CN1023567C - Process for extracting malignant tumor medicine from metabolite of staphylococcus aureus - Google Patents
Process for extracting malignant tumor medicine from metabolite of staphylococcus aureus Download PDFInfo
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- CN1023567C CN1023567C CN91106067A CN91106067A CN1023567C CN 1023567 C CN1023567 C CN 1023567C CN 91106067 A CN91106067 A CN 91106067A CN 91106067 A CN91106067 A CN 91106067A CN 1023567 C CN1023567 C CN 1023567C
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- pig heart
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Abstract
The present invention discloses a method for extracting medicine for treating malignant tumors using metabolites of staphylococcus aureus, and a product of the method is abbreviated as 'BM828' which has direct and indirect killing effects on malignant tumor cells. A culture medium comprises main components of peptone, pig heart leaching liquid and goat liver leaching liquid. Culture strains are reanimated with broth, and degreased milk is used as a protective agent of the strains. Bacteria are killed with high pressure without any preservative agents, and an injection is prepared through the methods of heat treatment, cold leaching, active carbon treatment, aseptic filtration, etc. The injection has obvious effects of greatly enhancing the immunological functions of human bodies.
Description
The present invention relates to biological response modifier and promptly extract the method for prevention and the medication of treatment malignant tumour with metabolic product of staphylococcus aureus.The product of this method has direct killing effect to malignant cell, can also increase substantially body's immunological function, suppresses indirectly and kills tumour cell, significantly repairs normal tissue cell, leukocyte increasing, hematoblastic quantity and function.Described biological response modifier is meant the material that can significantly strengthen, promote or recover human body defence capability (immunizing power), these material general designation biological response modifier, and the product of described this method abbreviates " BM828 " as.
Entrust auditor's (entrusting numbering 91019) of four ones of State Patent Office's examinations to propose following corresponding document by online information retrieval.JP55-8730 discloses a kind of medicament that is used for bronchial allergy, method is respectively to cultivate each Bacillaceae to gather up cell paste, and comes cell killing with heating, uses " formalin " sterilization again, then, be suspended in the normal saline solution, so just make the dead cell base soln, injection is made in the base soln mixing of gained, it just can be used as a kind of anti-virus formulation, its weak point, the quality product that this method makes is not good enough, poor stability, and certain side effect is arranged.
Another piece is that JP5445161 discloses a kind of immunopotentiator that is used to prevent and treat malignant tumour, and its method is that the component of gained is a kind of immunopotentiator with after golden cell of Staphylococcus or bacterial body pulverizing, filtering out solid ingredient.Can be used for preventing and treating effectively the symptom that the immunity function of cell is lowered, mainly treat viral infection.
Outside the deficiency: bacterium is pulverized earlier and refilters, and filtrate composition complexity filters out solid, contains the thin material of pulverizing in the liquid, is not difficult to expect that side effect is big, though immuno-potentiation is arranged, to the oncotherapy effect with brighten cytosis and do not have effect.
Purpose of the present invention discloses in order to overcome the deficiencies in the prior art part that a kind of technology is simple, reliable in quality, the minimum meta-bolites with streptococcus aureus of toxic side effect are extracted the method for treatment malignant tumour medication.
Purpose of the present invention realizes by following scheme:
Method of the present invention is described by technical process can divide three parts:
First part. the preparation of bacterial classification; Second section. medium preparation; Third part. merge the preparation of inoculation culture, fermentation, extraction, preparation.
The preparation of first part's bacterial classification:
1, recovery is cultivated: streptococcus aureus is inoculated in the nutrient agar medium inclined-plane of lixivium of pig heart preparation cultivated 18 hours for 35 ℃;
2. packing: it is an amount of to get slant culture, evenly grinds in the 1ml skimmed milk and makes the suspension branch aseptic bacterial classification pipe of packing into, every pipe 0.3ml;
3, freeze: the bacterial classification pipe that bacteria suspension will be housed was put in-30 ℃ of low temperature alcohol quick-frozen one hour;
4, low pressure is drained: the bacterial classification pipe after will freezing is put and is contained dry CaCl
2Connect vacuum pump in the vacuum drier, vacuum is drained (needing seven hours approximately);
5, sealing will the above-mentioned bacterial classification pipe of draining be connected on the many menifolds of rubber the about 10-20 of vacuum suction minute, and flame seals the mouth of pipe under vacuum condition.
The preparation of second section substratum:
Main component peptone 0.5-2g%, sodium-chlor 0.3-0.9g%, thiamines 0.0-0.05g%, pig heart infusion 5-20ml%, sheep liver soaking liquid 5-30ml%.
