CN115120604A - Application of apigenin 7-O-glucoside as preparation for reducing uric acid and gouty arthritis - Google Patents
Application of apigenin 7-O-glucoside as preparation for reducing uric acid and gouty arthritis Download PDFInfo
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Abstract
The invention relates to application of apigenin 7-O-glucoside as a preparation for reducing uric acid and gouty arthritis. Experiments prove that the apigenin 7-O glucoside can effectively reduce the serum uric acid level and has a certain blood pressure reducing effect; in addition, the anti-inflammatory and analgesic effects of the compound are further verified, and apigenin 7-O glucoside is expected to be developed for treating gouty arthritis.
Description
Technical Field
The invention belongs to the field of medicines, and particularly relates to application of apigenin 7-O-glucoside as a preparation for reducing uric acid and gouty arthritis.
Background
Apigenin 7-O-glucoside (apigenin-7-O-beta-D-glucopyranoside) belongs to plant flavonoid chemical components, and is a common component in many medicinal plants such as peony leaves, echinacea and ixeris sonchifolia. Apigenin-7-O-glucoside is also known as cosmosiin (Apigenin 7-O-glucoside, C) 21 H 20 O 10 (ii) a Molecular weight: 432.381, respectively; CAThe number S: 578-74-5). Apigenin 7-O-glucoside and its derivatives are known to be mainly used for anti-tumor, anti-inflammatory and anti-virus. However, no report is available at present about the preparation of apigenin 7-O-glucoside and derivatives thereof for treating hyperuricemia. The invention provides application of apigenin 7-O-glucoside as a uric acid reducing preparation.
Disclosure of Invention
The invention provides application of apigenin 7-O-glucoside in preparation of a medicine for reducing uric acid.
Another embodiment of the present invention provides the use of apigenin 7-O-glucoside in the prevention and/or treatment of hyperuricemia. The administration dosage of the apigenin 7-O-glucoside is selected from 0.1-1.6 mg/kg; preferably 0.8-1.6 mg/kg.
Another embodiment of the present invention provides the use of apigenin 7-O-glucoside in the prevention and/or treatment of gouty arthritis. The administration dosage of the apigenin 7-O-glucoside is selected from 0.1-1.6 mg/kg; preferably 0.8-1.6 mg/kg.
Another embodiment of the invention provides a uric acid lowering drug which is characterized in that the drug takes apigenin 7-O-glucoside as an active ingredient.
Another embodiment of the present invention provides a drug for preventing and/or treating hyperuricemia, characterized in that the drug comprises apigenin 7-O-glucoside as an active ingredient. The medicament optionally further comprises pharmaceutically acceptable adjuvants (such as carriers, diluents or excipients and the like). The dosage form of the medicament may be selected from solid, liquid or semi-solid formulations.
Another embodiment of the present invention provides a medicament for preventing and/or treating gouty arthritis, characterized in that the medicament comprises apigenin 7-O-glucoside as an active ingredient. The medicament optionally further comprises pharmaceutically acceptable adjuvants (such as carriers, diluents or excipients and the like). The dosage form of the drug can be selected from solid, liquid or semisolid formulations.
Compared with the prior art, the invention has the advantages that: (1) the invention discovers for the first time that the apigenin 7-O-glucoside can obviously reduce uric acid in serum and simultaneously shows a certain blood pressure reducing effect; (2) compared with other apigenin derivatives (such as CAS registration number: 1334524-22-9), the apigenin 7-O-glucoside has good solubility, remarkable uric acid reducing effect, anti-inflammatory effect and the like, and can be developed into medicaments for preventing and/or treating diseases such as hyperuricemia, gouty arthritis and the like.
Detailed Description
To facilitate a further understanding of the invention, the following examples are provided to illustrate it in more detail. However, these examples are only for better understanding of the present invention and are not intended to limit the scope or the principle of the present invention, and the embodiments of the present invention are not limited to the following. The method for establishing the disease animal model in each embodiment of the invention is a conventional general method in the field.
