A preparation method of total paeonol glycosides

Abstract

The present application relates to a kind of preparation methods of total paeonia lactiflora paeonia glycoside, the method includes the following steps: 1) extraction: take white peony root decoction piece, extract with ethanol, extract liquid filtration;2) concentration: extract liquid heating concentration, recover ethanol, obtain concentrated solution;3) extraction: a, alkalization: concentrated liquid is added lye, and pH is adjusted to 5~7;B, ethyl acetate extraction removes impurity: join ethyl acetate extraction, and retain mother liquor;C, n-butanol ethyl acetate mixed solution extraction: the above liquid is extracted with n-butanol ethyl acetate mixed solution, and n-butanol ethyl acetate extract is obtained;4) concentration: after n-butanol ethyl acetate extraction liquid is recovered, dissolve with water, heat concentration, and obtain concentrated solution;5) drying: the concentrated solution is carried out spray drying.

Description

TECHNICAL FIELD

[0001] The present application relates to an extraction method of traditional Chinese medicine, in particular to a method for extracting total glucosides of paeonia from paeonia lactiflora. BACKGROUND

[0002] Paeonia lactiflora Pall. is a kind of traditional Chinese medicine, which is the dried root of Paeonia lactiflora Pall. It is harvested in summer and autumn, washed, and then the head, tail and fine roots are removed. The root is boiled in water, and then the skin is removed or peeled. It is dried after being dried. It has the effects of nourishing blood, regulating menstruation, stopping sweating, soothing the liver and relieving pain, and suppressing liver yang. It is commonly used for blood deficiency, irregular menstruation, sweating, insomnia, dizziness, headache, and pain.

[0003] The mixture of paeoniflorin, albiflorin, benzoylpaeoniflorin and other physiological functional components obtained from the dried root of Paeonia lactiflora Pall. is called total glucosides of paeonia (TGP). According to the dried product, the content of paeoniflorin (C 23 H 28 O 11 ) should not be less than 40.0%, the content of albiflorin (C 23 H 28 O 11 ) should not be less than 10.0%, and the content of 1,2,3,4,6-O-penta-galloyl glucose (C 41 H 32 O 26 ) should not be less than 8.0%.

[0004] It is known that the preparation method of total glucosides of paeonia is as follows: taking Paeonia lactiflora Pall., extracting it with 75% ethanol solution for 3 times, 1.5 hours for the first and second times, and 1 hour for the third time, filtering, combining the filtrate, concentrating the filtrate to the relative density of 1.15-1.25 (50℃-65℃) of the extract, adding saturated sodium bicarbonate solution to adjust the pH value to 5.9-6.1, extracting with ethyl acetate twice, discarding the ethyl acetate layer, extracting the mother liquor with ethyl acetate-n-butanol mixed solvent for 3 times, combining the extract, concentrating the extract to the relative density of 1.13-1.20 (50℃-65℃), and then performing spray drying, crushing, sieving, and mixing.

[0005] The detection method of total glucosides of paeonia is as follows:

[0006] Test solution: about 50 mg of the product is accurately weighed, placed in a 100 ml volumetric flask, added with appropriate amount of methanol, ultrasonically dissolved, taken out, cooled, diluted with methanol to the mark, shaken, and centrifuged to obtain the supernatant.

[0007] Take paeonol reference substance, paeonolactone reference substance and 1,2,3,4,6-O-penta-O-galloyl-glucose reference substance, accurately weigh and determine, dissolve in methanol and dilute quantitatively to prepare a mixed solution containing about 0.2 mg of paeonol, 75 μg of paeonolactone and 30 μg of 1,2,3,4,6-O-penta-O-galloyl-glucose per 1 ml.

[0008] Chromatographic conditions: octadecylsilane-bonded silica gel as filler (4.6 mm x 250 mm, 5 μm or a performance equivalent chromatographic column), acetonitrile as mobile phase A, triethylamine phosphate solution [triethylamine-phosphoric acid-water (1:1:1000)] as mobile phase B, gradient elution according to the following table; flow rate is 0.8 ml per minute; detection wavelength is 230 nm; injection volume is 5 μl.

[0009]

[0010] System suitability requirements

[0011] The separation degree between the paeonolactone peak, the paeonol peak and the 1,2,3,4,6-O-penta-O-galloyl-glucose peak shall meet the requirements.

[0012] Determination method

[0013] Accurately take the test sample solution and the reference substance solution and inject into the liquid chromatograph, and record the chromatogram.

