A kind of Panax notoginseng puree and preparation method thereof

Abstract

The present invention proposes a Panax notoginseng puree and a preparation method and application thereof, and belongs to the field of medical technology. The preparation method comprises: washing the whole Panax notoginseng plant, sterilizing and inactivating enzymes, crushing and juicing, filtering and concentrating the juice to obtain a preliminary puree, adding a composite enzyme to hydrolyze the solid residue, inoculating Ganoderma lucidum seeds, selenium-enriched yeast and zinc-enriched yeast, fermenting and culturing, extracting and separating by an immobilized ionic liquid / tea saponin composite system, analyzing the solid residue to obtain a secondary puree, and concentrating the extract after dialysis to obtain a tertiary puree, which is uniformly mixed with the secondary puree and the preliminary puree, filtered, sterilized, and packaged to obtain the Panax notoginseng puree. The present invention can not only make full use of the whole plant resources of Panax notoginseng, improve the utilization rate of raw materials, but also improve the extraction rate of effective ingredients, enhance the medicinal value of the product, and has a good market application prospect. The puree prepared by the present invention has stable quality and is easy to store and transport.

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to a Panax notoginseng puree and a preparation method thereof. Background Art

[0002] Panax notoginseng is a perennial herbaceous plant of the genus Panax, Araliaceae, containing notoginseng total saponins (PNS), alcohols, flavonoids, nucleosides, alkaloids, proteins, vitamin C and other compounds and more than 18 kinds of inorganic elements, among which notoginseng total saponins are one of the main active ingredients. It has the effects of stopping bleeding, reducing swelling and relieving pain, and dispersing blood stasis; it mainly treats hemoptysis, traumatic bleeding, epistaxis, and traumatic injuries.

[0003] There are many processing technologies for Panax notoginseng in the prior art, but most of them use processes such as water extraction and microwave extraction to extract the total saponins in Panax notoginseng, and then prepare oral liquid or beverage with other additives or other raw materials. Some relevant documents in the prior art include:

[0004] CN1084369A, Panax notoginseng flower health drink; CN1185925A, Panax notoginseng distilled water health drink; CN1302565A, Panax notoginseng flower tea; CN1306780A, Panax notoginseng flower tea carbonated beverage; CN103082056A A fresh Panax notoginseng oral liquid and its preparation method; CN116098288A A Panax notoginseng oral liquid for lowering blood pressure and its preparation method; CN114748525A A preparation method of oral liquid containing Panax notoginseng and its product; Patent CN93102 The method disclosed in 0026 is to crush ginseng and squeeze it to obtain a pulp, which is not a whole plant pulp product. The ginseng residue left behind contains nutrients and effective ingredients; the method disclosed in patent CN105147588A is to mix ginseng dry powder, water and Bifidobacterium bifidum, and ferment to obtain ginseng biological fermentation liquid; the method disclosed in patent CN103446208A is to obtain ginseng liquid with partially hydrolyzed cellulose; the method disclosed in patent CN103446209A is to obtain ginseng pulp after enzymatic hydrolysis with α-amylase. The method disclosed in CN108904563B is a processing method and use of a whole plant fresh ginseng (forest mountain ginseng) pulp product, which is to directly pulp, filter and concentrate to obtain a pulp product.

[0005] The prior art has the following deficiencies and defects:

[0006] 1. The extraction process of Panax notoginseng total saponins is lengthy and cumbersome, the utilization rate of raw materials is extremely low, the waste is large, and the cost performance of the entire process is very low;

[0007] 2. Even in the existing technology of hot water extraction, the method of constant temperature and constant water addition is adopted, and the whole process is relatively lengthy and energy consumption is high;

[0008] 3. The final product has poor color and taste, and cannot fully utilize the nutrients and functional ingredients of the whole Panax notoginseng plant and completely retain the unique original flavor of Panax notoginseng. Summary of the invention

[0009] The purpose of the present invention is to propose a Panax notoginseng puree and a preparation method and application thereof, which can fully utilize the whole plant resources of Panax notoginseng, improve the extraction rate of effective ingredients, and obtain high-quality Panax notoginseng puree, which is rich in active substances, such as organic selenium, organic zinc, Ganoderma lucidum protein, Ganoderma lucidum polysaccharide, Panax notoginseng polysaccharide, Panax notoginseng saponin, flavonoids, triterpenes, saponin, polyphenols and other active substances, has good antioxidant, anti-inflammatory, anti-aging, immunity-enhancing, metabolism-promoting and other activities, greatly improves the physiological activity of the puree, has stable quality, and is easy to realize large-scale industrial production.

[0010] The technical solution of the present invention is achieved as follows: A method for preparing Panax notoginseng puree comprises the following steps:

[0011] S1. Select fresh and pure Panax notoginseng whole plant, including roots, stems, leaves and flowers, and wash to remove sand and impurities;

[0012] S2. The whole cleaned Panax notoginseng plant was treated with enzyme inactivation;

[0013] S3. The whole plant of Panax notoginseng was crushed and juiced after fixing, the juice was filtered, and the filtrate was concentrated to obtain a preliminary puree, and the solid residue was retained;

[0014] S4. Add the solid residue into water, add a composite enzyme for enzymolysis, then inoculate Ganoderma lucidum seed liquid, selenium-enriched yeast and zinc-enriched yeast seed liquid for fermentation and culture. After the fermentation is completed, filter the fermentation liquid to obtain a fermentation filtrate; add an immobilized ionic liquid / tea saponin composite system to the fermentation filtrate for extraction, and filter again to obtain a solid residue and an extract; add the solid residue to ethanol and heat it for desorption, and after filtering, respectively obtain a desorbed solid and a liquid, separate, wash and recycle the solid, and concentrate the liquid to obtain a secondary pulp; dialyze the extract, and concentrate the non-permeated liquid to obtain a tertiary pulp; the composite enzyme includes cellulase and neutral protease in a mass ratio of 10: (3-4);

