CN1872282A - Active extractive of Foliumnerviliae, preparation method and application - Google Patents

Abstract

An active extract of hairleaf nervilia is prepared from hairleaf nervilia through extracting by water, or supercritical CO2, or alcohol, and purifying by polyamide resin or macroreticular resin. It contains general flavone (10-90%) and general amino acid (10-90%) and can be used for treating fever, inflammation, pain, cough, etc, improving immunity and resisting against bacterium and virus.

Description

A kind of Folium Nerviliae fordii activity extract and its production and application

Technical field

The present invention relates to the field of Chinese medicines, be specifically related to Folium Nerviliae fordii extract and its production and application.

Technical background

Folium Nerviliae fordii derives from underground tuber and the herb of orchid Nervilia fordii Nerviliae Fordii (Hance) Schltr, be distributed in ground such as Guangdong, Guangxi, Sichuan, Yunnan, being the special product medical material in Guangxi, Guangdong, is China's Chinese medicine, also is the main Chinese medicinal materials of China's foreign exchange earning.Folium Nerviliae fordii energy nourishing the lung to arrest cough, heat-clearing and toxic substances removing, the silt pain relieving of loosing.Cure mainly the pulmonary tuberculosis spitting of blood, cough due to lung-heat, stomatitis, laryngopharynx swelling and pain, scrofula, sore swollen toxin, traumatic injury.The clinical research of Folium Nerviliae fordii in recent years at present is many based on the treatment pulmonary disease.Qiu Zhinan etc. (the clinical new usefulness of Folium Nerviliae fordii, Guangzhou Medical College journal, 02 phase of nineteen ninety-five) think Folium Nerviliae fordii except there being heat clearing away to also have the effect of dissipating phlegm and removing blood stasis pain relieving bringing down a fever, and the compatibility Radix Angelicae Sinensis also is usually used in the treatment of cold cough.(the clinical observation of Gekko Swinhonis tea treatment acute bronchitis such as history oats ice, current clinical medicine biotechnology magazine, 1997 02 phases) with the Folium Nerviliae fordii be principal agent, compatibility Flos Sauropi changiani leaf, Flos Farfarae, Ba Ji etc. form Gekko Swinhonis tea treatment acute bronchitis, and total effective rate reaches 95%.Folium Nerviliae fordii is applied to cough, expectorant, the card breathed heavily, and the power of eliminating the phlegm is greater than Bulbus Fritillariae Uninbracteatae, and the power of relievining asthma is better than Herba Ephedrae, and the effect of antitussive is better than Flos Farfarae, and the merit of transferring lung qi is arranged, and disease-resistant virulence is better than antivirus oral liquid, is first-selected good medicine to the cough with asthma patient of body exogenous pathogen victory.The Folium Nerviliae fordii activity extract is not reported at present.

Summary of the invention

The purpose of this invention is to provide a kind of Folium Nerviliae fordii activity extract.

Another object of the present invention provides a kind of preparation method of Folium Nerviliae fordii activity extract.

Another object of the present invention provides the application of a kind of Folium Nerviliae fordii activity extract in the viral respiratory system disease medicine of preparation treatment.

The Folium Nerviliae fordii of indication of the present invention refers in particular to the orchid Nervilia fordii, and formal name used at school is underground tuber and the herb of the plant of Nerviliae Fordii (Hance) Schltr.

Folium Nerviliae fordii activity extract of the present invention contains 10%～90% total flavones and 10%～90% total amino acids.

Folium Nerviliae fordii activity extract of the present invention is white in color to the yellowish-brown powder.The qualitative identification experiment shows: ninhydrin reaction is positive, and hydrochloric acid-magnesium powder reaction is positive, and the Molish reaction is positive.

Total amino acids detection method of content of the present invention adopts automatic amino acid analyzer (DIONEX) to detect.

It is reference substance that general flavone content detection method of the present invention adopts with the rutin, ultraviolet spectrophotometry.The general flavone content detection method is with reference to pharmacopeia version in 2005, appendix VA ultraviolet-visible spectrophotometry.

Folium Nerviliae fordii activity extract of the present invention, general flavone content preferably 20%, total amino acids content preferably 70%.

Described Folium Nerviliae fordii preparation method of the present invention: Folium Nerviliae fordii is extracted with pure water mixed solvent.The extracting method that extracting method can adopt those skilled in the art to use always.For example can be to soak ultrasonic extraction, heating and refluxing extraction.Alcohol of the present invention can be methanol, ethanol, propanol, isopropyl alcohol wherein one or more.Alcohol of the present invention is ethanol preferably.

The preparation method of Folium Nerviliae fordii activity extract of the present invention can also adopt supercritical carbon dioxide extraction (containing 2.5%～10% ethanol entrainer), the extract solvent removed by evaporation at reduced pressure, add the abundant degreasing decoloring of petroleum ether, petroleum ether does not dissolve partly to remove through the reduction vaporization lyophilization and desolvates, and gets product.

Folium Nerviliae fordii activity extract preparation method of the present invention, can adopt ethanol (determining alcohol is 0～80%) reflux, extract, 2～3 times that in the dry medical material of Folium Nerviliae fordii, adds 6～15 times of amounts, each 0.5～2 hour, merge extractive liquid,, concentrate, use macroporous adsorbent resin, polyamide, anion exchange resin, cation exchange resin purify the Folium Nerviliae fordii activity extract.

Folium Nerviliae fordii activity extract preparation method of the present invention better is, at the dry medical material of Folium Nerviliae fordii, the ethanol (determining alcohol is 30～60%) that adds 6～12 times of amounts refluxes each 1～2 hour 2～3 times, merge extractive liquid,, being concentrated into does not have the alcohol flavor, crosses macroporous adsorbent resin, is washed to that colourless (crossing post liquid and water lotion collects, get reserve liquid), 50～80% ethanol elutions are collected eluent then, and being evaporated to does not have the alcohol flavor, cross polyamide column, be washed to colourlessly, reuse 50～80% ethanol elutions are collected eluent, be evaporated to and do not have the alcohol flavor, dry Folium Nerviliae fordii total flavonoids extract: get reserve liquid and cross cation exchange resin, be washed to colourlessly, 50～80% ethanol elutions are to there not being ninhydrin reaction then, collect pure washing liquid, being evaporated to does not have the alcohol flavor, after anion exchange resin, is washed to colourless, reuse 50～80% ethanol elutions do not have ninhydrin reaction, collect ethanol elution, being evaporated to does not have the alcohol flavor, the dry Folium Nerviliae fordii total amino acids extract that gets; With Folium Nerviliae fordii total flavonoids extract and Folium Nerviliae fordii total amino acids extract mixing Folium Nerviliae fordii activity extract.

Macroporous resin of the present invention is nonpolar macroporous resin, can be AB-8, DP-100.AB-8 preferably.Ion exchange resin is anion exchange resin and cation exchange resin, and wherein anion exchange resin is better with 717 strong base, cation exchange resin is better with 732 strong acid type resins.Polyamide is pharmaceutical type, and granularity is better with 30～60 orders.

Folium Nerviliae fordii activity extract preparation method of the present invention can be to prepare Folium Nerviliae fordii total amino acids extract and Folium Nerviliae fordii total flavonoids extract at first respectively, then Folium Nerviliae fordii total amino acids extract and the mixing of Folium Nerviliae fordii total flavonoids extract is obtained.

Folium Nerviliae fordii total amino acids method for preparing extractive of the present invention is, in the dry medical material of Folium Nerviliae fordii, add 10～15 times of water gagings and soak 24～48h, be evaporated to small size, add ethanol and make precipitation, standing over night, supernatant are evaporated to does not have the alcohol flavor, crosses cation exchange resin, be washed to colourless, 50～80% ethanol elutions are collected pure washing liquid to there not being ninhydrin reaction then, and being evaporated to does not have the alcohol flavor, cross anion exchange resin, be washed to colourlessly, reuse 50～80% ethanol elutions are collected ethanol elution to there not being ninhydrin reaction, being evaporated to does not have the alcohol flavor, the dry Folium Nerviliae fordii total amino acids extract that gets.

Folium Nerviliae fordii total flavonoids method for preparing extractive of the present invention is, 80% alcohol reflux 2～3 times that in the dry medical material of Folium Nerviliae fordii, adds 6～12 times of amounts, each 1～2 hour, merge extractive liquid,, being evaporated to does not have the alcohol flavor, crosses macroporous adsorbent resin, is washed to colourless, 50～80% ethanol elutions are to colourless then, collect eluent, being evaporated to does not have the alcohol flavor, crosses polyamide column, be washed to colourless, reuse 50～80% ethanol elutions are collected eluent to colourless, and being evaporated to does not have the alcohol flavor, drying promptly gets the Folium Nerviliae fordii total flavonoids extract.

Folium Nerviliae fordii activity extract preparation method of the present invention can be, at first in the dry medical material of Folium Nerviliae fordii, add 10～15 times of water gagings and soak 24～48h, be evaporated to small size, add ethanol and make precipitation, standing over night, supernatant are evaporated to does not have the alcohol flavor, crosses 732 cation exchange resiies, be washed to colourless, 50～80% ethanol elutions are collected pure washing liquid to there not being ninhydrin reaction then, and being evaporated to does not have the alcohol flavor, cross 717 anion exchange resin, be washed to colourlessly, reuse 50～80% ethanol elutions are collected ethanol elution to there not being ninhydrin reaction, being evaporated to does not have the alcohol flavor, the dry Folium Nerviliae fordii total amino acids extract that gets; 80% alcohol reflux 2～3 times that in the dry medical material of Folium Nerviliae fordii, adds 6～12 times of amounts, each 1～2 hour, merge extractive liquid,, being evaporated to does not have the alcohol flavor, crosses the AB-8 macroporous adsorbent resin, is washed to colourless, 50～80% ethanol elutions are collected eluent to colourless then, and being evaporated to does not have the alcohol flavor, cross polyamide column, be washed to colourlessly, reuse 50～80% ethanol elutions are to colourless, collect eluent, being evaporated to does not have the alcohol flavor, and drying promptly gets the Folium Nerviliae fordii total flavonoids extract; Folium Nerviliae fordii total amino acids extract and Folium Nerviliae fordii total flavonoids extract are mixed.

Folium Nerviliae fordii activity extract preparation method of the present invention can be, in the dry medical material of Folium Nerviliae fordii, add 10～15 times of water gagings and soak 24～48h, be evaporated to small size, add ethanol and make precipitation, standing over night, supernatant are evaporated to does not have the alcohol flavor, crosses 732 type cation exchange resiies, be washed to colourless, 50～80% ethanol elutions are collected pure washing liquid to there not being ninhydrin reaction then, and being evaporated to does not have the alcohol flavor, cross 717 type anion exchange resin, be washed to colourlessly, reuse 50～80% ethanol elutions are collected ethanol elution to there not being ninhydrin reaction, being evaporated to does not have the alcohol flavor, standby; Medicinal residues after water is carried add ethanol (determining alcohol the is 80%) reflux, extract, 2～3 times of 6～12 times of amounts, each 1～2 hour, merge extractive liquid,, being evaporated to does not have the alcohol flavor, crosses macroporous adsorbent resin, be washed to colourlessly, 50～80% ethanol elutions are collected eluent to colourless then, being evaporated to does not have the alcohol flavor, crosses polyamide column, is washed to colourless, reuse 50～80% ethanol elutions are collected eluent to colourless, and being evaporated to does not have the alcohol flavor, mix the dry Folium Nerviliae fordii extract that gets with above-mentioned reserve liquid.

