CN1201805C - Combination of medication for reducing poison and synergic action in radiotherapy or chemotherapy as well as its preparing method - Google Patents
Combination of medication for reducing poison and synergic action in radiotherapy or chemotherapy as well as its preparing method Download PDFInfo
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Abstract
The present invention relates to a medical composition cooperated with radiotherapy or chemotherapy for increasing immunity, attenuating toxicity and increasing efficiency, which comprises the components of the proportion by weight: 1 to 40 parts of astragalus root, 1 to 15 parts of matrimony vine and 1 to 15 parts of ganoderma. The Chinese medical oral preparation with the advantages of conspicuous curative effect, stabilization and controllable quality is prepared by optimizing a preparation technology for enriching effective part groups.
Description
Technical field
The present invention relates to a kind of medicine that is used for the chemicotherapy attenuation synergistic and preparation method thereof, belong to the field of Chinese medicines.
Background technology
Malignant tumor is China's commonly encountered diseases and frequently-occurring disease.All antitumor drug nearly all belong to cytotoxic category at present, and cancer therapy drug also damages seriously normal structure in killing tumor cell, and this toxic and side effects that can't avoid has limited the application of cancer therapy drug and the performance of curative effect.The common medicine that is used for the chemicotherapy attenuation synergistic has ribonucleic acid, Chinese patent medicine that ZHENQI FUZHENG KELI, krestin capsule, smart Rhizoma Polygonati oral liquid (the accurate word of medicine (1993 No. 001612) is defended in Shan) etc. are arranged on the market a few days ago.The preparation process that this class Chinese patent medicine has is simple, and effective ingredient is indeterminate, and the action target spot that has disperses, thereby affect the treatment etc.Have data to show, be used for tumor radiotherapy, chemotherapy rise that white effect is obvious, toxicity is little and medicine that can platelet increasing seldom, the weixuening granule is a national Chinese medicine protection kind unique in the similar medicine.
Summary of the invention
The technical issues that need to address of the present invention are, how according to Chinese medical theory, the pharmaceutical composition with synergism and attenuation is used in development when chemicotherapy, simultaneously by optimizing preparation process, enrichment effective part group, be prepared into stable curative effect, quality controllable Chinese medicine preparation.
For addressing the above problem, the invention provides following technical proposals.
A kind of pharmaceutical composition that immunoregulation effect is arranged, it is characterized in that it is made by following bulk drugs basically: the Radix Astragali, Fructus Lycii, Ganoderma are 1-40 part: 1-15 part: 1-15 part.Wherein the consumption preferably of crude drug is: the Radix Astragali, Fructus Lycii, Ganoderma weight ratio are 1-30 part: 1-12 part: 1-12 part.Wherein the even more ideal consumption of crude drug is: the Radix Astragali, Fructus Lycii, Ganoderma weight ratio are 30 parts: 12 parts: 12 parts.The sweet temperature of the Radix Astragali wherein, strongly invigorating primordial QI is monarch.Fructus Lycii is sweet flat, nourishing the liver and kidney, and Ganoderma is sweet flat, and tonify deficiency lung benefiting, two medicines are minister altogether.All medicines share, and play the merit of supplementing QI and nourishing YIN, nourishing the liver and kidney altogether.
The preparation method of aforementioned pharmaceutical compositions comprises the following steps:
A) get the crude drug Radix Astragali, Fructus Lycii and Ganoderma, standby;
B) with the Radix Astragali of described weight proportion, with 70～90% ethanol extractions 1～4 time, each 1～2.5 hour, merge ethanol extract, reclaim ethanol, with removing macroporous resin column on the liquid of post precipitation, with 55～84% ethanol elutions, must Radix Astragali total saponins;
C) Fructus Lycii, Ganoderma and the above-mentioned ethanol extraction of described weight proportion are crossed Radix Astragali residue and merged, add 10～14 times of water gagings, merceration or decoction are extracted 2～4 times, each 1～2 hour, merge decoction liquor, be concentrated into relative density 1.0-1.4, adding ethanol makes and contains the alcohol amount and reach 70-85%, filter precipitate, dry sediment is after being dissolved in water, water liquid is concentrated into relative density 1.0-1.4, add ethanol and make and contain the alcohol amount and reach 70-80%, leaching precipitate, dry total polysaccharides;
D) Radix Astragali total saponins is mixed with total polysaccharides, get the active component of medicine of the present invention.This active component can be mixed with conventional adjuvant on the pharmaceutical formulations, also can not add conventional adjuvant, makes the conventional medicine preparation.
Pharmaceutical composition of the present invention preparation method preferably is:
B) with the Radix Astragali of described weight proportion, with 75～85% ethanol extractions 2～3 times, each 1.5～2.0 hours, merge ethanol extract, reclaim ethanol, with AB-8 and/or D101 model macroporous resin column on the liquid of removal post precipitation, with 60～80% ethanol elutions, get Radix Astragali total saponins;
C) Fructus Lycii, Ganoderma and the above-mentioned ethanol extraction of described weight proportion are crossed Radix Astragali residue and merged, add 11～13 times of water gagings, decoct 3 times, each 1.5 hours, merge decoction liquor, relative density 1.1-1.3 when being concentrated into 80 ℃, adding ethanol makes and contains the alcohol amount and reach 75-80%, filter precipitate, dry sediment is after being dissolved in water, relative density 1.1-1.3 when water liquid is concentrated into 80 ℃, add ethanol and make and contain the alcohol amount and reach 75-80%, leaching precipitate, dry total polysaccharides;
D) Radix Astragali total saponins is mixed with total polysaccharides, add pharmaceutic adjuvant, routine is made oral formulations.
The comparatively ideal technical characterictic of described preparation method is:
B) with the Radix Astragali of described weight proportion, with 80～82% ethanol extractions 3 times, each 1.5 hours, merge ethanol extract, reclaim ethanol, with removing AB-8 or D101 model macroporous resin column on the liquid of post precipitation, with 65～70% ethanol elutions, must Radix Astragali total saponins;
C) Fructus Lycii, Ganoderma and the above-mentioned ethanol extraction of described weight proportion being crossed Radix Astragali residue merges, adding 12 times of water gagings decocts, relative density 1.2 when decoction liquor was concentrated into 80 ℃, added ethanol and made and contain the alcohol amount and reach 80%, filter precipitate, dry sediment, after being dissolved in water, relative density 1.2 when water liquid was concentrated into 80 ℃, added ethanol and made and contain the alcohol amount and reach 75%, the leaching precipitate, the dry total polysaccharides that gets;
D) Radix Astragali total saponins, total polysaccharides are mixed with conventional adjuvant pharmaceutically, make capsule, tablet, powder, granule or oral liquid.
In the above-mentioned preparation method, it is characterized in that:
C) the water intaking decoction liquor concentrates precipitate with ethanol, filter behind the precipitate, taking precipitate is dissolved in water, and filters, and operates routinely with film and carries out ultrafiltration, remove macromole impurity with 300,000 molecular cut off membrane ultrafiltration earlier, and then remove moisture and part small molecular weight impurity with 1000 molecular cut off membrane ultrafiltration, and be concentrated into relative density 1.2, add ethanol and make and contain the alcohol amount and reach 75%, the leaching precipitation gets total polysaccharides.
Oral formulations adjuvant commonly used can select for use, and disintegrating agent is as hydroxypropyl starch, hyprolose, carboxymethyl starch sodium, carboxymethylcellulose calcium, polyvinylpolypyrrolidone, cross-linking sodium carboxymethyl cellulose etc.; Filler is as lactose, sucrose, mannitol, microcrystalline Cellulose, dextrin, starch, calcium phosphate, calcium hydrogen phosphate, calcium sulfate, calcium carbonate, cyclodextrin, micropowder cellulose etc.; Wetting agent and binding agent are as ethanol, pregelatinized Starch, polyvidone, sodium carboxymethyl cellulose, hypromellose; Lubricant is as Pulvis Talci, stearic acid, magnesium stearate, calcium stearate, micropowder silica gel, hydrogenated vegetable oil, Macrogol 4000 and 6000: wetting agent is as sodium lauryl sulphate, Tween 80.
Pharmaceutical composition of the present invention, the application in preparation raising immune drug.
Pharmaceutical composition of the present invention is used for the application of the medicine of chemicotherapy attenuation synergistic in preparation.
Pharmaceutical composition of the present invention is preparing the raising immunity of share with chemicotherapy, the application in the attenuation synergistic medicine.
After the crude drug prescription of pharmaceutical composition of the present invention forms, be the basis of the modern preparation of the preparation Chinese medicine that curative effect is accurate, component is clear and definite, quality controllable to the screening of effective site.
