CN109142571A - A method of measurement anthraquinone component content - Google Patents
A method of measurement anthraquinone component content Download PDFInfo
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- CN109142571A CN109142571A CN201810947817.8A CN201810947817A CN109142571A CN 109142571 A CN109142571 A CN 109142571A CN 201810947817 A CN201810947817 A CN 201810947817A CN 109142571 A CN109142571 A CN 109142571A
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- G—PHYSICS
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
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- G01N2030/062—Preparation extracting sample from raw material
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- G01N30/02—Column chromatography
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Abstract
The present invention provides a kind of methods for measuring anthraquinone component content, it is characterized by and is combined the adsorbent that crab shell powder is extracted as matrix solid phase dispersion with the target molecule absorption in Cassia Seed, so that target molecule is separated from Cassia Seed, then use environmentally protective ionic liquid as eluant, eluent, the eluent containing target molecule is obtained, eluent carries out separation detection using HPLC；The present invention, which collects, to be extracted and is located away from one, has many advantages, such as that simple to operate, extraction disengaging time is short, extraction efficiency is high, safety and environmental protection, eluant, eluent and amount of samples are few.
Description
(1) technical field
The present invention relates to a kind of Trace bio-element detection methods of anthraquinone component.
(2) background technique
Cassia seed is legume cassia (Cassia obtusifolia L.) or little cassia tora (Cassia tora L.)
Dry mature seed.Cassia seed has effects that heat-clearing improving eyesight, relax bowel and defecation, for treating the puckery pain of hot eyes, the more tears of photophobia, headache
The dark unknown, constipation of dizziness, mesh.Modern pharmacological studies have shown that cassia seed has preferable liver protection, reducing blood lipid, blood pressure lowering, resists
Oxidation, anti-aging, the effects of improving renal function, is antibacterial, are one of the herbal species of integration of drinking and medicinal herbs that national health department announces.
Contain Multiple components in cassia seed, mainly contains anthraquinone component, including aurantio-obtusin (aurantio-obtusin), yellow Cassia
Plain (chrysoobtusin), Obtusifolin (obtusifolin), obtusin (obtusin), aloe-emodin (aloe-
Emodin), Rhein (rhein), rheum emodin (emodin), Chrysophanol (chrysophanol), Physcion
(physcion) etc..Chinese Pharmacopoeia 2015 editions using Chrysophanol and aurantio-obtusin as index Composition Control Cassia obtusifolia L and
The quality of related preparations.The common method for extracting effective component in cassia seed, as heating and refluxing extraction method, ultrasonic extraction,
Microwave extraction method (MAE) etc., generally existing consumption of organic solvent is big, environmental pollution is big, time-consuming, cumbersome, extraction efficiency
Low disadvantage.
Matrix solid phase dispersion extraction (MSPD) is a kind of sample pre-treatments new technology established on the basis of Solid Phase Extraction, tool
Have the advantages that solvent usage is few, environmental pollution is few, extraction time is short, operation is succinct, extraction efficiency is high, is increasingly becoming in recent years
Popular research both domestic and external, and widely it has been applied to Analysis of Natural Products detection.Currently, being extracted in matrix solid phase dispersion
In technology, silica gel, aluminium oxide, C are mainly selected18, florisil silica, active carbon etc. be used as adsorbent.
Crab is as one of big marine product of consumption figure highest ten, however, the crab of only sub-fraction is as food quilt
Served dining table, remaining is typically considered rubbish and is arbitrarily abandoned, cause the waste of crab shell resource with since its is low
Environmental degradation caused by biodegrade.Therefore, exploitation crab shell is also significantly as biological adsorption agent.Crab shell is by shell
Glycan, calcium carbonate and some protein composition.Wherein, chitosan has free amino on its polymeric chain, shows higher
Metallic cohesion, further, since its inexpensive, high mechanical strength, rigid structure, be resistant to drastic conditions ability and height biology
The advantages that compatibility, is applied to absorption heavy metal and other substances such as phosphate and dyestuff.Since crab shell powder is safe and non-toxic, and
Matrix solid phase dispersion is used for good absorption property, therefore as adsorbent.