Folium Nerviliae fordii activity extract preparation method of the present invention can be, ethanol (determining alcohol the is 80%) reflux, extract, 2～3 times that in the dry medical material of Folium Nerviliae fordii, adds 6～12 times of amounts, each 1～2 hour, merge extractive liquid,, be evaporated to and do not have the alcohol flavor, cross the AB-8 macroporous adsorbent resin, be washed to colourlessly, 50～80% ethanol elutions are to colourless then, collect eluent, being evaporated to does not have the alcohol flavor, crosses polyamide column, is washed to colourless, reuse 50～80% ethanol elutions are to colourless, collect eluent, being evaporated to does not have the alcohol flavor, standby; Medicinal residues after the alcohol extraction, add 10～15 times of water gagings and soak 24～48h, be evaporated to small size, add ethanol and make precipitation, standing over night, supernatant is evaporated to does not have the alcohol flavor, cross 732 cation exchange resiies, be washed to colourlessly, 50～80% ethanol elutions are collected pure washing liquid to there not being ninhydrin reaction then, be evaporated to and do not have the alcohol flavor, cross 717 anion exchange resin, be washed to colourlessly, reuse 50～80% ethanol elutions are to there not being ninhydrin reaction, collect ethanol elution, being evaporated to does not have the alcohol flavor, mixes with above-mentioned reserve liquid, the dry Folium Nerviliae fordii extract that gets.

Pharmaceutical composition of the present invention contains the Folium Nerviliae fordii activity extract for the treatment of drug effect and acceptable adjuvant pharmaceutically, and this Folium Nerviliae fordii activity extract contains 10-90% total flavones and 10-90% total amino acids.

Total flavones and total amino acids content are mass percent in the Folium Nerviliae fordii activity extract of the present invention.

Multiple of the present invention is meant that solvent volume consumption (ml) is the multiple of quality of medicinal material consumption (g).

Concentration of alcohol of the present invention is meant volumetric concentration (v/v).

Pharmaceutical composition of the present invention can be made the oral Pharmaceutical dosage forms that meets the preparation requirement such as tablet, capsule, granule, oral liquid etc. according to common process.

Pharmaceutical composition of the present invention can require to make injection such as freeze-dried powder, injection etc. according to process for preparation of injection.

The application in the viral respiratory system disease medicine of preparation treatment of Folium Nerviliae fordii activity extract of the present invention and aforementioned pharmaceutical compositions.

Better understand the present invention, with medicine pharmacological testing of the present invention and result its purposes in drug worlds such as treatment respiratory tract disease is described below.

The pharmacologically active of Folium Nerviliae fordii activity extract of the present invention is as described below.

Pharmacodynamic experiment:

1. the antiinflammatory of Folium Nerviliae fordii activity extract and analgesic activity

(1) mice ear experiment

Material and reagent: dimethylbenzene (Tianjin Fu Yu Fine Chemical Co., Ltd, 041117), the 50ul micropipette (Chinese-foreign joint Qiujing Bio-Chemical Reagent Instrument Co., Ltd., Shanghai, Lu system: 02270112), card punch

Folium Nerviliae fordii injection, Folium Nerviliae fordii injecting drug use (total flavones and total amino acids mixed powder pin, FL+AM), (2ml/ props up QINKAILING ZHUSHEYE, Mingxing Pharmaceuticals Co., Ltd., Guangzhou City, 040415), prednisone acetate tablets (5mg/ sheet, Xianju, Zhejiang Pharmacy stock Co., Ltd, 050256), sodium chloride injection (500ml, Hunan Cologne Pharmaceutical Co., Ltd, 041221) laboratory animal: NIH mice, male and female half and half, 18～22g is provided by Traditional Chinese Medicine University Of Guangzhou's Experimental Animal Center.Make up a prescription and dosage: the tail vein injection administration is adopted in experiment, and injected dose is 0.1ml/10g, and wherein prednisone then adopts oral administration, irritates stomach (0.4ml/20g).Each medicine all gets with the preparation of sodium chloride injection dissolved dilution.(total flavones and total amino acids mixed powder pin FL+AM) are crossed microporous filter membrane before the administration for Folium Nerviliae fordii injection, Folium Nerviliae fordii injecting drug use.

Relevant drug level is as follows; Folium Nerviliae fordii injecting drug use (total flavones 0.51mg/ml, total amino acids 2.17mg/ml), Folium Nerviliae fordii injection (containing crude drug amount 0.1mg/ml)

Ear swelling experimental technique: 50 of NIH mices, random packet, every group 10, the normal saline group, the prednisolone acetate group, the QINKAILING ZHUSHEYE group, Folium Nerviliae fordii injection group, the Folium Nerviliae fordii combined group, prednisolone acetate group gastric infusion 0.2ml/10g, all the other respectively organize equal tail vein injection administration 0.1ml/10g, successive administration 3 days behind the last administration 30min, is applied to two sides, mouse right ear front and back with dimethylbenzene, 50ul/ only, behind the 1h dislocation of mice cervical vertebra is put to death, cut two ears, lay round auricle at same position respectively with the 8mm card punch along the auricle baseline, weigh, calculate inhibitory rate of intumesce.The results are shown in Table 1.

Swelling degree=auris dextra sheet weight-left auricle weight swelling rate=(auris dextra sheet weight-left auricle weight)/left auricle weight * 100%

Table 1 mice ear preliminary experiment result

	N	Dosage ml/10g	Swelling degree mg	Swelling rate %
					Normal saline group Folium Nerviliae fordii combined group Folium Nerviliae fordii injection group QINKAILING ZHUSHEYE group prednisolone acetate group	10 10 10 10 10	0.1 0.1 0.1 0.1 0.2	17.2±2.9 11.9±6.1 15.1±4.9 14.9±3.4 14.1±6.7	115.89±19.59 83.34±39.37* △ 110.68±26.77 105.28±27.70 102.79±48.91

Table 1 shows that with QINKAILING ZHUSHEYE and the positive contrast of prednisone acetate tablets, positive controls swelling rate is organized less than feminine gender.Folium Nerviliae fordii combined group and normal saline matched group and QINGKAILING group compare, swelling rate all lower (p＜0.05), and prompting has antiinflammatory action, and antiinflammatory action is better than QINGKAILING.The mice ear rate is lower in the Folium Nerviliae fordii injection group, but statistics meaningless (p＞0.05), prompting has antiinflammatory action trend.

(2) rat granuloma swelling experiment

Material and reagent: Folium Nerviliae fordii injecting drug use (total flavones and total amino acids mixed powder pin), Folium Nerviliae fordii total amino acids injection (AM), Folium Nerviliae fordii total flavonoids injection (FL), QINKAILING ZHUSHEYE (the bright emerging pharmacy in Guangzhou, 2ml/ props up).

Laboratory animal: the SD rat, male, 180～220g is provided by Traditional Chinese Medicine University Of Guangzhou's Experimental Animal Center.

Experimental technique: get 50 of male rats, random packet, is respectively matched group, positive group, Folium Nerviliae fordii injection drug group (FL+AM), Folium Nerviliae fordii total amino acids injection group (AM), Folium Nerviliae fordii total flavonoids injection (FL) group by 10 every group.Urethane is anaesthetized deeply, the sterile working, and it is subcutaneous that the 10mg rayon balls of autoclave sterilization is implanted rat side axillary fossa respectively, and the cotton balls shape should be the same with implant site.Postoperative is respectively organized rat tail vein injection (0.6ml/ only), negative control group gives the normal saline with volume, successive administration 7 days, rat is put to death in dislocation in the 8th day, takes out cotton balls, and it is roasting to constant weight to put 60 ℃ of baking boxs, deduct the former weight of cotton balls, be granuloma weight, granuloma weight is represented with g, respectively organizes granuloma weight.Experimental result sees Table 2.

Table 2 Folium Nerviliae fordii is to the bullate influence of rat granuloma (x ± s)

Group	Number of animals (only)	Concentration	Dosage	Granuloma weight (mg)
					Normal saline FL+AM FL AM QINKAILING ZHUSHEYE	10 10 10 10 10	-FL 0.48mg/ml, AM 2.09mg/ml 0.51mg/ml 2.13mg/ml stock solution	-0.6ml/ 0.6ml/ 0.6ml/ 0.6ml/ only	36.14±2.81 20.93±1.49**## 31.52±2.72**# 28.43±3.27** 25.67±2.21**

Table 2 is the result show, compare with the blank group: QINGKAILING positive controls, Folium Nerviliae fordii injection drug group (FL+AM), Folium Nerviliae fordii total amino acids injection group (AM), Folium Nerviliae fordii total flavonoids injection (FL) group, all can significantly suppress the growth (P all＜0.01) of cotton balls granulation tissue, point out each experimental group medicine that equal cotton balls granulation tissue is had good inhibitory effect.Compare with the QINGKAILING positive controls: Folium Nerviliae fordii injecting drug use group has significant difference (P＜0.01), and the antiinflammatory action of prompting Folium Nerviliae fordii injecting drug use group is stronger than QINGKAILING positive controls; Folium Nerviliae fordii total flavonoids injection group (FL) has significant difference (P＜0.05), the antiinflammatory action of prompting Folium Nerviliae fordii total flavonoids injection group than QINGKAILING positive controls a little less than; Folium Nerviliae fordii total amino acids injection group (AM) does not have significantly, and the antiinflammatory action of prompting Folium Nerviliae fordii total amino acids injection is suitable with the QINGKAILING positive controls; Experimental result shows that Folium Nerviliae fordii total flavonoids and total amino acids are mixed with the antiphlogistic synergism of enhancing.

(3) rat paw edema experiment

Material and reagent: Folium Nerviliae fordii injecting drug use (total flavones and total amino acids mixed powder pin, with the sodium chloride injection dissolving, concentration is for containing total flavones 0.48mg/ml, containing total amino acids 2.09mg/ml), QINKAILING ZHUSHEYE.

Laboratory animal: the SD rat, male, 180～220g is provided by Traditional Chinese Medicine University Of Guangzhou's Experimental Animal Center.

Experimental technique:

Get 30 of male rats, random packet, is respectively normal saline matched group, positive group, Folium Nerviliae fordii injecting drug use group by 10 every group.Tail vein injection administration (dosage is with the swollen experiment of rat granuloma), the blank group gives the isometric(al) normal saline, every day 1 time, successive administration 7 days.Respectively organize the right back sufficient normal volume of rat (making a clear graticule just down) with the measurement of improvement volumetric method at foot, after the last administration at the subcutaneous injection of right back sufficient sole of the foot 0.1ml carrageenin (concentration is 0.2%), mensuration causes scorching back 1,2,3,4, the right back sufficient volume of 5h, calculates swelling rate and inhibitory rate of intumesce.The results are shown in Table 3,4.

Table 3 Folium Nerviliae fordii is to the influence of rat paw edema rate (x ± s)

Group	N (only)	Foot swelling rate (%)
							1h	2h	3h	4h	5h
		Normal saline Folium Nerviliae fordii QINGKAILING	10 10 10	26.29±3.19 12.14±2.12 **## 14.68±1.24 **	32.23±3.78 13.43±2.18 **## 24.89±2.99 **	30.02±3.63 5.98±3.51 **## 21.60±2.63 **						27.00±3.94 2.95±5.01 **## 14.60±2.45 **	20.34±3.84 0.33±1.00 **## 9.53±1.82 **

Table 4 Folium Nerviliae fordii is to the influence of rat paw edema suppression ratio

Group	N (only)	Foot swelling suppression ratio %
							1h	2h	3h	4h	5h
		Normal saline Folium Nerviliae fordii QINGKAILING	10 10 10	53.82 43.46	58.34 22.95	80.07 28.02						89.06 45.94	98.36 53.15

Table 3 is the result show, with the blank group relatively: Folium Nerviliae fordii injecting drug use group and QINKAILING ZHUSHEYE group 1,2,3,4,5h all can significantly suppress carrageenin and cause rat paw edema (P value equal＜0.01), points out each experimental group medicine that tangible antiinflammatory action is all arranged.Compare with the QINKAILING ZHUSHEYE positive controls: Folium Nerviliae fordii injecting drug use group has significant difference (P＜0.01), and prompting Folium Nerviliae fordii injecting drug use group is better than QINKAILING ZHUSHEYE to acutely inflamed inhibitory action.

