluid, they release a large amount of active substances and have good pH responsiveness.

[0142] Test Example 2 DPPH free radical scavenging rate test

[0143] The extract 1 prepared in Examples 1-3 and Comparative Examples 1, 2, 6 and 7 was prepared into a 0.01 mg / mL sample solution.

[0144] Take 2.0mL of sample solution in a 10mL stoppered colorimetric tube, add 2.0mL of 0.0002mol / L DPPH solution, make up to volume, protect from light for 30min, and measure its absorbance A0 at a wavelength of 517nm. According to the above method, the absorbance value of the control group (the DPPH solution is replaced by an equal volume of ethanol solution) is A1, and the absorbance value of the blank group (2.0mL of each sample solution in the test group is replaced by 2.0mL of anhydrous ethanol) is A2. Prepare an equal concentration of VC solution as a control, and determine its clearance rate by the same method. The results are shown in Table 2. The calculation formula is as follows:

[0145] DPPH free radical scavenging rate (%) = 1-( A 0- A 1) A 2×100.

[0146] Table 2

[0147]

[0148] It can be seen from the above table that the extract 1 obtained by extracting in Examples 1-3 of the present invention has good antioxidant activity.

[0149] Test Example 3

[0150] SPF healthy male SD rats (200±20) g, 2 months old, were selected and after one week of adaptive feeding, they were randomly divided into blank control group, model group, positive drug group, Example 1-3 group, and Comparative Example 1-10 group, with 10 rats in each group. Except for the blank control group, the rats in the other groups were gavaged with potassium oxonate (750 mg / (kg·d)) daily and fed with yeast extract feed to establish a rat hyperuricemia model. At the same time, the blank control group and the model group were gavaged with normal saline, the positive drug group was gavaged with allopurinol (100 mg / (kg·d)), and the Example 1-3 and Comparative Example 1-10 groups were gavaged with Cordyceps militaris extract 1g / (kg·d) for 21 consecutive days. After the experiment, a sodium pentobarbital solution with a mass concentration of 1% was intraperitoneally injected, and the rats were anesthetized and blood was collected from the heart. The content of uric acid (UA) and serum creatinine (CRE) in the rat serum, the activity of xanthine oxidase (XOD) in the liver tissue, and the content of malondialdehyde (MDA), glutathione (GSH) and total antioxidant capacity (T-AOC) in the kidney tissue were determined according to the instructions of the kit. The results are shown in Table 3.

[0151] Table 3

[0152]

[0153] Note: * compared with the blank control group, P < 0.05; # compared with the model group, P < 0.05.

[0154] As can be seen from the above table, the uric acid-lowering composition containing Gardenia jasminoides prepared in Examples 1-3 of the present invention can significantly reduce the levels of serum uric acid and serum creatinine in rats, reduce xanthine oxidase in liver tissue, increase the glutathione (GSH) content and total antioxidant capacity (T-AOC) level in kidney tissue, and reduce the malondialdehyde (MDA) content.

