CN101757140B - Medicament with spermicidal effect in vitro and bacteriostatic action, and preparation method and application thereof - Google Patents
Medicament with spermicidal effect in vitro and bacteriostatic action, and preparation method and application thereof Download PDFInfo
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- CN101757140B CN101757140B CN2010101304824A CN201010130482A CN101757140B CN 101757140 B CN101757140 B CN 101757140B CN 2010101304824 A CN2010101304824 A CN 2010101304824A CN 201010130482 A CN201010130482 A CN 201010130482A CN 101757140 B CN101757140 B CN 101757140B
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Abstract
The invention provides a medicament with a spermicidal effect in vitro and a bacteriostatic action, which is characterized in that the medicament is an extract prepared by an ethanol heating reflux method after mixing radix sophorae flavescentis and root of Chinese pulsatilla in a mass ratio of 1:3 to 1:4. The invention discloses that: when the mass ratio of the radix sophorae flavescentis to the root of Chinese pulsatilla is 1 to 3 and 1 to 4, the spermicidal effect is stronger than a single medicament; and the spermicidal effect is strongest when the mass ratio is 1 to 3. The bacteriostatic action of the composite extract of the radix sophorae flavescentis and the root of Chinese pulsatilla is better than that of the extract of the single medicament. The medicament of the invention has the spermicidal effect in vitro and the bacteriostatic action which are better than that of the extract of the single medicament.
Description
Technical field
The invention belongs to field of medicaments, relate to medicine with the essence of killing and bacteriostasis and its production and application.
Background technology
China is populous, and practicing family planning is the fundamental state policy of country.Practicing family planning is based on contraception, and contraceptives commonly used at present have oral contraceptive, external applied contraceptive, condom, intrauterine device, sterilization operation etc.Oral contraceptive copies the human hormone to make more, often takes the homergy that can influence hormone in vivo, produces a series of side effect.Though continuous progress along with industrial level, the thickness of condom is also more and more littler, but the pleasant sensation may make sexual intercourse the time weakens, and also can cause in the sexual intercourse process that condom breaks or sexual intercourse is retained in cover in the vagina when finishing owing to improper use, make seminal fluid flow into vagina, thereby cause contraceptive failure.Therefore the research research and development of external vagina contraceptive that have the prevention and treatment of venereal diseases effect concurrently are significant, are subjected to extensive attention.
Summary of the invention
Task of the present invention provides a kind of medicine with external essence extremely and bacteriostasis, and the preparation method of this medicine is provided.
Realize that technical scheme of the present invention is:
This external medicine that kills smart and bacteriostasis that has provided by the invention is the Radix Sophorae Flavescentis and the Radix Pulsatillae by mass ratio is that 1: 3～1: 4 proportioning is mixed the extract that the back adopts ethanol extraction method to produce, the concrete employing of the embodiment of the invention be the alcohol heating reflux method.
This name provides thisly has external preparation method of killing smart and bacteriostasis medicine and is: after getting the Radix Sophorae Flavescentis and Radix Pulsatillae placement drying baker inner drying, is 1: 3～1: 4 with the Radix Sophorae Flavescentis and the Radix Pulsatillae by mass ratio, optimum ratio is that 1: 3 proportioning is mixed, get Radix Sophorae Flavescentis and Radix Pulsatillae mixture, measure heating and refluxing extraction 3 times, each 2 hours for 8 times with 60% ethanol; Collect 3 extracting solution of merging, filter, heating concentrates, and drips dilute NaOH solution setting pH to 7.2～7.5, and being settled to liquor strength is 1: 1, is equivalent to every ml medicinal liquid and contains crude drug in whole 1g, and at 121 ℃ of following high temperature sterilize 20min, packing is standby then.
