WO2022057499A1 - 一种组合物及其制备方法 - Google Patents

一种组合物及其制备方法 Download PDF

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Publication number
WO2022057499A1
WO2022057499A1 PCT/CN2021/110930 CN2021110930W WO2022057499A1 WO 2022057499 A1 WO2022057499 A1 WO 2022057499A1 CN 2021110930 W CN2021110930 W CN 2021110930W WO 2022057499 A1 WO2022057499 A1 WO 2022057499A1
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parts
composition
food
astragalus
medicine
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PCT/CN2021/110930
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English (en)
French (fr)
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肖伟
丁艳
王婧
邓辰辰
柏伟荣
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江苏康缘药业股份有限公司
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Publication of WO2022057499A1 publication Critical patent/WO2022057499A1/zh

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/64—Orobanchaceae (Broom-rape family)
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/105—Plant extracts, their artificial duplicates or their derivatives
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/48—Fabaceae or Leguminosae (Pea or Legume family); Caesalpiniaceae; Mimosaceae; Papilionaceae
    • A61K36/481—Astragalus (milkvetch)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/63—Oleaceae (Olive family), e.g. jasmine, lilac or ash tree
    • A61K36/638—Ligustrum, e.g. Chinese privet
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/81—Solanaceae (Potato family), e.g. tobacco, nightshade, tomato, belladonna, capsicum or jimsonweed
    • A61K36/815—Lycium (desert-thorn)
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
    • A61K2236/30—Extraction of the material
    • A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
    • A61K2236/331—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones using water, e.g. cold water, infusion, tea, steam distillation or decoction
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
    • A61K2236/30—Extraction of the material
    • A61K2236/39—Complex extraction schemes, e.g. fractionation or repeated extraction steps
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
    • A61K2236/50—Methods involving additional extraction steps
    • A61K2236/51—Concentration or drying of the extract, e.g. Lyophilisation, freeze-drying or spray-drying

Definitions

  • the invention relates to the technical fields of medicines, health products and food, in particular to a composition for enhancing immunity and a preparation method thereof.
  • Immunodeficiency is divided into primary and secondary categories. Primary is caused by congenital hypoplasia, most of which are related to heredity, and mostly occur in children; secondary is caused by viruses, bacteria, fungi and other infections or drugs, tumors, fatigue, insomnia, malnutrition, excessive stress, etc. Caused by people of all ages. Most of them are frail after illness and susceptible to infectious diseases are secondary immunocompromised.
  • the present invention provides a composition and a preparation method thereof.
  • the present invention proposes a composition, which can be directly ground into powder, or can be an extract or other forms prepared by conventional means, and includes, by weight: 1 to 8 parts of Cistanche deserticola , 1 to 8 parts of Astragalus, 1 to 8 parts of Ligustrum lucidum, and 1 to 8 parts of wolfberry.
  • the raw material drug of the present invention can be directly ground into powder, or can be an extract or other forms prepared by conventional means.
  • the composition comprises: 1-4 parts of Cistanche, 1-4 parts of Astragalus, 1-4 parts of Ligustrum lucidum, and 1-4 parts of Lycium barbarum.
  • the composition comprises: 1-2 parts of Cistanche, 1-2 parts of Astragalus, 1-2 parts of Ligustrum lucidum, and 1-2 parts of Lycium barbarum.
  • the composition includes 1 part of Cistanche deserticola, 1 part of Astragalus, 1 part of Ligustrum lucidum, and 1 part of Lycium barbarum.
  • the composition comprises: 2-3 parts of Cistanche deserticola, 2-3 parts of Astragalus, 2-3 parts of Ligustrum lucidum, and 2-3 parts of Lycium barbarum.
  • the composition comprises 2 parts of Cistanche, 3 parts of Astragalus, 2 parts of Ligustrum lucidum, and 3 parts of Lycium barbarum.
  • the present invention also provides a medicine, health care product or food containing the composition described in any of the foregoing.
  • the present invention also proposes the application of the composition as described in any one of the foregoing in the preparation of immunity-enhancing medicines, health-care products or food.
  • the medicine, health product or food further includes auxiliary materials; the auxiliary materials include diluents, disintegrants, and lubricants.
  • the diluent includes microcrystalline cellulose and corn starch.
  • the disintegrating agent includes one or more of sodium starch glycolate and croscarmellose sodium.
  • the lubricant includes one or more of magnesium stearate, talc and silicon dioxide.
  • the present invention also proposes a preparation method of any of the aforementioned compositions, comprising:
  • the medicine, food or health product is an oral preparation, preferably a capsule.
  • the present invention also proposes a preparation method of the aforementioned medicine, food or health product.
  • the preparation method comprises the following steps: (1) taking Cistanche, Astragalus, Ligustrum lucidum and Lycium barbarum, add water 8 to 12 times, reflux for 1 to 3 times, and filter for 0.5 to 2 hours each time to obtain a filtrate;
  • the granules are mixed with disintegrants and lubricants, and the capsules are filled after mixing.
  • the preparation method comprises the following steps:
  • Extraction, concentration and drying process remove impurities from the medicinal materials, weigh the medicinal materials of Cistanche deserticola, Astragalus, Ligustrum lucidum and Lycium barbarum according to the proportion, add 8 to 12 times the amount of water, reflux for 1 to 3 times, 0.5 to 2 hours each time, and combine The filtrate was filtered and concentrated under reduced pressure at 60-70°C to obtain a thick paste (60-70°C) with a relative density of 1.20-1.25. At 60 ⁇ 70 °C, dry under reduced pressure, pulverize into fine powder.
  • Granulation The prepared soft material is granulated by a rocking granulator.
  • Drying Spread the wet granules on the baking tray, put them on the drying car and push them into the box-type drying room, dry at 55 ⁇ 5°C, control the moisture content of the granules to be below 5.0%, and naturally cool to room temperature.
  • Granulation dry granules are granulated with a 15-25 mesh sieve using a rocking granulator. After granulation, add croscarmellose sodium and silicon dioxide to mix well.
  • the preparation method comprises the following steps:
  • Extraction, concentration and drying process remove impurities from the medicinal materials, weigh the medicinal materials of Cistanche deserticola, Astragalus, Ligustrum lucidum and Lycium barbarum according to the proportion, add 10 times the amount of water, reflux for extraction twice, 1.0h each time, combine the filtrates, filter, Concentrate under reduced pressure at 60-70°C to obtain a thick paste (60-70°C) with a relative density of 1.20-1.25. At 60 ⁇ 70 °C, dry under reduced pressure, pulverize into fine powder.
