Application of gynura bicolor extract in bacteriostasis
By studying different solvent extracts of *Indigofera tinctoria*, the antibacterial effects against various bacteria were verified, which solved the problem of the shortage of *Indigofera tinctoria* medicinal resources and provided a pharmacological basis for alternative medicinal materials.
Patent Information
- Application Number
- CN202511015738.X
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-07-23
- Publication Date
- 2025-10-17
AI Technical Summary
There is a shortage of *Indigofera tinctoria* medicinal resources, so it is necessary to find alternatives to ensure the supply of medicinal materials and maintain the antibacterial effect.
By studying different solvent extracts of *Lysimachia christinae*, including water extract, water-ethanol precipitate extract, and chloroform extract, we verified its antibacterial effects against *Bacillus subtilis*, *Staphylococcus aureus*, *Pseudomonas aeruginosa*, and *Escherichia coli*.
The extract of Guiyue Malan showed that it has a certain antibacterial effect against the above-mentioned bacteria, and has the potential to replace Malan medicinal material, providing pharmacological and efficacy support.
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Abstract
Description
TECHNICAL FIELD
[0001] The present application relates to the field of traditional Chinese medicine extraction, and particularly relates to application of Strobilanthes bantonensis Lindau extract in bacteriostasis. BACKGROUND
[0002] Baphicacanthus cusia (Nees) J.B.Imlay is a perennial herb of the family Acanthaceae, also known as Banlan, which is widely distributed in South China and West China, especially in Guangxi. The root and rhizome of Baphicacanthus cusia are used as medicine and called Banlan, which is bitter and cold in nature, and has the functions of clearing heat and detoxifying, and cooling blood and removing spots. The People's Republic of China Pharmacopoeia in 1999, 2000 and 2005 officially collects Banlan as Banlan and lists it as a kind of medicine. The leaf of Baphicacanthus cusia is also called Dacianye in many places in southern China, which is similar to the leaf of North Banlangen, and has the functions of clearing heat and detoxifying, cooling blood and stopping bleeding, and treating warm disease with heat, profuse thirst, influenza, acute infectious hepatitis, bacillary dysentery, acute gastroenteritis, acute pneumonia, erysipelas, hematemesis, epistaxis, jaundice, dysentery, laryngeal obstruction, aphtha, and carbuncle and sores. Pharmacological experiments show that the leaf of Baphicacanthus cusia has good bacteriostatic effect on Staphylococcus aureus and Klebsiella pneumoniae. As a commonly used traditional Chinese medicine, Baphicacanthus cusia has been widely used in various compound preparations. The compound Banlan granules have excellent effects in antiviral, anti-inflammatory and clearing aspects. Sun Xiaobing et al. [3] summarized the research on the chemical composition and pharmacological action of Baphicacanthus cusia in recent years, and Baphicacanthus cusia has significant effects in antibacterial, antitumor, antiviral, anti-inflammatory and immune promotion, and antipyretic aspects. Baphicacanthus cusia is widely used and in great demand. However, in recent years, investigations and researches show that the resources of Baphicacanthus cusia are rapidly decreasing, which is caused by unscientific harvesting of medicinal materials and planting of economic forests in various places, and has a great impact on the supply of medicinal resources for Baphicacanthus cusia related preparations. Therefore, the resource protection of Baphicacanthus cusia and the research and development of alternative medicinal materials are imminent.
[0003] Strobilanthes bantonensis Lindau, also known as purple back Banlangen, is an antiviral herb, mainly distributed in southern China and Vietnam. The medicinal characteristics and effects of Strobilanthes bantonensis Lindau and Baphicacanthus cusia are very similar, and Strobilanthes bantonensis Lindau is often used as a substitute for Baphicacanthus cusia in the treatment of wind-heat cold and viral infectious diseases in the Guangxi folk and the Sino-Vietnamese border area, and has good effects. At the same time, the planting technology of Strobilanthes bantonensis Lindau in the Guangxi region has become mature, and large-scale planting has been realized, so that the supply of medicinal resources is guaranteed. In addition, Strobilanthes bantonensis Lindau has greater advantages in price than Baphicacanthus cusia. In summary, Strobilanthes bantonensis Lindau has the basic conditions to replace Baphicacanthus cusia as a medicine. The present application intends to take Baphicacanthus cusia as a control, and investigate the bacteriostatic activity of Strobilanthes bantonensis Lindau extracts in different solvents, so as to provide theoretical basis and data support for its substitution as a medicine in the aspect of pharmacology and efficacy. SUMMARY
[0004] The present application intends to provide an application of Strobilanthes gueinzii extract in bacteriostasis to solve the problems in the background art.