Lycium barbarum polysaccharide in the pharmaceutical composition of the present invention and astragalus polysaccharides can both be in external enhancing LAK activity.Mouse peritoneal injection lycium barbarum polysaccharide 5g/L (7d) can significantly strengthen the killing ability of NK cell, and can partly resist the inhibitory action of cyclophosphamide to NK cells in mice.Chemical constituents such as the polysaccharide of Ganoderma, triterpenes, amino acid polypeptide class, adenosine, immune system, nervous system, E﹠M system, cardiovascular system etc. had regulating action, a kind of Gac-D triterpenic acid in the Ganoderma has the effect that is similar to paclitaxel, can make the cancerous cell cycle divide a word with a hyphen at the end of a line blocking-up, the overgrowing of blocking-up cancerous cell in the G2+M phase.Deep research shows that also ganoderan has good immunoregulation effect.And the ganoderan class is by combining with triterpene, can possess immunity identification and cell toxicant dual function, can optionally suppress tumor cell, guarantees that normal cell is without prejudice, can be comprehensively, the active adjustment body's immunity.With the chemicotherapy coupling, pharmaceutical composition of the present invention is that ganoderan, lycium barbarum polysaccharide and astragalus polysaccharides and astragaloside share, and has remarkable Synergistic attenuation potentiation, is in particular in cyclophosphamide is suppressed murine sarcoma S
180Remarkable synergism and attenuation is arranged, right
60The Co irradiation suppresses murine sarcoma S
180Tangible synergism and attenuation is arranged, normal mouse is had tangible Attenuation through the cyclophosphamide chemotherapy, right
60The caused toxic and side effects of Co radiotherapy has tangible attenuation efficacy effect, and the cellular immunization and the humoral immune function of tumor mice all had obvious potentiation, thereby has really realized " zero poisons treatment ".
One, effective site screening experiment
Investigate the difference of different extraction conditions and the pharmacologically active of different extract parts with pharmacodynamics test.Carry out the pharmacologically active screening of the different extract parts of the Radix Astragali with the test of normal mouse chemotherapy, different methods of extraction is to the influence of lycium barbarum polysaccharide pharmacodynamics simultaneously.The result shows that the Saponin in the Radix Astragali is an active site, and does not have obvious pharmacologically active by the aminoacid that the Radix Astragali extracts; Different extraction temperature does not have obvious influence to the pharmacologically active of lycium barbarum polysaccharide.
Animal: Kunming kind white mice, ♀ ♂ half and half, regular grade is purchased in animal reproduction field, Green Dragon mountain, Jiangning, Nanjing, the quality certification number: SCXK (Soviet Union) 2001-0014.Feeding environment is a regular grade, the quality certification number: SYXK (Soviet Union) 2001-0008.
Astragalus polysaccharides (medicinal liquid): contain crude drug 1g/ml, lot number: 20010609.Astragaloside (medicinal liquid); Contain crude drug 1g/ml, lot number: 20010426.Radix Astragali aminoacid (medicinal liquid): contain crude drug 1g/ml, lot number: 20010429, Chinese medicine preparation research department in Jiangsu Prov. Research Inst. Traditional Chinese Medical provides, and the time spent adds the suspension that an amount of distilled water is made into suitable concentration.
Lycium barbarum polysaccharide I (medicinal liquid): be the cold extraction gained, contain raw medicinal herbs 1g/ml, lot number: 20010425; Lycium barbarum polysaccharide II (medicinal liquid): carry gained for heat, contain raw medicinal herbs 1g/ml, lot number: 20010411, Chinese medicine preparation research department in Jiangsu Prov. Research Inst. Traditional Chinese Medical provides, and the time spent adds the suspension that an amount of distilled water is made into suitable concentration.
Cyclophosphamide for injection: 0.2g/ props up, and the accurate word (1996) of medicine is defended No. 403501 in the Shandong, lot number: 01010310, and Hengrui Medicine Co., Ltd., Jiangsu Prov. produces, and the time spent adds the medicinal liquid that an amount of physiological saline solution is made into desired concn.
The pharmacologically active of the different extract parts of the Radix Astragali relatively
50 of Kunming mouses, male and female half and half, body weight 18～22 grams, being divided into 5 groups at random is normal control group, cyclophosphamide chemotherapy group, astragalus polysaccharides group, Saponin group and aminoacid group.Astragalus polysaccharides, Saponin and aminoacid group are all irritated the respective sample that stomach is equivalent to raw medicinal herbs 11.0g/kg, and normal control and cyclophosphamide chemotherapy group filling stomach give the distilled water with volume, the equal gastric infusion of mice 10 days.Except that the normal control group, all the other mices all began in administration on the 8th day, when the filling stomach gives water or medicinal liquid, and intraperitoneal injection of cyclophosphamide every day (Cy) 40mg/kg, for three days on end.Behind last injection Cy 24 hours, each treated animal eye socket rear vein beard 30 μ l that take a blood sample injected diluent, and the homemade full-automatic animal blood counting instrument of usage is measured peripheral hemogram.
The result shows: normal mouse is after the cyclophosphamide chemotherapy, quantity of leucocyte significantly descends in the peripheral blood, compare with the cyclophosphamide group, the animal quantity of leucocyte that gives the treatment of astragalus polysaccharides and astragaloside all has obvious recovery (P＜0.05), and the murine interleukin quantity and the cyclophosphamide chemotherapy group that give the treatment of Radix Astragali aminoacid compare there was no significant difference, this quantity of leucocyte that shows that astragalus polysaccharides and astragaloside cause chemotherapeutic reduces significant improvement effect, and Radix Astragali aminoacid does not then have tangible leukogenic effect.
The lycium barbarum polysaccharide of Different Extraction Method gained is to the influence of normal mouse Cy peripheral blood after chemotherapy elephant
Experiment shows, compare with the cyclophosphamide chemotherapy group, the murine interleukin number average that gives lycium barbarum polysaccharide I and lycium barbarum polysaccharide II obviously raise (P＜0.05), and the hemoglobin number average of two treated animals increases (P＜0.01～0.001) to some extent than chemotherapy group, peripheral hemogram between these two groups is no significant difference then, show that the Fructus Lycii total polysaccharides of mainly being made up of lycium barbarum polysaccharide I and II all has significant improvement effect to the quantity of leucocyte minimizing that chemotherapy causes, prove that the different extraction temperature of lycium barbarum polysaccharide do not have obvious influence to its pharmacologically active.
The Radix Astragali total saponins and the Radix Astragali, Fructus Lycii and after the Ganoderma total polysaccharides is mixed, under the chemicotherapy situation, to improve laboratory animal such as immune function of mice, attenuation synergistic has remarkable effect.
Two, preparation of drug combination method of the present invention research
Milkvetch Root can adopt different concentration ethanol merceration or backflow and/or Soxhlet to extract with alcohol reflux; Can leave standstill during precipitation, the method for leaving standstill can leave standstill and/or room temperature leaves standstill with cold preservation; Separation method can adopt centrifugalize, mainly is to remove impurity and cold preservation standing separation, mainly is to remove the come-up oils and fats.During the resin column purifies and separates, can routinely macroporous resin be carried out pretreatment in advance.Reclaiming ethanol in the preparation method of the present invention can adopt normal pressure to reclaim and/or reclaim under reduced pressure.During dry isolating precipitation, can adopt oven drying, vacuum drying, lyophilization and/or spray drying.Water extraction such as the Radix Astragali can be used merceration and/or decoction.In the preparation method of the present invention, all when relating to concentration technology, can adopt concentrating under reduced pressure, normal pressure to concentrate and/or thin film evaporation concentrates.
All when relating to precipitation or precipitate and fluid separation applications in the inventive method, can adopt centrifugalize, membrance separation and/or normal pressure or filtration under diminished pressure.Pulverizing can be adopted independent pulverizing and/or mix and pulverize.Granulation can be adopted wet granulation and/or dry granulation.
To influencing the principal element of (+)-Astragenol reflux, extract: concentration of alcohol carries out orthogonal test by four factors, three horizontal L (3).Result of the test shows that the principal element that influences Radix Astragali total saponins is an extraction time; The principal element that influences astragaloside is alcoholic acid percentage concentration, is extraction time secondly, and solvent amount and extraction time influence are less.Comprehensive three, the many persons of total extract, it is also more promptly to bring impurity into, adds to the difficulties for next step purification, has selected invention preparation of drug combination method according to the result of analysis-by-synthesis.Because for the first time medical material is a dry product during alcohol reflux, itself will absorb the alcohol of about 2 times of amounts, and therefore alcohol adding amount is all added 2 times on the alcohol amount of basis for the first time.