  • Granulation The prepared soft material is granulated with a 20-mesh sieve with a rocking granulator.
  • Drying Spread the wet granules on the baking tray, put them on the drying car and push them into the box-type drying room, dry at 55 ⁇ 5°C, control the moisture content of the granules to be below 5.0%, and naturally cool to room temperature.
  • Granulation dry granules are granulated with a 20-mesh sieve with a rocking granulator, and after granulation, croscarmellose sodium and silicon dioxide are added to mix well.
  • the composition of the invention is prepared from Cistanche deserticola, Astragalus, Ligustrum lucidum and Lycium barbarum as main raw materials, and has the function of enhancing human immunity.
  • the word “immunity” first appeared in my country in the Ming Dynasty's “Immune Prescriptions”, which means “immune disease”.
  • the essence of traditional Chinese medicine “reinvigorating the righteousness and eliminating pathogenic factors” refers to people-oriented, and the selection of prescriptions with the effect of tonifying qi and nourishing blood to supplement the deficiency of yin and yang, qi and blood, and body fluid in the human body, thereby enhancing the immune function of the human body.
  • composition of the present invention follows the theory of traditional Chinese medicine in my country.
  • Cistanche deserticola is warm in nature, enters the kidney meridian and the large intestine meridian, directly enters the kidney family, is warm and moisturizing, warms the essence and blood and unclogs the yang qi, nourishes the five internal organs, strengthens the yin, and benefits Essence, for five strains and seven injuries, treats both the symptoms and root causes, and is the king medicine.
  • Astragalus is warm in nature, invigorating the qi of the lungs and spleen, invigorating the qi of the lung and spleen, invigorating the body surface to stop sweating, and tonifying the qi of the muscle surface, especially suitable for people with fatigue, lack of energy, loss of appetite, etc. caused by spleen deficiency; Liver and kidney meridians, soothe the five internal organs, treat waist and leg pain, mainly to clear the meridians and blood, and nourish the spirit; wolfberry is sweet and moisturizing, nourishing the nature, and can invigorate the kidney and qi. Thought to be medicine.
  • the above four herbs reflect the formula characteristics of nourishing the five organs, nourishing qi and nourishing blood, and strengthening the body.
  • composition of the invention is aimed at the pathogenesis of low immunity, combined with traditional Chinese medicine and modern medical knowledge, the formula has clear functions, no incompatibility, safe and effective consumption, and can play a good role in health care.
  • the raw medicinal materials of the composition of the present invention are scientifically compatible with the raw materials of the same origin of medicine and food issued by the State Administration or that can be used as health food.
  • the added amount of excipients is in line with the use standard of food additives and the Chinese Pharmacopoeia.
  • the operation process of the invention the extraction process is orthogonally verified, the operation is simple, the economy is environmentally friendly, and the process is stable.
  • the types and additions of excipients have been screened and tested, which are in line with the characteristics of this product and can improve the quality of the product.
  • This product has good safety and has been proved by animal experiments to enhance immunity. This product and process have broad application prospects.
  • the C factor is significant, and is determined as C2 according to the energy saving principle; the B factor is not significant, and it is determined as B1 according to the energy saving principle; the A factor is not significant, considering the transfer rate of each index component, it is respectively A2 and A3, according to the principle of energy saving, is determined as A2; therefore, the best extraction process is A2B1C2, that is, adding 10 times the amount of water each time, extracting twice, each time for 1.0 hours.
  • microcrystalline cellulose is easy to granulate, but the disintegration time is longer; the corn starch disintegration time is shorter. , but there is agglomeration during granulation, so microcrystalline cellulose + corn starch is selected as auxiliary materials at the same time.
  • auxiliary materials will affect the granulation process, so the dosage of microcrystalline cellulose and corn starch is investigated. According to weighing three parts of dry paste powder, each 200g, add different proportions of microcrystalline cellulose and corn starch respectively. The bulk density and angle of repose of the material are shown in Table 10.
  • the traditional Chinese medicine plaster becomes sticky when exposed to water, which affects the infiltration of water during the disintegration process of the capsule, resulting in difficulty in disintegration. Therefore, it is necessary to increase the disintegrant to improve the disintegration situation.
  • Sodium carboxymethyl cellulose is generally added in an amount of 2% to 5%, and 90% ethanol is used as a wetting agent to prepare a suitable soft material, granulated, dried, and granulated. The granules are added with disintegrants and lubricants. Mixing, filling capsules, and measuring the disintegration time limit, the results are shown in Table 12.
  • lubricant In order to ensure that the filling is carried out smoothly and the filling weight difference meets the regulations, it is necessary to add a certain amount of lubricant to the particles.
  • Commonly used lubricants are magnesium stearate, talc, and micropowder silica gel, and the general dosage is about 1-2%.
  • Other factors in the fixed formula were granulated according to the above-mentioned preferred formula, and 1% and 2% of magnesium stearate, talc, and silicon dioxide were added respectively, and the angle of repose and tablet smoothness were used as evaluation indicators. The results are shown in Table 13.
  • Table 13 shows that it is better to use 2% silica as the lubricant.
  • auxiliary materials added to this product are: 20% of microcrystalline cellulose and 10% of corn starch by mass of dry paste powder, 90% ethanol is used as a wetting agent for granulation, and 3% cross-linked carboxylate is used for granulation.
  • Sodium methylcellulose was used as disintegrant, and 2% silicon dioxide was used as lubricant.
  • the recommended dose of the sample prepared in Example 3 is 2 times a day, 2 capsules each time, and each capsule is 0.5 g, that is, the recommended human dose is 4 g/person/day. If the adult body weight is 60kg, the recommended dose of the sample is equivalent to 0.067g/kg ⁇ bw.
  • SPF grade ICR mice were purchased from Beijing Weitong Lihua Laboratory Animal Technology Co., Ltd., the experimental animal license: SCXK (Beijing) 2016-0006, and were raised in the SPF grade animal experimental center (license number: SYXK (Black) 2017-008 ).
  • the clean-grade feed used for the experimental animals was purchased from Beijing Keao Xieli Feed Co., Ltd. (license number: SCXK (Beijing) 2014-0010).
  • 40 healthy female mice, weighing 18-22 g were randomly divided into 4 groups with 10 mice in each group.
  • As the immune group carbon clearance test and animal viscera/body ratio were performed.
  • Forty healthy female mice, weighing 18-22 g were randomly divided into 4 groups, 10 mice in each group.
  • mice 40 healthy female mice, weighing 18-22 g, were randomly divided into 4 groups with 10 mice in each group.
  • the delayed-type allergic reaction test was performed.