[0014] Limits

[0015] In the chromatogram of the test sample solution, in addition to the solvent peak, the paeonol peak, the paeonolactone peak and the 1,2,3,4,6-O-penta-O-galloyl-glucose peak, the peak area of a single maximum component peak calculated by the peak area normalization method shall not be greater than 6% of the total peak area, and the sum of the peak areas of the components shall not be greater than 26% of the total peak area.

[0016] Baishao Zonggeng Capsules have the effects of inhibiting autoimmunity, anti-inflammation and pain relief, are the only plant-derived medicine for treating rheumatoid arthritis in China, are the exclusive patent product of Lihua Pharmaceutical, are the basic medicine for rheumatological diseases, and belong to the national medical insurance product. According to the clinical application for many years, Baishao Zonggeng has good effects on juvenile idiopathic arthritis, systemic lupus erythematosus, lupus nephritis, Sjogren's syndrome, ankylosing spondylitis, oral lichen planus, recurrent aphthous ulcer, inflammatory bowel disease, psoriasis, alopecia areata, vitiligo, eczema and the like.

[0017] In order to ensure product quality, according to relevant regulations, the total ginseng raw material and capsule need to be detected by fingerprint. In the established fingerprint, 14 common peaks are included. However, in the production detection, the small peaks often appear high and low, and the quality is not stable. As we all know, if the internal quality of traditional Chinese medicine is unstable, the difference of effective components is large, and the clinical effect is unstable. Therefore, based on the prior art, the preparation method of the total ginseng is optimized, and it is found that the small common peak detection result can be kept stable by the optimized preparation method. SUMMARY

[0018] The application provides a new preparation method of total ginseng, which comprises the following steps:

[0019] 1) extraction: taking ginseng slices, extracting with ethanol, and filtering the extract;

[0020] 2) concentration: heating and concentrating the extract, recovering ethanol, and obtaining a concentrated solution;

[0021] 3) extraction:

[0022] a, alkalization: adding alkali to the concentrated solution, and adjusting the pH value to 5-7;

[0023] b, ethyl acetate extraction: adding ethyl acetate for extraction, and retaining the mother liquor;

[0024] c, n-butanol-ethyl acetate mixed solution extraction: the above liquor is extracted with an n-butanol-ethyl acetate mixed solution to obtain an n-butanol-ethyl acetate extract;

[0025] 4) concentration: after recovering the n-butanol-ethyl acetate from the extract, dissolving in water, and heating and concentrating, a concentrated solution is obtained;

[0026] 5) drying: spray drying the concentrated solution.

[0027] Preferably, the method comprises the following steps:

[0028] 1) extraction: taking ginseng slices, extracting with 70-80% ethanol for 2-4 times, the amount of 70-80% ethanol is 2-5 times the weight of the ginseng slices, the extraction time is 1-2 hours, and the extraction temperature is controlled at 70-90 DEG C;

[0029] 2) concentration: heating and concentrating the extract, the concentration temperature is controlled at 50-60 DEG C, the vacuum degree is controlled at -0.06 to -0.09 Mpa, and the relative density is 1.15-1.25 at 50-65 DEG C;

[0030] 3) extraction:

[0031] a, alkalization: pH value of the concentrated solution is adjusted to 5.5-6.5 by adding sodium hydroxide, sodium carbonate or sodium bicarbonate solution;

[0032] b, ethyl acetate impurity removal: ethyl acetate is added to the above solution for extraction, and the ethyl acetate layer is discarded, and the mother liquor is retained;

[0033] c, extraction: the above mother liquor is extracted 2-4 times with a mixture of n-butanol: ethyl acetate = 3:7 (V / V), and the n-butanol ethyl acetate extract is retained;

[0034] 4) Concentration: after recovering the n-butanol ethyl acetate from the extract, water is added for dissolution, and heating and concentration are performed to obtain a concentrated solution;

[0035] 5) Drying: the concentrated solution is subjected to spray drying.

