A pharmaceutical composition capable of inhibiting Aspergillus niger, and its preparation and application

Abstract

The invention discloses a pharmaceutical composition capable of inhibiting Aspergillus niger, a preparation thereof and an application thereof. The pharmaceutical composition is composed of Atractylodes lancea, Clove, Zanthoxylum bungeanum, Artemisia argyi and Patchouli, and the pharmaceutical composition can be made into an extract, and further added with pharmaceutically acceptable excipients to prepare a preparation. The pharmaceutical composition, the extract and the preparation thereof can be used to prepare drugs and daily necessities for inhibiting Aspergillus niger.

Description

Technical Field

[0001] The invention belongs to the field of medicines, and in particular relates to a medicine composition capable of inhibiting Aspergillus niger, and a preparation and application thereof. Background Art

[0002] Aspergillus niger is a common species in the subphylum Ascomycota, class Hyphomycetes, orders Hyphomycetales, family Aspergillus. Aspergillus niger is an important industrial fermentation strain. It can also cause food to mold and spoil, and can secrete organic acids and toxins, which may cause human infection and illness. The harm of Aspergillus niger to the human body also includes reducing the body's resistance and immunity, and easily causing fungal infections. In patients with more severe symptoms, it can also cause fungal keratitis, fungal pneumonia, etc., and there are even reports of carcinogenesis.

[0003] At present, chemical mildew inhibitors are often used to treat Aspergillus niger, such as sulfur, dimethylphosphine, aluminum phosphide, pyrrolobenzene, imidazole and other reagents for fumigation and sterilization. Although these reagents have the effect of inhibiting Aspergillus niger, there are problems such as reagent residue and mold resistance. Therefore, the research and development of green and safe antibacterial alternatives has become one of the urgent problems to be solved. Summary of the invention

[0004] In view of the deficiencies in the prior art, the object of the present invention is to provide a pharmaceutical composition capable of inhibiting Aspergillus niger and its preparation and use. The pharmaceutical composition and its preparation have strong anti-Aspergillus niger activity and are safe and have no toxic side effects.

[0005] The objective of the present invention is achieved through the following technical solutions:

[0006] A pharmaceutical composition capable of inhibiting Aspergillus niger consists of the following components by weight: 40-80 parts of atractylodes, 40-80 parts of cloves, 40-80 parts of zanthoxylum bungeanum, 40-80 parts of wormwood leaves and 40-80 parts of patches of galangal.

[0007] Preferably, the pharmaceutical composition capable of inhibiting Aspergillus niger is composed of the following components by weight: 50-70 parts of Atractylodes lancea, 50-70 parts of Cloves, 50-70 parts of Zanthoxylum bungeanum, 50-70 parts of Artemisia argyi, and 50-70 parts of Patchouli.

[0008] More preferably, the pharmaceutical composition capable of inhibiting Aspergillus niger is composed of the following components by weight: 55-65 parts of Atractylodes lancea, 55-65 parts of Cloves, 55-65 parts of Zanthoxylum bungeanum, 55-65 parts of Artemisia argyi, and 55-65 parts of Patchouli.

[0009] The pharmaceutical composition of the present invention is composed of five Chinese medicines, each of which contains multiple active ingredients, such as terpenes, flavonoids, alkaloid compounds, etc. The numerous active ingredients interact with each other, can act on multiple targets at the same time, and exert a synergistic effect. Among them, wormwood volatile oil can block the invasion ability and drug resistance of Aspergillus niger by inhibiting the formation of Aspergillus niger biofilm or destroying the formed biofilm. After the whole prescription is compatible, it can act on the cell wall and cell membrane of Aspergillus niger, causing Aspergillus niger to shrink and collapse, so that Aspergillus niger cannot carry out normal cell division and reproduction; At the same time, it also affects the activity of DNA topoisomerase or ATP-related enzymes, thereby interfering with and inhibiting the metabolic ability of Aspergillus niger, making it unable to carry out normal metabolism and energy conversion, and finally causing the death or inactivation of Aspergillus niger. The pharmaceutical composition plays an inhibitory and killing effect on Aspergillus niger from many aspects, therefore, it has a good effect of inhibiting Aspergillus niger.