(4) influence that the mouse peritoneal capillary permeability is increased

Material and reagent: Folium Nerviliae fordii injecting drug use (concentration is 0.48mg/ml+2.09mg/ml for total flavones+total amino acids mixed powder pin, time spent sodium chloride injection dissolving), QINKAILING ZHUSHEYE.

Laboratory animal: the NIH mice, male, 18～22g is provided by Traditional Chinese Medicine University Of Guangzhou's Experimental Animal Center.

Experimental technique:

Get the NIH mice, body weight 1g～22g, male, be divided into three groups at random, 10 every group, be respectively matched group, positive group, Folium Nerviliae fordii injecting drug use group.The tail vein injection administration, the blank group gives the isometric(al) normal saline, every day 1 time, successive administration 2 times, 0.5h after the last administration, each caudal vein are injected 2% azovan blue normal saline solution 0.1ml/10g body weight, lumbar injection 0.8% acetic acid normal saline solution 0.2ml/ only immediately, the sacrificed by exsanguination mice is cut the abdominal cavity open behind the 20min, with the 10ml normal saline flushing for several times, collect washing liquid, centrifugal, get supernatant and survey OD value, relatively each group difference at spectrophotometer 590nm place.The results are shown in Table 5.

The influence that table 5 Folium Nerviliae fordii Dichlorodiphenyl Acetate induced mice abdominal cavity capillary permeability increases (x ± s)

Group	Number of animals (only)	Dosage (ml/10g)	The OD value
				Normal saline group Folium Nerviliae fordii medicine group QINGKAILING group	10 10 10	0.1 0.1 0.1	0.390±0.122 0.143±0.122 **△ 0.152±0.104 **

Table 5 result shows, Folium Nerviliae fordii is subjected to reagent group, QINKAILING ZHUSHEYE group and normal saline group relatively, all can suppress acetic acid induced mice capillary permeability and increase (p＜0.01), Folium Nerviliae fordii is subjected to reagent group and QINKAILING ZHUSHEYE group relatively, suppress effect that acetic acid induced mice capillary permeability increases strong (p＜0.05), showing that the Folium Nerviliae fordii activity extract has suppresses the effect that acetic acid induced mice capillary permeability increases preferably.

(5) influence of Dichlorodiphenyl Acetate induced mice pain (writhing method)

Material and reagent: the Folium Nerviliae fordii activity extract (rare alcohol extract, 2mg/ml), aspirin, normal saline.

Laboratory animal: the NIH mice, male and female half and half, 18～22g is provided by Traditional Chinese Medicine University Of Guangzhou's Experimental Animal Center.

Experimental technique:

Get the NIH mice, body weight 18～22g, male and female half and half are divided into three groups at random, 10 every group, are respectively normal saline matched group, positive group, Folium Nerviliae fordii activity extract group.During experiment, the fasting 12h of mice elder generation, gastric infusion then, every day 1 time, continuous 7 days, after the last administration 1h lumbar injection 0.8% acetic acid normal saline solution 0.2ml/ only, in the observed and recorded 20 minutes mice turn round the body number of times, respectively organize difference.The results are shown in Table 6.

The influence of table 6 Folium Nerviliae fordii activity extract Dichlorodiphenyl Acetate induced mice pain (x ± s)

Group	Number of animals (only)	Dosage	Turn round body number of times (inferior)
				Normal saline aspirin Folium Nerviliae fordii group	10 10 10	- 0.2g/kg 40mg/kg	34.30±11.95 4.50±3.54 ** 16.10±6.52 **

Table 6 result shows, compares with the blank group of normal saline, and the Folium Nerviliae fordii activity extract can significantly reduce the acetic acid induced mice and turn round body number of times (P＜0.01); What aspirin also can significantly reduce mice turns round body number of times (P＜0.01), and prompting Folium Nerviliae fordii activity extract has analgesic activity.

(6) influence of PARA FORMALDEHYDE PRILLS(91,95) foot plantar subcutaneous injection induced mice pain

Material and reagent: the Folium Nerviliae fordii activity extract (self-control, 2mg/ml), aspirin, normal saline.

Laboratory animal: the NIH mice, male and female half and half, 18～22g is provided by Traditional Chinese Medicine University Of Guangzhou's Experimental Animal Center.

Experimental technique:

Get the NIH mice, body weight 18～22g, male and female half and half are divided into three groups at random, 10 every group, be respectively normal saline matched group, positive group, Folium Nerviliae fordii activity extract group, the fasting 12h of mice elder generation, gastric infusion then, and behind medicine the left back sufficient plantar subcutaneous injection 2.5% formaldehyde normal saline solution of each Mus of 1h, observe the accumulated time that mice licks metapedes in the 5min immediately, relatively each group difference.The results are shown in Table 7.

The influence of table 7 Folium Nerviliae fordii activity extract PARA FORMALDEHYDE PRILLS(91,95) induced mice pain (x ± s)

Group	Number of animals (only)	Dosage (g/kg)	Lick sufficient accumulated time (s)
				Normal saline aspirin Folium Nerviliae fordii activity extract	10 10 10	- 0.2 1.05	49.61±17.10 17.96±10.01 ** 31.67±12.11 *

Table 7 results suggest, with the normal saline matched group relatively, what the Folium Nerviliae fordii activity extract can significantly reduce the pain caused mice of formaldehyde foot plantar subcutaneous injection licks sufficient accumulated time (P＜0.05); What aspirin also can significantly reduce the pain caused mice of formaldehyde foot plantar subcutaneous injection licks sufficient accumulated time (P＜0.01).Prompting Folium Nerviliae fordii activity extract can suppress formaldehyde induced mice pain.

2. the enhance immunity effect of Folium Nerviliae fordii activity extract

(1) the mice carbon granule is cleaned up experiment

Material and reagent: Folium Nerviliae fordii injecting drug use (total flavones and total amino acids mixed powder pin), SHENMAI ZHUSHEYE.0.25% prednisolone acetate solution, 0.1% sodium carbonate (Na ₂CO ₃) solution, one gets pavilion prepared Chinese ink.

Laboratory animal: the NIH mice, male and female half and half, 18～22g is provided by Traditional Chinese Medicine University Of Guangzhou's Experimental Animal Center.

Experimental technique:

Get the NIH mice, body weight 18～22g is divided into 4 groups at random, normal saline group, Folium Nerviliae fordii injecting drug use group (containing total flavones 0.51mg/ml, total amino acids 2.17mg/ml), injection containing ginseng extract group, prednisolone acetate group.Except that prednisolone acetate group gastric infusion 0.2ml/10g body weight, each organizes equal tail vein injection administration 0.1ml/10g body weight.Administration every day 1 time, successive administration 5 days, after the last administration 30 minutes, tail vein injection prepared Chinese ink (0.1ml/ only), get blood 50ul from the eye socket venous plexus in injection back 2 minutes and 12 minutes respectively with capillary tube (using liver rope solution wetted in advance), in 0.1% sodium carbonate liquor of solution 5ml, shake up, place the 600nm colorimetric, measure optical density (OD).Mice takes off cervical vertebra puts to death, and takes by weighing liver, spleen weight respectively, and index K is cleaned up in calculating or index α is cleaned up in correction.OD ₁

$K=\frac{\log {\mathrm{OD}}_1-\log {\mathrm{OD}}_2}{t_2-t_1}$

OD ₁, OD ₂Be the optical density of different time institute blood sampling, t ₂-t ₁For getting the time difference of two blood samples.The results are shown in following table:

The influence that table 8 medicine is cleaned up the mice carbon granule

Group	Number of animals	Phagocytic index (K)	Activate the phagocytic capacity (α)
				Normal saline Folium Nerviliae fordii medication (FL+AM) SHENMAI ZHUSHEYE prednisolone acetate	10 10 10 10	0.007±0.001 0.015±0.002**# 0.017±0.003**# 0.02±0.001**	23.54±3.69 21.86±2.77 23.56±3.04 27.78±4.50

Table 8 is the result show, compare with the blank group: Folium Nerviliae fordii injecting drug use group, SHENMAI ZHUSHEYE group, prednisolone acetate group, all can significantly improve the phagocytic function (the P value all＜0.01) of mice, the effect of pointing out each experimental group medicine all to be significantly improved immunologic function.Compare with the prednisolone acetate group: Folium Nerviliae fordii injecting drug use group, SHENMAI ZHUSHEYE group all have significant difference (the P value all＜0.05), and prompting Folium Nerviliae fordii injecting drug use and SHENMAI ZHUSHEYE all have the effect that improves immunologic function.Compare with the injection containing ginseng extract group: Folium Nerviliae fordii injecting drug use group zero difference (P＞0.05), action intensity and SHENMAI ZHUSHEYE that prompting Folium Nerviliae fordii activity extract injecting drug use improves immunity are suitable.

(2) humoral immunization experiment

Material: Folium Nerviliae fordii activity extract (60% ethanol extraction), SHENMAI ZHUSHEYE, 0.25% prednisolone acetate solution.5% normal saline chicken red blood cell suspension.

Laboratory animal: the NIH mice, male and female half and half, 18～22g is provided by Traditional Chinese Medicine University Of Guangzhou's Experimental Animal Center.

Experimental technique:

Get the NIH mice, body weight 18～22g is divided into 4 groups at random, normal saline group, Folium Nerviliae fordii activity extract group, SHENMAI ZHUSHEYE group, prednisolone acetate group.Each is organized mouse peritoneal and injects 5% normal saline chicken red blood cell suspension 0.2ml and carry out immunity, each group gastric infusion 0.2ml/10g body weight (the SHENMAI ZHUSHEYE group is tail vein injection administration 0.1ml/10g body weight then) respectively then, administration every day 1 time, successive administration 7 days.After the last administration 1 hour, each is organized the mouse orbit vein and gets blood, and was centrifugal, get serum with 100 times of normal saline dilutions, get dilute serum 1ml, mix with % normal saline chicken red blood cell suspension 0.5ml, 10% complement 0.5ml, after in 37 ℃ of calorstats, being incubated 30 minutes, stopped reaction in 0 ℃ of refrigerator.Centrifugal then, get supernatant in 540nm place colorimetric, measure optical density (OD), other establishes the not blank of increase serum, returns to zero when getting its supernatant as colorimetric.With the index of OD reading as the judgement serum hemolysin, relatively difference of each group.The results are shown in Table 9.

