CN116036217B - Traditional Chinese medicine compound granule for treating primary liver cancer and preparation method thereof - Google Patents

Traditional Chinese medicine compound granule for treating primary liver cancer and preparation method thereof Download PDF

Info

Publication number
CN116036217B
CN116036217B CN202211664377.8A CN202211664377A CN116036217B CN 116036217 B CN116036217 B CN 116036217B CN 202211664377 A CN202211664377 A CN 202211664377A CN 116036217 B CN116036217 B CN 116036217B
Authority
CN
China
Prior art keywords
parts
liver cancer
traditional chinese
primary liver
chinese medicine
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Active
Application number
CN202211664377.8A
Other languages
Chinese (zh)
Other versions
CN116036217A (en
Inventor
王明刚
李飞燕
毛德文
马家宝
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
First Affiliated Hospital of Guangxi Medical University
Original Assignee
First Affiliated Hospital of Guangxi Medical University
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by First Affiliated Hospital of Guangxi Medical University filed Critical First Affiliated Hospital of Guangxi Medical University
Priority to CN202211664377.8A priority Critical patent/CN116036217B/en
Publication of CN116036217A publication Critical patent/CN116036217A/en
Application granted granted Critical
Publication of CN116036217B publication Critical patent/CN116036217B/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/185Magnoliopsida (dicotyledons)
    • A61K36/80Scrophulariaceae (Figwort family)
    • A61K36/804Rehmannia
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/56Materials from animals other than mammals
    • A61K35/58Reptiles
    • A61K35/586Turtles; Tortoises, e.g. terrapins
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/56Materials from animals other than mammals
    • A61K35/62Leeches; Worms, e.g. cestodes, tapeworms, nematodes, roundworms, earth worms, ascarids, filarias, hookworms, trichinella or taenia
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/06Fungi, e.g. yeasts
    • A61K36/07Basidiomycota, e.g. Cryptococcus
    • A61K36/076Poria
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/185Magnoliopsida (dicotyledons)
    • A61K36/23Apiaceae or Umbelliferae (Carrot family), e.g. dill, chervil, coriander or cumin
    • A61K36/233Bupleurum
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/185Magnoliopsida (dicotyledons)
    • A61K36/25Araliaceae (Ginseng family), e.g. ivy, aralia, schefflera or tetrapanax
    • A61K36/258Panax (ginseng)
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/185Magnoliopsida (dicotyledons)
    • A61K36/28Asteraceae or Compositae (Aster or Sunflower family), e.g. chamomile, feverfew, yarrow or echinacea
    • A61K36/282Artemisia, e.g. wormwood or sagebrush
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/185Magnoliopsida (dicotyledons)
    • A61K36/28Asteraceae or Compositae (Aster or Sunflower family), e.g. chamomile, feverfew, yarrow or echinacea
    • A61K36/284Atractylodes
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/185Magnoliopsida (dicotyledons)
    • A61K36/48Fabaceae or Leguminosae (Pea or Legume family); Caesalpiniaceae; Mimosaceae; Papilionaceae
    • A61K36/484Glycyrrhiza (licorice)
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/185Magnoliopsida (dicotyledons)
    • A61K36/65Paeoniaceae (Peony family), e.g. Chinese peony
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/185Magnoliopsida (dicotyledons)
    • A61K36/71Ranunculaceae (Buttercup family), e.g. larkspur, hepatica, hydrastis, columbine or goldenseal
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/185Magnoliopsida (dicotyledons)
    • A61K36/73Rosaceae (Rose family), e.g. strawberry, chokeberry, blackberry, pear or firethorn
    • A61K36/736Prunus, e.g. plum, cherry, peach, apricot or almond
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/88Liliopsida (monocotyledons)
    • A61K36/896Liliaceae (Lily family), e.g. daylily, plantain lily, Hyacinth or narcissus
    • A61K36/8966Fritillaria, e.g. checker lily or mission bells
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/88Liliopsida (monocotyledons)
    • A61K36/896Liliaceae (Lily family), e.g. daylily, plantain lily, Hyacinth or narcissus
    • A61K36/8967Lilium, e.g. tiger lily or Easter lily
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/88Liliopsida (monocotyledons)
    • A61K36/902Sparganiaceae (Bur-reed family)
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/88Liliopsida (monocotyledons)
    • A61K36/906Zingiberaceae (Ginger family)
    • A61K36/9066Curcuma, e.g. common turmeric, East Indian arrowroot or mango ginger
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00Medicinal preparations characterised by special physical form
    • A61K9/14Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/16Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00Antineoplastic agents
    • YGENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02ATECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Landscapes

  • Health & Medical Sciences (AREA)
  • Natural Medicines & Medicinal Plants (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Veterinary Medicine (AREA)
  • Public Health (AREA)
  • General Health & Medical Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Animal Behavior & Ethology (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Epidemiology (AREA)
  • Engineering & Computer Science (AREA)
  • Mycology (AREA)
  • Biotechnology (AREA)
  • Microbiology (AREA)
  • Medical Informatics (AREA)
  • Botany (AREA)
  • Alternative & Traditional Medicine (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Tropical Medicine & Parasitology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Organic Chemistry (AREA)
  • Medicines Containing Plant Substances (AREA)

