Traditional Chinese medicine composition capable of relieving diarrhea caused by cryptosporidium parvum as well as preparation method and application of traditional Chinese medicine composition

Abstract

The invention discloses a traditional Chinese medicine composition capable of relieving diarrhea caused by cryptosporidium parvum as well as a preparation method and application of the traditional Chinese medicine composition, and belongs to the technical field of medicines. The traditional Chinese medicine composition consists of a prescription for clearing intestines and resolving dampness and allicin, the prescription for clearing intestines and resolving dampness is composed of scutellaria baicalensis, coptis chinensis, roasted radix aucklandiae, fried angelica sinensis, sanguisorba officinalis, parched white peony root, angelica dahurica and liquorice. The traditional Chinese medicine composition is prepared from the following raw materials in parts by weight: 10-15 parts of radix scutellariae, 6-10 parts of rhizoma coptidis, 6-10 parts of roasted radix aucklandiae, 10-15 parts of fried radix angelicae sinensis, 10-15 parts of radix sanguisorbae, 15-20 parts of fried radix paeoniae alba, 10-20 parts of radix angelicae, 6-10 parts of liquorice root and 0.02-0.07 part of allicin. The traditional Chinese medicine composition prepared by the invention is efficient, low in toxicity, safe and harmless, and can relieve diarrhea caused by cryptosporidium parvum, relieve intestinal injury, regulate the composition structure of intestinal flora and enhance intestinal barrier and immunity.

Description

Technical Field

[0001] The invention belongs to the technical field of medicines, and in particular relates to a traditional Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum, and a preparation method and application thereof. Background Art

[0002] Cryptosporidium parvum is a parasite that is ubiquitous worldwide and can infect a wide range of vertebrates, including humans, livestock, pets, and wild animals, causing cryptosporidiosis. The main symptom of the disease is severe diarrhea, which not only seriously impairs the host's health, but can even be fatal to individuals with compromised immune systems. The main route of transmission of Cryptosporidium parvum is the fecal-oral route, which means that it is mainly transmitted through water and food contaminated by the pathogen, but can also spread through direct contact between people or between people and animals. When animals are infected with Cryptosporidium parvum, they can not only directly transmit the pathogen to humans or other animals, but their excreta can also become a new source of infection if it contaminates water or food, leading to the widespread spread of this parasite between humans and animals and the environment. The World Health Organization (WHO) has listed Cryptosporidium parvum as one of the six common pathogens that cause infectious diarrhea, further highlighting its public health significance.

[0003] At present, there is still a lack of specific vaccines and highly effective therapeutic drugs for Cryptosporidiosis parvum. Nitazoxanide (NTZ), the only drug approved by the U.S. Food and Drug Administration (FDA) for the treatment of the disease, is not effective in newborns and immunocompromised patients. Traditional treatments such as chemical drugs, antibiotics and certain target inhibitors can alleviate the condition to a certain extent, but long-term or improper use of these drugs can easily lead to drug residues, increased drug resistance and toxic side effects, making the treatment effect unstable and prone to recurrence. Chinese herbal medicines have unique advantages in anti-inflammatory, immunomodulatory, anti-parasitic and antioxidant aspects, and they also have good efficacy in preventing and treating parasitic diseases; Chinese herbal medicines may not only play a comprehensive role through their multiple active ingredients to enhance the body's immune response, but also reduce the risk of drug side effects because they are not easy to develop drug resistance.

[0004] Therefore, developing a highly effective, safe, low-toxic and drug-resistant Chinese medicine composition for alleviating diseases such as diarrhea caused by Cryptosporidium parvum has become one of the problems that technicians in this field urgently need to solve. Summary of the invention

[0005] In view of the above-mentioned deficiencies in the prior art, the present invention discloses a Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum, and a preparation method and application thereof. The Chinese medicine composition prepared by the present invention is highly efficient, low in toxicity, safe and harmless, and can relieve diarrhea caused by Cryptosporidium parvum, relieve intestinal damage, regulate the composition structure of intestinal flora, and enhance intestinal barrier and immunity.

