CN101884651B - Water extract of turpinia formosana leaves and preparation method and application thereof - Google Patents
Water extract of turpinia formosana leaves and preparation method and application thereof Download PDFInfo
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Abstract
The invention relates to a water extract of turpinia formosana leaves, which is obtained from extracting the turpinia formosana leaves by adding water. The specific extraction method is as follows: taking the turpinia formosana leaves, and adding water to decoct twice or three times (1-3h for each time); filtering, merging, decompressing and concentrating the obtained decoction liquid into the relative density up to 1.18-1.27 (55-60 DEG C); and cooling, adding ethanol until the alcohol content reaches 40-60%, evenly stirring, keeping standing for 12-48h, filtering, decompressing and concentrating the obtained filtrate until the filtrate is dry to obtain the water extract of the turpinia formosana leaves. The extract can be taken as a neuraminidase inhibitor to prevent and treat influenza, and can be prepared into pharmaceutically acceptable dosage forms.
Description
Technical field
The present invention relates to a kind of water extract of turpinia formosana leaves.
Background technology
Influenza virus is serious day by day at present; And influenza virus and respiratory tract disease and the systemic disease that causes are closely related; China is one of influenza country occurred frequently, and is not only populous, and living habit also helps the relay of influenza virus; The number of times of falling ill for each person every year reaches 0.3～0.7 time not to be waited, and key population reaches 2～4 times.There is serious threat in influenza to the mankind, New Development kind influenza virus especially, and it not only causes other infected by microbes of Secondary cases, and can directly cause organ to destroy and the allergy causing death.But the method and the treatment means that do not have efficacious therapy influenza virus and disease (influenza) thereof at present as carry out the inflammatory reaction treatment to clinical symptoms, are cured the symptoms, not the disease.The method of present modal prevention and treatment influenza virus is to use influenza virus vaccine inoculation, non-specific anti virus herb or Western medicine oseltamivir phosphate (oseltamivir phosphate capsule)-neuraminidase inhibitor.Yet influenza virus vaccine prophylactic immunization drawback is that the inoculation crowd is selective, is not that everybody can inoculate.In addition, the influenza virus vaccine protective rate is not high, and guard time is also lacked (3～6 months).Though most of non-specific anti virus herbs lay claim to antivirus action at present, be not to influenza virus, and antivirus action mechanism are unclear.At many kind treatment affection of exogenous wind-cold of our former studies, the Chinese prescription of the flu that doctor trained in Western medicine is thought does not have the effect of direct resisiting influenza virus.Briefly, the Chinese prescription of not every treatment flu all has the effect of clear and definite inhibition influenza virus.Western medicine " oseltamivir phosphate (oseltamivir phosphate capsule; (3R; 4R, 5S)-amino-3 (1-third the 2-ethoxyethyl acetate)-1-cyclohexene-1 carboxylic acid, ethyl ester phosphate of 4-acetamide-5-) " be neuraminidase inhibitor, influenza virus is got into cell has specific inhibitory effect; But not only cost an arm and a leg, and have that some patients take that the back vomiting occurs, feels sick, side reactions such as insomnia, headache, stomachache, diarrhoea, dizziness, fatigue, nasal obstruction, pharyngalgia and cough.Oseltamivir phosphate capsule is that the esterase that is positioned at liver and intestinal in vivo is converted into active metabolite and plays the neuraminidase effect that suppresses, and also will influence its drug effect if patient's liver and intestinal organ dysfunction are undesired.In addition, the renal insufficiency patient also wants careful usefulness.
Summary of the invention
The purpose of this invention is to provide a kind of water extract of turpinia formosana leaves; It can suppress influenza neuraminidase hydrolysis cell surface sialic acid; Cause influenza virus not combine with cell surface receptor and to get in the cell; And reduce influenza virus in intracellular generation, and duplicate thereby suppress influenza virus effectively pointedly, more importantly be that the present invention provides a kind of water extract of turpinia formosana leaves can overcome the side reaction drawback of existing medicine.
Another object of the present invention has provided a kind of method for preparing of water extract of turpinia formosana leaves.
Another object of the present invention has provided the purposes of the medicine of water extract of turpinia formosana leaves preparation prevention and treatment influenza and complication disease thereof.
