A method for compounding plant extracts with antibacterial effects and application thereof

Abstract

The invention provides a compounding method and application of a plant extract with antibacterial effect, and relates to the technical field of natural plant extracts. The plant extracts are compounded in pairs by alopecuroides alopecuroides extract, a Mongolian allium extract and an oblique stem astragalus extract to prepare alopecuroides-Mongolian allium compound, a Mongolian allium-oblique stem astragalus compound and alopecuroides-oblique stem astragalus compound; the alopecuroides-Mongolian allium compound is prepared by alopecuroides extract and a Mongolian allium extract in a volume ratio of 1:1; the Mongolian allium-oblique stem astragalus compound is prepared by alopecuroides extract and an oblique stem astragalus extract in a volume ratio of 1:1; the alopecuroides-oblique stem astragalus compound is prepared by alopecuroides extract and an oblique stem astragalus extract in a volume ratio of 1:1; and the alopecuroides-oblique stem astragalus compound is prepared by alopecuroides extract and an oblique stem astragalus extract in a volume ratio of 1:1. After compounding the three plant extracts of alopecuroides, Mongolian allium and oblique stem astragalus in pairs, the bacterial concentration is significantly reduced, and the antibacterial efficiency of the three plant extracts is effectively improved.

Description

Technical Field

[0001] The invention relates to the technical field of natural plant extracts, in particular to a compound antibacterial agent capable of inhibiting the growth of main pathogenic bacteria of cow mastitis. Background Art

[0002] Cow mastitis is an inflammatory disease caused by contact with pathogenic microorganisms (such as Staphylococcus aureus, Streptococcus agalactiae) and environmental pathogens (Escherichia coli, Enterococcus, etc.), which can lead to a decrease in milk production and milk quality in dairy cows. The economic losses caused by cow mastitis in my country are as high as 10 billion yuan each year. At present, under the background of the country's advocacy of healthy breeding and antibiotic reduction and substitution, plant extracts have the advantages of being natural and without side effects, and have the potential to develop the prevention and treatment of cow mastitis.

[0003] Bitter beans ( Sophora alopecuroides L.): It is a common medicinal plant of the genus Sophora in the Leguminosae family. Its active ingredients are mainly alkaloids and flavonoids.

[0004] Mongolian Allium Mongolian garlic Regel): Also known as sand onion, it is a nutritious and uniquely flavored wild vegetable of the Amaryllis family Allium genus, mainly containing some polysaccharides, flavonoids and essential oils.

[0005] Astragalus obliquus Astragalus laxmannii Jacq.): Also known as Astragalus, it is an herbaceous plant of the genus Astragalus in the leguminous family, mainly containing flavonoids.

[0006] At present, relevant studies have reported that bitter bean extract has a good in vitro antibacterial effect on multidrug-resistant Escherichia coli, Mongolian leek extract can replace antibiotics and improve the quality of mutton, and Astragalus oblique stem extract has a good antibacterial effect on Escherichia coli and Staphylococcus aureus; however, there are few reports on the antibacterial effect of bitter bean extract, Mongolian leek extract, and Astragalus oblique stem extract after compounding. Summary of the invention

[0007] In order to solve the above technical problems, the present invention discloses a method for compounding plant extracts with antibacterial effects. The plant extracts are compounded in pairs by using a sophora alopecuroides extract, a Mongolian allium extract, and an astragalus oblique stem extract to prepare a sophora alopecuroides-mongolian allium compound, a Mongolian allium-oblique stem astragalus compound, and a sophora alopecuroides-oblique stem astragalus compound;

[0008] The sophora alopecuroides-allium mongolianum compound is prepared by mixing the sophora alopecuroides extract and the allium mongolianum extract in a volume ratio of 1:1;

[0009] The Mongolian Allium Mongolianum-Astragalus Oblique Stem compound is prepared by mixing Mongolian Allium Mongolianum extract and Astragalus Oblique Stem extract in a volume ratio of 1:1;

[0010] The sophora alopecuroides-astragalus oblique stem compound is prepared by mixing the sophora alopecuroides extract and the astragalus oblique stem extract in a volume ratio of 1:1;

[0011] The concentration of the sophora alopecuroides extract is 80 mg / L-160 mg / L;

[0012] The concentration of the Mongolian Allium extract is 80 mg / L-320 mg / L;

[0013] The concentration of the Astragalus obliquus extract is 80 mg / L-320 mg / L.

[0014] Moreover, the preparation method of the sophora alopecuroides extract / Allium mongolianum extract / Astragalus obliquus extract is as follows:

[0015] (a) Freshly collected Sophora alopecuroides plants / Allium mongolicum plants / Astragalus obliquus plants were naturally air-dried, cut into 1-2 cm, and made into powder. 10 g of plant powder was weighed, and 90% ethanol solution was added according to a solid-liquid ratio of 20:1. The mixture was heated and refluxed in a reflux device for 2 h. The supernatant was taken, and the extraction was repeated 3 times according to the above steps. The supernatants were combined. In the ethanol solution, V 乙醇 :V 水 =9:1;

[0016] (b) After decolorization with petroleum ether, the mixture was evaporated in an evaporator to a volume of 1 / 4 of the original volume, and concentrated freeze-dried to obtain the Sophora alopecuroides extract / Allium mongolicum extract / Astragalus obliquus extract, which were then stored at -20°C.

