Application of luteolin in bullfrog breeding, bullfrog tadpole metamorphosis development promotion and sex differentiation regulation

Abstract

The invention discloses application of luteolin in bullfrog culture, promotion of bullfrog tadpole metamorphosis development and sex differentiation regulation. The luteolin is utilized to effectively shorten the metamorphosis development period of bullfrog tadpoles into froglets, accelerate the metamorphosis development of the bullfrog tadpoles, improve the success rate of the metamorphosis development of the bullfrog tadpoles, improve the breeding efficiency, promote the male differentiation of the bullfrog tadpoles and increase the yield of male bullfrogs; the method has the remarkable advantages of greenness, safety, simplicity and convenience in operation and stable effect. The invention provides a new technical choice for bullfrog breeding industry, which is beneficial to optimizing germplasm resources of bullfrogs and improving economic benefits.

Description

Technical Field

[0001] The invention relates to the technical field of bullfrog breeding, and in particular to application of luteolin in bullfrog breeding and promoting the metamorphosis and regulating sex differentiation of bullfrog tadpoles. Background Art

[0002] Bullfrog is a large edible frog with tender meat, delicious taste, rich nutrition and strong fertility. It has been widely cultivated in China. Bullfrog is an amphibian, and its reproductive development process has significant metamorphosis characteristics. The sex determination and sex differentiation of amphibians occur during the metamorphosis stage, and genetic and environmental factors jointly affect the sex development of bullfrogs. The metamorphosis period of bullfrog tadpoles is a critical period in the breeding process, which will directly determine the success or failure of later adult frog breeding.

[0003] At present, due to the relatively backward development of bullfrog conservation and breeding technology, factors such as inbreeding and drug abuse have led to the degeneration of bullfrog parents. The metamorphosis of tadpoles bred takes a long time and the metamorphosis success rate is low, which restricts the healthy development of the bullfrog seedling, breeding and breeding industry chain. In addition, compared with female bullfrogs, male bullfrogs have the advantages of faster growth, heavier hind leg weight, and higher meat yield. It is necessary to induce as many bullfrog tadpoles as possible to develop into male bullfrogs. Therefore, accelerating metamorphosis and increasing the male rate are of great significance to optimizing the population structure of bullfrogs, accelerating the breeding efficiency of bullfrogs, and increasing the economic benefits of bullfrog farming.

[0004] Although there are some methods in the prior art to accelerate tadpole development and adjust the sex ratio, such as feeding hormones or chemical drugs, temperature control, light regulation, etc., these methods often have safety hazards such as drug residues, complex operations, high costs, and unstable effects. Therefore, it is urgent to develop a safer, simpler, more efficient and economical method to accelerate the metamorphosis of bullfrog tadpoles and increase the yield of male bullfrogs. Summary of the invention

[0005] In order to solve the problems of low metamorphosis efficiency, low metamorphosis success rate and low ratio of bred male bullfrogs in the prior art, and to explore a new breeding model for bullfrog tadpoles during metamorphosis that is greener, safer, simpler, more efficient and more economically beneficial, the present invention provides the application of luteolin in bullfrog breeding and in promoting the metamorphosis of bullfrog tadpoles and regulating sex differentiation.

[0006] The first object of the present invention is to provide the application of luteolin in accelerating the metamorphosis of bullfrog tadpoles.

[0007] The second object of the present invention is to provide the use of luteolin in improving the success rate of metamorphosis of bullfrog tadpoles.

[0008] The third object of the present invention is to provide the use of luteolin in promoting the male differentiation of bullfrog tadpoles.

[0009] The fourth object of the present invention is to provide an application in any two of accelerating the metamorphosis of bullfrog tadpoles, improving the success rate of the metamorphosis of bullfrog tadpoles, or promoting the male differentiation of bullfrog tadpoles.

