CN109975437A - The analysis method of sanguisorbin I in garden burnet or product containing garden burnet - Google Patents
The analysis method of sanguisorbin I in garden burnet or product containing garden burnet Download PDFInfo
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- CN109975437A CN109975437A CN201711449088.5A CN201711449088A CN109975437A CN 109975437 A CN109975437 A CN 109975437A CN 201711449088 A CN201711449088 A CN 201711449088A CN 109975437 A CN109975437 A CN 109975437A
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- sanguisorbin
- garden burnet
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- 244000173853 Sanguisorba officinalis Species 0.000 title claims abstract description 94
- 235000008291 Poterium sanguisorba Nutrition 0.000 title claims abstract description 92
- 235000008282 Sanguisorba officinalis Nutrition 0.000 title claims abstract description 92
- 229930190125 Sanguisorbin Natural products 0.000 title claims abstract description 91
- 238000004458 analytical method Methods 0.000 title abstract description 4
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims abstract description 204
- 238000000034 method Methods 0.000 claims abstract description 56
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims abstract description 54
- 238000001514 detection method Methods 0.000 claims abstract description 31
- 239000007864 aqueous solution Substances 0.000 claims abstract description 20
- 229910019142 PO4 Inorganic materials 0.000 claims abstract description 17
- 239000010452 phosphate Substances 0.000 claims abstract description 17
- 238000004128 high performance liquid chromatography Methods 0.000 claims abstract description 9
- 230000005526 G1 to G0 transition Effects 0.000 claims abstract description 6
- 125000002467 phosphate group Chemical group [H]OP(=O)(O[H])O[*] 0.000 claims abstract description 3
- 239000000243 solution Substances 0.000 claims description 57
- 239000013558 reference substance Substances 0.000 claims description 46
- 239000012085 test solution Substances 0.000 claims description 44
- 239000007788 liquid Substances 0.000 claims description 38
- 239000000047 product Substances 0.000 claims description 26
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims description 23
- 238000000926 separation method Methods 0.000 claims description 23
- 238000010812 external standard method Methods 0.000 claims description 15
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 claims description 14
- 239000012528 membrane Substances 0.000 claims description 13
- 239000006228 supernatant Substances 0.000 claims description 13
- 238000009210 therapy by ultrasound Methods 0.000 claims description 13
- 238000005259 measurement Methods 0.000 claims description 11
- 239000000377 silicon dioxide Substances 0.000 claims description 11
- 239000000706 filtrate Substances 0.000 claims description 6
- 238000003556 assay Methods 0.000 claims description 5
- 238000001914 filtration Methods 0.000 claims description 3
- -1 is shaken up Substances 0.000 claims description 3
- 125000004079 stearyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 3
- 239000007924 injection Substances 0.000 claims description 2
- 238000002347 injection Methods 0.000 claims description 2
- 238000010829 isocratic elution Methods 0.000 claims description 2
- 239000003292 glue Substances 0.000 claims 2
- 238000007689 inspection Methods 0.000 claims 1
- 239000003814 drug Substances 0.000 abstract description 28
- 229940079593 drug Drugs 0.000 abstract description 23
- YTJSFYQNRXLOIC-UHFFFAOYSA-N octadecylsilane Chemical compound CCCCCCCCCCCCCCCCCC[SiH3] YTJSFYQNRXLOIC-UHFFFAOYSA-N 0.000 abstract description 11
- 238000002360 preparation method Methods 0.000 description 26
- 239000012071 phase Substances 0.000 description 21
- 239000000843 powder Substances 0.000 description 20
- 238000004587 chromatography analysis Methods 0.000 description 14
- 230000000052 comparative effect Effects 0.000 description 13
- 238000005303 weighing Methods 0.000 description 11
- 230000014759 maintenance of location Effects 0.000 description 9
- 238000012545 processing Methods 0.000 description 9
- 239000003643 water by type Substances 0.000 description 9
- 229940126062 Compound A Drugs 0.000 description 8
- NLDMNSXOCDLTTB-UHFFFAOYSA-N Heterophylliin A Natural products O1C2COC(=O)C3=CC(O)=C(O)C(O)=C3C3=C(O)C(O)=C(O)C=C3C(=O)OC2C(OC(=O)C=2C=C(O)C(O)=C(O)C=2)C(O)C1OC(=O)C1=CC(O)=C(O)C(O)=C1 NLDMNSXOCDLTTB-UHFFFAOYSA-N 0.000 description 8
- 150000001875 compounds Chemical class 0.000 description 8
- LNTHITQWFMADLM-UHFFFAOYSA-N gallic acid Chemical compound OC(=O)C1=CC(O)=C(O)C(O)=C1 LNTHITQWFMADLM-UHFFFAOYSA-N 0.000 description 8
- 239000004615 ingredient Substances 0.000 description 8
- 238000012360 testing method Methods 0.000 description 8
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 6
- 201000002364 leukopenia Diseases 0.000 description 6
- 238000002512 chemotherapy Methods 0.000 description 5
- 231100001022 leukopenia Toxicity 0.000 description 5
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 4
- 238000010828 elution Methods 0.000 description 4
- 238000000105 evaporative light scattering detection Methods 0.000 description 4
- 235000004515 gallic acid Nutrition 0.000 description 4
- 229940074391 gallic acid Drugs 0.000 description 4
- 239000007791 liquid phase Substances 0.000 description 4
- IUCHKMAZAWJNBJ-RCYXVVTDSA-N oleanolic acid 3-O-beta-D-glucosiduronic acid Chemical compound O([C@H]1CC[C@]2(C)[C@H]3CC=C4[C@@]([C@@]3(CC[C@H]2C1(C)C)C)(C)CC[C@]1(CCC(C[C@H]14)(C)C)C(O)=O)[C@@H]1O[C@H](C(O)=O)[C@@H](O)[C@H](O)[C@H]1O IUCHKMAZAWJNBJ-RCYXVVTDSA-N 0.000 description 4
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 3