[0006] To achieve the above object, the present invention provides the following technical solutions:

[0007] The invention discloses a Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum, which is composed of an intestinal clearing and dampness-removing prescription and allicin; the intestinal clearing and dampness-removing prescription is composed of scutellaria baicalensis, coptis chinensis, stewed costusroot, stir-fried angelica sinensis, sanguisorba officinalis, stir-fried white peony root, angelica dahurica and liquorice; the Chinese medicine composition is prepared from the following raw materials in parts by weight: 10-15 parts of scutellaria baicalensis, 6-10 parts of coptis chinensis, 6-10 parts of stewed costusroot, 10-15 parts of stir-fried angelica sinensis, 10-15 parts of sanguisorba officinalis, 15-20 parts of stir-fried white peony root, 10-20 parts of angelica dahurica, 6-10 parts of liquorice and 0.02-0.07 parts of allicin.

[0008] Preferably, the Chinese medicine composition is made of the following raw materials in parts by weight: 10 parts of Scutellaria baicalensis, 6 parts of Coptis chinensis, 6 parts of stewed Aucklandia lappa, 10 parts of stir-fried Angelica sinensis, 10 parts of Sanguisorba officinalis, 15 parts of stir-fried White Peony Root, 10 parts of Angelica dahurica, 6 parts of Licorice, and 0.05 parts of Allicin.

[0009] The present invention also provides a method for preparing the above-mentioned Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum, comprising the following steps:

[0010] ① Weigh scutellaria baicalensis, coptis root, stewed costusroot, stir-fried angelica, sanguisorba officinalis, stir-fried white peony root, angelica dahurica, liquorice and allicin according to the weight ratio for later use;

[0011] ② Add the scutellaria baicalensis, coptis chinensis, stewed costusroot, fried angelica sinensis, sanguisorba officinalis, fried white peony root, angelica dahurica and liquorice weighed in step ① into water, soak at room temperature for 30 minutes, then decoct at 100° C. for 60 to 70 minutes, filter and obtain a liquid medicine for clearing intestines and removing dampness;

[0012] ③ After heating the intestinal clearing and dehumidifying prescription liquid prepared in step ② at 100° C. to 1 / 3 of the original volume, placing it in a refrigerator at -20° C. for 48 hours, and then placing it in a freeze dryer at -80° C. for 48 hours to remove moisture, to obtain the intestinal clearing and dehumidifying prescription freeze-dried powder, which is stored for later use;

[0013] ④ According to the weight ratio, the lyophilized powder of the formula for clearing intestines and removing dampness prepared in step ③ is evenly mixed with allicin to prepare a Chinese medicine composition.

[0014] The invention also discloses the use of the Chinese medicinal composition capable of relieving diarrhea caused by Cryptosporidium parvum in preparing drugs for resisting Cryptosporidium parvum and treating Cryptosporidium parvum diarrhea.

[0015] Preferably, the anti-Cryptosporidium parvum drug refers to one or more products directly used for the prevention, diagnosis, detection, protection, treatment and research of Cryptosporidium parvum and its related diseases.

[0016] Preferably, the drug for treating Cryptosporidium parvum diarrhea refers to one or more products directly used for preventing, diagnosing, detecting, protecting, treating and studying Cryptosporidium parvum diarrhea.

[0017] Compared with the prior art, the present invention has the following beneficial effects:

[0018] The invention provides a Chinese medicine composition (intestinal clearing and dampness-removing prescription + allicin) capable of relieving diarrhea caused by Cryptosporidium parvum. The toxicity of the Chinese medicine composition is lower than that of the existing commonly used potential anthelmintics, and experiments show that the Chinese medicine composition is safe and harmless to mice, and has a better inhibitory effect on Cryptosporidium parvum. After the animals infected with Cryptosporidium parvum use the Chinese medicine composition, the content of worm eggs in animal feces is significantly reduced, the composition structure of intestinal flora is regulated, the intestinal damage caused by the infection is alleviated, the expression amount of genes related to the barrier and immunity of the intestine (ZO-1, Occludin, Caspase-1, Claudin-1) is upregulated, the tight connection of the intestinal barrier of mice is enhanced, the intestinal immunity of mice is improved, and the diarrhea symptoms of model animal mice are alleviated, thereby providing a new reference for relieving diarrhea caused by Cryptosporidium parvum infection. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 It is an analysis chart of the expression levels of five intestinal barrier genes by RT-qPCR (in the figure: C is the control group, M is the infection and no treatment group, P is the infection + paromomycin treatment group, A is the infection + allicin treatment group, F is the infection + Qingchang Huashi prescription treatment group, AF is the infection + the Chinese medicine composition of the present invention treatment group; among them: a is Caspase-1, b is Claudin-1, c is NLRP3, d is Occludin, and e is ZO-1);