Abstract

The invention provides a traditional Chinese medicine compound granule for treating primary liver cancer and a preparation method thereof, belonging to the technical field of traditional Chinese medicines. The preparation method comprises the following steps: dried rehmannia root, capillary artemisia, turmeric, white paeony root, lily, turtle shell, rhizoma atractylodis, radix bupleuri, rhizoma curcumae, poria cocos, liquorice, ginseng, rhizoma sparganii, peach kernel, thunberg fritillary bulb and leech. Compared with the prior art, the preparation method of the traditional Chinese medicine compound granule for treating primary liver cancer can extract the medicine concentration which is 2 times of that of the prior art under the same condition; after the effective concentration of the medicine is obviously improved, the medicine effect of the medicine can be exerted to a greater extent.

Description

Traditional Chinese medicine compound granule for treating primary liver cancer and preparation method thereof
Technical Field
The invention relates to the technical field of traditional Chinese medicines, in particular to a traditional Chinese medicine compound granule for treating primary liver cancer and a preparation method thereof.
Background
According to statistics, the incidence rate of the primary liver cancer is on the 6 th place worldwide, the death rate is on the 3 rd place, the global liver cancer death cases 83.02 ten thousand in 2020, the death number in China accounts for 47% of the world, and the overall survival rate of the primary liver cancer is not more than 20% in 5 years. High morbidity and mortality are basic characteristics of PLC in epidemiology in China. For this reason, the discovery, and demonstration of new effective preventive/therapeutic methods or methods for primary liver cancer remains an urgent and serious scientific research.
Disclosure of Invention
The invention aims to provide a traditional Chinese medicine compound granule for treating primary liver cancer and a preparation method thereof, and compared with the prior art, the traditional Chinese medicine compound granule can extract the medicine concentration which is 2 times of that of the prior art under the same condition; after the effective concentration of the medicine is obviously improved, the medicine effect of the medicine can be exerted to a greater extent, and the weakness of spleen and stomach of a cancer patient is considered, so that the medicine is further subjected to an inclusion process, becomes enteric granules, and increases the tolerance of the patient.
The technical scheme of the invention is realized as follows:
the invention provides a traditional Chinese medicine compound granule for treating primary liver cancer, which is prepared from the following raw materials: dried rehmannia root, capillary artemisia, turmeric, white paeony root, lily, turtle shell, rhizoma atractylodis, radix bupleuri, rhizoma curcumae, poria cocos, liquorice, ginseng, rhizoma sparganii, peach kernel, thunberg fritillary bulb and leech.
As a further improvement of the invention, the invention is prepared from the following raw materials in parts by weight: 5-15 parts of dried rehmannia root, 5-15 parts of virgate wormwood herb, 5-15 parts of turmeric, 5-15 parts of white paeony root, 5-15 parts of lily, 5-15 parts of turtle shell, 5-15 parts of rhizoma atractylodis, 4-8 parts of radix bupleuri, 5-10 parts of rhizoma curcumae, 5-15 parts of poria cocos, 4-8 parts of liquorice, 5-10 parts of ginseng, 5-10 parts of common burreed rhizome, 3-6 parts of peach seed, 5-15 parts of fritillaria thunbergii and 4-8 parts of leech.
As a further improvement of the invention, the invention is prepared from the following raw materials in parts by weight: 10 parts of dried rehmannia root, 10 parts of virgate wormwood herb, 10 parts of turmeric, 10 parts of white paeony root, 10 parts of lily, 6 parts of bupleurum, 5 parts of peach seed, 10 parts of turtle shell, 10 parts of rhizoma atractylodis, 8 parts of zedoary, 10 parts of tuckahoe, 6 parts of liquorice, 8 parts of ginseng, 8 parts of common burreed rhizome, 10 parts of fritillary bulb and 6 parts of leech.
The invention further provides a preparation method of the traditional Chinese medicine compound granule for treating primary liver cancer, which comprises the following steps:
s1, water extraction: weighing dried rehmannia root, turmeric, white peony root, lily, turtle shell, rhizoma atractylodis, radix bupleuri, rhizoma curcumae, poria cocos, liquorice, ginseng, rhizoma sparganii, peach kernel, thunberg fritillary bulb and leech according to a proportion, cleaning, mixing and crushing to obtain traditional Chinese medicine powder, soaking in water for 1-2 hours, heating and boiling, extracting for 50-70 minutes, stopping heating after extraction, filtering to obtain an extract A, adding water into dregs, heating and boiling, extracting for 50-70 minutes, stopping heating after extraction, filtering to obtain an extract B, and mixing the extract A with the extract B to obtain an extract C;
s2, reflux extraction: weighing herba Artemisiae Scopariae, cleaning, mixing herba Artemisiae Scopariae with water, soaking for 1-2 hr, heating and refluxing, extracting for 50-70min, and stopping heating to obtain aromatic water D;
s3, preparation of traditional Chinese medicine: concentrating the extractive solution C under reduced pressure at low temperature to obtain extract, cooling, adding aromatic water D, granulating with adjuvant, and drying to obtain compound Chinese medicinal granule for treating primary liver cancer.
As a further improvement of the present invention, the solid-to-liquid ratio of the traditional Chinese medicine powder to water in step S1 is 1:10-12g/mL.