- GBMDVOWEEQVZKZ-UHFFFAOYSA-N methanol;hydrate Chemical compound O.OC GBMDVOWEEQVZKZ-UHFFFAOYSA-N 0.000 description 3
- 238000001959 radiotherapy Methods 0.000 description 3
- 229930182490 saponin Natural products 0.000 description 3
- 150000007949 saponins Chemical class 0.000 description 3
- 235000017709 saponins Nutrition 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- IAWPVNMEXASPFO-UHFFFAOYSA-N C(CCCCCCCCCCCCCCCCC)[SiH3].[C] Chemical compound C(CCCCCCCCCCCCCCCCC)[SiH3].[C] IAWPVNMEXASPFO-UHFFFAOYSA-N 0.000 description 2
- GZQIINDHMUJEAM-MUPFVPTMSA-N Sanguisorbin B Chemical compound O([C@H]1CC[C@]2(C)[C@H]3CC=C4[C@@]([C@@]3(CC[C@H]2C1(C)C)C)(C)CC[C@]1(CC[C@H]([C@@H]([C@H]14)C)C)C(O)=O)[C@H]1OC[C@H](O)[C@H](O)[C@H]1O GZQIINDHMUJEAM-MUPFVPTMSA-N 0.000 description 2
- MKEYDVIGPFSUDP-UHFFFAOYSA-N Sanguisorbin B Natural products CC1CCC(C2CCC3(C)C(=CCC4C5(C)CCC(OC6OCC(O)C(O)C6O)C(C)(C)C5CCC34C)C12)C(=O)O MKEYDVIGPFSUDP-UHFFFAOYSA-N 0.000 description 2
- 208000007502 anemia Diseases 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 239000000284 extract Substances 0.000 description 2
- 230000011132 hemopoiesis Effects 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 150000008442 polyphenolic compounds Chemical class 0.000 description 2
- 235000013824 polyphenols Nutrition 0.000 description 2
- 239000001397 quillaja saponaria molina bark Substances 0.000 description 2
- QEVHRUUCFGRFIF-MDEJGZGSSA-N reserpine Chemical compound O([C@H]1[C@@H]([C@H]([C@H]2C[C@@H]3C4=C(C5=CC=C(OC)C=C5N4)CCN3C[C@H]2C1)C(=O)OC)OC)C(=O)C1=CC(OC)=C(OC)C(OC)=C1 QEVHRUUCFGRFIF-MDEJGZGSSA-N 0.000 description 2
- 229910052594 sapphire Inorganic materials 0.000 description 2
- 239000010980 sapphire Substances 0.000 description 2
- PFTAWBLQPZVEMU-DZGCQCFKSA-N (+)-catechin Chemical compound C1([C@H]2OC3=CC(O)=CC(O)=C3C[C@@H]2O)=CC=C(O)C(O)=C1 PFTAWBLQPZVEMU-DZGCQCFKSA-N 0.000 description 1
- AFSDNFLWKVMVRB-UHFFFAOYSA-N Ellagic acid Chemical compound OC1=C(O)C(OC2=O)=C3C4=C2C=C(O)C(O)=C4OC(=O)C3=C1 AFSDNFLWKVMVRB-UHFFFAOYSA-N 0.000 description 1
- ATJXMQHAMYVHRX-CPCISQLKSA-N Ellagic acid Natural products OC1=C(O)[C@H]2OC(=O)c3cc(O)c(O)c4OC(=O)C(=C1)[C@H]2c34 ATJXMQHAMYVHRX-CPCISQLKSA-N 0.000 description 1
- 229920002079 Ellagic acid Polymers 0.000 description 1
- 102000001554 Hemoglobins Human genes 0.000 description 1
- 108010054147 Hemoglobins Proteins 0.000 description 1
- AFCARXCZXQIEQB-UHFFFAOYSA-N N-[3-oxo-3-(2,4,6,7-tetrahydrotriazolo[4,5-c]pyridin-5-yl)propyl]-2-[[3-(trifluoromethoxy)phenyl]methylamino]pyrimidine-5-carboxamide Chemical compound O=C(CCNC(=O)C=1C=NC(=NC=1)NCC1=CC(=CC=C1)OC(F)(F)F)N1CC2=C(CC1)NN=N2 AFCARXCZXQIEQB-UHFFFAOYSA-N 0.000 description 1
- 235000014220 Rhus chinensis Nutrition 0.000 description 1
- 240000003152 Rhus chinensis Species 0.000 description 1
- 241001593750 Turcica Species 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 239000002671 adjuvant Substances 0.000 description 1
- 201000011510 cancer Diseases 0.000 description 1
- 239000012159 carrier gas Substances 0.000 description 1
- ADRVNXBAWSRFAJ-UHFFFAOYSA-N catechin Natural products OC1Cc2cc(O)cc(O)c2OC1c3ccc(O)c(O)c3 ADRVNXBAWSRFAJ-UHFFFAOYSA-N 0.000 description 1
- 235000005487 catechin Nutrition 0.000 description 1
- ZPEIMTDSQAKGNT-UHFFFAOYSA-N chlorpromazine Chemical compound C1=C(Cl)C=C2N(CCCN(C)C)C3=CC=CC=C3SC2=C1 ZPEIMTDSQAKGNT-UHFFFAOYSA-N 0.000 description 1
- 229960001076 chlorpromazine Drugs 0.000 description 1
- 229950001002 cianidanol Drugs 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 235000004132 ellagic acid Nutrition 0.000 description 1
- 229960002852 ellagic acid Drugs 0.000 description 1
- 210000003743 erythrocyte Anatomy 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 239000012467 final product Substances 0.000 description 1
- 229930003944 flavone Natural products 0.000 description 1
- 150000002213 flavones Chemical class 0.000 description 1
- 235000011949 flavones Nutrition 0.000 description 1
- 239000007789 gas Substances 0.000 description 1
- 229930182470 glycoside Natural products 0.000 description 1
- 150000002338 glycosides Chemical class 0.000 description 1
- 238000000227 grinding Methods 0.000 description 1
- 208000002672 hepatitis B Diseases 0.000 description 1
- 210000000265 leukocyte Anatomy 0.000 description 1
- FAARLWTXUUQFSN-UHFFFAOYSA-N methylellagic acid Natural products O1C(=O)C2=CC(O)=C(O)C3=C2C2=C1C(OC)=C(O)C=C2C(=O)O3 FAARLWTXUUQFSN-UHFFFAOYSA-N 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 239000011148 porous material Substances 0.000 description 1
- 230000005855 radiation Effects 0.000 description 1
- 238000010992 reflux Methods 0.000 description 1
- 239000000741 silica gel Substances 0.000 description 1
- 229910002027 silica gel Inorganic materials 0.000 description 1
- 239000000344 soap Substances 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N2030/022—Column chromatography characterised by the kind of separation mechanism
- G01N2030/027—Liquid chromatography
Landscapes
- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The analysis method of sanguisorbin I in a kind of garden burnet or product containing garden burnet, the method includes the separating step of sanguisorbin I is carried out using high performance liquid chromatography, in the high performance liquid chromatography, using octadecyl silane as stationary phase, Detection wavelength is 203-216nm, and mobile phase is phosphate aqueous solution: acetonitrile: methanol=(28-48): (5-25): (37-57).The method of the present invention can efficiently separate, detect, quantitative determine garden burnet or the product containing garden burnet in sanguisorbin I guarantee the validity and safety of medication to preferably control the quality of garden burnet and the product containing garden burnet.