Another object of the present invention has provided the pharmaceutical preparation that contains water extract of turpinia formosana leaves.
Water extract of turpinia formosana leaves of the present invention, it is through obtaining the Turpinia pomifera(Roxb) D O. extracting in water, and concrete method for distilling is following: get leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 or 3 times, each 1～3 hour; Filter, merge decoction liquor, be evaporated to relative density 1.18～1.27 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 40～60%, stir; Left standstill 12～48 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Preferably, concrete method for distilling is following: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 times, each 1.5～3 hours; Filter, merge decoction liquor twice, be evaporated to relative density 1.20～1.25 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 45～60%, stir; Left standstill 12～36 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Preferred, concrete method for distilling is following: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 times, each 3 hours; Filter, merge decoction liquor twice, be evaporated to relative density 1.22 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 50～60%, stir; Left standstill 36 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Preferred, concrete method for distilling is following: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 times, each 2 hours; Filter, merge decoction liquor twice, be evaporated to relative density 1.23 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 50～55%, stir; Left standstill 24 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Preferred, concrete method for distilling is following: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 times, each 1.5 hours; Filter, merge decoction liquor twice, be evaporated to relative density 1.20 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 45～53%, stir; Left standstill 22 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Preferably, concrete method for distilling is following: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 3 times, each 1～1.5 hour; Filter, merge three times decoction liquor, be evaporated to relative density 1.18～1.20 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 45～55%, stir; Left standstill 12～20 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Preferred, concrete method for distilling is following: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 3 times, each 1 hour 20 minutes; Filter, merge three times decoction liquor, be evaporated to relative density 1.19 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 48%, stir; Left standstill 14 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Preferred, concrete method for distilling is following: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 3 times, each 1 hour; Filter, merge three times decoction liquor, be evaporated to relative density 1.18 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 47%, stir; Left standstill 20 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Preferably, wherein add 6～12 times of water gagings at every turn.
The purposes that the present invention also provides water extract of turpinia formosana leaves to be used to prepare prevention and to treat the medicine of influenza and complication disease thereof.
Preferably, water extract of turpinia formosana leaves is used to suppress influenza virus.
Preferably, water extract of turpinia formosana leaves is used to suppress influenza virus FM1.
Preferably, water extract of turpinia formosana leaves is used to suppress neuraminidase.
Preferably, its complication is meant renal failure.
Preferably, its complication is meant splenic trauma or/and injury of lung.
At last, the present invention also provides a kind of water extract of turpinia formosana leaves preparation, and said preparation is a main active with the above-mentioned water extract of turpinia formosana leaves of the present invention.
Experimental data proves; Water extract of turpinia formosana leaves of the present invention can extract effective inhibition neuraminic acid enzyme component, and water extract of turpinia formosana leaves is along with the using dosage size variation, and it suppresses the active ability of neuraminidase; Be also corresponding the changing of height of neuraminic acid enzyme inhibition rate; And one-tenth positive correlation, water extract of turpinia formosana leaves can be through suppressing influenza virus surface neuraminidase, and then suppress that influenza virus gets into the cell the inside, the influenza virus that suppresses to have got into the cell the inside duplicates, breeds; Thereby infection, the growth of influenza virus pair cell have been reduced; And prevention and treatment influenza and complication thereof, can also suppress influenza neuraminidase hydrolysis cell surface sialic acid, cause influenza virus not combine with cell surface receptor and to get in the cell; And reduce influenza virus in intracellular generation, more importantly be that the present invention provides a kind of water extract of turpinia formosana leaves can overcome the side reaction drawback of existing medicine.
And medical science and study of pharmacy personnel can't not do the inhibition influenza infection, duplicate in advance, or under the prerequisite of the experiment of inhibition neuraminidase, learn that in advance water extract of turpinia formosana leaves has prevention and treats the good result that the influenza virus sexuality is emitted.
In sum, water extract of turpinia formosana leaves provided by the invention and preparation method thereof and purposes are brought significant technique effect.
The specific embodiment
Water extract of turpinia formosana leaves of the present invention, it is through obtaining the Turpinia pomifera(Roxb) D O. extracting in water, and concrete method for distilling is following: get leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 or 3 times, each 1～3 hour; Filter, merge decoction liquor, be evaporated to relative density 1.18～1.27 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 40～60%, stir; Left standstill 12～48 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Preferably, wherein add 6～12 times of water gagings at every turn.