The extracting solution decompression recycling ethanol, centrifugal or leave standstill and remove the come-up oils and fats, AB-8 resin and/or D101 and/or D605 and/or D140 and/or NKA II purification by macroporous resin, ethanol elution, the eluent decompression recycling ethanol, drying, Radix Astragali total saponins.The results are shown in Table 1.
Radix Astragali total saponins extract usefulness AB-8 of the present invention and/or D101 and/or D605 and/or D140 and/or NKA II macroporous resin isolation and purification method are than traditional water alcohol method relatively, Radix Astragali total saponins content is higher in the sample that the present invention makes, and total saponin content accounts for total solid ratio height, show that AB-8 macroporous resin content is bigger, show the ratio height that has kept effective ingredient when removing impurity with the purification by macroporous resin method.With traditional handicraft be water alcohol method relatively, the content of total saponins improves 5 times as a result, the ratio that total saponins accounts for total solid matters is increased to more than 50% by original 2.3%.The results are shown in Table 2.
In the total saponins desorption process, higher with 85～55% ethanol elution total saponin contents, better with 75～60% ethanol elutions, best with 65% ethanol elution, total saponin content is 56.6% in its total extract, is higher than other concentration.The results are shown in Table 2.
To influencing the factor of the Radix Astragali, Fructus Lycii, the extraction of Ganoderma three medicine polysaccharide: amount of water, medical material soak time, extraction time are by four factors, three horizontal L
9(3
4) carry out orthogonal test.Routinely, the decocting condition of boiling of Chinese medicine is designed to by-level, gets two levels of height respectively again.Test shows that the principal element that influences the polysaccharide extraction is decocting time, and all to decoct three times, each 1.5 hours is good.The results are shown in Table 3.
Table 1 water alcohol method and purification by macroporous resin method sample size are relatively
In the total extraction of purification process total saponin content total extract
Mg/g mg/g total saponin content (%)
AB-8 73.3 10.5 69.8
Water alcohol method 14.7 63.0 2.3
Table 2 total saponins alcohol desorption concentration ratio
In the total extraction of concentration of alcohol % total saponin content total extract
Mg/g mg/g total saponin content (%)
35 1.5029 6.18 24.3
45 3.3896 7.09 47.8
55 4.2278 8.90 47.5
65 6.1688 10.09 56.6
75 5.7850 11.18 51.7
85 5.2486 10.09 52.0
95 3.5428 9.27 38.1
Table 3 water extraction orthogonal test
Total polyoses content that extracts
Sequence A B C D total polysaccharides %
Thing % mg/g crude drug
1 1 1 1 1 25.37 3.04 13.9704
2 1 2 2 2 33.36 9.24 50.5749
3 1 3 3 3 36.64 11.34 64.0512
4 2 1 2 3 34.52 10.05 58.6099
5 2 2 3 1 27.00 3.99 23.2983
6 2 3 1 2 33.98 9.43 50.0669
7 3 1 3 2 32.00 10.28 57.5368
8 3 2 1 3 35.33 8.75 49.7130
9 3 3 2 1 27.73 7.22 39.7752
Total K
195.37 91.89 94.68 80.10
Carry K
295.50 95.69 95.61 99.34
Get K
395.06 98.35 95.64 106.49
Thing R * 3 0.44 6.46 0.96 26.39
Total K
123.62 23.37 21.22 14.25
Many K
223.47 21.98 26.51 28.95
Sugar k
326.25 27.99 25.61 30.14
R×3 2.78 6.01 4.39 15.89
Many K
1128.5965 130.1171 113.753 77.0439
Sugar K
2131.9751 125.5862 148.960 158.1786
Contain K
3147.0250 153.8933 144.883 172.3741
Amount R * 3 18.4285 28.3071 31.1360 95.3302
Three, invention compositions main pharmacodynamics research data
1, cyclophosphamide is suppressed murine sarcoma S
180Synergism and attenuation
Pharmaceutical composition of the present invention is by 10.0,5.0 and 2.5g crude drug/kg gastric infusion, to S
180Tumor-bearing mice has tangible rising effect through caused leucocytes reduction of cyclophosphamide chemotherapy and marrow nucleated cell decreasing, caused spleen of chemotherapy and thymus index decline there is significant improvement effect, share its tumour inhibiting rate that significantly to raise with cyclophosphamide, show that this product suppresses murine sarcoma S to cyclophosphamide
180Tangible synergism and attenuation is arranged.
The present composition (dry powder): contain crude drug 37.0g/g, preparation method is seen embodiment 1.
The weixuening granule: the 8g/ bag, No. the 165801st, the accurate word (1993) of Su Wei medicine, lot number: 020712, strong people pharmaceutical factory of Changzhou Pharmaceutical limited company produces, and the time spent adds the suspension that an amount of distilled water is made into suitable concentration.(down together)
Cyclophosphamide for injection (being called for short Cy, down together): 0.2g/ props up, and the accurate word (1996) of medicine is defended No. 403501 in the Shandong, lot number: 02080421, and Hengrui Medicine Co., Ltd., Jiangsu Prov. produces, and the time spent adds the medicinal liquid that an amount of physiological saline solution is made into desired concn.(down together)
Tumor strain: sarcoma S
180Solid type moves from the institute of oncology, Jiangsu Province, is gone down to posterity to protect by Jiangsu Prov. Research Inst. Traditional Chinese Medical pharmacological room and plants.(down together)
The experiment detailed method is referring to new Chinese medicine pharmaceutical research guide. the pharmacodynamic study of treatment malignant tumor Chinese medicine. and Ministry of Public Health bureau of drug administration " study of tcm new drug guide ", 1994:104; Leaf should be lovely, Wang Yu is third-class. national Clinical Laboratory rule of operation. and the .1991:4 of publishing house of Southeast China University, 64; Wan Junmei, Zhang Yongzhong, Wu Wei etc. press down the experimentation that the tumor pill for curing improves chemotherapeutic efficacy. Chinese patent medicine, 2002,24 (9): 695～697
2, cyclophosphamide is suppressed murine sarcoma S
180Tumor heavily reaches the influence of tumour inhibiting rate
60 of Male Kunming strain mice, body weight 18-22 gram, being divided into 6 groups is the high, medium and low dosage group of lotus tumor matched group, cyclophosphamide chemotherapy group, weixuening groups of grains and the present composition.The administration group is irritated stomach respectively and is given present composition suspension 10.0,5.0 and 2.5g crude drug/kg, and the positive drug group is irritated stomach and given weixuening granule 4.8g/kg, and lotus tumor matched group filling stomach gives the distilled water with volume, the equal gastric infusion of mice 12 days.And, get abdominal part inoculation S in the 4th day of administration
1808 days tumor-bearing mice of tumor strain, the sterile working extracts down abdominal cavity tumor liquid, and (microscopy contains oncocyte several 5 * 10 to be diluted to the oncocyte liquid of 1: 4 concentration with sterile saline
7/ ml), in all mice right fore armpit subcutaneous vaccination 0.2ml/ Mus.Except that lotus tumor matched group, all the other mices are next day behind inoculated tumour all, when the filling stomach gives water or medicinal liquid, every day intraperitoneal injection of cyclophosphamide 15mg/kg, continuous 8 days, after the last administration 24 hours, mice takes off cervical vertebra puts to death, and peels off sarcoma, scales/electronic balance weighing.Calculate and respectively organize heavy meansigma methods of mouse tumor and tumour inhibiting rate.The results are shown in Table 1.
Table 4 present composition suppresses murine sarcoma S to cyclophosphamide
180The influence of tumor weight and tumour inhibiting rate (x ± SD, n=10)
The heavy tumour inhibiting rate of dosage Cy tumor
Group
(g/kg) (mg/kg×d) (g，
-X±SD) (％)
Lotus tumor matched group // 1.910 ± 0.402
Cy group/15 * 8 1.086 ± 0.305
* *43.12
Positive drug group 4.8 15 * 8 0.810 ± 0.258
* * Δ57.59
High dose group 10.0 15 * 8 0.708 ± 0.377
* * Δ62.93
Middle dosage group 5.0 15 * 8 0.735 ± 0.366
* * Δ61.52
Low dose group 2.5 15 * 8 0.904 ± 0.413
* *52.67
Annotate: 1. compare (t-check) with lotus tumor matched group,
* *P＜0.001;
2. compare (t-check) with the Cy group,
ΔP＜0.05;
Table 4 as seen, S
180Tumor-bearing mice is irritated stomach simultaneously in the Cy chemotherapy and is given the present composition, compare with lotus tumor matched group, each administration group tumor weight average has remarkable inhibitory action (P＜0.001), the heavy suppression ratio of tumor is between 52% to 62%, and compare with the Cy group with single, high, middle dosage group tumor is heavy obviously to alleviate (P＜0.05), shows that this product suppresses murine sarcoma S to Cy
180Obvious synergistic effect is arranged.
