AU2016313671A1

AU2016313671A1 - Health food and process for preparing the same

Info

Publication number: AU2016313671A1
Application number: AU2016313671A
Authority: AU
Inventors: Jiagang DENG; Zhengcai DU; Erwei HAO; Xiaotao HOU; Lizhen Huang; Zhongshang XIA; Yan Xie
Original assignee: Guangxi University of Chinese Medicine
Current assignee: Guangxi University of Chinese Medicine
Priority date: 2016-10-19
Filing date: 2016-12-08
Publication date: 2018-05-10
Also published as: CN106562415A; WO2018072276A1

Abstract

Abstract The invention discloses a health food and a process for preparing the same, wherein the active ingredients of the health food is composed of the following raw materials of Chinese traditional 5 medicine in parts by weight: 5 to 30 parts of Radix Puerariae (kudzuvine root); 2 to 20 portions of Gynostemma pentaphyllum; 0.5 to 8 parts of Panax notoginseng; I to 20 parts of Fructus Mori Albae (mulberry fruit); 1 to 10 parts of Flesh of Meretrix meretrix Linnaeus (clam meat); and 2 to 20 parts of Spirulina. The invention combines the traditional Chinese medicines of Radix Puerariae, Gynostemma pentaphyllum, Fructus Mori Albae and Panax notoginseng with Meretrix 10 meretrix Linnaeus and Spirulina to prepare a health food which can be helpful for lowering the blood glucose level and reducing the diabetic complications. The health food has an auxiliary hypoglycemic function and is safe without toxic side effects.

Description

TECHNICAL FIELD

The present invention relates to health food technology, in particular to a hypoglycemic health food and a process for preparing the same.

BACKGROUND

Diabetes mellitus (DM) is a non-infectious chronic endocrine metabolic disease with multiple complications. With the development of society and economy, people's dietary structure changes, so that the prevalence of diabetes increased year by year, seriously endangering human health. On December 1st, 2015, the Diabetes Atlas - the authoritative guide of the International Diabetes Federation (IDF) published new data showing that the current number of adult diabetes all the world has been more than 415 million, while China's adult diabetes came to 110 million and became the first diabetes' country in the world.

Diabetes can occur at any age, with the course of the disease, it is easy to occur complications such as systemic neurological, microvascular and microvascular diseases, and can result in the chronic progressive lesions of heart, brain, kidney, nerve, eye and other tissues and organs. Since many complications are developed and aggravated increasingly, the health and life of patients are seriously harmed. At present, the medications for treatment of diabetes mostly emphasis on stimulating islet to generate insulin, but the sensitivity of insulin target cells on insulin cannot be fundamentally improved, so that blood glucose levels often rebound again once stopping the medications for a period of time. At the same time, due to long-term drug stimulation on islet, the islet was subjected to a long-term stress and eventually the islet function fatigue was occurred, so that the diabetics eventually had to depend on exogenetic insulin to lower blood sugar. It will further result in the partial or complete loss of islet functions, and the repeated rebound of blood sugar levels causes the increasing blood viscosity and the microcirculation disturbance, so that it will be occurred of the complications of heart, brain, kidney, skin and nerve, etc. in the end.

SUMMARY OF THE INVENTION

In view of the above, it is an object of the present invention to provide a health food and a process for preparing the same to overcome the drawbacks of the prior art.

The active ingredients of the health food provided by the present invention are composed of the following raw materials of Chinese traditional medicine in parts by weight:

to 30 parts of Radix Puerariae (kudzuvine root);

to 20 portions of Gynostemma pentaphyllum;

0.5 to 8 parts of Panax notoginseng;

l to 20 parts of Fructus Mori Albae (mulberry fruit);

to 10 parts of Flesh of Meretrix meretrix Linnaeus (clam meat); and to 20 parts of Spirulina.

In some embodiments of the present invention, the active ingredients of the health food is composed of the following raw materials in parts by weight:

to 25 parts of Radix Puerariae;

to 15 portions of Gynostemma pentaphyllum;

to 5 portions of Panax notoginseng;

to 15 parts of Fructus Mori Albae;

to 8 parts of Flesh of Meretrix meretrix Linnaeus; and to 15 parts of Spirulina.

As a further embodiment of the present invention, the active ingredients of the health food provided by the invention is composed of the following raw materials in parts by weight:

to 30 portions of Radix Puerariae extract;

to 20 portions of Gynostemma pentaphyllum extract;

0.5 to 8 portions of Panax notoginseng extract;

to 20 parts of Fructus Mori Albae extract;

to 10 parts of Flesh of Meretrix meretrix Linnaeus extract;

to 20 portions of Spirulina extract.

to 25 parts of Radix Puerariae extract;

to 15 portions of Gynostemma pentaphyllum extract;

to 5 portions of Panax notoginseng extract;

to 15 portions of Fructus Mori Albae extract;

to 8 parts of Flesh of Meretrix meretrix Linnaeus extract;

to 15 parts of Spirulina extract.

In some embodiments of the present invention, the health food is used for preparing health food or medicine with hypoglycemic effect, and is safe without toxic side effects.

The present invention also provides a process for preparing the health food, comprising the following steps that:

the mixture of the Radix Puerariae and the Gynostemma pentaphyllum is extracted by refluxing alcohol, and then the extracting solution is concentrated, dried and crushed to obtain the extract I; the mixture of Fructus Mori Albae, the Meretrix meretrix Linnaeus and the Spirulina is extracted by boiling water and concentrated, and then the concentrated solution is added by ethanol and filtered to obtain the precipitate which is dried and crushed to obtain the extract II;

the Panax notoginseng is crushed into the Panax notoginseng powder as reserve; and the extract I, the extract II and the Panax notoginseng powder are mixed to obtain the health food.

In some embodiments of the present invention, the process for preparing the extract I comprises the steps that:

the mixture of the Radix Puerariae and the Gynostemma pentaphyllum is extracted for 1-4 times by refluxing 6 to 10 times amount of alcohol for 1 to 2 hours each time, and then the extracting solution is combined, concentrated, dried at 80~90°C, crushed and screened through 80-120 mesh sieve to obtain the extract I.

In some embodiments of the present invention, the process for preparing the extract I comprises the steps of:

the mixture of the Radix Puerariae and the Gynostemma pentaphyllum is soaked by 6-10 times amount of alcohol for 30-60 mins and extracted by reflux for 1-2 hours firstly, and then 6-8 times amount of alcohol is added to extract for 1-2 hours, and then the extracting solution is combined, concentrated, dried at 80~90°C, crushed and screened through 80-120 mesh sieve to obtain the extract I.

In some embodiments of the present invention, the process for preparing the extract II comprises the steps that:

the mixture of Fructus Mori Albae, the Meretrix meretrix Linnaeus and the Spirulina is extracted for 1-4 times by boiling 6 to 10 times amount of water for 1 to 2 hours each time, and then the extracting solution is combined and concentrated; and the concentrated solution is deposited by adding ethanol for 16-30 hours, and filtered to obtain the precipitate which is dried and crushed to obtain the extract II.

the mixture of Fructus Mori Albae, the Meretrix meretrix Linnaeus and the Spirulina is soaked by 6-10 times amount of water for 30-60 minutes and extracted by boiling for 1-2 hours firstly, and then 5-9 times amount of water is added to extract for 1-2 hours, and then the extracting solution is combined and concentrated; and the concentrated solution is added by 80~95vol% ethanol until the volume concentration of alcohol arriving 70-90% and then deposited for 16-30 hours, and filtered to obtain the precipitate which is dried at 60~75°C, crushed and screened through 80-120 mesh sieve to obtain the extract II.