[0010] The fifth objective of the present invention is to provide the use of luteolin in accelerating the metamorphosis of bullfrog tadpoles, improving the success rate of metamorphosis of bullfrog tadpoles and promoting the male differentiation of bullfrog tadpoles.

[0011] The sixth object of the present invention is to provide the use of luteolin in the preparation of a bullfrog feed additive and / or a bullfrog feed having any one, two or three functions of accelerating the metamorphosis of bullfrog tadpoles, improving the success rate of metamorphosis of bullfrog tadpoles or promoting the male differentiation of bullfrog tadpoles.

[0012] The seventh object of the present invention is to provide a bullfrog feed additive or bullfrog feed.

[0013] The eighth object of the present invention is to provide the use of the bullfrog feed additive or bullfrog feed in any one, two or three of accelerating the metamorphosis of bullfrog tadpoles, improving the success rate of metamorphosis of bullfrog tadpoles or promoting the male differentiation of bullfrog tadpoles.

[0014] In order to achieve the above object, the present invention is implemented by the following scheme:

[0015] Luteolin is widely found in many plants such as honeysuckle and perilla, and has multiple pharmacological activities such as anti-inflammatory, anti-allergic, anti-tumor, and antibacterial. As a natural plant extract, luteolin is environmentally friendly, highly safe, and will not pollute aquaculture water. It has been used in the breeding of poultry and livestock.

[0016] The present invention applies luteolin to bullfrog farming for the first time, and finds that luteolin has a direct and significant effect on the metamorphosis and sex differentiation of bullfrog tadpoles. By adding luteolin to the feed, it can not only promote the bullfrog tadpoles to complete metamorphosis quickly and efficiently, but also significantly increase the proportion of male individuals in the farmed bullfrogs.

[0017] Therefore, the present invention requests protection for the following contents:

[0018] The application of luteolin in accelerating the metamorphosis of bullfrog tadpoles. The accelerating the metamorphosis of bullfrog tadpoles is to shorten the metamorphosis period of bullfrog tadpoles.

[0019] Preferably, the luteolin is used for feeding bullfrog tadpoles and / or bullfrog larvae.

[0020] Preferably, the content of luteolin in the bullfrog tadpole feed is 0.5%wt to 2%wt.

[0021] More preferably, the content of luteolin in the feed is 0.5%wt, 1%wt or 2%wt.

[0022] Further preferably, the content of luteolin in the feed is 1%wt.

[0023] The application of luteolin in improving the success rate of metamorphosis of bullfrog tadpoles. The success rate of metamorphosis of bullfrog tadpoles is the ratio of bullfrog tadpoles that successfully develop into young frogs to all bullfrog tadpoles cultured, and the bullfrog tadpoles that successfully develop into young frogs do not include bullfrog tadpoles that die during metamorphosis and bullfrog tadpoles with deformed development.

[0024] Preferably, the luteolin is used for feeding bullfrog tadpoles and / or bullfrog larvae.

[0025] Preferably, the content of luteolin in the bullfrog tadpole feed is 0.5%wt to 2%wt.

[0026] More preferably, the content of luteolin in the feed is 0.5%wt, 1%wt or 2%wt.

[0027] Further preferably, the content of luteolin in the feed is 1%wt.

[0028] The application of luteolin in promoting the male differentiation of bullfrog tadpoles. The promotion of the male differentiation of bullfrog tadpoles is to promote the development of bullfrog tadpoles into male bullfrogs and increase the male rate of bullfrogs.

[0029] Preferably, the luteolin is used for feeding bullfrog tadpoles and / or bullfrog larvae.

[0030] Preferably, the content of luteolin in the bullfrog tadpole feed is 0.5%wt to 2%wt.

[0031] More preferably, the content of luteolin in the feed is 0.5%wt, 1%wt or 2%wt.

[0032] Further preferably, the content of luteolin in the feed is 1%wt.

[0033] The use of luteolin in any two of accelerating the metamorphosis of bullfrog tadpoles, improving the success rate of metamorphosis of bullfrog tadpoles, or promoting the male differentiation of bullfrog tadpoles.