3, to the influence of tumor-bearing mice Cy peripheral blood after chemotherapy elephant
70 of Male Kunming strain mice, body weight 18～22 grams, being divided into 7 groups at random is normal control group, lotus tumor matched group, cyclophosphamide chemotherapy group, positive drug group and the high, medium and low dosage group of administration.The administration group is irritated stomach respectively and is given present composition suspension 10.0,5.0 and 2.5g crude drug/kg, and the positive drug group is irritated stomach and given weixuening granule 4.8g/kg, normally reaches lotus tumor matched group and irritates stomach and give distilled water with volume, the equal gastric infusion of mice 15 days.And in the 4th day of administration, except that the normal control group, all the other mice right fore armpit subcutaneous vaccination S
180Oncocyte liquid 0.2ml/ Mus (method the same 2.1).Except that normal and lotus tumor matched group, all beginnings next day (being administration the 5th day) behind inoculated tumour of all the other mices, when the filling stomach gives water or medicinal liquid, intraperitoneal injection of cyclophosphamide every day (Cy) 15mg/kg, continuous 8 days.Behind last injection Cy second, four, six day, each treated animal eye socket rear vein beard 30 μ l that take a blood sample injected diluent, and the homemade full-automatic animal blood counting instrument of usage is measured peripheral hemogram.The results are shown in Table 5～8.
Table 5 present composition to the influence of WBC in the tumor-bearing mice Cy peripheral blood after chemotherapy liquid (x ± SD, n=10)
Cy WBC(10
9/L)
Group dosage (g/kg)
(mg/kg * d) for the third time for the second time for the first time
Normal control group // 17.0 ± 4.8 17.2 ± 2.7 18.1 ± 5.1
Lotus tumor matched group // 16.9 ± 5.5
* *16.0 ± 4.5
*18.1 ± 4.7
Cy group/15 * 8 7.3 ± 3.1 12.2 ± 2.6 18.3 ± 5.1
Positive drug group 4.8 15 * 8 11.9 ± 3.5
*15.9 ± 4.3
*19.8 ± 5.5
High dose group 10.0 15 * 8 12.2 ± 3.9
*17.1 ± 3.4
*19.7 ± 6.8
Middle dosage group 5.0 15 * 8 11.3 ± 4.5
*15.7 ± 3.9
*18.4 ± 3.9
Low dose group 2.5 15 * 8 13.2 ± 4.3
*15.1 ± 3.4
*20.8 ± 4.5
Annotate: with the Cy chemotherapy group than (t check),
*P＜0.05,
*P＜0.01,
* *P＜0.001.
Table 6 present composition to the influence of RBC in the tumor-bearing mice Cy peripheral blood after chemotherapy liquid (x ± SD, n=10)
Cy RBC(10
12/L)
Group dosage (g/kg)
(mg/kg * d) for the third time for the second time for the first time
Normal control group // 13.23 ± 1.53 12.72 ± 2.22 12.13 ± 1.15
Lotus tumor matched group // 11.96 ± 1.47 11.29 ± 1.01 11.98 ± 1.07
Cy group/15 * 8 10.92 ± 1.19 11.55 ± 1.10 11.04 ± 1.35
Positive drug group 4.8 15 * 8 11.50 ± 1.46 10.76 ± 1.28 10.13 ± 1.74
High dose group 10.0 15 * 8 12.52 ± 1.76
*12.43 ± 1.20 11.93 ± 1.66
Middle dosage group 5.0 15 * 8 11.99 ± 1.47 11.76 ± 2.34 11.11 ± 0.86
Low dose group 2.5 15 * 8 11.07 ± 1.39 11.55 ± 2.04 11.90 ± 1.60
Annotate: with the Cy chemotherapy group than (t check),
*P＜0.05.
Table 7 present composition to the influence of HGB in the tumor-bearing mice Cy peripheral blood after chemotherapy liquid (x ± SD, n=10)
Cy HGB(g/L)
Group dosage (g/kg)
(mg/kg * d) for the third time for the second time for the first time
Normal control group // 205.7 ± 19.1
Δ201.9 ± 37.0
Δ187.1 ± 18.0
Lotus tumor matched group // 182.3 ± 28.0 170.4 ± 15.5 185.7 ± 14.9
Cy group/15 * 8 169.6 ± 21.9 167.5 ± 19.2 171.4 ± 21.2
Positive drug 4.8 15 * 8 179.6 ± 26.8 161.1 ± 18.6 152.5 ± 22.6
High dose group 10.0 15 * 8 193.7 ± 24.9
*192.9 ± 21.9
*179.1 ± 25.0
Middle dosage group 5.0 15 * 8 187.4 ± 25.2 176.6 ± 32.1 167.8 ± 11.5
Low dose group 2.5 15 * 8 172.4 ± 22.2 173.6 ± 25.8 183.4 ± 26.8
Annotate: 1. with the Cy chemotherapy group than (t check),
*P＜0.05.
With lotus tumor group than (t check),
ΔP＜0.05.
Table 8 present composition to the influence of PLT in the tumor-bearing mice Cy peripheral blood after chemotherapy liquid (x ± SD, n=10)
Dosage Cy PLT (10
9/ L)
Group
(g/kg) (mg/kg * d) for the third time for the second time for the first time
Normal control group // 707.6 ± 132.1 793.2 ± 111.0 643.3 ± 79.7
Lotus tumor matched group // 627.6 ± 104.8 752.3 ± 202.4 697.4 ± 109.2
Cy group/15 * 8 649.5 ± 160.5 709.9 ± 96.0 720.0 ± 112.0
Positive drug 4.8 15 * 8 691.2 ± 123.4 778.5 ± 106.9 787.2 ± 192.7
High dose group 10.0 15 * 8 660.0 ± 89.9 771.1 ± 166.7 768.1 ± 82.1
Middle dosage group 5.0 15 * 8 693.4 ± 167.7 734.6 ± 118.6 766.7 ± 103.4
Low dose group 2.5 15 * 8 771.7 ± 198.1 748.5 ± 122.2 798.9 ± 68.9
By table 5～8 as seen, tumor-bearing mice is after the Cy chemotherapy, and WBC quantity significantly descends in the peripheral blood, and presents tangible time-effect relationship, and 24 hours is minimum point behind last injection Cy, recovers by wash rice later on.Compare with the Cy group, the WBC quantity that gives each treated animal of present composition treatment all has obvious recovery (P＜0.05～0.01) in different time sections, and chemotherapy group WBC quantity in minimum point time effect more remarkable, show that WBC number that this product causes chemotherapeutic reduces significant improvement effect is arranged.The RBC of chemotherapy group, HGB, PLT number then do not have significant change, but this product high dose group RBC and HGB are than chemotherapy group slightly raise (P＜0.05).
4, to the influence of bone marrow nucleated cell after the tumor-bearing mice Cy chemotherapy
70 of Male Kunming strain mice, body weight 18-22 gram, being divided into 7 groups is normal group, lotus tumor matched group, cyclophosphamide chemotherapy group, weixuening groups of grains and the high, medium and low dosage group of the present composition.The administration group is irritated stomach respectively and is given present composition suspension 10.0,5.0 and 2.5g crude drug/kg, and the positive drug group is irritated stomach and given weixuening granule 4.8g/kg, normally reaches lotus tumor matched group and irritates stomach and give distilled water with volume, the equal gastric infusion of mice 12 days.And in the 4th day of administration, except that the normal control group, all the other mice right fore armpit subcutaneous vaccination S
180Oncocyte liquid 0.2ml/ Mus (method the same 2.1).Except that lotus tumor matched group, all the other mices all behind inoculated tumour next day begin, when the filling stomach gives water or medicinal liquid, intraperitoneal injection of cyclophosphamide every day (Cy) 15mg/kg, continuous 8 days, after the last administration 24 hours, mice takes off cervical vertebra puts to death, and peels off the left side femur, goes out medullary cell with 3% acetic acid 10ml, by the numeration of leukocyte method, calculate the bone marrow nucleated cell quantity in 1 bone.The results are shown in Table 9.
