COMPOSITION FOR PROTECTING LIVER, PROTECTING INTESTINES AND ENHANCING IMMUNITY FOR FRESHWATER FISH, AND PREPARATION AND APPLICATION THEREOF

Abstract

A composition for protecting liver, protecting intestines and enhancing immunity for freshwater fish is disclosed, including the following raw materials in parts by weight: 15˜27 parts of Andrographis herba, 18˜22 parts of Isatidis folium, 10˜15 parts of Polygonum hydropiper, 8˜12 parts of Rheum palmatum L., and 8˜12 parts of Galla chinensis. And the preparation and the application of the composition are disclosed. The composition of the disclosure has the effect of resisting bacteria, scavenge free radicals, clearing heat, detoxifying, preventing and treating enteritis and the like, which can effectively improve the immunity of the freshwater fish and ensure the health of the organism.

Claims

1. A composition for protecting liver, protecting intestines and enhancing immunity for freshwater fish, comprising following raw materials in parts by weight: 15˜27 parts of Andrographis herba; 18˜22 parts of Isatidis folium; 10˜15 parts of Polygonum hydropiper; 8˜12 parts of Rheum palmatum L.; and 8˜12 parts of Galla chinensis.

2. The composition of claim 1, further comprising following raw materials in parts by weight: 7˜10 parts of taurine; and/or 15˜20 parts of diatomite.

3. A preparation method for the composition of claim 1, comprising: crushing each of the raw materials coarsely, sieving each of the crushed raw materials with a sieve of 80 mesh, pulverizing each of the sieved raw materials by an ultra-micro crusher with wall breaking at a low temperature, sieving each of the pulverized raw materials with a sieve of 300 mesh, and mixing all the raw materials well.

4. An application of the composition for protecting liver, protecting intestines and enhancing immunity for freshwater fish of claim 1 in preparation of a preparation for resisting bacteria and/or improving immunity and/or protecting liver and intestines.

5. An application of the composition for protecting liver, protecting intestines and enhancing immunity for freshwater fish of claim 1 in preparation of a functional feed.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0031] FIG. 1 shows the surviving crucian carp after challenge (using the compound Chinese herbal medicine composition for protecting liver, protecting intestines and enhancing immunity);

[0032] FIG. 2 shows the dead crucian carp after challenge (without using the compound Chinese herbal medicine composition for protecting liver, protecting intestines and enhancing immunity);

[0033] FIG. 3 shows the gill of the dead crucian carp after challenge;

[0034] FIG. 4 shows the abdomen of the dead crucian carp after challenge.

DETAILED DESCRIPTION OF EMBODIMENTS

[0035] Technical solutions of the present disclosure will be clearly and completely described below with reference to the embodiments. Obviously, the described embodiments are only part of the embodiments of the present disclosure, not all of them. Other embodiments made by those skilled in the art without sparing any creative effort should fall within the scope of the disclosure.

Embodiment 1

[0036] 20 parts by weight of Andrographis herba, 20 parts by weight of Isatidis folium, 10 parts by weight of Polygonum hydropiper, 10 parts by weight of Rheum palmatum L., 10 parts by weight of Galla chinensis, 10 parts by weight of taurine, and 20 parts by weight of diatomite were weighed. Each of the raw materials was crushed coarsely. Each of the crushed raw materials was pulverized by an ultra-micro crusher with wall breaking at −20° C. Each of the pulverized raw materials was sieved with a sieve of 300 mesh, and all the raw materials were mixed well to obtain a compound Chinese herbal medicine composition for protecting liver, protecting intestines and enhancing immunity.

Embodiment 2

[0037] Freshwater fish mainly rely on their non-specific immune system to protect themselves. The levels of lysozyme, immunoglobulin M, superoxide dismutase, malondialdehyde and total antioxidant capacity are directly related to the immune status of the body. The activities of aspartate aminotransferase and alanine aminotransferase can reflect the health status of liver. Intestinal health can be tested by the activities of digestive enzymes such as trypsin, lipase and amylase. Therefore, the embodiment evaluated the immunity and organism health of the crucian carp through the above indicators.

[0038] Growth test: nine crucian carp ponds (identical in size, density of crucian carp stocking, and mode of management) were randomly selected and divided into groups in Huangshagang area of Sheyang County, Jiangsu Province, China.

