CN112641809B - Application of wood frog seed and its extract in preparing weight-reducing medicine and food - Google Patents
Application of wood frog seed and its extract in preparing weight-reducing medicine and food Download PDFInfo
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/56—Materials from animals other than mammals
- A61K35/65—Amphibians, e.g. toads, frogs, salamanders or newts
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
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- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P3/00—Drugs for disorders of the metabolism
- A61P3/04—Anorexiants; Antiobesity agents
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- A61P3/06—Antihyperlipidemics
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Abstract
The invention relates to application of wood frog seeds and extracts thereof in preparation of weight-reducing medicines and foods, belonging to the field of natural medicines and foods. A Chinese forest frog seed extract for reducing weight and strengthening body constitution and physical strength, and its preparation method 2 And (3) extracting. Has the advantages that the application of different extracts of the wood frog seeds in the weight-losing function is researched, and the effect of enhancing the physical strength and controlling the weight is achieved; the weight-losing and slimming food can achieve the purpose of controlling weight, improving various functions of organisms, accelerating metabolism, removing excessive lipid accumulation, relieving fatigue and weakness caused by weight losing and weight control, and helping various crowds to keep a healthy body state while controlling weight; the wood frog seeds have low price and obvious weight-losing effect, and are an ideal weight-losing preparation.
Description
Technical Field
The invention belongs to the field of natural medicines and foods, and particularly relates to application of a wood frog seed extract with a weight-losing effect in the development of weight-controlling and weight-losing foods, health-care foods, weight-losing medicines and compositions containing the wood frog seed extract.
Background
Obesity refers to abnormal or excessive fat accumulation which impairs health, and is a chronic metabolic disease which is caused by genetic and environmental factors and has certain influence on health. With the continuous improvement of living standard of people, the incidence rate of obesity is increasing day by day, obesity has become epidemic diseases spreading in the world, the obesity population of our country is increasing rapidly in recent years, and many researches show that the obesity patients have higher risks of other diseases, and the obesity and the induced diseases seriously threaten and influence the life safety and the life quality of human beings, and the prevention and the control of the weight are reluctant.
The cause of excessive weight gain has not been fully elucidated so far, but is mainly due to the body's inability to consume excessive energy resulting in lipid accumulation. The current methods for treating obesity mainly change eating habits, exercise therapy, surgical treatment and drug intervention. However, each method for losing weight and losing weight has limitations and disadvantages, for example, the body is in a sub-health state, and fatigue and weakness are easily caused due to the reduction of the immunity of the body along with the insufficient energy intake when diet control is carried out. Exercise therapy is difficult to maintain, rebound is very easy to continue when the exercise is stopped, and rebound is easy to occur after the exercise is stopped. The operation treatment has large damage to the organism and overlong recovery period. The medicine intervention is mainly based on western medicines such as sibutramine and metformin, has increased side effects and is not beneficial to long-term administration.
The wood frog (Rana chensinensis) is commonly called Tianji, oviductus ranae and the like, is an amphibian with extremely high economic value and is widely distributed in Changbai mountain of Jilin province in China. The application history of wood frogs in China is long, and the wood frogs are recorded in Song Dynasty's herbal graph classic as early as the Ming Dynasty, especially in the Ming and Qing dynasty, and are widely pursued. The wood frog is used as a famous and precious nourishing delicacy on a dining table and enjoys a good reputation among people. The wood frog seeds are the eggs of female wood frog, are the main by-products of wood frog oil, have large annual output, are rich in various nutrient substances such as protein, steroid hormones, trace elements, fatty acid and the like, and have various curative effects. The Rana chensinensis seed extract comprises water extract, alcohol extract and supercritical CO 2 The extract and the like are widely concerned about the development of wood frog seeds at present, can be used for the development of medicines, foods and health-care foods, and has the effects of regulating immunity, delaying senility, tonifying yang, improving eyesight, regulating blood fat and preventing climacteric syndrome of women.
Disclosure of Invention
The invention provides application of wood frog seeds and extracts thereof in preparing weight-losing medicines and foods, and aims to control weight and keep a healthy body state.