1, pig heart infusion preparation: get fresh pig blood, remove clot, fat, manadesma cleans, cuts with scissors injection broken, that weigh, add qdx with tap water, fresh double distilled water and put refrigerator (4 ℃) 14 hours with fresh double distilled water, boil and used eight layers of husky cloth coarse filtration in two hours while hot, use B quantitative paper suction filtration again, it is standby to clarify yellow liquid.
2, sheep liver soaking liquid is used with quadrat method and is prepared;
3, the amount of making up a prescription as required takes by weighing peptone, sodium-chlor, with fresh double distilled water heated solution;
4, the amount of making up a prescription is measured (absorption) lixivium of pig heart, sheep liver leach liquor and thiamines as required;
5,3.4 solution are mixed, add injection with fresh double distilled water to aequum;
6, transfer pH to 3-4 to add the 0.5g% injection, boil (not stopping to stir) half an hour with powdered high-quality activated carbon;
7, with the decarburization of B quantitative paper suction filtration;
8, transferring pH to 7-9 to add 0.5% injection boils half an hour once more with Powdered high-quality activated carbon;
9, suction filtration decarburization;
10, transfer pH to 6.0-7.5 packing 500ml(cleaning liquor handle, double distilled water clean 180 ℃ do removed thermal source in roasting two hours) saline bottle adds aluminium lid and rolls mouth;
11,15 pounds of sterilizations in 30 minutes of high pressure are standby.
Third part
Inoculation concentration 10000-20 hundred million/ml, ratio 0.5-5ml% cultivates at 30 ℃-38 ℃, 18-72/ hour, adjusts pH6-7.6, millipore filtration 0.22 μ Sterile Filtration.
But the high poly-golden grape (Staphylcococcus aureus CGMCCNo.0165) of this bacterial strain called after is a kind of new bacterial strain of high-efficiency low-toxicity, had been sent to China Committee for Culture Collection of Microorganisms common micro-organisms center before the applying date microbiological specimens is provided.
1. the claimant that microbiological specimens is provided is that the biological system technology of the Shenyang synergism legal representative of company limited is huge surplus, address: No. 8, three good streets, peace zone, Shenyang City, 19 first, March 27 1991 time, preserving number CGMCCNO.0165.Brief description is as follows:
High poly-golden staphylococcus is to separate in the 105 strain staphylococcuses that obtain to screen from the various Pathologic specimens that each section of hospital clinical send Bacteriology Room to check; store method-the vacuum freeze-drying method of bacterial strain; protective material, skimmed milk; should prolonged preservation keep the constant purpose of bacterial strain genetics character again in order to reach; except selecting effectively stable method and cryodesiccated effect being detected at every turn, also to identify the bacterial strain of recovery comprehensively.
Toxicological test:
For reagent liquid is 20 times of concentrated wiring liquids of clinical usefulness " BM828 " injection liquid, small white mouse body weight 17-20 gram/only, female, male half and half is healthy no pregnant, get 20 sex body weight and be divided into two groups at random, every group 10, the normal control group is not given any medicine, after the above-mentioned test liquid 0.5ml of every mice by intraperitoneal injection of the test group administration, small white mouse does not have any upright reaction of sending out, observed seven days, small white mouse is strong deposits, and takes by weighing body weight, disconnected neck place row, dissect visual inspection, each main organs is not seen obvious pathological change, wins the heart, liver, spleen, two kidneys are weighed respectively, calculate the heart-body ratio of each animal, liver-body ratio, spleen-body ratio, related data to normal control group and administration group is carried out test of significance, above-mentioned internal organs by often fixedly making tissue slice, are observed each main organs (heart of control group and administration group under opticmicroscope, liver, spleen, kidney) there is no tangible Histological change.
Conclusion: administration is analyzed the no abnormal finding of each main internal organ of mouse and is compared with the normal control group after seven days, and the mouse speed of growth, the heart-body ratio, liver-body ratio, spleen-body ratio, kidney-body are than also no abnormality seen change of the histological examination of all not having significant difference (P770.05) conscience spleen kidney.
Every mouse of this experiment is all injected 0.5ml, restrains body weight calculating if press 20, and quite 20 times of concentrated wiring liquid 25ml/kg of mice by intraperitoneal injection are converted into injection liquid 500ml/kg, and this dosage is 50000 times of people's clinical dosage by kg body weight calculating.
Clinical trial:
Units such as Chinese Medical Sciences University institute of oncology, 307 hospitals of institute of section of army, tumour portion of Anshan City Central Hospital, the People's Armed Police of Liaoning Province hospital oncology have done clinical application respectively.
