Application of Ipomoea aquatica Extract in Anti-Helicobacter pylori

Abstract

The present invention belongs to the technical field of traditional Chinese medicine, and specifically relates to the application of the extract of Ipomoea aquatica in anti-Helicobacter pylori. Through research, it is found that the extract of Ipomoea aquatica has a strong inhibitory effect on the standard strain of Helicobacter pylori. Moreover, this extract is a traditional Chinese medicine extract that is both edible and medicinal, with a low drug resistance incidence and few adverse reactions, which helps to improve the compliance of patients. It is particularly suitable for Helicobacter pylori-infected patients and can supplement or replace the commonly used quadruple antibiotic therapy for treating Helicobacter pylori. The present invention broadens the application field of the traditional Chinese medicine Ipomoea aquatica that is both edible and medicinal, and provides a new direction for the treatment of Helicobacter pylori. The extract of Ipomoea aquatica prepared by the preparation method of the present invention has no residual organic solvents, retains more active ingredients, has obvious antibacterial effects in vitro, and the extraction process is simple.

Description

Technical Field

[0001] The present invention belongs to the technical field of traditional Chinese medicine, and particularly relates to the application of the extract of Ipomoea aquatica in anti-Helicobacter pylori. Background Art

[0002] Helicobacter pylori (HP) is a Gram-negative microaerophilic spiral bacterium, and the global infection rate is about 50%. Helicobacter pylori is a broad-spectrum pathogenic bacterium that can infect a variety of diseases, commonly including gastritis, peptic ulcer, gastric adenocarcinoma, mucosa-associated lymphoid tissue lymphoma, etc. The clinical eradication treatment plan for Helicobacter pylori is mainly through the combination of antibiotics and other non-antibiotic drugs, such as acid suppressants, traditional Chinese medicine, etc. The most commonly used eradication plan is quadruple therapy. However, due to the influence of factors such as antibiotic resistance, the drug combinations and drug doses in the Helicobacter pylori eradication treatment plan are constantly increasing, the treatment course is prolonged, the overall eradication rate is still showing a downward trend, and there is a certain recurrence rate, and there are certain side effects during the treatment process. Clinical studies have shown that some traditional Chinese medicine components can not only directly play a bactericidal role, but also have a very significant regulatory effect on the body, and the combination with antibiotics can reduce the occurrence of adverse reactions.

[0003] Ipomoea aquatica is a plant of the class Dicotyledoneae in the phylum Angiospermae, and has the reputation of "green health food". Ipomoea aquatica is rich in resveratrol, quercetin, rutin, phytosterols, linoleic acid and various trace elements, and these substances can play a role in regulating the three highs. In addition, Ipomoea aquatica is rich in dietary fiber, and after consumption, it can stimulate gastrointestinal peristalsis and accelerate the passage of food through the gastrointestinal tract, and has the effect of promoting defecation. However, there is currently no report on its anti-Helicobacter pylori effect. Summary of the Invention

[0004] In order to overcome the deficiencies of the above-mentioned prior art, the present invention proposes the application of the extract of Ipomoea aquatica in the preparation of anti-Helicobacter pylori products. The present invention has been confirmed by a large number of experiments that the extract of Ipomoea aquatica has a strong inhibitory effect on the standard strain of Helicobacter pylori, indicating that the extract of Ipomoea aquatica is expected to be applied in anti-Helicobacter pylori.

[0005] In order to achieve the above object, the technical solution adopted by the present invention is as follows:

[0006] The first aspect of the present invention provides the application of the extract of Ipomoea aquatica in the preparation of anti-Helicobacter pylori drugs.

[0007] It has been confirmed through a large number of experiments that the extract of Ipomoea aquatica has a strong inhibitory effect on the standard strain of Helicobacter pylori and can be used in the preparation of anti-Helicobacter pylori products. Moreover, Ipomoea aquatica is a substance with both medicinal and edible properties, with a mild nature, suitable for both men and women, a low incidence of drug resistance, and few adverse reactions, which helps to improve the compliance of patients and is especially suitable for Helicobacter pylori-infected patients. The present invention broadens the application field of the traditional Chinese medicine Ipomoea aquatica with both medicinal and edible properties and provides a new direction for the treatment of Helicobacter pylori.