Description
Technical field
The analysis method of sanguisorbin I in a kind of product the present invention relates to garden burnet or containing garden burnet.
Background technique
Generation rate of malignant tumour rises year by year, at present clinically frequently with treatment means have operation, radiation and chemotherapy
These three modes, and occurring toxicity in radiotherapy, chemotherapy is leukopenia an important factor for influencing clinical efficacy
(leucopenia) it is then wherein main toxic side effect, existing oneself becomes the most common complication after radiotherapy, chemotherapy.
Garden burnet in garden burnet and product containing garden burnet is rosaceous plant garden burnet Sanguisorba officinalis
L. or the dry root for garden burnet Sanguisorba officinalis L.Var.longifolia (Bert.) the Y ü et Li that comes into leaves, ground
Yuzhong mainly contains the chemical components such as Polyphenols, saponins and flavones, and polyphenol components mainly have gallic acid, (+)-catechu
Element, ellagic acid, nutgall catechin etc..Garden burnet has the function of increasing leukocyte quantity, clinically frequently as during chemotherapy
Adjuvant drug treats leukopenia caused by chemotherapy, it can also be used to treat leukopenia caused by chlorpromazine, do
Disturb leukopenia caused by extract for treating hepatitis B.
Sanguisorbin I is a main chemical compositions in garden burnet medicinal material, and available data shows that sanguisorbin I has one
Fixed hematopoiesis facilitation, can significantly raised red blood cell and hemoglobin, treat or prevent various anaemias, especially marrow hemopoiesis
Anaemia caused by hypofunction or failure has certain auxiliaring effect to treatment leukopenia.Therefore in detection garden burnet
Sanguisorbin I content, to control the product containing garden burnet quality play a significant role.
From the point of view of presently disclosed document, there are many analyzing detecting method report in relation to sanguisorbin.
Existing method 1: document " HPLC-ELSD method measurement Diyushengbai Tablet in sanguisorbin I content " (Chinese medicine with face
Bed, 2013,4 (1), 21-23 pages), using Garden Burnet by HPLC saponin I, with Sapphire C18 column (5 μm,
150 × 4.6mm) chromatography post separation, with methanol-water (64:36) for mobile phase, flow velocity 1.0ml/min, column temperature: room temperature；Drift tube
Temperature: 85 DEG C；Carrier gas: nitrogen, 2.8L/min；Not shunt mode；Sample volume: 20 μ l.
Existing method 2: patent CN101810705B also discloses the content assaying method of sanguisorbin I, using efficient liquid
Phase chromatography measures the content of sanguisorbin I, is stationary phase (4.6 × 150mm, 4 μm of partial size) with carbon octadecyl silane,
Methanol+Water gradient elution, elution program are (aqueous solution of 60% → 64% methanol of 0-7min, 7-13min64%
The aqueous solution of methanol), flow velocity 0.8ml/min, 25 DEG C of column temperature, ELSD drift tube temperature is 80 DEG C, gas (air) flow velocity 2.5L/
Min, shunt mode, theoretical cam curve should not be not less than 4 × 10 by sanguisorbin I calculating3。
Existing method 3: patent CN107064322A disclose " while measure garden burnet or or garden burnet class preparation in gallic acid
Switch method with the HPLC wavelength of sanguisorbin I content ", using Garden Burnet by HPLC saponin I, using C18 chromatography
Column, gradient elution, mobile phase A are acetonitrile, and Mobile phase B is 0.05% phosphate aqueous solution, detect galla turcica under the conditions of different wave length
The content of acid and sanguisorbin I.
Existing method 4: document " assay of sanguisorbin B and sanguisorbin-I in Chinese medicine the radix sanguisorbae total saponin " (north
Pharmacy the 1st phase of volume 14 in 2017, page 4), chromatographic column with Hypersil ODS chromatographic column (4.6mm × 250mm, 5.0 μm) into
Row experiment, mobile phase is acetonitrile: 0.02% phosphoric acid solution (82: 18), Detection wavelength 544nm, and the column temperature of chromatographic column is 25 DEG C, into
Sample amount is 15 μ L, flow velocity 0.8mL/min.
Present inventor when repeating above four kinds of methods for detecting the sanguisorbin I in garden burnet crude drug powder,
Have been surprisingly found that existing method 3 and existing method 4 relative to the testing result of existing method 1 and existing method 2 always higher percentage
20 or so.Existing method 1 and existing method 2 from the difference of existing method 3 and 4 other than chromatographic condition is different, detector
Also different.Existing method 1 and existing method 2 use ELSD detector, and existing method 3 and 4 uses UV detector, in general
The substance that different detectors can detect also has a difference, therefore present inventor speculates existing method 3 and existing method 4
Chromatographic condition can not separate other compounds in product of the sanguisorbin I with garden burnet or containing garden burnet.But it is existing
Can the chromatographic condition of method 1 and existing method 2 separate the sanguisorbin I in garden burnet or product containing garden burnet, and not
It obtains and knows.Present inventor is examined by using the chromatographic condition of existing method 1 and existing method 2 using UV detector
Sanguisorbin I in geodetic elm or product containing garden burnet, discovery testing result is also higher 20 percent.Therefore existing skill
The several method of art fails to for the sanguisorbin I in garden burnet or product containing garden burnet being kept completely separate with other compounds.Cause
This develops a kind of new chromatographic condition, so as to be very necessary by other compounds separate in sanguisorbin I and garden burnet.