[0005] To achieve the above-mentioned purpose, the present application provides the following technical solutions.
[0006] The application of Strobilanthes gueinzii extract in bacteriostasis, wherein the inhibited bacteria include Bacillus subtilis, Staphylococcus aureus, Pseudomonas aeruginosa and Escherichia coli.
[0007] Preferably, the application of Strobilanthes gueinzii extract in bacteriostasis, wherein the Strobilanthes gueinzii extract includes water extract, water extract alcohol precipitation extract and chloroform extract according to different extraction methods.
[0008] Compared with the prior art, the present application has the following beneficial effects:
[0009] The present application proves by experiments that Strobilanthes gueinzii has inhibitory effect on Bacillus subtilis, Staphylococcus aureus, Pseudomonas aeruginosa and Escherichia coli, and has certain bacteriostatic effect. Compared with Stachys sieboldii, the bacteriostatic activity of different solvent extracts of Strobilanthes gueinzii is investigated, and Strobilanthes gueinzii has the basic conditions to replace Stachys sieboldii. DETAILED DESCRIPTION
[0010] The present application will be further described in detail in combination with embodiments as follows:
[0011] The specific implementation process is as follows:
[0012] I. Test materials
[0013] 1.1. Instruments:
[0014] BSA124S type one-thousandth electronic analytical balance (Sartorius Company);
[0015] DZTM type temperature-controlled electric heating jacket (Jing City Yongguangming Medical Instrument Co., Ltd.);
[0016] RE-52AA rotary evaporator (Shanghai Yalong Biochemical Instrument Factory);
[0017] HH-6 digital display constant-temperature water bath (Jing City Yichen Instrument Manufacturing Co., Ltd.);
[0018] 70%-100% alcohol (Hebei Province (Jinghe City Jinghe Instrument and Meter Co., Ltd.);
[0019] YXQ-LS-Vertical pressure steam sterilizer (Shanghai Boxin Industrial Co., Ltd. Medical Equipment Factory);
[0020] SW-CJ-2FD clean bench (Suzhou Antai Air Technology Co., Ltd.);
[0021] LRH-150F biochemical incubator (Shanghai Qi Xin Scientific Instrument Co., Ltd.);
[0022] LQ-C6002 electronic balance (Rui'an Ante Weighing Equipment Co., Ltd.);
[0023] UV-1240 ultraviolet spectrophotometer (Shimadzu / China);
[0024] Trypticase soy broth liquid medium (TSB) (Guangdong Huan Kai Microbial Technology Co., Ltd., 1070791);
[0025] Trypticase soy agar medium (TSA) (Guangdong Huan Kai Microbial Technology Co., Ltd., 1070461)
[0026] 1.2, Reagents and drugs:
[0027] 0.9% sodium chloride solution (Shanghai Shenguo Bioengineering Co., Ltd.);
[0028] Bacillus subtilis [CMCC(B)63501], 2a23-1, Staphylococcus aureus [CMCC(B)26003], 5a30-2, Pseudomonas aeruginosa [CMCC(B)10104], 2a22-2, Escherichia coli [CMCC(B)44102], 3a26-2 were provided by China Food and Drug Inspection Institute, all of which were the 3rd generation strains;
[0029] Shrimp blue and guiyue shrimp blue were collected from Nanning suburb of Guangxi, and were identified as genuine products; The medicinal materials used in this study were dry leaves of shrimp blue and guiyue shrimp blue.
[0030] II. Test process and results
[0031] 2.1, Preparation of test samples
[0032] 2.1.1, Preparation of conventional samples of different extraction methods of shrimp blue and guiyue shrimp blue roots and leaves
[0033] 2.1.1.1, Water extract of shrimp blue and guiyue shrimp blue
[0034] Preparation of shrimp blue water extract sample: take 100g of shrimp blue medicinal material coarse powder, add water and reflux twice, the first time is 1.5h, the second time is 1h, filter the extract, combine the filtrate, concentrate to a certain volume, evaporate in a water bath (80℃) evaporating dish, and concentrate to a weight that does not decrease, i.e. shrimp blue water extract 39.06g, numbered 1;
[0035] Preparation of the water extract of the root and leaf of Strobilanthes gueinzii: 100 g of crude powder of Strobilanthes gueinzii was refluxed twice, 1.5 h for the first time and 1 h for the second time. The extract was filtered, the filtrates were combined, and concentrated to a certain volume. The concentrated extract was evaporated in a water bath (80°C) until the weight no longer decreased, and thus 34.13 g of the water extract of the root and leaf of Strobilanthes gueinzii was obtained, numbered 2.
