CN102552232B - Effective fraction of Xiao Ban Xia Tang, and preparation method and application thereof - Google Patents
Effective fraction of Xiao Ban Xia Tang, and preparation method and application thereof Download PDFInfo
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Abstract
The invention discloses an effective fraction of Xiao Ban Xia Tang. The effective fraction has physicochemical properties that: the effective fraction is a yellowish-brown extract, has certain moisture absorption and can be dissolved in hot water and ethanol; and the effective fraction mainly contains organic acid components, such as succinic acid, hydroxysuccinic acid, citric acid and linoleic acid. The invention also discloses a preparation method and application of the effective fraction of the Xiao Ban Xia Tang. An organic acid part extracted from the Xiao Ban Xia Tang has obvious pharmacological and pharmacodynamic effects. The effective fraction has an obvious effect of arresting vomiting; and through experimental models of pigeon emesis induced by both bluestone and cisplatin, results indicate that the organic acid part has obvious activity of arresting vomiting and can obviously reduce the vomiting frequency of pigeons; and the effective fraction has an effect of inhibiting gastric emptying of normal mice.
Description
Technical field
The invention belongs to technical field of Chinese medicines, be specifically related to effective site of a kind of Small Pinelliae Decoction and its preparation method and application.
Background technology
Vomiting is the disease that clinical common phenomenon, particularly Patients Receiving Chemotherapy cannot be avoided, and a lot of patients are postoperative vomiting and have to abandon chemotherapy or radiotherapy for fear often, thereby delays the state of an illness.Though it is many to be used for the treatment of the medicine of vomiting at present, but effect is unsatisfactory, though resemble ondansetron, this class medicine of granisetron has certain effect to emesis of chemotherapy, because these drug prices are expensive and certain side effect is arranged, also limited their use to a certain extent.
Vomiting belongs to " QI rising in reverse order " category in Chinese medicine, think that vomiting is because stomach-QI being unable to descend normally, adverse rising of stomach-QI are caused, and the treatment vomiting should be with regulating functional activities of qi, and stopping nausea and vomiting by lowering the adverse flow of QI is main.Chinese medicine has the irreplaceable advantage of chemical synthetic drug, low price, and toxic and side effects is little.But method and the related preparations of isolating at present drug effective region with good curing vomiting and active component from pure Chinese medicinal preparation are actually rare.
The present invention isolates the effective site with resisting emesis activity from the classical Chinese medicine name side " Small Pinelliae Decoction " with resisting emesis effect.
Summary of the invention
Technical problem to be solved by this invention provides the active site of Small Pinelliae Decoction.
The technical problem that the present invention also will solve provides the preparation method of the active site of above-mentioned Small Pinelliae Decoction.
The technical problem that the present invention will solve at last provides the application of the active site of above-mentioned Small Pinelliae Decoction.
For solving the problems of the technologies described above, the technical solution used in the present invention is as follows:
The effective site of Small Pinelliae Decoction, the physicochemical property of this effective site is: pale brown color extractum, have certain moisture, dissolve in hot water and ethanol; This effective site mainly contains the organic acid composition, and wherein, succinic acid content is 3.0～3.8%, hydroxyl succinic acid content is 21.6～26.8%, citric acid content is 30.6～38.6%, linoleic content is 0.92～1.26%.
Above-mentioned content is weight percentage, and 100% benchmark is the effective site gross weight, below identical.
The preparation method of the effective site of above-mentioned Small Pinelliae Decoction, with the Rhizoma Pinelliae and Rhizoma Zingiberis Recens by weight 1: 1 mix homogeneously, extracting in water 3～5h, filter extracting solution, medicinal residues continue to add water, extract 2～3h, merge extracted twice liquid; Extracting solution add ethanol to pure quality percentage composition be 60～80%, leave standstill 8～12h after, centrifugal, get supernatant, be concentrated into 1～2g crude drug mL
-1The concentrated solution extracted with diethyl ether obtains ether layer and water layer, and water layer is transferred to pH7 with NaOH solution, adsorb through anion exchange resin, wash with deionized water again and mix to colourless, with NaOH eluant solution resin, get eluent, this eluent transfers to pH3 with HCl, be evaporated to dried, again with anhydrous alcohol solution to remove NaCl, obtain the organic acid position namely.