[0010] The pharmaceutical composition capable of inhibiting Aspergillus niger can be prepared into an extract by adopting a low-temperature reduced-pressure reflux method or a steam distillation method.

[0011] The low-temperature, reduced-pressure reflux method comprises the following steps: taking all the medicinal materials, crushing them, adding 10 times the amount of solvent, intermittently ultrasonically oscillating, cold soaking overnight, extracting under reduced-pressure reflux at 30-40° C., and removing the solvent from the extract to obtain the obtained product.

[0012] The pharmaceutical composition capable of inhibiting Aspergillus niger and the extract thereof can be added with pharmaceutically acceptable excipients to prepare a preparation.

[0013] The pharmaceutical composition, extract and preparation thereof capable of inhibiting Aspergillus niger can be used for preparing drugs, daily necessities and the like capable of inhibiting Aspergillus niger.

[0014] Compared with the prior art, the present invention has the following beneficial effects:

[0015] 1. The pharmaceutical composition of the present invention has strong anti-Aspergillus niger activity, and the antibacterial effect of the pharmaceutical composition on Aspergillus niger is better than the antibacterial effect of any single herb in the pharmaceutical composition.

[0016] 2. Long-term clinical studies have shown that the pharmaceutical composition of the present invention also has a good inhibitory effect on Staphylococcus aureus, Escherichia coli, etc.

[0017] 3. The pharmaceutical composition of the present invention is composed of Chinese medicinal materials, which are essentially plants. Therefore, the pharmaceutical composition is of natural origin and has the advantages of safety, environmental protection, and non-toxicity.

[0018] 4. The extracts of the pharmaceutical composition of the present invention all have strong anti-Aspergillus niger activity, and the methanol extract has the strongest activity. So far, there is no report in the prior art on the use of methanol to extract aromatic Chinese medicine compounds.

[0019] 5. The present invention adopts 75% ethanol and glycerol as a mixed solvent in the dosage form preparation process, which can better meet the dissolution requirements of different compounds in the extract, so that the effective ingredients in the extract are fully dissolved. At the same time, the high viscosity and high moisturizing properties of glycerol are retained. The overall process is simpler and more stable, the cost is low, and the prepared preparation has a significant antibacterial effect. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 The inhibitory effects of the four extracts of different concentrations in Example 1 on Aspergillus niger are shown in Table 1 (wherein: Ⅰ is a steam distillation extract, Ⅱ is a petroleum ether extract, Ⅲ is an ethyl acetate extract, and Ⅳ is a methanol extract; A is 200 μL / mL, B is 100 μL / mL, C is 50 μL / mL, and D is a blank control).

[0021] Figure 2 The inhibitory effects of the pharmaceutical composition of the present invention and the methanol extracts of five medicinal materials on Aspergillus niger in Application Example 2 are compared (wherein: ① is a 200 μL / mL concentration group, ② is a 100 μL / mL concentration group, ③ is a 50 μL / mL concentration group; a is an extract of the composition, b is an extract of wormwood, c is an extract of cloves, d is an extract of Patchouli, e is an extract of Atractylodes, f is an extract of Zanthoxylum bungeanum, and g is a blank control group). DETAILED DESCRIPTION

[0022] The present invention will be further described below in conjunction with embodiments, but the present invention is not limited to the embodiments. Example 1

[0023] A pharmaceutical composition capable of inhibiting Aspergillus niger is composed of the following components: 40g of Atractylodes lancea, 40g of Clove, 40g of Zanthoxylum bungeanum, 40g of Artemisia argyi and 40g of Patchouli. Example 2

[0024] A pharmaceutical composition capable of inhibiting Aspergillus niger is composed of the following components: 40g of Atractylodes lancea, 50g of Cloves, 55g of Zanthoxylum bungeanum, 60g of Artemisia argyi and 65g of Patchouli. Example 3