[0015] The preparation method of the immobilized ionic liquid / tea saponin composite system is as follows:

[0016] T1. Soaking silica gel in acid solution, filtering, washing and drying to obtain activated silica gel;

[0017] T2. Adding activated silica gel to toluene, adding silane coupling agent KH560, silane coupling agent KH602 and pyridine under inert gas protection, heating and reflux stirring to react, filtering, washing and drying after the reaction to obtain modified silica gel;

[0018] T3. 1-methylimidazole was added to ethanol, concentrated sulfuric acid was added dropwise, modified silica gel was added, the reaction was heated and stirred, and after the reaction was completed, the imidazole-modified silica gel was obtained by filtering, washing and drying;

[0019] T4. The tea saponin was added to water, imidazole-modified silica gel was added, the reaction was heated and stirred, and after the reaction was completed, the immobilized ionic liquid / tea saponin composite system was obtained by filtering, washing and drying;

[0020] S5. The primary puree, the secondary puree and the tertiary puree are mixed evenly, the mixed solution is filtered and sterilized, and finally packaged to obtain Panax notoginseng puree.

[0021] As a further improvement of the present invention, the sterilization and enzyme inactivation treatment in step S2 is performed by using steam sterilization and enzyme inactivation at 95-100°C.

[0022] As a further improvement of the present invention, the density of the concentrated preliminary pulp relative to water in step S3 is 1.13-1.15.

[0023] As a further improvement of the present invention, the mass ratio of the solid residue and the composite enzyme in step S4 is 10:(1-2), the temperature of the enzymatic hydrolysis is 35-45°C, and the time is 1-3h; the bacterial content of the seed liquid is 108-109cfu / mL, the inoculation amounts of the Ganoderma lucidum seed liquid, selenium-enriched yeast and zinc-enriched yeast seed liquid are 2-3v / v%, 1-2v / v%, and 1.5-2.5v / v%, respectively, the fermentation and culture conditions are temperature 25-30°C, rotation speed 100-200r / min, and fermentation and culture time 24-36h; the addition amount of the immobilized ionic liquid / tea saponin composite system is 3-5wt% of the total mass of the fermentation filtrate, the temperature of the heating desorption is 40-50°C, the pore size of the dialysis bag used for dialysis is 3-5kDa, and the time is 10-12h.

[0024] As a further improvement of the present invention, the acid solution in step T1 is 5-10wt% HCl or sulfuric acid solution, and the immersion time is 3-5h; the mass ratio of activated silica gel, silane coupling agent KH560, silane coupling agent KH602 and pyridine in step T2 is 10:(2-3):(2-4):(0.5-1), and the heating reflux stirring reaction time is 3-5h; the mass ratio of 1-methylimidazole, concentrated sulfuric acid and modified silica gel in step T3 is (2.5-3.5):(1-2):10, the heating stirring reaction temperature is 60-70°C, and the time is 5-7h; the mass ratio of tea saponin and imidazole-modified silica gel in step T4 is (2-3):10, the heating stirring reaction temperature is 40-50°C, and the time is 1-2h.

[0025] As a further improvement of the present invention, the mass ratio of the primary puree, the secondary puree and the tertiary puree is 10:(3-5):(4-6).

[0026] The present invention also provides Panax notoginseng puree prepared by any one of the preparation methods described above.

[0027] The present invention has the following beneficial effects:

[0028] The method for preparing whole-plant Panax notoginseng essence puree provided by the present invention can not only fully utilize the whole-plant resources of Panax notoginseng, but also improve the extraction rate of effective ingredients, and has a good market application prospect. The present invention can fully utilize the whole-plant resources of Panax notoginseng and improve the utilization rate of raw materials. The present invention improves the extraction rate of effective ingredients in Panax notoginseng and enhances the medicinal value of the product. The puree prepared by the present invention has stable quality and is easy to store and transport;

[0029] Traditional Chinese medicine residues contain a large amount of nutrients, bioactive substances and some unknown growth-promoting substances. In particular, the residues produced after extracting medicinal ingredients from medicinal plants with rhizomes as medicine have a high starch content and are suitable for use as fermentation substrates. By selecting suitable microorganisms to ferment them, they can be converted into high value-added products, thus turning waste into treasure. The present invention uses solid residues of Panax notoginseng as the culture medium raw material. On the one hand, it can promote the proliferation of microorganisms and medicinal fungi Ganoderma lucidum. On the other hand, the microorganisms themselves can also promote the breaking of plant cell walls, thereby promoting the dissolution of highly active substances, such as polysaccharides, amino acids, proteins, flavonoids, triterpenes, saponins and other components. A part of the active components are converted by microorganisms into active substances produced by microorganisms, such as short-chain fatty acids, antimicrobial peptides, etc., and the other part enters the fermentation liquid, so that the obtained fermentation liquid contains rich active substances. The present invention adopts the "bidirectional fermentation" technology of fermenting traditional Chinese medicine residues with medicinal fungi to improve the utilization value of traditional Chinese medicine residues, which has important theoretical and practical significance for the economic industrial chain of derived traditional Chinese medicine resources and the realization of sustainable development of my country's traditional Chinese medicine industry.