[0034] Preferably, the luteolin is used for feeding bullfrog tadpoles and / or bullfrog larvae.

[0035] Preferably, the content of luteolin in the bullfrog tadpole feed is 0.5%wt to 2%wt.

[0036] More preferably, the content of luteolin in the feed is 0.5%wt, 1%wt or 2%wt.

[0037] Further preferably, the content of luteolin in the feed is 1%wt.

[0038] The application of luteolin in accelerating the metamorphosis of bullfrog tadpoles, improving the success rate of metamorphosis of bullfrog tadpoles and promoting the male differentiation of bullfrog tadpoles.

[0039] Preferably, the luteolin is used for feeding bullfrog tadpoles and / or bullfrog larvae.

[0040] Preferably, the content of luteolin in the bullfrog tadpole feed is 0.5%wt to 2%wt.

[0041] More preferably, the content of luteolin in the feed is 0.5%wt, 1%wt or 2%wt.

[0042] Further preferably, the content of luteolin in the feed is 1%wt.

[0043] The application of luteolin in the preparation of bullfrog feed additives and / or bullfrog feeds having any one, two or three functions of accelerating the metamorphosis of bullfrog tadpoles, improving the success rate of metamorphosis of bullfrog tadpoles or promoting the male differentiation of bullfrog tadpoles.

[0044] Preferably, the bullfrog feed additive and / or bullfrog feed is used to feed bullfrog tadpoles and / or bullfrog larvae.

[0045] A bullfrog feed additive comprises 0.5%wt-2%wt luteolin.

[0046] Preferably, the content of luteolin in the bullfrog feed additive is 0.5%wt, 1%wt or 2%wt.

[0047] More preferably, the content of luteolin in the bullfrog feed additive is 1%wt.

[0048] Preferably, it also contains an acceptable carrier.

[0049] The use of any one of the bullfrog feed additives in accelerating the metamorphosis of bullfrog tadpoles, improving the success rate of the metamorphosis of bullfrog tadpoles, or promoting the male differentiation of bullfrog tadpoles.

[0050] A bullfrog feed comprises 0.5%wt-2%wt luteolin.

[0051] Preferably, the content of luteolin in the bullfrog feed is 0.5%wt, 1%wt or 2%wt.

[0052] More preferably, the content of luteolin in the bullfrog feed is 1%wt.

[0053] Preferably, the bullfrog feed also contains protein feed, sugars and oils.

[0054] More preferably, the protein feed comprises animal protein feed and plant protein feed.

[0055] Further preferably, the animal protein feed comprises fish meal and chicken meal.

[0056] Further preferably, the plant protein feed includes soybean meal, rapeseed meal and cottonseed meal.

[0057] More preferably, the sugar comprises corn starch.

[0058] More preferably, the oils and fats include animal oils and fats and vegetable oils and fats.

[0059] More preferably, the animal fat comprises fish oil.

[0060] More preferably, the vegetable oil includes soybean oil.

[0061] More preferably, the bullfrog feed further comprises vitamins and minerals.

[0062] The vitamins are vitamins for frogs, including but not limited to vitamin A, vitamin B, vitamin C, vitamin D, vitamin E and vitamin K.

[0063] Further preferably, the vitamins are multivitamins for frogs, including vitamin A, vitamin D3, vitamin E2, vitamin K3, vitamin B1, vitamin B2, vitamin B6, vitamin B12, inositol, pantothenic acid, choline, folic acid, biotin and vitamin C1.

[0064] The minerals are frog-usable minerals, including but not limited to calcium salts, phosphorus salts, sodium salts, potassium salts, iodine salts, magnesium salts, iron salts and zinc salts.