Embodiment 1
Method for preparing: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 times, each 3 hours, add 8 times of water gagings for the first time, add 8 times of water gagings for the second time; Decoct secondary filter, merge decoction liquor twice, be evaporated to relative density 1.22 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 52%, stir; Left standstill 36 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Embodiment 2
Method for preparing: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 times, each 3 hours, add 8 times of water gagings for the first time, add 9 times of water gagings for the second time; Filter, merge decoction liquor twice, be evaporated to relative density 1.22 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 58%, stir; Left standstill 36 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Embodiment 3
Method for preparing: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 times, each 2 hours, add 9 times of water gagings for the first time, add 8 times of water gagings for the second time; Filter, merge decoction liquor twice, be evaporated to relative density 1.23 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 51%, stir; Left standstill 24 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Embodiment 4
Method for preparing: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 times, each 2 hours, add 9 times of water gagings for the first time, add 9 times of water gagings for the second time; Filter, merge decoction liquor twice, be evaporated to relative density 1.23 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 53%, stir; Left standstill 24 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Embodiment 5
Method for preparing: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 times, each 1.5 hours, add 7 times of water gagings for the first time, add 10 times of water gagings for the second time; Filter, merge decoction liquor twice, be evaporated to relative density 1.20 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 47%, stir; Left standstill 22 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Embodiment 6
Method for preparing: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 times, each 1.5 hours, add 6 times of water gagings for the first time, add 12 times of water gagings for the second time; Filter, merge decoction liquor twice, be evaporated to relative density 1.20 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 50%, stir; Left standstill 22 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Embodiment 7
Method for preparing: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 3 times, each 1 hour 20 minutes, add 9 times of water gagings for the first time, add 8 times of water gagings for the second time; Add 7 times of water gagings for the third time, filter, merge three times decoction liquor, be evaporated to relative density 1.19 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 48%, stir; Left standstill 14 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Embodiment 8
Method for preparing: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 3 times, each 1 hour, 8 times of water gagings added 7 times of water gagings for the second time for the first time; Add 6 times of water gagings for the third time, filter, merge three times decoction liquor, be evaporated to relative density 1.18 (55～60 ℃); Put coldly, add ethanol and make and contain the alcohol amount and reach 47%, stir; Left standstill 20 hours, and filtered, filtrate decompression is concentrated into dried water extract of turpinia formosana leaves.
Experimental data 1: the optimization research of Turpinia pomifera(Roxb) D O. water extraction process
Leaf of Turpinia pomifera (Roxb) D O. mainly contains chemical compounds such as flavonoid, organic acid, and the factor that influences the water extraction effect mainly contains amount of water, decocting time, decoction number of times, and therefore, we establish 3 levels to above factor, with L9 (3
4) show to carry out orthogonal test, investigating index is the total flavones extraction ratio, the result sees table 1, table 2.
Table 1 decocting process factor and water-glass
Table 2 decocting boils technology orthogonal test table and result
Variance analysis
Orthogonal experiments is carried out variance analysis, and the result sees table 3.
Table 3 total flavones extraction ratio analysis of variance table
*F
0.05(2，2)＝19.00；
**F
0.01(2，2)＝99.00
With total flavones extraction ratio in the water decoction serves as to investigate index, shows that by extreme difference R value size in the table 2 primary and secondary of each factor effect is C＞B＞A; The results of analysis of variance shows: the influence of C factor has utmost point significance meaning (P＜0.01), and the influence of A, B two factors there are no significant meaning (P＞0.05) is with A
3B
3C
3For good.Analyze in conjunction with the K value, select A
3B
3C
3Be optimised process, promptly get leaf of Turpinia pomifera (Roxb) D O., add 10 times of water gagings and decoct each 2 hours 3 times.
Experimental data 2: water extract of turpinia formosana leaves is to the active inhibitory action of neuraminidase
Get the water extract of turpinia formosana leaves that embodiment 3 method for preparinies are obtained, add suitable quantity of water and make dissolving, use tiring of neuraminidase inhibitor identification kit mensuration water extract of turpinia formosana leaves inhibition neuraminidase (N1) and see table 4.