In some embodiments of the invention, the extracting solution is concentrated to d=1.0—1.5 at 80°C.

From the above, it can be seen that the invention combines the traditional Chinese medicines of

Radix Puerariae, Gynostemma pentaphyllum, Fructus Mori Albae and Panax notoginseng with Meretrix meretrix Linnaeus and Spirulina to prepare a health food which can be helpful for lowering the blood sugar level and reducing the diabetic complications. The health food has an auxiliary hypoglycemic function and is safe without toxic side effects.

DESCRIPTION OF THE PREFERRED EMBODIMENTS

Hereinafter, the present invention will be further described in combination with the following examples in detail, so that the objects, technical solutions and advantages of the present invention will become more apparent.

Chinese medicine has a long history of diabetes prevention and treatment to accumulate rich experience and form a unique theoretical understanding. Diabetes is called as Xiaoke lesion in Chinese medicine, and there are many effective prescription which can significantly improve the symptoms of the diabetic patients and effectively reduce diabetic complications. After a longterm experimental research, in accordance with the theory of Differentiation syndrome and treatment of traditional Chinese medicine and in view of modern chemical compositions and biological activity researches, the inventors combined the traditional Chinese medicines of Radix Puerariae, Gynostemma pentaphyllum, Fructus Mori Albae and Panax notoginseng with Meretrix meretrix Linnaeus and Spirulina to prepare a health food which can be helpful for lowering the blood sugar level and reducing the diabetic complications. The health food has an auxiliary hypoglycemic function and is safe without toxic side effects to make up for the deficiencies of the prior arts.

Example 1: Preparation of health food capsules (1) 20 kg of Radix Puerariae and 12 kg of Gynostemma pentaphyllum are mixed and firstly added by 8 times amount of 75vol% edible alcohol to be soaked for 30 minutes and then extracted by reflux for 1 hour to collect the first extracting solution, and then the mixture is secondly added by 6 times amount of 75vol% edible alcohol to be extracted by reflux for 1 hour to collect the second extracting solution. After extracting twice, the first and second extracting solutions are combined, concentrated and dried at 85°C, and then crushed and screened through 100 mesh sieve to obtain the extract I.

(2) 3.5 kg of Meretrix meretrix Linnaeus is soaked by adding appropriate amount of water for 20 hours, and then mixed with 6 kg of Fructus Mori Albae and 15 kg of Spirulina. The mixture is soaked by adding 8 times amount of water for 30 minutes and extracted by boiling for 1 hour to collect the first exacting solution, and then the mixture is secondly added by 6 times amount of water to be extracted for 2 hours to collect the second extracting solution. After combining (filtering) the first and second extracting solutions, the combined extracting solution is concentrated as d~1.2 (80°C). The concentrated solution is added by 85vol% ethanol until the volume concentration of alcohol arriving 75% and then deposited for 24 hours, and filtered to obtain the precipitate (polysaccharide) which is dried in oven at 75°C, crushed and screened through 100 mesh sieve to obtain the extract II.

(3) 2 kg of Panax notoginseng is crushed into powder which is then screened through 100 mesh sieve, after moist heat sterilization, to obtain the Panax notoginseng powder.

(4) The extract I, extract II and Panax notoginseng powder are mixed for 30 minutes, and then mixed with appropriate excipients to fill into 0# capsules (0.5g/granule and 60 capsules/bottle), after sealing and labeling, to obtain the health food.

Example 2: Preparation of health food granules (1) 8 kg of Radix Puerariae and 10 kg of Gynostemma pentaphyllum are mixed and firstly added by 6 times amount of 78vol% edible alcohol to be soaked for 45 minutes and then extracted by reflux for 1.5 hours to collect the first extracting solution, and then the mixture is secondly added by 8 times amount of 70vol% edible alcohol to be extracted by reflux for 2 hours to collect the second extracting solution. After extracting twice, the first and second extracting solutions are combined, concentrated and dried at 88°C, and then crushed and screened through 100 mesh sieve to obtain the extract I.

(2) 8 kg of Meretrix meretrix Linnaeus is soaked by adding appropriate amount of water for 24 hours and then mixed with 11 kg of Fructus Mori Albae and 9 kg of Spirulina. The mixture is soaked by adding 7 times amount of water for 55 minutes and extracted by boiling for 1.2 hours to collect the first exacting solution, and then the mixture is secondly added by 9 times amount of water to be extracted by boiling for 1 hours to collect the second extracting solution. After combining (filtering) the first and second extracting solutions, the combined extracting solution is concentrated as d~l.3 (80°C). The concentrated solution is added by 90vol% ethanol until the volume concentration of alcohol arriving 86% and then deposited for 28 hours, and filtered to obtain the precipitate (polysaccharide) which is dried in oven at 65°C, crushed and screened through 120 mesh sieve to obtain the extract II.

(3) 3.5 kg of Panax notoginseng is crushed into powder which is then screened through 100 mesh sieve, after moist heat sterilization, to obtain the Panax notoginseng powder.

(4) The extract I, extract II and Panax notoginseng powder are mixed and then mixed with appropriate excipients to be prepared into granules by conventional preparation method, after packaging, to obtain the health food.

Example 3: Preparation of health food tablets (1) 5 kg of Radix Puerariae and 6 kg of Gynostemma pentaphyllum are mixed and firstly added by 6 times amount of 70vol% edible alcohol to be soaked for 40 minutes and then extracted by reflux for 1.2 hours to collect the first extracting solution, and then the mixture is secondly added by 8 times amount of 75vol% edible alcohol to be extracted by reflux for 2 hours to collect the second extracting solution, and then the mixture is thirdly added by 7 times amount of 75vol% edible alcohol to be extracted by reflux for 1 hour to collect the third extracting solution After extracting thrice, the first, second and third extracting solutions are combined, concentrated and dried at 88°C, and then crushed and screened through 80 mesh sieve to obtain the extract I.

(2) 1.5 kg of Meretrix meretrix Linnaeus is soaked by adding appropriate amount of water for 18 hours and then mixed with 15 kg of Fructus Mori Albae and 11 kg of Spirulina. The mixture is soaked by adding 6 times amount of water for 30 minutes and extracted by boiling for 1.3 hours to collect the first exacting solution, and then the mixture is secondly added by 5 times amount of water to be extracted by boiling for 1 hour to collect the second extracting solution. After combining (filtering) the first and second extracting solutions, the combined extracting solution is concentrated as d~l. 15 (80°C). The concentrated solution is added by 92vol% ethanol until the volume concentration of alcohol arriving 84% and then deposited for 20 hours, and filtered to obtain the precipitate (polysaccharide) which is dried in oven at 65°C, crushed and screened through 110 mesh sieve to obtain the extract II.

(3) 2.8 kg of Panax notoginseng is crushed into powder which is then screened through 120 mesh sieve, after moist heat sterilization, to obtain the Panax notoginseng powder.

(4) The extract I, extract II and Panax notoginseng powder are mixed and then mixed with appropriate excipients for 40 minutes, after tabletting and packaging, to obtain the health food.