Example 1
1 materials
1.1 animals
SPF grade male Kunming mice, 28-32g, were produced by the laboratory animal research center at the university of air force military medical sciences [ certification number: SCXK- (shan) 2015-007 ]. Drinking water is normally taken, the room temperature is kept at 25 ℃, the relative humidity is 40%, the light/dark cycle is carried out for 12 hours, and the test is carried out after the cultivation is carried out for one week in an adaptive feeding mode. The treatment of the animals in the experimental process conforms to the guidance opinions about the animals which are good at being tested issued by the Ministry of science and technology.
1.2 Instrument
A RT-9600 type semi-automatic biochemical analyzer (Shenzhen Ledu Life sciences GmbH); sartorius electronic analytical balance (Sartorius balance, germany); biofuge Primo R multipurpose table high speed centrifuge (Heraeus, Germany); SpectraMax model 190 microplate reader (Molecular Devices, USA); an XK96-A type rapid mixer (Xinkang medical instruments Co., Ltd., Jiangyan); BP2010 type non-invasive blood pressure instrument (Beijing softong biotechnology limited)
1.3 reagent
Yeast powder (OXOID, UK); adenine (beijing solibao science and technology ltd); potassium Oxazinate (Shanghai Michelin Biochemical technology, Inc.); apigenin 7-O glucoside (Baojichen photobiology, Ltd.); uric Acid (UA) determination kit (Shanghai Rongsheng biological pharmaceutical Co., Ltd.)
2 method
2.1 establishment, grouping and administration of hyperuricemia model
Prevention effect experiment: 100 male Kunming mice were randomly divided into a blank group, a model group, an apigenin 7-O glucoside group (0.025, 0.5, 0.1, 0.2, 0.4, 0.8, 1.6mg/kg) and an allopurinol group (10mg/kg), and 10 mice were each group. The injection is administered by intragastric administration at a rate of 10mL/kg every day for 7 days continuously, 1 hour after the last administration, the injection is administered to the abdominal cavity to the model group and each administration group at a dose of 300mg/kg of oteracil potassium, the blank group is administered with normal saline at an equal dose, and blood is taken after 1 hour, and the level of hematuric acid is detected.
Therapeutic effect experiments: mixing yeast dry powder and adenine in the crushed granulated feed, fully and uniformly mixing, and re-pressing and forming to prepare the high-purine feed, wherein the content of the yeast dry powder in the feed is 10 percent, and the content of the adenine in the feed is 0.1 percent. After the mice were bred adaptively for 1 week, they were randomly divided into a blank group, a model group, an apigenin 7-O glucoside group (0.1, 0.2, 0.4, 0.8, 1.6mg/kg), an allopurinol group (10mg/kg) and an apigenin derivative group (10mg/kg), each of which was 10 mice. The high-purine feed is fed to the model group and each administration group, the common feed is fed to the normal group, the medicines with different doses are fed to each administration group, the physiological saline with the same amount is fed to the blank group and the model group, and the administration is performed by intragastric administration according to 10mL/kg every day for 28 days continuously. After the last administration for 1h, 300mg/kg of Potassium Oxazinate was intraperitoneally injected into each of the other groups except the blank group. The apigenin derivative group used in this example was the CAS accession number: 1334524-22-9 parts of apigenin derivatives.
2.2 specimen Collection and index Observation
After 1h of potassium oxonate administration, each group of mice was bled from the eyeballs, left to stand at room temperature for 2h, centrifuged at 3500r/min at 4 ℃ for 15min, and the supernatant serum was aspirated. The content of Uric Acid (UA) in mouse serum is respectively detected by adopting corresponding kit instructions.
2.3 statistical analysis
Analysis was performed using SPSS 23.0 statistical software, data in
The comparison among the groups adopts a one-factor analysis of variance (LSD) method, wherein P is less than 0.05, and P is less than 0.01, so that the difference has a significant statistical difference.
3 results
3.1 prevention of the effects of apigenin 7-O glucoside on hyperuricemia mice
As shown in Table 1, compared with the blank group, the serum uric acid of the model group is remarkably increased (P is less than 0.01); compared with the model group, the apigenin 7-O glucoside group (0.1, 0.2, 0.4, 0.8, 1.6mg/kg) and the allopurinol group (10mg/kg) can both obviously reduce serum uric acid (P is less than 0.05 and P is less than 0.01).