[0008] Preferably, the extraction method of the Ipomoea aquatica extract is as follows: after the Ipomoea aquatica is crushed, water is added and soaked for 30 - 60 min, and then extracted by water bath heating at 90 - 100 °C for 1 - 3 h. After collecting the supernatant from the obtained extract, it is concentrated and dried to obtain the extract.

[0009] Preferably, the extraction method of the Ipomoea aquatica extract is as follows: ethanol is added to the crushed Ipomoea aquatica, and after ultrasonic extraction, the supernatant is collected from the obtained extract, and then concentrated and dried to obtain the extract.

[0010] More preferably, by volume, the amount of water added is 8 - 20 times that of the crushed Ipomoea aquatica.

[0011] More preferably, the concentration of ethanol is 70 - 90%, and the material-liquid ratio of the crushed Ipomoea aquatica to ethanol is 3 - 7 g: 150 - 250 mL.

[0012] More preferably, the power of ultrasonic extraction is 70 - 90 W, and the time is 45 - 80 min.

[0013] More preferably, the extraction is carried out three times in total, and all the supernatants are combined.

[0014] More preferably, the Ipomoea aquatica is crushed to 30 - 60 meshes.

[0015] The second aspect of the present invention provides an anti-Helicobacter pylori drug combination, and the drug combination includes the Ipomoea aquatica extract and amoxicillin.

[0016] Preferably, the drug combination further includes pharmaceutically acceptable excipients. Further, the excipients are functional excipients available in the field of pharmaceutical preparations, including surfactants, suspending agents, emulsifiers, and some novel pharmaceutical polymers, such as cyclodextrin, chitosan, polylactic acid (PLA), poly (lactic acid-glycolic acid) copolymer (PLGA), hyaluronic acid, etc.

[0017] Preferably, the dosage form of the drug combination is any dosage form acceptable in the field of pharmacy, including powder, granule, injection, capsule, tablet, oral liquid.

[0018] Compared with the prior art, the beneficial effects of the present invention are:

[0019] The present invention has found through research that the extract of Ipomoea aquatica has a strong inhibitory effect on the standard strain of Helicobacter pylori. Moreover, this extract is a traditional Chinese medicine extract that is both edible and medicinal, with a low drug resistance incidence and few adverse reactions, which helps to improve the compliance of patients. It is especially suitable for Helicobacter pylori-infected patients and can supplement or replace the commonly used quadruple antibiotic therapy for treating Helicobacter pylori. The present invention broadens the application field of the traditional Chinese medicine Ipomoea aquatica that is both edible and medicinal, and provides a new direction for the treatment of Helicobacter pylori. The extract of Ipomoea aquatica prepared by the preparation method of the present invention has no residual organic solvents, retains more active ingredients, has obvious antibacterial effects in vitro, and the extraction process is simple. Detailed implementation manners

[0020] The following further describes the detailed implementation manners of the present invention. It should be noted here that the description of these implementation manners is used to help understand the present invention, but does not constitute a limitation on the present invention. In addition, the technical features involved in the various implementation manners of the present invention described below can be combined with each other as long as they do not conflict with each other.

[0021] The experimental methods in the following examples are all conventional methods unless otherwise specified, and the test materials used in the following examples are all commercially available through conventional channels unless otherwise specified.

[0022] Example 1 Extraction of the extract of Ipomoea aquatica and study on its anti-Helicobacter pylori efficacy