Example 4: Preparation of a health food paste (1) 26 kg of Radix Puerariae and 9 kg of Gynostemma pentaphyllum are mixed and firstly added by 10 times amount of 70vol% edible alcohol to be soaked for 60 minutes and then extracted by reflux for 1 hour to collect the first extracting solution, and then the mixture is secondly added by 7 times amount of 70vol% edible alcohol to be extracted by reflux for 1 hour to collect the second extracting solution. After extracting twice, the first and second extracting solutions are combined, concentrated and dried at 80°C, and then crushed and screened through 80 mesh sieve to obtain the extract I.

(2) 7.5 kg of Meretrix meretrix Linnaeus is soaked by adding appropriate amount of water for 22 hours and then mixed with 13 kg of Fructus Mori Albae and 18 kg of Spirulina. The mixture is soaked by firstly adding 9 times amount of water for 38 minutes and extracted by boiling for 1.6 hours to collect the first exacting solution, and then the mixture is secondly added by 7 times amount of water to be extracted by boiling for 1.5 hours to collect the second extracting solution, and then the mixture is thirdly added by 8 times amount of water to be extracted by boiling for 1 hour to collect the third extracting solution. After combining (filtering) the first, second and third extracting solutions, the combined extracting solution is concentrated as d~1.25 (80°C). The concentrated solution is added by 80vol% ethanol until the volume concentration of alcohol arriving 72% and then deposited for 29 hours, and filtered to obtain the precipitate (polysaccharide) which is dried in oven at 68°C, crushed and screened through 100 mesh sieve to obtain the extract II.

(3) 4.5 kg of Panax notoginseng is crushed into powder which is then screened through 120 mesh sieve, after moist heat sterilization, to obtain the Panax notoginseng powder.

(4) The extract I, extract II and Panax notoginseng powder are mixed and then mixed with appropriate excipients to be prepared into paste by conventional preparation method, after packaging, to obtain the health food.

Example 5: Preparation of a health food oral solution (1) 14 kg of Radix Puerariae and 13 kg of Gynostemma pentaphyllum are mixed and firstly added by 7.5 times amount of 80vol% edible alcohol to be soaked for 44 minutes and then extracted by reflux for 1.8 hours to collect the first extracting solution, and then the mixture is secondly added by 6 times amount of 75vol% edible alcohol to be extracted by reflux for 2 hours to collect the second extracting solution. After extracting twice, the first and second extracting solutions are combined, concentrated and dried at 90°C, and then crushed and screened through 100 mesh sieve to obtain the extract I.

(2) 7 kg of Meretrix meretrix Linnaeus is soaked by adding appropriate amount of water for 17 hours and then mixed with 10 kg of Fructus Mori Albae and 5 kg of Spirulina. The mixture is soaked by adding 7 times amount of water for 55 minutes and extracted by boiling for 1.7 hours to collect the first exacting solution, and then the mixture is secondly added by 5.5 times amount of water to be extracted by boiling for 2 hours to collect the second extracting solution. After combining (filtering) the first and second extracting solutions, the combined extracting solution is concentrated as d~l.3 (80°C). The concentrated solution is added by 93vol% ethanol until the volume concentration of alcohol arriving 87% and then deposited for 20 hours, and filtered to obtain the precipitate (polysaccharide) which is dried in oven at 72°C, crushed and screened through 100 mesh sieve to obtain the extract II.

(4) The extract I, extract II and Panax notoginseng powder are mixed for 30 minutes and then mixed with appropriate excipients to be prepared into oral solution by conventional preparation method, after packaging, to obtain the health food.

Example 6: Preparation of health food pellets (1) 15 kg of Radix Puerariae and 13 kg of Gynostemma pentaphyllum are mixed and firstly added by 6.5 times amount of 76vol% edible alcohol to be soaked for 45 minutes and then extracted by reflux for 1.9 hours to collect the first extracting solution, and then the mixture is secondly added by 7.5 times amount of 72vol% edible alcohol to be extracted by reflux for 1.1 hours to collect the second extracting solution. After extracting twice, the first and second extracting solutions are combined, concentrated and dried at 82°C, and then crushed and screened through 100 mesh sieve to obtain the extract I.

(2) 9 kg of Meretrix meretrix Linnaeus is soaked by adding appropriate amount of water for 24 hours and then mixed with 3 kg of Fructus Mori Albae and 9 kg of Spirulina. The mixture is soaked by firstly adding 6 times amount of water for 58 minutes and extracted by boiling for 1 hour to collect the first exacting solution, and then the mixture is secondly added by 5.5 times amount of water to be extracted by boiling for 1 hour to collect the second extracting solution, and then the mixture is thirdly added by 5.5 times amount of water to be extracted by boiling for

1.5 hours to collect the third extracting solution, and then the mixture is fourthly added by 5 times amount of water to be extracted by boiling for 1.2 hours to collect the fourth extracting solution. After combining (filtering) the first, second, third and fourth extracting solutions, the combined extracting solution is concentrated as d~l.l (80°C). The concentrated solution is added by 78vol% ethanol until the volume concentration of alcohol arriving 70% and then deposited for 18 hours, and filtered to obtain the precipitate (polysaccharide) which is dried in oven at 73 °C, crushed and screened through 100 mesh sieve to obtain the extract II.

(3) 4 kg of Panax notoginseng is crushed into powder which is then screened through 100 mesh sieve, after moist heat sterilization, to obtain the Panax notoginseng powder.

(4) The extract I, extract II and Panax notoginseng powder are mixed and then mixed with appropriate excipients to be prepared into pellets by conventional preparation method, after packaging, to obtain the health food.

Animal experiments for testing the auxiliary hypoglycemic function of the health food

1. Materials and methods

1.1 Test samples:

The health food capsules of Example 1, specifications: 0.5g/capsule, product batch number: 150201, stored in dry place. Recommended oral dosage per person (adult): 2 times a day, 3 capsules each time, and such dosage is calculated by adult weight of 60kg, equivalent to a dosage of 50mg/kg BW. The capsule contents are taken for test.

1.2 Test animals:

144 healthy adult male Kunming mice, SPF level, were selected, which were bred by Guangdong Laboratory Animal Center, each weighing 24 to 28 grams, Laboratory Animal Production License: No. SCXK (Gui) 2013-0002, Laboratory Animal Quality Certificate: No. 44007200027357.

1.3 Laboratory animal room environmental conditions:

Barrier system, license number: SYXK (Gui) 2011-0005.

Animal laboratory temperature: 22 ~ 25°C, relative humidity: 55-70%

1.4 Dosage and administration of the test samples:

In accordance to the recommended dosage of human body, three test groups of 250, 500, lOOOmg/kgBW (equivalent to 5, 10, 20 times of the recommended dosage, respectively), a model control group, a normal animal test sample group (high dose) and a normal animal blank control group are set with 12 mice for each group.

1.25g, 2.50 and 5.00g of the health food capsules of Example 1 were weighted and added with water and mixed into lOOmL to prepare 12.5, 25.0 and 50.0 mg/mL of suspensions, respectively. Each suspension was administrated to corresponding test group by intragastric administration, 50.0 mg/mL suspension was administrated to the normal animal test sample group with 0.2mL/10gBW of perfusion volume, and equal volume pure water was administrated to the model control group and the normal animal blank control group. All mice were administrated once per day and lasting for 30 days by intragastric administration.

1.5 Major Instruments and reagents

Instrument: ACCU-CHEK performa blood glucose meter, electronic analytical balance, etc. Reagents: blood glucose test paper, etc.