The present invention discloses, and Radix Sophorae Flavescentis and Radix Pulsatillae proportioning are 1: 3～1: 4 o'clock, kill smart acting on and be better than single medicine, with proportioning be 1: 3 o'clock the strongest.Radix Sophorae Flavescentis, Radix Pulsatillae compound extract fungistatic effect are all good than single medicament extracting solution.Medicine of the present invention has external essence and the bacteriostasis of killing concurrently, and kills smart and bacteriostasis all is better than single medicament extracting solution.
Experimental data
1. method
1.1 Radix Sophorae Flavescentis, Radix Pulsatillae compound extract preparation
1.1.1 material: Radix Pulsatillae root powder, Radix Sophorae Flavescentis, NaOH solution, distilled water
1.1.2 preparation method:
9 groups of extracting solution that prepare following quality proportioning:
| Extracting solution | Radix Sophorae Flavescentis, Radix Pulsatillae proportioning (mass ratio) |
| 1 | 1∶0 |
| 2 | 1∶1 |
| 3 | 0∶1 |
| 4 | 1∶4 |
| 5 | 1∶3 |
| 6 | 1∶2 |
| 7 | 2∶1 |
| 8 | 3∶1 |
| 9 | 4∶1 |
Radix Pulsatillae root powder and Radix Sophorae Flavescentis are placed the drying baker inner drying, and each group is got 10g Radix Sophorae Flavescentis and Radix Pulsatillae mixture or single medicine by last table quality proportioning.Get 8 times of amounts of 60% ethanol heating and refluxing extraction 3 times, each 2h; Collect to merge 3 extracting solution, filter, heating drips dilute NaOH solution when being concentrated into not enough 10ml and sets up pH to 7.2～7.5, standardize solution 10ml, and this moment, liquor strength was 1: 1, was equivalent to every ml medicinal liquid and contained crude drug in whole 1g; Then at 121 ℃ of following high temperature sterilize 20min, packing standby [referring to Xu Jinning, Zhang Heming, Zhao Xiuhua. the progress of matrine extraction process and several formulations. the time precious traditional Chinese medical science traditional Chinese medicines, 2007,18 (9): 2287-2289; Liu Wen, Feng fruit, Liu Mengying etc. the research of Radix Pulsatillae Decoction technology for extracting effective component. the time precious traditional Chinese medical science traditional Chinese medicines, 2007,18 (9): 60-63; Zhang Kuiyuan, Yu Shichun, Xie Dongmei. the optimized choice of Radix Sophorae Flavescentis extraction process. basic unit's Chinese medicine magazine, 2002,16 (1): 13-14; Liu Lin, Jin Fengxie. the separation of pulchinenoside is purified. Dalian Polytechnic College journal, 23,1:18-21].
1.2 the external essence of killing acts on
1.2.1 sperm suspension is chosen: healthy male, the age is no more than 40 years old, ascetic 4～7d, 48h alcohol prohibition and coffee etc. before the sampling, the masturbation method is got essence.
1.2.2 with area of computer aided semen analysis (CASA) detection of drugs to motility of sperm.
10 selected routine semen samples are:
Sperm concentration:＞80*10^6/ml
Motility rate: a level＞30%, a level+b level＞50%
PH：7.2
Liquefying time:＜30min
Test method:
Seminal fluid and medicinal liquid are mixed and made into the mixed liquor of different liquor strengths with different volume ratios, observe 5 above visuals field at smear behind the mixing fast, observe terminal point for mixing back 20s, if each visual field sperm motility rate＜1%, think that then the medicinal liquid of this concentration has a moment and kills smart effect [referring to Mu Yi, Du person of outstanding talent. pulchinenoside is external to kill smart effect research. northwest pharmaceutical journal, 1996,11 (3): 119-120; Du person of outstanding talent, Guo Renyu, Liu Shengjia etc. EXPERIMENTAL STUDIES ON MATRINE KILLING SPERM IN VITRO. Xian Medical Univ's journal, 1996,3 (17): 338-339].