Wherein, during twice extracting in water, for the first time the weight of water is the Rhizoma Pinelliae and Rhizoma Zingiberis Recens gross weight 5～8 times, and the weight of water is the Rhizoma Pinelliae and Rhizoma Zingiberis Recens gross weight 3～5 times for the second time.
The another kind of preparation method of the effective site of above-mentioned Small Pinelliae Decoction by weight 1: 1 mix homogeneously, adds water with the Rhizoma Pinelliae and Rhizoma Zingiberis Recens, and the weight of water is the Rhizoma Pinelliae and Rhizoma Zingiberis Recens gross weight 5～8 times, and the dress volatile oil extractor extracts 3～5h, gets aqueous extract; Medicinal residues continue to add water, and the weight of water is the Rhizoma Pinelliae and Rhizoma Zingiberis Recens gross weight 3～5 times, and the dress volatile oil extractor extracts 2～3h, merge aqueous extract twice, add ethanol to pure quality percentage composition be 60～80%, leave standstill 8～12h after, centrifugal, get supernatant, be concentrated into 1～2g crude drug mL
-1The concentrated solution extracted with diethyl ether obtains ether layer and water layer, and water layer is transferred to pH7 with NaOH solution, adsorb through anion exchange resin, wash with deionized water again and mix to colourless, with NaOH eluant solution resin, get eluent, this eluent transfers to pH3 with HCl, be evaporated to dried, again with anhydrous alcohol solution to remove NaCl, obtain the organic acid position namely.
Wherein, during extracted with diethyl ether, extracting twice, the volume of each used ether is 0.8～1 times of amount of concentrated solution volume.
Wherein, ether layer concentrates when reclaiming ether, adopts the water-bath normal pressure, and bath temperature is 39～49 ℃.
Wherein, described anion exchange resin is D315 type anion exchange resin.
Wherein, the used NaOH solution concentration of eluting resin is 0.5～1molL
-1
Wherein, be evaporated to when doing, thickening temperature is 60～70 ℃.
The application of the effective site of above-mentioned Small Pinelliae Decoction in preparation preventing or arresting vomiting medicine.
Wherein, described effective site also can add acceptable accessories or excipient.
Wherein, described effective site can be made into acceptable forms clinically.
Wherein, the dosage form that can be made into of described effective site is any one in tablet, capsule, pill, granule, suspensoid, drop pill, oral liquid, injection, aerosol, suppository or the subcutaneous administration dosage form.
Beneficial effect: the present invention extracts the organic acid position that obtains and has tangible pharmacological effect effect from Small Pinelliae Decoction.
(1) the preventing or arresting vomiting effect is obvious.Causing pigeon vomiting and chemotherapeutics with copper sulfate---it is experimental model that cisplatin causes the pigeon vomiting, and the result shows that the organic acid position has significant preventing or arresting vomiting activity, can obviously reduce pigeon vomiting number of times.
(2) has the effect that suppresses the normal mouse gastric emptying.
Description of drawings
Figure 1A is the chromatogram that the GC/MS of reference substance detects.
Figure 1B is the chromatogram that the GC/MS of Rhizoma Zingiberis Recens sample detects.
Fig. 1 C is the chromatogram that the GC/MS of Rhizoma Pinelliae sample detects.
Fig. 1 D is the chromatogram that the GC/MS of Small Pinelliae Decoction sample detects.
Among Figure 1A to Fig. 1 D, the 1-dimethyl fumarate; 2-dimethyl succinate (dimethyl succinate); The 3-methyl laurate; 4-3-carbonyl methyl caproate; 5-3-hydroxy-2-methyl Glutaric Acid Dimethyl ester; 6-2,4-muconic acid dimethyl ester; 7-hydroxyl succinic acid dimethyl ester; 8-cinnamic acid methyl ester; The 9-dimethyl azelate; The 10-methyl hexadecanoate; 11-propylene-1,2,3-tricarboxylic acid trimethyl; The 12-dimethyl terephthalate (DMT); The 13-dimethyl phthalate; The 14-methyl linoleate; The 15-trimethyl citrate; 16-2-hydroxyl-methyl palmitate; 17-4-hydroxy 3-methoxybenzene methyl formate; 18-methyl para Toluic Acid methyl ester; The S-methyl salicylate.