(3) summary of the invention
It is an object of the invention to solve, amount of samples existing for conventional method is big, time-consuming, toxic organic solvents dosage
Big problem, the method for establishing anthraquinone component in more efficient, the more environmentally protective extraction of one kind and detection cassia seed.
The present invention is characterized by the mesh in the adsorbent and Cassia Seed for extracting crab shell powder as matrix solid phase dispersion
It marks Molecular Adsorption to combine, so that target molecule is separated from Cassia Seed, then uses environmentally protective ion
Liquid obtains the eluent containing target molecule as eluant, eluent.Eluent carries out separation detection using HPLC.It is mentioned compared to pharmacopeia
Method is taken, organic solvent usage amount of the invention greatly reduces, safety, economic, environmental protection.
Technical scheme is as follows:
A method of measurement anthraquinone component content, the anthraquinone component are aurantio-obtusin, yellow obtusin, Cassia
Anthraquinone, rheum emodin, the described method comprises the following steps:
(1) sample analysis
Sample to be tested powder is mixed with adsorbent, eluant, eluent is added after grinding uniformly, then carry out whirlpool with vortex mixer
Rotation mixing, then uses 0.22 μm of aperture membrane filtration, filtrate is taken to be centrifuged, and collecting supernatant is extracting solution, by gained extracting solution
Efficient liquid phase chromatographic analysis is carried out, sample spectrogram is obtained；
The mass ratio of the sample to be tested powder and adsorbent is 1:0.5~2, preferably 1:0.5；
The time of the grinding is 1~4min, preferably 3min；
The time of the vortex is 1~4min, preferably 3min；
The rate of the centrifugation is 13000rpm, time 6min；
The sample to be tested is, for example, cassia seed；
The adsorbent is crab shell powder；
The eluant, eluent is the aqueous solution of 150~300mM (preferably 250mM) ionic liquid, and the ionic liquid is selected from 1-
Butyl -3- methylimidazole hexafluorophosphate ([Bmim] PF6), chlorination 1- dodecyl -3- methylimidazole ([Domim] Cl), 1-
Dodecyl -3- methylimidazolium hydrogen sulphate salt ([Domim] HSO4) or 1- dodecyl -3- methylimidazolium nitrate ([Domim]
NO3), preferably 1- dodecyl -3- methylimidazolium hydrogen sulphate salt ([Domim] HSO4)；
The condition of the efficient liquid phase chromatographic analysis are as follows: chromatographic column: C18Column, 4.6mm × 250mm, 5 μm；Mobile phase: second
The mixed liquor of nitrile/water volume ratio 95:5 mixed liquor, acetonitrile/water volume ratio 5:95 contains volume fraction in the mixed liquor
0.1% formic acid；Flow velocity: 0.8~1.2mL/min, preferably 1mL/min；Column temperature: 20~30 DEG C, preferably 25 DEG C；Ultraviolet detection wave
It is long: 440nm；
(2) standard curve is established
The standard substance for weighing aurantio-obtusin, yellow obtusin, Obtusifolin, rheum emodin, is configured to by solvent of methanol
Mixed standard solution is detected under the conditions of by efficient liquid phase chromatographic analysis of the gained mixed standard solution in step (1), is obtained
It is corresponding in standard substance spectrogram using the sample volume of standard substance each in mixed standard solution as abscissa to standard substance spectrogram
The peak area of standard substance is ordinate, and it is bent to draw aurantio-obtusin, yellow obtusin, Obtusifolin, the standard of rheum emodin respectively
Line；
(3) testing result is obtained
Sample spectrogram obtained by step (1) is compareed with standard substance spectrogram obtained by step (2), obtains anthraquinone in sample
The qualitative results of constituents；
The peak area value of each anthraquinone component in sample spectrogram obtained by step (1) is substituted into respectively corresponding obtained by step (2)
Standard curve in, calculate obtain sample in anthraquinone component content.