As a further improvement of the present invention, the solid-to-liquid ratio of the residue to water in step S1 is 1:6-10g/mL.
As a further improvement of the present invention, in step S2, the solid-to-liquid ratio of the herba Artemisiae Scopariae medicinal material to water is 1:4-8g/mL.
As a further improvement of the invention, the low-temperature reduced-pressure concentration in the step S3 is carried out at a temperature of 50-70 ℃ and a pressure of 0.01-0.1MPa, and the concentration is carried out until the relative density is 1.5-1.7.
As a further improvement of the invention, the mass ratio of the extract to the aromatic water D in the step S3 is 1:2-3.
The invention further protects application of the traditional Chinese medicine compound granule for treating primary liver cancer in preparing a medicine for treating and assisting in treating primary liver cancer.
The invention has the following beneficial effects: the dry rehmannia root and the virgate wormwood herb are monarch drugs, the dry rehmannia root is sweet and bitter and cold in nature, and have the effects of expelling blood stasis and replenishing essence; herba Artemisiae Scopariae, bitter in flavor, has the actions of clearing heat and removing toxicity, promoting bile flow and relieving jaundice.
The ministerial drugs comprise carapax Trionycis, rhizoma Curcumae Longae, rhizoma Sparganii, rhizoma Curcumae and semen Persicae; turtle shell, carapax Trionycis, has the effects of relieving stuffy and salty taste, tonifying kidney and kidney, softening hardness and resolving hard mass; turmeric has pungent, bitter and warm taste and has the actions of dredging channels and relieving pain, breaking blood and promoting qi circulation; rhizoma sparganii and rhizoma zedoariae break blood and promote qi circulation; peach kernel has the functions of promoting blood circulation to disperse blood clots.
With the adjuvant materials including Bulbus Lilii, rhizoma Atractylodis, poria, radix Paeoniae alba, hirudo, and radix Ginseng; lily has sweet taste, and is effective in invigorating spleen, inhibiting liver, and relieving pain; rhizoma atractylodis, which can dry dampness and invigorate spleen; poria, having sweet and bland nature, strengthening spleen and excreting dampness; white peony root is bitter and flat, and has the effects of softening liver, relieving pain, removing blood stasis, breaking hard mass and tonifying qi; leech, which breaks blood, removes stasis and dredges channels; ginseng, radix ginseng, and it can strengthen primordial qi. So that bupleurum and liquorice are bitter in taste and enter liver and gall meridians to soothe liver and promote bile flow; the fritillary bulb is used for introducing the medicines in the prescription into focus; licorice root, radix Glycyrrhizae Praeparata, which relieves pain and regulates the drugs. The medicines are combined, so that the Chinese medicinal composition not only can tonify kidney and generate marrow, nourish liver and soften liver and remove liver, invigorate spleen and replenish qi and nourish blood, but also can eliminate toxin and dissipate stagnation, remove blood stasis and detumescence, and treat liver deficiency and liver stasis.
The compound Chinese medicine granule for treating primary liver cancer is a new medicine formed on the basis of the new knowledge of deficiency and liver stasis in the malignant progress of chronic liver disease, and the formula has the effects of tonifying liver deficiency and removing liver stasis, can improve the micro-environment state in/out of liver so as to further relieve or even block the continuous liver injury state, and has outstanding curative effects on treating primary liver cancer. The Chinese herbal compound granule for treating the primary liver cancer provides a new choice for treating the primary liver cancer, and the preparation method of the Chinese herbal compound granule for treating the primary liver cancer provides a better pure Chinese herbal granule preparation for patients with the primary liver cancer.
Compared with the prior art, the preparation method of the traditional Chinese medicine compound granule for treating primary liver cancer can extract the medicine concentration which is 2 times of that of the prior art under the same condition; after the effective concentration of the medicine is obviously improved, the medicine effect of the medicine can be exerted to a greater extent.
Drawings
In order to more clearly illustrate the embodiments of the invention or the technical solutions of the prior art, the drawings which are used in the description of the embodiments or the prior art will be briefly described, it being obvious that the drawings in the description below are only some embodiments of the invention, and that other drawings can be obtained according to these drawings without inventive faculty for a person skilled in the art.
FIG. 1 is a graph of ALT levels for each group in test example 1 of the present invention;
FIG. 2 is a view showing LN levels of each set in test example 1 of the invention;
FIG. 3 is a comparison chart of HE staining of each group in test example 1 according to the present invention;
FIG. 4 is a graph showing apoptosis results of test example 2 according to the present invention;
FIG. 5 is a graph showing the results of the cell migration test in test example 2 according to the present invention;
FIG. 6 is a graph showing the results of the cell invasion assay of test example 2 according to the present invention.
Detailed Description