Classes of compounds is more in garden burnet or product containing garden burnet, and up to tens kinds, by its in sanguisorbin I and garden burnet
He is kept completely separate compound, is very difficult.The present invention to solve the above-mentioned problems, develops a kind of new chromatographic condition,
Sanguisorbin I and other compounds in garden burnet can be separated, can from garden burnet the high sanguisorbin I of separation purity.And
Using the content of sanguisorbin I in this chromatographic condition quantitative detection garden burnet, sanguisorbin I can be fast and accurately detected.
Summary of the invention
In order to solve the above problems existing in the present technology, the present invention provides the separation method of new sanguisorbin I a kind of.
The method of the present invention can efficiently separate and detect the sanguisorbin I in garden burnet and product containing garden burnet.
The present invention provides the separation method of sanguisorbin I a kind of, which is characterized in that the method includes using efficient liquid phase
Chromatography carries out the separating step of sanguisorbin I, is to fix with octadecyl silane in the high performance liquid chromatography
Phase, Detection wavelength 203-216nm, mobile phase is phosphate aqueous solution: acetonitrile: methanol=(28-48): (5-25): (37-57).
Optionally, the phosphate aqueous solution concentration is 0.01%-0.1%.
Optionally, the flow velocity of the mobile phase is 0.5-2.0mL/min, is chosen as 0.5-1.5mL/min, or be chosen as
0.5-1.0mL/min；Column temperature is 20 DEG C -50 DEG C, is chosen as 20 DEG C -40 DEG C；Sample volume is 1-100 μ L, is chosen as 10-20 μ L.
The present invention also provides the purposes of above-mentioned separation method sanguisorbin I in separation garden burnet or product containing garden burnet.
The detection method of sanguisorbin I in a kind of product the present invention also provides garden burnet or containing garden burnet, which is characterized in that
The detection method includes using in the progress garden burnet of separation method described in any one of claim 1-3 or the product containing garden burnet
The step of detection of sanguisorbin I；
It is optionally, described that detection method includes the following steps:
(1) sanguisorbin I reference substance is taken, methanol is added to be configured to reference substance solution；
(2) sample to be tested is taken, methanol is added to extract, solution filters to obtain test solution；
(3) test solution and control solution are drawn respectively, high performance liquid chromatograph are injected, using claim 1-3
Any one of described in separation method be measured；
It is optionally, described that detection method includes the following steps:
(1) sanguisorbin I reference substance is taken, methanol is added to be configured to the solution that every 1mL contains 0.05mg-0.2mg；
(2) sample to be tested is taken, is pulverized, methanol is added, is ultrasonically treated, filtration takes filtrate；
(3) test solution and control solution are drawn respectively, high performance liquid chromatograph are injected, using claim 1-3
Any one of described in separation method be measured.
Optionally, the separation method includes:
(1) sanguisorbin I reference substance is taken, 50% methanol is added to be configured to the solution that every 1mL contains 0.08mg；
(2) sample to be tested is taken, is pulverized, is taken in right amount, 50% methanol is added and shakes up, weighed weight, ultrasonic treatment is let cool
To room temperature, then weighed weight, the weight of less loss is supplied with 50% methanol, is shaken up, supernatant is taken to be filtered with filter membrane, takes subsequent filtrate,
To obtain the final product.
(3) test solution and control solution, injection high performance liquid chromatograph measurement, with octadecyl key are drawn respectively
Conjunction silica gel is stationary phase, and Detection wavelength 203-216nm, mobile phase is 0.01%-0.1% phosphate aqueous solution: acetonitrile: methanol=
(28-48): (5-25): (37-57), isocratic elution, 20 DEG C -40 DEG C of column temperature, flow velocity 0.5-1.0mL/min；Sample volume is 10-
20μL。
The present invention also provides the use of above-mentioned detection method sanguisorbin I in detection garden burnet or product containing garden burnet
On the way.
The content assaying method of sanguisorbin I, feature in a kind of product the present invention also provides garden burnet or containing garden burnet
It is, the content assaying method includes the step that the assay of sanguisorbin I is carried out using detection method described in claim 5
Suddenly；
Optionally, the content assaying method is calculated using external standard method.
The separation, detection of sanguisorbin I and content assaying method tool in garden burnet of the invention or the product containing garden burnet
Other compounds in product of the sanguisorbin I with garden burnet or containing garden burnet can be efficiently separated by having the advantage that, while over the ground
The content of elm saponin I carries out Accurate Determining.To which the quality of the product to garden burnet or containing garden burnet controls, guarantee medication peace
It is complete and effective.
Detailed description of the invention
Fig. 1-7 is the high-efficient liquid phase chromatogram of test solution in embodiment 1-7；
Fig. 8-9 is respectively the high-efficient liquid phase chromatogram of test solution in comparative example 1-2.
Wherein, ordinate is response, and unit AU, abscissa is retention time, unit min.
Specific embodiment
Detailed description of the preferred embodiments below.It should be understood that described herein specific
Embodiment is merely to illustrate and explain the present invention, and is not intended to restrict the invention.
Garden burnet crude drug powder used in following embodiment and comparative example is the garden burnet medicinal material of same batch purchase, grinding
Cheng Fen crosses No. four sieves (65 mesh, sieve pore internal diameter: 250 μm ± 9.9 μm) and obtains.
Embodiment 1
Major experimental instrument and chromatographic condition:
Using Waters e2695 type high performance liquid chromatograph, it is equipped with waters2695 separative unit, 2489 UV, visible lights
Photodetector, chromatography processing system use Empower chromatographic work station.
Chromatographic condition:
Chromatographic column: octadecylsilane chemically bonded silica column
Mobile phase: 0.1% phosphate aqueous solution: acetonitrile: methanol=28:25:47
Wavelength: 210nm
Flow velocity: 0.6mL/min
Column temperature: 40 DEG C
Sample volume: 20 μ L
The preparation of reference substance solution: it is appropriate that precision weighs sanguisorbin I reference substance, adds 50% methanol that every 1ml is made containing 80 μ
G solution to get.