TABLE 1 preventive Effect of apigenin 7-O glucoside on hyperuricemia mice
Note: in comparison with the blank set, the results, ## p is less than 0.01; in comparison with the set of models, * P＜0.05， ** P＜0.01。
3.2 therapeutic Effect of apigenin 7-O glucoside on hyperuricemia mice
3.2.1 Effect on treating hyperuricemia
As can be seen in Table 2, the serum uric acid in the model group is significantly increased (P < 0.01) compared with that in the blank group; and model group
TABLE 2 serum uric acid lowering Effect of apigenin 7-O glucoside on hyperuricemia mice
Note: in comparison to the blank set, the results, ## p is less than 0.01; in comparison with the set of models, * P＜0.05， ** P＜0.01。
compared with the apigenin 7-O glucoside group (0.1, 0.2, 0.4, 0.8 and 1.6mg/kg) and the allopurinol group (10mg/kg), the traditional Chinese medicine composition can obviously reduce serum uric acid (P is less than 0.05 and less than 0.01), and the effective dose is 0.8 and 1.6 mg/kg.
3.2.2 Effect on the body weight of mice with hyperuricemia
Table 3 shows that there was no significant difference between the initial body weight and the body weight of the first week in each group of mice, the model group was significantly different from the blank group from the second week (P < 0.01), and the weight average of the model group was significantly increased in the third and fourth weeks (P < 0.01) compared to the blank group.
TABLE 3 weight changes in the groups of mice
Note: in comparison with the blank set, the results, ## P＜0.01。
3.2 Effect of apigenin 7-O glucoside on blood pressure in hyperuricemia mice
Compared with the blank group, the systolic pressure and the diastolic pressure of the model group are both obviously increased (P is less than 0.01); compared with the model group, the systolic pressure and diastolic pressure of each dose group of apigenin 7-O glucoside and the positive medicine group are reduced (P is less than 0.05, P is less than 0.01; Table 4).
TABLE 4 Effect of apigenin 7-O glucoside on the systolic and diastolic blood pressure in hyperuricemia mice
Note: in comparison with the blank set, the results, ## p is less than 0.01; in comparison with the set of models, * P＜0.05， ** P＜0.01。
example 2
1 Instrument and Material
1.1 instrument Sartorius electronic analytical balance (sidoris balance, germany); a micro-injection pump and a micro-injection needle (Shandongwei Gaoji medical polymer products, Inc.); RB-200 Intelligent hotplate Instrument (Chengdutai science and technology Co., Ltd.); surgical instruments (shanghai medical instruments ltd); 6mm punch.
1.2 reagent apigenin 7-O glucoside (batch No. 20191018, provided by Shanxi Paeonia ostii Zhengyuan Biotech Co., Ltd.); xylene (Fuyu Fine chemical Co., Tianjin); glacial acetic acid (sienna chemical reagent plant); chloral hydrate (Shanghai Shanpu chemical Co., Ltd.); dexamethasone tablets (Shanxi Tongda pharmaceutical Co., Ltd.); diclofenac sodium enteric sustained-release capsules (Hainanpril pharmaceutical Co., Ltd.); colchicine tablets (Guangzhou Pedi pharmaceutical Co., Ltd., Lot 20190921); sodium urate (Shanghai Yuanye Biotech Co., Ltd., batch No. 20190813), uric acid (UA; batch No. 20190811), xanthine oxidase (XOD; batch No. 20190814), IL-1 β (batch No. 20190410), and tumor necrosis factor α (TNF- α; batch No. 20190517) were purchased from Nanjing as Biotech Co., Ltd.