Table 9 present composition to tumor-bearing mice Cy chemotherapy after bone marrow nucleated cell quantity influence (x ± SD, n=10)
Group dosage (g/kg) Cy (the bone marrow nucleated cell number (10 of mg/kg * d)
9/ L)
Normal control group // 24.32 ± 3.75
Lotus tumor matched group // 23.70 ± 4.65
* *
Cy group/15 * 8 12.50 ± 2.86
Positive drug group 4.8 15 * 8 16.60 ± 3.66
*
High dose group 10.0 15 * 8 17.84 ± 3.05
* *
Middle dosage group 5.0 15 * 8 16.78 ± 3.33
*
Low dose group 2.5 15 * 8 15.29 ± 2.84
*
Annotate: with the Cy chemotherapy group than (t-check),
*P＜0.05,
*P＜0.01,
* *P＜0.001.
By table 9 as seen, with the Cy chemotherapy group relatively, the present composition is respectively organized mouse bone marrow cells nucleated cell number average and is obviously raise (P＜0.05～0.001), shows that bone marrow nucleated cell quantity that this product causes chemotherapy reduces to have significantly to increase effect.
5, to the influence of immune organ weight after the tumor-bearing mice Cy chemotherapy
70 of Male Kunming strain mice, body weight 18-22 gram, experimental technique is with 2.3.After the last administration 24 hours, take off cervical vertebra after mice is weighed and put to death, get spleen and thymus and weigh and calculate organ index, the results are shown in Table 10.
Table 10 present composition to tumor-bearing mice Cy chemotherapy after immune organ weight influence (x ± SD, n=10)
Group dosage (g/kg) Cy (index and spleen index (mg/g) thymus index (mg/g) of mg/kg * d)
Normal control group // 5.01 ± 1.16 2.70 ± 0.76
Lotus tumor matched group // 5.81 ± 0.74
* *2.64 ± 0.88
*
Cy group/15 * 8 3.27 ± 0.65 1.58 ± 0.44
Positive drug group 4.8 15 * 8 4.51 ± 1.70
*2.11 ± 0.64
*
High dose group 10.0 15 * 8 4.72 ± 1.34
*2.33 ± 0.54
*
Middle dosage group 5.0 15 * 8 4.13 ± 1.02
*1.93 ± 0.52
Low dose group 2.5 15 * 8 3.97 ± 1.13 1.67 ± 0.67
Annotate: with the Cy chemotherapy group than (t-check),
*P＜0.05,
*P＜0.01,
* *P＜0.001.
Table 10 as seen, compare with the Cy chemotherapy group, present composition height, middle dosage group mouse spleen index obviously increase (P＜0.05～0.01), and the high dose group thymus index significantly increases (P＜0.01), show that this product improves significantly to thymus and the index and spleen index reduction that the Cy chemotherapy causes.Experimental result shows that the present composition is to the caused S of cyclophosphamide chemotherapy
180Tumor-bearing mice peripheral white blood cell amount reduces and to improve significantly, the bone marrow nucleated cell quantity that can obviously raise simultaneously, and can increase spleen and the thymus index of lotus tumor chemotherapy mice; This product and cyclophosphamide share, and can obviously suppress tumor growth, and the tumor-inhibiting action of chemotherapeutic Cy is had obvious potentiation.Above result shows that the present composition and chemotherapeutic Cy share, and have tangible synergism and attenuation.
6, right
60The Co irradiation suppresses murine sarcoma S
180Synergism and attenuation
Pharmaceutical composition of the present invention is by 10.0,5.0 and 2.5g crude drug/kg gastric infusion, to S
180The tumor-bearing mice warp
60Caused leucocytes reduction of Co radiotherapy and marrow nucleated cell decreasing have tangible rising effect, and caused spleen of radiotherapy and thymus index decline are had some improvement, and the radiotherapy tumour inhibiting rate that can significantly raise shows that this product is right
60Co radiotherapy suppresses murine sarcoma S
180Tangible synergism and attenuation is arranged.
The present composition (dry powder): contain crude drug 37.0g/g, lot number: 20020629, Chinese medicine preparation research department in Jiangsu Prov. Research Inst. Traditional Chinese Medical provides.All the other are the same.
Experimental technique and result: right
60Co radiotherapy suppresses murine sarcoma S
180Tumor heavily reaches the influence of tumour inhibiting rate
60 of Male Kunming strain mice, body weight 18-22 gram, be divided into 6 groups be lotus tumor matched group,
60The high, medium and low dosage group of Co combination radiotherapy group, weixuening groups of grains and the present composition.The administration group is irritated stomach respectively and is given present composition suspension 10.0,5.0 and 2.5g crude drug/kg, and the positive drug group is irritated stomach and given weixuening granule 4.8g/kg, and lotus tumor matched group filling stomach gives the distilled water with volume, the equal gastric infusion of mice 12 days.And, get abdominal part inoculation S in the 4th day of administration
1808 days tumor-bearing mice of tumor strain, the sterile working extracts down abdominal cavity tumor liquid, and (microscopy contains oncocyte several 5 * 10 to be diluted to the oncocyte liquid of 1: 4 concentration with sterile saline
7/ ml), in all mice right fore armpit subcutaneous vaccination 0.2ml/ Mus.Except that lotus tumor matched group, all the other mices all after three days, once give 350Rad in inoculated tumour
60The Co irradiation.After the last administration 24 hours, mice takes off cervical vertebra put to death, and peels off sarcoma, scales/electronic balance weighing.Calculate and respectively organize heavy meansigma methods of mouse tumor and tumour inhibiting rate.The results are shown in Table 11.
Table 11 present composition is right
60Co radiotherapy suppresses murine sarcoma S
180The influence of tumor weight and tumour inhibiting rate (x ± SD, n=10)
Dosage
60The heavy tumour inhibiting rate of Co (Rad) tumor
Group
(g/kg) (g，
-X±SD) (％)
Lotus tumor matched group // 2.683 ± 0.695
60Co group/350 1.608 ± 0.467
* *40.06
Positive drug group 4.8 350 1.211 ± 0.0.365
* * Δ54.85
High dose group 10.0 350 1.153 ± 0.346
* * Δ57.02
Middle dosage group 5.0 350 1.179 ± 0.319
* * Δ56.05
Low dose group 2.5 350 1.332 ± 0.223
* *50.35
Annotate: 1. compare (t-check) with lotus tumor matched group,
* *P＜0.001;
With
60The Co combination radiotherapy group is (t-check) relatively,
ΔP＜0.05;
Table 11 as seen, S
180Tumor-bearing mice exists
60Co radiotherapy is irritated stomach simultaneously and is given the present composition, with lotus tumor matched group relatively, each administration group tumor weight average has remarkable inhibitory action (P＜0.001), the heavy suppression ratio of tumor between 50% to 57%, and with single usefulness
60The Co combination radiotherapy group compares, and high, middle dosage group tumor is heavy obviously to alleviate (P＜0.05), shows that this product is right
60Co radiotherapy suppresses murine sarcoma S
180Obvious synergistic effect is arranged.
7, to tumor-bearing mice
60The influence of peripheral hemogram after the Co radiotherapy
70 of Male Kunming strain mice, body weight 18-22 gram, be divided at random 7 groups be normal control group, lotus tumor matched group,
60The high, medium and low dosage group of Co combination radiotherapy group, positive drug group and administration.The administration group is irritated stomach respectively and is given present composition suspension 10.0,5.0 and 2.5g crude drug/kg, and the positive drug group is irritated stomach and given weixuening granule 4.8g/kg, normally reaches lotus tumor matched group and irritates stomach and give distilled water with volume, the equal gastric infusion of mice 15 days.And in the 4th day of administration, except that the normal control group, all the other mice right fore armpit subcutaneous vaccination S
180Oncocyte liquid 0.2ml/ Mus (method the same 2.1).Except that normal and lotus tumor matched group, all the other mices all in inoculated tumour after 3 days (being administration the 7th day) once give 350Rad
60The Co irradiation.In
60Co irradiation back the 3rd, six, the Ninth Heaven, each the treated animal eye socket rear vein beard 30 μ l that take a blood sample inject diluent, and the homemade full-automatic animal blood counting instrument of usage is measured peripheral hemogram.Experiment shows: the tumor-bearing mice warp
60After the Co radiotherapy, WBC quantity significantly descends in the peripheral blood, and continues to keep reduced levels, and the obviously minimizing on the 6th after irradiation of RBC, HGB and PLT quantity, to the 9th day, PLT still kept reduced levels.With
60The Co combination radiotherapy group relatively, the WBC quantity of present composition height, middle dosage treated animal daily had obvious recovery (P＜0.05～0.01) at the 6th day and the 9th, high dose group HGB and PLT quantity is obviously rising (P＜0.05～0.01) also, reduction to RBC has rising trend, but there was no significant difference.Above result shows that this product has significant improvement effect to the WBC number reduction that chemotherapeutic causes, and raise to a certain extent HGB and PLT.