The following description of the technical solutions in the embodiments of the present invention will be clear and complete, and it is obvious that the described embodiments are only some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
Example 1
The raw materials comprise the following components in parts by weight: 5 parts of dried rehmannia root, 5 parts of virgate wormwood herb, 5 parts of turmeric, 5 parts of white peony root, 5 parts of lily, 5 parts of turtle shell, 5 parts of rhizoma atractylodis, 4 parts of radix bupleuri, 5 parts of rhizoma curcumae, 5 parts of poria cocos, 4 parts of liquorice, 5 parts of ginseng, 5 parts of rhizoma sparganii, 3 parts of peach kernel, 5 parts of fritillaria thunbergii and 4 parts of leech.
The preparation method comprises the following steps:
s1, water extraction: weighing dried rehmannia root, turmeric, white paeony root, lily, turtle shell, rhizoma atractylodis, radix bupleuri, rhizoma curcumae, poria cocos, liquorice, ginseng, rhizoma sparganii, peach kernel, thunberg fritillary bulb and leech according to a proportion, cleaning, mixing and crushing to obtain traditional Chinese medicine powder, soaking the traditional Chinese medicine powder in water for 1h, wherein the solid-to-liquid ratio of the traditional Chinese medicine powder to water is 1:10g/mL, heating and boiling, extracting for 50min, stopping heating after extraction, filtering to obtain an extract A, adding water into residues, wherein the solid-liquid ratio of the residues to the water is 1:6g/mL, heating and boiling, extracting for 50min, stopping heating after extraction, filtering to obtain extract B, and mixing the extract A and the extract B to obtain extract C;
s2, reflux extraction: weighing herba Artemisiae Scopariae, cleaning, and mixing herba Artemisiae Scopariae with water, wherein the solid-liquid ratio of herba Artemisiae Scopariae to water is 1:4g/mL, soaking for 1h, heating and refluxing, extracting for 50min, and stopping heating after extraction to obtain aromatic water D;
s3, preparation of traditional Chinese medicine: concentrating the extract C at low temperature under reduced pressure at 50deg.C under 0.01MPa to obtain extract with relative density of 1.5, cooling, adding aromatic water D with mass ratio of 1:2, granulating with adjuvant, and drying to obtain compound Chinese medicinal granule for treating primary liver cancer.
Example 2
The raw materials comprise the following components in parts by weight: 15 parts of dried rehmannia root, 15 parts of virgate wormwood herb, 15 parts of turmeric, 15 parts of white paeony root, 15 parts of lily, 15 parts of turtle shell, 15 parts of rhizoma atractylodis, 8 parts of radix bupleuri, 10 parts of rhizoma curcumae, 15 parts of poria cocos, 8 parts of liquorice, 10 parts of ginseng, 10 parts of rhizoma sparganii, 6 parts of peach kernel, 15 parts of fritillaria thunbergii and 4-8 parts of leech.
The preparation method comprises the following steps:
s1, water extraction: weighing dried rehmannia root, turmeric, white paeony root, lily, turtle shell, rhizoma atractylodis, radix bupleuri, rhizoma curcumae, poria cocos, liquorice, ginseng, rhizoma sparganii, peach kernel, thunberg fritillary bulb and leech according to a proportion, cleaning, mixing and crushing to obtain traditional Chinese medicine powder, soaking the traditional Chinese medicine powder in water for 2 hours, wherein the solid-to-liquid ratio of the traditional Chinese medicine powder to water is 1:12g/mL, heating and boiling, extracting for 70min, stopping heating after extraction, filtering to obtain an extract A, adding water into residues, wherein the solid-liquid ratio of the residues to the water is 1:10g/mL, heating and boiling, extracting for 70min, stopping heating after extraction, filtering to obtain extract B, and mixing the extract A and the extract B to obtain extract C;
s2, reflux extraction: weighing herba Artemisiae Scopariae, cleaning, and mixing herba Artemisiae Scopariae with water, wherein the solid-liquid ratio of herba Artemisiae Scopariae to water is 1:8g/mL, soaking for 2h, heating and refluxing, extracting for 70min, and stopping heating after extraction to obtain aromatic water D;
s3, preparation of traditional Chinese medicine: concentrating the extract C at a low temperature under reduced pressure at 70deg.C under 0.1MPa to obtain extract with relative density of 1.7, cooling, adding aromatic water D with mass ratio of 1:3, granulating with adjuvant, and drying to obtain compound Chinese medicinal granule for treating primary liver cancer.
Example 3
The raw materials comprise the following components in parts by weight: 10 parts of dried rehmannia root, 10 parts of virgate wormwood herb, 10 parts of turmeric, 10 parts of white paeony root, 10 parts of lily, 6 parts of bupleurum, 5 parts of peach seed, 10 parts of turtle shell, 10 parts of rhizoma atractylodis, 8 parts of zedoary, 10 parts of tuckahoe, 6 parts of liquorice, 8 parts of ginseng, 8 parts of common burreed rhizome, 10 parts of fritillary bulb and 6 parts of leech.
The preparation method comprises the following steps:
s1, water extraction: weighing dried rehmannia root, turmeric, white peony root, lily, turtle shell, rhizoma atractylodis, radix bupleuri, rhizoma curcumae, poria cocos, liquorice, ginseng, rhizoma sparganii, peach kernel, thunberg fritillary bulb and leech according to a proportion, cleaning, mixing and crushing to obtain traditional Chinese medicine powder, soaking the traditional Chinese medicine powder in water for 1.5h, wherein the solid-to-liquid ratio of the traditional Chinese medicine powder to water is 1:11g/mL, heating and boiling, extracting for 60min, stopping heating after extraction, filtering to obtain an extract A, adding water into residues, wherein the solid-liquid ratio of the residues to the water is 1:8g/mL, heating and boiling, extracting for 60min, stopping heating after extraction, filtering to obtain extract B, and mixing the extract A and the extract B to obtain extract C;