1.3 observe colibacillary bacteriostasis with the susceptibility paper disk method
1.3.1 the preparation of drug sensitive test paper: common qualitative filter paper is broken into some circular papers with puncher, diameter is 6mm, place dried and clean in vitro, put into high-pressure sterilizing pot through 121 ℃ of autoclavings after 20 minutes, take out and put into thermostatic drying chamber and carry out drying for standby.
1.3.2 the preparation of agar culture medium: 1, prescription: Carnis Bovis seu Bubali cream 3.0g, peptone 10.0g, sodium chloride 5.0g, agar 18.0g, distilled water 1L, dilute NaOH solution.2, preparation method: to small beaker, add to pour in the large beaker that peptone and sodium chloride powder are housed after an amount of distilled water heating for dissolving and heat with Glass rod picking Carnis Bovis seu Bubali cream.Treat to put into agar after medicine dissolves fully, make it to dissolve, regulate PH7.4～7.6, be settled to 1L.The culture medium that makes is poured in the cultivation face while hot, makes agar height 4～5mm, solidifies back 121 ℃ of autoclaving 20min.After finishing, sterilization treats that it solidifies, puts into 37 ℃ of incubators and cultivate 24h that the 4 ℃ of refrigerators of putting into of no bacterial growth are preserved standby.
1.3.3 colibacillary preparation: obtain colibacillary type strain from Microbiological Lab and be inoculated in the agar culture medium surface with method of scoring and go down to posterity 2 times, make its activation.
Make suspension 1.3.4 will activate good coli strain with sterile purified water, at microscopically count fine bacteria concentration, adjust concentration to 10 with blood cell counting plate
5Individual/ml.Draw 100ul bacterium drop and media surface with micro sample adding appliance, even with the Glass rod shop.The scraps of paper of will sterilizing again soak with variant medicinal liquid in about 10 seconds, take out the scraps of paper and scrape even each agar culture medium surface that is affixed at interval, dried back slightly, put into 37 ℃ of constant incubators, take out after 24 hours, with the vernier caliper measurement inhibition zone [referring to Shi Weijing, Lu Zhenxiang, burnt female. the different extract extracorporeal bacteria inhibitor tests researchs of Radix Pulsatillae root. the Chinese veterinarian medical magazine, 2005,4:26-28; Li Wenguang, Zhan Zhinong, Zhou Zhenjian. the laboratory observation of Radix Sophorae Flavescentis effervescent tablet vitro antibacterial activity. Hainan Medical College's journal 2001,7 (2): 68～69].
2. result
2.1 externally kill smart result
Radix Sophorae Flavescentis, the Radix Pulsatillae extracting solution of different proportionings all have external moment to kill the effect of sperm.Its minimum is killed smart concentration and is slightly fluctuateed because of density, vigor, the individual variation of sperm.Wherein Radix Sophorae Flavescentis, Radix Pulsatillae proportioning are 1: 3 and 1: 4 o'clock, kill smart effect and be better than single medicine, with proportioning be 1: 3 o'clock the strongest.See Table 1.
Table 1 Radix Sophorae Flavescentis, Radix Pulsatillae compound extract be external to kill smart result
ED
80: the Cmin when expression is can be to 80% sample effective
2.2 bacteriostatic test result
Radix Sophorae Flavescentis, Radix Pulsatillae compound extract fungistatic effect are all good than single medicament extracting solution.The external fungistatic effect of single medicine is very little, and proportioning is that 1: 1 compound extract fungistatic effect is best, and proportioning is 1: 4～1: 2 taking second place.See Table 2, Fig. 1～8.Fungistatic effect still was 1: 1 compound extract far away from proportioning after Radix Sophorae Flavescentis, the single medicament extracting solution of the Radix Pulsatillae mixed by 1: 1.See Fig. 9.