  • the experiment of phagocytosis of chicken erythrocytes by mouse peritoneal macrophages was performed.
  • ConA-induced spleen lymphocyte transformation test and NK cell activity assay were performed.
  • the recommended dose of this sample is 2g per day, 4 capsules each time, each capsule is 0.5g, that is, the recommended human dose is 4g/person/day. If the adult body weight is 60kg, the recommended dose of the sample is equivalent to 0.067g/kg ⁇ bw. Taking 10 times the recommended human body, that is, 0.67g/kg body weight per day as the medium dose, there is a dose group at the top and bottom: 2.01g/kg body weight (30 times the recommended human body dose is the high dose) and 0.34g/kg body weight (human body weight). 5 times the recommended dose is the low dose).
  • Sheep red blood cells SRBC
  • chicken red blood cells normal saline, complement (guinea pig serum), SA buffer, Dulbecco's reagent, Indian ink, Na2CO3, Hank's solution, RPMI1640 complete medium, calf serum, penicillin, oxidized form Coenzyme, 0.2mol/L Tris-HCl, YAC-1 cells, 1% NP40, 2-mercaptoethanol (2-ME), penicillin, streptomycin, concanavalin A (ConA), hydrochloric acid, isopropanol, MTT, PBS buffer (pH7.2-7.4), acetone, methanol, Giemsa staining solution.
  • SRBC Sheep red blood cells
  • chicken red blood cells normal saline, complement (guinea pig serum), SA buffer, Dulbecco's reagent, Indian ink, Na2CO3, Hank's solution, RPMI1640 complete medium, calf serum, penicillin,
  • the spleen of the animals was aseptically taken, placed in a small dish containing an appropriate amount of sterile Hank's solution, and the spleen was gently torn with forceps to prepare a single cell suspension. Filtered through a 200-mesh sieve, washed twice with Hank's solution, centrifuged for 10 min each time (1000 r/min), discarded the supernatant, bounced the cytoplasm, added 0.5 mL of sterilized water for 20 s to lyse the red blood cells, and then added 0.5 mL2 times of Hank's solution and 8mL Hank's solution, 1000r/min, 10min centrifugation. Finally, the cells were suspended with 1.0 mL RPMI1640 complete medium, and the number of viable cells was counted by trypan blue staining (should be above 95%).
  • spleen cell stock solution prepared above was diluted with RPMI1640 complete culture medium to make the cell concentration 3 ⁇ 106 cells/mL.
  • the diluted cell suspension was added to a 24-well culture plate in two wells, 1 mL per well, 75 ⁇ L of ConA solution was added to one well, and the other well was used as a control, and cultured in 5% CO2 at 37°C for 72 hours.
  • NK% (reaction hole OD-natural release hole OD)/(maximum release hole OD-natural release hole OD) ⁇ 100%
  • Animals (immunization group 2) After 4 to 5 days of immunization, the eyeballs were removed and blood was collected in a centrifuge tube, placed for about 1 hour, the coagulated blood was stripped from the wall of the tube, the serum was separated, and the serum was collected by centrifugation at 2000 rpm for 10 minutes. Dilute the serum 250 times with SA buffer, take 1 mL of it and put it in a test tube, and add 10% (V/V, prepared with SA buffer) in turn by volume of 0.5 mL of SRBC and 1 mL of complement (diluted at 1:10 with SA buffer) . A control tube without serum was also set up (replaced with SA buffer).
  • Sample HC 50 sample optical density value/optical density value at half hemolysis of SRBC ⁇ dilution factor
  • mice Animals (immunized group) mice were injected with 0.1 mL/10 g body weight of Indian ink diluted 1:3.5 times by tail vein, and the timing was immediately after the ink was injected.
  • mice were weighed and sacrificed by cervical dislocation. The liver, spleen and thymus were removed to remove the fascia, and the blood stains on the surface of the organs were blotted with filter paper and weighed.
  • the 722 spectrophotometer was used to measure the optical density (OD) at a wavelength of 600 nm, and the Na 2 CO 3 solution was used as a blank control to calculate the phagocytic index ⁇ .
  • Animals (immune groups of four) were injected intraperitoneally with 1 mL of 20% chicken red blood cell suspension at 30min intervals. The animals were killed by cervical dislocation, and they were fixed on the mouse plate in supine position. The abdominal wall skin was cut in the middle, and normal saline was injected intraperitoneally.
  • the phagocytosis index of the middle and high dose groups was significantly different (P ⁇ 0.01), indicating that the samples could increase the carbon clearance function of mice.
  • the samples of different doses were administered orally to mice for 30 days, after statistical processing, the phagocytosis rate of the four groups was tested for the homogeneity of variance, and the variance was unequal, and the variables were transformed.
  • P is the phagocytosis rate, expressed as a decimal.
  • the phagocytosis index of each dose group was significantly different, P ⁇ 0.01. It shows that the sample can enhance the phagocytic function of mouse peritoneal macrophages.