Fig. 2 A is the GC collection of illustrative plates of reference substance.
The GC collection of illustrative plates of the negative sample of Fig. 2 B (lacking the Rhizoma Pinelliae).
Fig. 2 C is the GC collection of illustrative plates of sample.
Among Fig. 2 A to Fig. 2 C, the 1-dimethyl succinate; 2-hydroxyl succinic acid dimethyl ester; 3-methyl salicylate (interior mark); The 4-trimethyl citrate; The 5-methyl linoleate
The specific embodiment
According to following embodiment, the present invention may be better understood.Yet, those skilled in the art will readily understand that embodiment is described only to be used for explanation the present invention, and should also can not limit the present invention described in detail in claims.
Embodiment 1:
The Rhizoma Pinelliae and Rhizoma Zingiberis Recens by weight 1: 1 mix homogeneously, are added the water extraction 3～5h of 5～8 times of weight, filter extracting solution, medicinal residues continue to add the water of 3～5 times of weight, extract 2～3h, merge extracted twice liquid; Extracting solution add ethanol to pure mass content be 60～80%, leave standstill 8～12h after, 4500 commentaries on classics/min are centrifugal, get supernatant, are concentrated into 1～2g crude drug mL
-1, concentrated solution obtains ether layer and water layer with twice of extracted with diethyl ether (each is 0.8～1 times of volume of concentrated solution with the ether amount), water layer is transferred to pH7 with NaOH, after solution is clarified substantially, adsorb through anion exchange resin, wash assorted to colourless, with 0.5～1molL again with deionized water
-1NaOH eluant solution resin, get eluent, this eluent transfers to pH3 with HCl, is evaporated to driedly, dissolves 3 times repeatedly to remove NaCl with dehydrated alcohol again, obtains the organic acid position namely.
This active site is yellowish-brown extractum, has certain moisture, dissolves in hot water and ethanol.Analysis is detected at this position, find that this position mainly contains the organic acid composition, GC/MS detects 18 kinds of organic acid compositions altogether, and wherein content is higher citric acid, hydroxyl succinic acid, succinic acid, linoleic acid etc.Adopt the gas chromatogram internal standard method that these organic acid compositions have been carried out assay, found that succinic acid content is 3.0～3.8%, hydroxyl succinic acid content is 21.6～26.8%, citric acid content is 30.6～38.6%, linoleic content is 0.92～1.26%.
Perhaps, the Rhizoma Pinelliae and Rhizoma Zingiberis Recens by weight 1: 1 mix homogeneously, are added water, the weight of water is the Rhizoma Pinelliae and Rhizoma Zingiberis Recens gross weight 5～8 times, and the dress volatile oil extractor extracts 3～5h, gets aqueous extract; Medicinal residues continue to add water, the weight of water is the Rhizoma Pinelliae and Rhizoma Zingiberis Recens gross weight 3～5 times, the dress volatile oil extractor extracts 2～3h, merge aqueous extract twice, add ethanol to pure quality percentage composition be 60～80%, leave standstill 8～12h after, 4500 commentaries on classics/min are centrifugal, get supernatant, be concentrated into 1～2g crude drug mL
-1, concentrated solution obtains ether layer and water layer with twice of extracted with diethyl ether (each is 0.8～1 times of volume of concentrated solution with the ether amount), water layer is transferred to pH7 with NaOH, after solution is clarified substantially, adsorb through anion exchange resin, wash assorted to colourless, with 0.5～1molL again with deionized water
-1NaOH eluant solution resin, get eluent, this eluent transfers to pH3 with HCl, is evaporated to driedly, dissolves 3 times repeatedly to remove NaCl with dehydrated alcohol again, namely gets the organic acid position.The live part main component that obtains by this method is consistent with the said method result.