The technical scheme adopted by the invention is as follows:
application of Rana temporaria chensinensis David seed and its extract in preparing weight reducing medicine and food is provided.
The weight-reducing medicine and food of the invention are weight-reducing medicine and food for strengthening body and enhancing physical strength.
The wood frog seed extract comprises a wood frog seed aqueous extract, a wood frog seed alcohol extract and wood frog seed supercritical CO 2 And (3) extracting.
The invention relates to a wood frog seed supercritical CO 2 The preparation method of the extract comprises the following steps:
using supercritical CO 2 The parameters of the supercritical extraction equipment are set as follows: the pressure of the extraction kettle is set to 29MPa, the temperature of the extraction kettle is set to 50 ℃, the flow is set to 82L/h, the pressure of the separation kettle is set to 5MPa, the temperature of the separation kettle is set to 55 ℃, and after the extraction time is 132min, the rana chensinensis seed oil is collected from the outlet of the separation kettle.
The invention has the advantages that the application of different extracts of the wood frog seeds in the weight-losing function is researched, the body fatigue of the mouse can not be caused while the weight increase of the mouse is controlled, and the effect of enhancing the physical strength and controlling the weight is achieved; the weight-losing and slimming food can achieve the purpose of controlling weight, improving various functions of organisms, accelerating metabolism, removing excessive lipid accumulation, relieving fatigue and weakness caused by weight losing and weight control, and helping various crowds to keep a healthy body state while controlling weight; the wood frog seed extract is safe, the wood frog seeds are low in price, the weight losing effect is obvious, the weight can be effectively controlled, the wood frog seed extract is an ideal weight losing preparation, the wood frog seed extract can be used as food, health-care food or medicine for development, the product is suitable for weight losing of healthy people and weight control of sports personnel, is particularly suitable for people who are prone to fatigue and sub-healthy people to take, can enhance physical strength and relieve fatigue of people who engage in mental and physical labor for a long time, and has the function of assisting in blood fat reduction and health care.
Drawings
FIG. 1 is a section of epididymal adipose tissue from a blank control group of mice;
FIG. 2 is a slice of epididymis adipose tissue of a group of mice with supercritical extract of wood frog seeds;
FIG. 3 is a slice of epididymis adipose tissue of a high-dose group mouse extracted from Rana temporaria seed by supercritical extraction.
Detailed Description
The excessive weight increase of the mice is often accompanied by fat accumulation, and the examination of various physiological and biochemical indexes of the mice can help to internally evaluate the weight change of the mice. Provides a certain reference for the weight-losing mechanism of the wood frog seed extract. Examining various biochemical indicators of fatigue resistance in serum can help analyze whether the body health state of the mouse is influenced while the weight of the mouse is controlled.
Example 1
1. Preparation method of different extracts of experimental group wood frog seeds
1.1 supercritical CO of Rana temporaria chensinensis David seed 2 Preparation method of extract group
Using supercritical CO 2 The parameters of the supercritical extraction equipment are set as follows: the pressure of the extraction kettle is set to 29MPa, the temperature of the extraction kettle is set to 50 ℃, the flow is set to 82L/h, the pressure of the separation kettle is set to 5MPa, and the temperature of the separation kettle is set to 55 ℃. Collecting the Rana temporaria chensinensis seed oil from the outlet of the separation kettle after the extraction time is 132 min. Mixing with 1% Tween 80 and 0.5% CMC-Na before intragastric administration, and making into high and low concentrations of 3g/kg and 1g/kg (calculated according to crude drug amount), respectively, and intragastric administration according to body weight of mouse.
1.2 preparation method of alcohol extract group of wood frog seed
Heating and refluxing with 10 times of 95 ethanol twice in a thermostatic waterbath at 85 deg.C for 1.5 hr, filtering the extractive solution, and concentrating under reduced pressure. Mixing with 1% Tween 80 and 0.5% CMC-Na before intragastric administration, and making into high and low concentrations of 3g/kg and 1g/kg (calculated according to crude drug amount), respectively, and intragastric administration according to body weight of mouse.
1.3 preparation method of aqueous extract group of wood frog seeds
Extracting Rana temporaria chensinensis David seed with water, concentrating, making into high and low concentrations of 3g/kg and 1g/kg (calculated according to crude drug amount), and administering by intragastric administration according to mouse body weight.