In the present invention, the crab shell powder is prepared as follows:
Crab shell is rinsed with tap water, removes clast and mucus, then with pure water washing, is subsequently placed in 60~70 DEG C of baking
It dries in case to constant weight, dried crab shell is crushed and sieved with 100 mesh sieve, then with 20~25wt%'s at 90~100 DEG C
NaOH aqueous solution carries out the alkali process of 1h to sieving crab shell powder, to remove extra protein, later with pure water washing, and
Dry 12h, finally crushes again in 60~70 DEG C of electric furnace, crosses 200 meshes to get the crab shell powder as adsorbent.
Compared with prior art, the beneficial effects of the present invention are:
The present invention extracts the effective component in cassia seed using the technology that matrix solid phase dispersion is combined with vortex, and makes
The ionic liquid aqueous solution of green is used to replace traditional organic solvent as eluant, eluent, collection extracts and is located away from one, has behaviour
Make the advantages that simple and convenient, extraction disengaging time is short, extraction efficiency is high, safety and environmental protection, eluant, eluent and amount of samples are few.
Currently, crab shell powder is mainly concentrated use in the removal of heavy metal, phosphate and dyestuff in sewage, the present invention for the first time will
Extraction separation of the crab shell powder as adsorbent for complex matrices in study of semen cassia, has inexpensive, safe and non-toxic, high machinery
The advantages that intensity, rigid structure, ability for being resistant to drastic conditions, high-biocompatibility.
(4) Detailed description of the invention
Fig. 1 is the process flow chart of matrix solid phase dispersion extracting process of the present invention；
Fig. 2 is the characterization of crab shell powder: (a) scanning electron microscope image of (2000 times) of crab shell powder low power amplifications；(b) crab shell powder is high
The scanning electron microscope images of (50000 times) amplifications again；(c) the FT-IR spectrum of crab shell powder；
Fig. 3 is the effect of extracting histogram investigated in matrix solid phase dispersion extraction process；In figure, 1 aurantio-obtusin；2 is yellow
Obtusin；3 Obtusifolins；4 rheum emodins；(a) mass ratio of sample and adsorbent；(b) milling time；(c) vortex time；
Fig. 4 is to investigate eluant, eluent to the line chart of four kinds of anthraquinone extraction effects；In figure, 1 aurantio-obtusin；2 yellow obtusins；
3 Obtusifolins；4 rheum emodins；(a) type of ionic liquid；(b) concentration of ionic liquid.
(5) specific embodiment
The present invention is described further combined with specific embodiments below, but protection scope of the present invention is not limited in
This.
Cassia Seed used in the embodiment of the present invention is according to Chinese Pharmacopoeia version preparation in 2015.
The preparation of 1 crab shell powder of embodiment
Crab shell is rinsed with tap water, removes useless other clasts and mucus.They are placed on pure water washing again
It dries in 60 DEG C of baking oven to constant weight.Dried crab shell is ground into powder and is sieved with 100 mesh sieve.Then, it is used at 100 DEG C
20% (wt) NaOH carries out about 1 hour alkali process to sieving crab shell powder, removes extra protein.With pure water cleanser
End, and it is about 12 hours dry in 60 DEG C of electric furnace.Processed powder is crushed again, crosses 200 meshes to get as absorption
The crab shell powder of agent is used for sample pre-treatments.
The selection of the mass ratio of 2 sample of embodiment and adsorbent
Weigh 4 parts of Cassia Seed 20mg in parallel and be added separately in 4 groups of mortars, and weigh different quality (0mg, 10mg,
20mg, 40mg) crab shell powder, be separately added into 4 groups of mortars and Cassia Seed grinding 3min.4 10mL centrifuge tubes are taken, respectively
Ground solid mixture is transferred in centrifuge tube.It is separately added into [Domim] the Cl aqueous solution (250mM) of 1mL, uses vortex
Mixer carries out vortex mixed 3min, and crosses 0.22 μm of organic filter membrane, and 13000rpm is centrifuged 6min, and taking supernatant is Cassia
Sub- extracting solution fills sample, carries out sampling analysis with high performance liquid chromatograph, analysis condition is as follows:
The instrument that this experiment uses for Agilent highly effective liquid phase chromatographic system (Agilent 1260Infinity LC,
Agilent Technologies, Santa Clara, CA, USA) it is equipped with vacuum pump, binary mobile phase system, constant temperature is certainly
Dynamic sample injector, thermostatted column compartment.Chromatographic column Inersustain C18Column (4.6 × 250mm, 5 μm), 25 DEG C of column temperature.Flowing
Phase: the mixed liquor (Mobile phase B) of the mixed liquor (mobile phase A) of acetonitrile/water volume ratio 95:5, acetonitrile/water volume ratio 5:95, it is described
Formic acid containing volume fraction 0.1% in mixed liquor.Gradient elution: as shown in table 1, equilibration time 5min.Sample volume: 20 μ L,
Flow velocity: 1mL/min, Detection wavelength: 440nm.