[0030] The invention uses ganoderma lucidum seed liquid, selenium-enriched yeast and zinc-enriched yeast seed liquid for mixed fermentation, and while obtaining a large amount of ganoderma lucidum polysaccharides and ganoderma lucidum proteins, it also enriches organic selenium and organic zinc, thereby preparing ganoderma lucidum selenium / zinc polysaccharides and ganoderma lucidum selenium / zinc proteins, which greatly improves their pharmacological activity. The ganoderma lucidum selenium / zinc polysaccharides and ganoderma lucidum selenium / zinc proteins are mainly distributed in the tertiary puree (small molecular impurities are removed by dialysis), and their antioxidant, anti-inflammatory, anti-aging, immunity-enhancing, metabolism-promoting activities are improved, and they have excellent physiological value;

[0031] The present invention prepares an immobilized ionic liquid / tea saponin composite system, using silica gel as a carrier. Silica gel, as a stable and low-cost inorganic porous material, has good performance. It is modified by a silane coupling agent KH560 with an epoxy group and a silane coupling agent KH602 with an amino group, and the epoxy group reacts with the imidazole group, so that the imidazole-based ionic liquid is fixed on the silica gel. The amino group can form a hydrogen bond with the hydroxyl group of the tea saponin, so that the tea saponin is fixed on the silica gel. The immobilized ionic liquid / tea saponin composite system has a low-toxic, biodegradable new functional ionic liquid. At the same time, it has a good separation and purification effect, and forms a composite system with tea saponin, which can better separate active substances by forming micelles, thereby better promoting the separation and purification of active substances such as flavonoids, triterpenes, saponins, polyphenols, etc.

[0032] The Panax notoginseng puree prepared by the invention contains abundant active substances, such as organic selenium, organic zinc, ganoderma lucidum protein, ganoderma lucidum polysaccharide, Panax notoginseng polysaccharide, Panax notoginseng saponin, flavonoids, triterpenes, saponin, polyphenols and the like, has good activities such as anti-oxidation, anti-inflammatory, anti-aging, immunity-enhancing, metabolism-promoting and the like, greatly improves the physiological activity of the puree, has stable quality, and is easy to realize large-scale industrial production. DETAILED DESCRIPTION

[0033] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0034] Cellulase, 10,000 U / g, was purchased from Guangdong Mingtong Biotechnology Co., Ltd.; neutral protease, 5000 U / g, was purchased from Jiangsu Caiwei Biotechnology Co., Ltd.

[0035] Silica gel, 100 mesh, was purchased from Shandong Tenghai Chemical Co., Ltd.

[0036] Selenium-enriched yeast, 20 billion cfu / g, was purchased from Angel Yeast Co., Ltd.; zinc-enriched yeast, 20 billion cfu / g, was purchased from Shaanxi Rankang Biotechnology Co., Ltd.

[0037] Preparation of Ganoderma lucidum seed liquid: Take Ganoderma lucidum mycelium (National Product Identification Bacteria 2007045), add it into culture medium (20g / L potato, 2.0g / L glucose, 0.3g / L yeast powder, 0.2g / L peptone, 0.1g / L KH2PO4, 0.06g / LMgSO4, sterilized at 121℃ for 30min), and culture at 180r / min, 28℃ for 7d to obtain Ganoderma lucidum seed liquid.

[0038] Preparation method of bacterial seed liquid: inoculate the bacterial strain into Gao's medium, activate and culture for 18-24h at 30℃ and 100r / min, and obtain a liquid containing 10 8 -10 9 cfu / mL of bacterial seed solution.

[0039] Preparation Example 1 Preparation of immobilized ionic liquid / tea saponin composite system, the specific method is as follows:

[0040] T1. 10 g of silica gel was added to 100 mL of 5 wt% HCl solution and soaked for 3 h, filtered, washed, and dried to obtain activated silica gel;

[0041] T2. Add 10 g of activated silica gel to 200 mL of toluene, add 2 g of silane coupling agent KH560, 2 g of silane coupling agent KH602 and 0.5 g of pyridine under nitrogen protection, heat and reflux with stirring to react for 3 h, filter, wash and dry to obtain modified silica gel;

[0042] T3. Add 2.5 g of 1-methylimidazole to 200 mL of ethanol, add 1 g of concentrated sulfuric acid dropwise, add 10 g of modified silica gel, heat to 60°C, stir and react for 5 h, filter, wash, and dry to obtain imidazole-modified silica gel;

[0043] T4. Add 2 g of tea saponin into 200 mL of water, add 10 g of imidazole-modified silica gel, heat to 40°C, stir to react for 1 h, filter, wash, and dry to obtain an immobilized ionic liquid / tea saponin composite system.

[0044] Preparation Example 2 Preparation of immobilized ionic liquid / tea saponin composite system, the specific method is as follows:

[0045] T1. 10 g of silica gel was added to 100 mL of 10 wt% sulfuric acid solution and soaked for 5 h, filtered, washed, and dried to obtain activated silica gel;

[0046] T2. Add 10 g of activated silica gel to 200 mL of toluene, add 3 g of silane coupling agent KH560, 4 g of silane coupling agent KH602 and 1 g of pyridine under nitrogen protection, heat and reflux with stirring to react for 5 h, filter, wash and dry to obtain modified silica gel;

[0047] T3. Add 3.5 g of 1-methylimidazole to 200 mL of ethanol, add 2 g of concentrated sulfuric acid dropwise, add 10 g of modified silica gel, heat to 70°C, stir and react for 7 hours, filter, wash, and dry to obtain imidazole-modified silica gel;

[0048] T4. Add 3 g of tea saponin into 200 mL of water, add 10 g of imidazole-modified silica gel, heat to 50°C, stir and react for 2 h, filter, wash and dry to obtain an immobilized ionic liquid / tea saponin composite system.