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Abstract

一种增强免疫力的组合物及其制备方法以及含有该组合物的药物、保健品或食品，该组合物包括：肉苁蓉1～8份，黄芪1～8份，女贞子1～8份，枸杞子1～8份。该组合物可增强小鼠的迟发型变态反应及碳廓清吞噬指数，增强腹腔巨噬细胞的吞噬功能。

Description

一种组合物及其制备方法 技术领域
本发明涉及药品、保健品和食品技术领域，具体而言，涉及一种增强免疫力的组合物及其制备方法。
背景技术
免疫力低下分为原发性和继发性两类。原发性是先天发育不全所致，大多数与遗传有关，多发生于儿童；继发性则由病毒、细菌、真菌等感染或药物、肿瘤、疲劳、失眠、营养不良、压力过大等原因引起，可见于各种年龄的人群。多数病后体弱，易发感染性疾病者属继发性免疫力低下。
现代人生活和工作压力大，很多人处于亚健康状态，有些人出现了头晕、失眠、易疲劳等继发性免疫力低下的症状。表现为易生病，因经常患病，加重机体的消耗，所以一般有体质虚弱、营养不良、精神萎靡、疲乏无力、食欲降低、睡眠障碍等表现。每次生病都要很长时间才能恢复，而且常反复发作。除加强锻炼外，还可通过饮食调理提高人体免疫能力。
发明内容
为了解决上述存在的人体免疫力低下的问题，本发明提供了一种组合物及其制备方法。
具体来说，本发明提出了一种组合物，所述组合物可以直接研磨成粉，也可以是经过常规手段制得的提取物或其它形态等，以重量计，包括：肉苁蓉1～8份，黄芪1～8份，女贞子1～8份，枸杞子1～8份。 其中，本发明的原料药可以直接研磨成粉，也可以是经过常规手段制得的提取物或其它形态等。
优选地，所述组合物包括：肉苁蓉1～4份，黄芪1～4份，女贞子1～4份，枸杞子1～4份。
优选地，所述组合物包括：肉苁蓉1～2份，黄芪1～2份，女贞子1～2份，枸杞子1～2份。
优选的，所述组合物包括肉苁蓉1份，黄芪1份，女贞子1份，枸杞子1份。
优选地，所述组合物包括：肉苁蓉2～3份，黄芪2～3份，女贞子2～3份，枸杞子2～3份。
优选地，所述组合物包括肉苁蓉2份，黄芪3份，女贞子2份，枸杞子3份。
本发明还提出了一种含有如前任一所述的组合物的药物、保健品或食品。
本发明还提出了如前任一所述的组合物在制备增强免疫力药物、保健品或食品中的应用。
优选地，所述药物、保健品或食品还包括辅料；所述辅料包括稀释剂、崩解剂、润滑剂。
优选地，所述稀释剂包括微晶纤维素和玉米淀粉。所述崩解剂包括羧甲淀粉钠、交联羧甲基纤维素钠的一种或几种。所述润滑剂包括硬脂酸镁、滑石粉、二氧化硅的一种或几种。
本发明还提出了一种前述任一所述的组合物的制备方法，包括：
(1)取药材加入8～12倍量水回流提取1～3次，每次0.5～2.0小时，过滤，合并提取液，得到滤液；
(2)将所得滤液在55℃～85℃条件下减压浓缩，并浓缩至相对密度1.10～1.30，在55℃～85℃减压干燥，粉碎后过筛，制成干膏粉。所述药物、食品或保健品为口服制剂，优选为胶囊剂。
本发明还提出了一种如前所述的所述药物、食品或保健品的制备方法，所述药物、保健品或食品选自胶囊剂时，所述制备方法包括 以下步骤：(1)取肉苁蓉、黄芪、女贞子和枸杞子，加水8～12倍，回流提取1～3次，每次0.5～2h过滤，得滤液；
(2)将滤液减压浓缩，并浓缩至相对密度1.20～1.25，减压干燥，粉碎，过65目筛，制成干膏粉，加入稀释剂、90％～95％乙醇，制粒，干燥后整粒；
(3)整粒后颗粒加入崩解剂和润滑剂混合，混合后填充胶囊。
优选地，所述制备方法包括以下步骤：
1.提取浓缩和干燥工艺：药材除杂，按照比例称取肉苁蓉、黄芪、女贞子和枸杞子药材，加8～12倍量水，回流提取1～3次，每次0.5～2h，合并滤液，滤过，60～70℃减压浓缩，得相对密度为1.20～1.25的稠膏(60～70℃)。在60～70℃，减压干燥，粉碎成细粉。
2.混合：按配方称量干膏粉、微晶纤维素、玉米淀粉，投入湿法混合颗粒机中混合，加入适量90～95％乙醇，搅拌制成适宜软材。
3.制粒：制备好的软材用摇摆式颗粒机制粒。
4.干燥：湿颗粒平铺在烘盘上，置于烘车上推入箱式烘房中，55±5℃干燥，控制颗粒水分在5.0％以下，自然冷却至室温。
5.整粒：干颗粒用摇摆式颗粒机，以15～25目筛整粒，整粒后加入交联羧甲基纤维素钠、二氧化硅混匀。
6.充填：干颗粒先调节好填充量，然后填充胶囊。
优选地，所述制备方法包括以下步骤：
1.提取浓缩和干燥工艺：药材除杂，按照比例称取肉苁蓉、黄芪、女贞子和枸杞子药材，加10倍量水，回流提取2次，每次1.0h，合并滤液，滤过，60～70℃减压浓缩，得相对密度为1.20～1.25的稠膏(60～70℃)。在60～70℃，减压干燥，粉碎成细粉。
2.混合：按配方称量干膏粉、微晶纤维素、玉米淀粉，投入湿法混合颗粒机中混合，加入适量90％乙醇，搅拌制成适宜软材。
3.制粒：制备好的软材用摇摆式颗粒机，以20目筛网制粒。
4.干燥：湿颗粒平铺在烘盘上，置于烘车上推入箱式烘房中，55±5℃干燥，控制颗粒水分在5.0％以下，自然冷却至室温。
5.整粒：干颗粒用摇摆式颗粒机，以20目筛整粒，整粒后加入 交联羧甲基纤维素钠、二氧化硅混匀。
6.充填：干颗粒先调节好填充量，然后填充胶囊，胶囊为0.5g/粒。
本发明组合物以肉苁蓉、黄芪、女贞子和枸杞子为主要原料制成，具有增强人体免疫的功能。“免疫”一词我国最早出现于明朝《免疫类方》，是指“免除疫疡”的意思。中国传统医学精髓“扶正祛邪”即是指以人为本，选用具有补气益血作用的方药，以补充人体阴阳、气血、津液等不足，从而增强人体免疫功能。
本发明组合物遵循我国传统中医药理论，方中肉苁蓉性温，归肾经、大肠经，直入肾家，温而能润，能温养精血而通阳气，养五脏，强阴，益精气，针对五劳七伤，标本兼治，为君药。黄芪性温，补肺脾之气，益卫固表以止汗，且善补肌表之气，尤其适宜脾气虚引起的疲倦乏力、精神萎靡、食欲不振等人群；女贞子性平，入肝、肾经，安五脏，治腰腿疼，以通经和血、养精神为主；枸杞子甘平而润，性滋补、能补肾益气，乃平补之药；三药共助君药以为臣药。
以上四味药，体现养五腑，补气养血，扶正固本的配方特点。
本发明组合物是针对免疫力低下的发病机理，结合中药和现代医学知识，组方功能明确，无配伍禁忌，食用安全有效，能起到很好的保健作用。