8, to tumor-bearing mice
60The influence of bone marrow nucleated cell after the Co radiotherapy
70 of Male Kunming strain mice, body weight 18-22 gram, be divided into 7 groups be normal group, lotus tumor matched group,
60The high, medium and low dosage group of Co combination radiotherapy group, weixuening groups of grains and the present composition.The administration group is irritated stomach respectively and is given present composition suspension 10.0,5.0 and 2.5g crude drug/kg, and the positive drug group is irritated stomach and given weixuening granule 4.8g/kg, normally reaches lotus tumor matched group and irritates stomach and give distilled water with volume, the equal gastric infusion of mice 12 days.And in the 4th day of administration, except that the normal control group, all the other mice right fore armpit subcutaneous vaccination S
180Oncocyte liquid 0.2ml/ Mus (method the same 2.1).Except that lotus tumor matched group, all the other mices all once give 350Rad in inoculated tumour after 3 days
60The Co irradiation.After the last administration 24 hours, mice takes off cervical vertebra put to death, and peels off the left side femur, goes out medullary cell with 3% acetic acid 10ml, by the numeration of leukocyte method, calculated the interior bone marrow nucleated cell quantity of 1 bone.Experiment shows: with
60The Co combination radiotherapy group compares, and the present composition is right
60The caused S of Co radiotherapy
180Tumor-bearing mice peripheral white blood cell amount reduces and to improve significantly, and raises to a certain extent because the HGB that radiotherapy causes and the decline of PLT quantity, and bone marrow nucleated cell quantity can obviously raise simultaneously; This product with
60Co irradiation is share, and can obviously suppress tumor growth, and the tumor-inhibiting action of radiotherapy is had obvious potentiation.Above result shows that the present composition and radiotherapy share, and tangible synergism and attenuation is arranged.
9, to the Attenuation of normal mouse chemotherapy
Pharmaceutical composition of the present invention is by 10.0,5.0 and 2.5g crude drug/kg gastric infusion, normal mouse there is tangible rising effect through caused leucocytes reduction of cyclophosphamide chemotherapy and the minimizing of bone marrow nucleated cell number, caused spleen of chemotherapy and thymus index decline there are significant improvement effect, show that this product has tangible Attenuation to normal mouse through the cyclophosphamide chemotherapy.
10, to the tumor-bearing mice Immune Effects
Pharmaceutical composition of the present invention can make S by 10.0,5.0 and 2.5g crude drug/kg gastric infusion
180Tumor-bearing mice strengthens the lymphocyte transfer reaction that PHA stimulates, and mice peripheral blood T lymphocyte percentage ratio obviously raises, and tumor mice serum hemolysin level significantly improves.Above result shows that this product all has obvious potentiation to the cellular immunization and the humoral immune function of tumor mice.
The present composition (dry powder): contain crude drug 37.0g/g, lot number: 20020629, Chinese medicine preparation research department in Jiangsu Prov. Research Inst. Traditional Chinese Medical provides.
PHA is stimulated LT influence (revulsion in the body), referring to: Li Yikui etc. the herbal pharmacology experimental methodology. Shanghai: Shanghai science tech publishing house, 1991:164
70 of Kunming mouses, male and female half and half, body weight 18-22 gram, being divided into 7 groups is normal control group, lotus tumor matched group, cyclophosphamide model group, weixuening groups of grains and the high, medium and low dosage group of the present composition.Except that the normal control group, all the other mices are all in the S of right fore armpit subcutaneous vaccination dilution in 1: 4
180Oncocyte liquid 0.2ml/ Mus.Inoculated tumour begins administration next day, the administration group is irritated stomach respectively and is given present composition suspension 10.0,5.0 and 2.5g crude drug/kg, the positive drug group is irritated stomach and is given weixuening granule 4.8g/kg, normal group, lotus tumor matched group and model group filling stomach give the distilled water with volume, the equal gastric infusion of mice 8 days.And in the 1st day of administration, intramuscular injection every day PHA 10mg/kg once, continuous three times; Administration the 6th day is except that normal and lotus tumor matched group, the equal intraperitoneal injection of cyclophosphamide 40mg/kg of all the other each treated animals, the 7th day repeats once, after the last administration 1 hour, the blood sampling of mouse orbit rear vein beard, push jack, methanol is fixed, Ji's Albert'stain Albert, washing, dry, high power lens is counted lymphoblast number in 100 lymphocytes down, calculates lymhocyte transformation rate.The results are shown in Table 12.
Table 12, the present composition are to the influence of tumor-bearing mice lymphocyte transformation and T lymphocyte esterase dyeing rate
( x±SD，n＝10)
Group dosage Cy lymhocyte transformation rate ANAE rate of dyeing
(g/kg) (mg/kg×d) (％，
-X±SD) (％，
-X±SD)
Normal control group // 43.6 ± 5.6 59.3 ± 7.1
Lotus tumor matched group // 44.5 ± 3.9
* *58.7 ± 6.1
* *
Cy group/40 * 2 15.0 ± 4.8 32.2 ± 5.8
Positive drug group 4.8 40 * 2 22.7 ± 7.4
*40.8 ± 8.6
*
High dose group 10.0 40 * 2 24.8 ± 5.4
* *41.2 ± 7.2
*
Middle dosage group 5.0 40 * 2 22.5 ± 4.8
*39.0 ± 7.2
*
Low dose group 2.5 40 * 2 20.0 ± 3.9
*35.9 ± 7.5
Annotate: compare (t-check) with the Cy group,
*P＜0.05,
*P＜0.01,
* *P＜0.001;
Table 12 as seen, each dosage treated animal of the present composition strengthens the conversion reaction that PHA stimulates, lymphoblast percentage rate and cyclophosphamide group relatively are significantly increased (P＜0.05～0.001), show that this product can improve the lymphocytic answering of tumor mice T, promptly improves cellular immune function.
11, to the influence of tumor-bearing mice T-lymphocyte esterase dyeing rate
70 of Kunming mouses, male and female half and half, body weight 18-22 gram, being divided into 7 groups is normal control group, lotus tumor matched group, cyclophosphamide model group, weixuening groups of grains and the high, medium and low dosage group of the present composition.Except that the normal control group, all the other mices are all in the S of right fore armpit subcutaneous vaccination dilution in 1: 4
180Oncocyte liquid 0.2ml/ Mus.Inoculated tumour begins administration next day, the administration group is irritated stomach respectively and is given present composition suspension 10.0,5.0 and 2.5g crude drug/kg, the positive drug group is irritated stomach and is given weixuening granule 4.8g/kg, normal group, lotus tumor matched group and model group filling stomach give the distilled water with volume, the equal gastric infusion of mice 10 days.The 8th day except that normal and lotus tumor matched group, the equal intraperitoneal injection of cyclophosphamide 40mg/kg of all the other each treated animals, the 9th day repeats once, and after the last administration 1 hour, the blood sampling of mouse orbit rear vein beard, push jack, esterase dyeing dries, and oily mirror is 100 lymphocytes of counting down, wherein having brownish red particulate is the T cell, and no brownish red is particulate to be the B cell.By table 12 result as seen, present composition height, middle dosage group peripheral blood T lymphocyte percentage ratio are apparently higher than cyclophosphamide group (P＜0.05～0.01), illustrate that this product has obvious restitution to the tumor mice immunosuppressant of caused by cyclophosphamide, promptly the pair cell immunologic function has facilitation.
12, the influence (colorimetry) that the tumor-bearing mice hemolytic antibody is generated
70 of Kunming mouses, male and female half and half, body weight 18-22 gram, being divided into 7 groups is normal control group, lotus tumor matched group, cyclophosphamide model group, weixuening groups of grains and the high, medium and low dosage group of the present composition.Except that the normal control group, all the other mices are all in the S of right fore armpit subcutaneous vaccination dilution in 1: 4
180Oncocyte liquid 0.2ml/ Mus.Inoculated tumour begins administration next day, the administration group is irritated stomach respectively and is given present composition suspension 10.0,5.0 and 2.5g crude drug/kg, the positive drug group is irritated stomach and is given weixuening granule 4.8g/kg, normal group, lotus tumor matched group and model group filling stomach give the distilled water with volume, the equal gastric infusion of mice 10 days.The 3rd day each treated animal lumbar injection 5% chicken erythrocyte suspension 0.2ml/ only carries out immunity after administration, again after administration the 8th day except that normal and lotus tumor matched group, the equal intraperitoneal injection of cyclophosphamide 40mg/kg of all the other each treated animals, the 9th day repeats once, after the last administration 1 hour, the blood sampling of mouse orbit rear vein beard, centrifugal, get 10 μ l serum and dilute 100 times with normal saline, mix with 5% chicken erythrocyte suspension 0.5ml and 10% complement 0.5ml, after being incubated 30min in 37 ℃ of waters bath with thermostatic control, cessation reaction in 0 ℃ of ice-water bath, centrifugal, get supernatant in spectrophotometer 540nm place colorimetric, the photometry density value, other establishes the not blank of increase serum, the benchmark that returns to zero when getting supernatant as colorimetric.With the index of OD value as judgement serum hemolysin level.The present composition can obviously increase tumor-bearing mice serum hemolysin level (P＜0.05～0.001) in the experiment, makes the serum hemolysin content decline of caused by cyclophosphamide that obvious recovery be arranged, and shows that this product can improve the humoral immunization ability of tumor mice.Above experimental result shows that the present composition is to S
180Tumor-bearing mice has tangible restitution by the caused immunosuppressant of cyclophosphamide, T lymphocyte esterase dyeing rate, the inductive lymphocyte transfer reaction of enhancing PHA can significantly raise, and can obviously improve the serum hemolysin level, show that this product all has remarkable potentiation for the cellular immunization and the humoral immunization of tumor mice.