(1). standard curve is prepared: a. every hole in 96 hole luciferase targets adds 70 μ l neuraminidases and detects buffer; B. every hole adds 0,1,2,5,7.5,10 μ l H5N1 neuraminidases more respectively; C. every hole adds 0～20 μ l Milli-Q water again.
(2). the preparation of sample detection: a. every hole in 96 hole luciferase targets adds 70 μ l neuraminidases and detects buffer; B. every hole adds 10 μ l H5N1 neuraminidases again; C. every hole adds 0～10 μ l water extract of turpinia formosana leaves sample again; D. every hole adds 0～10 μ l Milli-Q water again.
(3). detect step:
A. vibrate the about 1min of mixing;
B.37 ℃ hatch 2min inhibitor and H5N1 neuraminidase are fully interacted, the sample of doing standard curve is also hatched together;
C. every hole adds 10 μ l neuraminidase fluorogenic substrates;
D. vibrate the about 1min of mixing again;
E.37 carry out fluoremetry after ℃ hatching 20～30min.Excitation wavelength is 360nm, and emission wavelength is 440nm.
(4). calculate the inhibition percentage ratio of sample according to standard curve, and calculate the IC50 of water extract of turpinia formosana leaves after doing concentration curve for the H5N1 neuraminidase for the H5N1 neuraminidase.The suppression ratio IC50 that water extract of turpinia formosana leaves reaches neuraminidase is 28.98mg/ml.See table 4.
Table 4. water extract of turpinia formosana leaves suppresses the activity of neuraminidase
Can be clear that according to above-mentioned experimental result:
(1). water extract of turpinia formosana leaves can extract effective inhibition neuraminic acid enzyme component;
(2). water extract of turpinia formosana leaves is along with the using dosage size variation, and it suppresses the active ability of neuraminidase, and promptly the height of neuraminic acid enzyme inhibition rate is also corresponding changes, and becomes positive correlation;
(3). visible by above-mentioned experiment; Water extract of turpinia formosana leaves can be through suppressing influenza virus surface neuraminidase; And then suppress that influenza virus gets into the cell the inside, the influenza virus that suppresses to have got into the cell the inside duplicates, breeds; Thereby reduced infection, the growth of influenza virus pair cell, and prevention and treatment influenza and complication thereof.
Medical science and study of pharmacy personnel can't not do the inhibition influenza infection, duplicate in advance, or under the prerequisite of the experiment of inhibition neuraminidase, learn that in advance water extract of turpinia formosana leaves has prevention and treats the good result that the influenza virus sexuality is emitted.
Experimental data 3: water extract of turpinia formosana leaves infects the inhibitory action of Embryo Gallus domesticus to influenza virus
Get water extract of turpinia formosana leaves, use influenza virus A-prime Mus lung adapted strain (FM1) and (H1N1) identify that water extract of turpinia formosana leaves suppresses the ability that the FM1 influenza virus is duplicated and suppresses in Embryo Gallus domesticus.(1). FM1 influenza virus liquid is inoculated in 10d no-special pathogen in age chick embryo allantois intracavity, and every embryo 0.2ml is hatched 72h for 37 ℃, observes and calculate half Embryo Gallus domesticus infective dose (EID50).(2). water extract of turpinia formosana leaves adopts the toxic action of Embryo Gallus domesticus; Physiological saline solution is done to be inoculated in 10d no-special pathogen in age chick embryo allantois intracavity behind the serial dilution to water extract of turpinia formosana leaves; Every embryo 0.2ml, each concentration is inoculated 6 embryos, hatches for 37 ℃; Observe Embryo Gallus domesticus growth promoter situation, can survive the Cmax of 96h as the TD of medicine with Embryo Gallus domesticus.(3). water extract of turpinia formosana leaves inhibitory action to influenza virus in Embryo Gallus domesticus adopts; 0.1ml influenza virus liquid and different dilution water extract of turpinia formosana leaves mix, 37 ℃ of effect 2h are inoculated in 10d no-special pathogen in age chick embryo allantoic cavity; Every winding kind 6 embryos are hatched 72h for 37 ℃.The virus attack amount is 50EID50, establishes virus control, physiological saline solution normal control simultaneously, calculates the median effective dose (ED50) of water extract of turpinia formosana leaves to viral inhibition.