[0039] Control group A (fed with a basic commercial feed);

[0040] Control group B (supplemented with 0.5% Andrographis herba in the basic commercial feed);

[0041] Control group C (supplemented with 0.5% Isatidis folium in the basic commercial feed);

[0042] Control group D (supplemented with 0.5% Polygonum hydropiper in the basic commercial feed);

[0043] Control group E (supplemented with 0.5% Rheum palmatum L. in the basic commercial feed);

[0044] Control group F (supplemented with 0.5% Galla chinensis in the basic commercial feed);

[0045] Control group G (supplemented with 0.15% Andrographis herba, 0.1% Isatidis folium, 0.05% Polygonum hydropiper, 0.05% Rheum palmatum L., 0.05% Galla chinensis and 0.1% diatomite in the basic commercial feed);

[0046] Control group H (supplemented 0.15% Andrographis herba, 0.1% Isatidis folium, 0.05% Polygonum hydropiper, 0.05% Rheum palmatum L., 0.05% Galla chinensis and 0.1% taurine in the basic commercial feed);

[0047] Test group (supplemented with 0.5% of the compound Chinese herbal medicine composition for protecting liver, protecting intestines and enhancing immunity prepared in embodiment 1 in the basic commercial feed).

[0048] Each group was fed once a day in the morning, middle and evening (07:00, 12:00, 19:00). Each group was fed continuously for 10 days/month, and the rest of the time was fed with the basic commercial feed. After 4 months of domestication, the samples were collected and the indexes were tested in late July (10 fish were randomly taken from each pond). The activities of lysozyme (LZM), immunoglobulin M (IgM), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in serum, superoxide dismutase (SOD), malondialdehyde (MDA) and total antioxidant capacity (T-AOC) in liver, and trypsin (TPS), lipase (LPS) and amylase (AMS) in intestine were detected. The test results are shown in Table 1.

TABLE-US-00001 TABLE 1 Group Control Control Control Control Control Control Control Control Test Index group A group B group C group D group E group F group G group H group LZM 11.94 18.16 14.17 16.11 14.03 15.07 22.22 27.78 33.33 (U/ml) IgM 1.32 1.49 1.42 1.46 1.51 1.41 1.91 2.10 2.32 (mg/ml) ALT 4.87 4.32 4.41 4.5 4.46 4.62 3.14 3.16 2.83 (U/L) AST 27.62 20.76 23.74 22.8 23.88 23.5 19.39 20.62 18.02 (U/L) SOD 904.5 967.15 961.15 948.58 945.06 932.96 957.16 977.23 1058.29 (U/mgprot) MDA 21.71 14.27 14.17 12.69 16.52 16.17 12.72 13.25 11.09 (nmol/ mgprot) T-AOC 0.1 0.15 0.15 0.14 0.15 0.13 0.16 0.18 0.19 (mmol/ gprot) TPS 30017.43 34885.14 36154.93 38673.52 35504.68 35221.45 47104.29 45049.62 49158.97 (U/mgprot) LPS 1.63 1.92 2.06 2.16 1.89 2.22 2.55 2.64 3.10 (U/mgprot) AMS 72.71 86.14 86.81 89.51 86.32 89.51 91.15 95.5 108.77 (U/mgprot)

[0049] The results showed that: compared with the control groups A-H, the levels of lysozyme (LZM) in serum in the test group were increased by 179.15%, 83.54%, 135.22%, 106.89%, 137.56%, 121.17%, 50.00% and 19.98%, respectively; the content of immunoglobulin M (IgM) in serum in the test group increased by 75.76%, 55.70%, 63.38%, 58.90%, 53.64%, 64.54%, 21.47% and 10.48% respectively; the activity of alanine aminotransferase (ALT) in serum in the test group decreased by 41.89%, 34.49%, 35.83%, 37.11%, 36.55%, 38.74%, 9.87% and 10.44%, respectively; the activity of aspartate aminotransferase (AST) in the test group decreased by 34.76%, 13.20%, 24.09%, 20.96%, 24.54%, 23.32%, 7.07% and 12.61%, respectively; the activity of superoxide dismutase (SOD) in the liver of fish in the test group increased by 17.00%, 9.42%, 10.11%, 10.57%, 11.98%, 13.43%, 10.57% and 8.29%, respectively; the content of malondialdehyde (MDA) in the test group decreased by 48.92%, 22.28%, 22.28%, 12.81%, 32.87%, 31.42%, 12.81% and 16.30%, respectively; the total antioxidant capacity (T-AOC) in the liver of fish in the test group increased by 90.00%, 26.67%, 26.67%, 35.71%, 26.67%, 46.15%, 18.75% and 5.56%, respectively; the activity of intestinal trypsin (TPS) in the test group increased by 63.77%, 40.92%, 35.97%, 27.11%, 38.46%, 39.57%, 4.36% and 9.12%, respectively; the activity of intestinal lipase (LPS) in the test group increased by 90.18%, 61.46%, 50.49%, 43.52%, 64.02%, 39.64%, 21.57% and 17.42%, respectively; the activity of intestinal amylase (AMS) in the test group increased by 49.59%, 26.27%, 25.30%, 21.52%, 26.01%, 21.52%, 19.33% and 13.90%, respectively. Therefore, compared with the control groups, the compound Chinese herbal medicine composition for protecting liver, protecting intestines and enhancing immunity with seven components can improve the immunity and antioxidant capacity of freshwater fish, and promote the health of liver and intestine.