1.6 Experimental methods:

According to the State Food and Drug Supervision and Protection [2012] 107, Notification for Issuance of 9 Health Function Evaluation Methods such as Antioxidant Function Evaluation Method, Annex 3 - Auxiliary Hypoglycemic Function Evaluation Method, released by the State Food and Drug Administration, the animal test program I - islet injury model of hyperglycemia.

1.6.1 Modeling of hyperglycemia model animals

Mice were administrated alloxan (120mg/kgBW) by intraperitoneal injection after fasting for 24 hours to establish a model. After 5 days, the mice was fasted for 4 hours, and then their eye canthus blood was taken to test the blood glucose, if the blood glucose value was fallen into 10-25 mmol/L, it can be considered as the successful modeling of hyperglycemia model. 120 mice were selected for modeling, and 102 of them were successful. 96 mice of the successful modeling were divided into two batches to test. The first batch of 48 mice were subjected to fasting blood glucose test and the second batch of 48 mice were subjected to glucose tolerance test.

1.6.2 The fasting blood glucose test

The first batch of hyperglycemia model animals were randomly divided into 4 groups according to the 4 hours fasting plasma glucose level (n = 12). The test groups were given different concentrations of sample solutions and the model control group was given pure water for 30 days. The fasting blood glucose levels (fasting as pre-test) were measured and compared the fasting blood glucose levels and the blood glucose decreasing ratio.

The decreasing ration of blood glucose = (pre-test blood glucose level - post-test blood glucose level) / pre-test blood glucose level x 100%

Under the modeling conditions, if the decline or the decreasing ration of the fasting blood glucose level of the test group relative with these of the model control group are statistically significant, then it can determined that the test results are positive.

1.6.3 Normal animals:

healthy adult mice were selected and randomly divided into the blank control group and the test sample group (high dose) with each group of 12 mice according to the 4 hours fasting blood glucose levels thereof. The other operations were the same as 1.6.2.

1.6.4 Glucose tolerance test:

The second batch of 48 hyperglycemia model mice were randomly divided into 4 groups according to the 4 hours fasting plasma glucose level (n = 12). The test groups were given different concentrations of sample solutions and the model control group was given pure water for 30 days just like 1.6.2. On the 31st day, after fasting for 4 hours, the test groups were given different concentrations of sample solutions, and the model control group was given equal volume of pure water. 15 minutes later, each group was orally administered with glucose solution (2.0g/kgBW). The blood glucose levels were measured at 0, 0.5 and 2 hours after administration of glucose. The change of area under blood glucose curve at each time point after glucose administration was observed for the model control group and the test groups.

Area under blood glucose curve = 0.25 x (blood glucose level at 0 hr + 4 x blood glucose level at 0.5 hr + 3 x blood glucose level at 2 hr)

Under the modeling conditions, if the decline of the area under blood glucose curve at 1, 0.5, 2 hr of the test group after administration of glucose relative with these of the model control group are statistically significant, than it can determined that the test results are positive.

io

1.7 Test data processing:

SPSS 11.0 statistical software was used for statistical analysis of variance analysis.

1.8 Results determination:

Under the modeling conditions, if any indicator of both indicators of the fasting blood glucose level and the glucose tolerance is positive and it is no effect on the normal animal fasting blood glucose level, then it can be determined that the auxiliary hypoglycemic animal test results of the test samples are positive.

2. Test results

2.1 The effect of the test samples on the mice body weights

Compared between the test groups and the model control group, there were no significant differences in initial body weight, mid-term body weight, final body weight and weight gain of the mice (P> 0.05), see Table 1. Compared between the normal animal test group and the normal animal blank control group, there were no significant difference in initial body weight, mid-term body weight, final body weight and weight gain (P> 0.05), see Table 2. The results showed that the test sample had no significant effect on the weight gain of the hyperglycemia model mice and the normal mice.

Table 1. The effect of the test sample on the hyperglycemia model mice (7 ± 5)

Test Groups I

Dosage (mg/kg BW)	Animal amount	Initial weight (g)	Mid-term weight (g)	Final weight (g)	Weight gain(g)
1000	12	28.3+1.5	33.8+2.2	37.0+2.3	8.7+1.5
500	12	29.0+1.5	33.8+1.9	37.7+1.9	8.7+1.4
250	12	28.2+1.1	32.9+1.7	36.9+1.8	8.7+1.4
Model control	12	28.4+1.3	33.0+1.4	36.7+2.0	8.4+1.4

Ps. Compared between the test groups and the model control group, there were no statistically significant differences in initial body weight, mid-term body weight, final body weight and weight gain of the mice (P> 0.05) _o

Test groups II

Dosage (mg/kg BW)	Animal amount	Initial weight (g)	Mid-term weight(g)	Final weight (g)	Weight gain (g)
1000	12	28.2+1.7	34.3+2.3	36.5+2.6	8.3+1.2
500	12	28.0+1.7	33.2+2.3	36.2+2.5	8.2+1.8
250	12	28.4+1.0	32.9+1.6	35.9+1.5	7.5+1.1
Model control	12	28.8+1.4	32.9+2.7	36.2+3.0	7.4+2.3

Table 2, The effect of the test sample on the normal mice (^x ± s)

Dosage (mg/kg BW)	Animal amount	Initial weight (g)	Mid-term weight(g)	Final weight (g)	Weight gain (g)
1000	12	26.3+0.6	31.6+1.3	39.7+2.4	13.4+2.5
Blank control	12	26.5+1.1	32.9+1.8	41.5+2.7	15.0+2.5

Ps. Compared between the normal animal test group and the normal animal blank control group, there were no statistically significant difference in initial body weight, mid-term body weight, final body weight and weight gain (P> 0.05).

2.2 The effect of the sample on fasting blood glucose in hyperglycemia model mice

Referring to Table 3, the model is established due to 4 hour fasting blood glucose > lOmmol/L. After 30 days of oral administration of different doses of the capsules of Example 1, the blood glucose level of the 30th day of each test group was lower than that of the model control group. The decline and the decreasing ratio of blood glucose level of each test group were higher than the model control group. There was significant difference (P <0.01 or P <0.05) between the blood glucose level, decline of blood glucose level and decreasing rate of blood glucose level in the high and middle dose test group and the model control group, which indicating that the test sample has the effect of lowering fasting blood glucose in hyperglycemia model mice.

_Table3, Results of the fasting blood glucose levels of hyperglycemia model mice (^x ±s)

Dosage (mg/kg BW)	Animal amount	Blood glucose level after modeling (mmol/L)	Blood glucose level on 30th day (mmol/L)	Decline of blood glucose level (mmol/L)	Decreasing ratio of blood glucose level (%)
1000	12	17.03+1.96	12.34+1.65**	4.69+0.86**	27.59+4.39**
500	12	17.19+1.71	12.80+1.53*	4.39+0.57**	25.63+3.01**
250	12	17.28+1.61	14.37+1.66	2.91+0.87	16.87+5.22
Model control	12	17.11+2.21	14.62+1.87	2.49+0.59	14.53+2.41
PS. * indicates	the difference	with the model	control group,	P<0.05 and	** indicates the

difference with the model control group, P<0.01.