Table 2 Radix Sophorae Flavescentis, Radix Pulsatillae compound extract fungistatic effect
" +++": inhibition zone D 〉=10.00mm " ++ ": inhibition zone D 〉=7.50mm, D＜10.00mm; "+": inhibition zone is arranged but D＜7.50mm; "-": no inhibition zone
China is populous, and practicing family planning is the fundamental state policy of country.Practicing family planning is based on contraception, and contraceptives commonly used at present have oral contraceptive, external applied contraceptive, condom, intrauterine device, sterilization operation etc.Oral contraceptive copies the human hormone to make more, often takes the homergy that can influence hormone in vivo, produces a series of side effect.Though continuous progress along with industrial level, the thickness of condom is also more and more littler, but the pleasant sensation may make sexual intercourse the time weakens, and also can cause in the sexual intercourse process that condom breaks or sexual intercourse is retained in cover in the vagina when finishing owing to improper use, make seminal fluid flow into vagina, thereby cause contraceptive failure.Therefore the research that has the external vagina contraceptive of prevention and treatment of venereal diseases effect concurrently just is being subjected to World Health Organization (WHO) and the attention of China State Family Planning Commission.China's traditional medicine has medication history and experience in several thousand, has the complete Chinese medical theory of a cover, also has abundant Chinese herbal medicine resource simultaneously.Therefore filtering out the external vagina contraceptive that has the prevention and treatment of venereal diseases effect concurrently from Chinese herbal medicine has bright prospects.
Radix Sophorae Flavescentis, the property hardship, cold.GUIXIN, liver, stomach, large intestine, urinary bladder channel.Function cures mainly: heat clearing and damp drying, parasite killing, diuresis.Be used for hematodiarrhoea, have blood in stool, jaundice urine retention, leucorrhea with red and white discharge, swelling of the vulva pudendal pruritus, eczema, eczema, skin pruritus, scabies leprosy; External treatment trichomonal vaginitis.The Radix Pulsatillae, the property hardship, cold.Return stomach, large intestine channel.Function cures mainly: heat-clearing and toxic substances removing, and eliminating pathogenic heat from blood to cure dysentery, the effect of removing dampness and killing parasites [referring to: Lu Fuer, Liu Peilin chief editor. basic Chinese medicine. Beijing: Science Press, 2003 (1): 265-273].Both share the same views and preferences, and return through unanimity, and mutual reinforcement between can improve curative effect.Modern Chinese medicine discovers that the Radix Sophorae Flavescentis and the Radix Pulsatillae all have antitumor, regulate immunity, antiinflammatory, antibiotic, anti-trichomonal, effect such as essence extremely, both pharmacological actions also be consistent [referring to Mo Shaohong. the chemical constituent of the Radix Pulsatillae and Advance on Pharmacological Activities. Chinese crude drug, 2001,24 (5): 385-387; Xu Jinning, Zhang Heming, Zhao Xiuhua. the pharmacology activity research progress of Radix Sophorae Flavescentis. Hebei North institute journal (medicine), 2007,24 (5): 67-69].