For example: tumour inpatient department of Anshan City Central Hospital uses " BM828 " patients such as primary hepatocarcinoma, mammary cancer, colorectal carcinoma, brain tumor, metastasis of cancer lymphoglandula is carried out immunotherapy, tracking monitor acquisition satisfactory result, as after treating 1 month, NK cell (improving index of immunologic function), ground adds the 162-312% solid tumor and dwindles 50%-75%.
1) case is selected:
Li Ming man 31 years old
Suffer from meningioma, perform the operation in total institute of Shenyang ground force in October, 1989, take out two 2 * 3 centimeters tumour, August nineteen ninety the disease relapse second operation, take out 3 * 4 centimeters tumours again, postoperative was less than 4 months, recurrence paralysis is for the third time haveed no alternative but perform the operation for the third time again, and cancer cells extensively shifted and only took down a part of cancer knurl this moment, when patient accepts BM828 after course of treatment, the state of an illness just is under control, and symptom obviously alleviates spirit and takes a turn for the better day by day, the same with the normal people now the feed, can leave the bed and walk, even do some light work.
Anshan 20 routine malignant tumor patients carry out immunotherapy and all obtain promising result.
2) treatment plan:
BM828 is little yellow, and clarifying sterilization biotechnological formulation does not contain sanitas, and is without any side effects, and injection liquid 2ml/ props up, and contains the intramuscular injection of 1000 units, injects once in 1-2 days, and 30 times is a course of treatment.
3) criterion of therapeutical effect:
Produce effects: malignant tumour there is direct killing effect, can also be significantly, improve body's immunological function and suppress indirectly and kill tumour cell, leukocyte increasing, hematoblastic effect.
The data that Anshan central hospital provides, efficient 90%, have no side effect.
Tumour hospital of PLA Academy of Military Sciences witnesses, through the animal test of pesticide effectiveness, and 1500 units of local application/once as a result, inhibitory rate 91%.
4) treatment result:
Xu Hongying: left breast cancer, left axillary lymph is carried down and is moved, and course of treatment of 4 * 3 kilograms of medications of lump, NK cell showed increased reaches 413%, and after lymphoglandula dwindled 2/3, two course of treatment, lump disappeared substantially.
Wang Guisheng: primary hepatocarcinoma, TTPV, high pressure, a large amount of ascites, edema of lower limbs, are all recovered normal ascites and are obviously reduced edema of pair of lower extremities and disappear at two courses of treatment of medication.
Luo Changman: esophagus cancer, thoracic vertebrae shift, after medication two courses of treatment, and NK cell showed increased, the coarse foci disappearance of the esophageal mucosa membrane of admiring.
Li Enquan: the nasopharyngeal carcinoma recurrence, bone shifts, after treating two courses of treatment, the NK cytosis, left rib lump disappears, pain relief, neck is movable to be increased.
Suffice to show that more than BM828 has significant curative effect, compare to have suitable superiority with chemotherapy.
The method for preparing biological regulator with streptococcus aureus having thus described the invention is a kind of method economically and reasonably, because the composition of this method is all got the fine kind, do not add any sanitas, it is again sterilization preparation, without any side effects, its reliable degree particularly adopts the BM828 parenteral solution of preparation of this law preparation considerably beyond general biotechnological formulation, quality and stable curative effect, it is good that lucifuge cool dark place preservation did not precipitate clarity in 2 years.Without any side effects fully through the toxicity Inspection Certificate, the difficult effect that not only confirms to have remarkable enhancing body immunologic function of pharmacology inspection, as normal mouse injection BM828(650 unit/mouse), the activity of natural killer cell (NK cell) increases by 232% before than administration, in suffering from lung cancer in mice, inject, the NK activity is than control group behind this medicine, increase by 304%, lymphocyte transformation rate increase by 158% etc., also very desirable to killing and wounding of malignant cell, the experimentation on animals result confirms lung cancer, cervical cancer, the malignant tumor of digestive tract tumour inhibiting rate has met or exceeded international standard between 30%-40%.