Embodiment 2: the detection method of organic acid composition in the Small Pinelliae Decoction.
One, instrument and reagent:
1, instrument
Electronic balance BP211D (German Sartorius company); Agilent6890/MSD5973 system (U.S. Hewlett-Packard Corporation); MICROFUGE microcentrifuge (U.S. BACKMAN company).
2, reference substance
Succinic acid (Succinic acid, lot number 090812, Shanghai Ling Feng chemical reagent company limited), hydroxyl succinic acid (Malic acid, lot number 20090328, the emerging biochemical reagents company limited of China's favour), citric acid (Citric acid, lot number 081245071, Nanjing Chemistry Reagent Co., Ltd.), linoleic acid (Linoleic acid, lot number 10129622, AlfaAesar), salicylic acid (Salicylic acid, lot number 060222, green grass or young crops is analysed Chemical Industry Science Co., Ltd in Shanghai), Palmic acid (Palmitic acid, lot number 081139075, Nanjing Chemistry Reagent Co., Ltd.), phthalic acid, gallic acid, the L-arginine, tartaric acid etc. are analytical pure.
3, reagent
Chloroform, methanol, concentrated sulphuric acid etc. are analytical pure.
Two, the preparation of reference substance and sample:
1, the preparation of sample
Take by weighing each 0.1g of sample at the organic acid position of the Rhizoma Pinelliae that obtains through D-315 type anion exchange resin process, Rhizoma Zingiberis Recens, Small Pinelliae Decoction respectively, place the 50mL reaction bulb, add the H of (7: 100)
2SO
4-CH
3OH 4mL shakes up, and places 60 ℃ of water-bath heating 24h esterification, transfers in the separatory funnel that fills the 10mL deionized water CHCl
3Extraction, CHCl
3Carrying out GC/MS behind the layer anhydrous sodium sulfate drying detects.
2, the preparation of reference substance
Reference substances such as citric acid, hydroxyl succinic acid, succinic acid, Palmic acid, phthalic acid, L-arginine, tartaric acid are carried out esterification by the sample step.
Three, GC/MS condition:
1, GC condition
Chromatographic column: HP-INNOWAX (0.32mm * 30m * 0.5 μ m).
Injector: split sampling, split ratio 10: 1.
Injector temperature: 250 ℃.
Carrier gas: nitrogen, flow velocity 5.0mLmin
-1
Sample size: 1 μ L.
Temperature programming: 100 ℃ of chromatographic column initial temperatures keep 3min, 5 ℃ of min of heating rate
-1, reach 220 ℃ and keep 5min, 10 ℃ of min of heating rate
-1, 260 ℃ of final temperatures keep 2min.
2, MS condition
The EI source, electron bombard energy: 70ev; Multiplier voltage: 1953v; Ion source temperature: 230 ℃; Quadrupole rod temperature: 150 ℃; Solvent delay: 1min.
Mass scanning scope: 55-550.
Four, experimental result:
Mix the reference substance gas chromatogram and see Fig. 1, Rhizoma Zingiberis Recens, the Rhizoma Pinelliae, Small Pinelliae Decoction organic acid position gas chromatogram are seen Fig. 1.
By mass spectral analysis and database retrieval, and with reference to the reference substance collection of illustrative plates, obtain chemical constitution and the title at each position, tie as shown in table 1.
The GC/MS analysis result at table 1 Small Pinelliae Decoction organic acid position
Experimental example 3: the mensuration of several organic acid contents in the organic acid position---adopt gas chromatogram internal standard method (salicylic acid is interior mark)
One, instrument and reagent:
1, instrument
Agilent 4890 gas chromatograpies, fid detector; Sartorius BP211D type electronic balance.
Organic acid reference substance: succinic acid (Succinic acid, lot number 090812, Shanghai Ling Feng chemical reagent company limited), hydroxyl succinic acid (Malic acid, lot number 20090328, the emerging biochemical reagents company limited of China's favour), citric acid (Citric acid monohydrate, lot number 081245071, Nanjing Chemistry Reagent Co., Ltd.), linoleic acid (Linoleic acid, lot number 10129622, Alfa Aesar), salicylic acid (Salicylic acid, lot number 060222, Shanghai green grass or young crops is analysed Chemical Industry Science Co., Ltd).
