CN101337000B - Solanum nigrum extract, preparation method and use thereof - Google Patents
Solanum nigrum extract, preparation method and use thereof Download PDFInfo
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Abstract
The invention relates to a solanum nigrum extractive, the preparation method and the medicament application thereof, wherein, the total content of solancarpine, solamargine and solasurine is 80wt% to 99wt%; the content of solancarpine is 30wt% to 50wt%; the content of solamargine is 10wt% to 30wt%; and the content of solasurine is 30wt% to 50wt%. The invention further relates to the preparation method of the solanum nigrum extractive and the application with the solanum nigrum extractive being the active components in the medicaments for preventing and curing diseases, such as tumor, diabetes, asthma, coronary heart disease, etc.
Description
Technical field
The present invention relates to the plant black nightshade extract, its preparation method and as the application of the medicine of diseases such as active component preparation prevention and treatment tumour, diabetes, asthma, coronary heart disease.
Background technology
Black nightshade Solalum Nigrum L. is annual Solanum herbaceous plant, and all there is growth most of at home provinces and regions.The herb hyoscine, cold in nature, bitter is hot little sweet, slightly poisonous, returns lung, urinary bladder channel.The ability eliminating stasis to subdue swelling is clearing heat and detoxicating.Gold-coloured staphylococci, typhoid bacillus, shigella dysenteriae, proteus, Escherichia coli, pseudomonas aeruginosa, hog cholera bacillus all there is stronger bacteriostasis; The cancer cell of inhibition and effects such as urinary system infection contamination and leukorrhea are arranged; Treatment carbuncle commonly used is swollen; Tumour, dysentery etc., it is among the people that to be used to treat diabetes curative effect remarkable.Black nightshade also has certain dietary function in addition, therefore has good medical value.Be rich in steroid alkaloid in the plant; ((δ-solanigrine), black nightshade are decided alkali (solanigridine), solasurine solasurine etc. for ε-solanigrine), δ-solanine like: solasonine (solasonine), solamargine (solamargine), the bright alkali of Australia eggplant (solasodamine), ε-solanine; These steroid alkaloids all are present in Solanum nigrum stem, leaf, root, really with the form of glycosides, and aglycon is solasodine (solasodine).
Solasonine solasonine solamargine solamargine
Solasurine solasurine
Solasonine and solamargine are two main in black nightshade alkaloid components; In in vitro test; Two compounds all show the destructiveness of cell membrane fat; Both show cooperative effect, and the destructiveness of solamargine is stronger, and rhamnose wherein possibly play important function in the combining of guiding glycoalkaloid and cell; Some researchs to solamargine show, thus solamargine can regulate the expression of Tumor Necrosis Factor Receptors and cause cancer cell-apoptosis, 2 rhamnose possibly play a decisive role in triggering Apoptosis.Other glycoalkaloid also shows approximate physiologically active.Therefore, find out suitable technology, from black nightshade, extract glucosides TA position, carry out drug efficacy study, can be used as a direction of anti-cancer agent research.To the research of prophylactic treatment diabetes, asthma and the coronary heart disease isoreactivity of black nightshade crude extract also is external focus at present, but at present also not research show it is that which chemical composition is in action in the crude extract definitely.
Water-soluble extract, its preparation method and the pharmaceutical compositions thereof of the disclosed nightshade of Chinese patent Granted publication CN1329038C, wherein water-soluble extract is to be that solasonine and solamargine by 60%-90% constitutes basically.
The black nightshade method for distilling cost that uses at present is higher, and active constituent content is lower, and impurity is more, and the pollution that produces in the preparation process is bigger.The product content of effective that after extracting, obtains has limited its use in pharmaceutical compositions, still can not satisfy the needs of efficient pharmaceutical compositions.
Summary of the invention
The object of the invention first aspect has provided a kind of black nightshade extract.
The object of the invention second aspect provides the preparation method of above-mentioned black nightshade extract.
The object of the invention third aspect provides the application of above-mentioned black nightshade extract in medicine.
As the black nightshade extract of first aspect present invention, comprise solasonine and solamargine, also comprise solasurine, wherein solasonine, solamargine and solasurine three's total content is 80wt%～99wt%.
Solasonine content is 30wt%～50wt%, and solamargine content is 10wt%～30wt%, and solasurine content is 30wt%～50wt%.
Preparation method as the black nightshade extract of second aspect present invention may further comprise the steps:
(1) black nightshade herb and fruit add alcohol extract, collect extract A;
(2) ethanol in the recovery extract gets concentrate B to there not being the alcohol flavor;
(3) concentrate B uses cationic exchange resin adsorption, and water, ethanol and contain alkali ethanol and carry out wash-out are successively collected and contained alkali ethanol elution portion C;
(4) contain alkali ethanol elution portion C, reclaim ethanol and get concentrate D;
(5) supernatant is abandoned in concentrate D centrifugation, gets sediment E;
(6) sediment E is washed with water to near-white and supernatant pH value less than 9, promptly gets black nightshade extract after the deposition drying.