Embodiment 3

[0050] Challenge test: after the 4-month growth test in embodiment 2, all groups of crucian carp were fasted for 24 hours, and the crucian carp with the same specifications were selected from each pond for Aeromonas hydrophila infection test.

[0051] The challenge test was carried out in a plastic box (1.0 m×0.4 m×0.4 m), with three replicates in each group and 30 fish in each replicate. Aeromonas hydrophila was provided by the fish disease laboratory of Jiangsu Freshwater Fisheries Research Institute. The final concentration of Aeromonas hydrophila was about 5×107 cells/ml diluted with sterile normal saline. The physiological activities of fish were observed after intraperitoneal injection of 1.0 ml bacterial solution per 100 g fish. The cumulative mortality of nine groups of fish at 0 h, 12 h, 24 h, 48 h and 96 h after challenge was counted, respectively. The results are shown in Table 2.

TABLE-US-00002 TABLE 2 Cumulative mortality (%) Group 0 h 12 h 24 h 48 h 96 h Control group A 0.00 10.00 21.11 41.11 61.11 Control group B 0.00 3.33 6.67 17.78 38.89 Control group C 0.00 6.67 10.00 23.33 48.89 Control group D 0.00 3.33 7.78 17.78 44.44 Control group E 0.00 5.56 13.33 22.22 46.67 Control group F 0.00 4.44 12.22 18.89 45.56 Control group G 0.00 2.22 5.56 14.44 31.11 Control group H 0.00 2.22 3.33 13.33 27.78 Test Group 0.00 1.11 2.22 7.78 18.89

[0052] The results showed that after the crucian carp was infected with Aeromonas hydrophila, the viability began to decrease, and the abdomen and gills of the dead crucian carp had bleeding symptoms in various degrees (the comparison of symptoms is shown in FIG. 1-4). Compared to the control groups A-H, the mortality of test fish 12 h after challenge was reduced by 88.90%, 66.70%, 83.35%, 66.70%, 80.02%, 75.03%, 50.05% and 50.05%, respectively; the mortality of test fish 24 h after challenge was reduced by 89.48%, 66.70%, 77.80%, 71.46%, 83.35%, 81.84%, 60.04% and 33.40%, respectively; the mortality of test fish 48 h after challenge was reduced by 81.08%, 56.24%, 6%, 6.66%, 56.24%, 64.99%, 58.81%, 46.14% and 41.65%, respectively; the mortality of test fish 96 h after challenge was reduced by 69.09%, 51.43%, 61.36%, 57.50%, 59.52%, 58.53%, 39.28% and 22.00%, respectively. Therefore, the compound Chinese herbal medicine composition for protecting liver, protecting intestines and enhancing immunity with seven components can improve freshwater fish's ability to resist Aeromonas hydrophila infection and reduce death caused by pathogenic bacteria.

Embodiment 4

[0053] 15 parts by weight of Andrographis herba, 18 parts by weight of Isatidis folium, 15 parts by weight of Polygonum hydropiper, 12 parts by weight of Rheum palmatum L., 12 parts by weight of Galla chinensis, 10 parts by weight of taurine, and 18 parts by weight of diatomite were weighed. Each of the raw materials was crushed coarsely. Each of the crushed raw materials was pulverized by an ultra-micro crusher with wall breaking at −20° C. Each of the pulverized raw materials was sieved with a sieve of 300 mesh, and all the raw materials were mixed well to obtain a compound Chinese herbal medicine composition for protecting liver, protecting intestines and enhancing immunity.

Embodiment 5

[0054] 25 parts by weight of Andrographis herba, 20 parts by weight of Isatidis folium, 10 parts by weight of Polygonum hydropiper, 8 parts by weight of Rheum palmatum L., 10 parts by weight of Galla chinensis, 8 parts by weight of taurine, and 19 parts by weight of diatomite were weighed. Each of the raw materials was crushed coarsely. Each of the crushed raw materials was pulverized by an ultra-micro crusher with wall breaking at −20° C. Each of the pulverized raw materials was sieved with a sieve of 300 mesh, and all the raw materials were mixed well to obtain a compound Chinese herbal medicine composition for protecting liver, protecting intestines and enhancing immunity.