Methanol is the HPLC level, and sulphuric acid, chloroform, anhydrous sodium sulfate etc. are analytical pure, and water is sub-boiling distillation water (self-control).
The organic acid position by weight 1: 1 mix homogeneously, adds the water extraction 3～5h of 5～8 times of weight with the Rhizoma Pinelliae and Rhizoma Zingiberis Recens, filter extracting solution, medicinal residues continue to add the water of 3～5 times of weight, extract 2～3h, merge extracted twice liquid; Extracting solution add ethanol to pure mass content be 60～80%, leave standstill 8～12h after, 4500 commentaries on classics/min are centrifugal, get supernatant, are concentrated into 1～2g crude drug mL
-1, concentrated solution obtains ether layer and water layer with twice of extracted with diethyl ether (each is 0.8～1 times of volume of concentrated solution with the ether amount), water layer is transferred to pH7 with NaOH, after solution is clarified substantially, adsorb through anion exchange resin, wash assorted to colourless, with 0.5～1molL again with deionized water
-1NaOH eluant solution resin, get eluent, this eluent transfers to pH3 with HCl, is evaporated to driedly, dissolves 3 times repeatedly to remove NaCl with dehydrated alcohol again, obtains the organic acid position.
2 methods and result
2.1 chromatographic condition and system suitability
Capillary chromatographic column: HP-5 quartz capillary column (30m * 0.53mm * 2.65 μ m); Carrier gas: nitrogen, purity 〉=99.999%; Column temperature: temperature programming, 100 ℃ kept 3 minutes, then with 5 ℃ of min
-1Be warming up to 220 ℃, keep 5min, at last with 10 ℃ of min
-1Be warming up to 280 ℃, keep 5min; Advance 250 ℃ of ocean mouthful temperature; 290 ℃ of detector temperatures; Carrier gas: nitrogen; Flow velocity: 1.0mLmin
-1Sample size: 1 μ L.By above-mentioned chromatographic condition, to analyzing by the mixing reference substance solution for preparing under the 2.2.2 item, four kinds of organic acid and internal standard substance can separate well, and chromatogram is seen Fig. 1.
2.2 the preparation of reference substance solution, need testing solution and negative sample solution
2.2.1 mix the preparation of reference substance stock solution: precision takes by weighing succinic acid 250.05mg, hydroxyl succinic acid 1.75010g respectively, citric acid 2.50165g, linoleic acid 76.62mg place the brown measuring bottle of same 50mL, add dissolve with methanol and be diluted to scale, shake up, namely get and mix the reference substance stock solution.
2.2.2 mix the preparation of reference substance solution: the accurate absorption mixed reference substance stock solution 0.1,0.3,0.5,0.7,0.9,11 respectively, and 1.3mL places the 50mL reaction bulb, adds salicylic acid (50.1824mgmL
-1) 0.5mL, add sulphuric acid-methanol solution 10mL of 7: 100, shake up, place 60 ℃ of water-bath heating 24h esterification, transfer in the separatory funnel that fills 10mL water chloroform extraction three times, each 6mL, get chloroform layer, place the 25mL measuring bottle, add chloroform and be settled to scale, shake up, behind anhydrous sodium sulfate drying, namely get 7 series and mix reference substance solution.
2.2.3 the about 72.0mg in organic acid position is got in the preparation of need testing solution, the accurate title, decide, and places the 50mL reaction bulb, adds salicylic acid (50.1824mgmL
-1) 0.5mL, with operating under the 2.2.2 item, namely get need testing solution.
2.2.4 the preparation of negative sample solution preparation lacks the organic acid position of the Rhizoma Pinelliae, with operating under the 2.2.3 item, namely gets negative sample solution.