Embodiment: 1 describe in conjunction with the accompanying drawings, accompanying drawing 1 is a process flow sheet of the present invention.Bright for instance step:
The preparation of embodiment 1 first part's bacterial classification:
1, recovery is cultivated: streptococcus aureus CGMCCNO/0165 is inoculated in the nutrient agar medium inclined-plane of lixivium of pig heart preparation cultivated 18 hours for 35 ℃;
2, packing: it is an amount of to get slant culture, evenly grinds in the 1ml skimmed milk and makes the suspension branch aseptic bacterial classification pipe of packing into, every 0.3ml;
3, freeze: the bacterial classification pipe that bacteria suspension will be housed was put in-30 ℃ of low temperature alcohol quick-frozen one hour;
4, low pressure is drained: the bacterial classification pipe after will freezing is put and is contained dry CaCl
2Connect vacuum pump in the vacuum drier, vacuum is drained (needing seven hours approximately);
5, sealing will the above-mentioned bacterial classification pipe of draining be connected on the many menifolds of rubber the about 10-20 of vacuum suction minute, and flame seals the mouth of pipe under vacuum condition.
The preparation of second section substratum:
Main component peptone 0.5g%, sodium-chlor 0.3g%, thiamines 0.0g%, pig heart infusion 5ml%, sheep liver soaking liquid 5ml%.
1, pig heart infusion preparation: get fresh pig blood, remove clot, fat, manadesma cleans, cuts with scissors injection broken, that weigh, add qdx with tap water, fresh double distilled water and put refrigerator (4 ℃) 14 hours with fresh double distilled water, boil two hours while hot with going into the husky cloth coarse filtration of layer, use B quantitative paper suction filtration again, it is standby to clarify yellow liquid.
2, sheep liver soaking liquid is used with quadrat method and is done;
3, the amount of making up a prescription as required takes by weighing peptone, sodium-chlor, with fresh double distilled water heated solution;
4, the amount of making up a prescription is measured (absorption) Pigs Hearts center of immersion liquid, sheep liver leach liquor and thiamines as required;
5,3.4 solution are mixed, add injection with fresh double distilled water to aequum;
6, transfer pH to 3-4 to add the 0.5g% injection, boil (not stopping to stir) half an hour with Powdered high-quality activated carbon;
7, with the decarburization of B quantitative paper suction filtration;
8, transferring pH to 7-9 to add the 0.5g% injection boils half an hour once more with Powdered high-quality activated carbon;
9, suction filtration decarburization;
10, transfer pH to 6.0-7.5 packing 500ml(cleaning liquor handle, double distilled water clean 180 ℃ do removed thermal source in roasting two hours) saline bottle adds aluminium lid and rolls mouth;
11,15 pounds of sterilizations in 30 minutes of high pressure are standby.
Third part
Concentration 10000-20 hundred million/ml is adopted in inoculation, and ratio 0.5ml% cultivates at 30 ℃-38 ℃, during 18-72/, adjusts pH6-7.6 with 0.22 μ millipore filtration degerming 2 times.
The preparation of embodiment 2 first part's bacterial classifications:
1, recovery is cultivated: streptococcus aureus (CGMCCNO.0165) is inoculated in the nutrient agar medium inclined-plane of lixivium of pig heart preparation cultivated 18 hours for 35 ℃;
2, packing: it is an amount of to get slant culture, evenly grinds in the 1ml skimmed milk and makes the suspension branch aseptic bacterial classification pipe of packing into, every 0.3ml;
3, freeze: the bacterial classification pipe that bacteria suspension will be housed was put in-30 ℃ of low temperature alcohol quick-frozen one hour;
4, low pressure is drained: the bacterial classification pipe after will freezing is put and is contained dry Cacl
2Connect vacuum pump in the vacuum drier, vacuum is drained (needing seven hours approximately);
5, sealing will the above-mentioned bacterial classification pipe of draining be connected on the many menifolds of rubber the about 10-20 of vacuum suction minute, and flame seals the mouth of pipe under vacuum condition.
The preparation of second section substratum:
Main component peptone 2g%, sodium-chlor 0.9g%, thiamines 0.05g%, pig heart infusion 20ml%, sheep liver soaking liquid 30ml%.
1, pig heart infusion preparation: get fresh pig blood, remove clot, fat, manadesma cleans, cuts with scissors injection broken, that weigh, add qdx with tap water, fresh double distilled water and put refrigerator (4 ℃) 14 hours with fresh double distilled water, boil and used eight layers of husky cloth coarse filtration in two hours while hot, use B quantitative paper suction filtration again, it is standby to clarify yellow liquid.