[0020] Figure 2 After mice were infected with Cryptosporidium parvum, their feces were collected for 8 consecutive days and the number of parasite oocysts shed in their feces was analyzed;

[0021] Figure 3It is a HE-stained section of mouse intestine (in the figure: a is a section, b is the analysis results of mouse intestinal villi and intestinal crypt depth and ratio, where: b1 is the length of intestinal villi, b2 is the depth of intestinal crypt, and b3 is the ratio of intestinal villi length / intestinal crypt depth);

[0022] Figure 4 The effect of different drugs on the intestinal flora of mice infected with Cryptosporidium parvum (where a is phylum, b is class, c is order, d is family, and e is genus);

[0023] Figure 5 To investigate the effects of different drugs on the α-diversity index of intestinal flora in mice infected with Cryptosporidium parvum.

[0024] Figure 6 The beta diversity analysis diagram of intestinal flora in mice infected with Cryptosporidium parvum and treated with different drugs (a is PCoA analysis, b is NMDS analysis);

[0025] Figure 7 This is a petal diagram of the intestinal flora of mice infected with Cryptosporidium parvum and treated with different drugs;

[0026] Figure 8 This is a LEfSe analysis diagram of the intestinal flora of mice infected with Cryptosporidium parvum and treated with different drugs (where a is the species evolution diagram of the marker flora, and b is the bar graph of the marker flora). DETAILED DESCRIPTION

[0027] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments in the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention. The experimental methods in the following embodiments are all conventional methods unless otherwise specified; the experimental materials used are all commercially available unless otherwise specified.

[0028] A traditional Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum is composed of a formula for clearing intestines and removing dampness and allicin; wherein the formula for clearing intestines and removing dampness is composed of scutellaria baicalensis, coptis chinensis, stewed costusroot, stir-fried angelica sinensis, sanguisorba officinalis, stir-fried white peony root, angelica dahurica and liquorice.

[0029] The above-mentioned Chinese medicine composition is made of the following raw materials in parts by weight: 10-15 parts of scutellaria baicalensis, 6-10 parts of coptis chinensis, 6-10 parts of stewed costusroot, 10-15 parts of stir-fried angelica sinensis, 10-15 parts of sanguisorba officinalis, 15-20 parts of stir-fried white peony root, 10-20 parts of angelica dahurica, 6-10 parts of licorice and 0.02-0.07 parts of allicin.

[0030] The most preferred ratio of the raw materials constituting the above-mentioned traditional Chinese medicine composition is: 10 parts of Scutellaria baicalensis, 6 parts of Coptis chinensis, 6 parts of stewed Aucklandia lappa, 10 parts of stir-fried Angelica sinensis, 10 parts of Sanguisorba officinalis, 15 parts of stir-fried White Peony Root, 10 parts of Angelica dahurica, 6 parts of Licorice, and 0.05 parts of Allicin.

[0031] The preparation method of the above-mentioned Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum is as follows:

[0032] ① Weigh out scutellaria baicalensis, coptis chinensis, stewed costusroot, stir-fried angelica, sanguisorba officinalis, stir-fried white peony root, angelica dahurica, liquorice and allicin according to the weight ratio and set aside.

[0033] ② Add the scutellaria baicalensis, coptis chinensis, stewed costusroot, fried angelica, sanguisorba officinalis, fried white peony root, angelica dahurica and liquorice weighed in step ① into water and soak at room temperature for 30 minutes, then boil at 100°C for 60-70 minutes, filter and obtain the intestinal clearing and dampness-removing prescription liquid.