[0065] Further preferably, the minerals are mixed minerals for frogs, including CaCO3, KH2PO4, MgSO4·7H2O, NaCl, Fe(II) gluconate, MnSO4·7H2O, ZnSO4·7H2O, CuSO4·5H2O, KI, CoCl2·6H2O, NH4 molybdate and NaSeO3.

[0066] More preferably, the feed also contains the fish meal, the chicken meal, the soybean meal, the rapeseed meal, the cottonseed meal, the corn starch, the fish oil, the soybean oil, the complex vitamins and the mixed minerals, and the mass ratios thereof are (40-60): (30-40): (140-160): (70-90): (60-80): (70-75): (6-9): (6-9): (2-4): (2-4).

[0067] Still further preferably, the feed also comprises the fish meal, the chicken meal, the soybean meal, the rapeseed meal, the cottonseed meal, the corn starch, the fish oil, the soybean oil, the complex vitamins and the mixed minerals, and the mass ratios thereof are 50:40:150:80:75:72.5:7.5:7.5:2.5:2.5 respectively.

[0068] The use of any one of the feeds in accelerating the metamorphosis of bullfrog tadpoles, improving the success rate of the metamorphosis of bullfrog tadpoles, or promoting the male differentiation of bullfrog tadpoles, or any one, two, or three of them.

[0069] Compared with the prior art, the present invention has the following beneficial effects:

[0070] The present invention utilizes luteolin to effectively shorten the metamorphosis period of bullfrog tadpoles to young frogs, accelerate the metamorphosis of bullfrog tadpoles, improve the success rate of metamorphosis of bullfrog tadpoles, improve the breeding efficiency, promote the male differentiation of bullfrog tadpoles, and increase the yield of male bullfrogs. The invention has the significant advantages of being green, safe, easy to operate, and having stable effects. The present invention provides a new technical option for the bullfrog breeding industry, helps to optimize the germplasm resources of bullfrogs, and improves economic benefits. DETAILED DESCRIPTION

[0071] The present invention is further described in detail below in conjunction with specific examples, which are only used to explain the present invention and are not used to limit the scope of the present invention. The test methods used in the following examples are conventional methods unless otherwise specified; the materials, reagents, etc. used are reagents and materials that can be obtained from commercial channels unless otherwise specified. The frog multivitamin used in the following examples is a commercially available vitamin premix (mg / kg), containing 500,000 IU of vitamin A, 3.5 million IU of vitamin D, 500 mg of vitamin E2, 1000 mg of vitamin K3, 5000 mg of vitamin B1, 5000 mg of vitamin B2, 5000 mg of vitamin B6, 5000 μg of vitamin B12, 25000 mg of inositol, 10000 mg of pantothenic acid, 10000 mg of choline, 10000 mg of folic acid, 250 mg of biotin and 10000 mg of vitamin C); the frog mixed mineral used is a commercially available mixed mineral (g / kg), containing 314.0 g of CaCO3, 469.3 g of KH2PO4, 147.4 g of MgSO4·7H2O, 49.8 g of NaCl, 10.9 g of Fe(II) gluconate, and 3.12 g of MnSO4·7H2O. g, ZnSO4·7H2O4.67 g, CuSO4·5H2O0.62 g, KI0.16 g, CoCl2·6H2 O0.08 g, NH4 molybdate 0.06 g and NaSeO30.02 g.

[0072] Embodiment 1 A kind of bullfrog feed

[0073] This embodiment provides a bullfrog feed, which is prepared by adding luteolin (CAS#: 491-70-3, purity ≥ 98%) to a standard feed for bullfrog tadpoles and mixing according to a conventional method. The specific formula is as follows:

[0074] Standard feed: by weight: 50 parts of fish meal, 40 parts of chicken meal, 150 parts of soybean meal, 80 parts of rapeseed meal, 75 parts of cottonseed meal, 72.5 parts of corn starch, 7.5 parts of fish oil, 7.5 parts of soybean oil, 2.5 parts of multivitamins for frogs; 2.5 parts of mixed minerals for frogs.