2. Establishment of animal model
Five-week-old male Kunming mice (n 70) were purchased from Liaoning Biotechnology GmbH, Inc., with a weight of 20 + -2 g and a license number SCXK (Liao) 2015-0001, and were bred in animal cages with 12h alternating light and shade in the room. During the experiment, all mice were allowed free intake of rodent chow and distilled water. The mice were fed continuously for 28 days, weighed every three days, and the weight change curve of the mice with feeding time was recorded.
3. Experimental group setting
After 7 days of adaptive feeding, mice were randomized into 7 groups:
test group 1(10 mice): administering a low dose of feed and a supercritical extract of wood frog seeds;
test group 2(10 mice): administering high doses of feed and oviductus Ranae supercritical extract;
test group 3(10 mice): administering a low dose of the feed and the wood frog seed alcohol extract;
test group 4(10 mice): administering high doses of feed and wood frog seed alcohol extract;
test group 5(10 mice): administering a low dose of feed and wood frog seed aqueous extract;
test group 6(10 mice): administering a high dose group of feed and wood frog seed aqueous extract;
blank control group (10 mice): feed and corresponding volume of solvent were administered.
4. Collection of samples
4.1 serum sample Collection
30min after the last administration of the medicine at 28d, swimming without load for 40min, and after resting for 30min, collecting blood from orbit after ether anesthesia. Standing whole blood at room temperature for 60min, centrifuging at 3000rpm/min for 15min, separating serum, packaging, and storing at-80 deg.C.
4.2 tissue sample Collection
After the blood sample is collected, the mouse is killed by dislocation of cervical vertebra, then liver and epididymis fat are quickly dissected and taken out on ice bath, the liver and epididymis fat are rinsed in a small amount of ice-cold physiological saline, blood is removed, filter paper is wiped dry, weighing is carried out, and tissues are stored at-80 ℃ for later use.
5. Content determination of mouse biochemical index
Each group of serum samples was thawed at room temperature. Detecting the content of lactic acid (LD), urea nitrogen (BUN), Malondialdehyde (MDA), Triglyceride (TC), cholesterol (TG), free fatty acid (NEFA) and superoxide dismutase (SOD) activity in blood serum by spectrophotometry. The procedures were performed with strict reference to kit instructions.
Liver and brain were measured by weight (g): volume (ml) ═ 1: 9, adding 9 times of physiological saline, grinding by an internal cutting type tissue refiner to prepare 10% homogenate under the ice bath condition, centrifuging at 1500r/min for 15min, and taking supernatant to be tested. Detecting the content of hepatic glycogen and the activity of ATP enzyme in the liver homogenate and detecting the content of malondialdehyde and the activity of superoxide dismutase in the brain homogenate by adopting a spectrophotometry.
6. Examination of mouse adipose tissue
Mouse epididymal fat was fixed in 4% paraformaldehyde for 24H, then dehydrated, cleared, paraffin-embedded and sectioned, and stained with hematoxylin and eosin (H & E). Images were obtained by microscopy at 200 x magnification.
7. Data statistics
All data were analyzed using SPSS 25.00 software, and comparative analysis of differences between groups was performed by one-way analysis of variance and LSD-t test, and data were considered statistically significant differences when P.ltoreq.0.05, with results expressed as mean. + -. standard error (means. + -. SD).
8. The experimental results are as follows:
8.1 weight changes in mice
The body weight changes of the mice in each test group and the blank control group within 28 days of feeding are shown in Table 1.
Table 1 effect of each experimental group on mouse body weight (X ± SD, n ═ 10)
Note: * P<0.05， ** P<0.01, comparison with blank control group
From table 1 it can be derived:
the average body weight of each group of mice over 28 days was compared. On day 0, there was no significant difference in the initial mean body weight of each group of mice, and the body weight of each group of mice tended to increase as the experiment proceeded.
The mean body weight in the second week was lower in the test group 1 than in the blank control group with significant difference (P < 0.05). And the weight mean at week four was much lower than the blank control group with significant differences (P < 0.01).