1 gradient elution process of table
The effect of extracting histogram of the mass ratio of different samples and adsorbent is as shown in a in Fig. 3, the results showed that, work as sample
When being raised to 2:1 from 1:0 with the mass ratio of adsorbent, effect of extracting enhancing, and when the mass ratio of sample and adsorbent is 2:1,
Anthraquinone adsorption effect for all researchs is optimal.However, effect of extracting is slow when mass ratio is increased to 1:1 or 1:2
Weaken, this may be because excessive adsorbent can make active constituent be difficult to desorb, to reduce effect of extracting.
The selection of 3 milling time of embodiment
It weighs 4 parts of Cassia Seed 20mg in parallel and 4 parts of 10mg crab shell powders is added separately in 4 groups of mortars, grind respectively
The different time (1min, 2min, 3min, 4min).4 10mL centrifuge tubes are taken, respectively shift ground solid mixture
Into centrifuge tube.It is separately added into [Domim] the Cl aqueous solution (250mM) of 1mL, carries out vortex mixed 3min with vortex mixer,
And organic filter membrane of 0.22 μm of mistake, 13000rpm are centrifuged 6min, taking supernatant is cassia seed extracting solution, sample is filled, with efficient liquid
Chromatography carries out sampling analysis (analysis condition is with embodiment 2).
The effect of extracting histogram of different milling times is as shown in b in Fig. 3, the results showed that, with the increase of milling time
Effect of extracting enhancing, after increasing to optimal value 3min, upon grinding between when extending to 4min, effect of extracting declines instead.This may
It is that active constituent is caused excessively to be extracted and be extruded in hole fine and close in adsorbent since milling time is too long, increases and wash
De- difficulty, so that effect of extracting be made to decline.
The selection of 4 vortex time of embodiment
It weighs 4 parts of Cassia Seed 20mg in parallel and 4 parts of 10mg crab shell powders is added separately in 4 groups of mortars, grind 3min.
4 10mL centrifuge tubes are taken, ground solid mixture is transferred in centrifuge tube respectively.It is separately added into [Domim] Cl of 1mL
Aqueous solution (250mM) is carried out the vortex mixed different time (1min, 2min, 3min, 4min) with vortex mixer respectively, and
0.22 μm of organic filter membrane is crossed, 13000rpm is centrifuged 6min, and taking supernatant is cassia seed extracting solution, fills sample, uses efficient liquid phase
Chromatograph carries out sampling analysis (analysis condition is with embodiment 2).
The effect of extracting histogram of different vortex times is as shown in c in Fig. 3, the results showed that, with the increase of vortex time
Effect of extracting enhancing, after increasing to optimal value 3min, when vortex time extends to 4min, effect of extracting increases there is no significant
Add, illustrates that vortex 3min enough elutes the active constituent in crab shell powder.
The selection of 5 ionic liquid type of embodiment
It weighs 4 parts of Cassia Seed 20mg in parallel and 4 parts of 10mg crab shell powders is added separately in 4 groups of mortars, grind 3min.
4 10mL centrifuge tubes are taken, ground solid mixture is transferred in centrifuge tube respectively.It is separately added into [Domim] Cl of 1mL
Aqueous solution (250mM), [Domim] HSO4Aqueous solution (250mM), [Domim] NO3Aqueous solution (250mM), [Bmim] PF6Aqueous solution
(250mM) carries out vortex mixed 3min with vortex mixer, and crosses 0.22 μm of organic filter membrane, and 13000rpm is centrifuged 6min, takes
Supernatant is cassia seed extracting solution, fills sample, carries out sampling analysis with high performance liquid chromatograph (analysis condition is with embodiment 2).