In above-mentioned steps (1), at every turn make a living 2～8 times of volumes of dose of consumption of ethanol; The weight percent concentration of ethanol is 50%～95%.Be preferably at every turn make a living 5 times of volumes of dose of consumption of ethanol; The weight percent concentration of ethanol is 90%.
In above-mentioned steps (1), adopt and reflux or the ultrasonic power extraction, extraction time is 2～5 times, each 1～3 hour.Be preferably and adopt reflux type to extract, extraction time is 3 times, each 1.5 hours.
When raw medicinal herbs is the black nightshade herb, in above-mentioned steps (2), increase by one to the step of concentrate B with the petroleum ether extraction degreasing.Be preferably concentrate B with 3 degreasings of equal-volume petroleum ether extraction.
In above-mentioned steps (3), make a living 0.5～5 times of volume of dose of the consumption of cationic ion-exchange resin.Be preferably 1 times of volume of crude drug amount.
The functional group of said cationic ion-exchange resin is a kind of among sulfonic group, pyrovinic acid base, phosphate, carboxyl and the phenolic hydroxyl group.Be preferably sulfonic group.
In above-mentioned steps (3), preferred elution process is closely colourless to flowing out liquid with washed resin for successively, and the weight percent concentration of 1～10 times of resin volume is 30%～90% ethanol elution, 0.5～10 times of resin volume contain the alkali ethanol elution.Preferred elution process is closely colourless to flowing out liquid with washed resin for successively; The weight percent concentration of 1～2 times of resin volume is 30%～60% ethanol elution; The weight percent concentration of 1～5 times of resin volume is 90% ethanol elution, 1～5 times of resin volume contain the alkali ethanol elution.Most preferred type of elution is closely colourless to flowing out liquid with washed resin for successively; The weight percent concentration of 1 times of resin volume is 40% ethanol elution; The weight percent concentration of 2 times of resin volumes is 90% ethanol elution, 3 times of resin volumes contain the alkali ethanol elution.
In above-mentioned steps (3), the said alkali ethanol that contains is that weight percent concentration is the mixed solution of 50～95% ethanol and alkali, and wherein alkali is a kind of among ammoniacal liquor, NaOH, sodium carbonate, sodium acid carbonate and other commercial production alkali commonly used.Preferably containing alkali ethanol is that weight percent concentration is that 90% ethanol is the solution that 25% ammoniacal liquor mixed in 10: 1 by volume with weight percent concentration.
In above-mentioned steps (6),, increase by a usefulness washing with acetone and be precipitated to subalbous step if sediment E color is darker.
In above-mentioned steps (3), described cationic ion-exchange resin can be regenerated, and the method for its regeneration is: add the heat soaking resin with 2% sodium hydrate aqueous solution; The dress post, the sodium hydrate aqueous solution 800ml wash-out with 2%; Water is washed till outflow liquid and is neutral; Using 1N salt pickling resin is 2 to flowing out liquid pH value; Using deionized water to wash resin is 6 to flowing out liquid pH value, can drop into repeated use.
Solasonine, solamargine and solasurine three's total content is 80wt%～99wt% in the black nightshade extract that method produced according to the present invention obtains; Solasonine content is 30wt%～50wt%; Solamargine content is 10wt%～30wt%, and solasurine content is 30wt%～50wt%.
As the medicinal usage of the above-mentioned black nightshade extract of third aspect present invention be with the solasonine in the described black nightshade extract, solamargine and solasurine as active component, add acceptable accessories, the medicine of any formulation of processing.Preferred formulation is an injection.
The application of black nightshade extract in the medicine that is preparing diseases such as prevention and treatment tumour, diabetes, asthma, heart disease as active component as third aspect present invention.
Above-mentioned black nightshade extract is following as the concrete medicinal usage of active component, but is not limited to following medicinal usage:
1, can in preparation prevention and treatment tumour medicine, use as active component with black nightshade extract.
2, can in preparation prevention and treatment diabetes medicament, use as active component with black nightshade extract.
3, can in preparation prevention and treatment asthma medicine, use as active component with black nightshade extract.
4, can in preparation prevention and treatment medicaments for coronary disease, use as active component with black nightshade extract.
Black nightshade extract of the present invention contains active component solasonine, solamargine and solasurine, and it has certain curative effect aspect diseases such as tumour, diabetes, asthma, coronary heart disease in prevention with treating.Preparation method of the present invention compared with prior art, this method technology is advanced, produces practically greatly, cost is relatively low.Organic solvent is mainly ethanol, and the soda acid after the use is discharging after treatment all, and environmental pollution is little.Extract obtained impurity is few, and active constituent content is high.
The specific embodiment
Below in conjunction with specific embodiment the present invention is done further explanation, but do not limit the present invention in any way.