Four, present composition acute toxicity test
The present composition does not see tangible acute toxic reaction for the mouse stomach administration, can't measure median lethal dose(LD 50) (LD according to a conventional method
50).The maximum dosage-feeding of a gastric infusion of mice is 740g crude drug/kg, and this dosage is equivalent to intend 822 times of clinical people's consumption, does not see tangible toxic reaction.
Five, present composition long term toxicity test
3 months long term toxicity test of gastric infusion studies show that continuously, give SD rat continuous irrigation stomach 90 days with the present composition that is equivalent to raw medicinal herbs 50.0,25.0 and 12.5g/kg dosage, wherein heavy dose ofly be about clinical people and intend 55.6 times with dosage, do not find that animal has phenomenons such as movable minimizing, Mao Songluan, listlessness be depressed, the body weight of rat, organ coefficient, blood biochemistry checking etc. be there is no tangible toxic reaction.Blood biochemical learn to check show heavy dose of group 3 months ALP and 6 months AST obviously raise, in, small dose group is not seen that overt toxicity is arranged, show that (25.0g crude drug/kg) is following to be safe dose for the middle dosage of this product long term toxicity test, heavy dose of group has certain infringement to liver function, notes blood biochemical is learned the monitoring of index when submitting clinical use to.
The specific embodiment
Embodiment 1
Get Radix Astragali 3.0kg, Fructus Lycii 1.2kg, Ganoderma 1.2kg.
Milkvetch Root is for the first time with 8 times of amounts, the second time measuring 80% alcohol reflux secondaries with 6 times, each 2 hours, merge backflow, filter, reclaim ethanol, mother solution leaves standstill 12 hours, and is centrifugal, handled the AB-8 purification by macroporous resin on the centrifugal liquid, 70% ethanol elution, reclaim ethanol, drying gets total saponins 30 grams.
Astragalus root dregs, prescription drugss such as Fructus Lycii, Ganoderma extract three times with 12 times of water gagings, each 1.5 hours, merge decoction liquor, be concentrated into relative density 1.2 (80 ℃), ethanol precipitation contains the alcohol amount and reaches 80%, leave standstill, filter, precipitate is flung to ethanol to doing, the adding distil water dissolving, centrifugal, centrifugal liquid is concentrated into relative density 1.2 (80 ℃), the reuse ethanol precipitation contains the alcohol amount and reaches 75%, leaves standstill, filter, fling to ethanol, get total polysaccharides 150 grams to doing.
Merge total Saponin and total polysaccharides, pulverize the snap fit capsule of packing into.Instructions about how to take medicine: one day 3 times, one time 2.
Embodiment 2
Get Radix Astragali 3.0kg, Fructus Lycii 1.2kg, Ganoderma 1.2kg.
Milkvetch Root is for the first time with 8 times of amounts, the second time measuring 80% alcohol reflux secondaries with 6 times, each 2 hours, merge backflow, filter, reclaim ethanol, mother solution leaves standstill 12 hours, and is centrifugal, handled the AB-8 purification by macroporous resin on the centrifugal liquid, 70% ethanol elution, reclaim ethanol, drying gets total saponins 30 grams.
Astragalus root dregs, prescription drugss such as Fructus Lycii, Ganoderma extract three times with 12 times of water gagings, each 1.5 hours, merge decoction liquor, be concentrated into relative density 1.2 (80 ℃), ethanol precipitation contains the alcohol amount and reaches 80%, leave standstill, filter, precipitate is flung to ethanol to doing, the adding distil water dissolving, centrifugal, centrifugal liquid is concentrated into relative density 1.2 (80 ℃), the reuse ethanol precipitation contains the alcohol amount and reaches 75%, leaves standstill, filter, fling to ethanol as for, total polysaccharides 150 grams.
Merge total Saponin and total polysaccharides, pulverize, add excipient, tabletting.Instructions about how to take medicine: one day 3 times, one time 3.
Embodiment 3
Get Radix Astragali 4.0kg, Fructus Lycii 1.5kg, Ganoderma 1.5kg.
With the Radix Astragali of described weight proportion, with 90% ethanol extraction 4 times, each 2.5 hours, merge ethanol extract, reclaim ethanol, with removing AB-8 macroporous resin column on the liquid of post precipitation, use 80% ethanol elution, Radix Astragali total saponins 40 restrains;
Fructus Lycii, Ganoderma and the above-mentioned ethanol extraction of described weight proportion are crossed Radix Astragali residue merge, add 10 times of water gagings, decoct 4 times, each 2 hours, merge decoction liquor, be concentrated into relative density 1.3, adding ethanol makes and contains the alcohol amount and reach 70%, filter precipitate, dry sediment is after being dissolved in water, water liquid is concentrated into relative density 1.1, add ethanol and make and contain the alcohol amount and reach 70%, leaching precipitate, dry total polysaccharides 196 grams; Merge total Saponin and total polysaccharides, pulverize, add ethanol and make binding agent, add starch and make filler, routine is pressed into granule.Instructions about how to take medicine: one day 3 times, one time 1 bag.
Embodiment 4
Get Radix Astragali 4.0kg, Fructus Lycii 3.5kg, Ganoderma 3.5kg.
With the Radix Astragali of described weight proportion, with 70% ethanol extraction 2 times, each 1 hour, merge ethanol extract, reclaim ethanol, with removing D101 macroporous resin column on the liquid of post precipitation, use 60% ethanol elution, Radix Astragali total saponins 40 restrains;
Fructus Lycii, Ganoderma and the above-mentioned ethanol extraction of described weight proportion are crossed Radix Astragali residue merge, add 14 times of water gagings, decoct 2 times, each 1 hour, merge decoction liquor, be concentrated into relative density 1.2, adding ethanol makes and contains the alcohol amount and reach 85%, filter precipitate, dry sediment is after being dissolved in water, water liquid is concentrated into relative density 1.3, add ethanol and make and contain the alcohol amount and reach 70%, leaching precipitate, dry total polysaccharides 276 grams; Radix Astragali total saponins is mixed with total polysaccharides, pulverize, add conventional adjuvant, make powder.Instructions about how to take medicine: one day 3 times, one time 1 bag.
Embodiment 5
Get Radix Astragali 4.0kg, Fructus Lycii 2.0kg, Ganoderma 2.0kg
With the Radix Astragali of described weight proportion, with 75% ethanol extraction 3 times, each 1 hour, merge ethanol extract, reclaim ethanol, with removing D101 macroporous resin column on the liquid of post precipitation, use 65% ethanol elution, Radix Astragali total saponins 40 restrains;
Fructus Lycii, Ganoderma and the above-mentioned ethanol extraction of described weight proportion are crossed Radix Astragali residue to be merged, adding 13 times of water gagings decocts, relative density 1.2 when decoction liquor was concentrated into 80 ℃, added ethanol and made and contain the alcohol amount and reach 80%, filter precipitate, dry sediment, after being dissolved in water, relative density 1.2 when water liquid was concentrated into 80 ℃, added ethanol and made and contain the alcohol amount and reach 75%, the leaching precipitate, dry that total polysaccharides 210 restrains; Merge total Saponin and total polysaccharides, pulverize, the conventional granulation, encapsulated.