[0034] ③ After heating the intestinal clearing and dehumidifying prescription liquid prepared in step ② at 100°C to 1 / 3 of the original volume, place it in a -20°C refrigerator and freeze it for 48 hours, then place it in a freeze dryer and freeze-dry it at -80°C for 48 hours to remove moisture, to obtain the intestinal clearing and dehumidifying prescription freeze-dried powder, which is stored for later use.

[0035] ④ According to the weight ratio, the lyophilized powder of the formula for clearing intestines and removing dampness prepared in step ③ is evenly mixed with allicin to prepare a Chinese medicine composition.

[0036] The Chinese medicine composition is used in preparing drugs for resisting Cryptosporidium parvum and treating Cryptosporidium parvum diarrhea.

[0037] Among them, the above-mentioned anti-Cryptosporidium parvum drugs refer to one or more products directly used for the prevention, diagnosis, detection, protection, treatment and research of Cryptosporidium parvum and its related diseases; drugs for treating Cryptosporidium parvum diarrhea refer to one or more products directly used for the prevention, diagnosis, detection, protection, treatment and research of Cryptosporidium parvum diarrhea.

[0038] The technical solution of the present invention is further described below in conjunction with specific embodiments and drawings:

[0039] Example 1

[0040] The preparation method of the Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum in this embodiment comprises the following specific steps:

[0041] ① According to the weight ratio, weigh 10 parts of Scutellaria baicalensis, 6 parts of Coptis chinensis, 6 parts of stewed costusroot, 10 parts of stir-fried angelica, 10 parts of Sanguisorba officinalis, 15 parts of stir-fried white peony root, 10 parts of Angelica dahurica, 6 parts of liquorice, and 0.05 parts of allicin for later use.

[0042] ② Add the scutellaria baicalensis, coptis chinensis, stewed costusroot, fried angelica, sanguisorba officinalis, fried white peony root, angelica dahurica and liquorice weighed in step ① into water and soak at room temperature for 30 minutes, then boil at 100°C for 60 minutes, filter and obtain the intestinal clearing and dampness-removing prescription liquid.

[0043] ③ After heating the intestinal clearing and dehumidifying prescription liquid prepared in step ② at 100°C to 1 / 3 of the original volume, place it in a -20°C refrigerator and freeze it for 48 hours, then place it in a freeze dryer and freeze-dry it at -80°C for 48 hours to remove moisture, to obtain the intestinal clearing and dehumidifying prescription freeze-dried powder, which is stored for later use.

[0044] ④ According to the weight ratio, the lyophilized powder of the formula for clearing intestines and removing dampness prepared in step ③ is evenly mixed with allicin to prepare a Chinese medicine composition, that is, a Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum.

[0045] Example 2

[0046] The preparation method of the Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum in this embodiment comprises the following specific steps:

[0047] ① According to the weight ratio, weigh 12 parts of Scutellaria baicalensis, 8 parts of Coptis chinensis, 8 parts of stewed costusroot, 12 parts of stir-fried angelica, 12 parts of Sanguisorba officinalis, 17 parts of stir-fried white peony root, 18 parts of Angelica dahurica, 8 parts of liquorice, and 0.05 parts of allicin for later use.

[0048] ② Add the scutellaria baicalensis, coptis chinensis, stewed costusroot, fried angelica, sanguisorba officinalis, fried white peony root, angelica dahurica and liquorice weighed in step ① into water and soak at room temperature for 30 minutes, then boil at 100°C for 70 minutes, filter and obtain the intestinal clearing and dampness-removing prescription liquid.

[0049] ③ After heating the intestinal clearing and dehumidifying prescription liquid prepared in step ② at 100°C to 1 / 3 of the original volume, place it in a -20°C refrigerator and freeze it for 48 hours, then place it in a freeze dryer and freeze-dry it at -80°C for 48 hours to remove moisture, to obtain the intestinal clearing and dehumidifying prescription freeze-dried powder, which is stored for later use.