Table 9 Folium Nerviliae fordii generates the influence of level to the mice serum hemolysin

Group	Dosage	Medication	Serum hemolysin (OD)
				Normal saline Folium Nerviliae fordii activity extract SHENMAI ZHUSHEYE prednisolone acetate	0.2ml/10g 0.2mg/10g 0.1ml/10g 0.2ml/10g	Irritate stomach and irritate stomach tail vein injection filling stomach	0.024±0.017 0.194±0.035 ** 0.186±0.023 ** 0.180±0.015 **

Table 9 is the result show, compare with the blank group: Folium Nerviliae fordii activity extract group, SHENMAI ZHUSHEYE group, prednisolone acetate group, all can significantly increase the erythrocyte antibody (EA) (the P value all＜0.01) of mice, the effect of pointing out each experimental group medicine all to be significantly improved immunologic function.With prednisolone acetate group relatively: Folium Nerviliae fordii activity extract group, SHENMAI ZHUSHEYE group there are no significant difference (P value is all＞0.05), point out Folium Nerviliae fordii injecting drug use and SHENMAI ZHUSHEYE all to have higher immunologic function.Compare with the injection containing ginseng extract group: Folium Nerviliae fordii activity extract group zero difference (P＞0.05), action intensity and SHENMAI ZHUSHEYE that prompting Folium Nerviliae fordii activity extract improves immunity are suitable.

(3) to mouse immune organ weight's influence

Material: Folium Nerviliae fordii activity extract (water extract), SHENMAI ZHUSHEYE, 0.25% prednisolone acetate solution.

Laboratory animal: the NIH mice, male and female half and half, 14～18g is provided by Traditional Chinese Medicine University Of Guangzhou's Experimental Animal Center.

Experimental technique: get the NIH mice, body weight 14～18g is divided into 4 groups at random, normal saline group, Folium Nerviliae fordii activity extract group, SHENMAI ZHUSHEYE group, prednisolone acetate group.Each group is gastric infusion 0.2ml/10g body weight (wheat injection group is tail vein injection administration 0.1ml/10g body weight then) respectively, administration every day 1 time, successive administration 7 days.After the last administration 2 hours, each group mice is all taken off cervical vertebra put to death, cut open and get thymus, spleen, its hetero-organization is peeled off totally, behind the filter paper suck dry moisture, weigh.Calculate thymic weight coefficient and spleen weight coefficient (thymic weight coefficient=thymus weight/body weight * 100, spleen weight coefficient=spleen weight/body weight * 100), the results are shown in Table 10.

Table 10 Folium Nerviliae fordii activity extract is to the influence of mouse immune organ thymus, spleen weight

Group	Dosage	Medication	The thymic weight coefficient	The spleen weight coefficient
					Normal saline Folium Nerviliae fordii activity extract SHENMAI ZHUSHEYE prednisolone acetate	0.2ml/10g 0.4mg/10g 0.1ml/10g 0.2ml/10g	Irritate stomach and irritate stomach tail vein injection filling stomach	0.324±0.118 0.590±0.054**△ 0.601±0.087**△ 0.191±0.032**	0.452±0.055 0.638±0.168**△ 0.589±0.075**△ 0.263±0.068**

Table 10 is the result show, compare with the normal saline matched group: Folium Nerviliae fordii activity extract group, the SHENMAI ZHUSHEYE group all can significantly improve mouse thymus coefficient, the heavy coefficient of spleen (the P value all＜0.01), the effect of pointing out each experimental group medicine all to be significantly improved immunologic function.Compare with the prednisolone acetate group: Folium Nerviliae fordii activity extract group, SHENMAI ZHUSHEYE group all have significant difference (the P value all＜0.05), and it is very strong that prompting Folium Nerviliae fordii activity extract and SHENMAI ZHUSHEYE improve the effect of immunologic function.The SHENMAI ZHUSHEYE group compares: Folium Nerviliae fordii activity extract zero difference (P＞0.05), action intensity and SHENMAI ZHUSHEYE that prompting Folium Nerviliae fordii activity extract improves immunity are suitable.

The refrigeration function of 3 Folium Nerviliae fordii activity extracts

Material: Folium Nerviliae fordii activity extract (rare alcohol extract), SHENMAI ZHUSHEYE, aspirin, yeast suspension.

Laboratory animal: the SD rat, male and female half and half, 220～250g is provided by Traditional Chinese Medicine University Of Guangzhou's Experimental Animal Center.

Experimental technique:

Get the SD rat, body weight 220～250g, 37 ± 1 ℃ of anus temperature are divided into 4 groups at random, (irritate stomach, 0.5ml/100g), Folium Nerviliae fordii activity extract group (is irritated stomach, 5mg/100g) to the normal saline group, (tail vein injection, 0.5ml/100g), the aspirin group (is irritated stomach, 0.1g/kg) to the SHENMAI ZHUSHEYE group.Except that the normal saline group, each organizes 15% beer yeast suspension (10ml/kg) of the new preparation of the equal subcutaneous injection of rat, injects administration in back 2 hours, surveys 1 anus temperature every 1 hour, surveys continuously 10 hours, injects timing behind the yeast certainly, calculates and respectively organizes average anus temperature.The results are shown in Table 11.

Table 11 Folium Nerviliae fordii activity extract is to the cooling effect of rat beer yeast pyrogenicity

Group	Basal body temperature (℃)	Body temperature behind the medicine (℃)
								3h	4h	5h	6h	8h	10h
		Normal saline Folium Nerviliae fordii activity extract SHENMAI ZHUSHEYE aspirin	37.4±0.2 36.9±0.3 37.2±0.2 37.1±0.4	38.5±0.2 37.8±0.2 38.0±0.3 * 37.7±0.2 *	38.8±0.3 38.2±0.2 * 38.3±0.2 * 38.2±0.3 *	39.2±0.3 38.4±0.3 * 38.5±0.3 * 38.5±0.4 *	39.8±0.2 38.5±0.2 * 38.8±0.3 * 38.4±0.3 *							39.5±0.2 38.0±0.4 * 38.1±0.2 * 37.9±0.3 *	38.1±0.2 37.1±0.3 * 37.7±0.3 * 37.0±0.2 *

Table 11 is the result show, compare with the normal saline matched group: Folium Nerviliae fordii activity extract group, SHENMAI ZHUSHEYE group, the aspirin group body temperature (the P value is all＜0.05) after each time point all can significantly reduce rat yeast pyrogenicity, point out each experimental group medicine that the better antipyretic effect is all arranged; Compare with the aspirin group: Folium Nerviliae fordii activity extract group, SHENMAI ZHUSHEYE group all do not have big difference (the P value all＞0.05), and the refrigeration function intensity of prompting Folium Nerviliae fordii activity extract and SHENMAI ZHUSHEYE approaches aspirin; Compare with the SHENMAI ZHUSHEYE group: Folium Nerviliae fordii activity extract group does not have statistical discrepancy, and the action intensity and the SHENMAI ZHUSHEYE of the rat temperature of prompting Folium Nerviliae fordii activity extract reduction yeast pyrogenicity are suitable.

The eliminating phlegm and relieving cough effect of 4 Folium Nerviliae fordii activity extracts

(1) mice ammonia is drawn the antitussive effect of coughing

Material: the Folium Nerviliae fordii activity extract (rare alcohol extract, 2mg/ml), 0.2% codeine phosphate, strong aqua ammonia, normal saline.

Laboratory animal: the NIH mice, male and female half and half, 18～22g is provided by Traditional Chinese Medicine University Of Guangzhou's Experimental Animal Center.

Experimental technique:

Get the NIH mice, body weight 18～22g is divided into 3 groups at random, normal saline group, the positive group of codeine, Folium Nerviliae fordii group.Positive group and Folium Nerviliae fordii group hour filling stomach 0.2ml/10g, the normal saline group then gives the equivalent normal saline.Be administered once every day, continuous 5 days.After the last administration 30 minutes, mice is put into glass bell jar, open the ultrasound atomizer that is attached thereto by rubber tube, spray into strong aqua ammonia in the glass bell jar equably, with the 53kPa constant voltage with ammonia even spraying 15 seconds, take out mice immediately, the cough latent period of observed and recorded mice and the number of times of coughing in 3 minutes (shrinking and magnify mouth with the mice abdominal muscle is cough action indication).The results are shown in Table 12.

Table 12 Folium Nerviliae fordii activity extract draws the antitussive action of coughing mice to ammonia

Group	Dosage	Medication	Cough latent period (s)	The cough number of times
					Normal saline Folium Nerviliae fordii activity extract 0.2% codeine phosphate	0.2ml/10g 0.4mg/10g 0.2ml/10g	Irritate stomach and irritate stomach filling stomach	35.0±7.1 59.5±6.7 **△△ 80.8±35.3 **	20.5±4.4 11.6±2.6 **△△ 8.2±2.2 **

Table 12 is the result show, with the blank group relatively: but the equal significant prolongation ammonia induced mice cough latent period (P＜0.01) of Folium Nerviliae fordii activity extract, codeine phosphate and can significantly suppress ammonia induced mice cough (P＜0.01).With the codeine phosphate positive controls relatively: the Folium Nerviliae fordii activity extract has remarkable statistical discrepancy (P＜0.01), and the prolongation mouse cough incubation period of prompting Folium Nerviliae fordii activity extract, the effect that suppresses mouse cough are stronger.

(2) to the phlegm-dispelling functions (the phenol red method of mice trachea) of mice

Material: the Folium Nerviliae fordii activity extract (rare alcohol extract, 2mg/ml), 0.5% phenol red solution, 5% sodium bicarbonate solution, Mucosolvan Ambroxol sheet (Shanghai Boehringer Ingelheim pharmaceutcal corporation, Ltd), normal saline.

Laboratory animal: the NIH mice, male and female half and half, 18～22g is provided by Traditional Chinese Medicine University Of Guangzhou's Experimental Animal Center.

Experimental technique:

Get the NIH mice, body weight 18～22g is divided into 3 groups at random, normal saline group, Mucosolvan Ambroxol sheet group, Folium Nerviliae fordii group.Each is organized the mice fasting and can't help water 12 hours.Folium Nerviliae fordii group gastric infusion (0.3ml/10g), Mucosolvan Ambroxol sheet group gives 30mg/kg, and the normal saline group then gives the normal saline of equivalent.Be administered once every day, continuous 4 days.After the last administration 30 minutes, lumbar injection 0.5% phenol red solution 0.5ml/ only.After spending 30 minutes, mice is put to death in the cervical vertebra dislocation, faces upward the position and is fixed on the operation plate, cut off throat skin, separate trachea, peel off the trachea surrounding tissue, cut one section trachea to the trachea bifurcation from thyroid cartilage, put in the test tube that fills the 3ml normal saline and wash, add 5% sodium bicarbonate solution 0.1ml again, the centrifugal 5min of 2000r/min gets supernatant, spectrophotometer 546nm place measures the OD value, calculates phenol red excretion according to phenol red standard curve.The results are shown in Table 13.

Table 13 Folium Nerviliae fordii activity extract is to the influence of the phenol red excretion amount of mice trachea

Group	Dosage	Medication	Phenol red excretion (mg/ml)
				Normal saline Folium Nerviliae fordii activity extract Mucosolvan Ambroxol	0.3ml/10g 0.3ml/10g 30mg/kg	Irritate stomach and irritate stomach filling stomach	1.12±0.23 2.90±0.29 **△△ 3.94±0.24 **

Table 13 is the result show, compare with the blank group: Folium Nerviliae fordii activity extract, Mucosolvan Ambroxol sheet all can significantly improve the phenol red excretion of mice trachea (P＜0.01); Compare with Mucosolvan Ambroxol: the phenol red excretion of Folium Nerviliae fordii activity extract group mice trachea has significant difference (P＜0.01); Prompting Folium Nerviliae fordii activity extract has the function that improves the phenol red excretion of mice trachea preferably, and significantly phlegm-dispelling functions is arranged.