1.3 animals SPF-grade kunming mice [ certification number: SCXK- (shan) 2015-007], the bulk mass (20 +/-2) g; male Wistar rats [ certification No.: SCXK- (shan) 2015-0010] and the body mass (180 +/-16) g, which are provided by the research center for experimental animals of the university of military medical science of air force. The feed is fed for 5 days in an adaptive way, and the water is drunk in a normal diet, the room temperature is kept at 22-26 ℃, and the humidity is 50% -70%. The treatment of the animals in the experimental process conforms to the guidance opinions on the animals which are good for the experiment issued by the ministry of science and technology.
2 Experimental methods
2.1 Effect of apigenin 7-O glucoside on mouse ear swelling healthy male Kunming mice with body mass of 18-22 g were randomly divided into 10 groups of 20 mice each, namely a model group (distilled water) and an apigenin 7-O glucoside group (0.05mg kg. multidot.kg) -1 ) Apigenin 7-O glucoside group (0.5mg kg) -1 ) Apigenin 7-O glucoside group (5mg kg) -1 ) And positive control group (dexamethasone 5 mg-kg) -1 ) Each group was administered by gavage 1 time daily for 5 days. 1h after the last administration, the left auricle of the mouse is placed on both sides with a micropipette0.04ml of xylene was spread evenly and the right ear was used as a blank. After 30min, the neck is removed, the mouse is killed, two ears are cut off along the auricle line, round ear pieces (diameter is 6mm) are punched at the same positions of the two ears by a puncher, the mass of the round ear pieces is precisely weighed, and the swelling inhibition rate is calculated by taking the weight difference of the two ear pieces as the swelling degree.
Inhibition rate (average swelling degree of ear in model group-average swelling degree of ear in administration group) ÷ average swelling degree of ear in model group × 100%
2.2 influence of apigenin 7-O glucoside on granuloma of cotton ball of mouse the cotton ball required for experiment was prepared, 10mg of sterile cotton ball (required for each mouse) was taken, weighed precisely with an electronic balance, autoclaved, and placed in an oven for drying for use. Taking 200 Kunming mice 18-22 g, each group comprises 20 mice, each half of the mice is female and male, and the grouping dose is under the same 2.1. With 5% chloral hydrate (0.1ml 10 g) -1 ) Intraperitoneal injection is performed for anesthesia, hair is removed from the back, 95% ethanol is used for sterilization, the median skin of the back is cut under the aseptic condition, the incision is 0.5cm-1cm long, subcutaneous tissues are carefully expanded by using forceps, a 10mg sterilized cotton ball is implanted into the subcutaneous part of a mouse, and the incision is closed and sterilized by using a suture line. Gavage administration (grouping, reagent and dose same as ear swelling experiment) was started on day 2 after the operation, and each group was gavage administered 1 time daily for 7 days continuously. And (3) taking off the neck of the mouse 2 days after the last administration to kill the mouse, separating and taking out a cotton ball by operation, removing surrounding fat tissues, drying at 100 ℃ for 10 hours, weighing the mass, and subtracting the mass of the original cotton ball from the mass of the cotton ball to obtain the mass of the granulation tissue of the cotton ball.
Inhibition rate (average granulation tissue mass in model group-average granulation tissue mass in administration group) ÷ average granulation tissue mass in control group × 100%
2.3 Effect of apigenin 7-O glucoside on acetic acid induced writhing experiment of mice 200 Kunming mice 18-22 g were selected, each group had 20 mice each with half male and female, and the mice were divided into model group (distilled water) and apigenin 7-O glucoside group (0.05mg kg. kg) -1 ) Apigenin 7-O glucoside group (0.5mg kg) -1 ) Apigenin 7-O glucoside group (5mg kg) -1 ) And positive control group (diclofenac sodium 5mg kg) -1 ). Each group was administered by gavage 1 time daily for 5 days. After the last administration for 1h, 0.6% glacial acetic acid 0 is injected into the abdominal cavity of each group of mice2ml, the inhibition rate was calculated by observing and recording the number of writhing of mice within 15min after acetic acid injection (a series of actions including repeated contraction in lumbar muscle, abdominal depression, straightening of trunk hind leg and rising of buttocks were used as a positive reaction).