具体实施方式
本发明组合物的原料药材选用国家局颁布的药食同源或可用保健食品的原料，科学配伍而成。辅料的添加量符合食品添加剂使用标准和中国药典。本发明的操作工艺，提取工艺经正交验证，操作简单，经济环保，工艺稳定。辅料的种类和添加量经筛选实验，符合本产品的特征，能改善产品的质量。本产品安全性良好，具有经动物实验证明，有增强免疫力功效。本产品和工艺具有广阔的应用前景。
实施例1
1.提取工艺选择
每份取肉苁蓉药材80g、黄芪药材80g、枸杞子药材80g、女贞子药材80g，共取9份，按因素水平表1相关参数要求进行提取，测定提取液中松果菊苷和毛蕊花糖苷、黄芪甲苷的转移率和粗多糖含量。以松果菊苷和毛蕊花糖苷、黄芪甲苷转移率和粗多糖含量为考察指标，采用L 9(3 4)正交试验，对提取溶剂量、提取时间、提取次数进行考察。
表1 因素水平
Figure PCTCN2021110930-appb-000001
表2 L 9(3 4)正交表(松果菊苷、毛蕊花糖苷)
Figure PCTCN2021110930-appb-000002
Figure PCTCN2021110930-appb-000003
表3 方差分析表(松果菊苷、毛蕊花糖苷)
Figure PCTCN2021110930-appb-000004
表4 L 9(3 4)正交表(黄芪甲苷)
Figure PCTCN2021110930-appb-000005
表5 方差分析表(黄芪甲苷)
Figure PCTCN2021110930-appb-000006
Figure PCTCN2021110930-appb-000007
表6 L 9(3 4)正交表(粗多糖)
Figure PCTCN2021110930-appb-000008
表7 方差分析表(粗多糖)
Figure PCTCN2021110930-appb-000009
从表2和3可以看出，以提取液中松果菊苷和毛蕊花糖苷转移率为指标进行考察，各因素对提取工艺的影响极差R值为；C>B>A，提取次数(因素C)的极差最大，各因素中C2>C1>C3、B3>B1>B2、A2>A3>A1，故直观分析最佳提取工艺为A2B3C2。方差分析显示， C因素显著，故最佳提取工艺为A2B1C2，即每次加10倍量水，回流提取2次，每次1.0小时。从表4和5可以看出，以提取液中黄芪甲苷转移率为指标进行考察，各因素对提取工艺的影响极差R值为；C>A>B，提取次数(因素C)的极差最大，各因素中C1>C2>C3、B3>B2>B1、A2>A3>A1，故直观分析最佳提取工艺为A2B3C1。方差分析显示，C因素显著，故最佳提取工艺为A2B1C1，即每次加10倍量水，回流提取3次，每次1.0小时。从表6和7可以看出，以提取液中粗多糖含量为指标进行考察，各因素对提取工艺的影响极差R值为；C>A>B，提取次数(因素C)的极差最大，各因素中C2>C1>C3、B1>B3>B2、A3>A2>A1，故直观分析最佳提取工艺为A3B1C2。方差分析显示，C因素显著，故最佳提取工艺为A3B1C2，即每次加12倍量水，回流提取2次，每次1.0小时。
三个指标成分综合考虑，C因素显著，根据节能原则，确定为C2；B因素均不显著，根据节能原则，确定为B1；A因素均不显著，考虑各指标成分转移率，分别为A2和A3，根据节能原则，确定为A2；故最佳提取工艺为A2B1C2，即每次加10倍量水，提取2次，每次1.0小时。
2.提取工艺验证试验
根据正交优选的结果，每份取肉苁蓉药材120g、黄芪药材120g、枸杞子药材120g、女贞子药材120g，平行3份，10倍量水，回流提取2次，每次1.0h，提取液滤过，合并滤液，测定提取液的松果菊苷和毛蕊花糖苷、黄芪甲苷、粗多糖的含量，以其转移率为指标验证提取工艺参数的可行性及稳定性，结果见表8。
表8提取工艺验证实验表
Figure PCTCN2021110930-appb-000010
验证实验结果表明，采用优选后的提取工艺对肉苁蓉和黄芪进行 提取，提取液中松果菊苷和毛蕊花糖苷、黄芪甲苷的转移率和粗多糖含量与正交试验结果接近，说明了该提取工艺稳定可行。
实施例2
1.药材提取浓缩和干燥工艺
按照比例称取肉苁蓉3.0kg、黄芪3.0kg、女贞子3.0kg和枸杞子3.0kg，加1200L的水，回流提取2次，每次1.0h。过滤，滤液在60℃条件下减压浓缩，得浓缩稠膏相对密度为1.25。继续在70℃条件下减压干燥，粉碎，过65目筛，制成干膏粉，干膏粉重量为7.95kg。
2.辅料种类和用量的选择
2.1辅料种类的选择
经过试验研究，需加入适当的辅料，以克服在制剂过程中结块、流动性差、粒重差异大、崩解困难等问题。在稀释剂的选择过程中，我们根据中间体的性质，选择微晶纤维素、玉米淀粉、糊精三种制药常用辅料，对三种辅料进行试验，实验结果见表9。
表9 辅料的选择及实验结果
Figure PCTCN2021110930-appb-000011
由实验结果表1可知，选用不同的辅料，对制剂的制粒后物料状态产生一定的影响，采用微晶纤维素的容易制粒，但崩解时间较长；采用玉米淀粉崩解时间较短，但制粒时有结块现象，因此同时选择微晶纤维素+玉米淀粉作辅料。
2.2辅料用量的选择
辅料用量会影响制粒工艺的进行，因此对微晶纤维素和玉米淀粉的用量进行考察，按照称取三份干膏粉，各200g，分别加入不同比 例的微晶纤维素和玉米淀粉，考察物料的堆密度、休止角，实验结果见表10。
表10 辅料用量选择及试验结果
Figure PCTCN2021110930-appb-000012
由实验结果表10可知，加入干膏粉质量的20％的微晶纤维素和10％的玉米淀粉时，制粒情况最好，且崩解时间合格。
2.3润湿剂的筛选
分别以体积分数70％、80％、90％、95％的乙醇为润湿剂制软材，20目筛挤压制粒，60℃干燥，再用20目筛整粒，收集颗粒。以制软材情况、软材过筛情况、颗粒状态为指标，考察不同体积分数乙醇的制粒效果，结果见表3。
表11 润湿剂考察结果
Figure PCTCN2021110930-appb-000013
由实验结果表11可知，采用90％乙醇作为润湿剂制粒最优。
2.4崩解剂的选择