[0075] The mass ratio of luteolin to standard feed was 5:1000.

[0076] Embodiment 2 A kind of bullfrog feed

[0077] This embodiment provides a bullfrog feed, which is prepared by adding luteolin (CAS#: 491-70-3, purity ≥ 98%) to a standard feed for bullfrog tadpoles and mixing according to a conventional method. The specific formula is as follows:

[0078] Standard feed: by weight: 50 parts of fish meal, 40 parts of chicken meal, 150 parts of soybean meal, 80 parts of rapeseed meal, 75 parts of cottonseed meal, 72.5 parts of corn starch, 7.5 parts of fish oil, 7.5 parts of soybean oil, 2.5 parts of multivitamins; 2.5 parts of mixed minerals.

[0079] The mass ratio of luteolin to standard feed was 10:1000.

[0080] Embodiment 3 A kind of bullfrog feed

[0081] This embodiment provides a bullfrog feed, which is prepared by adding luteolin (CAS#: 491-70-3, purity ≥ 98%) to a standard feed for bullfrog tadpoles and mixing according to a conventional method. The specific formula is as follows:

[0082] Standard feed: by weight: 50 parts of fish meal, 40 parts of chicken meal, 150 parts of soybean meal, 80 parts of rapeseed meal, 75 parts of cottonseed meal, 72.5 parts of corn starch, 7.5 parts of fish oil, 7.5 parts of soybean oil, 2.5 parts of multivitamins for frogs; 2.5 parts of mixed minerals for frogs.

[0083] The mass ratio of luteolin to standard feed was 20:1000.

[0084] Comparative Example 1: Standard feed for bullfrog

[0085] This comparative example provides a standard feed for bullfrogs, and the specific formula is as follows:

[0086] Calculated by weight, 50 parts of fish meal, 40 parts of chicken meal, 150 parts of soybean meal, 80 parts of rapeseed meal, 75 parts of cottonseed meal, 72.5 parts of corn starch, 7.5 parts of fish oil, 7.5 parts of soybean oil, 2.5 parts of multivitamins for frogs; 2.5 parts of mixed minerals for frogs.

[0087] Comparative Example 2: A bullfrog feed

[0088] This comparative example provides a bullfrog feed, which is prepared by adding thyroid hormone (CAS#: 300-30-1, purity ≥ 98%) to the standard feed of bullfrog tadpoles. The specific formula is as follows:

[0089] Standard feed: by weight: 50 parts of fish meal, 40 parts of chicken meal, 150 parts of soybean meal, 80 parts of rapeseed meal, 75 parts of cottonseed meal, 72.5 parts of corn starch, 7.5 parts of fish oil, 7.5 parts of soybean oil, 2.5 parts of multivitamins for frogs; 2.5 parts of mixed minerals for frogs.

[0090] The mass ratio of thyroid hormone to standard feed is 30:1000000.

[0091] Comparative Example 3: A bullfrog feed

[0092] This comparative example provides a bullfrog feed, which is prepared by adding estrogen (CAS#: 35950-87-9, purity ≥ 98%) to the standard feed of bullfrog tadpoles. The specific formula is as follows:

[0093] Standard feed: by weight: 50 parts of fish meal, 40 parts of chicken meal, 150 parts of soybean meal, 80 parts of rapeseed meal, 75 parts of cottonseed meal, 72.5 parts of corn starch, 7.5 parts of fish oil, 7.5 parts of soybean oil, 2.5 parts of multivitamins for frogs; 2.5 parts of mixed minerals for frogs.

[0094] The mass ratio of estrogen to standard feed is 30:1000000.