(1) the .FM1 influenza virus is calculated through the Reed-Muench method the virulence of Embryo Gallus domesticus, and its EID50 is 10
-5.07(2). after water extract of turpinia formosana leaves is inoculated in Embryo Gallus domesticus, its growth promoter and normal control group basically identical.The 96h Embryo Gallus domesticus is all survived.Embryo Gallus domesticus gives water extract of turpinia formosana leaves stock solution and does not see chicken embryo death, so can think that TD0 is 17.0g/100ml.(3). water extract of turpinia formosana leaves inhibitory action to influenza virus in Embryo Gallus domesticus is seen table 5.
Table 5. water extract of turpinia formosana leaves infects the inhibitory action of Embryo Gallus domesticus to influenza virus
Compare with the virus control group:
*P＜0.05
Can know that by table 5 water extract of turpinia formosana leaves has significant inhibitory effect (P＜0.05), ED at 10.0～160.0mg/ml to influenza virus
50Be 43.82mg ± 0.16mg/ml, TI is 5.17 ± 1.69.
Experimental data 4: water extract of turpinia formosana leaves influences the FM1 influenza virus
Get water extract of turpinia formosana leaves, use influenza virus A-prime Mus lung adapted strain (FM1) and (H1N1) identify that water extract of turpinia formosana leaves suppresses the ability of FM1 influenza virus virulence.(1) .FM1 adopts cell ID 50, median infective dose (TCID50) micromethod to the toxicity test of Testis et Pentis Canis passage cell (MDCK).(2). water extract of turpinia formosana leaves adopts the DMEM of serum-free that water extract of turpinia formosana leaves is done to be inoculated in the mdck cell hole that forms monolayer behind the serial dilution to the toxicity test of mdck cell; Every hole 100 μ l; Each dilution factor repeats 4 holes, establishes the normal cell contrast simultaneously.Culture plate is put 37 ℃, 5%CO
2Cultivate in the incubator, observation of cell pathological changes every day (CPE) is observed 3d continuously, with " +～++ ++ " the record result, press the Reed-Muench method and calculate medicine median toxic concentration (TD50) and maximal non-toxic concentration (TD0).(3). water extract of turpinia formosana leaves suppresses the effect of FM1 influenza virus and measures: mdck cell 5 * 10
5/ ml, every hole 100 μ l, in 96 orifice plates, 37 ℃, the interior cultivation of 5%CO2 incubator are inhaled and remove culture fluid in the hole next day, add 100TCID50 influenza virus liquid, every hole 100 μ l, supernatant is removed in suction behind 37 ℃ of absorption 1h.Wash 2 times with phosphate buffer (PBS); TD0 with medicine is the 1st hole, and the DMEM liquid of reuse serum-free is made serial dilution to water extract of turpinia formosana leaves, adds respectively in the cell of above-mentioned infective virus; Establish virus control and normal control group simultaneously, 37 ℃, 5%CO
2Cultivate in the incubator, observe the mdck cell characteristics of lesion that influenza virus produces every day, i.e. cell monolayer degeneration becomes circle etc., and 3d calculates 50% of medicine and suppresses pathological changes concentration (IC50) and therapeutic index (TI) continuously.The calculating of TI: TI=TD50/IC50, the TI value is big more, shows that the safety range of medicine is big more.With Kruskal-Walis and Mann-Whitney method of inspection comparative test group and the cytopathic difference of virus control group; Drug dose and suppression ratio that virus infected cell is avoided cytopathy (CPE) takes place are carried out correlation analysis, judge whether amount validity response relation.
(1) the .FM1 influenza virus is calculated through the Reed-Muench method the virulence of mdck cell, and its TCID50 is 10
-4.81(2). the TD0 of water extract of turpinia formosana leaves mdck cell is respectively 1681.1mg ± 40.22g/100ml.(3). after water extract of turpinia formosana leaves made serial dilution, the 100TCID50 influenza virus is carried out inhibition test, median effective dose IC50 and the TI value of calculating medicine are big or small, and the result sees table 6.