[0017] The three plant extracts prepared according to the above method were made into tryptone soy broth culture media containing 80 mg / L-320 mg / L of the extracts, respectively. They were compounded in pairs according to different concentrations, and finally the optimal concentration combination for inhibiting Escherichia coli, Staphylococcus aureus, and Streptococcus agalactiae was obtained.

[0018] On the other hand, the present invention provides an application of a method for compounding a plant extract with antibacterial efficacy, which is used to inhibit Escherichia coli, Staphylococcus aureus, and Streptococcus agalactiae.

[0019] Moreover, the three plant extracts were mixed in pairs according to a volume ratio of 1:1, and the minimum inhibitory concentration combination for Escherichia coli was:

[0020] The minimum inhibitory concentration of the sophora alopecuroides-astragalus oblique stem compound is: 160 mg / L sophora alopecuroides extract-160 mg / L astragalus oblique stem extract;

[0021] The minimum inhibitory concentration of the bitter bean-Mongolian allium compound is: 160 mg / L bitter bean extract-80 mg / L Mongolian allium extract;

[0022] The minimum inhibitory concentration of the Mongolian Allium Mongolianum-Astragalus oblique stem compound is: 160 mg / L Mongolian Allium Mongolianum extract-160 mg / L Astragalus oblique stem extract.

[0023] Moreover, the three plant extracts were mixed in pairs according to a volume ratio of 1:1, and the minimum inhibitory concentration combination for Staphylococcus aureus was:

[0024] The minimum inhibitory concentration of the sophora alopecuroides-astragalus oblique stem compound is: 160 mg / L sophora alopecuroides extract-160 mg / L astragalus oblique stem extract;

[0025] The minimum inhibitory concentration of the bitter bean-Mongolian allium compound is: 160 mg / L bitter bean extract-320 mg / L Mongolian allium extract;

[0026] The minimum inhibitory concentration of the Mongolian Allium Mongolianum-Astragalus oblique stem compound is: 160 mg / L Mongolian Allium Mongolianum extract-320 mg / L Astragalus oblique stem extract.

[0027] Moreover, the three plant extracts were mixed in pairs according to a volume ratio of 1:1, and the minimum inhibitory concentration combination for Streptococcus agalactiae was:

[0028] The minimum inhibitory concentration of the sophora alopecuroides-astragalus oblique stem compound is: 160 mg / L sophora alopecuroides extract-160 mg / L astragalus oblique stem extract;

[0029] The minimum inhibitory concentration of the bitter bean-Mongolian allium compound is: 160 mg / L bitter bean extract-160 mg / L Mongolian allium extract;

[0030] The minimum inhibitory concentration of the Mongolian Allium Mongolianum-Astragalus oblique stem compound is: 160 mg / L Mongolian Allium Mongolianum extract-160 mg / L Astragalus oblique stem extract.

[0031] Moreover, the extract of bitter bean is pseudomorphine, atropine, theophylline, sophora flavescens, deoxytubule glycoside, lupine, quinidine, subicillin, spartinine, aloperidine, retamine, arecaline, fetidine, and thebaine;

[0032] The Mongolian allium extract is gelatin and schisandra saponin;

[0033] The extract of Astragalus obliquus contains isocarpioside, beta-ecdysone, atractylodesin, phytolaccaside and naringin.

[0034] The beneficial effects of the present invention are:

[0035] 1. The effect of the compound preparation of Sophora alopecuroides and Astragalus obliquus provided by the present invention on Escherichia coli ( E. coli ), Staphylococcus aureus ( S.aureus )、Streptococcus agalactiae( S. agalactiae ) The inhibitory effect of the tested bacteria was additive; the bitter bean-Mongolian allium compound agent had an inhibitory effect on Escherichia coli ( E. coli ) and Streptococcus agalactiae ( S. agalactiae ) The inhibitory effect of the tested bacteria was additive; the Mongolian Allium-Astragalus oblique stem compound had an inhibitory effect on Escherichia coli ( E. coli ) and Streptococcus agalactiae ( S. agalactiae ) The inhibitory effect on the test bacteria was additive.

[0036] 2. The antibacterial concentration of the three plant extracts of Sophora alopecuroides, Allium mongolicum and Astragalus oblique stems after compounding them in pairs in the present invention is significantly reduced, effectively improving the antibacterial efficiency of the three plant extracts of Sophora alopecuroides, Allium mongolicum and Astragalus oblique stems.

[0037] 3. The antibacterial effect of the plant extract provided by the present invention is equivalent to that of the antibiotics gentamicin and norfloxacin, and can replace traditional antibiotics. Moreover, compared with traditional antibiotics, the plant extract used in the present invention has the advantages of being natural and having no side effects, which not only conforms to the current concept of healthy breeding and reducing and replacing antibiotics, but also can effectively reduce antibiotic residues and improve the safety and quality of dairy products.

[0038] 4. The plant extract preparation method of the present invention is simple, easy to operate, and has high extraction efficiency. High-concentration plant extracts can be obtained through reflux extraction, decolorization, concentration freeze drying and other steps, which provides strong support for subsequent compounding and application.