[0095] Comparative Example 4: A bullfrog feed

[0096] This comparative example provides a bullfrog feed, which is obtained by adding progesterone (CAS#: 3562-63-8, purity ≥ 98%) to the standard feed of bullfrog tadpoles. The specific formula is as follows:

[0097] Standard feed: by weight: 50 parts of fish meal, 40 parts of chicken meal, 150 parts of soybean meal, 80 parts of rapeseed meal, 75 parts of cottonseed meal, 72.5 parts of corn starch, 7.5 parts of fish oil, 7.5 parts of soybean oil, 2.5 parts of multivitamins for frogs; 2.5 parts of mixed minerals for frogs.

[0098] The mass ratio of progesterone to standard feed is 30:1000000.

[0099] Application Example 1 Evaluation of the feeding effect of bullfrog tadpoles

[0100] 1. Breeding test

[0101] The bullfrog tadpoles used in this experiment were all from Guangzhou Guanxing Agricultural Technology Co., Ltd., and the experiment was carried out at the Zhongshan Innovation Base of South China Agricultural University.

[0102] Bullfrog tadpoles (weighing about 0.02 g) with similar development, good health and no obvious deformities were randomly divided into 7 groups, each with 120 tadpoles. Each group was fed with the feeds of Examples 1 to 3 and Comparative Examples 1 to 4 in turn, and conventional feeding management was carried out under the same environmental conditions, and feeding was carried out according to conventional methods, and continued to be raised until complete metamorphosis.

[0103] During the rearing period, the metamorphosis of bullfrog tadpoles in each group was observed regularly, and the growth stage of the tadpoles was determined by using the Gosner staging method. When the tail of the bullfrog tadpole completely disappeared and turned into a young frog, the metamorphosis was completed.

[0104] The body length, weight, date of becoming a young frog and the sex of the young frog of the bullfrog tadpoles were recorded in time. When the bullfrogs were raised to 80 days (the metamorphosis rate of half of the groups exceeded 50%), the final weight gain, weight gain rate, specific growth rate, fatness, feed coefficient and frog rate of each group of bullfrogs were counted. When the end point of the feeding was reached (the tadpoles of all groups completed metamorphosis), the metamorphosis success rate and male rate of each group of bullfrogs were counted. Thyroid hormone plays a key role in the growth and development process of bullfrogs, especially in the metamorphosis stage, so a commercial test kit was also used to detect the thyroid hormone level in the young frogs at the end point of the feeding.

[0105] The calculation formula involved in the above test method is as follows:

[0106] Final weight gain (g) = final weight - initial weight; Weight gain rate (%) = final weight gain ÷ initial weight × 100%; Specific growth rate (% / day) = {ln (final weight) - ln (initial weight)} ÷ number of days of feeding × 100%; Fatness (g / cm 3 ) = weight ÷ body length 3 ; Feed coefficient (%) = weight of feed ingested during the feeding period ÷ final weight gain × 100%; Adult frog rate (%) = number of young frogs at 80 days ÷ total number of tadpoles at the beginning × 100%; Metamorphosis success rate (%) = number of young frogs at the end of feeding ÷ total number of tadpoles at the beginning × 100%; Mortality rate (%) = number of dead tadpoles at the end of feeding ÷ total number of tadpoles at the beginning × 100%; Deformity rate (%) = number of deformed tadpoles at the end of feeding ÷ total number of tadpoles at the beginning × 100%; Male rate (%) = number of male frogs at the end of feeding ÷ total number of tadpoles at the beginning × 100%.

[0107] 2. Test results

[0108] Table 1 Effects of different feeds on the growth and development of bullfrog tadpoles fed for 80 days

[0109]

[0110] As shown in Table 1, when fed for 80 days, the final weight gain, weight gain rate, specific growth rate and fatness of groups 1 to 3 were significantly higher and the feed conversion rate was significantly lower than those of groups 4 to 7. This indicates that the standard feed with luteolin added (Examples 1 to 3) can promote the growth and development of bullfrog tadpoles and improve meat production performance more than the conventional standard feed (Comparative Example 1) and the standard feed with hormone added (Comparative Examples 2 to 4).