The present invention tests external essence extremely and all more single medicine of fungistatic effect of finding 7 kinds of compound extracts obvious enhancing.Wherein to be that 1: 3 extracting solution kills smart effect best for Radix Sophorae Flavescentis, Radix Pulsatillae proportioning; Proportioning is that 1: 1 extracting solution bactericidal effect is best, and proportioning is 1: 4～1: 2 taking second place.Illustrate that two kinds of drug combinations can produce synergism and promote curative effect.Kill the extracting solution before the first six specimen semen sample use high temperature sterilize in the smart test, back four semen samples use the extracting solution behind the high temperature sterilize.Behind the high temperature action, kill smart effect and obviously strengthened finally, illustrate that variation has taken place medicine liquid ingredient under high temperature action.The mixed liquor that with the proportioning of same concentrations is in test 1: 1 extracting solution and two kinds of single medicament extracting solutions is cooked external bacteriostatic experiment, and the former has the obvious sterilization effect (inhibition zone＞10mm) latter does not then have bacteriostasis as a result.The synergism that two kinds of medicines are described is not only that different pharmaceutical components act on different loci, but has generated new pharmaceutical component, and this composition is antibacterial, and to kill smart effect all fairly obvious.Reason may be to contain N → O coordinate bond in the structure of oxymatrine, N-oxysophocarpine, close when frying in shallow oil with the triterpenoid saponin of groups such as hydroxyl, two keys, oxymatrine, N-oxysophocarpine are reduced to matrine, sophocarpine [referring to Wu Di, Liu Fajin, Liao Caizhen. drug matching is to the influence of chemical constituent. Chinese patent medicine, 2003,25 (2): 152-154] or other chemical reactions.Do not see the research of relevant Radix Sophorae Flavescentis and Radix Pulsatillae mixed extraction in the existing document, relevant single medicine extracting method of experimental basis of the present invention and cost are taken all factors into consideration the alcohol heating reflux method of having taked.Radix Sophorae Flavescentis and Radix Pulsatillae proportioning are that to kill smart effect at 1: 3 o'clock best, and proportioning is that 1: 1～1: 4 o'clock fungistatic effect is better.So Radix Sophorae Flavescentis, the external essence extremely of Radix Pulsatillae compound extract and antibacterial best proportioning are 1: 3.The pharmacological action of considering medicine medicine of the present invention is had both sterilization for killing essence, and Radix Sophorae Flavescentis, Radix Pulsatillae proportioning be externally to kill smart, fungistatic effect all clearly at 1: 3 o'clock, so prescription is decided to be Radix Sophorae Flavescentis and Radix Pulsatillae proportioning is 1: 3 (mass ratio).
Description of drawings
Fig. 1 extracting solution 1﹠amp; 3: do not have bacteriostasis substantially;
Fig. 2 extracting solution 2: fungistatic effect is best, and the inhibition zone size reduces with concentration and reduces;
Fig. 3 extracting solution 4: concentration does not have obvious inhibition zone during for 0.1g/ml;
Fig. 4 extracting solution 5: inhibition zone is slightly little than extracting solution 4;
Fig. 5 extracting solution 6: concentration does not have obvious inhibition zone during for 0.1g/ml);
Fig. 6 extracting solution 7: inhibition zone was very little when concentration was 0.2g/ml;
Fig. 7 extracting solution 8: concentration is 0.1, do not have obvious inhibition zone during 0.2g/ml;
When being 0.1～0.3g/ml, Fig. 8 extracting solution 9 concentration do not have obvious inhibition zone;
Fig. 9 left side: extracting solution 2; Right: extracting solution 1+3: with concentration proportioning is that 1: 1 the extracting solution (left side) and mixed liquor (right side) fungistatic effect of two kinds of single medicament extracting solutions compare, and there is the obvious sterilization effect in the left side, and the right side does not then have.
The specific embodiment
Embodiment 1
Preparation Radix Sophorae Flavescentis, Radix Pulsatillae compound extract
Material: Radix Pulsatillae root powder, Radix Sophorae Flavescentis, NaOH solution, distilled water
Preparation method: Radix Pulsatillae root powder and Radix Sophorae Flavescentis are placed the drying baker inner drying.Get 10g Radix Sophorae Flavescentis and Radix Pulsatillae mixture, Radix Sophorae Flavescentis and Radix Pulsatillae proportioning are 1: 3 (mass ratio), get 8 times of amounts of 60% ethanol heating and refluxing extraction 3 times, each 2h; Collect to merge 3 extracting solution, filter, heating drips dilute NaOH solution when being concentrated into not enough 10ml and sets up pH to 7.2～7.5, standardize solution 10ml, this moment, liquor strength was 1: 1, was equivalent to every ml medicinal liquid and contained crude drug in whole 1g, at 121 ℃ of following high temperature sterilize 20min, packing is standby then.