Different ionic liquid type to the line chart of four kinds of anthraquinone extraction effects as shown in a in Fig. 4, [Bmim] PF6To orange
Yellow obtusin has optimal elution efficiency, but to yellow obtusin, the extraction efficiency of Obtusifolin and rheum emodin be not very well,
This may be due to [Bmim] PF6Polarity between three kinds of active constituents is different.[Domim] Cl, [Domim] HSO4With
[Domim]NO3It is different to the elution efficiency of four kinds of active constituents although long chain cation having the same, wherein [Domim]
HSO4[Domim] NO3Extraction efficiency it is better than [Domim] Cl, this may be because of anion Cl-Have between analyte
Weaker electrostatic.Comprehensively consider, with [Domim] HSO4Better extraction efficiency can be obtained as eluting solvent.
The selection of 6 ionic liquid concentration of embodiment
It weighs 4 parts of Cassia Seed 20mg in parallel and 4 parts of 10mg crab shell powders is added separately in 4 groups of mortars, grind 3min.
4 10mL centrifuge tubes are taken, ground solid mixture is transferred in centrifuge tube respectively.It is separately added into the 1mL of various concentration
[Domim] HSO4Aqueous solution (150mM, 200mM, 250mM, 300mM) carries out vortex mixed 3min with vortex mixer, and
0.22 μm of organic filter membrane is crossed, 13000rpm is centrifuged 6min, and taking supernatant is cassia seed extracting solution, fills sample, uses efficient liquid phase
Chromatograph carries out sampling analysis (analysis condition is with embodiment 2).
Different ionic liquid concentration is to the line chart of four kinds of anthraquinone extraction effects as shown in b in Fig. 4, the results showed that, with
[Domim]HSO4Increase elution effect enhancing, after increasing to optimal value 250mM, as [Domim] HSO4Concentration increases to 300mM
When, elution effect increases there is no significant, this may be to cause to be difficult to by anthraquinone from cassia seed since solution viscosity increases
It is transferred to [Domim] HSO4In aqueous solution.
Anthraquinone component quantifies in 75 kinds of different batches cassia seeds of embodiment
1, each 20mg and 5 part of 10mg crab shell powder of 5 kinds of different batches Cassia Seeds is weighed respectively is added separately to 5 groups of mortars
In, grind 3min.5 10mL centrifuge tubes are taken, ground solid mixture is transferred in centrifuge tube respectively.It is added 1mL's
[Domim]HSO4Aqueous solution (250mM) carries out vortex mixed 3min with vortex mixer, and crosses 0.22 μm of organic filter membrane,
13000rpm is centrifuged 6min, takes supernatant to fill sample, carries out sampling analysis (the same embodiment of analysis condition with high performance liquid chromatograph
2)。
Anthraquinone component is quantitative as shown in table 2 in 5 kinds of different batches cassia seeds:
The measurement result (mg/g) of four kinds of active constituents in 25 kinds of different batches cassia seeds of table
aRaw cassia seed
bFried cassia
2, the formulation of standard curve:
The accurate reference substance for weighing aurantio-obtusin, yellow obtusin, Obtusifolin and rheum emodin respectively, it is molten with methanol respectively
Solution is made into the reference substance solution that concentration is 1.03mg/mL, 0.5mg/mL, 0.52mg/mL and 0.22mg/mL.By control obtained
Product solution is configured to containing 123.6 μ g/mL of aurantio-obtusin, 40.0 μ g/mL of yellow obtusin, 41.6 μ g/mL of Obtusifolin, rheum emodin
The mixed reference substance solution of 13.2 μ g/mL.The mixing control of 0.5 μ L, 1 μ L, 2 μ L, 5 μ L, 10 μ L, 15 μ L and 20 μ L are drawn respectively
Product solution injection liquid chromatograph is analyzed, and peak area is measured.Using sample volume as abscissa, peak area is ordinate, is drawn
Standard curve.