2.3 The effect of the test sample on glucose tolerance in hyperglycemia model mice

Table4, Results of the glucose tolerance of hyperglycemia model mice (^x + s)

Dosage (mg/kg BW)	Animal amount	Blood glucose level after modeling (mmol/L)	Blood glucose level after gastric perfusion of glucose (mmol/L)	Area under blood glucose curve
Oh 0.5h	2h
1000	12	17.31±1.74	12.35+1.51 18.63+2.57	16.11+2.35	33.80+4.60**
500	12	17.36+1.90	12.68±1.67 18.99+2.57 12	16.96+2.43	34.88+4.64**

250 12

Model control 12

16.95±2.04 14.26±1.98 21.40±2.91 18.88±2.90 39.13±5.38 16.97±1.84 15.20±1.90 22.86±2.94 19.40±2.62 41.21±5.25

PS. ** indicates the difference with the model control group, P<0.01.

It can be seen from Table 4 that after oral administration of different doses of the capsules of Example 1 for 30 days, the area under blood glucose curve in each test group was lower than that in the model control group, and the differences between the high and middle dose test groups and the model control group were significant (P<0.01), which indicating that the test sample has the effect of reducing the area under blood glucose curve at each time point in the glucose tolerance test of hyperglycemia mice, and the glucose tolerance test results are positive.

2.4 The effect of the test sample on the fasting blood glucose level of normal mice

As shown in Table 5, after oral administration of 1000 mg/kg BW of the health food capsule (Example 1) for 30 days, there was no significant difference (P>0.05) in the fasting blood glucose level between the normal animal test group and the normal animal blank control group (P>0.05), which indicating that the test sample has no significant effect on the fasting blood glucose level in normal mice.

Table 5. The results of fasting blood glucose level in the normal mice (χ ± .sj

Dosage (mg/kg BW)	Animal amount	Blood glucose level before test (mmol/L)	Blood glucose level on 30th day (mmol/L)
1000	12	5.30 ±0.26	5.33 ±0.29
Blank control	12	5.61 ±0.37	5.43 ± 0.24

PS. There are no statistically significant difference between the blood glucose levels before and after test of the normal animal test group and the blood glucose level of the blank control group (P>0.05) .

3. Conclusion:

After gastric perfusion of the test capsules in 250, 500, 1000 mg/kg BW (equivalent to 5, 10, and 20 times of recommended dose of human body) for 30 days in the hyperglycemia model mice, the fasting blood glucose and the area under blood glucose curve at each time point of the glucose tolerance test can be decreased. The glucose tolerance test results were positive and the weight gain of the mice was not significantly affected. After gastric perfusion of the test capsules in 1000 mg/kg BW dose for 30 days in the normal mice, it has no significant effect on the fasting blood glucose level and the weight gain of the normal mice. As a result, the test sample has an auxiliary hypoglycemic function.

Studies on the acute and chronic toxicity

1. Materials and methods

1.1 Test samples:

The health food capsules of Example 1, specifications: 0.5g/capsule, product batch number: 150201, stored in dry place. Recommended oral dosage per person (adult): 2 times a day, 3 capsules each time, and such dosage is calculated by adult weight of 60kg, equivalent to a dosage of 50mg/kg BW.

1.2 Test animals and environmental conditions:

SPF level healthy adult male Kunming mice and SD rats were bred by Guangdong Laboratory Animal Center, Laboratory Animal Production License: No. SCXK (Gui) 2014-0002, Laboratory Animal Quality Certificate: No. 45000300000170, 45000300000223, 45000300000379. The laboratory animal room is a barrier system, license number: SYXK (Gui) 2011-0005. Animal laboratory temperature: 22 ~ 25°C, relative humidity: 55-70%.

1.3 Test on acute oral toxicity:

The maximum tolerated dose (MTD) test is used by selecting 20 Kunming mice of the body weight of 18 - 22g, half male and half female. The mice were fasted for 16 hours before the test without limit to drink water. 20.Og of the test samples were weighed, and then added with pure water to 40mL, mixed to prepare 500mg/mL concentration of suspension, and then fed to the mice by gastric perfusion twice (interval of 6h) with the dose of 0.4mL/20gBW, and the total dose was 20000 mg/kg BW. After gastric perfusion, the poisoning manifestations of the mice were observed and recorded. The mice were weighed once a week and observed for two weeks. At the end of the test, all the mice were generally observed by anatomy. The acute toxicity of the test sample was evaluated according to the toxicity grading standard.

1.4 Thirty days feeding test in rats

1.4.1 Dosage and administration of the test samples:

SD rats in each weight of 60-80g were selected, half male and half female. The rats were randomly divided into four groups, namely negative control group and three test groups, each group of 20, half female and half male. The doses of the three test groups were set at 5000, 2500 and 1250 mg/kg BW, which were equivalent to 100, 50 and 25 times of the human recommended dose, respectively. 50.0, 25.0, 12.5 g of the test sample were weighed and added with pure water to lOOmL, and mixed to prepare 500, 250, 125 mg/mL of suspensions, respectively. Each suspension in volume of l.OmL/lOOg BW was administrated to the corresponding test group by intragastric administration, and equal volume of pure water was administrated to the negative control group. All rats were administrated once per day and lasting for 30 days by intragastric administration.

1.4.2 Test Method:

All animals during the test were given with general feed separately by voluntary feeding and water drinking. The activity and growth of the animals were observed daily, and the animals were additionally fed twice a week and weighed once a week to record the food intake per week and food utilization rate. At the end of the test, the animals were fasted overnight (for 16 h, not limited to drinking water) and weighted to obtain the fasted weight. After the animals were sacrificed, two blood samples were taken in which one blood sample was added with anticoagulant and used to detect Hb, RBC, WBC, PLT, etc., and the other was not added with anticoagulant to isolated serum and detect AST, ALT, BUN, Cr, TC, TG, Glu, TP, Alb and other items by ELISA kit and automatic biochemical analyzer. After blood sampling, the animals were dissected to collect and weight the liver, kidney, spleen, stomach, duodenum, testis, ovary and other organs and calculate the ratio of viscera/body. The liver, kidney, spleen and testis were examined by histopathological method. If the general examination of each test group did not show significant changes in lesions and biochemical indicators, then only the main organs of the high-dose test group and the control group were examined by histopathological method; if the significant changes in lesions and biochemical indicators were found, then the corresponding organs of the middle and low dose groups were examined too.

1.4.3 Statistical analysis for the test data:

The SPSS statistical software was used for one-way analysis of variance. In the statistical analysis, the data were performed by homogeneity test of variance, if there was homogeneous variance, an overall comparison would be performed by using one-way analysis of variance. If there was difference, then a pairwise comparison between the test groups and the control group would be performed by Dunnett test. If there was inhomogeneous variance, then the data would be converted to the appropriate variables to meet the variance homogeneity test, and then the converted data would be used for statistical analysis. If the converted data had not yet reached the variance homogeneity requirement, then the statistical would be performed by using the rank sum test.

2. Results

2.1 Acute oral toxicity test

Table 6 the acute toxicity test results of the test sample on mice ( ^x ± s)

Gender	Number of animals	Administration	Dose (mg/kg BW)	Initial weight	Weight on 7th day	Weight on 14th day	Number of animals with toxic symptom	Number of dead animals
Male	10	oral	20000	20.6±l.l	27.1±1.5	32.7±1.8	0	0
Female	10	oral	20000	19.7±1.2	25.6±1.6	30.3±2.0	0	0

As shown in Table 6, after gastric perfusion of 20000 mg/kg BW of the test sample, the animals grew well and there is no effect on the body weight of animals. All tested mice did not show any toxic symptom and no animals death were observed for 14 days. At the end of the test, the animals were dissected and the liver, kidney, spleen, heart, lung, stomach, intestine and other major organs were no significant abnormal changes in general observation. The results showed that the acute oral toxicity MTD of mice was more than 20000 mg/kg BW and the acute oral toxicity was non-toxic.