[0025] A pharmaceutical composition capable of inhibiting Aspergillus niger is composed of the following components: 50 g of Atractylodes lancea, 65 g of Cloves, 60 g of Zanthoxylum bungeanum, 55 g of Artemisia argyi and 70 g of Patchouli. Example 4

[0026] A pharmaceutical composition capable of inhibiting Aspergillus niger is composed of the following components: 55 g of Atractylodes lancea, 70 g of Cloves, 65 g of Zanthoxylum bungeanum, 60 g of Artemisia argyi and 80 g of Patchouli. Example 5

[0027] A pharmaceutical composition capable of inhibiting Aspergillus niger is composed of the following components: 60g of Atractylodes lancea, 80g of Cloves, 70g of Zanthoxylum bungeanum, 65g of Artemisia argyi and 40g of Patchouli. Example 6

[0028] A pharmaceutical composition capable of inhibiting Aspergillus niger is composed of the following components: 65g of Atractylodes lancea, 40g of Cloves, 80g of Zanthoxylum bungeanum, 70g of Artemisia argyi and 50g of Patchouli. Example 7

[0029] A pharmaceutical composition capable of inhibiting Aspergillus niger is composed of the following components: 70 g of Atractylodes lancea, 50 g of Cloves, 40 g of Zanthoxylum bungeanum, 80 g of Artemisia argyi and 55 g of Patchouli. Example 8

[0030] A pharmaceutical composition capable of inhibiting Aspergillus niger is composed of the following components: 80g of Atractylodes lancea, 55g of Cloves, 50g of Zanthoxylum bungeanum, 40g of Artemisia argyi and 60g of Patchouli. Example 9

[0031] An extract of a pharmaceutical composition capable of inhibiting Aspergillus niger is prepared by weighing medicinal materials according to the weight of the pharmaceutical composition described in any one of Examples 1-8, and performing the following steps: crushing all the medicinal materials into coarse powder and passing through a 10-mesh sieve, adding 10 times the amount of methanol, ultrasonicating for 20 minutes each time, performing intermittent ultrasonic oscillation 3 times, cold soaking overnight, extracting under reduced pressure and reflux at 40°C, and finally recovering the solvent. Example 10

[0032] An extract of a pharmaceutical composition capable of inhibiting Aspergillus niger is prepared by weighing medicinal materials according to the weight of the pharmaceutical composition described in any one of Examples 1-8, and performing the preparation according to the following steps: crushing all the medicinal materials into coarse powder and passing through a 10-mesh sieve, adding 10 times the amount of petroleum ether, ultrasonicating for 20 minutes each time, performing intermittent ultrasonic oscillation 3 times, cold soaking overnight, extracting under reduced pressure and reflux at 30°C, and finally recovering the solvent. Embodiment 11

[0033] An extract of a pharmaceutical composition capable of inhibiting Aspergillus niger is prepared by weighing medicinal materials according to the weight of the pharmaceutical composition described in any one of Examples 1-8, and performing the following steps: crushing all the medicinal materials into coarse powder and passing through a 10-mesh sieve, adding 10 times the amount of ethyl acetate, ultrasonicating for 20 minutes each time, performing intermittent ultrasonic oscillation for 3 times, cold soaking overnight, extracting under reduced pressure and reflux at 40°C, and finally recovering the solvent. Example 12

[0034] An extract of a pharmaceutical composition capable of inhibiting Aspergillus niger is prepared by weighing medicinal materials according to the weight of the pharmaceutical composition described in any one of Examples 1-8 and performing the following steps: crushing all the medicinal materials into coarse powder, passing through a 10-mesh sieve, adding 10 times the amount of water, soaking for 2 hours, extracting by steam distillation for 5 hours, standing for 1 hour, adding excess anhydrous sodium sulfate, and centrifuging at 6000 r / min for 10 minutes. Embodiment 13

[0035] An antibacterial liquid capable of inhibiting Aspergillus niger is prepared by taking an extract of the pharmaceutical composition capable of inhibiting Aspergillus niger described in any one of Examples 9 to 12 according to the following steps: adding 5%-8% (volume percentage) of glycerol to the extract of the pharmaceutical composition capable of inhibiting Aspergillus niger and stirring evenly, then adding 15%-20% (volume percentage) of 75% alcohol and mixing evenly, and filtering through a 0.22 μm filter to obtain the antibacterial liquid capable of inhibiting Aspergillus niger.