[0023] I. Method

[0024] 1. Preparation of medicinal materials

[0025] 1.1 Preparation of water extract: Crush Ipomoea aquatica with a pulverizer and pass through a 40-mesh sieve to obtain the crude medicinal powder. Weigh an appropriate amount of the crude medicinal powder into a round-bottom flask, add 10 times the amount (by volume) of secondary water and soak for 30 min, then heat and extract in a water bath at 90-100 °C for 1 h. After the obtained extract is cooled to room temperature, centrifuge at 4000 rpm for 10 min, and collect the supernatant. Pour the precipitate in the lower layer back into the round-bottom flask and continue to extract with 10 times the amount of secondary water. Extract a total of three times, and combine all the supernatants. The obtained supernatant is concentrated to an extract paste with a rotary evaporator, transferred to a glass dish, placed in a -80 °C refrigerator and frozen overnight, and then dried in a freeze dryer for 24 h to obtain the water extract of Ipomoea aquatica. Transfer the obtained extract to a dry sample bottle and store it in a -20 °C refrigerator for standby. Prepare at least three batches of water extracts of Ipomoea aquatica according to the same method. As shown in Table 1, the extraction rate of the water extract is 24.63%.

[0026] 1.2 Preparation of ethanol extract: Weigh 5 g of the coarsely powdered Xue Tongcai herbs crushed in step 1.1 into a 250 mL beaker, add 200 mL of 80% ethanol, extract by ultrasonic wave (80 W) for 61 min, filter by suction, collect the filtrate, pour the filter residue back into the beaker and continue extraction twice. Combine the filtrates from the three extractions, concentrate to dryness in a rotary evaporator under a water bath at 55 °C, redissolve with a small amount of secondary water, transfer the solution to a glass petri dish, seal it with plastic wrap, make holes, pre-freeze at -80 °C overnight, and then dry in a freeze dryer for 24 h to obtain the freeze-dried powder of the ethanol extract. Put the obtained freeze-dried powder into a brown sample bottle and store it in a -20 °C refrigerator for standby after drying. As shown in Table 1, the extraction rate of the ethanol extract is 8.35%.

[0027] 2. Preliminary antibacterial experiment of Xue Tongcai extract

[0028] The broth microdilution method was used for the determination of MIC. First, add 50.0 μL of the extract (the final concentration in the first well is 2560.0 μg / mL) / clarithromycin solution (the highest final concentration in the first well is 0.128 μg / mL) diluted two-fold with BHI to the 96-well plate, and then add 50.0 μL of the bacterial (Helicobacter pylori standard strain ATCC700392) suspension (the final concentration is 1×10 6 CFU / mL). Among them, clarithromycin was used as a positive control, and the medium containing only the extract BHI solution was used as a negative control. Place the 96-well plate in a three-gas incubator and shake and culture it under microaerophilic conditions at 37 °C and 150 rpm for 3 d. Define the drug concentration at which the bacterial turbidity decreases significantly as the MIC of the drug against H. pylori. The experiment was repeated three times.

[0029] As shown in Table 2, the MICs of the water extract and ethanol extract of Xue Tongcai against the Helicobacter pylori standard strain ATCC700392 were both 2560 μg / mL.

[0030] Table 1 Extraction rates of different extracts

[0031]

[0032] Table 2 Antibacterial effects of Xue Tongcai extract against Helicobacter pylori standard strain ATCC700392

[0033]

[0034]

[0035] Example 2 Combined effects of Xue Tongcai extract and antibiotics

[0036] The checkerboard dilution method was used to determine the combined effects of the extracts of Ipomoea aquatica Forsk. with antibiotics commonly used in clinical treatment of Helicobacter pylori infection. The specific experimental steps are as follows:

[0037] Prepare the medicinal liquid of Ipomoea aquatica Forsk. extracts at 80 - 1280 μg / mL by the two-fold dilution method (denoted as XTC). Prepare antibiotics at different concentrations in the same way [amoxicillin (AMO) at 0.125 - 8 μg / mL, clarithromycin (CLR) at 0.004 - 0.256 μg / mL, metronidazole (MTZ) at 0.05 - 3.2 μg / mL, levofloxacin (LEF) at 0.025 - 1.6 μg / mL]. Using Helicobacter pylori ATCC 43504 as a representative, scrape the bacteria with a moistened sterile cotton swab, adjust the turbidity of the bacterial suspension to 1 McF with brain heart infusion containing 20% FBS, and after diluting 10 times, according to the microbroth checkerboard dilution method, adjust the volume ratio of the bacterial suspension: the medicinal liquid of extracts: the antibiotic to 2:1:1, and inoculate 120 μL per well into a 96-well plate. Place it in a triple-gas incubator and culture it with shaking at 37 °C, in a microaerophilic environment, and at 150 rpm for 72 h. The well with obvious clarification observed with the naked eye is the minimum combined inhibitory concentration.