[0111] Table 2 Effects of different feeds on the metamorphosis of bullfrog tadpoles fed for 80 days

[0112]

[0113]

[0114] As shown in Table 2, after 80 days of feeding, the frog-making rate of bullfrog tadpoles in groups 1 to 3 was above 60%, which was 3 to 4 times that of group 4 and significantly higher than that of groups 5 to 7; the earliest frog-making days of groups 1 to 3 were significantly less than those of groups 4, 6 to 7, and equivalent to those of group 5. This indicates that under intensive breeding conditions, the standard feed with luteolin added (Examples 1 to 3) can accelerate the overall metamorphosis process of bullfrog tadpoles and improve the breeding efficiency compared with the conventional standard feed (Comparative Example 1) and the standard feed with hormone added (Comparative Examples 2 to 4).

[0115] Table 3 Effects of different feeds on the metamorphosis and sex development of bullfrog tadpoles until the end of feeding

[0116]

[0117] As shown in Table 3, when the feeding end point was reached, the metamorphosis success rate of bullfrog tadpoles in groups 1 to 3 was above 90%, which was significantly higher than that of bullfrog tadpoles in groups 4 to 7, and the mortality rate and deformity rate were significantly lower than those of bullfrog tadpoles in groups 4 to 7; the male rate of bullfrogs in groups 2 to 3 was significantly higher than that of groups 4 to 7, and although the male rate of bullfrogs in group 1 was lower than that of group 6, it was still as high as more than 50%, which was significantly higher than that of groups 4 to 5 and 7; the thyroid hormone levels of bullfrogs in groups 1 to 3 were significantly higher than those in groups 4 to 7.

[0118] The above results show that the standard feed with added luteolin (Examples 1 to 3) can promote the successful metamorphosis of bullfrog tadpoles, upregulate thyroid hormone levels, and regulate sex differentiation, promoting more bullfrog tadpoles to develop into male frogs, compared with the conventional standard feed (Comparative Example 1) and the standard feed with added hormones (Comparative Examples 2 to 4).

[0119] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention rather than to limit the protection scope of the present invention. For ordinary technicians in this field, other different forms of changes or modifications can be made based on the above descriptions and ideas. It is not necessary and impossible to list all the implementation methods here. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in the protection scope of the claims of the present invention.

Claims

1. Application of luteolin in accelerating the metamorphosis of bullfrog tadpoles.

2. Application of luteolin in improving the success rate of metamorphosis of bullfrog tadpoles.

3. Application of luteolin in promoting male differentiation of bullfrog tadpoles.

4. Use of luteolin in any two of accelerating the metamorphosis of bullfrog tadpoles, improving the success rate of metamorphosis of bullfrog tadpoles, or promoting the male differentiation of bullfrog tadpoles.

5. Application of luteolin in accelerating the metamorphosis of bullfrog tadpoles, improving the success rate of metamorphosis of bullfrog tadpoles and promoting the male differentiation of bullfrog tadpoles.

6. The use according to any one of claims 1 to 5, It is characterized in that The content of the luteolin in the bullfrog feed or the bullfrog feed additive is 0.5%wt-2%wt.

7. Use of luteolin in the preparation of a bullfrog feed additive and / or a bullfrog feed having any one, two or three functions of accelerating the metamorphosis of bullfrog tadpoles, improving the success rate of metamorphosis of bullfrog tadpoles or promoting the male differentiation of bullfrog tadpoles.

8. A bullfrog feed additive or bullfrog feed, It is characterized in that Contains 0.5%wt-2%wt luteolin.

9. The bullfrog feed additive or bullfrog feed according to claim 8, It is characterized in that The bullfrog feed also contains protein feed, sugar and oil.

10. Use of the bullfrog feed additive or bullfrog feed according to claim 8 or 9 in any one, two or three of accelerating the metamorphosis of bullfrog tadpoles, improving the success rate of metamorphosis of bullfrog tadpoles or promoting the male differentiation of bullfrog tadpoles.

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