[0036] Application Example 1 - Experiment on the antibacterial effect of the extract of the pharmaceutical composition of the present invention on Aspergillus niger

[0037] The medicinal materials were weighed according to the pharmaceutical composition described in Example 1, and methanol extract, petroleum ether extract, ethyl acetate extract and steam distillation extract were prepared according to the methods described in Examples 9-12.

[0038] Prepare PDA agar plates, use a sterile pipette to transfer 200 μL of Aspergillus niger liquid onto the plate on an ultra-clean workbench, and spread it evenly. Mix the four extracts with Tween and then dilute them with sterile water to prepare three concentrations: 200 μL / mL, 100 μL / mL, and 50 μL / mL. Use blank drug-sensitive tablets with a diameter of 6.0 mm to soak the extracts of different concentrations. The blank control group is soaked in a blank solvent mixed with Tween and sterile water for 30 minutes, and then the culture dish is placed in a biochemical incubator at 26±2℃ for 48 hours, and the diameter of the inhibition zone is measured. The experiment was repeated 3 times, and the results are shown in Table 1 and Figure 1 shown.

[0039] SPSS 27.0 statistical software was used for analysis. When the data conform to normal distribution, one-way ANOVA test is used to judge the overall significant difference. When the variance is equal, Tukey method is used, and when the variance is unequal, Dunnett T3 method is used. When the data do not conform to normal distribution, non-parametric test is used. t test, P The difference was statistically significant when <0.05.

[0040]

[0041] Conclusion: The extracts obtained by extracting the pharmaceutical composition of the present invention with four different solvents all have inhibitory effects on Aspergillus niger, and the inhibitory effects are ranked from strong to weak as follows: methanol extract > petroleum ether extract > steam distillation extract > ethyl acetate extract, among which the methanol extract has the strongest inhibitory effect, which is significantly better than the antibacterial effects of the other three extracts.

[0042] Application Example 2 - Comparative Experiment on the Antibacterial Effect of the Extract of the Pharmaceutical Composition of the Present Invention and the Extract of a Single Herbal Material on Aspergillus Niger

[0043] The medicinal materials were weighed according to the pharmaceutical composition described in Example 1, and the same amount of wormwood, cloves, Patchouli, Atractylodes, and Zanthoxylum bungeanum was weighed respectively to prepare according to the method described in Example 9 to obtain the methanol extract of the pharmaceutical composition and the methanol extracts of the five medicinal materials respectively.

[0044] Prepare a PDA agar plate, use a sterile pipette to transfer 200 μL of Aspergillus niger liquid to the plate on a clean bench, and spread it evenly. The drug composition and the methanol extracts of the five medicinal materials were prepared into three concentrations (the method is the same as that of Application Example 1): 200 μL / mL, 100 μL / mL, and 50 μL / mL. For the blank control group, blank drug-sensitive sheets with a diameter of 6.0 mm were soaked in extracts of different concentrations for 30 minutes, and then the culture dishes were placed in a biochemical incubator at 26±2°C for 48 hours, and the diameter of the inhibition zone was measured. The experiment was repeated 3 times, and the results are shown in Table 2 and Figure 2 shown.

[0045]

[0046] The experimental results show that the antibacterial effect of the composition of the present invention on Aspergillus niger is better than that of any single herbal medicine of Artemisia argyi, clove, Patchouli, Atractylodes lancea and Zanthoxylum bungeanum. P <0.01, the difference is statistically significant.