[0039] 5. Since the plant extract compound provided by the present invention has significant antibacterial effect and is natural and has no side effects, it can not only be used for the prevention and treatment of cow mastitis, but can also be expanded to the prevention and treatment of other animal diseases as well as food preservation, sanitation and disinfection, and has broad market application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] Figure 1 The effects of different concentrations of three plant extracts on Escherichia coli ( E.coli )’s antibacterial effect;

[0041] Figure 2 The results of different concentrations of three plant extracts on Staphylococcus aureus ( S.aureus )’s antibacterial effect;

[0042] Figure 3 The results of different concentrations of three plant extracts on Streptococcus agalactiae ( S. agalactiae )'s antibacterial effect. DETAILED DESCRIPTION

[0043] The technical scheme of the invention will be clearly and completely described below in conjunction with the drawings in the embodiments of the invention. The described examples are not all examples of the invention. Based on the embodiments of the present invention, other inventions without substantial technical breakthroughs belong to the protection scope of the present invention.

[0044] Example 1

[0045] Preparation method of Sophora alopecuroides extract:

[0046] (a) Freshly collected sophora alopecuroides plants were air-dried, and the dried plants were cut into 1-2 cm pieces and powdered using a high-speed grinder. 10 g of plant powder was weighed and added with 200 mL of 90% ethanol solution (V 乙醇 :V 水 =9:1), heat and reflux in a reflux device for 2 h, take the supernatant, repeat the extraction three times according to the above steps, and combine the supernatants;

[0047] (b) adding petroleum ether for decolorization, and evaporating the mixture on a rotary evaporator until the volume is 1 / 4 of the original volume;

[0048] (c) The powder extract was prepared by freeze-drying and stored at -20°C.

[0049] Furthermore, the extract of bitter bean is pseudomorphine, atropine, theophylline, sophora flavescens, deoxytubule glycoside, lupine, quinidine, subicillin, spartinine, aloperidine, retamine, arecaline, fetidine, and thebaine.

[0050] Example 2

[0051] Preparation method of Mongolian allium extract:

[0052] (a) The naturally dried Mongolian leek plants were cut into pieces of 1-2 cm and powdered using a high-speed grinder. 10 g of plant powder was weighed and added with 90% ethanol solution (V 乙醇 :V 水 =9:1), heat and reflux in a reflux device for 2 h, take the supernatant, repeat the extraction three times according to the above steps, and combine the supernatants;

[0053] (b) decolorizing with petroleum ether and evaporating on a rotary evaporator to a volume of 1 / 4 of the original volume;

[0054] (c) The powder extract was prepared by freeze-drying and stored at -20°C.

[0055] Furthermore, the Mongolian Allium extract is gelatin and Schisandra saponin.

[0056] Example 3

[0057] Preparation method of Astragalus obliquus extract:

[0058] (a) The naturally dried Astragalus obliquus plants were cut into pieces of 1-2 cm and powdered using a high-speed grinder. 10 g of plant powder was weighed and added with 90% ethanol solution (V 2.5) at a solid-liquid ratio of 20:1. 乙醇 :V 水 =9:1) and refluxed in a reflux device for 2 h, taking the supernatant, repeating the above steps for 3 times, and combining the supernatants;

[0059] (b) decolorizing with petroleum ether and evaporating on a rotary evaporator to a volume of 1 / 4 of the original volume;

[0060] (c) The powder extract was prepared by freeze-drying and stored at -20°C.

[0061] Furthermore, the Astragalus obliquus extract is isocarpioside, β-ecdysone, atractylodesin, phytolaccaside, and naringin.

[0062] Example 4

[0063] A method for compounding plant extracts with antibacterial effects, comprising compounding plant extracts of Sophora alopecuroides extract, Allium mongolicum extract, and Astragalus oblique stem extract in pairs to prepare Sophora alopecuroides-Allium mongolicum compound, Allium mongolicum-Astragalus oblique stem compound, and Sophora alopecuroides-Astragalus oblique stem compound;

[0064] The sophora alopecuroides-allium mongolianum compound is prepared by mixing the sophora alopecuroides extract and the allium mongolianum extract in a volume ratio of 1:1;

[0065] The Mongolian Allium Mongolianum-Astragalus Oblique Stem compound is prepared by mixing Mongolian Allium Mongolianum extract and Astragalus Oblique Stem extract in a volume ratio of 1:1;

[0066] The sophora alopecuroides-astragalus oblique stem compound is prepared by mixing the sophora alopecuroides extract and the astragalus oblique stem extract in a volume ratio of 1:1;

[0067] The concentration of the sophora alopecuroides extract is 80 mg / L-160 mg / L;

[0068] The concentration of the Mongolian Allium extract is 80 mg / L-320 mg / L;

[0069] The concentration of the Astragalus obliquus extract is 80 mg / L-320 mg / L.

[0070] Furthermore, the preparation methods of the Sophora alopecuroides extract, the Allium mongolicum extract, and the Astragalus obliquus extract are shown in Examples 1-3.