2.2 Thirty days feeding test in rats

2.2.1 Animal general performance:

During the test, the animals in each group grown and developed well, and the animal abnormal behavior and poisoning performance were not observed. There was no death in each group of animals.

2.2.2 The effect of the test sample on body weight and food utilization rate in rats

The results are shown in Table 7 and 8. The rats were orally administered with the test sample at the doses of 5000, 2500 and 1250 mg/kg BW for 30 days. During the test, there was no statistically significant difference in the body weight every week, weight gain, food intake every week and overall food utilization rate in male and female rats between the test groups and the control group (P> 0.05), which indicated that the test sample has no significant effect on the weight gain and food utilization rate in rats.

Table 7. The effects of the test sample on weight of rats (χ ± .s)

Gender	Group	Number of animals	Initial weight (g)	First week (g)	Second week (g)	Third week (g)	Final weight (g)
	High-dose	10	73.2+5.4	139.9+6.6	201.7+7.4	261.4+12.0	326.6+18.7
Male	Mid-dose	10	73.6+4.0	140.9+4.6	202.5+5.4	262.7+10.9	328.1+17.8
	Low-dose	10	73.5+4.4	140.4+3.7	203.1+5.7	261.7+10.5	329.8+15.6
	Control	10	74.3+5.1	140.7+5.6	201.3+6.9	261.5+11.0	325.9+15.7
	High-dose	10	71.7+4.9	119.0+9.7	159.1+13.4	190.2+16.7	219.3+15.9
Female	Mid-dose	10	71.3+5.1	120.2+8.8	162.0+11.3	193.5+15.0	221.7+17.8
	Low-dose	10	71.9+4.3	119.7+4.7	159.9+8.4	192.1+7.0	223.0+11.5
	Control	10	72.3+3.8	119.7+9.3	161.1+13.4	192.4+12.2	220.9+9.8

PS. There were no statistically significant differences between the test groups and the negative 20 control group in Table 7 (P>0.05).

Table 8. The effects of the test sample on the overall food utilization rate in rats (χ ± .s)

Gender	Group	Number of animals	Overall weight gain (g)	Overall food intake (g)	Overall food utilization rate (%)
	High-dose	10	253.4+16.9	783.0+31.3	32.4+1.9
Male	Mid-dose	10	254.5+18.8	783.1+20.5	32.5+2.7
	Low-dose	10	256.3+15.6	778.1+19.3	32.9+2.0
	Control	10	251.6+14.7	770.0+31.3	32.7+2.0
	High-dose	10	147.6+12.9	530.7+41.7	27.8+1.4

Female	Mid-dose Low-dose Control	10 10 10	150.4+15.6 151.1+10.6 148.6+7.8	542.0+35.4 544.3+17.8 537.5+34.4	27.8+2.2 27.7+1.4 27.7+1.6
PS. There	were no	statistically significant differences between the test groups and the negative
control group in Table 7 (P>0.05).
2.5.3 The effects of the test sample on blood routine index in rats
Table 9	The results of blood routine indexes in rats at the end of thirty days feeding test of the
			test sample (^χ ± s)
Gender	Group	Number of	Hemoglobin	Erythrocyte	Platelet
		animals	(g/L)	(1012/L)	(109/L)
	High-dose	10	146.8+9.0	7.48+0.47	982.8+182.9
Male	Mid-dose	10	147.2+8.7	7.46+0.46	948.4+160.5
	Low-dose	10	148.0+5.0	7.60+0.31	990.1+125.3
	Control	10	149.6+9.0	7.66+0.51	958.2+161.0
	High-dose	10	145.6+6.7	7.52+0.41	988.8+163.8
Female	Mid-dose	10	148.4+8.9	7.63+0.45	963.2+167.4
	Low-dose	10	147.9+6.5	7.58+0.38	961.4+120.7
	Control	10	148.6+6.1	7.66+0.36	976.6+128.7

PS. There were no statistically significant differences between the test groups and the negative control group in Table 7 (P>0.05).

Table 10. The results of blood routine indexes in rats at the end of thirty days feeding test of the test sample (^χ ± s)

Gender	Group	Number of animals	Leukocyte (109/L)	Leukocyte classification (%)
Lymphocyte	Neutrophil	Monocyte	Eosinocyte	Basophil
	High-dose	10	7.50+0.90	83.9+1.3	10.4+1.2	4.66+.20	0.94+0.13	0.13+0.13
Male	Mid-dose	10	7.40+0.81	83.8+1.4	10.5+1.2	4.67+0.49	0.92+0.11	0.12+0.15
	Low-dose	10	7.62+0.69	83.6+1.5	10.6+1.2	4.74+0.49	0.94+0.20	0.14+0.11
	Control	10	7.40+0.70	83.7+1.9	10.6+1.9	4.60+0.20	0.97+0.28	0.13+0.14
	High-dose	10	7.58+0.76	83.7+1.9	10.5+1.8	4.69+0.50	0.97+0.20	0.13+0.13
Female	Mid-dose	10	7.44+0.67	84.0+1.3	10.3+1.1	4.62+0.50	0.94+0.22	0.13+0.16
	Low-dose	10	7.66+1.06	83.8+1.2	10.6+1.0	4.53+0.61	0.91+0.17	0.15+0.14
	Control	10	7.44+0.93	83.6+1.3	10.7+1.6	4.73+0.75	0.93+0.37	0.12+0.13

As shown in Tables 9 and 10, the rats were orally administrated with the test sample at doses of 5000, 2500 and 1250 mg/kgBW for 30 days. There were no statistically significant differences in hemoglobin, total red blood cell count and total WBC count of male and female rats between the test groups and the control group (P> 0.05), which indicated that the test sample had no obvious effect on the blood routine indexes in rats.

2.5.4 Effect of the test sample on blood biochemical indexes in rats 5 The results are shown in Tables 11 and 12. The rats were orally administrated with the test sample at doses of 5000, 2500, and 1250 mg/kg BW for 30 days. There were no statistically significant differences in serum aspartate aminotransferase (SGOT), alanine aminotransferase (ALT), blood urea nitrogen (BUN), creatinine (Cre), cholesterol, triglycerides, total protein, albumin, blood glucose of male and female rats between the test groups and the control group (P> 0.05), which indicated that the test sample had no obvious effect on the blood biochemical indexes of rats.