[0050] ④ According to the weight ratio, the lyophilized powder of the formula for clearing intestines and removing dampness prepared in step ③ is evenly mixed with allicin to prepare a Chinese medicine composition, that is, a Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum.

[0051] Example 3

[0052] The preparation method of the Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum in this embodiment comprises the following specific steps:

[0053] ① According to the weight ratio, weigh 15 parts of Scutellaria baicalensis, 10 parts of Coptis chinensis, 10 parts of stewed costusroot, 15 parts of stir-fried angelica, 15 parts of Sanguisorba officinalis, 20 parts of stir-fried white peony root, 20 parts of Angelica dahurica, 10 parts of liquorice, and 0.07 parts of allicin for later use.

[0054] ② Add the scutellaria baicalensis, coptis chinensis, stewed costusroot, fried angelica, sanguisorba officinalis, fried white peony root, angelica dahurica and liquorice weighed in step ① into water and soak at room temperature for 30 minutes, then boil at 100°C for 65 minutes, filter and obtain the intestinal clearing and dampness-removing prescription liquid.

[0055] ③ After heating the intestinal clearing and dehumidifying prescription liquid prepared in step ② at 100°C to 1 / 3 of the original volume, place it in a -20°C refrigerator and freeze it for 48 hours, then place it in a freeze dryer and freeze-dry it at -80°C for 48 hours to remove moisture, to obtain the intestinal clearing and dehumidifying prescription freeze-dried powder, which is stored for later use.

[0056] ④ According to the weight ratio, the lyophilized powder of the formula for clearing intestines and removing dampness prepared in step ③ is evenly mixed with allicin to prepare a Chinese medicine composition, that is, a Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum.

[0057] Example 4

[0058] The preparation method of the Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum in this embodiment comprises the following specific steps:

[0059] ① According to the weight ratio, weigh 10 parts of Scutellaria baicalensis, 6 parts of Coptis chinensis, 6 parts of stewed costusroot, 10 parts of stir-fried angelica, 10 parts of Sanguisorba officinalis, 15 parts of stir-fried white peony root, 10 parts of Angelica dahurica, 6 parts of liquorice, and 0.02 parts of allicin for later use.

[0060] ② Add the scutellaria baicalensis, coptis chinensis, stewed costusroot, fried angelica, sanguisorba officinalis, fried white peony root, angelica dahurica and liquorice weighed in step ① into water and soak at room temperature for 30 minutes, then boil at 100°C for 70 minutes, filter and obtain the intestinal clearing and dampness-removing prescription liquid.

[0061] ③ After heating the intestinal clearing and dehumidifying prescription liquid prepared in step ② at 100°C to 1 / 3 of the original volume, place it in a -20°C refrigerator and freeze it for 48 hours, then place it in a freeze dryer and freeze-dry it at -80°C for 48 hours to remove moisture, to obtain the intestinal clearing and dehumidifying prescription freeze-dried powder, which is stored for later use.

[0062] ④ According to the weight ratio, the lyophilized powder of the formula for clearing intestines and removing dampness prepared in step ③ is evenly mixed with allicin to prepare a Chinese medicine composition, that is, a Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum.

[0063] Example 5

[0064] This example is a test of the effect of the Chinese medicine composition prepared in the above Example 1 on the intestinal barrier and fecal oocyst count of mice.

[0065] 60 ICR mice were selected to study the inhibitory effect of the derivatives on C.parvum. The experimental mice were randomly divided into 6 groups, each with 10 mice, namely group C (control group without infection), group M (infection without treatment), group P (infection + paromomycin treatment group), group A (infection + allicin treatment group), group F (infection + Qingchang Huashi prescription treatment group), and group AF (Chinese medicine composition treatment group prepared in Example 1). The insecticidal effect of C.parvum was studied for 7 consecutive days. 5 mg / L dexamethasone sodium phosphate injection was added to the drinking water of the mice every day to suppress the immune function of the mice. On the first day of the experiment, 4000 C.parvum oocysts were gavaged to each mouse in each group; treatment began on the second day and continued for 7 days. The feces of each mouse in each group were collected every day, and an equal amount of penicillin-streptomycin-amphotericin solution was added and stored at 4°C. After the experiment, all mice were euthanized, the small intestine was collected, and it was immersed and stored in 4% formalin solution. Subsequently, HE staining analysis and RT-qPCR were used to quantify the oocysts in the feces, and five genes related to the intestinal barrier, namely Caspase-1, Claudin-1, NLRP3, Occludin, and ZO-1, were quantitatively analyzed to evaluate the therapeutic effect of each group in the experiment on mice infected with Cryptosporidium parvum. The specific analysis is as follows.