5. the antibiosis and antiviral functions of Folium Nerviliae fordii activity extract

(1) extracorporeal antivirus effect effect (the outer method of Embryo Gallus domesticus)

Material: Folium Nerviliae fordii activity extract (rare alcohol extract, self-control), Embryo Gallus domesticus (the prosperous Embryo Gallus domesticus of planting of 10 age in days Lays).Influenza virus A type (PR8, a 57-4, strains such as the 68-1 of capital section, Shanghai 74-5), influenza B (Lee strain), influenza third type (1233 strain), parainfluenza virus I type (Sand strain) are from national preventing and controlling influenza research center.

Experimental technique:

Medicinal liquid was begun to do doubling dilution with the PH7.0 buffer saline from 1: 10, each dilution factor injects 3-5 chick embryo allantoic cavity respectively, and 0.3ml/ only establishes the normal control Embryo Gallus domesticus simultaneously.Observe its growing state every day, when the pastille Embryo Gallus domesticus do not occur dead least concentration then for this medicine to Embryo Gallus domesticus avirulence boundary, adopt this concentration when carrying out formal test.

With dilution medicinal liquid of difference and viral liquid (1 HAU) equivalent mixing, matched group replaces medicinal liquid with buffer saline.Put above-mentioned each pipe in 24 ℃ of water-bath 1h, each is managed test solution and injects 3-5 chick embryo allantoic cavity respectively, and 0.3ml/ is only hatched 48h for 35 ℃.Under matched group Embryo Gallus domesticus survival condition, collect the chick embryo allantoic liquid of each survival, carry out the Sanguis Gallus domesticus hemagglutination test (HA test), can anti-anti-viral duplicate and the lowest concentration of drug of Sanguis Gallus domesticus red cell agglutination do not occur, be the minimum antiviral concentration of this medicine.

The result shows that the Folium Nerviliae fordii activity extract has the effect of significantly anti-various influenza virus and I type parainfluenza virus outside Embryo Gallus domesticus, and its minimum antiviral concentration sees Table 14.

The minimum antiviral concentration of table 14 Folium Nerviliae fordii activity extract resisiting influenza virus and I type parainfluenza virus

Virus Type	Influenza A (PR 8)	Influenza A (opening 57-4)	Influenza A (68-1 of capital section)	Influenza A (Shanghai 74-5)	Influenza B (Lee)	Influenza C (1233)	I type parainfluenza virus
								Cmin	1∶2560	1∶1536	1∶1024	1∶1024	1∶800	1∶500	1∶1024

Table 14 results suggest, the Folium Nerviliae fordii activity extract all has in various degree inhibitory action to try virus, and is wherein stronger to influenza A virus, I type parainfluenza virus inhibitory action.

(2) vitro antibacterial activity

Material: Folium Nerviliae fordii activity extract, nutrient broth (institute of microbiology of Guangdong Academy of Sciences product), husky Bao Shi culture medium (this chamber prepares routinely).

Staphylococcus aureus, beta hemolytic streptococcus, Bing Xinglianqiujun, viridans streptococci, shigella flexneri, streptococcus pneumoniae, micrococcus catarrhalis, hemophilus influenza, Bacillus typhi, paratyphoid bacillus A, bacillus pyocyaneus, pneumobacillus, escherichia coli and proteus vulgaris.

Experimental technique:

Get 11 of sterile test tube and be arranged in order, every pipe adds meat extract soup 1ml, adds 1ml experiment medicinal liquid in the 1st pipe, and mixing is drawn 1ml and added the 2nd pipe, mixing.Draw 1ml and add the 3rd pipe, so operation is added to the 10th pipe successively, and mixing discards 1ml, and the 11st pipe is made as antibacterial control tube (not adding medicinal liquid).In above-mentioned test tube, add 1: 2000 for examination bacterium liquid (8 hours cultures) 0.05ml, cultivated 24 hours for 37 ℃, get each 1 ring of experimental liquid in each test tube successively, transferred species is on the Sanguis Leporis seu oryctolagi agar plate, cultivate after 24 hours for 37 ℃, observed result is judged minimal inhibitory concentration (MIC).The results are shown in Table 15.

Table 15 Folium Nerviliae fordii minimal inhibitory concentration

The bacterium name	MIC	The bacterium name	MIC
				Staphylococcus aureus beta hemolytic streptococcus γ-streptococcus viridans streptococci, shigella flexneri micrococcus catarrhalis pneumococcus	1∶800 1∶128 1∶96 1∶32 1∶16 1∶1024 1∶512	Bacillus influenzae typhoid bacillus paratyphoid bacillus A pyocyanic pneumonia bacillus Escherichia coli proteus vulgaris	1∶94 1∶32 1∶64 1∶32 1∶256 1∶64 1∶32

Table 15 result shows, the Folium Nerviliae fordii activity extract all has in various degree antibacterial action to the examination strain, and wherein the bacteriostasis to staphylococcus aureus, beta hemolytic streptococcus, micrococcus catarrhalis, streptococcus pneumoniae and pneumobacillus etc. is stronger.

6. the resisting acute lung injury effect of Folium Nerviliae fordii activity extract

Material: Folium Nerviliae fordii injecting drug use (total flavones and total amino acids mixed powder pin), SHENMAI ZHUSHEYE, endotoxin (LPS, Sigma), normal saline.

Laboratory animal: the SD rat, male, 200 ± 20 grams, are provided by Traditional Chinese Medicine University Of Guangzhou's Experimental Animal Center by 40.

Experimental technique:

Get the SD rat and divide 4 groups at random, tail intravenously administrable before the operation test, Folium Nerviliae fordii injecting drug use group is by (2mg total flavones+8mg total amino acids)/kg administration, and the SHENMAI ZHUSHEYE dosage is 10mg/kg; Normal saline group and model group be the about 1ml of tail vein injection saline (4ml/kg) then.Each organizes rat operation test administration the previous day 1 time, and normal saline group and model group are then pressed 4ml/kg injection, fasting 12 hours.During test, animal with the pentobarbital sodium intraperitoneal injection of anesthesia, is distinguished circulation of qi promoting pipe, right side external jugular vein, left carotid intubate according to the 45mg/kg body weight, and each group is handled as follows afterwards:

Model group: from the right side external jugular vein, inject LPS (O111B4) 15mg/kg, carry out modeling.The normal saline of injecting afterwards (4ml/kg).The normal saline group is with the physiologic saline for substitute endotoxin of equivalent, in identical time point, administration in the same manner.The treatment group: in the right side external jugular vein, that gives 1/3 dosage in advance is subjected to the reagent injection, injects 15mg/kg LPS subsequently, and the reagent injection that is subjected to of 2/3 amount of surplusing is injected after modeling.Matched group: the right side external jugular vein, the SHENMAI ZHUSHEYE of giving 1/3 therapeutic dose is in advance injected 15mg/kg LPS subsequently, and 2/3 SHENMAI ZHUSHEYE of measuring of surplusing is injected after modeling, and the injection containing ginseng extract liquid measure is 10mg/kg.

Each treated animal is all in injection endotoxin while vena femoralis injection Evans indigo plant (50mg/kg), animal is opened breast take out lungs, cut off surrounding tissue, lungs are immersed in (the Methanamide consumption is pressed the heavy 20mg/100g of animal body) in the formamide solution, put incubation 72h in the 45-50 ℃ of incubator, pigment all leaches in waiting to organize, take out tissue, centrifugal, get supernatant, the light photometer carries out colorimetric at 620nm, estimates the variation of pulmonary vascular permeability according to the OD value; Another batch animal opens breast taking-up lungs, and after the title weight in wet base, (80 ℃, 20h) roasting to constant weight, the title dry weight is calculated dried/weight in wet base ratio and be the results are shown in Table 16 to put baking oven.

Table 16 Folium Nerviliae fordii injecting drug use is done/influence of weight in wet base ratio induced lung vascular permeability and lung

Group	N (only)	Dosage	The OD value	Do/wet ratio
					Normal saline group model group Folium Nerviliae fordii injecting drug use group SHENMAI ZHUSHEYE group	10 10 10 10	--total flavones 2mg/kg total amino acids 8mg/kg 10mg/kg	0.054±0.021 0.497±0.073 ** 0.285±0.061 **△△ 0.293±0.074 **△△	0.242±0.067 0.138±0.053 ** 0.197±0.068△ 0.188±0.052△

Table 16 result shows, with normal saline blank group relatively: model group, Folium Nerviliae fordii injecting drug use group, SHENMAI ZHUSHEYE group, all there were significant differences for OD value (P＜0.01), the model group induced lung does/weight in wet base is than variant (P＜0.01), the prompting modeling is successful; Compare with model group: Folium Nerviliae fordii injecting drug use group, SHENMAI ZHUSHEYE group OD value all can significantly reduce (P＜0.01), and prompting can reduce the induced lung vascular permeability; Induced lung does/and weight in wet base is than variant (P＜0.05), and prompting Folium Nerviliae fordii injecting drug use, SHENMAI ZHUSHEYE can increase the SD induced lung and do/the weight in wet base ratio.Experimental result prompting Folium Nerviliae fordii injecting drug use and SHENMAI ZHUSHEYE have the effect of SD induced lung damage due to the protection endotoxin.With the SHENMAI ZHUSHEYE group relatively: Folium Nerviliae fordii injecting drug use group does not have remarkable statistical discrepancy (P＞0.05), and the effect and the SHENMAI ZHUSHEYE of SD induced lung damage is suitable due to the prompting Folium Nerviliae fordii activity extract injecting drug use protection endotoxin.

Folium Nerviliae fordii activity extract of the present invention has: antibiosis and antiviral functions, suppress various pathogens, virus; Acute lung injury has protective effect to rat endotoxin type; The effect that improves animal immunizing power is arranged; Have refrigeration function, can reduce the body temperature (endotoxin or yeast cause heating) of rat or rabbit; Effect with anti-inflammatory and antalgic; Effect with eliminating phlegm and relieving cough.

Folium Nerviliae fordii activity extract preparation method abundant raw material of the present invention, the processing technology simple and convenient.

Description of drawings

Fig. 1 Folium Nerviliae fordii total flavonoids extract UV scanning collection of illustrative plates.

The aminoacid HLPC finger printing of Fig. 2 total amino acids extract.

The specific embodiment

Related method is the technological means that those skilled in the art can grasp and use, but following exemplifying embodiment must not be interpreted as the restriction to claim of the present invention of going up in all senses.

Embodiment 1

The dry medical material 1kg of Folium Nerviliae fordii, add 80% alcohol reflux 2 times (10 times of amounts, 8 times of amounts for the second time for the first time), each 1 hour, merge extractive liquid,, being evaporated to does not have the alcohol flavor, adds the dilution of equimultiple water, crosses AB-8 type macroporous adsorbent resin, be washed to colourless, 70% ethanol elution is collected eluent to colourless, and being evaporated to does not have the alcohol flavor, add the dilution of equimultiple water, cross polyamide column (30～60 order), be washed to colourlessly, 70% ethanol elution is to colourless, collect eluent, being evaporated to does not have the alcohol flavor, and lyophilization promptly gets the Folium Nerviliae fordii total flavonoids extract.Folium Nerviliae fordii total flavonoids extract UV scanning collection of illustrative plates as shown in Figure 1.

As seen from Figure 1, be the characteristic absorption peak that there is flavone compound at 270nm and 360nm place at wavelength respectively.