[0036] More preferably, the method of the present application comprises the following steps:

[0037] 1) Extraction: Bai Shao decoction pieces are taken, and 75±1% ethanol is used for extraction for 3 times, with the amount of 75±1% ethanol being 4 times, 3 times and 3 times the weight of the decoction pieces, respectively, and the extraction time being 1.5 hours, 1.5 hours and 1 hour, respectively. The extraction temperature is controlled at 80±5°C for extraction. The three extraction solutions are filtered and subjected to concentration process;

[0038] 2) Concentration: the concentration temperature is controlled at 50-60°C, the vacuum degree is controlled at -0.06 to -0.09 Mpa, and the concentrated solution is concentrated to a relative density of 1.15-1.25 at 50-65°C, and the concentration of recovered ethanol is controlled to be less than 5%, and then the solution is subjected to extraction process;

[0039] 3) Extraction:

[0040] a, alkalization: saturated NaHC03 solution is added to the concentrated solution to adjust the pH value to 5.9-6.1;

[0041] b, ethyl acetate impurity removal: ethyl acetate is added to the above solution for extraction, and the amount of ethyl acetate is 0.6 times the volume of the solution, which is stirred uniformly, and then is allowed to stand until it is separated into layers. The lower mother liquor is extracted again in the same way, and the mother liquor is retained;

[0042] c, extraction: the above solution is extracted with a mixture of n-butanol: ethyl acetate = 3:7 (V / V). The amount of the first extract is 0.8-1.2 times the volume of the solution, which is stirred uniformly and then is allowed to stand until it is separated into layers. The amount of the second extract is 0.6-0.8 times the volume of the solution, which is stirred uniformly and then is allowed to stand until it is separated into layers. The amount of the third extract is 0.4-1.0 times the volume of the solution, which is stirred uniformly and then is allowed to stand until it is separated into layers. The obtained n-butanol ethyl acetate extract is combined;

[0043] 4) Concentration: After recovering n-butanol and ethyl acetate from the extract under reduced pressure, dissolve in water, then heat and concentrate to 50-65°C, relative density 1.13-1.20, concentration temperature controlled at 40-60°C, vacuum degree controlled at -0.05 to -0.09 Mpa;

[0044] 5) Drying: spray dry the concentrated extract.

[0045] Most preferably, the method of the present application comprises the following steps:

[0046] 1) Extraction: take Baishao decoction pieces, extract with 75±1% ethanol for 3 times, 75±1% ethanol added amount is 4 times, 3 times and 3 times of the weight of the decoction pieces respectively, time is 1.5 hours, 1.5 hours and 1 hour respectively, extract at 80±5°C, filter the three times of extract liquid, and enter the concentration process;

[0047] 2) Concentration: concentration temperature controlled at 50-60°C, vacuum degree controlled at -0.06 to -0.09 Mpa, concentrated to 50-65°C, relative density 1.15-1.25, and control the concentration of recovered ethanol to be less than 5%, deliver to the extraction process;

[0048] 3) Extraction:

[0049] a, alkalization: add saturated NaHC03 solution to the concentrated liquid to adjust pH to 5.9-6.1;

[0050] b, ethyl acetate impurity removal: add ethyl acetate to the above liquid for extraction, the amount of ethyl acetate is 0.6 times of the volume of the liquid, stir uniformly, stand until stratification, extract the lower mother liquor again according to the above method, and reserve the mother liquor;

[0051] c, extraction: extract the above liquid with n-butanol: ethyl acetate = 3:7 (V / V) mixed liquid, the first extraction amount is 0.9 times of the volume of the liquid, stir uniformly, stand until stratification; the second extraction amount is 0.6 times of the volume of the liquid, stir uniformly, stand until stratification; the third extraction amount is 0.6 times of the volume of the liquid, stir uniformly, stand until stratification; combine the obtained n-butanol and ethyl acetate extract;

[0052] 4) Concentration: After recovering n-butanol and ethyl acetate from the extract under reduced pressure, dissolve in water, then heat and concentrate to 50-65°C, relative density 1.13-1.20, concentration temperature controlled at 40-60°C, vacuum degree controlled at -0.05 to -0.09 Mpa;

[0053] 5) Drying: spray dry the concentrated extract.

[0054] The present application also provides a traditional Chinese medicine extract, which is total ginsenoside prepared according to any one of the above schemes.

[0055] This invention provides a traditional Chinese medicine preparation, which is prepared from the extract of the traditional Chinese medicine described in this invention, with or without pharmaceutically acceptable excipients. Preferably, the traditional Chinese medicine preparation can be a decoction, pill, tablet, capsule, or granule.

[0056] Finally, this invention also provides the application of the traditional Chinese medicine extract described herein in the preparation of drugs for treating immune diseases. The immune diseases may include rheumatoid arthritis, juvenile idiopathic arthritis, systemic lupus erythematosus, lupus nephritis, Sjögren's syndrome, ankylosing spondylitis, oral lichen planus, recurrent aphthous ulcers, inflammatory bowel disease, psoriasis, alopecia areata, vitiligo, and eczema. Preferably, the Sjögren's syndrome is primary Sjögren's syndrome; the inflammatory bowel disease is ulcerative colitis and Crohn's disease.