[0066] Figure 1 This is the analysis of the expression of five intestinal barrier genes by RT-qPCR. Figure 1 It can be seen that the analysis of Caspase-1, Claudin-1, NLRP3, Occludin and ZO-1, five genes related to the intestinal barrier, by RT-qPCR showed that after infection with Cryptosporidium parvum, the expression levels of Caspase-1, Claudin-1, Occludin and ZO-1 in the mice of the M group were downregulated compared with those in the control group C group; after treatment with paromomycin, allicin, Qingchang Huashi prescription and the Chinese medicine composition prepared in Example 1, the corresponding P group, A group, F group and AF group The expression levels of Caspase-1, Claudin-1, Occludin and ZO-1 in mice were significantly higher than those in group M, and the treatment effect of group AF was the most prominent. After infection with Cryptosporidium parvum, the expression level of NLRP3 gene in group M was higher than that in group C. After treatment with paromomycin, allicin, Qingchang Huashi recipe and the Chinese medicine composition prepared in Example 1, the expression levels of NLRP3 gene in mice in the corresponding groups P, A, F and AF were significantly downregulated compared with those in group M, and the treatment effect of group AF was the most prominent.

[0067] Figure 2 After mice were infected with Cryptosporidium parvum, the feces of the mice were collected for 8 consecutive days, and the number of parasite oocysts shed in the feces was shown in the figure. Figure 2 It can be seen that compared with the mice in group M, the number of C.parvum oocysts in the feces of mice in groups P, A, F and AF could be reduced after treatment; among them, the effect of treatment in group AF was the most prominent, and the number of C.parvum oocysts in the feces of mice was reduced by about 2300 times.

[0068] Figure 3 This is a HE-stained section of mouse intestine. Figure 3 It can be seen that: by HE staining of the intestines of the above-mentioned groups of mice, the results showed that the mice in group C (uninfected group) maintained the integrity of the intestinal mucosa and intestinal villi, the mice in group M (infected but not treated) showed pathological changes characterized by villous atrophy, shedding and mucosal erosion, while the mice in groups P, A, F and AF that were treated maintained the complete intestinal mucosa and intestinal villus structure, and its integrity was similar to that of the mice treated with paromomycin.

[0069] This shows that after the mice infected with Cryptosporidium parvum were treated with the Chinese medicine composition prepared in Example 1 of the present invention, not only the interaction between different cell proteins was improved, but also the integrity of the mouse intestinal structure was promoted, the tight connection of the mouse intestinal barrier was enhanced, and the intestinal immune ability of the mouse was improved.

[0070] Example 6

[0071] This example is an experiment on the effect of the Chinese medicine composition prepared in the above Example 1 on the intestinal flora of mice.

[0072] 60 ICR mice were selected to study the inhibitory effect of the derivatives on C.parvum. The experimental mice were randomly divided into 6 groups, each with 10 mice, namely group C (control group uninfected group), group M (infected untreated group), group P (infected + paromomycin treatment group), group A (infected + allicin treatment group), group F (infected + Qingchang Huashi prescription treatment group), and group AF (Chinese medicine composition treatment group prepared in Example 1). The insecticidal effect of C.parvum was studied for 7 consecutive days. 5 mg / L dexamethasone sodium phosphate injection was added to the drinking water of the mice every day to suppress the immune function of the mice. On the first day of the experiment, 4000 oocysts of C.parvum were gavaged to each mouse in each group; treatment began on the second day and continued for 7 days. After the experiment, all mice were euthanized, the small intestine of the mice was collected for 16SrRNA high-throughput sequencing, and the effect on the intestinal flora of the mice was analyzed to evaluate the therapeutic effect of each group on mice infected with Cryptosporidium parvum in the experiment. The specific analysis is as follows.