[0071] Example 5

[0072] An application of a compounding method of plant extracts with antibacterial efficacy for inhibiting Escherichia coli; the minimum inhibitory concentration combination is:

[0073] The minimum inhibitory concentration of the sophora alopecuroides-astragalus oblique stem compound is: 160 mg / L sophora alopecuroides extract-160 mg / L astragalus oblique stem extract;

[0074] The minimum inhibitory concentration of the bitter bean-Mongolian allium compound is: 160 mg / L bitter bean extract-80 mg / L Mongolian allium extract;

[0075] The minimum inhibitory concentration of the Mongolian Allium Mongolianum-Astragalus Oblique Stem compound is: 160 mg / L Mongolian Allium Mongolianum extract-160 mg / L Astragalus Oblique Stem extract.

[0076] Example 6

[0077] An application of a compounding method of plant extracts with antibacterial efficacy, used to inhibit Staphylococcus aureus; the minimum inhibitory concentration combination is:

[0078] The minimum inhibitory concentration of the sophora alopecuroides-astragalus oblique stem compound is: 160 mg / L sophora alopecuroides extract-160 mg / L astragalus oblique stem extract;

[0079] The minimum inhibitory concentration of the bitter bean-Mongolian allium compound is: 160 mg / L bitter bean extract-320 mg / L Mongolian allium extract;

[0080] The minimum inhibitory concentration of the Mongolian Allium Mongolianum-Astragalus oblique stem compound is: 160 mg / L Mongolian Allium Mongolianum extract-320 mg / L Astragalus oblique stem extract.

[0081] Example 7

[0082] An application of a compounding method of plant extracts with antibacterial efficacy, which is used to inhibit Streptococcus agalactiae; the minimum inhibitory concentration combination is:

[0083] The minimum inhibitory concentration of the sophora alopecuroides-astragalus oblique stem compound is: 160 mg / L sophora alopecuroides extract-160 mg / L astragalus oblique stem extract;

[0084] The minimum inhibitory concentration of the bitter bean-Mongolian allium compound is: 160 mg / L bitter bean extract-160 mg / L Mongolian allium extract;

[0085] The minimum inhibitory concentration of the Mongolian Allium Mongolianum-Astragalus Oblique Stem compound is: 160 mg / L Mongolian Allium Mongolianum extract-160 mg / L Astragalus Oblique Stem extract.

[0086] Experimental Section

[0087] Experiment 1

[0088] Determination of antibacterial effect: The antibacterial rates of Sophora alopecuroides extract, Allium mongolicum extract and Astragalus obliquus extract at different concentrations were determined using a 96-well plate, and the absorbance was measured at 600 nm to determine the minimum inhibitory concentration (MIC) of the extract.

[0089] Culture medium preparation: Take 15 g of tryptone soy broth and tryptone soy agar, dissolve them in ultrapure water, and make up to 500 mL. Sterilize them with high-pressure steam at 121°C for 20 min to prepare 30 g / L tryptone soy broth culture medium and tryptone soy agar culture medium, respectively.

[0090] Preparation of bacterial solution: The test bacteria were purchased from Shanghai Xinyu Biotechnology Co., Ltd. The three purchased bacterial solutions (Escherichia coli, Staphylococcus aureus, and Streptococcus agalactiae) were thawed and inoculated on tryptone soy agar plate medium respectively. After culturing in a 37°C incubator for 24 h, a single colony was picked and placed in tryptone soy broth medium (5% calf serum was added to the Streptococcus agalactiae medium). After culturing in a 37°C incubator for 24 h, 10 uL of the bacterial solution was taken from it and placed in a centrifuge tube. Sterile water was added to dilute it to 100 times. 10 uL of the diluted bacterial solution was taken and counted in a hemocytometer. The bacterial concentration after dilution was 1.0×10 6 cfu / mL.

[0091] Antibacterial test of single plant extract: Three plant extract solutions (sweet bean extract, Mongolian Allium extract, and Astragalus obliquus extract) were mixed with tryptone soy broth medium (containing 1% Tween 80 and 2% DMSO) at a volume ratio of 3:2 to prepare a plant extract medium dilution with a concentration of 2400 mg / L; 120 uL of plant extract medium dilution was added to the first and second wells of a 96-well plate, and 120 uL of broth medium was added to wells 2-8. After mixing the plant extract medium dilution and broth medium in well 2, 120 uL of liquid was transferred to well 3. After mixing the solution in well 3, 120 uL of liquid was transferred to well 4. uL of liquid was transferred to the 4th well, and the plant extract was diluted step by step until the 8th well. 120uL of liquid was removed from the 8th well and discarded; the concentrations of plant extracts in wells 1-8 were 2400, 1200, 600, 300, 150, 75, 37.5, and 18.75 mg / L, respectively, and 30 uL of bacterial solution was added to each well, so that the volume of each well was 150uL and the bacterial solution concentration was about 1.0×10 6cfu / mL, 90% ethanol was used as blank control, gentamicin and norfloxacin solution were used as positive control, and cultured in a 37°C incubator for 24 hours. After the culture was completed, the growth of microorganisms in each well was observed, and the minimum antibiotic concentration that inhibited the growth of microorganisms, i.e., the minimum inhibitory concentration (MIC), was recorded. The absorbance of the culture solution at 600 nm was measured, and the inhibition rate was calculated according to the following formula.