s2, reflux extraction: weighing herba Artemisiae Scopariae, cleaning, and mixing herba Artemisiae Scopariae with water, wherein the solid-liquid ratio of herba Artemisiae Scopariae to water is 1:6g/mL, soaking for 1.5h, heating and refluxing, extracting for 60min, and stopping heating after extraction to obtain aromatic water D;
s3, preparation of traditional Chinese medicine: concentrating the extract C at low temperature under reduced pressure at 60deg.C under 0.05MPa to obtain extract with relative density of 1.6, cooling, adding aromatic water D with mass ratio of 1:2.5, granulating with adjuvant, and drying to obtain compound Chinese medicinal granule for treating primary liver cancer.
Test example 1 treatment of chronic liver injury to primary liver cancer in animal model
1.1 model building, grouping scheme, and pharmaceutical intervention
After 65 SD rats are acclimatized, 5 rats are randomly selected as normal group, and the rest 60 rats adopt CCl 4 (40%) rats were intraperitoneally injected to replicate an animal model of persistent chronic liver injury (2 ml/kg), 1 injection every 3 days for 6 weeks. After 6 weeks the remaining rats were randomly divided into: model control, low drug dose, medium drug dose, high drug dose. Low, medium and high doses of drug were administered by intragastric administration beginning at week 6 and drug was administered continuously for 4 weeks. During this period, the liver injury status of rats in the control group and the drug intervention group were maintained according to the model quantity of CCl 4.
1.2 modeling success criteria and general behavioral observations
After molding, rats in the control group had decreased appetite, listlessness, diarrhea, and a part of rats had distended abdomen and significantly reduced body weight. The appetite of rats in the traditional Chinese medicine treatment group is improved, no obvious diarrhea symptom exists, and the overall weight of the rats is increased obviously compared with the control group.
1.3 serological index detection
ELSA detects ALT and LN levels in the periphery of rats in each group; the detection of the levels of glutamic pyruvic transaminase (ALT) and Laminin (LN) in peripheral blood of rats is carried out according to the ELSAkit requirements and the operation steps.
1.4HE detection of pathological changes in liver tissue
Taking paraformaldehyde fixed liver tissue, embedding paraffin after dehydration, slicing (the thickness is about 4 mu m), flattening, pasting on a glass slide, dewaxing, soaking in hematoxylin solution, dyeing with 0.1% eosin dye, soaking in alcohol with different concentration gradients, airing, sealing, and observing under an optical microscope.
1.5 experimental results
The results are shown in FIGS. 1-3. ALT and LN levels in serum of rats in the liver injury group are significantly increased, and the difference compared with the normal group has statistical significance P <0.05; ALT and LN levels were reduced after treatment with the traditional Chinese medicine, and the reduction was evident in the medium dose group and the high dose group, and the difference was statistically significant P <0.05 compared with the control group of the model group. Meanwhile, in the HE staining of liver tissues, the liver lobule structure of a model group rat is observed to be damaged, and liver cells are divided and packed into circular or oval liver cell clusters with different sizes by widely proliferated fibrous tissues, so that the connective tissues are increased to cause the formation of a pseudo lobule structure to replace normal liver tissues. The arrangement of liver cells in the prosthetic lobules is disordered, the liver cells are swollen and deformed, few cell nucleus fragments are involved, a large amount of inflammatory cells infiltrate and gather, fibrous tissue hyperplasia of rat liver tissues after treatment by traditional Chinese medicines is less, the structure of the prosthetic lobules is hidden, the liver cells are arranged neatly, the deformation is less, the infiltration of inflammatory cells is less, and the fibrosis and hardening degree of the model group are obviously reduced.
Annotation: p <0.05 compared to the normal group, +.p <0.05 compared to the model group.
Test example 2 experimental study of inhibition of tumor cell proliferation, invasion and metastasis by drugs
2.1 cell culture
Taking WLR68 cells and HepG2 cell lines, putting the WLR68 cells and the HepG2 cell lines into liquid nitrogen, rapidly putting the WLR68 cells and the HepG2 cell lines into a water bath at 37 ℃, and slightly shaking a freezing tube to dissolve out frozen solution; after lysis, the cells were transferred into a centrifuge tube containing 5ml of medium, collected by centrifugation and centrifuged at 1000rmp for 5min at room temperature, and the supernatant was discarded; culture was carried out with DMEM medium containing 10% fetal bovine serum at 37℃and saturated humidity of 5% CO2, and the medium was changed every two days for subculture 1 time.
2.2 cell treatment
Taking logarithmic growth phase, inoculating 3×10 cells in WLR68 cells and HepG2 cells in 96 well plate 3 Individual/well plates 37 ℃ and 5% co 2 Overnight in the incubator of (a).
2.3 cell CCK8 assay
After the time required for cell culture is over, 10 mu LCCK8 and 37 ℃ are added to each well to culture for 2 hours; the absorbance of each well was measured by a microplate reader OD 450.
2.4 flow apoptosis experiments