Embodiment 1
The preparation of black nightshade extract:
(1) black nightshade herb 400g adds 90% ethanol 2000ml, refluxing extraction 3 times, each 1.5 hours;
(2) extract recovery ethanol gets the about 300ml of concentrate to there not being the alcohol flavor, petroleum ether extraction 3 times, and each consumption is 300ml;
(3) concentrate is processed into HD-8 strongly acidic cation-exchange (Shanghai Huazhen Science and Technology Co., Ltd.) the dress post absorption of H+ type with 400ml; It is closely colourless that water is eluted to outflow liquid successively; 40% ethanol 400ml wash-out; 90% ethanol 800ml wash-out contains ammoniacal liquor ethanol (90% ethanol: 25% ammoniacal liquor is 10: 1) 1200ml wash-out;
(4) collection contains ammoniacal liquor ethanol elution part, reclaims ethanol;
(5) concentrate 5000rpm is centrifugal 10 minutes, abandons supernatant;
(6) low amounts of water washing precipitation, centrifugal 10 minutes of 5000rpm so uses water washing 2 times, again with the small amount of acetone washing once, promptly gets black nightshade extract 0.17g, yield 0.04% behind the deposition drying under reduced pressure.
Wherein resin regeneration process is following: add the heat soaking resin with 2% sodium hydrate aqueous solution 800ml; The dress post, the sodium hydrate aqueous solution 800ml wash-out with 2%; Water is washed till outflow liquid and is neutral; Using 1N salt pickling resin is 2 to flowing out liquid pH value; Using deionized water to wash resin is 6 to flowing out liquid pH value, can drop into repeated use.
Embodiment 2
The preparation of black nightshade extract:
(1) Solanum nigrum fruit 1000g adds 90% ethanol 5000ml, refluxing extraction 3 times, each 1.5 hours;
(2) extract recovery ethanol gets the about 800ml of concentrate to there not being the alcohol flavor;
(3) concentrate is processed into HD-8 strongly acidic cation-exchange (Shanghai Huazhen Science and Technology Co., Ltd.) the dress post absorption of H+ type with 1000ml; It is closely colourless that water is eluted to outflow liquid successively; 40% ethanol 1000ml wash-out; 90% ethanol 2000ml wash-out contains ammoniacal liquor ethanol (90% ethanol: 25% ammoniacal liquor is 10: 1) 3000ml wash-out;
(4) collection contains ammoniacal liquor ethanol elution part, reclaims ethanol;
(5) concentrate 5000rpm is centrifugal 10 minutes, abandons supernatant;
(6) low amounts of water washing precipitation, centrifugal 10 minutes of 5000rpm so uses water washing 2 times, again with the small amount of acetone washing once, promptly gets black nightshade extract 1.5g, yield 0.15% behind the deposition drying under reduced pressure.
Wherein resin regeneration process is following: add the heat soaking resin with 2% sodium hydrate aqueous solution 2000ml; The dress post, the sodium hydrate aqueous solution 2000ml wash-out with 2%; Water is washed till outflow liquid and is neutral; Using 1N salt pickling resin is 2 to flowing out liquid pH value; Using deionized water to wash resin is 6 to flowing out liquid pH value, can drop into repeated use.
Embodiment 3
The preparation of black nightshade extract:
(1) black nightshade herb 400g adds 90% ethanol 2000ml, ultrasonic Extraction 3 times, each 1.5 hours;
(2) extract recovery ethanol gets the about 300ml of concentrate to there not being the alcohol flavor, petroleum ether extraction 3 times, and each consumption is 300ml;
(3) concentrate is processed into HD-8 strongly acidic cation-exchange (Shanghai Huazhen Science and Technology Co., Ltd.) the dress post absorption of H+ type with 400ml; It is closely colourless that water is eluted to outflow liquid successively; 40% ethanol 400ml wash-out; 90% ethanol 800ml wash-out contains 80% ethanol 1200ml wash-out of 0.1% NaOH;
(4) collection contains NaOH ethanol elution part, reclaims ethanol;
(5) concentrate 5000rpm is centrifugal 10 minutes, abandons supernatant;
(6) low amounts of water washing precipitation, centrifugal 10 minutes of 5000rpm.So with after the water washing 4 times, supernatant pH value is about 8.Promptly get black nightshade extract 0.13g, yield 0.03% after the deposition drying.
Wherein resin regeneration process is following: add the heat soaking resin with 2% sodium hydrate aqueous solution 800ml; The dress post, the sodium hydrate aqueous solution 800ml wash-out with 2%; Water is washed till outflow liquid and is neutral; Using 1N salt pickling resin is 2 to flowing out liquid pH value; Using deionized water to wash resin is 6 to flowing out liquid pH value, can drop into repeated use.
Embodiment 4
(LC-MS method) measured in alkaloidal discriminating in the black nightshade extract
Laboratory apparatus:
The LC-MS appearance, Agilent 1200 LC/MSD/VL
EISD, SoftA 300A
Enlightening horse C18 jewel post, 250 * 4.6mm
The HPLC condition:
Methyl alcohol: 0.2% acetate, gradient 0min, 5: 95; 10min, 5: 95; 15min, 90: 10; 25min, 100: 0.Flow velocity 0.8ml/min, 35 ℃ of column temperatures.