[0036] 2.1.1.2. Water extract of Strobilanthes dyerianus and Strobilanthes gueinzii
[0037] Preparation of the water extract of Strobilanthes dyerianus: 100 g of crude powder of Strobilanthes dyerianus was refluxed twice, 1.5 h for the first time and 1 h for the second time. The extract was filtered, the filtrates were combined, and concentrated to a certain volume. The concentrated extract was evaporated in a water bath (80°C) until the weight no longer decreased, and thus 21.37 g of the water extract of Strobilanthes dyerianus was obtained, numbered 3.
[0038] Preparation of the water extract of Strobilanthes gueinzii: 100 g of crude powder of Strobilanthes gueinzii was refluxed twice, 1.5 h for the first time and 1 h for the second time. The extract was filtered, the filtrates were combined, and concentrated to a certain volume. The concentrated extract was evaporated in a water bath (80°C) until the weight no longer decreased, and thus 20.66 g of the water extract of the root of Strobilanthes gueinzii was obtained, numbered 4.
[0039] 2.1.1.3. 70% ethanol extract of Strobilanthes dyerianus and Strobilanthes gueinzii
[0040] Preparation of the 70% ethanol extract of Strobilanthes dyerianus: 100 g of crude powder of Strobilanthes dyerianus was repeatedly refluxed and extracted with 70% ethanol (4000 ml of 70% ethanol was added each time, and refluxed for 1 h). The extraction was repeated until the extract was basically colorless. The extract was filtered with a medium-speed filter paper, and the ethanol was recovered by rotary evaporation under reduced pressure. The concentrated extract was evaporated in a water bath (80°C) until the weight no longer decreased, and thus 15.72 g of the 70% ethanol extract of Strobilanthes dyerianus was obtained, numbered 5.
[0041] Preparation of the 70% ethanol extract of Strobilanthes gueinzii: 100 g of crude powder of Strobilanthes gueinzii was repeatedly refluxed and extracted with 70% ethanol (4000 ml of 70% ethanol was added each time, and refluxed for 1 h). The extraction was repeated until the extract was basically colorless. The extract was filtered with a medium-speed filter paper, and the ethanol was recovered by rotary evaporation under reduced pressure. The concentrated extract was evaporated in a water bath (80°C) until the weight no longer decreased, and thus 16.26 g of the 70% ethanol extract of Strobilanthes gueinzii was obtained, numbered 6.
[0042] 2.1.1.4. Petroleum ether extract of Strobilanthes dyerianus and Strobilanthes gueinzii
[0043] Preparation of petroleum ether extract of Indigofera kirilowii: Take 100g of crude powder of Indigofera kirilowii, repeatedly reflux extract with petroleum ether (add 4000ml of petroleum ether each time, reflux for 1h), until the extract is basically colorless, filter, get the residue (I). Combine the filtrate, recover the petroleum ether by rotary evaporation under reduced pressure, evaporate in evaporating dish in pure water bath (80℃) until the weight no longer decrease, get the petroleum ether extract of Indigofera kirilowii 1.56g, numbered as 7;
[0044] Preparation of petroleum ether extract of Indigofera kirilowii: Take 100g of crude powder of Indigofera kirilowii, repeatedly reflux extract with petroleum ether (add 4000ml of petroleum ether each time, reflux for 1h), until the extract is basically colorless, filter, get the residue (I). Combine the filtrate, recover the petroleum ether by rotary evaporation under reduced pressure, evaporate in evaporating dish in pure water bath (80℃) until the weight no longer decrease, get the petroleum ether extract of Indigofera kirilowii 1.56g, numbered as 7;
[0045] 2.1.1.5, chloroform extract of Indigofera kirilowii and Indigofera kirilowii var. sinensis
[0046] Preparation of chloroform extract of Indigofera kirilowii: Take the residue (I) under item "2.1.1.4", repeatedly reflux extract with chloroform (add 4000ml of chloroform each time, reflux for 1h), until the extract is basically colorless, filter, get the residue (III). Combine the filtrate, recover the chloroform by rotary evaporation under reduced pressure, evaporate in evaporating dish in pure water bath (80℃) until the weight no longer decrease, get the chloroform extract of Indigofera kirilowii 6.09g, numbered as 9;