[0073] Figure 4 This is a diagram showing the effect of different drugs on the intestinal flora of mice infected with Cryptosporidium parvum (where a is phylum, b is class, c is order, d is family, and e is genus). Figure 4 It can be seen that the intestinal flora of mice differed at the phylum, class, order, family, genus, and species levels; at the phylum level, the relative abundance of Clostridia (Clostridia_258483) and Campylobacteria in the intestines of group M increased after infection with Cryptosporidium parvum, while the relative abundance of Clostridia (Clostridia_258483) and Campylobacteria in the intestines of groups P, A, F, and AF after treatment decreased (e.g. Figure 4 a); From the genus level, the relative abundance of Helicobacter pylori (Helicobacter_C_479931) in the intestine of mice in group M increased significantly after infection with Cryptosporidium parvum, while the relative abundance of Helicobacter pylori (Helicobacter_C_479931) and Bacteroides (Bacteroides_H) in the intestine of mice in groups P, A, F, and AF after treatment decreased significantly (e.g. Figure 4 e); among them, the relative abundance of Bacteroidota and Aloprevotela in group P increased significantly, the relative abundance of Campylobacterota and Helicobacter in group A increased significantly, and the relative abundance of Firmicutes in group F increased significantly.

[0074] Figure 5 The effect of different drugs on the α-diversity index of intestinal flora in mice infected with Cryptosporidium parvum. Figure 5 It can be seen that there were no significant differences in Chao 1 index, Faith_pd index, Goods_coverage index and Observed_cpecies index among groups C, M, P, A, F and AF (p>0.05), but there were significant differences in Pielou_e index (p=0.0018), Shannon index (p=0.006) and Simpson index (p=0.015) among the groups, indicating that after treatment with groups P, A, F and AF, the richness, uniformity and diversity of the intestinal flora of mice were increased.

[0075] Figure 6This is a graph showing the β diversity of intestinal flora in mice infected with Cryptosporidium parvum and treated with different drugs (a is PCoA analysis, b is NMDS analysis). Figure 6 It can be seen that the points of group C, group M, group P, group A, group F, and group AF all showed different degrees of separation, indicating that after treatment with group P, group A, group F, and group AF, the intestinal flora structure of mice changed; and the stress value of NMDS analysis (<0.2) also provided reliable results for further research and analysis ( Figure 6 a, 6b).

[0076] Figure 7 This is a petal diagram of the intestinal flora of mice infected with Cryptosporidium parvum and treated with different drugs. Figure 7 It can be seen that there are 107 common ASVs in the above 6 groups, of which group C has 3864 unique ASVs, group M has 1245 unique ASVs; groups P, A, F, and AF have 703, 1285, 1156, and 1522 unique ASVs, respectively.

[0077] Figure 8 This is a graph showing the LEfSe analysis of intestinal flora in mice infected with Cryptosporidium parvum and treated with different drugs. Figure 8 It can be seen that the intestinal flora in the above six groups differed in composition and structure, with significantly different phyla and genera; the characteristic flora of group C was Clostridium; the characteristic flora of group M was Ligilactobacillus, and significant enrichment of UBA3282, Burkholderia, Corynebacterium, and Mycobacterium was also detected; the characteristic flora of group P was Bacteroides and Parabacteroides, and Tannerelaceae, Mediterrane The characteristic flora of group A were Campylobacter and Helicobacter, and the enrichment of Mucispirillum and Deferribacterota was also detected; the characteristic flora of group F was Helicobacter, and the enrichment of Rikenellaceae and UBA11471 was also detected; the characteristic flora of group AF was Limosilactobacillus, and the enrichment of Desulfovibrionia, Nanosyncoccaceae and other flora was also detected.