The EISD testing conditions:
DT＝70℃，SC＝35℃，GAS＝50psi
Experimental technique and result:
Get black nightshade extract 10mg, the accurate title, decide, put in the 25ml measuring bottle, and the small amount of methanol dissolving, methyl alcohol is diluted to scale, 0.45 μ m filtering with microporous membrane, 10 μ l sample introductions.The one group peak of retention time between 16～20 minutes is glycoalkaloid, and the peak area sum accounts for 85% of total peak area.HPLC-MS detects molecular ion peak 17.5min, 884 (solasonine, M+1); 17.8,868 (solamargine, M+1); 18.2,722 (solasurine, M+1).
Embodiment 5
The mensuration of alkaloid (thin layer chromatography scanning) in the black nightshade extract
Laboratory apparatus:
Tianjin, island CS-9310 type dual-wavelength lamellar scanning appearance
The TLC condition:
High-efficient silica gel G plate 10cm * 20cm; Solvent is a n-butanol: ammoniacal liquor (4: 1) upper strata liquid; The bismuth potassium iodide colour developing; λ S=468nm scanning.
Experimental technique and result:
Get black nightshade extract 10.05mg, the accurate title, decide, and puts in the 5ml measuring bottle small amount of methanol dissolving; Methyl alcohol is diluted to scale, shakes up, and gets 5 μ l point samples, launches; Colour developing, scanning, the area percentage of solasonine, solamargine and three points of solasurine is 40.5%, 25.4% and 34.1%.
Embodiment 6
The preparation of black nightshade extract injection
Get black nightshade extract 2000mg, add injection water 900ml, 1,2-propane diols 50ml, sodium chloride 9g with the hydrochloric acid adjust pH to 4.0 of 0.1N, adds the injection water to 1000ml, with 0.2 μ m filtering with microporous membrane, embedding, moist heat sterilization gets the black nightshade extract injection.
Embodiment 7
The black nightshade extract gastric infusion is to the experimental study of rat liver cancer H22 influence
Choose the female C57BL/6 mouse of 18～20g, get well-grown rat liver cancer H22 ascites, be diluted to 1～2 * 10 with physiological saline
7The cell suspension of individual/ml concentration, every the right armpit subcutaneous vaccination of mouse 0.2ml, random packet, several 10 of every treated animal, 15 of control groups, establish:
Blank group (coordinative solvent, 25ml/kg, ig * 7)
Syklofosfamid ampoule (30mg/kg, ip * 7)
Black nightshade extract (50mg/kg, ig * 7)
Black nightshade extract (100mg/kg, ig * 7)
Black nightshade extract (200mg/kg, ig * 7)
Black nightshade extract is made into suspension with 0.5%CMC after with several Tween-80 hydrotropies, and syklofosfamid ampoule is mixed with desired concn with physiological saline.
Administration is played next day in the inoculation back, and the administration volume is the 0.5ml/20g body weight, and per os was irritated stomach 7 days continuously.Inoculate back 10 days and take off neck execution animal, dissect and get the knurl piece, claim that knurl is heavy, calculate tumour inhibiting rate.
Result of the test is seen table 1, and black nightshade extract can obviously reduce rat liver cancer H22 tissue level 50,100, on the level of 200mg/kg during the per os gastric infusion.After the administration 7 days, each administration group is compared with control group, and knurl is heavy, and there were significant differences (P＜0.01).
Table 1. black nightshade extract gastric infusion is to the tumor-inhibiting action of rat liver cancer H22
Compare with control group:
*P＜0.05,
*P＜0.01.
Embodiment 8
The black nightshade extract gastric infusion is to the experimental study of Mice Bearing Lewis Lung Cancer influence
Choose the female C57BL/6 mouse of 18～20g, get well-grown Mice Bearing Lewis Lung Cancer knurl piece, be prepared into 1～2 * 10 with physiological saline homogenate method
7The cell suspension of individual/ml concentration, every the right armpit subcutaneous vaccination of mouse 0.2ml, random packet, several 6 of each treated animal, 12 of control groups, establish:
The blank group (coordinative solvent, 25ml/kg)
Syklofosfamid ampoule (30mg/kg, ip * 7)
Black nightshade extract (50mg/kg, ig * 10)
Black nightshade extract (100mg/kg, ig * l0)
Black nightshade extract (200mg/kg, ig * 10)
Black nightshade extract is made into suspension with 0.5%CMC after with several Tween-80 hydrotropies, and syklofosfamid ampoule is mixed with desired concn with physiological saline.
Inoculation plays administration next day, and the administration volume is the 0.5ml/20g body weight, and per os was irritated stomach 10 days continuously.Inoculate back 14 days and take off neck execution animal, dissect and get the knurl piece, claim that knurl is heavy, calculate tumour inhibiting rate.