Embodiment 6
Get Radix Astragali 4.0kg, Fructus Lycii 1.0kg, Ganoderma 1.0kg
With the Radix Astragali of described weight proportion, with 75% ethanol extraction 3 times, each 1 hour, merge ethanol extract, reclaim ethanol, with removing D101 macroporous resin column on the liquid of post precipitation, use 65% ethanol elution, Radix Astragali total saponins 40 restrains;
Fructus Lycii, Ganoderma and the above-mentioned ethanol extraction of described weight proportion are crossed Radix Astragali residue to be merged, adding 13 times of water gagings decocts, the water intaking decoction liquor, concentrate, precipitate with ethanol, filter behind the precipitate, taking precipitate is dissolved in water, and filters, and operates routinely with film and carries out ultrafiltration, remove macromole impurity with 300,000 molecular cut off membrane ultrafiltration earlier, and then remove moisture and part small molecular weight impurity with 1000 molecular cut off membrane ultrafiltration, and be concentrated into relative density 1.2, add ethanol and make and contain the alcohol amount and reach 75%, the leaching precipitation gets total polysaccharides 176 grams.Merge total Saponin and total polysaccharides, pulverize, encapsulated.
Embodiment 7
Get Radix Astragali 3.0kg, Fructus Lycii 1.2kg, Ganoderma 1.2kg.
Milkvetch Root is for the first time with 8 times of amounts, the second time measuring 80% alcohol reflux secondaries with 6 times, each 2 hours, merge backflow, filter, reclaim ethanol, mother solution leaves standstill 12 hours, and is centrifugal, handled the AB-8 purification by macroporous resin on the centrifugal liquid, 70% ethanol elution, reclaim ethanol, drying gets total saponins 30 grams.
Astragalus root dregs, prescription drugss such as Fructus Lycii, Ganoderma extract three times with 12 times of water gagings, each 1.5 hours, merge decoction liquor, be concentrated into relative density 1.2 (80 ℃), ethanol precipitation contains the alcohol amount and reaches 80%, leave standstill, filter, precipitate is flung to ethanol to doing, the adding distil water dissolving, centrifugal, centrifugal liquid is concentrated into relative density 1.2 (80 ℃), the reuse ethanol precipitation contains the alcohol amount and reaches 75%, leaves standstill, filter, fling to ethanol, get total polysaccharides 150 grams to doing.Merge total Saponin and total polysaccharides, the adding distil water dissolving is filtered, and makes every 1ml and contains 1.8g crude drug solution, and fill is sterilized in 10ml control oral liquid bottle.
Claims (10)
1, a kind of pharmaceutical composition that immunoregulation effect is arranged is characterized in that it is made by following bulk drugs basically:
The Radix Astragali, Fructus Lycii, Ganoderma are 1-40 part: 1-15 part: 1-15 part.
2, the pharmaceutical composition of claim 1, wherein the consumption of crude drug is: the Radix Astragali, Fructus Lycii, Ganoderma weight ratio are 1-30 part: 1-12 part: 1-12 part.
3, the pharmaceutical composition of claim 2, wherein the consumption of crude drug is: the Radix Astragali, Fructus Lycii, Ganoderma weight ratio are 30 parts: 12 parts: 12 parts.
4, one of claim 1～3 preparation of drug combination method comprises the following steps:
A) get the crude drug Radix Astragali, Fructus Lycii and Ganoderma, standby;
B) with the Radix Astragali of described weight proportion, with 70～90% ethanol extractions 1～4 time, each 1～2.5 hour, merge ethanol extract, reclaim ethanol, with removing macroporous resin column on the liquid of post precipitation, with 55～84% ethanol elutions, must Radix Astragali total saponins;
C) Fructus Lycii, Ganoderma and the above-mentioned ethanol extraction of described weight proportion are crossed Radix Astragali residue and merged, add 10～14 times of water gagings, merceration or decoction are extracted 2～4 times, each 1～2 hour, merge decoction liquor, be concentrated into relative density 1.0-1.4, adding ethanol makes and contains the alcohol amount and reach 70-85%, filter precipitate, dry sediment is after being dissolved in water, water liquid is concentrated into relative density 1.0-1.4, add ethanol and make and contain the alcohol amount and reach 70-80%, leaching precipitate, dry total polysaccharides;
D) Radix Astragali total saponins is mixed with total polysaccharides, get the active component of medicine of the present invention.
5, the preparation method of claim 4 is characterized in that:
B) with the Radix Astragali of described weight proportion, with 75～85% ethanol extractions 2～3 times, each 1.5～2.0 hours, merge ethanol extract, reclaim ethanol, with AB-8 and/or D101 model macroporous resin column on the liquid of removal post precipitation, with 60～80% ethanol elutions, get Radix Astragali total saponins;
C) Fructus Lycii, Ganoderma and the above-mentioned ethanol extraction of described weight proportion are crossed Radix Astragali residue and merged, add 11～13 times of water gagings, decoct 3 times, each 1.5 hours, merge decoction liquor, relative density 1.1-1.3 when being concentrated into 80 ℃, adding ethanol makes and contains the alcohol amount and reach 75-80%, filter precipitate, dry sediment is after being dissolved in water, relative density 1.1-1.3 when water liquid is concentrated into 80 ℃, add ethanol and make and contain the alcohol amount and reach 75-80%, leaching precipitate, dry total polysaccharides;
D) with Radix Astragali total saponins with after total polysaccharides is mixed, add pharmaceutic adjuvant, routine is made oral formulations.
6, the preparation method of claim 5 is characterized in that;
B) with the Radix Astragali of described weight proportion, with 80～82% ethanol extractions 3 times, each 1.5 hours, merge ethanol extract, reclaim ethanol, with removing AB-8 or D101 model macroporous resin column on the liquid of post precipitation, with 65～70% ethanol elutions, must Radix Astragali total saponins;
C) Fructus Lycii, Ganoderma and the above-mentioned ethanol extraction of described weight proportion being crossed Radix Astragali residue merges, adding 12 times of water gagings decocts, relative density 1.2 when decoction liquor was concentrated into 80 ℃, added ethanol and made and contain the alcohol amount and reach 80%, filter precipitate, dry sediment, after being dissolved in water, relative density 1.2 when water liquid was concentrated into 80 ℃, added ethanol and made and contain the alcohol amount and reach 75%, the leaching precipitate, the dry total polysaccharides that gets;
D) Radix Astragali total saponins, total polysaccharides are mixed with conventional adjuvant pharmaceutically, make capsule, tablet, powder, granule or oral liquid.
7, the preparation method of claim 6 is characterized in that:
C) the water intaking decoction liquor concentrates precipitate with ethanol, filter behind the precipitate, taking precipitate is dissolved in water, and filters, and operates routinely with film and carries out ultrafiltration, remove macromole impurity with 300,000 molecular cut off membrane ultrafiltration earlier, and then remove moisture and part small molecular weight impurity with 1000 molecular cut off membrane ultrafiltration, and be concentrated into relative density 1.2, add ethanol and make and contain the alcohol amount and reach 75%, the leaching precipitation gets total polysaccharides.
8, the application of the pharmaceutical composition of claim 1 in preparation raising immune drug.
9, the pharmaceutical composition of claim 1 is used for the application of the medicine of chemicotherapy attenuation synergistic in preparation.
10, the pharmaceutical composition of claim 1 is preparing the raising immunity of share with chemicotherapy, the application in the attenuation synergistic medicine.
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| CN 03132059 CN1201805C (en) | 2003-07-18 | 2003-07-18 | Combination of medication for reducing poison and synergic action in radiotherapy or chemotherapy as well as its preparing method |
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| WO2006128378A1 (en) * | 2005-06-01 | 2006-12-07 | East China University Of Science And Technology | Use of ganoderic acid in treating tumour |
| CN103191179A (en) * | 2013-03-27 | 2013-07-10 | 张宗升 | Traditional chinese medicine extract and preparation method thereof |
| CN105031301A (en) * | 2015-08-24 | 2015-11-11 | 周伟刚 | Anti-tumor traditional Chinese medicine composition and preparation method thereof |
| CN107510813A (en) * | 2016-06-15 | 2017-12-26 | 郭道明 | Pharmaceutical composition for reducing side effects of cancer treatment drugs, preparation method and application thereof |
| TWI634901B (en) * | 2017-12-14 | 2018-09-11 | 郭道明 | Pharmaceutical composition for decreasing the side effects of pancreatic cancer drug, and manufacturing method and uses thereof |
| CN108392543A (en) * | 2018-05-08 | 2018-08-14 | 东莞市亚洲制药有限公司 | A kind of health composition and its preparation method and application |
| CN109260312A (en) * | 2018-09-12 | 2019-01-25 | 江苏医药职业学院 | A kind of medicinal herb tea that improving immunity and medicinal herb tea drink and tea bag and beverage |
| CN112022897A (en) * | 2020-10-10 | 2020-12-04 | 邓述才 | A Chinese medicinal composition for preventing cancer, and its preparation method |
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