[0049] Preparation Example 3 Preparation of immobilized ionic liquid / tea saponin composite system, the specific method is as follows:

[0050] T1. 10 g of silica gel was added to 100 mL of 7 wt % HCl solution and soaked for 4 h, filtered, washed, and dried to obtain activated silica gel;

[0051] T2. Add 10 g of activated silica gel to 200 mL of toluene, add 2.5 g of silane coupling agent KH560, 3 g of silane coupling agent KH602 and 0.7 g of pyridine under nitrogen protection, heat and reflux with stirring to react for 4 h, filter, wash and dry to obtain modified silica gel;

[0052] T3. Add 3 g of 1-methylimidazole to 200 mL of ethanol, add 1.5 g of concentrated sulfuric acid dropwise, add 10 g of modified silica gel, heat to 65 ° C, stir and react for 6 hours, filter, wash, and dry to obtain imidazole-modified silica gel;

[0053] T4. Add 2.5 g tea saponin into 200 mL water, add 10 g imidazole-modified silica gel, heat to 45°C, stir to react for 1.5 h, filter, wash, and dry to obtain an immobilized ionic liquid / tea saponin composite system.

[0054] Comparative Preparation Example 1

[0055] Compared with Preparation Example 3, the difference is that the silane coupling agent KH560 is not added in step T2.

[0056] The details are as follows:

[0057] T1. 10 g of silica gel was added to 100 mL of 7 wt % HCl solution and soaked for 4 h, filtered, washed, and dried to obtain activated silica gel;

[0058] T2. 10 g of activated silica gel was added to 200 mL of toluene, and under nitrogen protection, 5.5 g of silane coupling agent KH602 and 0.7 g of pyridine were added, and the mixture was heated to reflux and stirred for 4 h, filtered, washed, and dried to obtain modified silica gel;

[0059] T3. Add 3 g of 1-methylimidazole to 200 mL of ethanol, add 1.5 g of concentrated sulfuric acid dropwise, add 10 g of modified silica gel, heat to 65 ° C, stir and react for 6 hours, filter, wash, and dry to obtain imidazole-modified silica gel;

[0060] T4. Add 2.5 g tea saponin into 200 mL water, add 10 g imidazole-modified silica gel, heat to 45°C, stir to react for 1.5 h, filter, wash, and dry to obtain an immobilized ionic liquid / tea saponin composite system.

[0061] Comparative Preparation Example 2

[0062] Compared with Preparation Example 3, the difference is that the silane coupling agent KH602 is not added in step T2.

[0063] The details are as follows:

[0064] T1. 10 g of silica gel was added to 100 mL of 7 wt % HCl solution and soaked for 4 h, filtered, washed, and dried to obtain activated silica gel;

[0065] T2. 10 g of activated silica gel was added to 200 mL of toluene, and under nitrogen protection, 5.5 g of silane coupling agent KH560 and 0.7 g of pyridine were added, and the mixture was heated to reflux and stirred for 4 h, filtered, washed, and dried to obtain modified silica gel;

[0066] T3. Add 3 g of 1-methylimidazole to 200 mL of ethanol, add 1.5 g of concentrated sulfuric acid dropwise, add 10 g of modified silica gel, heat to 65 ° C, stir and react for 6 hours, filter, wash, and dry to obtain imidazole-modified silica gel;

[0067] T4. Add 2.5 g tea saponin into 200 mL water, add 10 g imidazole-modified silica gel, heat to 45°C, stir to react for 1.5 h, filter, wash, and dry to obtain an immobilized ionic liquid / tea saponin composite system.

[0068] Comparative Preparation Example 3

[0069] Compared with Preparation Example 3, the difference is that step T2 is not performed.

[0070] The details are as follows:

[0071] T1. 10 g of silica gel was added to 100 mL of 7 wt % HCl solution and soaked for 4 h, filtered, washed, and dried to obtain activated silica gel;

[0072] T2. Add 3 g of 1-methylimidazole to 200 mL of ethanol, add 1.5 g of concentrated sulfuric acid dropwise, add 10 g of activated silica gel, heat to 65 ° C, stir and react for 6 hours, filter, wash, and dry to obtain imidazole-modified silica gel;

[0073] T3. Add 2.5 g tea saponin into 200 mL water, add 10 g imidazole-modified silica gel, heat to 45°C, stir to react for 1.5 h, filter, wash, and dry to obtain an immobilized ionic liquid / tea saponin composite system.

[0074] Comparative Preparation Example 4

[0075] Compared with Preparation Example 3, the difference is that step T4 is not performed.

[0076] The details are as follows:

[0077] T1. 10 g of silica gel was added to 100 mL of 7 wt % HCl solution and soaked for 4 h, filtered, washed, and dried to obtain activated silica gel;

[0078] T2. Add 10 g of activated silica gel to 200 mL of toluene, add 2.5 g of silane coupling agent KH560, 3 g of silane coupling agent KH602 and 0.7 g of pyridine under nitrogen protection, heat and reflux with stirring to react for 4 h, filter, wash and dry to obtain modified silica gel;

[0079] T3. Add 3 g of 1-methylimidazole to 200 mL of ethanol, add 1.5 g of concentrated sulfuric acid dropwise, add 10 g of modified silica gel, heat to 65°C, stir and react for 6 hours, filter, wash and dry to obtain imidazole-modified silica gel.

[0080] Test Example 1

[0081] The specific surface area and total pore volume of the products obtained in Examples 1-3 and Comparative Examples 1-7 were measured using a 3-FLEX 3500 multi-station high-throughput gas adsorption instrument. The results are shown in Table 1.

[0082] Table 1

[0083]

[0084] It can be seen from the above table that the products prepared in Preparation Examples 1-3 of the present invention have higher specific surface areas and total pore volumes.