Result of the test is seen table 2, and black nightshade extract can obviously reduce the Mice Bearing Lewis Lung Cancer tissue level 50,100, on the level of 200mg/kg during the per os gastric infusion.After the administration 10 days, each administration group is compared with control group, and knurl is heavy, and there were significant differences (P＜0.01).
Table 2. black nightshade extract gastric infusion is to the tumor-inhibiting action of Mice Bearing Lewis Lung Cancer
Compare with control group:
*P＜0.05,
*P＜0.01.
Embodiment 9
The experimental study that black nightshade extract is subcutaneous, intraperitoneal administration influences rat liver cancer H22
Choose the female C57BL/6 mouse of 18～20g, get well-grown rat liver cancer H22 ascites, be prepared into 1～2 * 10 with physiological saline homogenate method
7The cell suspension of individual/ml concentration, every the right armpit subcutaneous vaccination of mouse 0.2ml, random packet, several 10 of each treated animal, 15 of control groups, establish:
The blank group (coordinative solvent, 25ml/kg)
Syklofosfamid ampoule (30mg/kg, ip * 7)
Black nightshade extract (40mg/kg, sc * 10)
Black nightshade extract (40mg/kg, ip * 10)
Black nightshade extract is made into suspension with 0.5%CMC after with several Tween-80 hydrotropies, and syklofosfamid ampoule is mixed with desired concn with physiological saline.
Inoculation plays administration next day, and the administration volume is the 0.5ml/20g body weight, subcutaneous continuously, intraperitoneal administration 7 days.Inoculate back 10 days and take off neck execution animal, dissect and get the knurl piece, claim that knurl is heavy, calculate tumour inhibiting rate.
Result of the test is seen table 3, and black nightshade extract can obviously reduce rat liver cancer H22 tissue level when subcutaneous and intraperitoneal administration on the level of 40mg/kg.After the administration 7 days, black nightshade extract 40mg/kg subcutaneous administration group, intraperitoneal injection group are compared with control group, and knurl is heavy, and there were significant differences (P＜0.01).
Table 3. black nightshade extract is subcutaneous, intraperitoneal administration is to the tumor-inhibiting action of rat liver cancer H22
Compare with control group:
*P＜0.05,
*P＜0.01.
Embodiment 10
The experimental study that black nightshade extract is subcutaneous, intraperitoneal administration influences Mice Bearing Lewis Lung Cancer
Choose the male C57BL/6 mouse of 18～20g, get well-grown mice lung cancer knurl piece, be prepared into 1～2 * 10 with physiological saline homogenate method
7The cell suspension of individual/ml concentration, every the right armpit subcutaneous vaccination of mouse 0.2ml, random packet, several 10 of each treated animal, 15 of control groups, establish:
The blank group (coordinative solvent, 25ml/kg)
Syklofosfamid ampoule (30mg/kg, ip * 7)
Black nightshade extract (40mg/kg, sc * 10)
Black nightshade extract (40mg/kg, ip * 10)
Black nightshade extract is made into suspension with 0.5%CMC after with several Tween-80 hydrotropies, and syklofosfamid ampoule is mixed with desired concn with physiological saline.
Inoculation plays administration next day, and the administration volume is the 0.5ml/20g body weight, subcutaneous continuously, intraperitoneal administration 10 days.Inoculate back 14 days and take off neck execution animal, dissect and get the knurl piece, claim that knurl is heavy, calculate tumour inhibiting rate.
Result of the test is seen table 4, and black nightshade extract can obviously reduce the Mice Bearing Lewis Lung Cancer tissue level when subcutaneous and intraperitoneal administration on the level of 40mg/kg.After the administration 10 days, black nightshade extract 40mg/kg intraperitoneal injection group is compared with control group, and knurl is heavy, and there were significant differences (P＜0.01).
Table 4. black nightshade extract is subcutaneous, intraperitoneal administration is to the tumor-inhibiting action of Mice Bearing Lewis Lung Cancer
Compare with control group:
*P＜0.05,
*P＜0.01.
Embodiment 11
Black nightshade extract is to the influence of normal mouse postprandial blood sugar
Choose 18～20g male mice in kunming; Be divided into blank group (physiological saline), positive control group (glibenclamide 2.5mg/kg), administration group I (black nightshade extract 10mg/kg) and administration group II (black nightshade extract 20mg/kg) at random by body weight; Every group each 6, water is can't help in the 18h fasting before the test, behind the gastric infusion 1,2,4,8, the 10h tail vein blood; Behind the centrifugal separation plasma, measure glucose level with glucose oxidase-peroxidase (GOD-POD) method.
Result of the test is seen table 5, and black nightshade extract can obviously reduce the normal mouse level of postprandial blood sugar 10, on the level of 20mg/kg.Behind administration 8h, the 10h, the 20mg/kg dose groups is compared with control group, and there were significant differences for blood glucose value (P＜0.01).