The dry medical material of Folium Nerviliae fordii, be ground into coarse powder, get 1kg and add 10 times of water gagings immersion 24h, be evaporated to every ml and contain the about 1g of crude drug, adding ethanol is 80% to containing the alcohol amount, standing over night, supernatant is evaporated to does not have the alcohol flavor, crosses 732 strongly acidic cation-exchanges, is washed to colourless, 70% ethanol elution is to there not being ninhydrin reaction, collect pure washing liquid, being evaporated to does not have the alcohol flavor, crosses 717 strong basic type anion-exchange resins, be washed to colourless, 70% ethanol elution is collected ethanol elution to there not being ninhydrin reaction, and being evaporated to does not have the alcohol flavor, activated carbon decolorizing, lyophilization get the Folium Nerviliae fordii total amino acids extract.The aminoacid HLPC finger printing of total amino acids extract as shown in Figure 2.

As seen from Figure 2,18 seed amino acids are contained at the Folium Nerviliae fordii total amino acids position, and the relative peak area at each aminoacid peak is different with the peak area of standard 18 seed amino acids, and other has the unknown amino acid peak except that standard 18 seed amino acids.

Under aseptic condition, operate, with above-mentioned Folium Nerviliae fordii total flavonoids extract and Folium Nerviliae fordii total amino acids extract mixed according to 1: 4, packing, sterilization promptly gets Folium Nerviliae fordii powder pin.General flavone content is about 18% in the Folium Nerviliae fordii activity extract, and the content of total amino acids is about 72%.

Embodiment 2

The dry medical material 1kg of Folium Nerviliae fordii adds 60% alcohol reflux 2 times (10 times of amounts, 8 times of amounts for the second time for the first time), and each 1 hour, merge extractive liquid, was evaporated to extractum (relative density 1.05～1.25).Add the dextrin of equimultiple, the starch mix homogeneously of 4 times of amounts, make tablet by the requirement of tablet.General flavone content is about 4% in the Folium Nerviliae fordii activity extract, and total amino acids is about 16%.

Embodiment 3

The dry medical material 1kg of Folium Nerviliae fordii, add 80% alcohol reflux 2 times (12 times of amounts, 8 times of amounts for the second time for the first time), each 1 hour, merge extractive liquid,, being evaporated to does not have the alcohol flavor, adds the dilution of equimultiple water, crosses AB-8 type macroporous adsorbent resin, be washed to colourless, 50% ethanol elution is collected eluent to colourless, and being evaporated to does not have the alcohol flavor, add the dilution of equimultiple water, cross polyamide column (30～60 order), be washed to colourlessly, 80% ethanol elution is to colourless, collect eluent, being evaporated to does not have the alcohol flavor, the dry Folium Nerviliae fordii activity extract that gets, and wherein general flavone content is about 83%.

Above-mentioned blue sky extractable is dissolved with water for injection, transfer pH value to equal 9 with 10% sodium hydroxide solution, packing, sterilization promptly gets the Folium Nerviliae fordii total flavonoids injection.

Embodiment 4

Get the dry medical material 500g of Folium Nerviliae fordii, add the water reflux, extract, 2 times (15 times of amounts, 10 times of amounts for the second time for the first time), each 1 hour, merge extractive liquid, was crossed ultrafilter membrane (molecular weight 1000), be concentrated into about 1000ml, adding ethanol is 80% to containing the alcohol amount, standing over night, get supernatant concentration to about 500ml, add 0.1% active carbon and boil half an hour, filter, filtrate is crossed the 0.2um microporous filter membrane, obtain the Folium Nerviliae fordii activity extract, wherein general flavone content be about 12% and total amino acids content be about 50%.

Above-mentioned Folium Nerviliae fordii activity extract is added the injection water to 1000ml, packing (every bottle of 100ml), sterilization promptly gets the Folium Nerviliae fordii injection.

Embodiment 5

The dry medical material 1kg of Folium Nerviliae fordii, add 10 times of water gagings and soak 48h, be evaporated to 1000ml, add ethanol and make precipitation, standing over night, supernatant are evaporated to about 1000ml, cross 732 strongly acidic cation-exchanges, be washed to colourless, 80% ethanol elution is collected pure washing liquid to there not being ninhydrin reaction then, and being evaporated to does not have the alcohol flavor, cross 717 strong basic type anion-exchange resins, be washed to colourlessly, reuse 80% ethanol elution is collected ethanol elution to there not being ninhydrin reaction, being evaporated to does not have the alcohol flavor, gets reserve liquid;

Medicinal residues after water is carried add 80% alcohol reflux 2 times (10 times of amounts, 8 times of amounts for the second time for the first time), each 1 hour, merge extractive liquid,, being evaporated to does not have the alcohol flavor, add the dilution of equimultiple water, cross DP-100 type macroporous adsorbent resin, be washed to colourlessly, 65% ethanol elution is to colourless, collect eluent, being evaporated to does not have the alcohol flavor, adds the dilution of equimultiple water, crosses polyamide column (30～60 order), be washed to colourless, 60% ethanol elution is collected eluent to colourless, and being evaporated to does not have the alcohol flavor, mix with above-mentioned reserve liquid, the dry Folium Nerviliae fordii activity extract that gets is yellow to the yellowish-brown powder, and the qualitative identification experiment shows: ninhydrin reaction is positive, hydrochloric acid-magnesium powder reaction is positive, the Molish reaction is positive detection by quantitative: automatic amino acid analyzer (DIONEX) detects, and total amino acids content is about 70%; With the rutin is reference substance, and the determined by ultraviolet spectrophotometry general flavone content is about 16%.The Folium Nerviliae fordii activity extract adds the equimultiple carboxymethyl starch sodium, mixing, and dry-pressing is granulated, and is encapsulated, gets capsule.

Embodiment 6

The dry medical material 1kg of Folium Nerviliae fordii, add 80% alcohol reflux 2 times (10 times of amounts for the first time, 8 times of amounts for the second time), each 1 hour, merge extractive liquid,, be evaporated to and do not have the alcohol flavor, add the dilution of equimultiple water, cross AB-8 type macroporous adsorbent resin, be washed to colourless, 80% ethanol elution is to colourless, collect eluent, being evaporated to does not have the alcohol flavor, adds the dilution of equimultiple water, cross polyamide column (30～60 order), be washed to colourlessly, 70% ethanol elution is collected eluent to colourless, being evaporated to does not have the alcohol flavor, gets reserve liquid;

Medicinal residues after the alcohol extraction, add 10 times of water gagings and soak 36h, be evaporated to every ml and contain the about 1g of crude drug, adding ethanol is 80% to containing the alcohol amount, standing over night, and supernatant is evaporated to does not have the alcohol flavor, cross 732 strongly acidic cation-exchanges, be washed to colourlessly, 70% ethanol elution is collected pure washing liquid to there not being ninhydrin reaction, be evaporated to and do not have the alcohol flavor, cross 717 strong basic type anion-exchange resins, be washed to colourlessly, 75% ethanol elution is to there not being ninhydrin reaction, collect ethanol elution, being evaporated to does not have the alcohol flavor, mixes with reserve liquid, the dry Folium Nerviliae fordii activity extract that gets.Automatic amino acid analyzer (DIONEX) detects, and total amino acids content is about 65%; With the rutin is reference substance, and the determined by ultraviolet spectrophotometry general flavone content is about 18%.The Folium Nerviliae fordii activity extract adds equimultiple sucrose, equimultiple dextrin and appropriate amount of starch, makes granule, drying, and granulate is made 1000g, is distributed into the every bag of 10 grams.

Embodiment 7

The dry medicinal material coarse powder 1kg of Folium Nerviliae fordii, supercritical carbon dioxide fluid (including about 5% ethanol is entrainer) extracted 2 hours under 45 ℃, the condition of 20MPa, and gained extractum is the supercritical carbon dioxide extraction extract of Folium Nerviliae fordii.Get this extract, add fully degreasing decoloring of petroleum ether (60-90 ℃), the insoluble part concentrating under reduced pressure of petroleum ether, lyophilization gets the Folium Nerviliae fordii activity extract to eliminate solvent, pulverizes, cross 100 mesh sieves, promptly get the Folium Nerviliae fordii activity extract, wherein general flavone content is about 11%, and total amino acids content is about 46%.

Embodiment 8

The dry medical material 1kg of Folium Nerviliae fordii, add 70% alcohol reflux 2 times (10 times of amounts, 8 times of amounts for the second time for the first time), each 1 hour, merge extractive liquid,, being evaporated to does not have the alcohol flavor, adds the dilution of equimultiple water, crosses AB-8 type macroporous adsorbent resin, be washed to colourless, 80% ethanol elution is collected eluent to colourless, and being evaporated to does not have the alcohol flavor, add the dilution of equimultiple water, cross polyamide column (30-60 order), be washed to colourlessly, 80% ethanol elution is to colourless, collect eluent, being evaporated to does not have the alcohol flavor, and lyophilization promptly gets the Folium Nerviliae fordii total flavonoids extract.

The dry medical material of Folium Nerviliae fordii, be ground into coarse powder, get 1kg and add 10 times of water gagings immersion 24-48h, be evaporated to every ml and contain the about 1g of crude drug, adding ethanol is 80% to containing the alcohol amount, standing over night, supernatant is evaporated to does not have the alcohol flavor, crosses 732 type cation exchange resiies, is washed to colourless, 60% ethanol elution is to there not being ninhydrin reaction, collect pure washing liquid, being evaporated to does not have the alcohol flavor, crosses 717 type anion exchange resin, be washed to colourless, 50% ethanol elution is collected ethanol elution to there not being ninhydrin reaction, and being evaporated to does not have the alcohol flavor, activated carbon decolorizing, lyophilization get the Folium Nerviliae fordii total amino acids extract.

Above-mentioned Folium Nerviliae fordii total flavonoids extract that obtains and Folium Nerviliae fordii total amino acids extract are obtained the Folium Nerviliae fordii activity extract with weight ratio mixing in 2: 1, and wherein general flavone content is about 58%, and total amino acids content is about 30%.

Embodiment 9

The dry medical material 1kg of Folium Nerviliae fordii adds 10 times of water gagings and soaks 48h, filter water extract and Folium Nerviliae fordii medicinal residues.Water extraction solution is evaporated to every ml contains the about 1g of crude drug, adding ethanol is 80% to containing the alcohol amount, standing over night, supernatant is evaporated to does not have the alcohol flavor, cross 732 type cation exchange resiies, be washed to colourlessly, 75% ethanol elution is collected pure washing liquid to there not being ninhydrin reaction, be evaporated to and do not have the alcohol flavor, cross 717 type anion exchange resin, be washed to colourlessly, 80% ethanol elution is to there not being ninhydrin reaction, collect ethanol elution, being evaporated to does not have the alcohol flavor, activated carbon decolorizing, and lyophilization gets the Folium Nerviliae fordii total amino acids extract.

With the Folium Nerviliae fordii medicinal residues drying behind the above-mentioned water extraction, add 80% alcohol reflux 2 times (10 times of amounts, 8 times of amounts for the second time for the first time), each 1 hour, merge extractive liquid,, being evaporated to does not have the alcohol flavor, add the dilution of equimultiple water, cross polyamide column (30-60 order), be washed to colourlessly, 70% ethanol elution is to colourless, collect eluent, being evaporated to does not have the alcohol flavor, and lyophilization promptly gets extractive of general flavone.

With above-mentioned extractive of general flavone that obtains and total amino acids extract mixed in equal amounts, obtain the Folium Nerviliae fordii activity extract, wherein general flavone content is about 41%, and total amino acids content is about 44%.

Embodiment 10

Folium Nerviliae fordii drying medical material 1kg adds 80% alcohol reflux 2 times (10 times of amounts, 8 times of amounts for the second time for the first time), and each 1 hour, filter, merge extracted twice liquid, get alcohol extraction solution and Folium Nerviliae fordii medicinal residues.Alcohol extract is evaporated to does not have the alcohol flavor, adds the dilution of equimultiple water, crosses AB-8 type macroporous adsorbent resin, is washed to colourlessly, and 70% ethanol elution is collected eluent to colourless, and being evaporated to does not have the alcohol flavor, and lyophilization promptly gets extractive of general flavone.