The inhibition rate is (average number of writhing in model group-average number of writhing in drug group) ÷ average number of writhing in model group × 100%
2.4 Effect of apigenin 7-O glucoside on mouse hotplate experiments 100 Kunming female mice with weight of 20 +/-2 g and group dose of 2.3 were taken, placed on a hotplate with temperature of 55 +/-0.5 ℃ by using an intelligent hotplate apparatus, and the time from the contact of the mouse with the hotplate apparatus chassis to the occurrence of foot licking was recorded as the pain threshold. Pre-measuring the basic pain threshold of each mouse, measuring for 2 times at intervals of at least 5 minutes (min), removing those with pleasure jump or insensitive pain sensation expression, and selecting mice with average basic pain threshold of 5-30s for formal experiment. Each group is administered by intragastric administration (grouping, reagent and dosage are same with writhing experiment) every day, and is continuously administered for 5 days, pain threshold values of 15min, 30min, 45min, 60min, 90min and 120min after the last administration are respectively measured, so that in order to avoid scalding hind feet of the mouse, if the hind feet of the mouse is not licked for more than 60s on a hot plate instrument, the hind feet of the mouse are immediately taken out, and differences among the groups are compared according to 60 seconds.
2.5 Effect of apigenin 7-O glucoside on gouty arthritis in rats 10 rats were selected as the normal group; the other 50 gouty arthritis models are prepared according to the literature method (Chinese animal research report, 2017, 25 (5): 494- -1 Sodium urate solution. The rats are fixed on an anatomical plate in a supine mode, 75% ethanol is used for disinfecting ankle joints of right hind limbs and crus, needles are inserted from the outer sides of the ankle joints, the inclined planes of needle openings face upwards and form an angle of 45 degrees with shin bones to penetrate into ankle joint cavities, 0.2mL of sodium urate solution is injected, and 0.2mL of 0.9% sodium chloride solution is injected into the same positions of the rats in a normal group. The model rats were randomly divided into models of 0.04 mg-kg -1 、0.40mg·kg -1 And 4.00 mg/kg -1 Apigenin 7-O glucoside and 1.0 mg/kg -1 Colchicine 4 groups of 10 per group. At the same time of molding, 6 groups of animals were administered the test product by gavage, wherein the normal group and the model control group were administered 0.9% sodium chloride solution continuously for 8 days. Measuring the volume of the right hind sole of each rat by using a toe volume measuring instrument 1, 8, 12, 24 and 48 hours after the model is made, and calculating the swelling rate of the sole; the swelling rate (%) of the foot sole is (volume of the molded hind foot-volume of the molded forefoot)/volume of the molded forefoot x 100%. 30min after the last administration, injecting 10% chloral hydrate into abdominal cavity of rat for anesthesia, fixing the dorsal position on a dissection board, taking abdominal main venous blood, separating serum, collecting joint fluid, storing in a-80 ℃ ultra-low temperature refrigerator, and detecting the contents of serum UA, XOD, IL-1 beta and TNF-alpha according to the operation of a related kit.
2.6 data statistics and analysis SPSS 23.0 statistical software was used for analysis, data analysis
The comparison among the groups adopts a one-factor analysis of variance (LSD) method, wherein P is less than 0.05, and P is less than 0.01, so that the difference has a significant statistical difference.
3 results
3.1 apigenin 7-O glucoside has significant anti-inflammatory effect, and it is observed that after the left ear of a mouse is smeared with dimethylbenzene, obvious red swelling appears immediately compared with the right ear. See table 5. As can be seen from the table 5, compared with the model group, the medium and high dose apigenin 7-O glucoside group has obvious inhibition effect (P is less than 0.01) on mouse ear swelling caused by xylene, the inhibition rate is stronger than that of a positive control group, and the result shows that the apigenin 7-O glucoside has good anti-inflammatory effect.
TABLE 5 Effect of apigenin 7-O glucoside on mouse ear swelling by para-xylene
Note: comparison with model groups: p < 0.05, P < 0.01. "-" indicates gavage with physiological saline.