[0038] The combined inhibitory index FICI = MIC XTC (in combination) / MIC XTC (alone) + MIC antibiotic (in combination) / MIC antibiotic (alone).

[0039] When FICI ≤ 0.5, it indicates a synergistic effect; when 0.5 < FICI ≤ 1, it is an additive effect; when 1 < FICI ≤ 2, it is an irrelevant effect; when FICI > 2, it is an antagonistic effect.

[0040] The results are shown in Table 2. It can be seen from Table 2 that the extracts of Ipomoea aquatica Forsk. do not produce inhibitory effects on the antibiotics commonly used in clinical treatment of Helicobacter pylori infection. Among them, the combination of the extracts of Ipomoea aquatica Forsk. and amoxicillin shows an additive effect, which not only reduces the MIC of the antibiotic but also can reduce the clinical dosage of the antibiotic, reduce the occurrence of adverse reactions, and contribute to the improvement of the antibiotic resistance situation.

[0041] Table 2 The minimum combined inhibitory concentration of the combination of the extracts of Ipomoea aquatica Forsk. and antibiotics

[0042]

[0043] The above has made a detailed description of the embodiments of the present invention, but the present invention is not limited to the described embodiments. For those skilled in the art, without departing from the principles and spirit of the present invention, various changes, modifications, substitutions, and variations made to these embodiments still fall within the protection scope of the present invention.

Claims

1. Use of the extract of Ipomoea aquatica Forssk. in the preparation of an anti-Helicobacter pylori drug, characterized in that, The extraction method of the Ipomoea aquatica extract is selected from any of the following: (1) The Ipomoea aquatica is crushed to 30 - 60 meshes, then water is added for soaking for 30 - 60 min, followed by water bath heating extraction at 90 - 100 °C for 1 - 3 h. The supernatant is collected from the obtained extract, and the extraction is carried out three times in total. All the supernatants are combined, and then concentrated and dried to obtain the product; calculated by volume, the water addition amount is 8 - 20 times that of the crushed Ipomoea aquatica. (2) Ethanol is added to the Ipomoea aquatica crushed to 30 - 60 meshes, and after ultrasonic extraction, the supernatant is collected from the obtained extract. The extraction is carried out three times in total, and all the supernatants are combined, and then concentrated and dried to obtain the product; the concentration of ethanol is 70 - 90%, and the material - liquid ratio of the crushed Ipomoea aquatica to ethanol is 3 - 7 g: 150 - 250 mL; the power of ultrasonic extraction is 70 - 90 W, and the time is 45 - 80 min.

2. A drug combination against Helicobacter pylori, characterized in that, The pharmaceutical combination includes the Ipomoea aquatica extract and amoxicillin; the extraction method of the Ipomoea aquatica extract is selected from any of the following: (1) The Ipomoea aquatica is crushed to 30 - 60 meshes, then water is added for soaking for 30 - 60 min, followed by water bath heating extraction at 90 - 100 °C for 1 - 3 h. The supernatant is collected from the obtained extract, and the extraction is carried out three times in total. All the supernatants are combined, and then concentrated and dried to obtain the product; calculated by volume, the water addition amount is 8 - 20 times that of the crushed Ipomoea aquatica. (2) Ethanol is added to the Ipomoea aquatica crushed to 30 - 60 meshes, and after ultrasonic extraction, the supernatant is collected from the obtained extract. The extraction is carried out three times in total, and all the supernatants are combined, and then concentrated and dried to obtain the product; the concentration of ethanol is 70 - 90%, and the material - liquid ratio of the crushed Ipomoea aquatica to ethanol is 3 - 7 g: 150 - 250 mL; the power of ultrasonic extraction is 70 - 90 W, and the time is 45 - 80 min.

3. The anti-Helicobacter pylori drug combination according to claim 2, characterized in that, The pharmaceutical combination further includes pharmaceutically acceptable excipients.

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