Table 6. water extract of turpinia formosana leaves is to the IC of FM1 influenza virus
50(mg/100ml) and TI (x ± s)
Can be known that by table 6 IC50 of water extract of turpinia formosana leaves is low, TI is high.Water extract of turpinia formosana leaves suppresses the cytopathogenic effect of FM1 influenza virus all to be strengthened along with the increase of drug dose.Drug dose and medicine are shown that to the correlation analysis that the suppression ratio of CPE carries out the dosage of water extract of turpinia formosana leaves and medicine are to being tangible positive correlation between the CPE suppression ratio.
Experimental data 5: water extract of turpinia formosana leaves is to the spleen index and the exponential influence of lung of influenza virus infection FM1 strain in the mice body
Get water extract of turpinia formosana leaves, use influenza virus A-prime Mus lung adapted strain (FM1) and (H1N1) identify the dead protective effect of water extract of turpinia formosana leaves influenza virus infection FM1 strain in the mice body.(1). influenza virus FM1 strain virus is inoculated every group of 10 BALB/C mices respectively, male and female half and half after doing 10 times of doubling dilutions.After the slight anesthesia of ether, give and different dilution viruses respectively for every group, every mice collunarium is inoculated 20 μ l.Observe the dead mouse situation of 10d, calculating LD50 by the Reed-Muench method is 10
-1.36So confirm that the used modeling concentration of experiment is 10LD50.(2). water extract of turpinia formosana leaves is to the dead protective effect of influenza virus infection FM1 strain in the mice body: normal control group, influenza virus FM1 strain virus matched group, water extract of turpinia formosana leaves 5.0mg/ml, 10.0mg/ml, 20.0mg/ml, 40.0mg/ml dose groups etc. are irritated stomach respectively, irritate gastric capacity and only are 0.4ml/.Behind the 3d, each is organized under the slight anesthesia of ether with 10LD50 influenza virus FM1 strain collunarium infecting mouse 20 μ l/ except that the normal control group.The normal control group gives the normal saline with volume simultaneously.4 groups of administration are continued administration, normal control group and influenza virus FM1 strain virus control group administered physiological saline 8 days, and dosage is the same.Day by day observe animal morbidity and record death toll, observed altogether 14 days, calculate mortality rate (mortality rate=every group of death toll/every group of total mice * 100%), the result sees table 7.(3). water extract of turpinia formosana leaves is to the exponential influence of influenza virus infection FM1 strain lung in the mice body: normal control group, influenza virus FM1 strain virus matched group, water extract of turpinia formosana leaves 20.0mg/ml, 40.0mg/ml, 80.0mg/ml, 160.0mg/ml dose groups etc. are irritated stomach respectively, irritate gastric capacity and only are 0.4ml/.After 3 days, each is organized under the slight anesthesia of ether with 1.0LD50 influenza virus FM1 strain collunarium infecting mouse 20 μ l/ except that the normal control group.The normal control group gives the normal saline with volume simultaneously.4 groups of administration are continued administration, normal control group and influenza virus FM1 strain virus control group administered physiological saline 8 days, and dosage is the same.Behind viral infection, put to death mice on the 8th day, weigh, get lung and claim that lung is heavy, calculates lung index (lung index=lung quality/body constitution amount * 100%); In addition, get spleen and claim that spleen is heavy, calculate spleen index (spleen index=spleen quality/body constitution amount * 100%), the result sees table 7.
Table 7. water extract of turpinia formosana leaves is to the death protection result of influenza virus infection FM1 strain in the mice body
Annotate: ※ ※ P＜0.01VS influenza virus model group ※ P＜0.05VS influenza virus model group
Table 8. water extract of turpinia formosana leaves is to the spleen index and the exponential influence of lung of influenza virus infection FM1 strain in the mice body
Annotate: #P＜0.05 VS normal control group is annotated: ##P＜0.01 VS normal control group
※ ※ p＜0.001 VS influenza virus model group ※ p＜0.05 VS influenza virus model group
(1). can know that by table 7 water extract of turpinia formosana leaves has significant protective effect (p＜0.01-p＜0.05) at 40.0～160.0mg/ml to the influenza virus infecting mouse.
(2). can know that by table 8 water extract of turpinia formosana leaves has significant effect (p＜0.01-p＜0.05) at 80.0～160.0mg/ml to the lung index suppression ratio of influenza virus infecting mouse.