The preparation of test solution: weighing 100mg garden burnet crude drug powder, accurately weighed, sets in 100ml stuffed conical flask, essence
50% methanol 25ml of close addition shakes up, weighed weight, and ultrasonic treatment (power 300W, frequency 40KHz) 10 minutes is let cool to room
Temperature, then weighed weight, the weight of less loss is supplied with 50% methanol, is shaken up, and is taken supernatant to be filtered with 0.22 μm of filter membrane, is taken continuous filter
Liquid to get.
Measuring method: it is accurate respectively to measure above-mentioned reference substance solution and each 20 μ l of test solution, inject liquid chromatograph, note
Record chromatogram.It is calculated according to external standard method.
Test solution measurement result:
| Ingredient | Retention time | Content |
| Sanguisorbin I | 22.394 | 2.15% |
| Unknown compound A | 21.156 | 0.46% |
Embodiment 2
Major experimental instrument and chromatographic condition:
Using Waters e2695 type high performance liquid chromatograph, it is equipped with waters2695 separative unit, 2489 UV, visible lights
Photodetector, chromatography processing system use Empower chromatographic work station.
Chromatographic condition:
Chromatographic column: octadecylsilane chemically bonded silica column
Mobile phase: 0.07% phosphate aqueous solution: acetonitrile: methanol=35:15:50
Wavelength: 216nm
Flow velocity: 0.8mL/min
Column temperature: 20 DEG C
Sample volume: 20 μ L
The preparation of reference substance solution: it is appropriate that precision weighs sanguisorbin I reference substance, adds 50% methanol that every 1ml is made containing 80 μ
G solution to get.
The preparation of test solution: weighing 100mg garden burnet crude drug powder, accurately weighed, sets in 100ml stuffed conical flask, essence
50% methanol 25ml of close addition shakes up, weighed weight, and ultrasonic treatment (power 300W, frequency 40KHz) 10 minutes is let cool to room
Temperature, then weighed weight, the weight of less loss is supplied with 50% methanol, is shaken up, and is taken supernatant to be filtered with 0.22 μm of filter membrane, is taken continuous filter
Liquid to get.
Measuring method: it is accurate respectively to measure above-mentioned reference substance solution and each 20 μ l of test solution, inject liquid chromatograph, note
Record chromatogram.It is calculated according to external standard method.
Test solution measurement result:
| Ingredient | Retention time | Content |
| Sanguisorbin I | 29.299 | 2.23% |
| Unknown compound A | 27.633 | 0.45% |
Embodiment 3
Major experimental instrument and chromatographic condition:
Using Waters e2695 type high performance liquid chromatograph, it is equipped with waters2695 separative unit, 2489 UV, visible lights
Photodetector, chromatography processing system use Empower chromatographic work station.
Chromatographic condition:
Chromatographic column: octadecylsilane chemically bonded silica column
Mobile phase: 0.01% phosphate aqueous solution: acetonitrile: methanol=48:15:37
Wavelength: 203nm
Flow velocity: 0.7mL/min
Column temperature: 30 DEG C
Sample volume: 10 μ L
The preparation of reference substance solution: it is appropriate that precision weighs sanguisorbin I reference substance, adds 50% methanol that every 1ml is made containing 80 μ
G solution to get.
The preparation of test solution: weighing 100mg garden burnet crude drug powder, accurately weighed, sets in 100ml stuffed conical flask, essence
50% methanol 25ml of close addition shakes up, weighed weight, and ultrasonic treatment (power 300W, frequency 40KHz) 10 minutes is let cool to room
Temperature, then weighed weight, the weight of less loss is supplied with 50% methanol, is shaken up, and is taken supernatant to be filtered with 0.22 μm of filter membrane, is taken continuous filter
Liquid to get.
Measuring method: it is accurate respectively to measure above-mentioned reference substance solution and each 10 μ l of test solution, inject liquid chromatograph, note
Record chromatogram.It is calculated according to external standard method.
Test solution measurement result:
| Ingredient | Retention time | Content |
| Sanguisorbin I | 19.583 | 2.10% |
| Unknown compound A | 18.503 | 0.43% |
Embodiment 4
Major experimental instrument and chromatographic condition:
Using Waters e2695 type high performance liquid chromatograph, it is equipped with waters2695 separative unit, 2489 UV, visible lights
Photodetector, chromatography processing system use Empower chromatographic work station.
Chromatographic condition:
Chromatographic column: octadecylsilane chemically bonded silica column
Mobile phase: 0.05% phosphate aqueous solution: acetonitrile: methanol=38:5:57
Wavelength: 210nm
Flow velocity: 0.5mL/min
Column temperature: 40 DEG C
Sample volume: 10 μ L
The preparation of reference substance solution: it is appropriate that precision weighs sanguisorbin I reference substance, adds 50% methanol that every 1ml is made containing 80 μ
G solution to get.
The preparation of test solution: weighing 100mg garden burnet crude drug powder, accurately weighed, sets in 100ml stuffed conical flask, essence
50% methanol 25ml of close addition shakes up, weighed weight, and ultrasonic treatment (power 300W, frequency 40KHz) 10 minutes is let cool to room
Temperature, then weighed weight, the weight of less loss is supplied with 50% methanol, is shaken up, and is taken supernatant to be filtered with 0.22 μm of filter membrane, is taken continuous filter
Liquid to get.
Measuring method: it is accurate respectively to measure above-mentioned reference substance solution and each 10 μ l of test solution, inject liquid chromatograph, note
Record chromatogram.It is calculated according to external standard method.
Test solution measurement result:
| Ingredient | Retention time | Content |
| Sanguisorbin I | 27.736 | 2.27% |
| Unknown compound A | 26.196 | 0.52% |
Embodiment 5
Major experimental instrument and chromatographic condition:
Using Waters e2695 type high performance liquid chromatograph, it is equipped with waters2695 separative unit, 2489 UV, visible lights
Photodetector, chromatography processing system use Empower chromatographic work station.