[0057] The applicant discovered during production that the existing preparation method for total paeoniflorin was not perfect. Therefore, the applicant conducted a detailed study of each key operational step in the existing technology and improved it. Ultimately, it was found that by strictly controlling the amount of the extraction solvent, a mixed solution of n-butanol and ethyl acetate, the content of not only the main components of total paeoniflorin remained stable, but other components present in smaller amounts also maintained stability to a certain extent, effectively improving the intrinsic quality stability of the product.

[0058] Screening experiment

[0059] The following experimental work is a process of improving the existing technology in the extraction process. The amount of mixed extraction solvents n-butanol and ethyl acetate was screened, and the technical solution of the present invention was obtained, which solves the problems in the existing technology.

[0060] This invention uses fingerprint patterns (see...) Figure 1 The stability of the peak areas of 1, 2, 3, 6, 7, 8, 10, 11, 12, 13, and 14 among the 14 common peaks was used as an evaluation index to screen the amount of n-butanol:ethyl acetate = 3:7 (V / V) mixture used in the extraction step of the preparation method.

[0061] (1) First screening experiment on extraction dosage

[0062] Preparation of the extraction solution: Mix n-butanol and ethyl acetate at a volume ratio of 3:7.

[0063] Extraction process: the first extraction, the prepared extraction solvent was added to the extracted material, stirred and mixed, stirred for 2 minutes, and separated for more than 1.5 hours, wherein the amount of extraction solvent was 1.4 times the volume of extraction solvent, i.e. 1.4:1 = extraction solvent: extracted material (V / V); and 1.2 times the volume of extraction solvent, i.e. 1.2:1 = extraction solvent: extracted material (V / V); and 1.0 times the volume of extraction solvent, i.e. 1.0:1 = extraction solvent: extracted material (V / V); and 0.8 times the volume of extraction solvent, i.e. 0.8:1 = extraction solvent: extracted material (V / V); and 0.6 times the volume of extraction solvent, i.e. 0.6:1 = extraction solvent: extracted material (V / V);

[0064] The second extraction amount was 0.8:1 (V / V), stirred for 2 minutes, and separated for more than 1.5 hours;

[0065] The third extraction amount was 0.8:1 (V / V), stirred for 2 minutes, and separated for more than 1.5 hours;

[0066] The combined n-butanol ethyl acetate extract was recovered, and then the subsequent steps of Example 1 were followed to obtain white peony root total glycoside powder.

[0067] Five batches of products were prepared by the above method, and the five batches of products were detected by the fingerprint detection method of Example 4, and the peak areas of 14 common peaks of the five batches of products were statistically analyzed, and the analysis results are shown in Table 1:

[0068] Table 1 Peak area statistics table of first extraction screening process

[0069]

[0070]

[0071]

[0072] As can be seen from Table 1, in the case of the volume ratio of extraction solvent to extracted material being 0.8-1.2:1 in the first extraction, the common peak area of the product, including small common peaks 1, 2, 3, 6, 7, 8, 10, 11, 12, 13 and 14, has a smaller fluctuation range.

[0073] (2) Second extraction amount screening experiment

[0074] Further, the amount of the extraction solvent in the second extraction process was parameter screened.

[0075] Preparation of extraction solution: n-butanol and ethyl acetate were mixed in a volume ratio of 3:7.

[0076] Extraction process:

[0077] The first extraction amount is fixed at 1:1 (V / V), stirring for 2 minutes, and standing for more than 1.5 hours to separate layers;

[0078] The third extraction amount is fixed at 0.8:1 (V / V), stirring for 2 minutes, and standing for more than 1.5 hours to separate layers;

[0079] The second extraction method is as follows:

[0080] The prepared extraction solvent is added to the extracted material, stirred and mixed, stirred for 2 minutes, and stood for more than 1.5 hours to separate layers, wherein the extraction solvent amount is 1.0 times the volume of the extraction solvent, i.e. 1.0:1.0 = extraction solvent:extracted material (V / V); and 0.8 times the volume of the extraction solvent, i.e. 0.8:1.0 = extraction solvent:extracted material (V / V); and 0.6 times the volume of the extraction solvent, i.e. 0.6:1.0 = extraction solvent:extracted material (V / V); and 0.4 times the volume of the extraction solvent, i.e. 0.4:1.0 = extraction solvent:extracted material (V / V), stirred for 2 minutes, and stood for more than 1.5 hours to separate layers;

[0081] The n-butanol and ethyl acetate extract is collected and recovered, and then the subsequent steps of Example 1 are followed to obtain the total paeonol glycoside powder.