中药膏粉遇水变粘，影响胶囊崩解过程中水分渗入，造成崩解困难，因此需要增加崩解剂以改善崩解情况，用制剂生产中常用的崩解剂羧甲淀粉钠和交联羧甲基纤维素钠，加入量一般为2％～5％，以90％乙醇为润湿剂，制成适宜的软材，制粒，干燥，整粒，颗粒加入崩解剂和润滑剂，混匀，充填胶囊，测定崩解时限，结果见表12。
表12 崩解剂考察情况
Figure PCTCN2021110930-appb-000014
实验结果表12表明，在相同辅料用量情况下，交联羧甲基纤维素钠对胶囊的崩解效果较好，因此综合辅料对充填胶囊及崩解的影响，优选3％添加量交联羧甲基纤维素钠作为崩解剂。
2.5润滑剂的筛选
为了保证充填顺利进行，充填重差异符合规定，需要在颗粒中加入一定量的的润滑剂。常用的润滑剂为硬脂酸镁、滑石粉、微粉硅胶，一般用量约为1～2％。固定配方中其它因素，按照上述优选的配方制粒，分别加入1％和2％硬脂酸镁、滑石粉、二氧化硅，以颗粒休止角和片剂光洁度为评价指标，结果见表13。
表13 润滑剂筛选表
Figure PCTCN2021110930-appb-000015
由实验结果表13表明，以2％二氧化硅作为润滑剂较好。
因此，通过实验可知，本次产品添加辅料为：干膏粉质量的20％的微晶纤维素和10％的玉米淀粉，采用90％乙醇作为润湿剂来制粒， 以3％交联羧甲基纤维素钠作为崩解剂，以2％二氧化硅作为润滑剂。
实施例3
按照比例称取肉苁蓉54kg、黄芪54kg、女贞子54kg和枸杞子54kg，加2160L的水，回流提取2次，每次1.0h。过滤，滤液在70℃条件下减压浓缩，得浓缩稠膏相对密度为1.20。继续在70℃条件下减压干燥5小时，粉碎，过65目筛，制成干膏粉，干膏粉重量为47.73kg，然后再在干膏粉中按照比例加入辅料，加入微晶纤维素9.82kg、加入玉米淀粉5.04kg，混合均匀后，用90％乙醇湿润，过20目筛，制粒，60℃干燥5小时，过20目筛，整粒，颗粒加入崩解剂交联羧甲基纤维素钠1.92kg和润滑剂1.30kg，混合，得颗粒重量约63.24kg，颗粒得率约为96.1％。灌装胶囊，即得胶囊剂，胶囊每粒重0.5g。
实施例4
增强免疫力功能动物实验
1.材料与方法
1.1样品
实施例三制得的样品推荐剂量为每日2次，每次2粒，每粒0.5g，即人体推荐剂量为4g/人/天。成人体重为60kg计，则样品推荐剂量相当于0.067g/kg·bw。
1.2试验动物与条件
SPF级ICR小鼠购自北京维通利华实验动物技术有限公司，实验动物许可证：SCXK(京)2016-0006，在SPF级动物实验中心饲养(许可证号：SYXK(黑)2017-008)。试验动物所用的清洁级饲料购自北京科澳协力饲料有限公司(许可证号：SCXK(京)2014-0010)。选择健康雌性小鼠40只，体重18～22g，随机分为4组，每组10只，作为免疫一组，进行碳廓清试验及动物脏/体比值测定。健康雌性小鼠鼠40只，体重18～22g，随机分为4组，每组10只，作为免疫二组，进行半数溶血值(HC50)测定和抗体生成细胞检测。健康雌性小鼠 40只，体重18～22g，随机分为4组，每组10只，作为免疫三组，进行迟发型变态反应试验。健康雌性小鼠40只，体重18～22g，随机分为4组，每组10只，作为免疫四组，进行小鼠腹腔巨噬细胞吞噬鸡红细胞试验。健康雌性小鼠40只，体重18～22g，随机分为4组，每组10只，作为免疫五组，进行ConA诱导的小鼠脾淋巴细胞转化试验和NK细胞活性测定。
1.3剂量设计
该样品推荐剂量为每日2g，每次4粒，每粒0.5g，即人体推荐剂量为4g/人/天。成人体重为60kg计，则样品推荐剂量相当于0.067g/kg·bw。以人体推荐的10倍，即每日0.67g/kg体重为中剂量，上下各设一个剂量组：2.01g/kg体重(人体推荐量的30倍为高剂量)和0.34g/kg体重(人体推荐量的5倍为低剂量)。分别称取样品5.025g、1.675g、0.85g，用蒸馏水溶解并定容至50mL；同时设置对照组(蒸馏水)，动物则按照体重的2％灌胃容积进行灌胃，试验组溶剂均为蒸馏水，连续灌胃30天后测定各项免疫指标。
1.4仪器与试剂
动物天平、分析天平、冰箱、离心机、722分光光度计、显微镜、洁净工作台、二氧化碳培养箱、酶标仪等。无菌手术器械、游标卡尺、微量注射器(25μL)、细胞计数器、纱布、试管、计时器、血色素吸管、24孔培养板等。
绵羊红细胞(SRBC)、鸡红细胞、生理盐水、补体(豚鼠血清)、SA缓冲液、都氏试剂、印度墨汁、Na2CO3、Hank’s液、RPMI1640完全培养液、小牛血清、青链霉素、氧化型辅酶、0.2mol/L的Tris-HCl、YAC-1细胞、1％NP40、2-巯基乙醇(2-ME)、青霉素、链霉素、刀豆蛋白A(ConA)、盐酸、异丙醇、MTT、PBS缓冲液(pH7.2～7.4)、丙酮、甲醇、Giemsa染液。
1.5试验方法
1.5.1 ConA诱导的小鼠脾淋巴细胞转化试验和NK细胞活性测定
1.5.1.1脾细胞悬浮液制备
动物(免疫五组)无菌取脾，置于盛有适量无菌Hank’s液的小平皿中，用镊子轻轻将脾撕碎，制成单个细胞悬液。经200目筛网过滤，用Hank’s液洗2次，每次离心10min(1000r/min)，弃上清液，将细胞浆弹起，加入0.5mL灭菌水20s，裂解红细胞，然后再加入0.5mL2倍的Hank’s液及8mLHank’s液，1000r/min，10min离心。最后用1.0mLRPMI1640完全培养液将细胞悬浮，用台酚兰染色计数活细胞数(应在95％以上)。
1.5.1.2淋巴细胞增殖反应(MTT法)
用上述制备的脾细胞原液适量，用RPMI1640完全培养液稀释使细胞浓度为3×106个/mL。将稀释后细胞悬液分两孔加入24孔培养板中，每孔1mL，一孔加入75μLConA液，另一孔作为对照，置5％CO2，37℃培养72h。培养结束前4h，每孔轻轻吸取上清液0.7mL，加入0.7mL不含小牛血清的RPMI1640完全培养液，同时加入MTT(5mg/mL)完全溶解，然后分装到96孔培养板中，每个孔做3个平行孔，用酶标仪，以570nm波长测定值。
1.5.1.3 NK细胞活性测定(LDH测定法)
另取上述脾细胞原液适量，用RPMI1640完全培养液稀释，使细胞浓度为2×107个/mL作为效应细胞浓度，再据此调整靶细胞(YAC-1)浓度，使效靶比为50:1。