[0085] Example 1

[0086] This embodiment provides a method for preparing Panax notoginseng puree, comprising the following steps:

[0087] S1. Select fresh and pure Panax notoginseng whole plant, including roots, stems, leaves and flowers, wash and remove sand and impurities;

[0088] S2. The whole cleaned Panax notoginseng plant was sterilized and enzymes were inactivated by steam at 95°C for 10 min;

[0089] S3. The whole plant of Panax notoginseng was crushed and juiced after fixing, the juice was filtered, and the filtrate was concentrated to obtain a preliminary puree, the initial puree relative to water density was 1.13, and the solid residue was retained;

[0090] S4. Add 10g of solid residue to 200mL of water, add 1g of complex enzyme, perform enzymolysis at 35℃ for 1h, inoculate Ganoderma lucidum seed liquid, selenium-enriched yeast and zinc-enriched yeast seed liquid, the inoculation amounts are 2v / v%, 1v / v%, 1.5v / v%, respectively, at 25℃, 100r / min, ferment and culture for 36h, filter to obtain fermentation filtrate, add the immobilized ionic liquid / tea saponin composite system prepared in Preparation Example 1 to the fermentation filtrate for extraction, the immobilized ionic liquid / tea saponin composite system added in an amount of 1.5v / v% to obtain the fermentation filtrate, and extract the fermentation filtrate. 3wt% of the total mass of the fermentation filtrate, after filtering again, a solid residue and an extract were obtained, the solid residue was added to ethanol and heated to 40°C, desorbed for 1h, filtered, and desorbed solid and liquid were obtained respectively, the solid was separated, washed and recycled, the liquid was concentrated to obtain a secondary stock, the density of the secondary stock relative to water was 1.14, the extract was dialyzed for 10h using a dialysis bag with a pore size of 3kDa, and the non-permeated liquid was concentrated to obtain a tertiary stock, the density of the tertiary stock relative to water was 1.12;

[0091] Wherein, the complex enzyme includes cellulase and neutral protease in a mass ratio of 10:3;

[0092] S5. Stir and mix 10 g of primary puree, 3 g of secondary puree, and 4 g of tertiary puree for 10 min, filter, sterilize, and package to obtain Panax notoginseng puree.

[0093] Example 2

[0094] This embodiment provides a method for preparing Panax notoginseng puree, comprising the following steps:

[0095] S1. Select fresh and pure Panax notoginseng whole plant, including roots, stems, leaves and flowers, wash and remove sand and impurities;

[0096] S2. The whole cleaned Panax notoginseng plant was sterilized and enzymes were inactivated by steam at 100°C for 10 minutes;

[0097] S3. The whole plant of Panax notoginseng was crushed and juiced after fixing, the juice was filtered, and the filtrate was concentrated to obtain a preliminary puree, the initial puree relative to water density was 1.15, and the solid residue was retained;

[0098] S4. Add 10g of solid residue to 200mL of water, add 2g of complex enzyme, enzymolysis at 45℃ for 3h, inoculate Ganoderma lucidum seed liquid, selenium-enriched yeast and zinc-enriched yeast seed liquid, the inoculation amount is 3v / v%, 2v / v%, 2.5v / v%, respectively, at 30℃, 200r / min, ferment and culture for 24h, filter to obtain fermentation filtrate, add the immobilized ionic liquid / tea saponin composite system prepared in Preparation Example 2 to the fermentation filtrate for extraction, the addition amount of the immobilized ionic liquid / tea saponin composite system is The solid residue was 5wt% of the total mass of the fermentation filtrate. After filtering again, a solid residue and an extract were obtained. The solid residue was added to ethanol and heated to 50°C for desorption for 1 hour. After filtering, the desorbed solid and liquid were obtained respectively. The solid was separated, washed and recycled. The liquid was concentrated to obtain a secondary stock. The density of the secondary stock relative to water was 1.14. The extract was dialyzed for 12 hours using a dialysis bag with a pore size of 5kDa. The non-permeated liquid was concentrated to obtain a tertiary stock. The density of the tertiary stock relative to water was 1.12.

[0099] Wherein, the composite enzyme comprises cellulase and neutral protease in a mass ratio of 10:4;

[0100] S5. Stir and mix 10 g of the primary puree, 5 g of the secondary puree, and 6 g of the tertiary puree for 10 min, filter, sterilize, and package to obtain Panax notoginseng puree.

[0101] Example 3

[0102] This embodiment provides a method for preparing Panax notoginseng puree, comprising the following steps:

[0103] S1. Select fresh and pure Panax notoginseng whole plant, including roots, stems, leaves and flowers, wash and remove sand and impurities;

[0104] S2. The whole cleaned Panax notoginseng plant was sterilized and enzymes were inactivated by steam at 100°C for 10 minutes;

[0105] S3. The whole plant of Panax notoginseng was crushed and juiced after fixing, the juice was filtered, and the filtrate was concentrated to obtain a preliminary puree, the initial puree relative to water density was 1.14, and the solid residue was retained;

[0106] S4. 10 g of the solid residue was added to 200 mL of water, 1.5 g of the complex enzyme was added, and the enzyme was hydrolyzed at 40 ° C for 2 h. The seed liquid of Ganoderma lucidum, selenium-enriched yeast and zinc-enriched yeast were inoculated at an inoculation rate of 2.5 v / v%, 1.5 v / v% and 2 v / v%, respectively. The fermentation was cultured at 27 ° C and 150 r / min for 30 h. After filtration, the fermentation filtrate was obtained. The immobilized ionic liquid / tea saponin composite system prepared in Preparation Example 3 was added to the fermentation filtrate for extraction. The immobilized ionic liquid / tea saponin composite system was added The amount added is 4wt% of the total mass of the fermentation filtrate. After filtering again, a solid residue and an extract are obtained. The solid residue is added to ethanol and heated to 45°C for desorption for 1 hour. After filtering, the desorbed solid and liquid are obtained respectively. The solid is separated, washed and recycled. The liquid is concentrated to obtain a secondary stock. The density of the secondary stock relative to water is 1.12. The extract is dialyzed for 11 hours using a dialysis bag with a pore size of 4kDa. The non-permeated liquid is concentrated to obtain a tertiary stock. The density of the tertiary stock relative to water is 1.15.