Cell collection; cell centrifugation was performed by PBS, and the operation of the Annexin V-FITICPI apoptosis detection kit was described as follows: adding 500 mu LBinding buffer re-suspended cell, adding 5 mu LAnnexinV-FITC, mixing, adding 5 mu LPI, mixing, reacting at room temperature in dark for 5-15min (negative control is set at the same time, namely normal cell is not added with annexinV-FITC and PI), detecting by flow cytometry.
2.5 cell migration experiments
Collecting cells, centrifuging to remove supernatant and washing with PBS; the serum-free MEM medium was resuspended and counted by a cell counting plate to dilute the cell concentration in the serum-free MEM medium to 3X 10 5 A cellml is used; 700. Mu.L of MEM medium (containing double antibody) with 10% FBS was previously added to a 24-well plate, and placed in a small room of a transwell, 200. Mu.L each of the cell suspensions was connected to the upper room of the transwell, and cultured in a CO2 incubator at 37℃for 24 hours; taking out transwell, flushing the cell with PBS, and fixing the cell with 70% ice-ethanol solution for 1h; the cells were not removed on the upper chamber side by wiping with clean cotton balls after staining with 0.5% crystal violet dye solution for 20min at normal temperature and washing with PBS.
2.6 cell invasion assay
Collecting cells, centrifuging to remove supernatant and washing with PBS; the MEM medium resuspended cells and the 1640 medium diluted the cell concentration to 3X 105ml; thawing Matrigel at 4deg.C, transwell chamber, 24-well culture plate, and gun head overnight at-20deg.C; matrigel was diluted to a concentration of 1mgml in medium and run on ice; mu.L of 10% FBSMEM medium (containing double antibody) at 4℃was added to a 24-well plate, and placed in a small room of a transwell, 100. Mu.L of Matrigel at 1mgml concentration was added to the bottom of the upper chamber of the transwell, incubated at 37℃for 4-5 hours, matrigel was dried to gel, the upper chamber of the transwell was connected to 200. Mu.L of cell suspension, and cultured in an incubator at 37℃and 5% CO 2; taking out the transwell, washing the cell by PBS, and fixing the cell in 70% ice-ethanol solution for 1h; and (3) dyeing by using 0.5% crystal violet dye solution, washing by using PBS (phosphate buffer solution) at normal temperature for 20min, and wiping the side surface of the upper chamber by using a cleaning cotton ball, wherein cells are not removed.
3.7 results of the study
The flow apoptosis results show that the apoptosis rate of the 5% drug-containing serogroup is obviously increased and increased with time compared with that of the blank group and the 5% control serogroup, and the apoptosis is obvious. The differences were statistically significant (P < 0.01) (FIG. 4). The medicine is suggested to have promoting effect on apoptosis of primary liver cancer cells.
Cell migration experiment results show that compared with a blank group and a 5% control serum group, the cell number of the 5% drug-containing serum group is the lowest, and the cell migration capacity of the 5% drug-containing serum group is obviously reduced in 48 hours; the difference was statistically significant (P < 0.05) in the 5% drug-containing serogroup compared to the blank (FIG. 5). The medicine has the inhibiting effect on the cell migration of primary liver cancer.
Cell invasion experimental results show that compared with a blank group and a 5% control serogroup, the cell number of the 5% drug-containing serogroup is the lowest, and compared with the 5% drug-containing serogroup at 24 hours, the cell invasion capacity of the 5% drug-containing serogroup at 48 hours is obviously reduced; the difference was statistically significant (P < 0.05) in the 5% drug-containing serogroup compared to the blank group (FIG. 6). The medicine has the inhibiting effect on the invasion of primary liver cancer cells.
Test example 3 cases
Huang Mou A, 57 years old, guangxi Guihong Kong people have the history of primary liver cancer, debilitation, yellow eyes, poor sleep, and after the comprehensive treatment in hospitals, the jaundice is not obvious, the debilitation and yellow eyes are still caused after discharge, and the traditional Chinese medicine preparation prepared in the example 3 is taken 3 times a day, 100g each time, for 2 weeks continuously, the spirit is improved, and the jaundice is fading.
Chen Mou A, 51S, shangxi Shanglin, has a history of primary liver cancer, and has the symptoms of sallow complexion, exacerbation of sudden debilitation, deepening of yellow eye staining, and after the treatment is carried out on the foundation support by tunneling to a hospital, the treatment is continuously taken for more than 1 month after leaving the hospital, the sallow complexion of the patient is relieved after 1 month follow-up, debilitation symptoms are improved, and yellow eye staining is reduced.
Li Mou female, 55 year old Guangxi nan Ning people have history of primary liver cancer, dark complexion, sudden feeling and weakness, yellow complexion, and can be eaten after being taken for one week, the weakness is improved, and the yellow complexion is resolved.
Zhang Mou A, 59 years old, guangxi Yulin, has history of primary liver cancer, pale yellow complexion, yellow eyes, chest and hypochondrium pain, and after administration for 1 week, chest and hypochondrium pain is improved, yellow eyes are relieved, and after administration for 1 month, the spirit is better.
The foregoing description of the preferred embodiments of the invention is not intended to be limiting, but rather is intended to cover all modifications, equivalents, alternatives, and improvements that fall within the spirit and scope of the invention.