After the Folium Nerviliae fordii medicinal residues drying after the above-mentioned alcohol extraction, 10 times of water gagings soak 48h, filter the Folium Nerviliae fordii water extraction solution.Water extraction solution is evaporated to every ml contains the about 1g of crude drug, adding ethanol is 80% to containing the alcohol amount, standing over night, supernatant is evaporated to does not have the alcohol flavor, cross 732 type cation exchange resiies, be washed to colourlessly, 70% ethanol elution is collected pure washing liquid to there not being ninhydrin reaction, be evaporated to and do not have the alcohol flavor, cross 717 type anion exchange resin, be washed to colourlessly, 70% ethanol elution is to there not being ninhydrin reaction, collect ethanol elution, being evaporated to does not have the alcohol flavor, activated carbon decolorizing, and lyophilization gets the Folium Nerviliae fordii total amino acids extract.

Above-mentioned extractive of general flavone that obtains and total amino acids extract are obtained the Folium Nerviliae fordii activity extract according to mixing in 4: 1, and wherein general flavone content is about 71%, and total amino acids content is about 12%.

Embodiment 11

Get the dry medical material 500g of Folium Nerviliae fordii, add the water reflux, extract, 2 times (10 times of amounts, 8 times of amounts for the second time for the first time), each 1 hour, merge extractive liquid, was crossed ultrafilter membrane (molecular weight 1000), be concentrated into about 800ml, adding ethanol is 85% to containing the alcohol amount, standing over night, get supernatant concentration to about 500ml, add 0.2% activated carbon decolorizing, filter, filtrate is crossed the 0.2um microporous filter membrane, concentrating under reduced pressure, drying gets the Folium Nerviliae fordii activity extract.Wherein general flavone content be about 10% and total amino acids content be about 48%.

Embodiment 12

The dry medical material 1kg of Folium Nerviliae fordii adds 80% alcohol reflux 3 times (10 times of amounts, 6 times of amounts for the second time for the first time, 6 times of amounts for the third time), each 0.5 hour, merge extractive liquid,, being evaporated to does not have the alcohol flavor, add the dilution of equimultiple water, cross DP-100 type macroporous adsorbent resin, be washed to colourlessly, 80% ethanol elution is to colourless, collect eluent, being evaporated to does not have the alcohol flavor, adds the dilution of equimultiple water, crosses polyamide column (30～60 order), be washed to colourless, 80% ethanol elution is collected eluent to colourless, and being evaporated to does not have the alcohol flavor, drying promptly gets the Folium Nerviliae fordii total flavonoids extract.

The dry medical material of Folium Nerviliae fordii, be ground into coarse powder, get 1kg and add 15 times of water gagings immersion 40h, be evaporated to every ml and contain the about 1g of crude drug, adding ethanol is 80% to containing the alcohol amount, standing over night, supernatant is evaporated to does not have the alcohol flavor, crosses 732 strongly acidic cation-exchanges, is washed to colourless, 70% ethanol elution is to there not being ninhydrin reaction, collect pure washing liquid, being evaporated to does not have the alcohol flavor, crosses 717 strong basic type anion-exchange resins, be washed to colourless, 70% ethanol elution is collected ethanol elution to there not being ninhydrin reaction, and being evaporated to does not have the alcohol flavor, activated carbon decolorizing, the dry Folium Nerviliae fordii total amino acids extract that gets.

With above-mentioned Folium Nerviliae fordii total flavonoids extract and Folium Nerviliae fordii total amino acids extract according to weight ratio mix at 1: 1 the Folium Nerviliae fordii activity extract, wherein general flavone content is about 39%, the content of total amino acids is about 45%.

Embodiment 13

The dry medical material 1kg of Folium Nerviliae fordii, add 80% alcohol reflux 2 times (10 times of amounts, 8 times of amounts for the second time for the first time), each 1 hour, merge extractive liquid,, being evaporated to does not have the alcohol flavor, adds the dilution of equimultiple water, crosses AB-8 type macroporous adsorbent resin, be washed to colourless, 80% ethanol elution is collected eluent to colourless, and being evaporated to does not have the alcohol flavor, add the dilution of equimultiple water, cross polyamide column (30～60 order), be washed to colourlessly, 80% ethanol elution is to colourless, collect eluent, being evaporated to does not have the alcohol flavor, and lyophilization promptly gets the Folium Nerviliae fordii total flavonoids extract.

The dry medical material of Folium Nerviliae fordii, be ground into coarse powder, get 1kg and add 10 times of water gagings immersion 24h, be evaporated to every ml and contain the about 1g of crude drug, adding ethanol is 80% to containing the alcohol amount, standing over night, supernatant is evaporated to does not have the alcohol flavor, crosses 732 strongly acidic cation-exchanges, is washed to colourless, 65% ethanol elution is to there not being ninhydrin reaction, collect pure washing liquid, being evaporated to does not have the alcohol flavor, crosses 717 strong basic type anion-exchange resins, be washed to colourless, 70% ethanol elution is collected ethanol elution to there not being ninhydrin reaction, and being evaporated to does not have the alcohol flavor, activated carbon decolorizing, lyophilization get the Folium Nerviliae fordii total amino acids extract.

Above-mentioned Folium Nerviliae fordii total flavonoids extract and Folium Nerviliae fordii total amino acids extract are obtained the Folium Nerviliae fordii activity extract according to weight ratio mixing in 10: 1, and wherein general flavone content is about 77%, and the content of total amino acids is about 9%.

Embodiment 14

The dry medical material 1kg of Folium Nerviliae fordii, add 15 times of water gagings and soak 48h, be evaporated to 1000ml, add ethanol and make precipitation, standing over night, supernatant are evaporated to about 1000ml, cross 732 strongly acidic cation-exchanges, be washed to colourless, 80% ethanol elution is collected pure washing liquid to there not being ninhydrin reaction then, and being evaporated to does not have the alcohol flavor, cross 717 strong basic type anion-exchange resins, be washed to colourlessly, reuse 80% ethanol elution is collected ethanol elution to there not being ninhydrin reaction, being evaporated to does not have the alcohol flavor, the dry Folium Nerviliae fordii total amino acids extract that gets.

Medicinal residues after water is carried, add 80% alcohol reflux 2 times (12 times of amounts for the first time, 10 times of amounts for the second time), each 1 hour, merge extractive liquid,, be evaporated to and do not have the alcohol flavor, add the dilution of equimultiple water, cross DP-100 type macroporous adsorbent resin, be washed to colourless, 60% ethanol elution is to colourless, collect eluent, being evaporated to does not have the alcohol flavor, adds the dilution of equimultiple water, cross polyamide column (30～60 order), be washed to colourlessly, 70% ethanol elution is collected eluent to colourless, being evaporated to does not have the alcohol flavor, the dry Folium Nerviliae fordii total flavonoids extract that gets.

Obtain to such an extent that Folium Nerviliae fordii total flavonoids extract and total amino acids extract one weight ratio are mixed and obtained the Folium Nerviliae fordii activity extract at 1: 10 with above-mentioned, wherein total amino acids content is about 81%; General flavone content is 8%.

Embodiment 15

The dry medical material 1kg of Folium Nerviliae fordii, add 80% alcohol reflux 2 times (10 times of amounts for the first time, 8 times of amounts for the second time), each 1 hour, merge extractive liquid,, be evaporated to and do not have the alcohol flavor, add the dilution of equimultiple water, cross AB-8 type macroporous adsorbent resin, be washed to colourless, 70% ethanol elution is to colourless, collect eluent, being evaporated to does not have the alcohol flavor, adds the dilution of equimultiple water, cross polyamide column (30～60 order), be washed to colourlessly, 70% ethanol elution is collected eluent to colourless, being evaporated to does not have the alcohol flavor, and drying obtains extractive of general flavone.

Medicinal residues after the alcohol extraction, add 10 times of water gagings and soak 36h, be evaporated to every ml and contain the about 1g of crude drug, adding ethanol is 80% to containing the alcohol amount, standing over night, supernatant is evaporated to does not have the alcohol flavor, crosses 732 strongly acidic cation-exchanges, is washed to colourless, 70% ethanol elution is to there not being ninhydrin reaction, collect pure washing liquid, being evaporated to does not have the alcohol flavor, crosses 717 strong basic type anion-exchange resins, be washed to colourless, 70% ethanol elution is collected ethanol elution to there not being ninhydrin reaction, the dry total amino acids extract that gets.

With above-mentioned extractive of general flavone and total amino acids extract with weight ratio mix at 1: 5 the Folium Nerviliae fordii activity extract, wherein total amino acids content is about 75%; General flavone content is about 14%.

Embodiment 16

The dry medical material 1kg of Folium Nerviliae fordii, add 80% alcohol reflux 2 times (6 times of amounts, 6 times of amounts for the second time for the first time), each 2 hours, merge extractive liquid,, being evaporated to does not have the alcohol flavor, adds the dilution of equimultiple water, crosses AB-8 type macroporous adsorbent resin, be washed to colourless, 70% ethanol elution is collected eluent to colourless, and being evaporated to does not have the alcohol flavor, add the dilution of equimultiple water, cross polyamide column (30～60 order), be washed to colourlessly, 70% ethanol elution is to colourless, collect eluent, being evaporated to does not have the alcohol flavor, and lyophilization promptly gets the Folium Nerviliae fordii total flavonoids extract.

The dry medical material of Folium Nerviliae fordii, be ground into coarse powder, get 1kg and add 12 times of water gagings immersion 24h, be evaporated to every ml and contain the about 1g of crude drug, adding ethanol is 80% to containing the alcohol amount, standing over night, supernatant is evaporated to does not have the alcohol flavor, crosses 732 strongly acidic cation-exchanges, is washed to colourless, 70% ethanol elution is to there not being ninhydrin reaction, collect pure washing liquid, being evaporated to does not have the alcohol flavor, crosses 717 strong basic type anion-exchange resins, be washed to colourless, 70% ethanol elution is collected ethanol elution to there not being ninhydrin reaction, and being evaporated to does not have the alcohol flavor, activated carbon decolorizing, lyophilization get the Folium Nerviliae fordii total amino acids extract.

Operate under aseptic condition, above-mentioned Folium Nerviliae fordii total flavonoids extract and Folium Nerviliae fordii total amino acids extract are obtained the Folium Nerviliae fordii activity extract according to weight ratio mixing in 3: 10, wherein general flavone content is about 20%, and total amino acids content is about 70%.

Embodiment 17

The dry medical material 1kg of Folium Nerviliae fordii, add 15 times of water gagings and soak 48h, be evaporated to 1000ml, add ethanol and make precipitation, standing over night, supernatant are evaporated to about 1000ml, cross 732 strongly acidic cation-exchanges, be washed to colourless, 80% ethanol elution is collected pure washing liquid to there not being ninhydrin reaction then, and being evaporated to does not have the alcohol flavor, cross 717 strong basic type anion-exchange resins, be washed to colourlessly, reuse 80% ethanol elution is collected ethanol elution to there not being ninhydrin reaction, being evaporated to does not have the alcohol flavor, the dry Folium Nerviliae fordii total amino acids extract that gets.