Experimental data 6: water extract of turpinia formosana leaves is to the influence of chronic renal failure
1) SD rat, 200～240g, Shanghai west pul-Bi Kai laboratory animal responsibility company limited, the animal quality certification number: SCXK (Shanghai) 2003-0002
2) reagent and medicine adenine, adenine, (content＞98% is the import packing, Chinese Shanghai, lot number 20010520), the water extract of turpinia formosana leaves that embodiment 3 method for preparinies are obtained;
3) test method: male SD rat, about body weight 220g, earlier feed 10 days normal growths with normal diet after, be divided into normal control group, administration experimental group and modeling matched group at random by body weight, 13～15 every group.Administration experimental group and modeling matched group are irritated stomach with adenine and are made chronic renal failure (CRF) model, irritate stomach with adenine 320mg/ (kgd), only process the about 2ml/ of suspension with distilled water, totally 20 days; Administration after 20 days; The administration experimental group is irritated stomach with water extract of turpinia formosana leaves 2.4g/ (kgd), and water extract of turpinia formosana leaves is processed suspension solution (0.4g/ml) with distilled water, and about at every turn 2ml/ only; Stomach is irritated in gradation; Normal control group and modeling matched group are irritated stomach with equal-volume water, and administration was used etherization with rat after 35 days, and each item index is observed in the blood sampling of Mus arteria caudalis.
4) result:
Table 9. water extract of turpinia formosana leaves is to the influence of CRF kidney of rats function
Annotate: through the T check, administration experimental group and the comparison of modeling matched group "
*" expression P＜0.05, "
*" expression P＜0.01
This experiment is looked sidelong at purine with gland and is irritated after stomach sets up rat CRF model, modeling control rats lethargy, and body weight obviously reduces; Serum Bun, Ser obviously raise, and Hb obviously descends, and Ret obviously raises; Showing the rat impaired renal function, after giving the treatment of Turpinia pomifera(Roxb) D O. ethanol extraction. rat spirit is obviously bestirred oneself, and body weight is recovered gradually; Improve creatinine clearance rate, promote the discharge of metabolite, reduce serum BUN, SCr concentration; Renal function and anemia are obviously improved, and show that water extract of turpinia formosana leaves has preventive and therapeutic effect to kidney of rats damage due to the adenine.
Can be clear that according to above-mentioned experimental result:
(1). water extract of turpinia formosana leaves can extract effectively preventing chronic renal failure composition;
(2). water extract of turpinia formosana leaves suppresses the active effect of neuraminidase simultaneously having, also have the effect of control chronic renal failure, has overcome oseltamivir phosphate capsule sternly to clear ability of creatinine and renal failure crowd's side effect, on the contrary prevention effect in addition.
Claims (12)
1. water extract of turpinia formosana leaves is used to prepare the purposes of the medicine of prevention and treatment influenza and complication disease thereof, and said complication is renal failure, splenic trauma or/and injury of lung, and wherein said water extract of turpinia formosana leaves is to obtain through following method for distilling: get leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 or 3 times; Each 1～3 hour, filter, merge decoction liquor; Be evaporated to relative density and be determined as 1.18～1.27 for 55～60 ℃, put coldly, add ethanol and make and contain the alcohol amount and reach 40～60%; Stir, left standstill 12～48 hours, filter; Filtrate decompression is concentrated into dried, gets water extract of turpinia formosana leaves.
2. purposes as claimed in claim 1, wherein water extract of turpinia formosana leaves obtains through following method for distilling: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 times, each 1.5～3 hours, filter; Merge decoction liquor twice, be evaporated to relative density and be determined as 1.20～1.25 for 55～60 ℃, put cold; Add ethanol and make and contain the alcohol amount and reach 45～60%, stir, left standstill 12～36 hours; Filter, filtrate decompression is concentrated into dried, gets water extract of turpinia formosana leaves.
3. purposes as claimed in claim 2, wherein water extract of turpinia formosana leaves obtains through following method for distilling: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 times, each 3 hours, filter; Merge decoction liquor twice, be evaporated to relative density and be determined as 1.22 for 55～60 ℃, put cold; Add ethanol and make and contain the alcohol amount and reach 50～60%, stir, left standstill 36 hours; Filter, filtrate decompression is concentrated into dried, gets water extract of turpinia formosana leaves.