[0107] Wherein, the complex enzyme includes cellulase and neutral protease in a mass ratio of 10:3.5;

[0108] S5. Stir and mix 10 g of the primary puree, 4 g of the secondary puree, and 5 g of the tertiary puree for 10 min, filter, sterilize, and package to obtain Panax notoginseng puree.

[0109] Example 4

[0110] Compared with Example 3, the difference is that the complex enzyme is replaced by a single cellulase.

[0111] Example 5

[0112] Compared with Example 3, the difference is that the complex enzyme is replaced by a single neutral protease.

[0113] Comparative Example 1

[0114] Compared with Example 3, the difference is that no complex enzyme is added.

[0115] The details are as follows:

[0116] S4. 10 g of solid residue was added to 200 mL of water, and inoculated with Ganoderma lucidum seed liquid, selenium-enriched yeast and zinc-enriched yeast seed liquid, with inoculation amounts of 2.5 v / v%, 1.5 v / v% and 2 v / v%, respectively. The fermentation culture was carried out at 27°C and 150 r / min for 30 h. After filtration, a fermentation filtrate was obtained. The immobilized ionic liquid / tea saponin composite system prepared in Preparation Example 3 was added to the fermentation filtrate for extraction. The immobilized ionic liquid / tea saponin composite system was added in an amount of 4 wt% of the total mass of the fermentation filtrate. After filtration again, solid residue and extract were obtained. The solid residue was added to ethanol and heated to 45°C for desorption for 1 h. After filtration, the desorbed solid and liquid were obtained respectively. The solid was separated, washed and recycled. The liquid was concentrated to obtain a secondary stock, and the density of the secondary stock relative to water was 1.12. The extract was dialyzed for 11 h using a dialysis bag with a pore size of 4 kDa. The non-permeated liquid was concentrated to obtain a tertiary stock, and the density of the tertiary stock relative to water was 1.15.

[0117] Comparative Example 2

[0118] Compared with Example 3, the difference is that the Ganoderma lucidum seed liquid is not inoculated.

[0119] The details are as follows:

[0120] S4. 10 g of solid residue was added to 200 mL of water, 1.5 g of composite enzyme was added, and enzymolysis was carried out at 40 ° C for 2 h. The seed liquid of selenium-enriched yeast and zinc-enriched yeast was inoculated, and the inoculation amount was 1.5 v / v%, 2 v / v%, 27 ° C, 150 r / min, and the fermentation culture was carried out for 30 h. After filtering, a filtrate was obtained. The immobilized ionic liquid / tea saponin composite system prepared in Preparation Example 3 was added to the filtrate for extraction. The addition amount of the immobilized ionic liquid / tea saponin composite system was 4 wt% of the total mass of the filtrate. After filtering again, a solid residue and an extract were obtained. The solid residue was added to ethanol and heated to 45 ° C. Desorption was carried out for 1 h. After filtering, the desorbed solid and liquid were obtained respectively. The solid was separated, washed and recycled. The liquid was concentrated to obtain a secondary stock. The density of the secondary stock relative to water was 1.12. The extract was dialyzed for 11 h using a dialysis bag with a pore size of 4 kDa. The non-permeated liquid was concentrated to obtain a tertiary stock. The density of the tertiary stock relative to water was 1.15.

[0121] Wherein, the complex enzyme comprises cellulase and neutral protease, and the mass ratio is 10:3.5.

[0122] Comparative Example 3

[0123] Compared with Example 3, the difference is that the selenium-enriched yeast seed liquid is not inoculated.

[0124] The details are as follows:

[0125] S4. 10 g of solid residue was added to 200 mL of water, 1.5 g of complex enzyme was added, and enzymolysis was carried out at 40 ° C for 2 h. Ganoderma lucidum seed liquid and zinc-rich yeast seed liquid were inoculated at an inoculation rate of 2.5 v / v% and 3.5 v / v%, respectively. The fermentation was carried out at 27 ° C and 150 r / min for 30 h. After filtering, a filtrate was obtained. The immobilized ionic liquid / tea saponin composite system prepared in Preparation Example 3 was added to the filtrate for extraction. The addition amount of the immobilized ionic liquid / tea saponin composite system was 4 wt% of the total mass of the filtrate. After filtering, solid residue and extract were obtained. The solid residue was added to ethanol and heated to 45 ° C. Desorption was carried out for 1 h. After filtering, the desorbed solid and liquid were obtained respectively. The solid was separated, washed and recycled. The liquid was concentrated to obtain a secondary stock. The density of the secondary stock relative to water was 1.12. The extract was dialyzed for 11 h using a dialysis bag with a pore size of 4 kDa. The non-permeated liquid was concentrated to obtain a tertiary stock. The density of the tertiary stock relative to water was 1.15.

[0126] Wherein, the complex enzyme comprises cellulase and neutral protease, and the mass ratio is 10:3.5.

[0127] Comparative Example 4

[0128] Compared with Example 3, the difference is that the zinc-enriched yeast seed liquid is not inoculated.

[0129] The details are as follows:

[0130] S4. 10 g of solid residue was added to 200 mL of water, 1.5 g of complex enzyme was added, and enzymolysis was carried out at 40 ° C for 2 h. Ganoderma lucidum seed liquid and selenium-enriched yeast seed liquid were inoculated at an inoculation rate of 2.5 v / v% and 3.5 v / v%, respectively. The fermentation was carried out at 27 ° C and 150 r / min for 30 h. After filtering, a filtrate was obtained. The immobilized ionic liquid / tea saponin composite system prepared in Preparation Example 3 was added to the filtrate for extraction. The amount of the immobilized ionic liquid / tea saponin composite system added was 4 wt% of the total mass of the filtrate. After filtering, solid residue and extract were obtained. The solid residue was added to ethanol and heated to 45 ° C for desorption for 1 h. After filtering, the desorbed solid and liquid were obtained respectively. The solid was separated, washed and recycled. The liquid was concentrated to obtain a secondary stock. The density of the secondary stock relative to water was 1.12. The extract was dialyzed for 11 h using a dialysis bag with a pore size of 4 kDa. The non-permeated liquid was concentrated to obtain a tertiary stock. The density of the tertiary stock relative to water was 1.15.