2.4 the preparation of standard correction curve
Accurate each 1 μ L of mixing reference substance solution that draws above-mentioned 7 series tests by the chromatographic condition under 2.1, and the record peak area is vertical coordinate with reference substance and interior mark peak area ratio Y, is abscissa with mass concentration X, carries out linear regression, gets regression equation.The results are shown in Table 2.As seen this method is good in scope of experiment internal linear relation.
Each component linear relationship measurement result of table 2
2.6 precision test
Get the about 72.0mg in organic acid position, the accurate title, decide, place the 50mL reaction bulb, according to the operation under " 2.2.3 " item, sample size 1 μ L, continuous sample introduction 6 times, calculate precision, succinic acid, hydroxyl succinic acid, citric acid, linoleic RSD value are respectively 1.86%, 1.82% as a result, 1.66%, 1.81%.
2.7 replica test
Get the about 72.0mg in organic acid position, the accurate title, decide, and parallel 6 parts, according to the operation under " 2.2.3 " item, sample size 1 μ L, inner mark method ration, the repeatability of computational methods.The every 1mL Small Pinelliae Decoction of result preparation contains succinic acid, hydroxyl succinic acid, citric acid, linoleic amount and is respectively 3.29%, 24.82%, 34.61%, 1.06%, RSD and is respectively 1.92%, 1.73%, 1.82%, 1.96%.
2.8 stability test
Get the about 72.0mg in organic acid position, accurate claim fixed, according to " 2.2.3 " operation down, respectively after preparation 0,4,8,12,16,20,24h sample introduction 1 μ L, analyze, investigate stable.Succinic acid, hydroxyl succinic acid, citric acid, linoleic RSD are respectively 1.26%, 1.62%, 1.82%, 1.95% as a result.Show that need testing solution keeps stable in 24h.
2.9 average recovery test
Get the about 36.0mg in organic acid position, the accurate title, decided parallel 6 parts, add respectively and mix reference substance stock solution 0.25mL, reclaim liquid according to preparation application of sample under the 2.2.3 item, measure content, calculate average average recovery, succinic acid, hydroxyl succinic acid, citric acid, linoleic average average recovery are respectively 99.96%, 99.95% as a result, 100.61%, 99.76%, RSD is respectively 1.38%, 1.47%, 1.40%, 1.04%.See Table 3.
Table 3 determination of recovery rates is .n=6 as a result
2.10 sample size is measured
Get the organic acid position of three lot numbers, according to " 2.2.3 " operation down, make need testing solution, get 1 μ L sample introduction, calculate content with internal standard method, parallel three parts of each lot number, every part of sample introduction 2 times.The results are shown in Table 4.
The measurement result of several organic acid contents (n=6) in table 4 Small Pinelliae Decoction
Experimental example 3: effective site of the present invention is to the pharmacological action of vomiting treatment
One, to the influence of pigeon vomiting due to copper sulfate and the cisplatin
1. experiment material
1.1 sample, 1. the active site that makes of embodiment 1 is the organic acid position, faces the time spent to be mixed with certain density solution.Compound method: get organic acid extractum or an amount of, precision claims fixed, adds a certain amount of 0.5%CMC-Na, and ultrasonic 10min stirs, till abundant suspendible; 2. metoclopramide; 3. ondansetron.
1.2 reagent, copper sulfate (analytical pure, Nanjing chemical reagent factory); Normal saline (Nanjing Xiaoying Pharmaceutical Factory); (Jinan, Shandong Qilu Pharmaceutical Co., Ltd. produces cisplatin for inj (freeze-dried type), and 20mg/ props up, lot number: 6040151), face the time spent to be mixed with 1.0mgmL with physiological saline solution
-1Solution; CMC-Na, analytical pure.
1.3 animal, the healthy adult pigeon, the male and female dual-purpose, 450～500g, animal reproduction field, Jiangning District Green Dragon mountain, Nanjing provides.