Claims (8)

1. A traditional Chinese medicine compound granule for treating primary liver cancer is characterized by being prepared from the following raw materials in parts by weight: 5-15 parts of dried rehmannia root, 5-15 parts of virgate wormwood herb, 5-15 parts of turmeric, 5-15 parts of white paeony root, 5-15 parts of lily, 5-15 parts of turtle shell, 5-15 parts of rhizoma atractylodis, 4-8 parts of radix bupleuri, 5-10 parts of rhizoma curcumae, 5-15 parts of poria cocos, 4-8 parts of liquorice, 5-10 parts of ginseng, 5-10 parts of common burreed rhizome, 3-6 parts of peach seed, 5-15 parts of fritillaria thunbergii and 4-8 parts of leech;
the preparation method comprises the following steps:
s1, water extraction: weighing dried rehmannia root, turmeric, white peony root, lily, turtle shell, rhizoma atractylodis, radix bupleuri, rhizoma curcumae, poria cocos, liquorice, ginseng, rhizoma sparganii, peach kernel, thunberg fritillary bulb and leech according to a proportion, cleaning, mixing and crushing to obtain traditional Chinese medicine powder, soaking in water for 1-2 hours, heating and boiling, extracting for 50-70 minutes, stopping heating after extraction, filtering to obtain an extract A, adding water into dregs, heating and boiling, extracting for 50-70 minutes, stopping heating after extraction, filtering to obtain an extract B, and mixing the extract A with the extract B to obtain an extract C;
s2, reflux extraction: weighing herba Artemisiae Scopariae, cleaning, mixing herba Artemisiae Scopariae with water, soaking for 1-2 hr, heating and refluxing, extracting for 50-70min, and stopping heating to obtain aromatic water D;
s3, preparation of traditional Chinese medicine: concentrating the extractive solution C under reduced pressure at low temperature to obtain extract, cooling, adding aromatic water D, granulating with adjuvant, and drying to obtain compound Chinese medicinal granule for treating primary liver cancer.
2. The traditional Chinese medicine compound granule for treating primary liver cancer according to claim 1, which is characterized by being prepared from the following raw materials in parts by weight: 10 parts of dried rehmannia root, 10 parts of virgate wormwood herb, 10 parts of turmeric, 10 parts of white paeony root, 10 parts of lily, 6 parts of bupleurum, 5 parts of peach seed, 10 parts of turtle shell, 10 parts of rhizoma atractylodis, 8 parts of zedoary, 10 parts of tuckahoe, 6 parts of liquorice, 8 parts of ginseng, 8 parts of common burreed rhizome, 10 parts of fritillary bulb and 6 parts of leech.
3. The compound traditional Chinese medicine granule for treating primary liver cancer according to claim 1, wherein the solid-to-liquid ratio of the traditional Chinese medicine powder to water in the step S1 is 1:10-12g/mL.
4. The compound traditional Chinese medicine granule for treating primary liver cancer according to claim 1, wherein the solid-to-liquid ratio of the dregs to water in the step S1 is 1:6-10g/mL.
5. The compound traditional Chinese medicine granule for treating primary liver cancer according to claim 1, wherein the solid-to-liquid ratio of the herba Artemisiae Scopariae medicinal material to water in the step S2 is 1:4-8g/mL.
6. The compound Chinese medicinal granule for treating primary liver cancer according to claim 1, wherein the low-temperature reduced-pressure concentration in step S3 is performed at 50-70 ℃ and 0.01-0.1MPa under a relative density of 1.5-1.7.
7. The compound Chinese medicinal granule for treating primary liver cancer according to claim 1, wherein the mass ratio of the extract to the aromatic water D in the step S3 is 1:2-3.
8. Use of the compound Chinese medicinal granule for treating primary liver cancer according to any one of claims 1-7 in preparing medicaments for treating and assisting in treating primary liver cancer.
CN202211664377.8A 2022-12-23 2022-12-23 Traditional Chinese medicine compound granule for treating primary liver cancer and preparation method thereof Active CN116036217B (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN202211664377.8A CN116036217B (en) 2022-12-23 2022-12-23 Traditional Chinese medicine compound granule for treating primary liver cancer and preparation method thereof