[0092]

[0093] Antibacterial test after compounding of Sophora alopecuroides extract, Allium mongolicum extract and Astragalus obliquus extract: The operation was similar to the above steps, and the concentration gradient of plant extracts in the 96-well plate was adjusted to 5120, 2560, 1280, 640, 320, 160, 80, 40, 20, 10, and 5 mg / L. The three plant extracts were compounded in pairs using the chessboard method, and the MIC values ​​of each extract were 1 / 8, 1 / 4, 1 / 2, 1 times, and 2 times respectively. The schematic table is as follows, 1# and 2# represent two different extracts, respectively.

[0094] Checkerboard Dilution Method:

[0095]

[0096] Ensure that the volume of liquid added to each well of the 96-well plate is consistent, and make 3 parallels for each well. Calculate the fractional inhibition concentration (FIC) according to the following formula to judge the antibacterial effect after compounding.

[0097]

[0098] When FIC≤0.5, it means that the antibacterial effects of the two plant extracts are synergistic; when 0.5<FIC≤1, it means that the antibacterial effects of the two plant extracts are additive; when 1<FIC≤2, it means that the antibacterial effects of the two plant extracts are unrelated; when FIC>2, it means that the antibacterial effects of the two plant extracts are antagonistic.

[0099] Antibacterial test of mixed use of three extracts of Sophora alopecuroides, Allium mongolicum and Astragalus obliquus: The operation was similar to the above steps. The concentration gradient of plant extracts in the 96-well plate was set to 5120, 2560, 1280, 640, 320, 160, 80, 40, 20, 10, and 5 mg / L. The three plant extracts were compounded using the chessboard method and combined at 1 / 8, 1 / 4, 1 / 2, 1 times, and 2 times the MIC value of each extract. The schematic table is as follows, 1#, 2#, and 3# represent three different extracts, respectively.

[0100] Checkerboard Dilution Method:

[0101]

[0102] Table 1 Minimum inhibitory concentrations (MICs) of extracts from Sophora alopecuroides, Allium mongolicum and Astragalus obliquus against three tested strains

[0103]

[0104] As can be seen from Table 1, when the concentration of bitter bean extract, Mongolian leek extract, and Astragalus oblique stem extract is 320 mg / L, they have a strong inhibitory effect on Escherichia coli, Staphylococcus aureus, and Streptococcus agalactiae, and their minimum inhibitory concentration is 320 mg / L. At 300 mg / L, the inhibition rates of norfloxacin and gentamicin on Escherichia coli were 93.92% and 93.86%, respectively, on Staphylococcus aureus, and on Streptococcus agalactiae, respectively. It can be seen that when the concentration of bitter bean extract, Mongolian leek extract, and Astragalus oblique stem extract is 300 mg / L, it is comparable to the antibiotics norfloxacin and gentamicin in the inhibition of Escherichia coli ( E. coli ), Staphylococcus aureus ( S.aureus ) and Streptococcus agalactiae ( S. agalactiae ) had a similar inhibitory effect, and the best antibacterial state was achieved when the concentration was 320 mg / L.

[0105] The antibacterial effects of the combination of Sophora alopecuroides and Allium mongolianum extracts are shown in Tables 2.1-2.3.

[0106] Table 2.1 Antibacterial results of the combination of Sophora alopecuroides and Astragalus obliquus (Escherichia coli)

[0107]

[0108] Table 2.2 Antibacterial results of bitter bean-astragalus obliquus compound (Staphylococcus aureus)

[0109]

[0110] Table 2.3 Antibacterial results of bitter bean-astragalus complex (Streptococcus agalactiae)

[0111]

[0112] Tables 2.1-2.3 show that the combination of Sophora alopecuroides and Astragalus obliquus has a significant effect on Escherichia coli ( E. coli ), Staphylococcus aureus ( S.aureus ) and Streptococcus agalactiae ( S. agalactiae ) with a volume ratio of 1:1. When the combination of bitter bean and astragalus obliquus was used, the inhibitory effect on Escherichia coli ( E. coli ) has a minimum inhibitory concentration of 160 mg / L (1 / 2 MIC) of Sophora alopecuroides -160 mg / L (1 / 2 MIC) of Astragalus obliquus, S.aureus The minimum inhibitory concentration was 160 mg / L (1 / 2 MIC) Sophora alopecuroides-160 mg / L (1 / 2 MIC) Astragalus obliquus, which was effective against Streptococcus agalactiae ( S. agalactiae ) was 160 mg / L (1 / 2 MIC) Sophora alopecuroides - 160 mg / L (1 / 2 MIC) Astragalus obliquus.

[0113] Table 2.4 shows the composite FIC index of Sophora alopecuroides-Astragalus obliquus extract.

[0114] Table 2.4 FIC index of the combination of Sophora alopecuroides and Astragalus obliquus

[0115]

[0116] The FIC criteria show that the FIC values ​​of the combination of Sophora alopecuroides and Astragalus obliquus against the three tested bacteria were all >0.5 and ≤1, so the combined effect of the two extracts showed a single additive effect. Therefore, the minimum inhibitory concentration of the two extracts when combined was reduced to 1 / 2MIC (160 mg / L), which was at least doubled compared to the concentration when used alone, improving its inhibitory effect on Escherichia coli ( E. coli ), Staphylococcus aureus ( S.aureus )、Streptococcus agalactiae( S. agalactiae )’s inhibitory effect.