Chromatographic condition:
Chromatographic column: octadecylsilane chemically bonded silica column
Mobile phase: 0.03% phosphate aqueous solution: acetonitrile: methanol=40:15:45
Wavelength: 216nm
Flow velocity: 1.0mL/min
Column temperature: 30 DEG C
Sample volume: 20 μ L
The preparation of reference substance solution: it is appropriate that precision weighs sanguisorbin I reference substance, adds 50% methanol that every 1ml is made containing 80 μ
G solution to get.
The preparation of test solution: weighing 100mg garden burnet crude drug powder, accurately weighed, sets in 100ml stuffed conical flask, essence
50% methanol 25ml of close addition shakes up, weighed weight, and ultrasonic treatment (power 300W, frequency 40KHz) 10 minutes is let cool to room
Temperature, then weighed weight, the weight of less loss is supplied with 50% methanol, is shaken up, and is taken supernatant to be filtered with 0.22 μm of filter membrane, is taken continuous filter
Liquid to get.
Measuring method: it is accurate respectively to measure above-mentioned reference substance solution and each 20 μ l of test solution, inject liquid chromatograph, note
Record chromatogram.It is calculated according to external standard method.
Test solution measurement result:
| Ingredient | Retention time | Content |
| Sanguisorbin I | 18.131 | 2.09% |
| Unknown compound A | 17.098 | 0.41% |
Embodiment 6
Major experimental instrument and chromatographic condition:
Using Waters e2695 type high performance liquid chromatograph, it is equipped with waters2695 separative unit, 2489 UV, visible lights
Photodetector, chromatography processing system use Empower chromatographic work station.
Chromatographic condition:
Chromatographic column: octadecylsilane chemically bonded silica column
Mobile phase: 0.05% phosphate aqueous solution: acetonitrile: methanol=36:14:50
Wavelength: 203nm
Flow velocity: 0.9mL/min
Column temperature: 20 DEG C
Sample volume: 10 μ L
The preparation of reference substance solution: it is appropriate that precision weighs sanguisorbin I reference substance, adds 50% methanol that every 1ml is made containing 80 μ
G solution to get.
The preparation of test solution: weighing 100mg garden burnet crude drug powder, accurately weighed, sets in 100ml stuffed conical flask, essence
50% methanol 25ml of close addition shakes up, weighed weight, and ultrasonic treatment (power 300W, frequency 40KHz) 10 minutes is let cool to room
Temperature, then weighed weight, the weight of less loss is supplied with 50% methanol, is shaken up, and is taken supernatant to be filtered with 0.22 μm of filter membrane, is taken continuous filter
Liquid to get.
Measuring method: it is accurate respectively to measure above-mentioned reference substance solution and each 10 μ l of test solution, inject liquid chromatograph, note
Record chromatogram.It is calculated according to external standard method.
Test solution measurement result:
| Ingredient | Retention time | Content |
| Sanguisorbin I | 12.298 | 2.27% |
| Unknown compound A | 11.621 | 0.54% |
Embodiment 7
Major experimental instrument and chromatographic condition:
Using Waters e2695 type high performance liquid chromatograph, it is equipped with waters2695 separative unit, 2489 UV, visible lights
Photodetector, chromatography processing system use Empower chromatographic work station.
Chromatographic condition:
Chromatographic column: octadecylsilane chemically bonded silica column
Mobile phase: 0.06% phosphate aqueous solution: acetonitrile: methanol=35:13:52
Wavelength: 216nm
Flow velocity: 0.7mL/min
Column temperature: 30 DEG C
Sample volume: 10 μ L
The preparation of reference substance solution: it is appropriate that precision weighs sanguisorbin I reference substance, adds 50% methanol that every 1ml is made containing 80 μ
G solution to get.
The preparation of test solution: weighing 100mg garden burnet crude drug powder, accurately weighed, sets in 100ml stuffed conical flask, essence
50% methanol 25ml of close addition shakes up, weighed weight, and ultrasonic treatment (power 300W, frequency 40KHz) 10 minutes is let cool to room
Temperature, then weighed weight, the weight of less loss is supplied with 50% methanol, is shaken up, and is taken supernatant to be filtered with 0.22 μm of filter membrane, is taken continuous filter
Liquid to get.
Measuring method: it is accurate respectively to measure above-mentioned reference substance solution and each 10 μ l of test solution, inject liquid chromatograph, note
Record chromatogram.It is calculated according to external standard method.
Test solution measurement result:
| Ingredient | Retention time | Content |
| Sanguisorbin I | 20.762 | 2.26% |
| Unknown compound A | 19.546 | 0.50% |
Comparative example 1
Major experimental instrument and chromatographic condition:
Using Waters e2695 type high performance liquid chromatograph, it is equipped with waters2695 separative unit, 2489 UV, visible lights
Photodetector, chromatography processing system use Empower chromatographic work station.
Chromatographic condition:
Chromatographic column: octadecylsilane chemically bonded silica column
Mobile phase: 0.01% phosphate aqueous solution: acetonitrile: methanol=46:27:27
Wavelength: 216nm
Flow velocity: 1.0mL/min
Column temperature: 30 DEG C
Sample volume: 20 μ L
The preparation of reference substance solution: it is appropriate that precision weighs sanguisorbin I reference substance, adds 50% methanol that every 1ml is made containing 80 μ
G solution to get.
The preparation of test solution: weighing 100mg garden burnet crude drug powder, accurately weighed, sets in 100ml stuffed conical flask, essence
50% methanol 25ml of close addition shakes up, weighed weight, and ultrasonic treatment (power 300W, frequency 40KHz) 10 minutes is let cool to room
Temperature, then weighed weight, the weight of less loss is supplied with 50% methanol, is shaken up, and is taken supernatant to be filtered with 0.22 μm of filter membrane, is taken continuous filter
Liquid to get.
Measuring method: it is accurate respectively to measure above-mentioned reference substance solution and each 20 μ l of test solution, inject liquid chromatograph, note
Record chromatogram.It is calculated according to external standard method.
Test solution chromatogram such as Fig. 8: knowing with this condition, and sanguisorbin I and unknown compound A cannot divide completely
From.
Comparative example 2
Major experimental instrument and chromatographic condition:
Using Waters e2695 type high performance liquid chromatograph, it is equipped with waters2695 separative unit, 2489 UV, visible lights
Photodetector, chromatography processing system use Empower chromatographic work station.