取靶细胞和效应细胞各100μL，靶细胞最大释放孔加靶细胞和1％NP40各100μL，上述各项均设三个平行孔，37℃、5％CO2培养箱中培养4h，将96孔板以1500rpm离心5分钟，每孔吸取上清液100μL置酶标板上，加入LDH基质液100μL，反应10min，然后每孔加入1mol/L的HCl溶液30μL终止反应，在酶标仪490nm波长处测定OD值计算NK细胞活性。
NK％＝(反应孔OD-自然释放孔OD)/(最大释放孔OD-自然释放孔OD)×100％
1.5.2迟发型变态反应(DTH)
将免疫三组动物每只鼠经腹腔注射2％(V/V，用生理盐水配制)压积SRBC(2000rpm，10分钟)0.2mL，致敏后4天，测量左后足跎部厚度，用同一部位测量三次，取平均值。然后在测量部位皮下注射20％(V/V，用生理盐水配制)压积SRBC20μL，于注射后24小时测量左后足跎部厚度，以攻击前后足跎厚度的差值(足跎肿胀度)来表示DTH程度。
1.5.3半数溶血值(HC 50)的测定和抗体生成细胞检测
1.5.3.1动物免疫
在试验结束前4～5天进行动物免疫。取脱纤维的羊血，用生理盐水洗涤3次，每次离心2000r/min，10min。将压积SRBC用生理盐水配成2％(V/V)的细胞悬液，每只鼠腹腔注射0.2mL。
1.5.3.2半数溶血值(HC 50)的测定
动物(免疫二组)动物免疫4～5天后，摘除眼球取血于离心管中，放置约1小时，将凝固血与管壁剥离，使血清充分析出，2000rpm离心10分钟，收集血清。用SA缓冲液将血清稀释250倍，取1mL置试管内，依次加入10％(V/V，用SA缓冲液配制)压积SRBC0.5mL、补体1mL(用SA缓冲液按1:10稀释)。另设不加血清的对照管(以SA缓冲液代替)。置37℃恒温水浴中保温30min后，冰浴终止反应。2000rpm离心10min，取上清液1mL、加都氏试剂3mL，同时取10％(V/V，用生理盐水配制)压积SRBC0.25mL、加都氏试剂至4mL于另一试管中，充分混匀，放置10min，于540nm波长处以对照管作为空白，分别测定各管光密度值。溶血素的量以半数溶血值(HC 50)表示，按下式计算。
样品HC 50＝样品光密度值/SRBC半数溶血时的光密度值×稀释倍数
1.5.3.3抗体生成细胞检测
1.5.3.3.1脾细胞悬浮液制备(同前1.5.1.1)
1.5.3.3.2空斑的测定
将表层培养基(1g琼脂糖加双蒸水至100mL)加热溶解后，放45℃～50℃水浴保温，与等量pH7.2～7.4 2倍浓度的Hank’s液混合，分装小试管，每管0.5mL，再向管内加50μL 10％SRBC(V/V，用SA缓冲液配制)，20μL脾细胞悬液(5×106个/mL)，迅速混匀，倾倒于已刷琼脂糖薄层的玻璃片上，待琼脂凝固后，将玻片水平扣放再片架上，放入二氧化碳培养箱中温育1～1.5h，然后用SA缓冲液稀释的补体(1:8)加入到玻片架凹槽内，继续温育1～1.5h后，计数溶血空斑数。
1.5.4小鼠碳廓清试验和脏/体比值的测定
1.5.4.1小鼠碳廓清试验
动物(免疫一组)小鼠按0.1mL/10g体重尾静脉注射1:3.5倍稀释的印度墨汁，待墨汁注入，立即计时。
注入墨汁后2、10min，分别从内眦静脉丛取血20μL，将其加到2mL0.1％Na 2CO 3溶液中。将小鼠称重后颈椎脱臼处死，取肝脏、脾脏和胸腺去尽筋膜，用滤纸吸干脏器表面血污，称重。
用722分光光度计在600nm波长处测光密度值(OD)，以Na 2CO 3溶液作空白对照，计算吞噬指数α。
1.5.5小鼠腹腔巨噬细胞吞噬鸡红细胞试验
1.5.5.1鸡红细胞悬液制备
取鸡血置于有玻璃珠的锥形瓶中，朝一个方向充分摇动，以脱纤维。用生理盐水洗涤2～3次，离心(2000r/min，10min)，去上清，用生理盐水配成20％(V/V)的鸡红细胞悬液。
1.5.5.2吞噬功能的测定
将动物(免疫四组)每只鼠腹腔注射20％的鸡红细胞悬液1mL，间隔30min，颈椎脱臼处死动物，将其仰位固定于鼠板上，正中剪开腹壁皮肤，经腹腔注入生理盐水2mL，转动鼠板1min，然后吸入腹腔洗液1mL，平均滴于2片载玻片上，放入垫有湿纱布的搪瓷盒内，移置37℃孵箱温育30min，孵毕，于生理盐水中漂洗，以除去未贴片 细胞，晾干，以1:1丙酮甲醇溶液固定，4％(V/V)Giemsa-磷酸缓冲液染色3min，再用蒸馏水漂洗晾干。结果以吞噬百分率或吞噬指数来表示小鼠巨噬细胞的吞噬功能。
2.1对小鼠体重的影响
表14 对小鼠体重的影响(免疫一组)
Figure PCTCN2021110930-appb-000016
Figure PCTCN2021110930-appb-000017
表15 对小鼠体重的影响(免疫二组)
Figure PCTCN2021110930-appb-000018
Figure PCTCN2021110930-appb-000019
表16 对小鼠体重的影响(免疫三组)
Figure PCTCN2021110930-appb-000020
Figure PCTCN2021110930-appb-000021
表17 对小鼠体重的影响(免疫四组)
Figure PCTCN2021110930-appb-000022
Figure PCTCN2021110930-appb-000023
表18 对小鼠体重的影响(免疫五组)
Figure PCTCN2021110930-appb-000024
Figure PCTCN2021110930-appb-000025
由表14、15、16、17、18结果可见，各免疫组实验中，低、中、高三个剂量组的初始体重和终末体重与对照组相比较均无显著性差异(P＞0.05)，表明本实验条件下，样品对小鼠体重增长无影响。
表19 对小鼠脏器/体重比值的影响
Figure PCTCN2021110930-appb-000026
Figure PCTCN2021110930-appb-000027
经口给予不同剂量的样品30天，脾脏/体重比值、胸腺/体重比值 在低、中、高剂量组与对照组间均无显著性差异(P＞0.05)，表明对脏器/体重比值无影响。
表20 对小鼠迟发型变态反应(DTH)的影响
Figure PCTCN2021110930-appb-000028
Figure PCTCN2021110930-appb-000029