[0117] The antibacterial effects of the combination of Sophora alopecuroides and Allium mongolianum extracts are shown in Tables 3.1-3.3.

[0118] Table 3.1 Antibacterial results of bitter bean-Mongolian leek compound (Escherichia coli)

[0119]

[0120] Table 3.2 Results of the combination of bitter bean and Mongolian leek (Staphylococcus aureus)

[0121]

[0122] Table 3.3 Results of the combination of bitter bean and Mongolian leek (Streptococcus agalactiae)

[0123]

[0124] Table 3.1-3.3 shows the effects of the compound extract of Sophora alopecuroides and Allium mongolianum on Escherichia coli ( E. coli ), Staphylococcus aureus ( S. aureus ) and Streptococcus agalactiae ( S. agalactiae ) with a volume ratio of 1:1; the combined effect of bitter bean-Mongolian allium compound on Escherichia coli ( E. coli ) has a MIC of 160 mg / L (1 / 2 MIC) of Sophora alopecuroides - 80 mg / L (1 / 4 MIC) of Allium mongolicum, against Staphylococcus aureus ( S. aureus ) has a MIC of 160 mg / L (1 / 2 MIC) Sophora alopecuroides - 320 mg / L (1 MIC) Mongolian leek, against Streptococcus agalactiae ( S. agalactiae ) is 160 mg / L (1 / 2 MIC) Sophora alopecuroides - 160 mg / L (1 / 2MIC) Mongolian leek.

[0125] The FIC index of the compound extracts of Sophora alopecuroides and Allium mongolianum is shown in Table 3.4:

[0126] Table 3.4 Antibacterial FIC index of bitter bean-Mongolian leek compound

[0127]

[0128] Combined with the FIC criteria, it can be seen that the bitter bean-Mongolian allium compound agent has a strong inhibitory effect on Escherichia coli ( E. coli ) and Streptococcus agalactiae ( S. agalactiae ) The FIC value of the tested bacteria was >0.5 and ≤1, so the inhibitory effect was additive. S. aureus ) The FIC value of the tested bacteria is greater than 1, and the inhibitory effect is no relevant effect.

[0129] The antibacterial effects of the compounding of Astragalus obliquus and Allium mongolicum extracts are shown in Tables 4.1-4.4.

[0130] Table 4.1 Antibacterial results of Mongolian chives-Astragalus oblique stem compound (Escherichia coli)

[0131]

[0132] Table 4.2 Antibacterial results of Mongolian chive-Astragalus oblique stem compound (Staphylococcus aureus)

[0133]

[0134] Table 4.3 Antibacterial results of Mongolian chives-Astragalus obliquus compound (Streptococcus agalactiae)

[0135]

[0136] Tables 4.1 - 4.3 show the effects of Mongolian Allium and Astragalus obliquus compound on Escherichia coli ( E. coli ), Staphylococcus aureus ( S.aureus ) and Streptococcus agalactiae ( S. agalactiae) with a volume ratio of 1:1; the combined effect of Mongolian Allium and Astragalus obliquus compound on Escherichia coli ( E. coli ) has a MIC of 160 mg / L (1 / 2MIC) Mongolian Allium - 160 mg / L (1 / 2MIC) Astragalus oblique stem, against Staphylococcus aureus ( S. aureus ) has a MIC of 160 mg / L (1 / 2MIC) Mongolian Allium - 320 mg / L (1MIC) Astragalus obliquus, which is effective against Streptococcus agalactiae ( S. agalactiae ) has a MIC of 160 mg / L (1 / 2MIC) Mongolian Allium - 160 mg / L (1 / 2MIC) Astragalus obliquus.

[0137] The FIC index of Mongolian Allium and Astragalus obliquus compound is shown in Table 4.4.

[0138] Table 4.4 FIC index of Mongolian Allium and Astragalus obliquus compound

[0139]

[0140] Combined with the FIC value judgment standard, it can be seen that the Mongolian Allium-Astragalus oblique stem compound has a good effect on Staphylococcus aureus ( S. golden ) The FIC value of the tested bacteria was greater than 1, so the combined effect of the two extracts showed an irrelevant effect. The FIC values ​​of the other two bacteria were greater than 0.5 and ≤ 1, showing an additive effect, and when the two extracts acted together, their MIC dropped to 1 / 2MIC, effectively improving the antibacterial effect of the two plant extracts.

[0141] The antibacterial effects of the compounding of the three plant extracts of Sophora alopecuroides, Allium mongolicum and Astragalus obliquus are shown in Tables 5.1-5.3.

[0142] Table 5.1 Antibacterial results of bitter bean-Mongolian leek-Astragalus obliquus compound (Escherichia coli)

[0143]

[0144] Table 5.2 Antibacterial results of bitter bean-Mongolian leek-Astragalus obliquus compound (Staphylococcus aureus)

[0145]

[0146] Table 5.3 Antibacterial results of bitter bean-Mongolian leek-Astragalus obliquus compound (Streptococcus agalactiae)

[0147]

[0148] Table 5.1 shows the effects of the compound of Sophora alopecuroides-Allium Mongolianum-Astragalus obliquus on Escherichia coli ( E. coli) and the combined effect of bitter bean-Mongolian leek-Astragalus obliquus on Escherichia coli ( E. coli ) was 320 mg / L (1MIC) Sophora alopecuroides - 320 mg / L (1MIC) Mongolian Allium - 160 mg / L (1 / 2MIC) Astragalus obliquus. Compared with the three plant extracts mixed in pairs, the E. coli ) The MIC combinations of: 160 mg / L (1 / 2MIC) Sophora alopecuroides-80 mg / L (1 / 4MIC) Allium mongolicum, 160 mg / L (1 / 2MIC) Sophora alopecuroides-320 mg / L (1MIC) Astragalus oblique stem, 160 mg / L (1 / 2MIC) Allium mongolicum-160mg / L (1 / 2MIC) Astragalus oblique stem, the MIC of the three after combination did not decrease significantly.