Chromatographic condition:
Chromatographic column: octadecylsilane chemically bonded silica column
Mobile phase: 0.05% phosphate aqueous solution: acetonitrile=70:30
Wavelength: 216nm
Flow velocity: 1.0mL/min
Column temperature: 30 DEG C
Sample volume: 20 μ L
The preparation of reference substance solution: it is appropriate that precision weighs sanguisorbin I reference substance, adds 50% methanol that every 1ml is made containing 80 μ
G solution to get.
The preparation of test solution: weighing 100mg garden burnet crude drug powder, accurately weighed, sets in 100ml stuffed conical flask, essence
50% methanol 25ml of close addition shakes up, weighed weight, and ultrasonic treatment (power 300W, frequency 40KHz) 10 minutes is let cool to room
Temperature, then weighed weight, the weight of less loss is supplied with 50% methanol, is shaken up, and is taken supernatant to be filtered with 0.22 μm of filter membrane, is taken continuous filter
Liquid to get.
Measuring method: it is accurate respectively to measure above-mentioned reference substance solution and each 20 μ l of test solution, inject liquid chromatograph, note
Record chromatogram.It is calculated according to external standard method.
Test solution measurement result:
| Ingredient | Retention time | Content |
| Sanguisorbin I | 30.083 | 2.76% |
Comparative example 3:
Using document " content that HPLC-ELSD method measures sanguisorbin I in Diyushengbai Tablet " (Chinese medicine and clinical, 2013,
4 (1), 21-23 pages) chromatographic condition, with UV detector, Detection wavelength 210nm detects the garden burnet soap in garden burnet crude drug powder
Glycosides I.
Chromatographic condition: with Sapphire C18 column (5 μm, 150 × 4.6mm) chromatography post separation, with methanol-water (64 ︰ 36)
For mobile phase, flow velocity 1.0ml/min, column temperature: room temperature；Sample volume: 20 μ l.
The preparation of reference substance solution: it is appropriate that precision weighs sanguisorbin I reference substance, adds 50% methanol that every 1ml is made containing 80 μ
G solution to get.
The preparation of test solution: weighing 100mg garden burnet crude drug powder, accurately weighed, sets in 100ml stuffed conical flask, essence
50% methanol 25ml of close addition shakes up, weighed weight, and ultrasonic treatment (power 300W, frequency 40KHz) 10 minutes is let cool to room
Temperature, then weighed weight, the weight of less loss is supplied with 50% methanol, is shaken up, and is taken supernatant to be filtered with 0.22 μm of filter membrane, is taken continuous filter
Liquid to get.
Measuring method: it is accurate respectively to measure above-mentioned reference substance solution and each 20 μ L of test solution, inject liquid chromatograph, note
Record chromatogram.It is calculated according to external standard method.Testing result is that the content of sanguisorbin I is 2.72%.
Comparative example 4:
Wave is detected with UV detector using the chromatographic condition of the assay of patent CN101810705B sanguisorbin I
A length of 210nm detects the sanguisorbin I in garden burnet crude drug powder.
Chromatographic condition: being stationary phase (4.6 × 150mm, 4 μm of partial size), methanol-water mixing with carbon octadecyl silane
Solvent gradient elution, elution program be (aqueous solution of 60% → 64% methanol of 0-7min, 64% methanol of 7-13min it is water-soluble
Liquid), flow velocity 0.8ml/min, 25 DEG C of column temperature, theoretical cam curve is calculated by sanguisorbin I should be not less than 4 × 103。
The preparation of reference substance solution: it is appropriate that precision weighs sanguisorbin I reference substance, adds 50% methanol that every 1ml is made containing 80 μ
G solution to get.
The preparation of test solution: weighing 100mg garden burnet crude drug powder, accurately weighed, sets in 100ml stuffed conical flask, essence
50% methanol 25ml of close addition shakes up, weighed weight, and ultrasonic treatment (power 300W, frequency 40KHz) 10 minutes is let cool to room
Temperature, then weighed weight, the weight of less loss is supplied with 50% methanol, is shaken up, and is taken supernatant to be filtered with 0.22 μm of filter membrane, is taken continuous filter
Liquid to get.
Measuring method: it is accurate respectively to measure above-mentioned reference substance solution and each 20 μ L of test solution, inject liquid chromatograph, note
Record chromatogram.It is calculated according to external standard method.Testing result is that the content of sanguisorbin I is 2.84%.
Comparative example 5
According to document " assay of sanguisorbin B and sanguisorbin-I in Chinese medicine radix sanguisorbae total saponin " (northern pharmacy
The 1st phase of volume 14 in 2017, page 4) method detection garden burnet crude drug powder in sanguisorbin I content, operate as follows:
The selection of chromatographic condition: chromatographic column is carried out real with Hypersil ODS chromatographic column (4.6mm × 250mm, 5.0 μm)
Test, mobile phase is acetonitrile: 0.02% phosphoric acid solution (82: 18), Detection wavelength 544nm, the column temperature of chromatographic column are 25 DEG C, sample introduction
Amount is 15 μ L, flow velocity 0.8mL/min, using sanguisorbin I chromatographic peak theoretical cam curve as reference, theoretical cam curve > 5000.
Mixed reference substance solution is prepared: sanguisorbin I reference substance is appropriate, after accurately weighed, be placed in 10mL volumetric flask, add
Enter after methanol solution is dissolved to scale and shake up, the mixed reference substance solution of known concentration content is obtained after standing, is placed in -20 DEG C of ice
It is stored in case, it is spare.
The preparation of test solution: garden burnet Chinese medicine 10g is weighed, crosses 20 after being ground into fine powder in accurately weighed rear pulverizer
Mesh is added in the ethanol solution ultrasonic instrument of 10 volumes and extracts 120min, filters, and filter residue is washed using ethanol solution
It is dissolved to after merging filtrate in 100mL volumetric flask afterwards, insufficient section solution is supplied after crossing 0.45 μm of miillpore filter, is shaken up rear quiet
It sets to get test solution.
It is calculated with external standard method, testing result is that the content of sanguisorbin I is 2.79%.