经口给予不同剂量的样品30天，其足跎肿胀度在注射SRBC前后差值，中、高剂量组与对照组间比较有极显著性差异(P＜0.01)，表明能增强小鼠的迟发型变态反应。
表21 对小鼠脾淋巴细胞转化的影响
Figure PCTCN2021110930-appb-000030
Figure PCTCN2021110930-appb-000031
经口给予不同剂量的样品30天，各剂量组的小鼠脾淋巴细胞转化功能与对照组比较无显著性差异(P＞0.05)，表明样品不能增强小鼠脾淋巴细胞转化功能。
表22 对小鼠版数溶血值(HC 50)的影响
Figure PCTCN2021110930-appb-000032
Figure PCTCN2021110930-appb-000033
经口给予不同剂量的样品30天，中、高剂量组的半数溶血值(HC 50)与对照组比较有极显著性差异(P＜0.01)，低剂量组的半数溶血值(HC 50)与对照组比较有显著性差异(P＜0.05)，表明样品能提高小鼠半数溶血值(HC 50)。
表23 对小鼠抗体生成细胞的影响
Figure PCTCN2021110930-appb-000034
Figure PCTCN2021110930-appb-000035
经口给予不同剂量的样品30天，各剂量组空斑数与对照组比较无显著性差异(P＞0.05)，表明样品不能增强小鼠抗体生成细胞功能。
表24 对小鼠碳廓清功能的影响
Figure PCTCN2021110930-appb-000036
Figure PCTCN2021110930-appb-000037
中高剂量组吞噬指数与对照组比较有极其显著性差异(P＜0.01)，表明样品能增加小鼠碳廓清功能。
表25 对小鼠巨噬细胞吞噬鸡红细胞的影响
Figure PCTCN2021110930-appb-000038
Figure PCTCN2021110930-appb-000039
经口给予小鼠不同剂量的样品30天经统计学处理，四组吞噬率进行方差齐性检验，方差不齐，进行变量变换
Figure PCTCN2021110930-appb-000040
式中P为吞噬率，用小数表示。各剂量组的吞噬指数与对照组比较有极显著性差异P＜0.01。表明样品能增强小鼠腹腔巨噬细胞的吞噬功能。
表26 对小鼠小鼠NK细胞活性的影响
Figure PCTCN2021110930-appb-000041
Figure PCTCN2021110930-appb-000042
经口给予小鼠不同剂量的实施例三制得的样品30天，将NK细胞活性进行变量变换
Figure PCTCN2021110930-appb-000043
式中P为NK细胞活性，用小数表示。经统计学处理，各剂量的NK细胞活性与对照组比较均无显著性差异(P＞0.05)，表明样品对小鼠NK细胞活性功能无影响。
结论：经口投予小鼠2.01g/kg·bw、0.67g/kg·bw、0.34g/kg·bw(分别为人体推荐剂量的30倍、10倍和5倍)的样品30天，结果表明，中、高剂量组的样品可显著增强小鼠的迟发型变态反应及碳廓清吞噬指数，低、中、高剂量组的样品可提高小鼠的半数溶血值和高剂量组能增强腹腔巨噬细胞的吞噬功能。根据“保健食品功能学评价程序和检验方法”中的判定标准，认为样品具有增强免疫力的功能。
以上所述仅为本申请的优选实施例而已，并不用于限制本申请，对于本领域的技术人员来说，本申请可以有各种更改和变化。凡在本申请的精神和原则之内，所作的任何修改、等同替换、改进等，均应包含在本申请的保护范围之内。

Claims (10)

  1. 一种组合物，其特征在于，以重量计，所述组合物包括：肉苁蓉1～8份，黄芪1～8份，女贞子1～8份，枸杞子1～8份。
  2. 根据权利要求1所述的组合物，其特征在于，以重量计，所述组合物包括：肉苁蓉1～4份，黄芪1～4份，女贞子1～4份，枸杞子1～4份。。
  3. 根据权利要求1所述的组合物，其特征在于，以重量计，所述组合物包括：肉苁蓉1份、黄芪1份、女贞子1份，枸杞子1份。
  4. 根据权利要求1所述的组合物，其特征在于，以重量计，所述组合物包括肉苁蓉2份、黄芪3份、女贞子2份，枸杞子3份。
  5. 根据权利要求1-4任一所述的组合物在制备增强免疫力药物、保健品或食品中的应用。
  6. 一种含有权利要求1-4任一所述的组合物的药物、保健品或食品。
  7. 根据权利要求6所述的药物、保健品或食品，其特征在于，所述药物、保健品或食品还包括辅料；所述辅料包括稀释剂、崩解剂、润滑剂。
  8. 根据权利要求7所述的药物、保健品或食品，其特征在于，
    所述稀释剂包括微晶纤维素和玉米淀粉；
    所述崩解剂包括羧甲淀粉钠、交联羧甲基纤维素钠的一种或几种；
    所述润滑剂包括硬脂酸镁、滑石粉、二氧化硅的一种或几种。
  9. 一种如权利要求1-4任一所述的组合物的制备方法，其特征在于，包括：
    (1)取药材加入8～12倍量水回流提取1～3次，每次0.5～2.0小时，过滤，合并提取液，得到滤液；
    (2)将所得滤液在55℃～85℃条件下减压浓缩，并浓缩至相对密度1.10～1.30，在55℃～85℃减压干燥，粉碎后过筛。
  10. 一种如权利要求6所述药物、保健品或食品的制备方法，所述药物、保健品或食品选自胶囊剂时，其特征在于，所述制备方法包括以下步骤：
    (1)取肉苁蓉、黄芪、女贞子和枸杞子，加水8～12倍，回流提取1～3次，每次0.5～2h过滤，得滤液；
    (2)将滤液减压浓缩，并浓缩至相对密度1.20～1.25，减压干燥，粉碎，过65目筛，制成干膏粉，加入稀释剂、90％～95％乙醇，制粒，干燥后整粒；
    (3)整粒后颗粒加入崩解剂和润滑剂混合，混合后填充胶囊。
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CN111529598A (zh) * 2020-06-29 2020-08-14 王亚科 一种养元补气中药组合物及其制备方法
CN112057522A (zh) * 2020-09-18 2020-12-11 江苏康缘药业股份有限公司 一种组合物及其制备方法

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