Ester of aminoglycan and uses thereof

Abstract

A new biomaterial is disclosed. The biomaterial is prepared by mixing glycosaminoglycan with a phosphoric acid, phosphate ester, or salt or derivative thereof under the action of a catalyst in a liquid reaction medium at a pH of 2.1 to 4.9. The obtained material is capable of enhancing the immunological resistance in human and in animals, treating diseases in human and in animals caused by viruses or bacteria, strengthening the anti-stress ability of animals, improving the appetite of animals, promoting the growth of animals, inhibiting the growth of tumor, lowering blood fat level, as well as preventing and treating viral diseases in plants.

Claims

1. An ester of aminoglycan, wherein the ester of aminoglycan comprises at least one of the following structural units: ##STR00007## where X is OH, OCH.sub.2CH.sub.2N(CH.sub.3).sub.3, OCH.sub.2CH.sub.2NH.sub.2, metal element or non-metal element independently; and Y is OH, ##STR00008## metal element or non-metal element independently.

2. A method of enhancing the immune resistance in human and animals, treating diseases in human and in animals caused by viruses or bacteria, strengthening the anti-stress ability of animals, improving the appetite of animals, promoting the growth of animals, inhibiting the growth of tumor, reducing blood lipid level and preventing and controlling viral diseases of plants comprising administering the ester of aminoglycan in claim 1 to a human, animal, or plant in need thereof.

3. The method according to claim 2, wherein the diseases in human or in animals comprise diseases harmful to human health, diseases caused by viruses, diseases caused by influenza virus, diseases caused by hepatitis A virus, diseases caused by hepatitis B virus, diseases caused by hepatitis C virus, diseases caused by HIV, diseases caused by herpes virus; cancerous diseases; diseases harmful to animals, viral and bacterial diseases harmful to livestock, poultry, aquatic product of fish, shrimp and crab, avian influenza, swine influenza, picornvirus disease, saprolegniasis, and streptococcusis.

Description

EMBODIMENTS

(1) The present invention makes further description based on the following examples. Those skilled in the art know that the following examples merely illustrate the present invention. On the premise of not departing from the spirit of the invention, any improvements and substitutions made to the invention are within the protection scope of the invention.

Example 1

(2) Preparing the preparation containing ester of aminoglycan of the present invention.

(3) The liquid fermentation medium is prepared with soluble starch 25 g, calcium carbonate 5 g, potassium dihydrogen phosphate 2 g and ammonium sulfate 5 g are mixed in 1000 mL water, then add 4.0 mL corn oil and mix well without adjusting the pH. The medium is sterilized at 121 C. for 20 min. The spores of purified Trichoderma harzianum (purchased from Chengdu Tepu Bio-Tech Co., Ltd.) are collected from a test tube containing sterile water using an inoculation loop. The spores are dispersed from hyphae and filtered through a sterile gauze to remove hyphae. After the spore concentration is adjusted to 10.sup.7 cfu/mL the spores are inoculated to the liquid fermentation medium in an inoculum size of 1%. After cultured at 28 C. and 150 r/min for 48 hours, 10 g sterilized deacetylated chitins (purchased from Sigma-Aldrich Company) and 5 g lecithin (purchased from Sigma-Aldrich Company) and 5 g of disodium hydrogen phosphate are added. The culture is further incubated at 28 C. and 150 r/min for 72 hours. The culture is filtered to collect the solution that is precipitated with 2-3 times of absolute ethanol. Carry out a suction filtration and the precipitates are diluted in water to make a microemulsion of ester of aminoglycan. The microemulsion is further adjusted to achieve the content of ester of aminoglycan at 0.01 mg ml. Upon detection, the microemulsion of ester of aminoglycan obtained in this example has an ester of aminoglycan which comprises the following structural unit,

(4) ##STR00005##
(wherein X is OCH.sub.2CH.sub.2N(CH.sub.3).sub.3, and Y is

(5) ##STR00006##
).

Example 2

(6) Enhanced immune resistance effect of the preparation containing ester of aminoglycan of the present invention.

(7) Macaca fascicularis (Yunnan Primate Laboratory Animal Co., Ltd.) are managed according to the country standard Feeding and Management and Operating Guideline for Experimental Animals. Fifteen Macaca fascicularis are divided into 3 groups with each group of 5 Macaca fascicularis. The group A is given sterile water as control. The group B is given the ester of aminoglycan obtained in Example 1 at a rate of 3 mg/3 ml/kg. The group C is given the ester of aminoglycan obtained in Example 1 at a rate of 9 mg/3 ml/kg. The administration to the animals is carried out at 10:00 am (30 min). The drug and dose given in each group are shown in the table below:

(8) TABLE-US-00001 TABLE 1 Arrangement of the Test Groups Group Treatment Dose (dose/water/body weight) Group A Sterile water 0 mg/3 ml/kg Group B Ester of aminoglycan 3 mg/3 ml/kg Group C Ester of aminoglycan 9 mg/3 ml/kg

(9) The detection in the antibody demonstrated that immunoglobulins IgG and IgM in blood increased significantly in Groups B and C that were orally given the ester of aminoglycan obtained in Example 1, especially, the ester of aminoglycan had a significant effect on increase of IgM. After 4 weeks under the administration, the level of IgM reached the maximum in both Group B and Group C that is 17% and 28% higher than that in Group A (control group), respectively.