Aurantio-obtusin, yellow obtusin, Obtusifolin, the standard curve of rheum emodin, linearly dependent coefficient, the range of linearity and
Detection limit such as the following table 3:
Each ingredient standard curvilinear equation of table 3, linearly dependent coefficient, the range of linearity and detection limit
3, repeatability is investigated
Cassia Seed 20mg and 10mg crab shell powder is weighed in mortar, grinds 3min.By ground solid mixture
It is transferred in the centrifuge tube of 10mL.[Domim] HSO of 1mL is added4Aqueous solution (250mM) be vortexed with vortex mixer mixed
3min is closed, and crosses 0.22 μm of organic filter membrane, 13000rpm is centrifuged 6min, and supernatant is taken to fill sample.Parallel system according to the method described above
6, sample, its repeatability is investigated under above-mentioned chromatographic condition, the results are shown in Table 4.
4, precision is investigated
Under above-mentioned chromatographic condition, precision draws mixed reference substance solution interior repetition sample introduction 6 times on the same day, is in a few days accurate
Degree.Under above-mentioned chromatographic condition, precision draws standard solution, and daily sample introduction 2 times for three days on end are day to day precision.As a result see
Table 4.
5, study on the stability
Cassia Seed 20mg and 10mg crab shell powder is weighed in mortar, grinds 3min.By ground solid mixture
It is transferred in the centrifuge tube of 10mL.[Domim] HSO of 1mL is added4Aqueous solution (250mM) be vortexed with vortex mixer mixed
3min is closed, and crosses 0.22 μm of organic filter membrane, 13000rpm is centrifuged 6min, and take supernatant to fill sample, under above-mentioned chromatographic condition, point
Not in 2,4,8,16, measure peak area for 24 hours, investigate its stability, the results are shown in Table 4.
4 repeatability of table, precision, stability experiment result
Data are shown in table, and the method for the present invention is reproducible, stability is good, and the precision of instrument is good.
6, sample-adding recycling is investigated
Cassia Seed 10mg and 10mg crab shell powder is weighed in mortar, grinds 3min.By ground solid mixture
It is transferred in the centrifuge tube of 10mL.[Domim] HSO of 1mL is added4Aqueous solution (250mM) (containing 9.11 μ g/mL aurantio-obtusins,
4.20 μ g/mL Huang obtusins, 2.93 μ g/mL Obtusifolins, 0.77 μ g/mL rheum emodin), vortex mixed is carried out with vortex mixer
3min, and 0.22 μm of organic filter membrane is crossed, 13000rpm is centrifuged 6min, takes supernatant to fill sample, carries out under above-mentioned chromatographic condition
Analysis.
Cassia Seed 10mg and 10mg crab shell powder is weighed in mortar, grinds 3min.By ground solid mixture
It is transferred in the centrifuge tube of 10mL.[Domim] HSO of 1mL is added4Aqueous solution (250mM) (contains the 18.22 orange Cassia of μ g/mL
Element, 8.40 μ g/mL Huang obtusins, 5.86 μ g/mL Obtusifolins, 1.54 μ g/mL rheum emodins), it is vortexed with vortex mixer
3min is mixed, and crosses 0.22 μm of organic filter membrane, 13000rpm is centrifuged 6min, takes supernatant to fill sample, under above-mentioned chromatographic condition
It is analyzed.
Cassia Seed 10mg and 10mg crab shell powder is weighed in mortar, grinds 3min.By ground solid mixture
It is transferred in the centrifuge tube of 10mL.[Domim] HSO of 1mL is added4Aqueous solution (250mM) (contains the 27.33 orange Cassia of μ g/mL
Element, 12.40 μ g/mL Huang obtusins, 8.79 μ g/mL Obtusifolins, 2.31 μ g/mL rheum emodins), it is vortexed with vortex mixer
3min is mixed, and crosses 0.22 μm of organic filter membrane, 13000rpm is centrifuged 6min, takes supernatant to fill sample, under above-mentioned chromatographic condition
It is analyzed.
Sample recovery rate experimental result such as the following table 5:
The experiment of 5 sample recovery rate of table
Data are shown in table, and the method for the present invention rate of recovery is high.
The above embodiment is a preferred embodiment of the present invention, but embodiments of the present invention are not by above-described embodiment
Limitation, other any changes, modifications, substitutions, combinations, simplifications made without departing from the spirit and principles of the present invention,
It should be equivalent substitute mode, be included within the scope of the present invention.