Table 5. black nightshade extract is to the influence of normal mouse postprandial blood sugar
| Group | Dosage (mg/kg) | Blood sugar level (mg/dl) | |||||
| 0h | 1h | 2h | 4h | 8h | 10h | ||
| Blank | 2ml/kg | 98.11±5.25 | 98.77±5.76 | 97.07±6.22 | 94.20±5.02 | 90.47±4.82 | 91.44±5.02 |
| Glibenclamide | 2.5 | 97.54±6.21 | 97.29±6.34 | 88.94±5.73 | 83.99±3.94 | 70.04±3.21 ** | 56.84±3.00 ** |
| The black nightshade TA | 10 | 98.79±6.95 | 99.20±5.21 | 93.05±7.26 | 88.47±4.21 | 83.56±2.57 | 77.28±2.56 * |
| 20 | 99.64±6.28 | 98.49±6.32 | 89.53±4.77 | 84.55±4.50 | 80.03±2.12 ** | 69.14±3.11 ** | |
Compare with control group:
*P＜0.05,
*P＜0.01 (n=6).
Embodiment 12
Black nightshade extract is to the influence of tissue of experimental diabetic mice postprandial blood sugar
Choose 18～20g male mice in kunming; Water is can't help in the 24h fasting, presses 120mg/kg lumbar injection alloxan physiological salt liquid, feeding behind the 1h; Tail vein blood is measured blood sugar level behind the 48h, and the mouse that reaches more than the 200mg/100ml when blood glucose value is decided to be the diabetes model mouse.It is divided into blank group (physiological saline), positive control group (glibenclamide 2.5mg/kg), administration group I (black nightshade extract 10mg/kg) and administration group II (black nightshade extract 20mg/kg) at random by body weight; Every group each 6; Behind the gastric infusion 1,2,4,8, the 10h tail vein blood; Behind the centrifugal separation plasma, measure glucose level with glucose oxidase-peroxidase (GOD-POD) method.
Result of the test is seen table 6, and black nightshade extract can obviously reduce the tissue of experimental diabetic mice level of postprandial blood sugar 10, on the level of 20mg/kg.Behind administration 4h, 8h and the 10h, each dose groups is compared with control group, and there were significant differences for blood glucose value (P＜0.01).
Table 6. black nightshade extract is to the influence of tissue of experimental diabetic mice postprandial blood sugar
| Group | Dosage (mg/kg) | Blood sugar level (mg/dl) | |||||
| 0h | 1h | 2h | 4h | 8h | 10h | ||
| Blank | 2ml/kg | 274.11±10.44 | 278.19±10.20 | 288.41±9.73 | 299.10±10.02 | 505.21±11.82 | 320.85±5.02 |
| Glibenclamide | 2.5 | 269.48±11.22 | 261.33±12.54 | 214.85± 8.62 ** | 164.33± 9.03 ** | 119.53± 8.21 ** | 91.44±9.08 ** |
| The black nightshade TA | 10 | 285.02±12.75 | 281.73±10.02 | 273.99±7.06 | 220.49± 8.04 ** | 152.11± 8.57 ** | 127.28± 6.59 ** |
| 20 | 284.33±10.46 | 269.44±9.21 | 250.18±9.21 | 181.40± 7.67 ** | 139.40± 7.10 ** | 108.14± 5.71 ** | |
Compare with control group:
*P＜0.05,
*P＜0.01 (n=6).
Embodiment 13
Black nightshade extract is to the influence of cavy experimental asthma
Choose 200～250g healthy guinea pig and be divided into normal control group, asthmatic model group, administration group I (black nightshade extract 10mg/kg) and administration group II (black nightshade extract 20mg/kg) at random, every group each 6.Set up the asthmatic guinea pigs model with ovalbumin (OVA) lumbar injection sensitization with atomizing to suck to excite.Detect serum NO level, T-NOS, iNOS level.
Result of the test is seen table 7, and black nightshade extract can obviously reduce serum NO level, T-NOS and iNOS level 10, on the level of 20mg/kg, and relatively there were significant differences (P＜0.05) with model group.
Table 7. black nightshade extract is to the influence of cavy experimental asthma
| Group | Dosage (mg/kg) | NO(μmol/L) | T-NOS(U/ml) | iNOS(U/ml) |
| Normal control | 68.3±35.6 | 28.11±2.06 | 5.78±0.32 | |
| Asthmatic model | 150.44±49.65 | 43.96±3.22 | 10.28±1.99 | |
| Black nightshade extract | 10 | 84.22±35.74 * | 33.61±3.09 * | 7.59±1.92 * |
| 20 | 69.18±23.11 * | 31.28±1.50 * | 5.58±1.42 * |
Compare with model group:
*P＜0.05 (n=6).