[0155] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A method for preparing a uric acid-lowering composition containing Gardenia jasminoides, characterized in that: Mulberry leaves, chicory, gardenia and kudzu root are fermented with Lactobacillus rhamnosus and Bifidobacterium longum, and the probiotics and the fermentation liquid are separated. Immobilized ionic liquid is added to the fermentation liquid, carbon dioxide is introduced for adsorption, and the broth is filtered. The solid is added to ethanol, heated for desorption, and concentrated to obtain extract 1. The filtrate is ethanol precipitated to obtain extract 2, and the probiotics are embedded with sodium alginate to obtain a uric acid-lowering embedding material, which is uniformly mixed with extract 1, vitamin C, and tea polyphenols to obtain a uric acid-lowering composition containing gardenia. The preparation method of the uric acid-lowering composition containing gardenia comprises the following steps: S1. The mulberry leaves, chicory, gardenia and Pueraria root were washed, dried and crushed to obtain a mixed powder; the mass ratio of the mulberry leaves, chicory, gardenia and Pueraria root was 7-10:12-15:14-20:4-7; S2. activating Lactobacillus rhamnosus and Bifidobacterium longum to obtain a bacterial seed solution; S3. The mixed powder is added to water, sterilized, Lactobacillus rhamnosus and Bifidobacterium longum seed solution is added, fermented, filtered, washed with solids, the culture solution is collected, freeze-dried, and probiotics are obtained, and the filtrate is retained; S4. The immobilized ionic liquid was added to the filtrate, carbon dioxide was adsorbed, filtered, the filtrate was retained, the solid was added to ethanol, heated for desorption, filtered, the solid was washed, dried, and reused, and the filtrate was decompressed to remove the solvent to obtain an extract 1; the mass ratio of the filtrate to the immobilized ionic liquid was 10:2-3; The preparation method of the immobilized ionic liquid is as follows: T1. Adding silica gel to hydrochloric acid, heating to reflux, filtering, washing, and drying to obtain activated silica gel; T2. Add activated silica gel to water, add tannic acid and a catalyst, heat and stir to react, filter, wash, and dry to obtain modified silica gel; T3. Add the modified silica gel to water, add NHS and EDC, stir to activate, add 1-butyl-3-methylimidazole L-proline, stir to react, filter, wash, and dry to obtain an amino acid imidazole immobilized material; T4. The amino acid imidazole immobilized material was mixed with a salt and added to acetonitrile, the reaction was stirred, filtered, washed, and dried to obtain an immobilized ionic liquid; S5. The filtrate was precipitated by adding ethanol, washed, and dried to obtain extract 2; S6. The extract 2 and sodium alginate are dissolved in water, probiotics and lecithin are added, stirred and mixed, added dropwise to the fish oil, emulsified, calcium chloride solution is added dropwise, solidified at room temperature, filtered, washed, and dried to obtain a uric acid-lowering embedding material; the mass ratio of the extract 2, sodium alginate, probiotics and lecithin is 7-10:8-12:10-15:1-2; S7. The extract 1, the uric acid-lowering embedding material, vitamin C and tea polyphenols are mixed evenly to prepare a uric acid-lowering composition containing gardenia; the mass ratio of the extract 1, the uric acid-lowering embedding material, vitamin C and tea polyphenols is 10-12:15-20:2-3:1-3.

2. The preparation method according to claim 1, characterized in that: The activation conditions in step S2 are 28-33°C, 100-150r / min, anaerobic conditions, activation treatment for 18-24h, and the bacterial content of the bacterial seed liquid is 10 8 -10 9 cfu / mL.

3. The preparation method according to claim 1, characterized in that: The solid-to-liquid ratio of the mixed powder and water in step S3 is 1:7-10 g / mL, the inoculation amounts of the Lactobacillus rhamnosus and Bifidobacterium longum seed solutions are 1-3 v / v% and 2-4 v / v%, respectively, and the fermentation culture conditions are 35-37° C., 100-150 r / min, and anaerobic conditions for fermentation culture for 36-48 hours.

4. The preparation method according to claim 1, characterized in that: The concentration of hydrochloric acid in step T1 is 1-2 mol / L, and the heating reflux time is 3-4 hours; the mass ratio of activated silica gel, tannic acid and catalyst in step T2 is 10:3-5:0.2-0.3, the temperature of heating and stirring reaction is 40-50°C, the time is 2-4 hours, and the catalyst is Tris-HCl solution with pH=8.5-9.5; the mass ratio of modified silica gel, NHS, EDC and 1-butyl-3-methylimidazole L-proline in step T3 is 10:2-3:1-3:3-4, the stirring activation time is 20-30 minutes, and the stirring reaction time is 7-10 hours; the mass ratio of amino acid imidazole immobilization material and salt in step T4 is 1:2-3, the salt is NaBF4 or KPF6, and the stirring reaction time is 20-24 hours.

5. The preparation method according to claim 1, characterized in that: In step S4, the ventilation volume of carbon dioxide is 3-5 mL / min, the adsorption time is 30-50 min, and the heating desorption temperature is 40-50°C.

6. The preparation method according to claim 1, characterized in that: In step S5, ethanol is added until the ethanol content of the system is 70-80wt%, and precipitation is carried out for 1-2h.

7. The preparation method according to claim 1, characterized in that: The time for room temperature curing in step S6 is 20-30 minutes.

8. A uric acid-lowering composition containing Gardenia jasminoides obtained by the preparation method according to any one of claims 1 to 7.

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