Claims (9)
1. a kind of method for measuring anthraquinone component content, the anthraquinone component is aurantio-obtusin, yellow obtusin, Cassia anthracene
Quinone, rheum emodin, which is characterized in that the described method comprises the following steps:
(1) sample analysis
Sample to be tested powder is mixed with adsorbent, eluant, eluent is added after grinding uniformly, then be vortexed with vortex mixer and mix
It closes, then uses 0.22 μm of aperture membrane filtration, filtrate is taken to be centrifuged, collecting supernatant is extracting solution, and gained extracting solution is carried out
Efficient liquid phase chromatographic analysis obtains sample spectrogram；
The adsorbent is crab shell powder；
The eluant, eluent is the aqueous solution of 150~300mM ionic liquid, and the ionic liquid is selected from 1- butyl -3- methylimidazole
Hexafluorophosphate, chlorination 1- dodecyl -3- methylimidazole, 1- dodecyl -3- methylimidazolium hydrogen sulphate salt or 1- dodecane
Base -3- methylimidazolium nitrate；
The condition of the efficient liquid phase chromatographic analysis are as follows: chromatographic column: C18Column, 4.6mm × 250mm, 5 μm；Mobile phase: acetonitrile/water
The mixed liquor of the mixed liquor of volume ratio 95:5, acetonitrile/water volume ratio 5:95 contains volume fraction 0.1% in the mixed liquor
Formic acid；Flow velocity: 0.8~1.2mL/min；Column temperature: 20~30 DEG C；Ultraviolet detection wavelength: 410nm；
(2) standard curve is established
The standard substance for weighing aurantio-obtusin, yellow obtusin, Obtusifolin, rheum emodin, is configured to mix using methanol as solvent
Standard solution is detected under the conditions of by efficient liquid phase chromatographic analysis of the gained mixed standard solution in step (1), is marked
Quasi- substance spectrogram, using the sample volume of standard substance each in mixed standard solution as abscissa, respective standard in standard substance spectrogram
The peak area of substance is ordinate, draws the standard curve of aurantio-obtusin, yellow obtusin, Obtusifolin, rheum emodin respectively；
(3) testing result is obtained
Sample spectrogram obtained by step (1) is compareed with standard substance spectrogram obtained by step (2), in acquisition sample Anthraquinones at
The qualitative results divided；
The peak area value of each anthraquinone component in sample spectrogram obtained by step (1) is substituted into corresponding mark obtained by step (2) respectively
In directrix curve, the content for obtaining anthraquinone component in sample is calculated.
2. the method as described in claim 1, which is characterized in that the crab shell powder is prepared as follows:
Crab shell is rinsed with tap water, removes clast and mucus, then with pure water washing, is subsequently placed in 60~70 DEG C of baking oven
Dried crab shell is crushed and is sieved with 100 mesh sieve, then with the NaOH of 20~25wt% at 90~100 DEG C to constant weight by drying
Aqueous solution carries out the alkali process of 1h to sieving crab shell powder, to remove extra protein, later with pure water washing, and 60
Dry 12h, finally crushes again in~70 DEG C of electric furnace, crosses 200 meshes to get the crab shell powder as adsorbent.
3. the method as described in claim 1, which is characterized in that in step (1), the sample to be tested is cassia seed.
4. the method as described in claim 1, which is characterized in that in step (1), the matter of the sample to be tested powder and adsorbent
Amount is than being 1:0.5~2.
5. the method as described in claim 1, which is characterized in that in step (1), the time of the grinding is 1~4min.
6. the method as described in claim 1, which is characterized in that in step (1), the time of the vortex is 1~4min.
7. the method as described in claim 1, which is characterized in that in step (1), the rate of the centrifugation is 13000rpm, when
Between be 6min.
8. the method as described in claim 1, which is characterized in that in step (1), the eluant, eluent is 250mM ionic liquid
Aqueous solution.
9. the method as described in claim 1, which is characterized in that in step (1), the ionic liquid is selected from 1- dodecyl-
3- methylimidazolium hydrogen sulphate salt.
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