2, sheep liver soaking liquid is used with quadrat method and is done;
3, the amount of making up a prescription as required takes by weighing peptone, sodium-chlor, with fresh double distilled water heated solution;
4, the amount of making up a prescription is measured (absorption) pig heart infusion, sheep liver leach liquor and thiamines as required;
5,3.4 solution are mixed, add injection with fresh double distilled water to aequum;
6, transfer pH to 3-4 to add the 0.5g% injection, boil (not stopping to stir) half an hour with Powdered high-quality activated carbon;
7, with the decarburization of B quantitative paper suction filtration;
8, transferring pH to 7-9 to add the 0.5g% injection boils half an hour once more with Powdered high-quality activated carbon;
9, suction filtration decarburization;
10, transfer pH to 6.0-7.5 packing 500ml(cleaning liquor handle, double distilled water clean 180 ℃ do removed thermal source in roasting two hours) saline bottle adds aluminium lid and rolls mouth;
11,15 pounds of sterilizations in 30 minutes of high pressure are standby.
Third part
Concentration 10000-20 hundred million/ml is adopted in inoculation, and ratio 5ml% cultivates at 30 ℃-38 ℃, 18-72 hour, adjusts pH6-7.6, with 0.22 μ millipore filtration degerming secondary.
Claims (2)
1, a kind of meta-bolites with streptococcus aureus prepares the method for microbial preparation, it is characterized in that staphylococcus aureus strains (Staphylococcus aureus CGmCCNO.0165) is inoculated in the lixivium of pig heart nutrient agar, cultivated 18 hours for 35 ℃, to plant daughter bacteria again is inoculated in the fermention medium with the concentration of 10000-20 hundred million/ml, cultivated 18-72 hour for 30 ℃-38 ℃, end to cultivate, adjust pH6-7.6, with 0.22 μ millipore filtration degerming secondary, the packing of bacterium liquid;
2,, it is characterized in that used substratum is peptone 2g%, sodium-chlor 0.9g%, thiamines 0.05g%, pig heart infusion 20ml/%, sheep liver soaking liquid 30ml% according to the described method of claim 1.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN91106067A CN1023567C (en) | 1991-04-01 | 1991-04-01 | Process for extracting malignant tumor medicine from metabolite of staphylococcus aureus |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN91106067A CN1023567C (en) | 1991-04-01 | 1991-04-01 | Process for extracting malignant tumor medicine from metabolite of staphylococcus aureus |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1054190A CN1054190A (en) | 1991-09-04 |
| CN1023567C true CN1023567C (en) | 1994-01-19 |
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Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN91106067A Expired - Lifetime CN1023567C (en) | 1991-04-01 | 1991-04-01 | Process for extracting malignant tumor medicine from metabolite of staphylococcus aureus |
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| Country | Link |
|---|---|
| CN (1) | CN1023567C (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1108170C (en) * | 1996-10-11 | 2003-05-14 | 李绍光 | Buffering liquid for staphylococcus Aureus Injection and stephylococcus Aureus Injection with buffering liquid |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1061537C (en) * | 1996-09-26 | 2001-02-07 | 张茂良 | Monascus preparation for hyperlipemia and relevant cardiac and cerebral diseases |
-
1991
- 1991-04-01 CN CN91106067A patent/CN1023567C/en not_active Expired - Lifetime
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1108170C (en) * | 1996-10-11 | 2003-05-14 | 李绍光 | Buffering liquid for staphylococcus Aureus Injection and stephylococcus Aureus Injection with buffering liquid |
Also Published As
| Publication number | Publication date |
|---|---|
| CN1054190A (en) | 1991-09-04 |
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Free format text: CORRECT: PATENTEE; FROM: XIEHE BIOLOGICAL AGENT TECHNOLOGY CO., LTD., SHENYANG TO: SHENYANG XIEHE GROUP CO.,LTD. |
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Patentee after: Xiehe Group Co., Ltd., Shenyang Patentee before: Xiehe Biological Agent Technology Co., Ltd., Shenyang |
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| C15 | Extension of patent right duration from 15 to 20 years for appl. with date before 31.12.1992 and still valid on 11.12.2001 (patent law change 1993) | ||
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Assignee: Xiehe Biological Pharmaceutical Co., Ltd., Shenyang Assignor: Xiehe Group Co., Ltd., Shenyang Contract fulfillment period: 2009.11.6 to 2011.3.31 Contract record no.: 2009210000271 Denomination of invention: Process for extracting malignant tumor medicine from metabolite of staphylococcus aureus Granted publication date: 19931225 License type: General permission Record date: 20091110 |
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Free format text: COMMON LICENSE; TIME LIMIT OF IMPLEMENTING CONTACT: 2009.11.6 TO 2011.3.31; CHANGE OF CONTRACT Name of requester: SHENYANG XIEHE BIOLOGICAL PHARMACEUTICAL CO., LTD. Effective date: 20091110 |
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Expiration termination date: 20110401 Granted publication date: 19940119 |