[0082] Five batches of products are prepared by the above method, and the five batches of products are detected by the fingerprint detection method of Example 4, and statistical analysis is performed on the 14 common peaks of the five batches of products. The analysis results are shown in Table 2:

[0083] Table 2 Peak area statistics table of second extraction screening process

[0084]

[0085]

[0086]

[0087] As can be seen from Table 2, in the second extraction, when the volume ratio of extraction solvent to extracted material is 0.6-0.8:1, the common peaks of total paeonol glycoside, including small common peaks 1, 2, 3, 6, 7, 8, 10, 11, 12, 13 and 14, have a smaller fluctuation range of peak area.

[0088] (3) Third extraction amount screening experiment

[0089] The amount of the extraction solvent in the third extraction process is further screened.

[0090] The extraction solution is prepared by mixing n-butanol and ethyl acetate at a volume ratio of 3:7.

[0091] The first extraction amount is fixed at 1:1 (V / V), stirring for 2 minutes, and standing for more than 1.5 hours for layering;

[0092] The second extraction amount is fixed at 0.8:1 (V / V), stirring for 2 minutes, and standing for more than 1.5 hours for layering;

[0093] The third extraction method is as follows:

[0094] The prepared extraction solvent is added to the extracted material, stirred and mixed, stirred for 2 minutes, and stood for more than 1.5 hours for layering, wherein the extraction solvent amount is 1.0 times the volume of the extraction solvent, i.e. 1.0:1.0 = extraction solvent: extracted material (V / V); and 0.8 times the volume of the extraction solvent, i.e. 0.8:1 = extraction solvent: extracted material (V / V); and 0.6 times the volume of the extraction solvent, i.e. 0.6:1 = extraction solvent: extracted material (V / V); and 0.4 times the volume of the extraction solvent, i.e. 0.4:1 = extraction solvent: extracted material (V / V), stirred for 2 minutes, and stood for more than 1.5 hours for layering;

[0095] The n-butanol and ethyl acetate extraction solution is collected and recovered, and then the subsequent steps of Example 1 are followed to obtain the total ginsenoside powder.

[0096] Five batches of products are prepared by the above method, and the five batches of products are detected by the fingerprint detection method of Example 4, and statistical analysis is performed on the 14 common peaks of the five batches of products. The analysis results are shown in Table 3:

[0097] Table 3 Peak area statistics table of third extraction screening process

[0098]

[0099]

[0100]

[0101] From Table 3, it can be seen that under the condition of fixed first and second extraction amounts, the volume ratio of the third extraction extraction solvent to the extracted material is 0.4-1:1, and the peak area fluctuation range of the small common peaks 1, 2, 3, 6, 7, 8, 10, 11, 12, 13, and 14 is smaller.

[0102] Compared with the prior art, the present application has the following beneficial effects:

[0103] The total paeony glycoside fingerprint established by the inventor includes 14 common peaks, but when the existing product is detected by using the detection method, the small common peaks often appear high and low, and the product quality between batches is not very stable. The preparation process of the total paeony glycoside is refined, the content instability of the small common peak components is solved, and it is proved through multiple tests that the preparation method can ensure the batch stability of the product. BRIEF DESCRIPTION OF DRAWINGS

[0104] Figure 1 The total paeony glycoside standard fingerprint. DETAILED DESCRIPTION

[0105] The application is further illustrated by the following examples, but is not limited to the application.

[0106] Example 1

[0107] 1) Extraction: 1000g of paeony root slices is taken, and 75±1% ethanol is used for extraction for 3 times, the ethanol is added in a quantity of 4 times, 3 times and 3 times of the weight of the paeony root slices respectively, the time is 1.5 hours, 1.5 hours and 1 hour respectively, the extraction temperature is controlled at 80±5℃, the extraction is carried out for 3 times, and the filtrate is filtered into the concentration process, and the residue is dried;

[0108] 2) Concentration: the concentrated solution is concentrated under reduced pressure, the concentration temperature is controlled at 50-60℃, the vacuum degree is controlled at -0.06 to -0.09 Mpa, the concentrated solution is concentrated to the relative density of 1.15-1.25 at 50-65℃, and the concentration of the recovered ethanol is controlled to be less than 5%, and the extraction process is delivered;