[0047] Preparation of chloroform extract of Indigofera kirilowii: Take the residue (I) under item "2.1.1.4", repeatedly reflux extract with chloroform (add 4000ml of chloroform each time, reflux for 1h), until the extract is basically colorless, filter, get the residue (III). Combine the filtrate, recover the chloroform by rotary evaporation under reduced pressure, evaporate in evaporating dish in pure water bath (80℃) until the weight no longer decrease, get the chloroform extract of Indigofera kirilowii 6.09g, numbered as 9;
[0048] 2.1.1.5, chloroform extract of Indigofera kirilowii and Indigofera kirilowii var. sinensis
[0049] Preparation of acetone extract of Indigofera kirilowii: Take the residue (III) under item "2.1.1.5", repeatedly reflux extract with acetone (add 4000ml of acetone each time, reflux for 1h), until the extract is basically colorless, filter, get the residue (V). Combine the filtrate, recover the acetone by rotary evaporation under reduced pressure, evaporate in evaporating dish in pure water bath (80℃) until the weight no longer decrease, get the acetone extract of Indigofera kirilowii 11.32g, numbered as 11;
[0050] Preparation of the acetone extract of Strobilanthes guilgus: Take the residue (IV) in "2.1.1.5", repeatedly reflux extract with acetone (4000 ml of acetone is added each time, and reflux for 1 hour), until the extract is basically colorless, and then extract with acetone. Filter, combine the filtrate, and recover the acetone by rotary evaporation under reduced pressure. Evaporate and concentrate in a water bath (80°C) until the weight no longer decreases, to obtain the acetone extract of Strobilanthes guilgus 10.55 g, numbered 12.
[0051] 2.1.1.7, 95% ethanol extract of Strobilanthes jamaicensis
[0052] Preparation of the 95% ethanol extract of Strobilanthes jamaicensis: Take the residue (V) in "2.1.6", repeatedly reflux extract with 95% ethanol (4000 ml of 95% ethanol is added each time, and reflux for 1 hour), until the extract is basically colorless, and then extract with 95% ethanol. Filter, combine the filtrate, and recover the ethanol by rotary evaporation under reduced pressure. Evaporate and concentrate in a water bath (80°C) until the weight no longer decreases, to obtain the 95% ethanol extract of Strobilanthes jamaicensis 19.12 g, numbered 13.
[0053] Preparation of the 95% ethanol extract of Strobilanthes guilgus: Take the residue (VI) in "2.1.6", repeatedly reflux extract with 95% ethanol (4000 ml of 95% ethanol is added each time, and reflux for 1 hour), until the extract is basically colorless, and then extract with 95% ethanol. Filter, combine the filtrate, and recover the ethanol by rotary evaporation under reduced pressure. Evaporate and concentrate in a water bath (80°C) until the weight no longer decreases, to obtain the 95% ethanol extract of Strobilanthes guilgus 18.36 g, numbered 14.
[0054] 2.2, Bacteriostatic test
[0055] The commonly used methods for in vitro bacteriostatic test include the paper disc diffusion method and the agar dilution method. The agar plate punching method is similar in principle to the paper disc diffusion method. After the quantitative drug dissolves in the water in the agar plate, it continuously diffuses to the surrounding area of the paper disc, forming a gradient concentration, and the growth of bacteria in the bacteriostatic concentration range around the drug is inhibited, forming a transparent and smooth bacteriostatic ring. This method can observe the sensitivity or drug resistance of one bacterium to multiple traditional Chinese medicine ingredients on the same plate, and the experimental results are very intuitive, and it is easy to judge and compare the inhibition effects of multiple traditional Chinese medicine ingredients on the same bacterium. The agar plate punching method is used in the test.
[0056] 2.2.1, Preparation of culture medium
[0057] 2.2.1.1, Preparation of liquid culture medium:
[0058] An appropriate amount of TSB culture medium powder is weighed and added to the corresponding amount of distilled water in a ratio of 40 g:1000 ml, shaken, plugged, placed in a steam sterilization pot, sterilized at 121°C for 15 min, and used at room temperature.
[0059] 2.2.1.2. Preparation of solid culture medium:
[0060] An appropriate amount of TSA medium powder was weighed and added to the corresponding amount of distilled water at a ratio of 40 g: 1000 ml, shaken, stoppered, placed in a steam sterilization pot, sterilized at 121°C for 15 min, poured into a flat plate, and refrigerated for standby use.