2. method and result
Healthy pigeon, male and female half and half, by 8 one group, random packet.By dosage gastric infusion in the table, model group gives 0.5%CMC-Na every day 2 times, successive administration 3 days., positive controls gives metoclopramide 1.0mgkg
-1, for three days on end.Fasting 12h before the last administration, behind the administration 1h, every Columba livia gavages 2% copper sulfate 0.2gKg
-1Every pigeon the time (vomiting incubation period) of the 1st vomiting occurred and gives copper sulfate 1.5h interior vomiting number of times after copper sulfate given in record.The results are shown in Table 5.
Other gets healthy pigeon, by 8 one group, and random packet.By dosage gastric infusion in the table, model group gives 0.5%CMC-Na, every day 2 times, successive administration 3 days.Intravenous injection ondansetron 0.3mgkg under the every pigeon wing of positive controls
-1, for three days on end.Fasting 12h before the last administration, behind the administration 30min, intravenous injection cisplatin 1.0mgKg under every pigeon wing
-1The time (vomiting incubation period) of the 1st vomiting and the vomiting number of times in the injection cisplatin 6h appear in every pigeon behind the record injection cisplatin.The results are shown in Table 6.
Table 5 organic acid position is to the influence of pigeon vomiting due to the copper sulfate
Annotate: adopt variance analysis to compare in twos: compare with model group,
*P＜0.05.
Table 6 organic acid position is to the influence of pigeon vomiting due to the cisplatin
Annotate: adopt variance analysis to compare in twos, compare with model group,
*P＜0.05.
The result shows that the high low dose group at organic acid position all can obviously reduce the pigeon vomiting number of times due to copper sulfate and the cisplatin, and the preventing or arresting vomiting effect is obvious.Compare with model group, significant difference is arranged.
Two, to the influence of normal mouse gastric emptying
1. experiment material
1.1 sample, 1. organic acid position is faced the time spent to be mixed with certain density solution.Compound method is the same.2. Small Pinelliae Decoction is got Rhizoma Zingiberis Recens and Rhizoma Pinelliae equivalent, adds 5 times of water gagings and decocts 30min, filters, and residue adds 4 times of water gagings again and fries in shallow oil 30min.Merge filtrate twice, be concentrated into 2g crude drug mL
-1
1.2 reagent, phenol red (Shanghai chemical reagent purchasing and supply station, 86-12-17), NaHCO
3, CMC-Na, be analytical pure.
1.3 animal, ICR mice, body weight 18～22g, male and female dual-purpose, Shanghai Slac Experimental Animal Co., Ltd. (SCXK (Shanghai) 2007-0005).
1.4 instrument: SPECTRA MAX 190 UV, visible light microwell plate detectors (U.S. MOLECULAR DEVICES); GS-15R type tabletop refrigerated centrifuge (U.S. BECKMAN company); PHS series pH meter (Lida Instrument Factory, Shanghai).
2. method and result
Only get healthy mice, male and female half and half, random packet, 10 every group.Each group respectively gastric infusion or etc. capacity 0.5%CMC-Na solution (10mlkg
-1), every day twice, for three days on end, water 12h is can't help in fasting before administration in the 4th day.Behind the 40min, every mouse stomach 0.04% phenol red solution 0.2ml puts to death behind the 15min, gets the small beaker that stomach places the 5ml distilled water, cuts off stomach with eye scissors along greater gastric curvature, and gastric content is fully washed in distilled water, uses NaHCO after the administration
3Solution is regulated pH value to 6.0～6.5, gets supernatant with 12000rmin
-1Centrifugal 3 minutes, the accurate supernatant 200 μ l that draw measured light absorption value (wavelength 560nm) with microplate reader in 96 orifice plates.The light absorption value that records is phenol red light absorption value residual in the stomach.The height of phenol red residual rate can reflect the speed of gastric emptying.The results are shown in Table 7.
Stomach residual rate (%)=(the phenol red optical density of phenol red optical density/benchmark in the stomach) * 100%
Table 7 organic acid position is to the influence of normal mouse gastric emptying
Annotate: adopt variance analysis to compare in twos, compare with the normal saline group,
*P＜0.05.