Embodiment 14
Black nightshade extract is to the protective effect of myocardial ischemia-reperfusion injury
Adopt isolated rat perfusion heart (Langendorff heart) ischemia-reperfusion injury model; With the K-HShi buffer solution perfused hearts that contains black nightshade extract; Detect in the cardiac muscular tissue MDA content and to irritate in the 15min perfusate LDH more active, this two indexes can reflect and the closely-related oxygen radical level variation of myocardial damage indirectly.Chamber incidence of quivering of each group of record.The result that causes of perfusion is like table 8,9, shown in 10 again.
Table 8. black nightshade extract is to the quiver influence of incidence of isolated rat perfusion heart chamber
| Group | Number quivers in the chamber | The chamber incidence (%) of quivering |
| Negative control | 11 | 92.5 |
| Black nightshade extract (0.2g/L) | 5 | 46.7 |
| Black nightshade extract (2.0g/L) | 2 | 18.2 ** |
Annotate: compare with negative control group,
*P＜0.05,
*P＜0.01 (n=12).
Table 9. black nightshade extract is organized the influence of MDA content to isolated myocardium
| Group | MDA (mmol/g cardiac muscular tissue) |
| Negative control | 0.33±0.05 |
| Normal control | 0.16±0.02 ** |
| Black nightshade extract (0.2g/L) | 0.24±0.03 * |
| Black nightshade extract (2.0g/L) | 0.18±0.02 ** |
Annotate: compare with negative control group,
*P＜0.05,
*P＜0.01 (n=12).
Table 10. black nightshade extract extract is to irritate the active influence of LDH in the perfusate again
| Group | LDH(units/100ml) |
| Negative control | 144.2±11.7 |
| Black nightshade extract (0.2g/L) | 105.5±7.9 * |
| Black nightshade extract (2.0g/L) | 64.3±4.7 ** |
Annotate: compare with negative control group,
*P＜0.05,
*P＜0.01 (n=12).
More than show and described basic principle of the present invention, principal character and advantage of the present invention.The technical staff of the industry should understand; The present invention is not restricted to the described embodiments; That describes in the foregoing description and the specification just explains principle of the present invention; The present invention also has various changes and modifications under the prerequisite that does not break away from spirit and scope of the invention, and these variations and improvement all fall in the scope of the invention that requires protection.The present invention requires protection domain to be defined by appending claims and equivalent thereof.
Claims (21)
1. the preparation method of a black nightshade extract is characterized in that, comprises the steps:
(1) black nightshade and fruit add alcohol extract, collect extract A;
(2) ethanol in the recovery extract gets concentrate B to there not being the alcohol flavor;
(3) concentrate B uses cationic exchange resin adsorption, and water, ethanol and contain alkali ethanol and carry out wash-out are successively collected and contained alkali ethanol elution portion C;
(4) contain alkali ethanol elution portion C, reclaim ethanol and get concentrate D;
(5) supernatant is abandoned in concentrate D centrifugation, gets sediment E;
(6) sediment E is washed with water to near-white and supernatant pH value less than 9, promptly gets black nightshade extract after the deposition drying.
2. the method for claim 1 is characterized in that, in above-mentioned steps (1), and at every turn make a living 2～8 times of volumes of dose of consumption of ethanol; The weight percent concentration of ethanol is 50%～90%.
3. the method for claim 1 is characterized in that, in above-mentioned steps (1), and at every turn make a living 5 times of volumes of dose of consumption of ethanol; The weight percent concentration of ethanol is 90%.
4. the method for claim 1 is characterized in that, in above-mentioned steps (1), adopts and refluxes or the ultrasonic power extraction, and extraction time is 2～5 times, each 1～3 hour.
5. the method for claim 1 is characterized in that, in above-mentioned steps (1), adopts and refluxes or the ultrasonic power extraction, and extraction time is 3 times, each 1.5 hours.
6. the method for claim 1 is characterized in that, when raw medicinal herbs is the black nightshade herb, in above-mentioned steps (2), increases the step of a pair of concentrate B with the petroleum ether extraction degreasing.
7. the method for claim 1 is characterized in that, when raw medicinal herbs is the black nightshade herb, in above-mentioned steps (2), increases the step of a pair of concentrate B with 3 degreasings of equal-volume petroleum ether extraction.
8. the method for claim 1 is characterized in that, in above-mentioned steps (3), and make a living 0.5～5 times of volume of dose of the consumption of said cationic ion-exchange resin.
9. the method for claim 1 is characterized in that, in above-mentioned steps (3), and make a living 1 times of volume of dose of the consumption of said cationic ion-exchange resin.
10. the method for claim 1 is characterized in that, in above-mentioned steps (3), the functional group of said cationic ion-exchange resin is a kind of among sulfonic group, pyrovinic acid base, phosphate, carboxyl and the phenolic hydroxyl group.
11. the method for claim 1 is characterized in that, in above-mentioned steps (3), the functional group of said cationic ion-exchange resin is a sulfonic group.