Embodiment 6

[0055] 23 parts by weight of Andrographis herba, 22 parts by weight of Isatidis folium, 14 parts by weight of Polygonum hydropiper, 9 parts by weight of Rheum palmatum L., 8 parts by weight of Galla chinensis, 9 parts by weight of taurine, and 15 parts by weight of diatomite were weighed. Each of the raw materials was crushed coarsely. Each of the crushed raw materials was pulverized by an ultra-micro crusher with wall breaking at −20° C. Each of the pulverized raw materials was sieved with a sieve of 300 mesh, and all the raw materials were mixed well to obtain a compound Chinese herbal medicine composition for protecting liver, protecting intestines and enhancing immunity.

Embodiment 7

[0056] 21 parts by weight of Andrographis herba, 21 parts by weight of Isatidis folium, 12 parts by weight of Polygonum hydropiper, 11 parts by weight of Rheum palmatum L., 11 parts by weight of Galla chinensis, 7 parts by weight of taurine, and 17 parts by weight of diatomite were weighed. Each of the raw materials was crushed coarsely. Each of the crushed raw materials was pulverized by an ultra-micro crusher with wall breaking at −15° C. Each of the pulverized raw materials was sieved with a sieve of 300 mesh, and all the raw materials were mixed well to obtain a compound Chinese herbal medicine composition for protecting liver, protecting intestines and enhancing immunity.

Embodiment 8

[0057] 19 parts by weight of Andrographis herba, 19 parts by weight of Isatidis folium, 13 parts by weight of Polygonum hydropiper, 12 parts by weight of Rheum palmatum L., 12 parts by weight of Galla chinensis, 9 parts by weight of taurine, and 16 parts by weight of diatomite were weighed. Each of the raw materials was crushed coarsely. Each of the crushed raw materials was pulverized by an ultra-micro crusher with wall breaking at −15° C. Each of the pulverized raw materials was sieved with a sieve of 300 mesh, and all the raw materials were mixed well to obtain a compound Chinese herbal medicine composition for protecting liver, protecting intestines and enhancing immunity.

Embodiment 9

[0058] Following the method of the growth test in embodiment 2 and the challenge test in embodiment 3, the physiological and biochemical indexes and the cumulative mortality (only 24 h and 48 h after challenge) of crucian carp fed with the compound Chinese herbal medicine composition for protecting liver, protecting intestines and enhancing immunity prepared in embodiments 4-9 were determined, respectively. The control group was fed with the basic commercial feed, and each test group was fed with the basic commercial feed added with 0.5% of the compound Chinese herbal medicine composition for protecting liver, protecting intestines and enhancing immunity prepared in embodiments 4-9, respectively (test group A, test group B, test group C, test group D, and test group E, respectively). The results are presented in Tables 3 and Tables 4.

TABLE-US-00003 TABLES 3 Group Control Test Test Test Test Test Index group group A group B group C group D group E LZM 11.94 29.17 37.55 36.21 32.76 30.55 (U/ml) IgM 1.32 2.25 2.75 2.54 2.16 2.37 (mg/ml) ALT 4.87 2.85 2.56 2.74 2.95 2.93 (U/L) AST 27.62 20.10 19.47 20.02 19.25 19.72 (U/L) SOD 904.50 992.48 1104.78 1099.45 988.45 1033.51 (U/mgprot) MDA 21.71 13.46 10.95 12.44 12.60 13.68 (nmol/mgprot) T-AOC 0.10 0.18 0.19 0.19 0.18 0.17 (nmol/gprot) TPS 30017.43 49186.06 48457.12 49020.45 49798.34 50280.21 (U/mgprot) LPS 1.63 3.05 2.84 2.78 2.94 3.35 (U/mgprot) AMS 72.71 107.51 100.24 98.24 100.84 109.39 (U/mgprot)

TABLE-US-00004 TABLES 4 Cumulative mortality (%) Group 24 h 48 h Control group 21.11 41.11 Test group A 6.67 8.89 Test group B 3.33 5.56 Test group C 5.56 7.78 Test group D 6.67 8.89 Test group E 5.56 7.78

[0059] The general principle as defined herein may be achieved in other embodiments without departing from the spirit or scope of the disclosure. The present disclosure will therefore not be restricted to these embodiments shown herein, but rather to comply with the broadest scope consistent with the principles and novel features disclosed herein.