The body weight of the test group 2 from the first week to the end of feeding at the fourth week was much lower than that of the blank control group, with a very significant difference (P < 0.01).
The weight gain of the test group 1 and the test group 2 during the feeding period of the mice is far lower than that of the blank control group, and both the groups have statistical significance. The results show that the high and low doses of the supercritical extract of the wood frog seeds have the effect of inhibiting the weight gain of mice, and the high dose group has better effect of controlling the weight of the mice. Can be used for controlling weight loss.
The test group 4 is a wood frog seed alcohol extract high-dose group, the weight gain is lower than that of a blank control group after the mice are fed for 28 days, and the statistical significance is achieved (P is less than 0.05). The results show that the composition also has the effect of controlling the weight gain of the mice, but the effect is slightly lower than that of the groups 1 and 2.
8.2 mouse organ index
The liver index and epididymal fat index of each group of mice are shown in table 2.
Table 2 organ index (X ± SD, n ═ 10) of mice in each experimental group
Note: * P<0.05， ** P<0.01, comparison with blank control group
The liver and epididymis fat of the dissected mouse are weighed (table 2), and the result shows that the epididymis fat index of the control group is highest, the epididymis fat index of the test group 1 and the epididymis fat index of the test group 2 are lower than that of the blank control group, and the significant difference (P <0.05) is achieved, so that the fat accumulation in the mouse body can be reduced to a certain extent by the high-low dose group of the supercritical extract of the wood frog seeds.
8.3 Biochemical index level of blood fat in mice
Serum biochemical indicators of each group of mice were determined and the results are shown in table 3.
TABLE 3 Effect of administration on the hypolipidemic index Components of mice (X + -SD, n ═ 10)
Note: * P<0.05， ** P<0.01, comparison with blank solvent group
As can be seen from Table 3:
the blank control group has the highest indexes TG and TC levels, and the TG levels of the test groups 1, 2, 3 and 4 are reduced to a certain degree, so that the statistical significance is achieved. The TG reduction of the test group 2 group compared with the blank control group is the largest, and the obvious difference is very obvious (P < 0.01). The blank control group has the highest index TC level, and the TC levels of the test group 1 and the test group 2 are reduced and have obvious difference (P < 0.05). The free fatty acid levels in the test groups 1, 2 and 4 were all decreased compared to the blank control group with significant differences (P < 0.05). By combining the results of Table 3, the high and low dose groups of the supercritical extract of wood frog seeds and the high dose group of the alcohol extract can obviously reduce the blood fat of mice, and the effect of the high dose group of the supercritical extract of wood frog seeds is superior to that of the other two groups.
8.4 Biochemical index level of fatigue of mice
After each group of mice are fed for 28 days for swimming experiments and blood is taken, the biochemical index data in the serum of each group of mice are shown in a table 4.
Table 4 mouse liver glycogen, blood lactic acid and serum urea nitrogen levels (X ± SD, n ═ 10)
Note: * P<0.05， ** P<0.01, comparison with blank solvent group
As can be seen from the liver glycogen index data in Table 4, the level of glycogen in the blank control group is the lowest, and the average level of glycogen in the groups 1 to 4 in the test group is higher than that in the blank control group, so that the statistical significance is achieved. The results show that the other test groups except the wood frog seed water extract groups 5 and 6 can achieve the effect of relieving the fatigue by increasing the glycogen storage of the body or reducing the glycogen consumption when possible. The index result of the blood lactic acid shows that the lactic acid level of the blank control group is the highest, and the lactic acid levels of the groups 1, 2 and 4 are all lower than that of the blank control group, wherein the lactic acid level of the group 2, namely the high-dose group of the supercritical extract of the wood frog seeds, is the lowest, and has very significant difference (P <0.01), which indicates that the 3 groups of the test groups can relieve the fatigue of the mice by reducing the accumulation of the lactic acid.
The results of the serum urea nitrogen show that the urea nitrogen levels of the 2 groups and the 4 groups of the test groups are reduced in different degrees compared with the blank control group, and have significant difference (P <0.05), which indicates that the 2 groups of the test groups have the efficacy of relieving physical fatigue.