[0149] Table 5.2 shows the combined effect of bitter bean-Mongolian leek-Astragalus obliquus compound on Staphylococcus aureus ( S. golden ) had an antibacterial effect. The MIC of the combination of bitter bean, Mongolian leek and oblique stem astragalus was 320 mg / L (1MIC) bitter bean - 320mg / L (1MIC) Mongolian leek - 320 mg / L (1MIC) oblique stem astragalus. Compared with the combination of the three plant extracts, the antibacterial effect of the three plant extracts on Staphylococcus aureus ( S. aureus ) The MIC combinations of: 160 mg / L (1 / 2MIC) Sophora alopecuroides-320 mg / L (1MIC) Allium mongolicum, 160 mg / L (1 / 2MIC) Sophora alopecuroides-160 mg / L (1 / 2MIC) Astragalus oblique stem, 160 mg / L (1 / 2MIC) Allium mongolicum-320 mg / L (1 / 2MIC) Astragalus oblique stem, the MIC of the three after combination did not decrease significantly.

[0150] Table 5.3 shows the combined effect of bitter bean-Mongolian leek-Astragalus obliquus compound on Streptococcus agalactiae ( S. agalactia ), the combined effect of bitter bean-Mongolian leek-Astragalus oblique stem had an MIC of 160 mg / L (1 / 2MIC) against Streptococcus agalactiae, bitter bean-320 mg / L (1MIC) Mongolian leek-160 mg / L (1 / 2MIC) Astragalus oblique stem, compared with the three plant extracts combined in pairs against Streptococcus agalactiae ( S. agalactiae ) MIC combinations of: 160 mg / L (1 / 2MIC) Sophora alopecuroides-160 mg / L (1 / 2MIC) Allium mongolicum, 160 mg / L (1 / 2MIC) Sophora alopecuroides-160 mg / L (1 / 2MIC) Astragalus oblique stem, 160mg / L (1 / 2MIC) Allium mongolicum-160 mg / L (1 / 2MIC) Astragalus oblique stem. The MIC of the three after combination did not decrease significantly.

[0151] Experiment 2

[0152] The antibiotics gentamicin and norfloxacin were used as positive controls to preliminarily screen the antibacterial effects of the three plant extracts. Figure 1-3 .

[0153] Figure 1 The results showed that the extracts of Sophora alopecuroides, Allium mongolicum and Astragalus obliquus at different concentrations could inhibit Escherichia coli ( E.coli ), and with antibiotics norfloxacin and gentamicin as positive controls, it can be seen from the figure that there are certain differences in the inhibitory effects of the three plant extracts on Escherichia coli (E. coli) at different concentrations. When the concentration of the bitter bean, Mongolian leek, and Astragalus oblique stem extracts was 300 mg / L, the inhibition rates of E. coli were 93.52%, 93.11%, and 93.07%, respectively. The inhibition rates of norfloxacin and gentamicin on E. coli were 93.92% and 93.86%, respectively, at 300 mg / L.

[0154] Figure 2 The results showed that the extracts of Sophora alopecuroides, Allium mongolicum and Astragalus obliquus at different concentrations could inhibit Staphylococcus aureus ( S.aureus ) inhibition rate, and with antibiotics norfloxacin and gentamicin as positive controls, it can be seen from the figure that the three plant extracts at different concentrations inhibited Staphylococcus aureus ( S.aureus ) had certain differences in the inhibitory effects. When the concentration of sophora alopecuroides, leek Mongolian and Astragalus obliquus extracts was 300 mg / L, the inhibition rates against Staphylococcus aureus were 94.73%, 94.41% and 94.37%, respectively. The inhibition rates of norfloxacin and gentamicin against Staphylococcus aureus were 95.06% and 95.01%, respectively, at 300 mg / L.

[0155] Figure 3 The results showed that the extracts of Sophora alopecuroides, Allium mongolicum and Astragalus obliquus at different concentrations could inhibit Streptococcus agalactiae ( S. agalactiae ) inhibition rate, and with antibiotics norfloxacin and gentamicin as positive controls, it can be seen from the figure that the three plant extracts at different concentrations inhibited Streptococcus agalactiae ( S. agalactiae ) had certain differences in their inhibitory effects. When the concentration of extracts from Sophora alopecuroides, Allium mongolicum, and Astragalus obliquus was 300 mg / L, they had no significant effects on Streptococcus agalactiae ( S. agalactiae ) were 90.71%, 90.69% and 90.80% respectively, and the inhibition rates of norfloxacin and gentamicin against Streptococcus agalactiae at 300 mg / L were 90.19% and 91.61% respectively.