Comparative example 6
According to the content of sanguisorbin I in the method detection garden burnet crude drug powder of patent CN107064322A embodiment 1.
Chromatographic condition: InertsilODS-3 chromatographic column (250mm × 4.6mm, 5 μm, GL Sciences Inc.)；Flowing
It is mutually -0.05% phosphoric acid of acetonitrile, and gradient elution (0-12min, 5%；12-14min, 5%-36%；14-28min, 36%-
45%；28-30min, 45%-5%)；(0-26min detects gallic acid at a wavelength of 280 nm for wavelength switching；26-30min,
Sanguisorbin I is detected under 210nm wavelength)；Flow velocity 1mLmin-1；30 DEG C of column temperature；Sample volume is 20 μ L.In above-mentioned chromatostrip
Under part, between 0.95-1.05, theoretical cam curve is all larger than the symmetrical factor of gallic acid and sanguisorbin I chromatographic peak
5000。
The preparation of reference substance solution: it is appropriate that precision weighs sanguisorbin I reference substance, 50% methanol dilution to sanguisorbin I
Concentration be 1.503mgmL-1 to get.
The preparation of test solution: taking garden burnet crude drug powder 100mg, accurately weighed, and 50% methanol 25mL is added in precision, weighed
Weight is heated to reflux 1h, lets cool to room temperature, then weighed weight, supplies weight, shakes up, filtration, take subsequent filtrate to get.
It is calculated with external standard method, testing result is that the content of sanguisorbin I is 2.86%.
1-7 of the embodiment of the present invention and comparative example 2-6, with different high performance liquid chromatography method separation detections with a batch
Sanguisorbin I in secondary garden burnet crude drug powder, the result that the method for comparative example 2-6 detected are higher than embodiment 1-7
20% or more.Embodiment 1-7 and the chromatogram of comparative example 1,2 are compared, it is known that, comparative example 1,2 cannot be by garden burnet
Saponin I unknown compound proximate to it separate, so as to cause comparative example 2 testing result more higher than actual value 20% with
On.
Therefore, separation of the invention, detection and content assaying method can efficiently separate detection garden burnet or contain garden burnet
Product in sanguisorbin I, advantageously ensure that the drug safety and validity of garden burnet or the product containing garden burnet.
The preferred embodiment of the present invention has been described above in detail, still, during present invention is not limited to the embodiments described above
Detail within the scope of the technical concept of the present invention can be with various simple variants of the technical solution of the present invention are made, this
A little simple variants all belong to the scope of protection of the present invention.
Claims (7)
1. the separation method of sanguisorbin I in a kind of garden burnet or product containing garden burnet, which is characterized in that the method includes adopting
The separating step of sanguisorbin I is carried out with high performance liquid chromatography, in the high performance liquid chromatography, with octadecyl bonded silica
Glue is stationary phase, and Detection wavelength 203-216nm, mobile phase is phosphate aqueous solution: acetonitrile: methanol=(28-48): (5-25):
(37-57)。
2. separation method according to claim 1, it is characterised in that: the phosphate aqueous solution concentration is 0.01%-
0.1%.
3. separation method according to claim 1 or 2, it is characterised in that: the flow velocity of the mobile phase is 0.5-2.0mL/
Min is chosen as 0.5-1.5mL/min, or is chosen as 0.5-1.0mL/min；
Column temperature is 20 DEG C -50 DEG C, is chosen as 20 DEG C -40 DEG C；
Sample volume is 1-100 μ L, is chosen as 10-20 μ L.
4. the sanguisorbin I in separation garden burnet or product containing garden burnet of separation method described in any one of claim 1-3
Purposes.
5. the detection method of sanguisorbin I in a kind of garden burnet or product containing garden burnet, which is characterized in that the detection method packet
Include the inspection using sanguisorbin I in the progress garden burnet of separation method described in any one of claim 1-3 or the product containing garden burnet
The step of survey；
It is optionally, described that detection method includes the following steps:
(1) sanguisorbin I reference substance is taken, methanol is added to be configured to reference substance solution；
(2) sample to be tested is taken, methanol is added to extract, solution filters to obtain test solution；
(3) test solution and control solution are drawn respectively, inject high performance liquid chromatograph, are appointed using in claim 1-3
One separation method is measured；
It is optionally, described that detection method includes the following steps:
(1) sanguisorbin I reference substance is taken, methanol is added to be configured to the solution that every 1mL contains 0.05mg-0.2mg；
(2) sample to be tested is taken, is pulverized, methanol is added, is ultrasonically treated, filtration takes filtrate；
(3) test solution and control solution are drawn respectively, inject high performance liquid chromatograph, are appointed using in claim 1-3
One separation method is measured.
Optionally, the separation method includes:
(1) sanguisorbin I reference substance is taken, 50% methanol is added to be configured to the solution that every 1mL contains 0.08mg；
(2) sample to be tested is taken, is pulverized, is taken in right amount, 50% methanol is added and shakes up, weighed weight, ultrasonic treatment is let cool to room
Temperature, then weighed weight, the weight of less loss is supplied with 50% methanol, is shaken up, supernatant is taken to be filtered with filter membrane, take subsequent filtrate to get.
(3) test solution and control solution, injection high performance liquid chromatograph measurement, with octadecyl bonded silica are drawn respectively
Glue is stationary phase, and Detection wavelength 203-216nm, mobile phase is 0.01%-0.1% phosphate aqueous solution: acetonitrile: methanol=(28-
48): (5-25): (37-57), isocratic elution, 20 DEG C -40 DEG C of column temperature, flow velocity 0.5-1.0mL/min；Sample volume is 10-20 μ
L。
6. the purposes of the sanguisorbin I in detection garden burnet or product containing garden burnet of detection method described in claim 5.
7. the content assaying method of sanguisorbin I in a kind of garden burnet or product containing garden burnet, which is characterized in that described containing measuring
The method of determining includes the steps that the assay that sanguisorbin I is carried out using detection method described in claim 5；
Optionally, the content assaying method is calculated using external standard method.
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| CN116359355A (en) * | 2021-12-27 | 2023-06-30 | 江阴天江药业有限公司 | Application of sanguisorbin I in quality detection of holly leaf samples and corresponding quality detection method |
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