As can be seen from Table 6, the mass of the cotton balls in the model group was increased as compared with that of the original cotton balls, indicating that the cotton balls implanted in the mice had granuloma and hyperplasia. Compared with a model group, the apigenin 7-O glucoside with medium and high doses has obvious inhibition effect on the formation of mouse cotton ball granulation (P is less than 0.05, and P is less than 0.01).
TABLE 6 Effect of apigenin 7-O glucoside on mouse Cotton boll granuloma
Note: comparison with model group: p < 0.05, P < 0.01. "-" indicates the gavage with physiological saline.
3.2 apigenin 7-O glucoside has significant analgesic effect as shown in Table 7, and positive reaction appears in each group of mice in the experiment within 15min after 0.6% glacial acetic acid is injected into the abdominal cavity. Compared with the model group, each administration group can effectively reduce the writhing frequency of the mice (P is less than 0.01). Wherein the positive control group and the medium and high dose apigenin 7-O glucoside group can obviously inhibit the pain reaction caused by glacial acetic acid stimulating mice, and the inhibition rate is over 50 percent.
TABLE 7 Effect of apigenin 7-O glucoside on acetic acid induced writhing in mice experiments
Note: comparison with model groups: p < 0.01. "-" indicates gavage with physiological saline.
As can be seen from Table 8, compared with the model group, the medium-high apigenin 7-O glucoside group can obviously prolong the pain tolerance time of the mice to the thermal stimulation 15min, 30min, 45min, 60min and 120min after administration, and the pain threshold value reaches the peak (P < 0.05 or P < 0.01) 120min after administration.
3.3 apigenin 7-O glucoside has obvious treatment effect on gouty arthritis, the rate of foot sole swelling of rats in the table 9 is known, and compared with a normal control group, the rate of foot sole swelling at each time point of the model control group is obviously increased (P is less than 0.01), which indicates that the molding is successful. Compared with a model control group, the swelling rate of 12-48h of the small-dose group, the medium-dose group and the large-dose group of the apigenin 7-O glucoside is obviously reduced (P is less than 0.05 or P is less than 0.01), and the swelling rate of 8-48h of the foot sole of the colchicine group is obviously reduced (P is less than 0.05 or P is less than 0.01) in a dose-effect relationship.
As can be seen in Table 10, the levels of UA, XOD, IL-1 beta and TNF-alpha are significantly increased (P < 0.01) in the model control group compared with the normal control group. Compared with a model control group, the contents of UA, XOD, IL-1 beta and TNF-alpha in medium and high dose apigenin 7-O glucoside and colchicine groups are obviously reduced (P is less than 0.05 or P is less than 0.01).
TABLE 10 Effect of apigenin 7-O glucoside on rat serum UA, XOD, IL-1 beta, TNF-a levels
Note: comparison with normal control group: # p is less than 0.01; comparison with model groups: p < 0.05.
Claims (10)
1. Application of apigenin 7-O-glucoside in preparing medicine for reducing uric acid is provided.
2. Application of apigenin 7-O-glucoside in preventing and/or treating hyperuricemia is provided.
3. Application of apigenin 7-O-glucoside in preventing and/or treating gouty arthritis is provided.
4. Use as claimed in any one of claims 2 to 3, characterized in that the administered dose of apigenin 7-O-glucoside is selected from 0.1-1.6 mg/kg.
5. The use as claimed in claim 4, characterized in that the administered dose of apigenin 7-O-glucoside is selected from 0.8-1.6 mg/kg.
6. A medicine for reducing uric acid is characterized in that the medicine takes apigenin 7-O-glucoside as an effective component.
7. A drug for preventing and/or treating hyperuricemia is characterized in that the drug takes apigenin 7-O-glucoside as an active ingredient.
8. A medicament for preventing and/or treating gouty arthritis is characterized in that the medicament takes apigenin 7-O-glucoside as an effective component.
9. Pharmaceutical according to any one of claims 6 to 8, characterized in that it optionally further comprises pharmaceutically acceptable adjuvants (such as carriers, diluents or excipients, etc.).
10. The pharmaceutical according to claim 9, wherein the pharmaceutical is in a dosage form selected from the group consisting of a solid formulation, a liquid formulation and a semi-solid formulation.
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