[0109] 3) Extraction:

[0110] a, alkalization: saturated NaHC03 solution is added to the concentrated solution to adjust the pH value to 5.9-6.1;

[0111] b, ethyl acetate impurity removal: ethyl acetate is added to the above liquid for extraction, the extraction amount is 0.6:1 (V / V), the stirring time is about 2-3 minutes, and the liquid is left to separate into layers for a specific time according to the separation condition, the ethyl acetate layer is taken out and placed separately, the lower layer mother liquor is extracted again by the above method, the ethyl acetate impurity removal liquid is combined, the ethyl acetate is recovered, and the concentrated solution is discarded;

[0112] c, extraction: the above liquid is extracted with a mixture of n-butanol and ethyl acetate in a ratio of 3:7 (V / V) in a reaction tank, the first extraction amount is 1.2:1 (V / V), the stirring time is 2 minutes, the liquid is left to separate into layers for more than 1.5 hours, the second and third extraction amounts are 0.6:1 (V / V), the stirring time is 2 minutes, and the liquid is left to separate into layers for more than 1.5 hours, and the n-butanol and ethyl acetate extraction liquids separated are combined;

[0113] 4) Concentration: recovering the n-butanol ethyl acetate extract under reduced pressure, adding water and heating to concentrate to a relative density of 1.13-1.20 at 50-65°C, the concentration temperature being controlled at 40-60°C and the vacuum degree at -0.05 to -0.09 MPa, to obtain a concentrated solution in an amount of 15% (M / M) of the crude drug;

[0114] 5) Drying: spray-drying the extracted concentrated solution to obtain 576 g of powder.

[0115] Example 2

[0116] 1) Extraction: taking 1000 g of white peony root decoction pieces, extracting with 75±1% ethanol for 3 times, the ethanol being added in an amount of 4, 3 and 3 times of the weight of the white peony root decoction pieces respectively, the time being 1.5 hours, 1.5 hours and 1 hour respectively, the extraction temperature being controlled at 80±5°C, the three times of extraction being filtered into the concentration process, and the residue being dried;

[0117] 2) Concentration: concentrating under reduced pressure, the concentration temperature being controlled at 50-60°C, the vacuum degree at -0.06 to -0.09 MPa, the concentrated solution being concentrated to a relative density of 1.15-1.25 at 50-65°C, and the concentration of the recovered ethanol being controlled to be less than 5%, to deliver the extraction process;

[0118] 3) Extraction:

[0119] a, Alkalization: adding saturated NaHC03 solution to the concentrated solution to adjust the pH value to 5.9-6.1;

[0120] b, Ethyl acetate impurity removal: adding ethyl acetate to the above solution for extraction, the extraction amount being 0.6:1 (V / V), the stirring time being about 2-3 minutes, and the standing time being determined according to the separation condition, the ethyl acetate layer being taken out and placed separately, and the lower layer mother liquor being extracted again according to the above method, the ethyl acetate impurity removal solution being combined, the ethyl acetate being recovered, and the concentrated solution being discarded;

[0121] c, Extraction: the above solution being extracted with a mixture of n-butanol and ethyl acetate = 3:7 (V / V) in a reaction tank, the first extraction amount being 1.0:1 (V / V), the stirring time being 2 minutes, and the standing time being more than 1.5 hours, the second and third extraction amounts being 0.8:1 (V / V), the stirring time being 2 minutes, and the standing time being more than 1.5 hours, and the n-butanol ethyl acetate extract obtained being combined;

[0122] 4) Concentration: recovering the n-butanol ethyl acetate extract under reduced pressure, adding water and heating to concentrate to a relative density of 1.13-1.20 at 50-65°C, the concentration temperature being controlled at 40-60°C and the vacuum degree at -0.05 to -0.09 MPa, to obtain a concentrated solution in an amount of 15% (M / M) of the crude drug;

[0123] 5) Drying: The extraction concentrate was spray dried to obtain 552 g of powder.