Claims

1. A method for compounding a plant extract having antibacterial effect, characterized in that: The plant extract is prepared by compounding a bitter bean extract, a Mongolian allium extract and an oblique stem astragalus extract in pairs to prepare a bitter bean-Mongolian allium compound, a Mongolian allium-oblique stem astragalus compound and a bitter bean-oblique stem astragalus compound; The sophora alopecuroides-allium mongolianum compound is prepared by mixing the sophora alopecuroides extract and the allium mongolianum extract in a volume ratio of 1:1; The Mongolian Allium Mongolianum-Astragalus Oblique Stem compound is prepared by mixing Mongolian Allium Mongolianum extract and Astragalus Oblique Stem extract in a volume ratio of 1:1; The sophora alopecuroides-astragalus oblique stem compound is prepared by mixing the sophora alopecuroides extract and the astragalus oblique stem extract in a volume ratio of 1:1; The concentration of the sophora alopecuroides extract is 80 mg / L-160 mg / L; The concentration of the Mongolian Allium extract is 80 mg / L-320 mg / L; The concentration of the Astragalus obliquus extract is 80 mg / L-320 mg / L; The preparation method of the sophora alopecuroides extract / Allium mongolianum extract / Astragalus obliquus extract is as follows: (a) Freshly collected Sophora alopecuroides plants / Allium mongolicum plants / Astragalus obliquus plants were naturally air-dried, cut into 1-2 cm, and made into powder. 10 g of plant powder was weighed, and 90% ethanol solution was added according to a solid-liquid ratio of 20:

1. The mixture was heated and refluxed in a reflux device for 2 h. The supernatant was taken, and the above steps were repeated for 3 times, and the supernatants were combined. In the ethanol solution, V 乙醇 :V 水 =9:1; (b) After decolorization with petroleum ether, the mixture was evaporated in an evaporator to a volume of 1 / 4 of the original volume, and concentrated freeze-dried to obtain the Sophora alopecuroides extract / Allium mongolicum extract / Astragalus obliquus extract, which were stored at -20°C.

2. A method for compounding a plant extract with antibacterial effect as claimed in claim 1, characterized in that: The extract of bitter bean is pseudomorphine, atropine, theophylline, sophora flavescens, deoxytubule glycoside, lupine, quinidine, ciprofloxacin, spartinine, aloperidine, retamine, arecaline, fetidine, and thebaine; The Mongolian allium extract is gelatin and schisandra saponin; The extract of Astragalus obliquus contains isocarpioside, beta-ecdysone, atractylodesin, phytolaccaside and naringin.

3. The use of a product prepared by the compounding method of a plant extract with antibacterial effect as claimed in claim 2, characterized in that: Used to prepare drugs for inhibiting Escherichia coli, Staphylococcus aureus and Streptococcus agalactiae.

4. The use of a product prepared by the compounding method of a plant extract with antibacterial effect as claimed in claim 3, characterized in that: The three plant extracts were mixed in pairs according to a volume ratio of 1:1, and the minimum inhibitory concentration combination for Escherichia coli was: The minimum inhibitory concentration of the sophora alopecuroides-astragalus oblique stem compound is: 160 mg / L sophora alopecuroides extract-160 mg / L astragalus oblique stem extract; The minimum inhibitory concentration of the bitter bean-Mongolian allium compound is: 160 mg / L bitter bean extract-80 mg / L Mongolian allium extract; The minimum inhibitory concentration of the Mongolian Allium Mongolianum-Astragalus oblique stem compound is: 160 mg / L Mongolian Allium Mongolianum extract-160 mg / L Astragalus oblique stem extract.

5. The use of the product prepared by the compounding method of a plant extract with antibacterial effect as claimed in claim 3, characterized in that: The three plant extracts were mixed in pairs according to a volume ratio of 1:1, and the minimum inhibitory concentration combination for Staphylococcus aureus was: The minimum inhibitory concentration of the sophora alopecuroides-astragalus oblique stem compound is: 160 mg / L sophora alopecuroides extract-160 mg / L astragalus oblique stem extract; The minimum inhibitory concentration of the bitter bean-Mongolian allium compound is: 160 mg / L bitter bean extract-320 mg / L Mongolian allium extract; The minimum inhibitory concentration of the Mongolian Allium Mongolianum-Astragalus oblique stem compound is: 160 mg / L Mongolian Allium Mongolianum extract-320 mg / L Astragalus oblique stem extract.

6. The use of a product prepared by the compounding method of a plant extract with antibacterial effect as claimed in claim 3, characterized in that: The three plant extracts were mixed in pairs according to a volume ratio of 1:1, and the minimum inhibitory concentration combination for Streptococcus agalactiae was: The minimum inhibitory concentration of the sophora alopecuroides-astragalus oblique stem compound is: 160 mg / L sophora alopecuroides extract-160 mg / L astragalus oblique stem extract; The minimum inhibitory concentration of the bitter bean-Mongolian allium compound is: 160 mg / L bitter bean extract-160 mg / L Mongolian allium extract; The minimum inhibitory concentration of the Mongolian Allium Mongolianum-Astragalus Oblique Stem compound is: 160 mg / L Mongolian Allium Mongolianum extract-160 mg / L Astragalus Oblique Stem extract.

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