[0078] This shows that after treatment with groups P, A, F, and AF, the main bacterial flora structure in the intestines of mice changed, and the relative abundance of beneficial bacteria increased, which had a significant effect on the recovery of the immune function of mice and the increase of the intestinal barrier.

[0079] The above is a detailed introduction to a Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum, its preparation method and application disclosed in the present invention. Specific examples are used herein to illustrate the principles and implementation methods of the present invention, and the description of the above embodiments is only used to help understand the method of the present invention and its core idea. It should be pointed out that for ordinary technicians in this technical field, without departing from the principle of the present invention, the present invention can also be improved and modified in a number of ways, and these improvements and modifications also fall within the scope of protection of the claims of the present invention.

Claims

1. A Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum, characterized in that: The Chinese medicine composition is composed of a formula for clearing intestines and removing dampness and allicin; the formula for clearing intestines and removing dampness is composed of scutellaria baicalensis, coptis chinensis, stewed costus root, stir-fried angelica sinensis, sanguisorba officinalis, stir-fried white peony root, angelica dahurica and liquorice; the Chinese medicine composition is prepared from the following raw materials in parts by weight: 10-15 parts of scutellaria baicalensis, 6-10 parts of coptis chinensis, 6-10 parts of stewed costus root, 10-15 parts of stir-fried angelica sinensis, 10-15 parts of sanguisorba officinalis, 15-20 parts of stir-fried white peony root, 10-20 parts of angelica dahurica, 6-10 parts of liquorice and 0.02-0.07 parts of allicin.

2. A Chinese medicinal composition capable of relieving diarrhea caused by Cryptosporidium parvum according to claim 1, characterized in that: The Chinese medicine composition is prepared from the following raw materials in parts by weight: 10 parts of scutellaria baicalensis, 6 parts of coptis chinensis, 6 parts of stewed costusroot, 10 parts of stir-fried angelica sinensis, 10 parts of sanguisorba officinalis, 15 parts of stir-fried white peony root, 10 parts of angelica dahurica, 6 parts of liquorice and 0.05 parts of allicin.

3. A method for preparing the Chinese medicinal composition capable of relieving diarrhea caused by Cryptosporidium parvum according to any one of claims 1 to 2, characterized in that: The following steps are involved: ① Weigh scutellaria baicalensis, coptis root, stewed costusroot, stir-fried angelica, sanguisorba officinalis, stir-fried white peony root, angelica dahurica, liquorice and allicin according to the weight ratio for later use; ② Add the scutellaria baicalensis, coptis chinensis, stewed costusroot, fried angelica sinensis, sanguisorba officinalis, fried white peony root, angelica dahurica and liquorice weighed in step ① into water, soak at room temperature for 30 minutes, then decoct at 100° C. for 60 to 70 minutes, filter and obtain a liquid medicine for clearing intestines and removing dampness; ③ After heating the intestinal clearing and dehumidifying prescription liquid prepared in step ② at 100° C. to 1 / 3 of the original volume, placing it in a refrigerator at -20° C. for 48 hours, and then placing it in a freeze dryer at -80° C. for 48 hours to remove moisture, to obtain the intestinal clearing and dehumidifying prescription freeze-dried powder, which is stored for later use; ④ According to the weight ratio, the lyophilized powder of the formula for clearing intestines and removing dampness prepared in step ③ is evenly mixed with allicin to prepare a Chinese medicine composition.

4. Use of the traditional Chinese medicine composition capable of relieving diarrhea caused by Cryptosporidium parvum according to any one of claims 1 to 2 in the preparation of drugs for resisting Cryptosporidium parvum and treating Cryptosporidium parvum diarrhea.

5. The use of the Chinese medicine composition according to claim 4, characterized in that: The anti-Cryptosporidium parvum drug refers to one or more products directly used for the prevention, diagnosis, detection, protection, treatment and research of Cryptosporidium parvum and its related diseases.

6. The use of the Chinese medicine composition according to claim 4, characterized in that: The drug for treating Cryptosporidium parvum diarrhea refers to one or more products directly used for preventing, diagnosing, detecting, protecting, treating and studying Cryptosporidium parvum diarrhea.

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