Claims (1)
1. one kind has the external medicine that kills smart and bacteriostasis, it is characterized in that, it is the medicine for preparing by the following method: after getting the Radix Sophorae Flavescentis and Radix Pulsatillae placement drying baker inner drying, is that the proportioning of 1 ﹕, 3～1 ﹕ 4 is mixed with Radix Sophorae Flavescentis and the Radix Pulsatillae by mass ratio, get Radix Sophorae Flavescentis and Radix Pulsatillae mixture, measure heating and refluxing extraction 3 times, each 2 hours for 8 times with 60% ethanol; Collect 3 extracting solution of merging, filter, heating concentrates, and drips dilute NaOH solution setting pH to 7.2～7.5, and being settled to liquor strength is 1 ﹕ 1, is equivalent to every ml medicinal liquid and contains crude drug in whole 1g, and at 121 ℃ of following high temperature sterilize 20min, packing is standby then.
2. the medicine with external essence extremely and bacteriostasis according to claim 1 is characterized in that the quality proportioning of the Radix Sophorae Flavescentis and the Radix Pulsatillae is 1 ﹕ 3.
3. the described preparation method with external essence extremely and bacteriostasis medicine of claim 1 is: after getting the Radix Sophorae Flavescentis and Radix Pulsatillae placement drying baker inner drying, is that the proportioning of 1 ﹕, 3～1 ﹕ 4 is mixed with Radix Sophorae Flavescentis and the Radix Pulsatillae by mass ratio, get Radix Sophorae Flavescentis and Radix Pulsatillae mixture, measure heating and refluxing extraction 3 times, each 2 hours for 8 times with 60% ethanol; Collect 3 extracting solution of merging, filter, heating concentrates, and drips dilute NaOH solution setting pH to 7.2～7.5, and being settled to liquor strength is 1 ﹕ 1, is equivalent to every ml medicinal liquid and contains crude drug in whole 1g, and at 121 ℃ of following high temperature sterilize 20min, packing is standby then.
4. preparation method according to claim 3 is characterized in that, the quality proportioning of the Radix Sophorae Flavescentis and the Radix Pulsatillae is 1 ﹕ 3.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US9884049B2 (en) | 2014-07-14 | 2018-02-06 | Orion Biotechnology Canada Ltd. | Microbicidal composition comprising an octoxynol and a quinolizidine alkaloid compound or a source thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN108186552A (en) * | 2018-01-09 | 2018-06-22 | 大连大学 | A kind of Traditional Chinese medicine compound gel suppository for being used for contraception and preparation method thereof |
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| CN1113797A (en) * | 1994-06-14 | 1995-12-27 | 蔡健 | Fuyanjing lubricant and its preparing method |
| CN1056076C (en) * | 1996-05-22 | 2000-09-06 | 延边天池兽药有限公司 | Prepn of injection with Radix Sophorae flavescentis and Radix Pulsatillae |
| CN1053380C (en) * | 1996-11-19 | 2000-06-14 | 沈阳市奇生生物研究所 | Injection for curing AIDS and preparation method thereof |
| CN1206606A (en) * | 1997-07-25 | 1999-02-03 | 初新东 | External traditional Chinese medicine and preparation process thereof |
| CN1273035A (en) * | 1999-05-07 | 2000-11-15 | 初新东 | Chinese-medicinal liquid extract for sterilization |
| CN101417018B (en) * | 2007-10-22 | 2011-08-10 | 洛阳普莱柯生物工程有限公司 | Preparation method of animal injection capable of clearing away the heat-evil and expelling superficial evils, cooling blood and relieving dysentery |
| CN101181464A (en) * | 2007-11-08 | 2008-05-21 | 刘玉美 | Chinese medicine composition for curing women's leukorrhagia |
| CN101564362A (en) * | 2008-04-21 | 2009-10-28 | 曲奕 | Chinese medical herb antibacterial lotion |
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| US9884049B2 (en) | 2014-07-14 | 2018-02-06 | Orion Biotechnology Canada Ltd. | Microbicidal composition comprising an octoxynol and a quinolizidine alkaloid compound or a source thereof |
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