The result shows, extracts the organic acid position that obtains from Small Pinelliae Decoction, can obviously suppress the gastric emptying of normal mouse, compares with the normal control product, and significant difference is arranged; Compare organic acid position unknown significance difference with classical preventing or arresting vomiting side Small Pinelliae Decoction.Illustrate that the organic acid position is identical with the action effect of Small Pinelliae Decoction.
Claims (8)
1. the effective site of Small Pinelliae Decoction is characterized in that, the physicochemical property of this effective site is: yellowish-brown extractum, have certain moisture, and dissolve in hot water and ethanol; This effective site mainly contains the organic acid composition, and wherein, succinic acid content is 3.0～3.8%, hydroxyl succinic acid content is 21.6～26.8%, citric acid content is 30.6～38.6%, linoleic content is 0.92～1.26%;
This effective site prepares as follows: with the Rhizoma Pinelliae and Rhizoma Zingiberis Recens by weight the 1:1 mix homogeneously, extracting in water 3～5h, filter extracting solution, medicinal residues continue to add water, extract 2～3h, merge extracted twice liquid; Extracting solution add ethanol to pure quality percentage composition be 60～80%, leave standstill 8～12h after, centrifugal, get supernatant, be concentrated into 1～2g crude drug mL
-1The concentrated solution extracted with diethyl ether obtains ether layer and water layer, and water layer is transferred to pH7 with NaOH solution, through anion exchange resin, wash with deionized water again and mix to colourless, with NaOH eluant solution resin, get eluent, this eluent transfers to pH3 with HCl, be evaporated to dried, again with anhydrous alcohol solution to remove NaCl, obtain the organic acid position namely.
2. the preparation method of the effective site of the described Small Pinelliae Decoction of claim 1 is characterized in that, with the Rhizoma Pinelliae and Rhizoma Zingiberis Recens by weight the 1:1 mix homogeneously, extracting in water 3～5h, filter extracting solution, medicinal residues continue to add water, extract 2～3h, merge extracted twice liquid; Extracting solution add ethanol to pure quality percentage composition be 60～80%, leave standstill 8～12h after, centrifugal, get supernatant, be concentrated into 1～2g crude drug mL
-1The concentrated solution extracted with diethyl ether obtains ether layer and water layer, and water layer is transferred to pH7 with NaOH solution, through anion exchange resin, wash with deionized water again and mix to colourless, with NaOH eluant solution resin, get eluent, this eluent transfers to pH3 with HCl, be evaporated to dried, again with anhydrous alcohol solution to remove NaCl, obtain the organic acid position namely.
3. the preparation method of the effective site of Small Pinelliae Decoction according to claim 2 is characterized in that, during twice extracting in water, for the first time the weight of water is the Rhizoma Pinelliae and Rhizoma Zingiberis Recens gross weight 5 ~ 8 times, and the weight of water is the Rhizoma Pinelliae and Rhizoma Zingiberis Recens gross weight 3 ~ 5 times for the second time.
4. the preparation method of the effective site of Small Pinelliae Decoction according to claim 2 is characterized in that, during extracted with diethyl ether, and extracting twice, the volume of twice used ether is 0.8～1 times of amount of concentrated solution volume.
5. the preparation method of the effective site of Small Pinelliae Decoction according to claim 2 is characterized in that, ether layer concentrates when reclaiming ether, adopts the water-bath normal pressure, and bath temperature is 39～49 ℃.
6. the preparation method of the effective site of Small Pinelliae Decoction according to claim 2 is characterized in that, described anion exchange resin is D315 type anion exchange resin.
7. the preparation method of the effective site of Small Pinelliae Decoction according to claim 2 is characterized in that, the used NaOH solution concentration of eluting resin is 0.5 ~ 1molL
-1
8. the preparation method of the effective site of Small Pinelliae Decoction according to claim 2 is characterized in that, is evaporated to when doing, and thickening temperature is 60～70 ℃.
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| 半夏及不同炮制品中微量元素分析;杨玉琴等;《微量元素与健康研究》;20020626;第19卷(第02期);33 * |
| 半夏药材中总有机酸的定量方法和含量测定研究;许风清等;《南京中医药大学学报(自然科学版)》;20050220;第21卷(第01期);34-36 * |
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