8.5 mouse liver adipose histology evaluation
H & E stained sections of mouse epididymis adipose tissues are prepared, and the influence of a blank control group and high and low dose groups of the supercritical extract of the wood frog seeds on the mouse adipose tissues is observed.
As shown in figure 1, the arrangement of epididymal adipose tissue cells of the blank control group mice is irregular, the cells have larger volume and are obviously vacuolated, and occasionally, the fusion phenomenon is seen at the intersection of the cells. As shown in fig. 2 and 3, the slice of the high-low dose group of the supercritical extract from wood frog seeds shows that the epididymal adipose tissue cells are regularly arranged, uniform in size and clear in shape, and the high-dose group of the supercritical extract from wood frog seeds has the best shape, so that the shape of the adipose cells can be effectively improved, the accumulation of adipose tissue lipid can be inhibited, and the experimental data is consistent with the experimental data.
Therefore, combining the analysis of the above data results, the test groups 1 and 2, i.e. the groups with high and low doses of the supercritical extract of wood frog seeds, have significant effects on controlling the body weight of mice, possibly by regulating lipid metabolism in vivo to promote fat accumulation in vivo. And the weight of the mouse is controlled to increase, the body fatigue of the mouse is not caused, and the effect of enhancing the physical strength and controlling the weight is achieved.
Example 2 supercritical CO of Rana temporaria chensinensis David seeds of the invention 2 The extract preparation can be used in food, medicine and health food.
Preparation I, wood frog seed supercritical extract emulsion
The preparation process comprises the following steps: weighing Rana temporaria chensinensis David seed protein extract, dissolving in ultrapure water, and stirring overnight to obtain 5% Rana temporaria chensinensis David seed protein solution. Adding wood frog seed oil into protein solution according to a certain proportion, and homogenizing by a homogenizer to form a coarse emulsion. Then, a uniform fine emulsion is formed by using an ultrasonic crusher and ultrasonic waves.
Preparation II, Chinese forest frog seed supercritical extract microcapsule
And (3) using a spray dryer to set the inlet temperature and the outlet temperature, and spray-drying the fine emulsion prepared in the preparation I to obtain the wood frog seed microcapsule powder.
Claims (1)
1. An application of a wood frog seed extract in preparing weight-losing medicines and foods is characterized in that: the Rana temporaria chensinensis David seed extract is supercritical CO of Rana temporaria chensinensis David seed 2 Extract of Rana temporaria chensinensis David seed supercritical CO 2 The preparation method of the extract comprises the following steps:
using supercritical CO 2 The parameters of the supercritical extraction equipment are set as follows: the pressure of the extraction kettle is set to 29MPa, the temperature of the extraction kettle is set to 50 ℃, the flow is set to 82L/h, the pressure of the separation kettle is set to 5MPa, the temperature of the separation kettle is set to 55 ℃, and after the extraction time is 132min, the rana chensinensis seed oil is collected from the outlet of the separation kettle.
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| JPH06345656A (en) * | 1993-06-08 | 1994-12-20 | Toreede Uindo Kk | Antiobestic food and its preparation |
| CN102919853A (en) * | 2012-11-12 | 2013-02-13 | 刘书新 | Rana japonica oil soft capsule |
| CN102948663A (en) * | 2012-12-19 | 2013-03-06 | 刘书新 | Rana japonica oil granules and preparation method thereof |
| CN103027297A (en) * | 2013-01-08 | 2013-04-10 | 吉林大学 | Rana oil polypeptide granules and preparation method thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH06345656A (en) * | 1993-06-08 | 1994-12-20 | Toreede Uindo Kk | Antiobestic food and its preparation |
| CN102919853A (en) * | 2012-11-12 | 2013-02-13 | 刘书新 | Rana japonica oil soft capsule |
| CN102948663A (en) * | 2012-12-19 | 2013-03-06 | 刘书新 | Rana japonica oil granules and preparation method thereof |
| CN103027297A (en) * | 2013-01-08 | 2013-04-10 | 吉林大学 | Rana oil polypeptide granules and preparation method thereof |
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| 两种方法提取林蛙籽脂肪酸的气-质联用分析;李铁纯等;《食品科学》;20120925;第33卷(第18期);195-197 * |
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