[0124] Example 3

[0125] 1) Extraction: 1000 g of Bai-shao decoction pieces were extracted with 75±1% ethanol for 3 times, the ethanol was added in a quantity of 4 times, 3 times and 3 times of the weight of Bai-shao decoction pieces respectively, the extraction time was 1.5 hours, 1.5 hours and 1 hour respectively, the extraction temperature was controlled at 80±5°C, the extraction was performed for 3 times, and the filtrate was subjected to the concentration process, and the residue was dried;

[0126] 2) Concentration: The concentrated solution was concentrated under reduced pressure, the concentration temperature was controlled at 50-60°C, the vacuum degree was controlled at -0.06 to -0.09 Mpa, the concentrated solution was concentrated to a relative density of 1.15-1.25 at 50-65°C, and the concentration of recovered ethanol was controlled to be less than 5%, and the concentrated solution was delivered to the extraction process;

[0127] 3) Extraction:

[0128] a, Alkalization: The pH value of the concentrated solution was adjusted to 5.9-6.1 by adding saturated NaHC03 solution;

[0129] b, Ethyl acetate impurity removal: Ethyl acetate was added to the above solution for extraction, the extraction amount was 0.6:1 (V / V), the stirring time was about 2-3 minutes, and the solution was allowed to stand until it was separated into layers, the specific time was determined according to the separation condition, the ethyl acetate layer was taken out and placed separately, and the lower layer mother liquor was extracted again according to the above method, the ethyl acetate impurity removal liquid was combined, the ethyl acetate was recovered, and the concentrated solution was discarded;

[0130] c, Extraction: The above solution was extracted with a mixture of n-butanol and ethyl acetate (3:7, V / V) in a reaction tank, the first extraction amount was 0.9:1 (V / V), the stirring time was 2 minutes, and the standing time was more than 1.5 hours until the solution was separated into layers, the second and third extraction amounts were 0.6:1 (V / V), the stirring time was 2 minutes, and the standing time was more than 1.5 hours until the solution was separated into layers, and the n-butanol and ethyl acetate extraction liquids obtained were combined;

[0131] 4) Concentration: The n-butanol and ethyl acetate extraction liquid was recovered under reduced pressure, and after adding water, it was heated and concentrated to a relative density of 1.13-1.20 at 50-65°C, the concentration temperature was controlled at 40-60°C, the vacuum degree was controlled at -0.05 to -0.09 Mpa, and the amount of concentrated solution obtained was 12% (M / M) of the crude drug amount;

[0132] 5) Drying: The extraction concentrate was spray dried to obtain 541 g of powder.

[0133] Example 4

[0134] Establishment of Total Glucoside Standard Fingerprint of Bai-shao

[0135] Determination by high performance liquid chromatography (Chinese Pharmacopoeia 2015 edition General 0512).

[0136] Chromatographic conditions and system suitability test with octadecylsilane-bonded silica gel as the filler (column length 25 cm, inner diameter 4.6 mm, particle size 5 μm); 0.05% phosphoric acid aqueous solution as mobile phase A, acetonitrile as mobile phase B, gradient elution according to the provisions in the following table; flow rate 1.0 mL / min; detection wavelength 230 nm; column temperature 20°C. The theoretical plate number should not be less than 8000 calculated by the peak of paeoniflorin.

[0137]

[0138] Reference solution preparation: take paeoniflorin, paeoniflorin, gallic acid, catechin, 1,2,3,4,6-O-penta-galloyl glucose control product, accurately weigh, add methanol to make 1 mL containing paeoniflorin 200 μg, paeoniflorin 40 μg, gallic acid 8 μg, catechin 10 μg, 1,2,3,4,6-O-penta-galloyl glucose 20 μg of mixed solution, shake well, filter, take the filtrate, get.

[0139] Test solution preparation: take 10 batches of qualified paeoniflorin powder prepared by the method of the application, about 25 mg per batch, accurately weigh, put into a 25 mL volumetric flask, add methanol, ultrasonic treatment (300 W, 40 kHz) to dissolve, take out, cool, dilute to the mark with methanol, shake well, filter, take the filtrate, get the test solution.

[0140] Determination method: accurately take 5 μL of blank solution, reference solution and test solution respectively, inject into the liquid chromatograph, determine, and record the chromatogram respectively.

[0141] The chromatogram of each batch of paeoniflorin obtained by 10 batches or more is corrected by computer simulation, calculated and output to obtain the standard fingerprint spectrum of the application, which is used as a standard control fingerprint spectrum for comparison with the fingerprint spectrum of any batch of product obtained in the production process. Figure 1 The standard fingerprint spectrum of the application.