(10) Immunocytokine assays also demonstrated that the new material ester of aminoglycan from the present invention significantly improved the level of cytokines IL10 and TNF-a in blood (Table 2). The increase of the level was related to the rate of the ester of aminoglycan concentration.

(11) TABLE-US-00002 TABLE 2 Effect of ester of aminoglycan on immunocytokines level IL-10 (pg/ml) TNF-a (pg/ml) Treatment The 9t.sup.h day The 15.sup.th day The 30.sup.th day The 9.sup.th day The 15.sup.th day The 30.sup.th day Group A 31.40 189.34 74.85 560.52 1057.46 592.20 Group B 145.61 181.16 118.08 250.63 1366.14 1037.07 Group C 146.55 805.80 536.43 830.60 1587.15 1730.95

Example 3

(12) Effect on treating swine viral illness by the preparation containing ester of aminoglycan of the present invention.

(13) Twenty pigs with symptom of hog cholera were selected in a hog farm. They were divided into treatment group by ester of aminoglycan and control group without treatment. Two groups of ill pigs was consistent in age, clinical manifestation and degree of illness. There were 10 pigs in treatment with the ester of aminoglycan and 10 in control respectively. The ester of aminoglycan was added in the feeding materials uniformly based on a dose of 3 mg per kg body weight in treatment. The control group is separated from the treatment and received the conventional management. The mental status, body temperature and food intake of the pigs were observed every day. Five days after the test started, the ill pigs fully recovered from body temperature and appetite and mental status and all clinic symptoms disappeared were recorded as cured. The ill pigs partially recovered from body temperature and appetite and mental status were recorded as improved. Both improvement and cure were regarded to be effective. Those with inactivity, anorexia or even death after 10 days from the test were recorded to be ineffective.

(14) TABLE-US-00003 TABLE 3 Effect of ester of aminoglycan on treating viral disease of swine Cured Improved Effective Ineffective Number Number Cured Number Improved Number Effective Number Ineffective Groups of cases of swine Rate % of swine Rate % of swine Rate % of swine Rate % Control group 10 0 0 1 10 1 10 9 90 Aminoglycan 10 8 80 7 20 10 100 0 0 ester group

(15) By adding ester of aminoglycan into the feeding materials, 8 ill pigs were cured, 2 ill pigs were improved. In the control group received the conventional treatment, 5 pigs died and only 1 pig was improved. This demonstrated that the ester of aminoglycan had a significant effect on the treatment of viral diseases in pigs. It was proved that the ester of aminoglycan from the present invention had a remarkable feature of simplicity in operation and excellent effect compared with the conventional management in which antibiotic drugs were injected.

Example 4

(16) The effect of the preparation containing ester of aminoglycan from the present invention on the prevention and control of plant virus diseases.

(17) The ester of aminoglycan prepared in Example 1 was used to prevent and control mosaic virus disease in Capsicum annum. Capsicum annuum which had viral symptom was selected for this test. The test were arranged as 5 treatments. Each treatment was designed with 4 blocks with 50 plants in each block. Treatment 1 was a dilution of the ester of aminoglycan at 1000 times. Treatment 2 was a dilution of the ester of aminoglycan at 500 times. Treatment 3 was a dilution of the ester of aminoglycan at 300 times. Treatment 4 was designed with 6% oligosaccharide-combined with protein at a dilution of 1000 times (Beijing Green Agricultural Science and Technology Group Co., Ltd.) as recommended. Treatment 5 was water control. The treatment was applied as leaf spray every 7 days. The incidence was investigated after three applications. The disease index was recorded to distinguish the effect of treatment as shown in Table 4.

(18) The classification of disease index:

(19) Level 0: symptom free in the whole plant;

(20) Level 1: Number of scabs is less than 2;

(21) Level 3: Number of scabs is 3-5;

(22) Level 5: Number of scabs is 6-8;

(23) Level 7: Number of scabs is more than 9;

(24) Level 9: The scabs are densely covered, and leaves are severely withered.
Disease index=(quantity of leaves at every levelrelative level value)/(total quantity of leaves9)100%
Control effect (%)=(disease index in controldisease index in treatment)/disease index in control100%.

(25) TABLE-US-00004 TABLE 4 Effect of ester of aminoglycan on prevention and control of virus diseases in Capsicum annuum Disease Control Treatment index effect (%) Ester of aminoglycan diluted 1000 times 3.65 81.16 Ester of aminoglycan diluted 500 times 2.25 88.38 Ester of aminoglycan diluted 300 times 1.74 91.02 oligosaccharides-protein diluted 1000 times 5.21 73.10 water control 19.37

(26) The water control plants were seriously infected by disease. The control effect by the ester of aminoglycan obtained in Example 1 was significant in control of the disease as the symptom of mosaic virus in young leaves significantly reduced. The effect was significantly improved when the ester of aminoglycan concentration was increased from a dilution of 1000 times to 500 or 300 times.

(27) Overall, it is clear from the results shown in the above tests that the preparation containing ester of aminoglycan in the present invention can effectively enhance the immune resistance in human and in animals, prevent and control illness in human and in animals that are caused by viruses or bacteria, and prevent plant diseases as well.