12. the method for claim 1; It is characterized in that; In above-mentioned steps (3); The elution process that adopts is closely colourless to flowing out liquid with washed resin for successively, and the weight percent concentration of 1～10 times of resin volume is 30%～90% ethanol elution, 0.5～10 times of resin volume contain the alkali ethanol elution.
13. the method for claim 1; It is characterized in that; In above-mentioned steps (3), the elution process of employing is closely colourless to flowing out liquid with washed resin for successively, and the weight percent concentration of 1～2 times of resin volume is 30%～60% ethanol elution; The weight percent concentration of 1～5 times of resin volume is 90% ethanol elution, 1～5 times of resin volume contain the alkali ethanol elution.
14. the method for claim 1; It is characterized in that; In above-mentioned steps (3), the elution process of employing is closely colourless to flowing out liquid with washed resin for successively, and the weight percent concentration of 1 times of resin volume is 40% ethanol elution; The weight percent concentration of 2 times of resin volumes is 90% ethanol elution, 3 times of resin volumes contain the alkali ethanol elution.
15., it is characterized in that the described alkali ethanol that contains is that weight percent concentration is the mixed solution of 50%～95% ethanol and alkali like claim 3 or 10 or 11 or 12 described methods, wherein alkali is a kind of in ammoniacal liquor, NaOH, sodium carbonate, the sodium acid carbonate.
16., it is characterized in that the described alkali ethanol that contains is that weight percent concentration is that 90% ethanol is the solution that 25% ammoniacal liquor mixed in 10: 1 by volume with weight percent concentration like claim 3 or 10 or 11 or 12 described methods.
17. the method for claim 1 is characterized in that, in above-mentioned steps (6) if in sediment E color darker, increase by a usefulness washing with acetone and be precipitated to subalbous step.
18. method as claimed in claim 3 is characterized in that, described cationic ion-exchange resin can be regenerated, and the method for its regeneration is: add the heat soaking resin with 2% sodium hydrate aqueous solution; The dress post is with 2% sodium hydrate aqueous solution 800ml wash-out; Water is washed till outflow liquid and is neutral; Using 1N salt pickling resin is 2 to flowing out liquid pH value; Using deionized water to wash resin is 6 to flowing out liquid pH value, can drop into repeated use.
19. the black nightshade extract that a preparation method as claimed in claim 1 prepares is as the application of active component in preparation prevention and treatment diabetes medicament.
20. the black nightshade extract that a preparation method as claimed in claim 1 prepares is as the application of active component in preparation prevention and treatment asthmatic medicament.
21. the black nightshade extract that a preparation method as claimed in claim 1 prepares is as the application of active component in the protection medicine of preparation myocardial ischemia-reperfusion injury.
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| TWI414304B (en) * | 2011-05-04 | 2013-11-11 | G & E Herbal Biotechnology Co Ltd | Treatment of wart with a water soluble extract from plant of solanum genus |
| TWI476012B (en) | 2011-05-12 | 2015-03-11 | G & E Herbal Biotechnology Co Ltd | Treatment and/or prevention of inflammation and cutaneous photodamage and photoprotection of the skin with a water soluble extract from plant of solanum genus |
| CN102872260A (en) * | 2011-06-10 | 2013-01-16 | 德英生物科技股份有限公司 | Method for treating and/or preventing inflammation and cutaneous photodamage by using water soluble extract from plant of Solanum genus and having photoprotective effect |
| CN106577844B (en) * | 2016-11-15 | 2019-08-30 | 中国农业科学院麻类研究所 | Prevent and treat capsicum epidemic disease preparation, its preparation and application |
| CN106509579B (en) * | 2016-11-24 | 2017-06-30 | 张锦超 | A kind of preparation method of black nightshade fruit product |
| CN107260685B (en) * | 2017-06-02 | 2020-11-24 | 神威药业集团有限公司 | Preparation method of solanum nigrum fruit formula granules |
| CN108324798A (en) * | 2018-05-10 | 2018-07-27 | 李明慧 | A kind of converted products and purposes of S. photeinocarpum |
| CN111214581A (en) * | 2018-11-26 | 2020-06-02 | 天津中新药业集团股份有限公司研究院分公司 | Preparation method and application of solanum nigrum fruit total alkaloids |
| CN109674884A (en) * | 2019-02-01 | 2019-04-26 | 李明慧 | Black nightshade product and processing method and purposes |
| CN109620885A (en) * | 2019-02-02 | 2019-04-16 | 李明慧 | A kind of black nightshade converted products and application thereof |
| CN110501200B (en) * | 2019-09-23 | 2022-05-03 | 吉林师范大学 | Method for extracting and separating effective components of recyclable black nightshade |
| CN111493255A (en) * | 2020-04-15 | 2020-08-07 | 吉林创岐生态农业技术开发有限公司 | Solanum torvum solid beverage and application thereof |
| CN114767766A (en) * | 2022-04-13 | 2022-07-22 | 陕西海斯夫生物工程有限公司 | Extraction method of black nightshade fruit extract |
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