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN202211664377.8A CN116036217B (en) 2022-12-23 2022-12-23 Traditional Chinese medicine compound granule for treating primary liver cancer and preparation method thereof

Publications (2)

Publication Number Publication Date
CN116036217A CN116036217A (en) 2023-05-02
CN116036217B true CN116036217B (en) 2023-11-21

Family

ID=86112464

Family Applications (1)

Application Number Title Priority Date Filing Date
CN202211664377.8A Active CN116036217B (en) 2022-12-23 2022-12-23 Traditional Chinese medicine compound granule for treating primary liver cancer and preparation method thereof

Country Status (1)

Country Link
CN (1) CN116036217B (en)

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101757560A (en) * 2009-12-16 2010-06-30 张�林 Medicament for treating liver cancer and preparation method thereof
CN109172781A (en) * 2018-11-22 2019-01-11 致宁医药(肇庆)股份有限公司 A kind of Chinese medicine composition for treating hepatitis B, cirrhosis
CN110269896A (en) * 2019-08-07 2019-09-24 广西中医药大学第一附属医院 A kind of Chinese medicine composition for preventing and treating chronic liver disease and jaundice
CN110420304A (en) * 2019-08-26 2019-11-08 贵州中医药大学第一附属医院 A kind of drug and preparation method thereof for the benign tumors sexual stage

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101757560A (en) * 2009-12-16 2010-06-30 张�林 Medicament for treating liver cancer and preparation method thereof
CN109172781A (en) * 2018-11-22 2019-01-11 致宁医药(肇庆)股份有限公司 A kind of Chinese medicine composition for treating hepatitis B, cirrhosis
CN110269896A (en) * 2019-08-07 2019-09-24 广西中医药大学第一附属医院 A kind of Chinese medicine composition for preventing and treating chronic liver disease and jaundice
CN110420304A (en) * 2019-08-26 2019-11-08 贵州中医药大学第一附属医院 A kind of drug and preparation method thereof for the benign tumors sexual stage

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
中西医结合治疗早期乙肝肝硬化的疗效观察;岑成灿 等;广西中医药;第39卷(第03期);34-35 *
自拟疏肝健脾化瘀散结方治疗肝癌2例;王涵;内蒙古中医药;-(第05期);第48页左栏第1段、右栏第2-3段 *

Also Published As

Publication number Publication date
CN116036217A (en) 2023-05-02

Similar Documents

Publication Publication Date Title
CN102078473B (en) Traditional Chinese medicine preparation for treating chronic hepatitis b and preparation method thereof
CN101596230B (en) Medicinal composition for treating hepatopathy
CN102048902B (en) Hepatitis treating traditional Chinese medicine composition, extract and preparation method, application and formulation
CN116036217B (en) Traditional Chinese medicine compound granule for treating primary liver cancer and preparation method thereof
CN104784549A (en) Traditional Chinese medicine for treating allergic rhinitis
CN103705578A (en) Traditional Chinese medicine preparation with functions of reducing blood fat and inhibiting blood glucose increase and preparation method thereof
CN103877323B (en) Medicinal composition for treating non-alcoholic fatty liver disease
CN104383050B (en) A kind of medicine of combination of inner and outside treatment rheumatic arthritis and preparation method
CN113116976A (en) Application of traditional Chinese medicine composition in preparation of medicine for preventing or treating digestive internal diseases
CN101642541B (en) Composition with hepatoprotective effect, preparation method and application thereof
CN102198251B (en) Chinese medicinal composition for treating climacteric syndrome and preparation method thereof
CN105663439A (en) Pharmaceutical composition for preventing and treating diabetes and preparation method thereof
CN111569017A (en) Suppository for treating ulcerative colitis
CN105194427A (en) Traditional Chinese medicine composition for treating xerophthalmia and preparation method of traditional Chinese composition
Zhang et al. Progress in the treatment of chronic glomerulonephritis with traditional Chinese medicine
CN104083548A (en) Traditional Chinese medicinal composition for treating ovarian cyst and preparation method thereof
CN116421675B (en) Traditional Chinese medicine composition for treating non-alcoholic fatty liver disease and application thereof
CN115475200B (en) Chinese herbal compound preparation for treating liver fibrosis and preparation method thereof
CN107224557A (en) It is a kind of to be used to treat decoction medicine of chronic prostatitis and preparation method thereof
CN114796417B (en) Blood sugar reducing traditional Chinese medicine formula and preparation method thereof
CN101966324A (en) Chinese medicinal compound preparation for treating chronic hepatitis B liver fibrosis and preparation method thereof
CN102526548B (en) Traditional Chinese medicine composition for treating chronic hepatitis and hepatic fibrosis and preparation method thereof
CN101766749A (en) Chinese patent medicine for treating chronic rhinitis and nasosinusitis
CN105943860A (en) Medicine preparation for treating diabetes mellitus and preparation method thereof
CN117180365A (en) Pharmaceutical composition for preventing and/or treating breast cancer, preparation method and application thereof

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
GR01 Patent grant
GR01 Patent grant