[0131] Wherein, the complex enzyme comprises cellulase and neutral protease, and the mass ratio is 10:3.5.

[0132] Comparative Example 5

[0133] Compared with Example 3, the difference is that the seed liquid of selenium-enriched yeast and zinc-enriched yeast is not inoculated.

[0134] The details are as follows:

[0135] S4. 10 g of solid residue was added to 200 mL of water, 1.5 g of complex enzyme was added, and enzymolysis was carried out at 40 ° C for 2 h. Ganoderma lucidum seed liquid was inoculated with an inoculation amount of 2.5 v / v%, 27 ° C, 150 r / min, and fermentation was carried out for 30 h. After filtering, a filtrate was obtained. The immobilized ionic liquid / tea saponin composite system prepared in Preparation Example 3 was added to the filtrate for extraction. The addition amount of the immobilized ionic liquid / tea saponin composite system was 4 wt% of the total mass of the filtrate. After filtering, solid residue and extract were obtained. The solid residue was added to ethanol and heated to 45 ° C. Desorption was carried out for 1 h. After filtering, the desorbed solid and liquid were obtained respectively. The solid was separated, washed and recycled. The liquid was concentrated to obtain a secondary pulp. The density of the secondary pulp relative to water was 1.12. The extract was dialyzed for 11 h using a dialysis bag with a pore size of 4 kDa. The non-permeated liquid was concentrated to obtain a tertiary pulp. The density of the tertiary pulp relative to water was 1.15.

[0136] Wherein, the complex enzyme comprises cellulase and neutral protease, and the mass ratio is 10:3.5.

[0137] Comparative Example 6

[0138] Compared with Example 3, the difference is that the immobilized ionic liquid / tea saponin composite system is prepared by Comparative Preparation Example 1.

[0139] Comparative Example 7

[0140] Compared with Example 3, the difference is that the immobilized ionic liquid / tea saponin composite system is prepared by Comparative Preparation Example 2.

[0141] Comparative Example 8

[0142] Compared with Example 3, the difference is that the immobilized ionic liquid / tea saponin composite system is prepared by Comparative Preparation Example 3.

[0143] Comparative Example 9

[0144] Compared with Example 3, the difference is that the immobilized ionic liquid / tea saponin composite system is prepared by Comparative Preparation Example 4.

[0145] Comparative Example 10

[0146] Compared with Example 3, the difference is that no immobilized ionic liquid / tea saponin composite system is added.

[0147] The details are as follows:

[0148] S4. Add 10g of solid residue to 200mL of water, add 1.5g of complex enzyme, enzymolysis at 40℃ for 2h, inoculate Ganoderma lucidum seed liquid, selenium-enriched yeast and zinc-enriched yeast seed liquid, the inoculation amount is 2.5v / v%, 1.5v / v%, 2v / v%, respectively, at 27℃, 150r / min, ferment and culture for 30h, filter to obtain filtrate, concentrate the filtrate to obtain secondary puree, the density of secondary puree relative to water is 1.12;

[0149] Wherein, the complex enzyme comprises cellulase and neutral protease, and the mass ratio is 10:3.5.

[0150] S5. Stir and mix 10 g of the primary puree and 9 g of the secondary puree for 10 min, filter, sterilize, and package to obtain Panax notoginseng puree.

[0151] Test Example 2

[0152] The Panax notoginseng puree prepared in Example 1-5 or Comparative Example 1-10 was diluted 100 times to prepare a sample solution.

[0153] Take 2 mL of sample solution or ascorbic acid solution (0.5 g / L), add 2 mL of DPPH· (0.2 mmol / L, prepared with anhydrous ethanol) solution in a test tube and mix well (absolute ethanol is used instead of DPPH· as sample blank), place it at room temperature and avoid light for 30 minutes, and then detect the absorbance value at a wavelength of 517 nm, and each concentration should be measured 3 times. The results are shown in Table 2.

[0154] Clearance rate (%) = [1-(A i -A j ) / A c ]×100%, where A i A is the absorbance of a mixture of 2 mL DPPH· solution and 2 mL of the sample solution to be determined; j A is the absorbance of a mixture of 2 mL of anhydrous ethanol and 2 mL of the sample solution to be measured; c It is the absorbance of a mixture of 2 mL of DPPH solution and 2 mL of anhydrous ethanol.

[0155] Table 2

[0156]

[0157] It can be seen from the above table that the Panax notoginseng puree prepared in Examples 1-3 of the present invention has good antioxidant properties.

[0158] Test Example 3

[0159] SPF female ICR mice with a body weight of 18-22g were selected and randomly divided into a negative control group, Example 1-5 groups, and Comparative Example 1-10 groups, with 10 mice in each group. The experimental group was given 0.5 mg / kg·d of the corresponding prepared Panax notoginseng puree by oral gavage, and the negative control group was given an equal amount of normal saline. After continuous oral gavage for 30 days, 1 mL of 20% sheep red blood cell solution was injected into the mouse peritoneal cavity on the 31st day, and the mice were killed 0.5 hours later and placed in a supine position. 2 mL of normal saline wash solution was injected into the mouse peritoneal cavity. After massaging the abdomen for 1 minute, 1 mL of normal saline wash solution was aspirated and evenly dropped on 2 slides, respectively, and moved to a 37°C incubator for incubation for 30 minutes. The slides were then rinsed with normal saline and dried. After being fixed in a methanol solution for 1 minute, they were stained with 4% Giemsa for 3 minutes, rinsed with distilled water, and dried. The morphology of macrophages was observed and counted under a 40× microscope, and the calculation was performed according to formula (1). The results are shown in Table 3.