[0047] Application Example 3 - Experiment on the antibacterial effect of the antibacterial liquid prepared from the extract of the pharmaceutical composition of the present invention on Aspergillus niger

[0048] The medicinal materials were weighed according to the pharmaceutical composition described in Example 1, and the methanol extract was prepared according to the preparation method described in Example 9. The methanol extract was taken and weighed, and then the antibacterial solution was prepared according to the preparation method described in Example 13, so that the mass volume concentration of the methanol extract in the antibacterial solution was 100 mg / mL.

[0049] Prepare PDA agar plates, transfer 200 μL of Aspergillus niger liquid onto the plates with a sterile pipette on the clean bench, and spread evenly. Berberine hydrochloride solution (100 mg / mL) was used as a positive control, and a mixture of glycerol and 75% alcohol in the same amount as the antibacterial liquid was used as a blank control. Soak blank drug-sensitive tablets with a diameter of 6.0 mm for 30 minutes, then place the culture dish in a 26±2℃ biochemical incubator for 48 hours, and measure the diameter of the antibacterial zone. The experiment was repeated 10 times, and the results are shown in Table 3.

[0050]

[0051] The experimental results show that the mixture of glycerol and 75% alcohol has no antibacterial effect. The antibacterial liquid of the pharmaceutical composition of the present invention has a very significant inhibitory effect on Aspergillus niger, and the inhibitory effect is even stronger than that of berberine hydrochloride commonly used at present. P <0.01, the difference is statistically significant.

Claims

1. A pharmaceutical composition capable of inhibiting Aspergillus niger, characterized in that: The pharmaceutical composition is prepared from the methanol extracts of the following raw materials by weight: 40-80 parts of Atractylodes lancea, 40-80 parts of cloves, 40-80 parts of Zanthoxylum bungeanum, 40-80 parts of Artemisia argyi, and 40-80 parts of Patchouli; the methanol extract is prepared by a low-temperature, reduced-pressure, and reflux extraction method, comprising the following steps: taking all the medicinal materials, crushing them, adding methanol, intermittent ultrasonic oscillation, cold soaking overnight, extracting under reduced pressure and reflux at 30-40°C, and recovering the solvent from the extract.

2. The pharmaceutical composition capable of inhibiting Aspergillus niger according to claim 1, characterized in that: The pharmaceutical composition is prepared from the methanol extracts of the following raw materials by weight: 50-70 parts of Atractylodes lancea, 50-70 parts of Cloves, 50-70 parts of Zanthoxylum bungeanum, 50-70 parts of Artemisia argyi, and 50-70 parts of Patchouli.

3. The pharmaceutical composition capable of inhibiting Aspergillus niger according to claim 2, characterized in that: The pharmaceutical composition is prepared from the methanol extracts of the following raw materials by weight: 55-65 parts of Atractylodes lancea, 55-65 parts of Cloves, 55-65 parts of Zanthoxylum bungeanum, 55-65 parts of Artemisia argyi, and 55-65 parts of Patchouli.

4. The preparation of the pharmaceutical composition capable of inhibiting Aspergillus niger according to any one of claims 1 to 3, characterized in that: The preparation is prepared by adding pharmaceutically acceptable excipients to the pharmaceutical composition.

5. The preparation of the pharmaceutical composition capable of inhibiting Aspergillus niger according to claim 4, characterized in that: The preparation method of the preparation comprises the following steps: adding 5%-8% by volume of glycerol to the pharmaceutical composition, stirring evenly, then adding 15%-20% by volume of 75% alcohol, mixing evenly, and filtering through 0.22 μm to obtain the preparation.

6. Use of the pharmaceutical composition capable of inhibiting Aspergillus niger according to any one of claims 1 to 3, characterized in that: The pharmaceutical composition capable of inhibiting Aspergillus niger is used for preparing medicines and daily necessities for inhibiting Aspergillus niger.

7. The use of the pharmaceutical composition for inhibiting Aspergillus niger according to claim 4, characterized in that: The preparation of the pharmaceutical composition capable of inhibiting Aspergillus niger is used for preparing medicines and daily necessities for inhibiting Aspergillus niger.

Images