Table 11. The results of blood biochemical indexes in rats at the end of thirty days feeding test of the test sample (^χ ± s)

Gender	Group	Number of animals	SGOT (U/L)	ALT (U/L)	BUN (mmol/L)	Cre (pmol/L)
			124.1+13.1	49.1+11.1	5.90+0.42	42.8+2.5
	High-dose	10	129.2+28.7	49.2+9.3	5.79+0.48	42.9+2.7
Male	Mid-dose	10	130.0+28.1	50.6+9.2	5.85+0.44	42.4+2.0
	Low-dose	10	129.4+15.4	47.8+7.5	5.87+0.47	43.3+2.8
	Control	10	125.3+19.0	48.6+8.2	6.07+0.57	41.0+3.5
	High-dose	10	127.6+17.1	47.0+8.9	5.97+0.67	42.6+4.3
Female	Mid-dose	10	128.0+17.9	48.0+9.6	6.10+0.47	42.0+4.0
	Low-dose	10	123.9+16.1	48.7+7.9	5.92+0.42	41.3+2.5

Table 12. The results of blood biochemical indexes in rats at the end of thirty days feeding test of the test sample (^χ ± s)

Gender	Group	Number of animals	cholesterol (mmol/L)	triglycerides (mmol/L)	total protein (g/L)	albumin (g/L)	blood glucose (mmol/L)
			2.02+0.15	1.18+0.27	71.0+3.3	37.4+1.4	5.55+0.65
	High-dose	10	2.04+0.12	1.13+0.43	70.9+7.5	36.2+2.9	5.40+0.66
Male	Mid-dose	10	2.09+0.14	1.15+0.35	71.2+6.9	36.9+2.6	5.51+0.49
	Low-dose	10	2.07+0.14	1.13+0.43	69.9+6.9	36.4+2.7	5.42+0.65
	Control	10	2.16+0.34	1.17+0.31	71.0+7.1	38.1+2.8	5.73+0.51
	High-dose	10	2.12+0.45	1.07+0.36	73.2+5.9	37.1+2.4	5.83+0.44
Female	Mid-dose	10	2.19+0.41	1.12+0.28	72.6+5.3	37.2+1.9	5.78+0.35
	Low-dose	10	2.07+0.40	1.24+0.27	71.4+8.3	37.7+4.3	5.84+0.48

2.5.5 The effect of the test sample on organ weight and organ/body weight ratio in rats

The rats were orally administrated with the test sample at the dose of 5000, 2500, and 1250 18 mg/kg BW for 30 days. There were no statistically significant differences in the weights of liver, kidney, spleen, testicle and the weight ratio of liver/body, kidney/body, spleen/body, testicle/body in the test groups and the control group (P> 0.05), which indicated that the test sample has no significant effects on the was no significant difference in the organ weight and organ / body weight ratio between organ weight and organ/body weight ratio in rats.

2.5.6 Results of general anatomy observation and histological examination

All the animals were dissected at the end of the test, and no obvious lesions were observed. Therefore, only the high-dose test group and the negative control group were selected to detect the major organs of animals by histopathological examination method, and the results were shown in Tables 18-24. The results showed that there were one male in the high-dose group and one male and one female in the control group with the vacuolar degeneration of liver cells in hepatic lobule; one male and one female in the high-dose group and one male and two females in the control group with the point necrosis of liver cells in hepatic lobule; one male and two females in the high-dose group and one male and two females in the control group with little infiltration of inflammatory cells in liver portal area; and two males and one female in the highdose group and one male and one female in the control group with little infiltration of inflammatory cells in interstitial cells of renal cortex. The above-mentioned histological lesions belong to the spontaneous mild lesions of animals and the degree of tissue lesions between the two groups of animals were similar, so that it can be ruled out that the lesions were caused by the test sample. Since the pathological changes in other organs were not observed, which indicated that the test sample has no damages on the organs in rats.

3. Conclusion

There were no toxic symptoms and death in the tested animals after orally administration of the sample of the invention at the dose of 20000 mg/kg BW, so that the acute oral toxicity MTD of the sample of the invention is more than 20,000 mg/kg BW, and the acute oral toxicity is nontoxic level. During 30 days feeding test (chronic toxicity test), the rats were fed with the sample at the three doses of 5000, 2500, 1250 mg/kg BW (100, 50 and 25 times of the human recommended dosage) for 30 days, and the animals grown and developed well. There were no statistically significant differences in the body weight, weight gain, food intake, food utilization rate, blood routine indexes, blood biochemical indexes, organ weight and organ/body weight ratio of the animals between each test group and the control group (P > 0.05). There were no abnormal changes related to the sample in general anatomic observation and histopathological examination. No significant adverse effects of the sample were observed in the range of the tested dose.

Three genetic toxicity tests:

1. Ames test

1.1 Methods:

Four Salmonella typhimurium strains of histidine-deficient TA97a, TA98, TAI00, TAI02 identified to meet the requirements were used in the test. 5.0 g of the sample of the present invention was weighed and added with pure water to 100 mL, and then mixed into a 50 mg/mL solution, followed by a 5-fold dilution (10 mL to 50 mL of pure water) in turn to prepare 10, 2, 0.4, 0.08 mg/mL solutions, respectively. The prepared solutions were treated by autoclaving (0.103Mpa, 20min) for test. The rat liver microsomal enzyme (S-9) induced by polychlorinated biphenyls (PCBs) was used as an in vitro metabolic activation system. By the plate incorporation method, 0.1 mL of the test strains enrichment broths, 0.1 mL of the test sample solutions and 0.5 mL of the S-9 mixture (when metabolic activation was required) were added into the incubated top medium plate, and then mixed and poured into the bottom medium plate. The 5 tested doses were 5000, 1000, 200, 40, 8qg/plate respectively, and the spontaneous revertant control groups, solvent control groups and positive mutant control groups were prepared at the same time. The spontaneous change control groups had the same conditions as the test sample groups, except without the sample. The solvent control groups has the same conditions as the test sample groups except that the sample was replaced with sterile pure water. Three parallel plates were made for each strain in each group. The strains were incubated at 37 °C for 48 hours, and the number of colonies per plate was counted. The whole test was repeated twice under the same conditions. If the number of revertant colonies of the test sample was increased by more than 2 times of the number of spontaneous revertant colonies and there was a dose-response relationship, then that is the positive mutagenic test.

1.2 Results:

As for the test strains of TA97a, TA98, TAI00, TAI02, with or without addition of S-9, the number of revertant colonies of the test sample in each dose test group was increased by more than 2 times of the number of spontaneous revertant colonies without any dose-response relationship, which indicating that the mutagenicity test results for the sample of the present invention were negative.

2. Micronucleus test of mouse bone marrow cells

2.1 Methods:

The test was performed by oral perfusion twice at 24 hours interval. Fifty Kunming mice with weighting 25 ~ 30g were randomly divided into 5 groups, 10 in each group. The three test groups were administrated with the test sample at the doses of 10000, 5000 and 2500 mg/kgBW, respectively. The pure water was used as the negative control and 40 mg/kgBW dose of cyclophosphamide (cp) was used as the positive control. 20.0, 10.0, 5.0g of the test sample were weighed and added with pure water to 40 mL, and mixed to prepare 500, 250, 125 mg/mL suspension. The test groups were administrated at the volume of 0.4mL/20g BW, the negative control group was given equal volume of pure water, and the positive control group was given equal volume of 2 mg/mL cyclophosphamide solution. All the animals were sacrificed at 6 hour after the second administration. The sternum marrow was diluted with calf serum on smear, fixed with methanol and stained with Giemsa. Under light microscope, 1000 polychromatic erythrocytes (PCE) per each animal were counted to observe the number of PCE containing micronucleus and calculate the micronucleus rate (%o); while 200 polychromatic erythrocytes were counted to calculate the ratio of polychromatic erythrocytes to mature red blood cells (PCE/NCE). The data was statistically analyzed by using the Poisson distribution mean comparison method in SPSS statistical software. If the micronucleus rate of the test group was higher than the negative control group and there is an obvious dose-response relationship and statistical significance, then the test had a positive result.

2.2 Results:

There was no significantly difference between the micronucleus rate of mouse bone marrow cells in each dose test group of the test sample of the present invention and that in the negative control group (P> 0.05), and the PCE/NCE value in each dose test group was not less than 20% so that there was no significant difference between the test group and the negative control group (P<0.01). But there was a significant difference between the micronucleus rate in the cyclophosphamide positive control group and that in the negative control group (P <0.01). Such results indicated that the test sample of the invention has no damage and inhibition effects on mouse bone marrow cells.