[0160] Phagocytic rate (%) = number of macrophages phagocytosing sheep red blood cells / total number of macrophages × 100% (1).

[0161] Table 3

[0162]

[0163] Note: * Compared with the negative control group, P < 0.05.

[0164] It can be seen from the above table that the Panax notoginseng puree prepared in Examples 1-3 of the present invention enhances the phagocytic activity of mouse peritoneal macrophages and improves the function of mouse monocyte-macrophages.

[0165] The above contents are merely examples and explanations of the structure of the present invention. The technicians in this technical field may make various modifications or additions to the specific embodiments described or replace them in a similar manner. As long as they do not deviate from the structure of the present invention or exceed the scope defined by the claims, they should all fall within the protection scope of the present invention.

Claims

1. A method for preparing Panax notoginseng puree, characterized in that: The following steps are involved: S1. Select fresh and pure Panax notoginseng whole plant, including roots, stems, leaves and flowers, and wash to remove sand and impurities; S2. The whole cleaned Panax notoginseng plant was treated with enzyme inactivation; S3. The whole plant of Panax notoginseng was crushed and juiced after fixing, the juice was filtered, and the filtrate was concentrated to obtain a preliminary puree, and the solid residue was retained; S4. The solid residue is added to water, and a composite enzyme is added for enzymatic hydrolysis, followed by inoculation of Ganoderma lucidum seed solution, selenium-enriched yeast and zinc-enriched yeast seed solution for fermentation. After the fermentation is completed, the fermentation liquid is filtered to obtain a fermentation filtrate; the fermentation filtrate is extracted by adding an immobilized ionic liquid / tea saponin composite system, and filtered again to obtain a solid residue and an extract; The solid residue is added to ethanol and heated for desorption treatment. After filtering, the desorbed solid and liquid are obtained respectively. The solid is separated, washed and recycled, and the liquid is concentrated to obtain a secondary pulp; the extract is dialyzed, and the undialyzed liquid is concentrated to obtain a tertiary pulp; the composite enzyme includes cellulase and neutral protease, and the mass ratio is 10: (3-4); The preparation method of the immobilized ionic liquid / tea saponin composite system is as follows: T1. Soaking silica gel in acid solution, filtering, washing and drying to obtain activated silica gel; T2. Adding activated silica gel to toluene, adding silane coupling agent KH560, silane coupling agent KH602 and pyridine under inert gas protection, heating and reflux stirring to react, filtering, washing and drying after the reaction to obtain modified silica gel; T3. 1-methylimidazole was added to ethanol, concentrated sulfuric acid was added dropwise, modified silica gel was added, the reaction was heated and stirred, and after the reaction was completed, the imidazole-modified silica gel was obtained by filtering, washing and drying; T4. The tea saponin was added to water, imidazole-modified silica gel was added, the reaction was heated and stirred, and after the reaction was completed, the immobilized ionic liquid / tea saponin composite system was obtained by filtering, washing and drying; S5. The primary puree, the secondary puree and the tertiary puree are mixed evenly, the mixed solution is filtered and sterilized, and finally packaged to obtain Panax notoginseng puree.

2. The preparation method according to claim 1, characterized in that: The sterilization and enzyme inactivation treatment in step S2 is carried out by using steam at 95-100°C to sterilize the enzyme.

3. The preparation method according to claim 1, characterized in that: The density of the concentrated preliminary pulp relative to water in step S3 is 1.13-1.

15.

4. The preparation method according to claim 1, characterized in that: In step S4, the mass ratio of the solid residue to the composite enzyme is 10:(1-2), the temperature of the enzymolysis is 35-45°C, and the time is 1-3h; the bacterial content of the seed solution is 108-109cfu / mL, the inoculation amounts of the ganoderma lucidum seed solution, selenium-enriched yeast and zinc-enriched yeast seed solution are 2-3v / v%, 1-2v / v%, and 1.5-2.5v / v%, respectively; the fermentation culture conditions are a temperature of 25-30°C, a rotation speed of 100-200r / min, and a fermentation culture time of 24-36h; the addition amount of the immobilized ionic liquid / tea saponin composite system is 3-5wt% of the total mass of the fermentation filtrate, the temperature of the heating desorption is 40-50°C, the dialysis bag pore size used in the dialysis is 3-5kDa, and the time is 10-12h.

5. The preparation method according to claim 1, characterized in that: The acid solution in step T1 is 5-10wt% HCl or sulfuric acid solution, and the immersion time is 3-5h; the mass ratio of activated silica gel, silane coupling agent KH560, silane coupling agent KH602 and pyridine in step T2 is 10:(2-3):(2-4):(0.5-1), and the heating reflux stirring reaction time is 3-5h; the mass ratio of 1-methylimidazole, concentrated sulfuric acid and modified silica gel in step T3 is (2.5-3.5):(1-2):10, the heating stirring reaction temperature is 60-70°C, and the time is 5-7h; the mass ratio of tea saponin and imidazole-modified silica gel in step T4 is (2-3):10, the heating stirring reaction temperature is 40-50°C, and the time is 1-2h.

6. The preparation method according to claim 1, characterized in that: The mass ratio of the primary puree, the secondary puree and the tertiary puree is 10:(3-5):(4-6).

7. A Panax notoginseng puree obtained by the preparation method according to any one of claims 1 to 6.

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