[0061] 2.2.2. Preparation of test drugs
[0062] The extracts 1-14 under item “2.1.1” were freeze-dried, taken out, and ground into fine powder. An appropriate amount of each extract powder was taken, water was added to prepare a solution or suspension of 50 mg / mL, which was used as a test drug.
[0063] 2.2.3. Measurement of the diameter of the inhibition zone
[0064] The diameters (d) of the inhibition zones of the extracts 1-14 were measured by the agar plate punching method. 100 μL of bacterial solution with a concentration of 1 x 108CFU / mL was evenly applied to the plate, then two sample holes were punched in the same plate using a puncher (diameter: 6 mm), the agar block was picked out, and 35 μL of the traditional Chinese medicine extract sample solution was injected into the hole using a syringe. Each bacterium was repeated 3 times for parallel test. After the plate was placed in a constant temperature incubator at 37°C for 18 h, the diameter of the inhibition zone was measured using a digital vernier caliper (crossed repeatedly for 3 times), and the average value was taken. Pure water was used as the negative control, and penicillin (0.005 mg / mL) was used as the positive control. The data were processed by SPSS 28.0 statistical software, the differences in the antibacterial effects of different extracts of Indigofera tinctoria and I. decora were compared, Mann-Whitney U test was used, and P<0.05 indicated that the difference was statistically significant. The test results are shown in Table 1.
[0065] Table 1. In vitro antibacterial test results of extracts 1-14
[0066]
[0067]
[0068] Note: A-Indigofera tinctoria; B-I. decora
[0069] As shown in Table 1, the two plant water extracts, water extraction alcohol precipitation extracts and chloroform extracts have certain inhibitory effect on the three bacteria of Bacillus subtilis, Staphylococcus aureus and Pseudomonas aeruginosa, the minimum inhibitory diameter is (5.41±0.74) mm, the maximum is (16.99±0.12) mm, and the inhibitory effect has no significant difference (P>0.05); the two plant petroleum ether extracts have inhibitory effect on Staphylococcus aureus and Pseudomonas aeruginosa, the minimum inhibitory diameter is (5.32±0.34) mm, the maximum is (7.70±0.36) mm, and the inhibitory effect has no significant difference (P>0.05); the two plant acetone extracts and different concentration ethanol extracts have inhibitory effect on Bacillus subtilis and Staphylococcus aureus, the minimum inhibitory diameter is (5.94±0.18) mm, the maximum is (11.45±0.54) mm, and the inhibitory effect has no significant difference (P>0.05); the 70% ethanol extract and 95% ethanol extract of E. guettardii have inhibitory effect on Escherichia coli, the inhibitory diameters are (6.42±0.54) mm and (7.74±0.55) mm respectively, and the other extracts have no obvious inhibitory effect on Escherichia coli.
[0070] 2.3、Conclusion
[0071] The bacteriostatic effect test is carried out based on different extracts of E. guettardii and E. guettardii. In order to avoid the interference of solvent on the bacteriostatic effect, pure water is used as negative control. The volatile solvent is completely removed in the preparation of extract, so as to ensure the reliability of bacteriostatic data. The present study proves that the different solvent extracts of the two plants have weak inhibitory effect on Escherichia coli, and have certain inhibitory effect on the three bacteria of Bacillus subtilis, Staphylococcus aureus and Pseudomonas aeruginosa, and the inhibitory effect has no obvious difference.
[0072] The above is only an embodiment of the present application, and the specific technical solutions and / or characteristics of the scheme are not described in detail. It should be noted that for those skilled in the art, without departing from the technical solutions of the present application, some modifications and improvements can be made, which should also be regarded as the protection scope of the present application, which will not affect the effect and practicality of the patent. The protection scope of the present application should be subject to the content of its claims, and the specific implementation mode and the like in the description can be used to explain the content of the claims.
Claims
1. An application of a Kuiyuemai indigo extract in antibacterial treatment, characterized in that: The inhibited bacterial species include Bacillus subtilis, Staphylococcus aureus, Pseudomonas aeruginosa and Escherichia coli.
2. The use of the indigofera truncatula extract in antibacterial treatment according to claim 1, wherein: The indigofera indigofera extract includes water extract, water-extracted alcohol-precipitated extract and chloroform extract according to different extraction methods.