Claims

1. A preparation method of total paeonol glycosides, the method comprising the following steps: 1) extraction: taking white peony root decoction pieces, extracting with 75±1% ethanol for 3 times, the amount of 75±1% ethanol added is 4 times, 3 times and 3 times of the weight of the decoction pieces respectively, the time is 1.5 hours, 1.5 hours and 1 hour respectively, the extraction temperature is controlled at 80±5℃, the three extraction liquids are filtered and enter the concentration process; 2) concentration: the concentration temperature is controlled at 50-60℃, the vacuum degree is controlled at -0.06 to -0.09 Mpa, the relative density is 1.15-1.25 at 50-65℃ after concentration, and the concentration of recovered ethanol is controlled to be less than 5%, and then the extraction process is performed; 3) extraction: a, alkalization: saturated NaHC03 solution is added to the concentrated liquid to adjust the pH value to 5.9-6.1; b, ethyl acetate impurity removal: ethyl acetate is added to the above liquid for extraction, the amount of ethyl acetate is 0.6 times of the volume of the liquid, it is stirred uniformly, and it is left to separate into layers, the lower mother liquor is extracted again by the above method, and the mother liquor is reserved; c, extraction: the above liquid is extracted with a mixture of n-butanol and ethyl acetate in a ratio of 3:7 (V / V), the amount of the first extraction liquid is 0.8-1.2 times of the volume of the liquid, it is stirred uniformly, left to separate into layers; the amount of the second extraction liquid is 0.6-0.8 times of the volume of the liquid, it is stirred uniformly, left to separate into layers; the amount of the third extraction liquid is 0.4-1.0 times of the volume of the liquid, it is stirred uniformly, left to separate into layers; the n-butanol and ethyl acetate extraction liquids obtained are combined; 4) concentration: after the n-butanol and ethyl acetate extraction liquid is recovered under reduced pressure, it is dissolved with water, heated and concentrated to a relative density of 1.13-1.20 at 50-65℃, the concentration temperature is controlled at 40-60℃, and the vacuum degree is controlled at -0.05 to -0.09 Mpa; 5) drying: the extraction and concentrated liquid is spray dried.

2. The method according to claim 1, comprising the following steps: 1) extraction: taking white peony root decoction pieces, extracting with 75±1% ethanol for 3 times, the amount of 75±1% ethanol added is 4 times, 3 times and 3 times of the weight of the decoction pieces respectively, the time is 1.5 hours, 1.5 hours and 1 hour respectively, the extraction temperature is controlled at 80±5℃, the three extraction liquids are filtered and enter the concentration process; 2) concentration: the concentration temperature is controlled at 50-60℃, the vacuum degree is controlled at -0.06 to -0.09 Mpa, the relative density is 1.15-1.25 at 50-65℃ after concentration, and the concentration of recovered ethanol is controlled to be less than 5%, and the extraction process is performed; 3) extraction: a, alkalization: saturated NaHC03 solution is added to the concentrated liquid to adjust the pH value to 5.9-6.1; b, ethyl acetate impurity removal: ethyl acetate is added to the above liquid for extraction, the amount of ethyl acetate is 0.6 times of the volume of the liquid, it is stirred uniformly, and it is left to separate into layers, the lower mother liquor is extracted again by the above method, and the mother liquor is reserved; ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ c, extraction: the above-mentioned medicinal liquid is extracted with a mixture of n-butanol and ethyl acetate in a ratio of 3:7 (V / V), the first extraction amount is 0.9 times the volume of the medicinal liquid, stirring is uniform, and it is left to separate into layers; the second extraction amount is 0.6 times the volume of the medicinal liquid, stirring is uniform, and it is left to separate into layers; the third extraction amount is 0.6 times the volume of the medicinal liquid, stirring is uniform, and it is left to separate into layers; the obtained n-butanol and ethyl acetate extract is combined; 4) concentration: after recovering the n-butanol and ethyl acetate from the extract under reduced pressure, it is dissolved in water and then concentrated to a relative density of 1.13-1.20 at 50-65°C, the concentration temperature is controlled at 40-60°C, and the vacuum degree is controlled at -0.05 to -0.09 Mpa; 5) drying: the extract concentrate is subjected to spray drying.

3. A traditional Chinese medicine extract, characterized in that The traditional Chinese medicine extract is prepared by any one of the methods of claims 1-2.

4. A traditional Chinese medicine preparation, characterized in that The preparation is prepared by adding or not adding pharmaceutically acceptable adjuvants to the traditional Chinese medicine extract of claim 3.

5. The traditional Chinese medicine preparation according to claim 4, characterized in that The traditional Chinese medicine preparation is selected from the group consisting of decoction, pill, tablet, capsule and granule.

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