3. Mouse sperm abnormality test

3.1 Methods:

Fifty Kunming male mice with weight of 25-35 g were selected and randomly divided into 5 groups, 10 in each group. Three test groups were administrated with the test sample at the doses of 10000, 5000 and 2500 mg/kgBW, respectively. The pure water was used as the negative control and 40 mg/kgBW dose of cyclophosphamide (cp) was used as the positive control. 20.0, 10.0, 5.0g of the test sample were weighed and added with pure water to 40 mL, and mixed to prepare 500, 250, 125 mg/mL suspension. The test groups were administrated at the volume of 0.4mL/20g BW, the negative control group was given equal volume of pure water, and the positive control group was given equal volume of 2 mg/mL cyclophosphamide solution. All the animals were administrated by gavage once a day for 5 days. All the animals were sacrificed at 30 days after the final administration. The epididymal sperm was taken on smear, fixed with methanol and stained with Eosin. Under light microscope, 1000 intact sperms per each animal were counted to calculate the sperm deformity rate. The data was statistically analyzed by using the Wilcoxon rank sum test in SPSS statistical software. If the sperm deformity rate in the test group was higher than the negative control group and there is an obvious dose-response relationship and statistical significance, then the test had a positive result.

3.2 Results:

The sample of the present invention does not cause a significant change for the sperm deformity rate in mice. There was no significant difference between the sperm deformity rate in each dose test group and that in the negative control group (P> 0.05), and there was a significant difference between the sperm deformity rate in the cyclophosphamide positive control group and the negative control group (P <0.01), which indicating that the sample of the present invention has no teratogenic effect on the sperm of male mice.

4. Conclusion:

The three genetic toxicity tests (Ames test, mouse bone marrow cell micronucleus test, mouse sperm abnormality test) were all negative, which indicating that the sample of the present invention has no genetic toxicity.

The experimental studies described herein confirmed that the health food provided by the invention has obvious hypoglycemic health care function and is safe and non-toxic.

It should be understood by those skilled in the art that the foregoing discussion of any embodiment is intended to be illustrative and not intended to suggest that the scope of the present disclosure (including the claims) is limited to the examples. According to the idea of the present invention, the technical features in various embodiments may also be combined, and there are many other variations of the various aspects of the invention as described above, which are not described in details for the sake of brevity. Therefore, any omissions, modifications, equivalents, improvements and the like within the spirit and principles of the present invention are intended to be fallen within the scope of the present invention.

Claims

To be claimed:

1. A health food, characterized in that the active ingredients of the health food is composed of the following raw materials of Chinese traditional medicine in parts by weight:

5 to 30 parts of Radix Puerariae;
2 to 20 portions of Gynostemma pentaphyllum;

0.5 to 8 parts of Panax notoginseng;

1 to 20 parts of Fructus Mori Albae;

1 to 10 parts of Flesh of Meretrix meretrix Finnaeus; and

2 to 20 parts of Spirulina.

2. The health food as claimed in claim 1, wherein the active ingredients of the health food is composed of the following raw materials of Chinese traditional medicine in parts by weight:

7 to 25 parts of Radix Puerariae;

5 to 15 portions of Gynostemma pentaphyllum;

1 to 5 portions of Panax notoginseng;

4 to 15 parts of Fructus Mori Albae;

2 to 8 parts of Flesh of Meretrix meretrix Finnaeus; and

4 to 15 parts of Spirulina.
3. A health food, characterized in that the active ingredients of the health food is composed of the following raw materials of Chinese traditional medicine in parts by weight:

5 to 30 portions of Radix Puerariae extract;

2 to 20 portions of Gynostemma pentaphyllum extract;

0.5 to 8 portions of Panax notoginseng extract;

1 to 20 parts of Fructus Mori Albae extract;

1 to 10 parts of Flesh of Meretrix meretrix Finnaeus extract;

2 to 20 portions of Spirulina extract.
4. The health food as claimed in claim 3, wherein the active ingredients of the health food is composed of the following raw materials of Chinese traditional medicine in parts by weight:

7 to 25 parts of Radix Puerariae extract;
5 to 15 portions of Gynostemma pentaphyllum extract;

1 to 5 portions of Panax notoginseng extract;

4 to 15 portions of Fructus Mori Albae extract;

2 to 8 parts of Flesh of Meretrix meretrix Finnaeus extract;

4 to 15 parts of Spirulina extract.

5. The health food as claimed in any one of claims 1 to 4, wherein the health food is used for preparing health food or medication with hypoglycemic effect which is safe without toxic side effects, and the dosage form comprises oral solid, oral semi-solid and oral solution.
6. A process for preparing the health food as claimed in any one of claims 1 to 4, comprising the following steps that:

the mixture of the Radix Puerariae and the Gynostemma pentaphyllum is extracted by refluxing alcohol, and then the extracting solution is concentrated, dried and crushed to obtain the extract I;

the mixture of Fructus Mori Albae, the Meretrix meretrix Linnaeus and the Spirulina is extracted by boiling water and concentrated, and then the concentrated solution is added by ethanol and filtered to obtain the precipitate which is dried and crushed to obtain the extract II; the Panax notoginseng is crushed into the Panax notoginseng powder as reserve; and the extract I, the extract II and the Panax notoginseng powder are mixed to obtain the health food.
7. The process as claimed in claim 6, wherein the step of preparing the extract I comprises the steps that:

the mixture of the Radix Puerariae and the Gynostemma pentaphyllum is extracted for 1-4 times by refluxing 6 to 10 times amount of alcohol for 1 to 2 hours each time, and then the extracting solution is combined, concentrated, dried at 80~90°C, crushed and screened through 80-120 mesh sieve to obtain the extract I.
8. The process as claimed in claim 6, wherein the step of preparing the extract I comprises the steps that:

the mixture of the Radix Puerariae and the Gynostemma pentaphyllum is soaked by 6-10 times amount of alcohol for 30-60 minutes and extracted by reflux for 1-2 hours firstly, and then 6-8 times amount of alcohol is added to extract for 1-2 hours, and then the extracting solution is combined, concentrated, dried at 80~90°C, crushed and screened through 80-120 mesh sieve to obtain the extract I.
9. The process as claimed in claim 6, wherein the step of preparing the extract II comprises the steps that:

the mixture of Fructus Mori Albae, the Meretrix meretrix Linnaeus and the Spirulina is extracted for 1-4 times by boiling 6 to 10 times amount of water for 1 to 2 hours each time, and then the extracting solution is combined and concentrated; and the concentrated solution is deposited by adding ethanol for 16-30 hours, and filtered to obtain the precipitate which is dried and crushed to obtain the extract II.
10. The process as claimed in claim 9, wherein the step of preparing the extract II comprises the steps that:

the mixture of Fructus Mori Albae, the Meretrix meretrix Linnaeus and the Spirulina is soaked by 6-10 times amount of water for 30-60 mins and extracted by boiling for 1-2 hours firstly, and then 5-9 times amount of water is added to extract for 1-2 hours, and then the extracting solution is combined and concentrated; and the concentrated solution is added by 80~95vol% ethanol until the volume concentration of alcohol arriving 70-90% and then deposited for 16-30 hours, and filtered to obtain the precipitate which is dried at 60~75°C, crushed and screened through 80-120 mesh sieve to obtain the extract

5 II.