4. purposes as claimed in claim 2, wherein water extract of turpinia formosana leaves obtains through following method for distilling: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 times, each 2 hours, filter; Merge decoction liquor twice, be evaporated to relative density and be determined as 1.23 for 55～60 ℃, put cold; Add ethanol and make and contain the alcohol amount and reach 50～55%, stir, left standstill 24 hours; Filter, filtrate decompression is concentrated into dried, gets water extract of turpinia formosana leaves.
5. purposes as claimed in claim 2, wherein water extract of turpinia formosana leaves obtains through following method for distilling: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 2 times, each 1.5 hours, filter; Merge decoction liquor twice, be evaporated to relative density and be determined as 1.20 for 55～60 ℃, put cold; Add ethanol and make and contain the alcohol amount and reach 45～53%, stir, left standstill 22 hours; Filter, filtrate decompression is concentrated into dried, gets water extract of turpinia formosana leaves.
6. purposes as claimed in claim 1, wherein water extract of turpinia formosana leaves obtains through following method for distilling: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 3 times, each 1～1.5 hour, filter; Merge three times decoction liquor, be evaporated to relative density and be determined as 1.18～1.20 for 55～60 ℃, put cold; Add ethanol and make and contain the alcohol amount and reach 45～55%, stir, left standstill 12～20 hours; Filter, filtrate decompression is concentrated into dried, gets water extract of turpinia formosana leaves.
7. purposes as claimed in claim 6, wherein water extract of turpinia formosana leaves obtains through following method for distilling: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 3 times, each 1 hour 20 minutes, filter; Merge three times decoction liquor, be evaporated to relative density and be determined as 1.19 for 55～60 ℃, put cold; Add ethanol and make and contain the alcohol amount and reach 48%, stir, left standstill 14 hours; Filter, filtrate decompression is concentrated into dried, gets water extract of turpinia formosana leaves.
8. purposes as claimed in claim 6, wherein water extract of turpinia formosana leaves obtains through following method for distilling: get the leaf of Turpinia pomifera (Roxb) D O. decocte with water 3 times, each 1 hour, filter; Merge three times decoction liquor, be evaporated to relative density and be determined as 1.18 for 55～60 ℃, put cold; Add ethanol and make and contain the alcohol amount and reach 47%, stir, left standstill 20 hours; Filter, filtrate decompression is concentrated into dried, gets water extract of turpinia formosana leaves.
9. purposes as claimed in claim 1, wherein water extract of turpinia formosana leaves extracts through add 6～12 times of water gagings at every turn.
10. each described purposes of claim 1～9 is characterized in that suppressing influenza virus.
11. each described purposes of claim 1～9 is characterized in that suppressing influenza virus FM1.
12. each described purposes of claim 1～9 is characterized in that suppressing neuraminidase.
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| CN106279459B (en) * | 2016-08-25 | 2018-09-04 | 湖南中医药大学 | The extracting method of polysaccharide and its application in a kind of acute turpinia leaf |
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| CN1857343A (en) * | 2006-04-11 | 2006-11-08 | 徐蕾 | Chinese medicine composition for preventing and treating viral cold and throat disease and its preparing method |
| CN1872106A (en) * | 2006-04-18 | 2006-12-06 | 徐蕾 | Application of wild basil circle leaves in treating disease of virulence cold |
| CN101249118A (en) * | 2007-02-25 | 2008-08-27 | 胡军 | Chinese medicine extract and medicine use thereof |
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| CN1843326A (en) * | 2006-02-13 | 2006-10-11 | 徐蕾 | Pharmaceutical toothpaste containing Turpinia arguta Seem |
| CN1857343A (en) * | 2006-04-11 | 2006-11-08 | 徐蕾 | Chinese medicine composition for preventing and treating viral cold and throat disease and its preparing method |
| CN1872106A (en) * | 2006-04-18 | 2006-12-06 | 徐蕾 | Application of wild basil circle leaves in treating disease of virulence cold |
| CN101249118A (en) * | 2007-02-25 | 2008-08-27 | 胡军 | Chinese medicine extract and medicine use thereof |
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