Medicinal residues after water is carried, add 80% alcohol reflux 2 times (12 times of amounts for the first time, 8 times of amounts for the second time), each 1.5 hours, merge extractive liquid,, be evaporated to and do not have the alcohol flavor, add the dilution of equimultiple water, cross DP-100 type macroporous adsorbent resin, be washed to colourless, 50% ethanol elution is to colourless, collect eluent, being evaporated to does not have the alcohol flavor, adds the dilution of equimultiple water, cross polyamide column (30～60 order), be washed to colourlessly, 70% ethanol elution is collected eluent to colourless, being evaporated to does not have the alcohol flavor, the dry Folium Nerviliae fordii total flavonoids extract that gets.

Obtain to such an extent that the weight ratio of Folium Nerviliae fordii total flavonoids extract and total amino acids extract is mixed and obtained the Folium Nerviliae fordii activity extract at 3: 10 with above-mentioned, wherein total amino acids content is 70%; General flavone content is 20%.

Embodiment 18

The dry medical material 1kg of Folium Nerviliae fordii, add 80% alcohol reflux 2 times (6 times of amounts for the first time, 6 times of amounts for the second time), each 2 hours, merge extractive liquid,, be evaporated to and do not have the alcohol flavor, add the dilution of equimultiple water, cross AB-8 type macroporous adsorbent resin, be washed to colourless, 70% ethanol elution is to colourless, collect eluent, being evaporated to does not have the alcohol flavor, adds the dilution of equimultiple water, cross polyamide column (30～60 order), be washed to colourlessly, 70% ethanol elution is collected eluent to colourless, being evaporated to does not have the alcohol flavor, and drying obtains extractive of general flavone.

Medicinal residues after the alcohol extraction, add 12 times of water gagings and soak 36h, be evaporated to every ml and contain the about 1g of crude drug, adding ethanol is 80% to containing the alcohol amount, standing over night, supernatant is evaporated to does not have the alcohol flavor, crosses 732 strongly acidic cation-exchanges, is washed to colourless, 70% ethanol elution is to there not being ninhydrin reaction, collect pure washing liquid, being evaporated to does not have the alcohol flavor, crosses 717 strong basic type anion-exchange resins, be washed to colourless, 70% ethanol elution is collected ethanol elution to there not being ninhydrin reaction, and drying obtains the total amino acids extract.

With above-mentioned extractive of general flavone and total amino acids extract with weight ratio mix at 3: 10 the Folium Nerviliae fordii activity extract, wherein total amino acids content is about 70%; General flavone content is about 20%.

Embodiment 19

The dry medical material 1kg of Folium Nerviliae fordii adds 70% alcohol reflux 2 times (12 times of amounts, 6 times of amounts for the second time for the first time), and each 0.5 hour, merge extractive liquid, was evaporated to extractum (relative density 1.05～1.25).Add the dextrin of equimultiple, the starch mix homogeneously of 4 times of amounts, make tablet by the requirement of tablet.General flavone content is about 5% in the Folium Nerviliae fordii activity extract, and total amino acids is about 15%.

Embodiment 20

The dry medical material 1kg of Folium Nerviliae fordii adds 10% alcohol reflux 2 times (10 times of amounts, 6 times of amounts for the second time for the first time), and each 1.5 hours, merge extractive liquid, was evaporated to extractum (relative density 1.05～1.25).Add the dextrin of equimultiple, the starch mix homogeneously of 4 times of amounts, make tablet by the requirement of tablet.General flavone content is about 4% in the Folium Nerviliae fordii activity extract, and total amino acids is about 18%.

Embodiment 21

The dry medical material 1kg of Folium Nerviliae fordii adds 50% alcohol reflux 2 times (10 times of amounts, 8 times of amounts, 6 times of amounts for the third time for the second time for the first time), and each 1 hour, merge extractive liquid, was evaporated to extractum (relative density 1.05～1.25).Add equimultiple sucrose, equimultiple dextrin and appropriate amount of starch, make granule, drying, granulate is made granule.General flavone content is about 4% in the Folium Nerviliae fordii activity extract, and total amino acids is about 16%.

Embodiment 22

The dry medical material 1kg of Folium Nerviliae fordii adds 30% alcohol reflux 2 times (10 times of amounts, 8 times of amounts for the second time for the first time), and each 1 hour, merge extractive liquid, was evaporated to extractum (relative density 1.05～1.25).Add the dextrin of equimultiple, the starch mix homogeneously of 4 times of amounts, make tablet by the requirement of tablet.General flavone content is about 4% in the Folium Nerviliae fordii activity extract, and total amino acids is about 16%.

Claims (12)

1, a kind of Folium Nerviliae fordii activity extract contains 10～90% (mass percent) total flavones and 10～90% (mass percent) total amino acids.

2, the described Folium Nerviliae fordii activity extract of claim 1 wherein contains 70% (mass percent) total amino acids and 20% (mass percent) total flavones.

3, a kind of method for preparing claim 1 or 2 described Folium Nerviliae fordii activity extracts, 0～80% alcohol reflux 2～3 times that adds 6～15 times of amounts in the dry medical material of Folium Nerviliae fordii, each 0.5～2 hour, merge extractive liquid,, concentrate, use macroporous adsorbent resin, polyamide, anion exchange resin, cation exchange resin purify the Folium Nerviliae fordii activity extract.

4, the described Folium Nerviliae fordii activity extract of claim 3 preparation method, it is characterized in that at the dry medical material of Folium Nerviliae fordii, 10～70% alcohol refluxs 2～3 times that add 6～12 times of amounts, each 1～2 hour, merge extractive liquid,, being concentrated into does not have the alcohol flavor, cross the AB-8 macroporous adsorbent resin, be washed to colourlessly, collected post liquid and water lotion and get reserve liquid, 50～80% ethanol elutions are to colourless then, collect eluent, being evaporated to does not have the alcohol flavor, crosses polyamide column, is washed to colourless, reuse 50～80% ethanol elutions are to colourless, collect eluent, being evaporated to does not have the alcohol flavor, the dry Folium Nerviliae fordii total flavonoids extract that gets; Get reserve liquid and cross 732 type cation exchange resiies, be washed to colourless, 50～80% ethanol elutions are collected pure washing liquid to there not being ninhydrin reaction then, and being evaporated to does not have the alcohol flavor, after 717 type anion exchange resin, be washed to colourlessly, reuse 50～80% ethanol elutions are collected ethanol elution to there not being ninhydrin reaction, being evaporated to does not have the alcohol flavor, the dry Folium Nerviliae fordii total amino acids extract that gets; Merge the Folium Nerviliae fordii activity extract.

5. method for preparing claim 1 or 2 described Folium Nerviliae fordii activity extracts, use contains the supercritical carbon dioxide extraction Folium Nerviliae fordii of 2.5%～10% ethanol entrainer, the extract solvent removed by evaporation at reduced pressure, add the defat with petroleum ether decolouring, petroleum ether does not dissolve partly to remove through the reduction vaporization lyophilization and desolvates, and gets product.

6. a method for preparing claim 1 or 2 described Folium Nerviliae fordii activity extracts at first prepares Folium Nerviliae fordii total amino acids extract and Folium Nerviliae fordii total flavonoids extract, then Folium Nerviliae fordii total amino acids extract and the mixing of Folium Nerviliae fordii total flavonoids extract is obtained.

7. the preparation method of the described Folium Nerviliae fordii activity extract of claim 6, it is characterized in that the Folium Nerviliae fordii total amino acids extract prepares according to following method: in the dry medical material of Folium Nerviliae fordii, add 10～15 times of water gagings and soak 24～48h, be evaporated to small size, add 80% ethanol and make precipitation, standing over night, supernatant is evaporated to does not have the alcohol flavor, cross 732 cation exchange resiies, be washed to colourlessly, 50～80% ethanol elutions are to there not being ninhydrin reaction then, collect pure washing liquid, being evaporated to does not have the alcohol flavor, crosses 717 anion exchange resin, is washed to colourless, reuse 50～80% ethanol elutions do not have ninhydrin reaction, collect ethanol elution concentrating under reduced pressure drying.

8. the preparation method of the described Folium Nerviliae fordii activity extract of claim 6, it is characterized in that the Folium Nerviliae fordii total flavonoids extract is according to the preparation of following method: 80% alcohol reflux 2～3 times that in the dry medical material of Folium Nerviliae fordii, adds 6～12 times of amounts, each 1～2 hour, merge extractive liquid,, be evaporated to and do not have the alcohol flavor, cross the AB-8 macroporous adsorbent resin, be washed to colourlessly, 50～80% ethanol elutions are collected eluent to colourless then, be evaporated to and do not have the alcohol flavor, cross polyamide column, be washed to colourlessly, reuse 50～80% ethanol elutions are to colourless, collect eluent, the concentrating under reduced pressure drying.

9. a method for preparing claim 1 or 2 described Folium Nerviliae fordii activity extracts is, in the dry medical material of Folium Nerviliae fordii, add 10～15 times of water gagings and soak 24～48h, be evaporated to small size, add ethanol and make precipitation, standing over night, supernatant are evaporated to does not have the alcohol flavor, crosses 732 type cation exchange resiies, be washed to colourless, 50～80% ethanol elutions are collected pure washing liquid to there not being ninhydrin reaction then, and being evaporated to does not have the alcohol flavor, cross 717 type anion exchange resin, be washed to colourlessly, reuse 50～80% ethanol elutions are collected ethanol elution to there not being ninhydrin reaction, being evaporated to does not have the alcohol flavor, standby; Medicinal residues after water is carried add 80% alcohol reflux 2～3 times of 6～12 times of amounts, each 1～2 hour, merge extractive liquid,, being evaporated to does not have the alcohol flavor, crosses macroporous adsorbent resin, be washed to colourlessly, 50～80% ethanol elutions are collected eluent to colourless then, being evaporated to does not have the alcohol flavor, crosses polyamide column, is washed to colourless, reuse 50～80% ethanol elutions are collected eluent to colourless, and being evaporated to does not have the alcohol flavor, mix the dry Folium Nerviliae fordii extract that gets with above-mentioned reserve liquid.

10. a method for preparing claim 1 or 2 described Folium Nerviliae fordii activity extracts is, 80% alcohol reflux 2～3 times that in the dry medical material of Folium Nerviliae fordii, adds 6～12 times of amounts, each 1～2 hour, merge extractive liquid,, be evaporated to and do not have the alcohol flavor, cross the AB-8 macroporous adsorbent resin, be washed to colourlessly, 50～80% ethanol elutions are collected eluent to colourless then, be evaporated to and do not have the alcohol flavor, cross polyamide column, be washed to colourlessly, reuse 50～80% ethanol elutions are to colourless, collect eluent, be evaporated to nothing alcohol and distinguish the flavor of standby; Medicinal residues after the alcohol extraction, add 10～15 times of water gagings and soak 24～48h, be evaporated to small size, add ethanol and make precipitation, standing over night, supernatant is evaporated to does not have the alcohol flavor, cross 732 cation exchange resiies, be washed to colourlessly, 50～80% ethanol elutions are collected pure washing liquid to there not being ninhydrin reaction then, be evaporated to and do not have the alcohol flavor, cross 717 anion exchange resin, be washed to colourlessly, reuse 50～80% ethanol elutions are to there not being ninhydrin reaction, collect ethanol elution, being evaporated to does not have the alcohol flavor, mixes with above-mentioned reserve liquid, the dry Folium Nerviliae fordii extract that gets.

11. a pharmaceutical composition contains the claim 1 for the treatment of drug effect or 2 described Folium Nerviliae fordii activity extracts and acceptable adjuvant pharmaceutically.

12. claim 1 or the 2 described Folium Nerviliae fordii activity extracts application in the viral respiratory system disease medicine of preparation treatment.

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