CN106442777A - Method for identifying Fujian Guanxi honey pomelos by chromatography method - Google Patents
Method for identifying Fujian Guanxi honey pomelos by chromatography method Download PDFInfo
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Abstract
The invention discloses a method for identifying Fujian Guanxi honey pomelos by a chromatography method. According to the method, firstly, acetonitrile is used for extracting chemical ingredients in each position of outer peels, endodermis, endocarp, pulp and the like of the honey pomelos; then, a gas chromatograph, a hydrogen flame ionization detector and a liquid chromatogram-ultraviolet detector is used at the same time for detection; chemical ingredients in each position of the Fujian Guanxi honey pomelos are subjected to chromatographic qualitative analysis; specific substances of cnidium lactone, limonene, alpha-linoleic acid, palmitic acid, sitosterol, oleamide and nootkatone and the like in the Fujian Guanxi honey pomelos can be accurately detected; a honey pomelo specific substance chromatographic fingerprint chromatogram is successfully built. The method provided by the invention has the advantages that the operation is simple; the qualitative detection is accurate; the detection is fast; the method belongs to a fire-new fast, accurate and simple honey pomelo origin traceability and quality identification method.
Description
Technical field
The invention belongs to analytical chemistry field of chromatography.Specifically a kind of employing chromatography identifies the side of Fujian small stream honey Fructus Citri grandiss
Method, this method carries out qualitative analyses using gas chromatogram and liquid chromatography to the chemical constituent in Fujian small stream honey Fructus Citri grandiss simultaneously.
Background technology
Fructus Citri grandiss (citrus junos tanaka), also known as pomelo, are Rutaceae Aurantioideae Fructus Citri grandiss platymisciums, and China is
Fructus Citri grandiss class cultivated area is maximum in the world, and yield occupies the big producing country of the 2nd.China's Fructus Citri grandiss veriety aboundresources, kind (strain)
Various, current main breed has sweet Fructus Citri grandiss, small stream honey Fructus Citri grandiss, Yuhuan pomelo etc., accounts for the 80% about of national Fructus Citri grandiss cultivated area, growing surface
Amass and all reach more than 0.37 ten thousand hectares, annual production, more than 1.2 ten thousand tons, is distributed mainly on Fujian, Zhejiang, Taiwan, Guangxi, Guangdong etc. many
Individual province.Small stream honey Fructus Citri grandiss originate in Fujian Province Pinghe County, existing more than 500 year cultivation history.Nineteen ninety-five passes through national farming article
Plant validation board's authorization.Pinghe County plants more than 50 ten thousand hectares within 2009, and gross annual output amount reaches more than 100 ten thousand tons, and year creates the output value 1,500,000,000
Polynary, become Fujian Province Pinghe County fruit industry pillar industry, be also the important channel that orchard worker gets rich.Honey Fructus Citri grandiss successively have registered small stream
" small stream " trade mark, " gentle small stream honey Fructus Citri grandiss " certification mark, obtain country, " pollution-free food " certification and State General Administration for Quality Supervision and originate in
Ground Taken authentication, carries out international registration of trademarks in 17 countries and regions such as the U.S., Britain, France respectively, is proposed as and Europe
One of ten big geography symbol products of alliance's exchange protection, are identified as " Chinese brand name agricultural product ", but are as the nutrition of Fructus Citri grandiss
It is worth and medical value is increasingly subject to pay attention to, the phenomenon palming off famous-brand and high-quality kind Fructus Citri grandiss happens occasionally, and quality grade divides and originates in
The identification technology on ground is just particularly important.The present invention utilizes gas chromatogram and liquid chromatography technology means to Fu Jianxi simultaneously
The chemical constituent of sweet Fructus Citri grandiss carries out qualitative analyses, is successfully established sweet Fructus Citri grandiss specificity substance chromatographic fingerprinting.Therefore, set up a kind of letter
List, accurately and rapidly chromatogram analysis method, trace to the source to the original producton location of Fujian small stream honey Fructus Citri grandiss and identification have great importance.
For the identification technology of Fructus Citri grandiss, at present, headspace solid-phase microextraction-Gas chromatographyMass spectrometry (Chinese agriculture
College journal, 2011,16 (6):52), sensory evaluation and low-polarity components (modern food science and technology, 2014,30 (10):274),
Chemical composition Systematic pretest method (China National folk medicine, 2010,5:35-36), HPLC-MS technology
(chromatograph, 2007,25 (5):770-771), chromatograph-mass spectrometer coupling technology (chromatograph, 2016,34 (6):558-566) etc. method has been
It is used for the qualitative analyses of Fructus Citri grandiss.Headspace solid-phase microextraction-Gas chromatographyMass spectrometry is simple and quick, is suitable for detection Fructus Citri grandiss
Volatile ingredient in son, but the involatile constituent detection in improper Fructus Citri grandiss；HPLC-MS technology is high
Accurately, the involatile constituent of suitable Fructus Citri grandiss detects effect, but is not suitable for the detection of volatile ingredient.Currently without discovery report
Using gas chromatogram or liquid chromatography, Testing and appraisal is carried out to sweet Fructus Citri grandiss.
Content of the invention
The present invention, in order to overcome the shortcomings of existing identification technology, provides a kind of employing chromatography to identify Fujian small stream honey Fructus Citri grandiss
Method, to the chemical constituent Qualitative Identification in Fujian small stream honey Fructus Citri grandiss.
The technical scheme that the present invention solves above-mentioned technical problem is as follows：
A kind of method that employing chromatography identifies Fujian small stream honey Fructus Citri grandiss, initially with acetonitrile to sweet Fructus Citri grandiss exocuticle, endepidermis,
Organic substance in each position such as endocarp and sarcocarp is extracted, and then adopts gas chromatograph, hydrogen flame ion detection simultaneously
Device and liquid chromatograph-UV-detector are detected, carry out chromatographic qualitative analysis to Organic substance in the Fujian small stream honey each position of Fructus Citri grandiss,
Detect exactly osthole in Fujian small stream honey Fructus Citri grandiss, limonene, α-linoleic acid, Palmic acid, sitosterol, oleamide and
The specificity substances such as (+)-Nootkatone, are successfully established sweet Fructus Citri grandiss specificity substance chromatographic fingerprinting：
1) adopt the method that Fujian small stream honey Fructus Citri grandiss identified by gas chromatograph, hydrogen flame ionization detector, operating procedure is as follows：
1.1) the chromatographic column initial temperature of gas chromatograph is room temperature, and 10 DEG C/min of heating rate rises to temperature 50～280
DEG C, the peak for meeting all the components in sample is kept completely separate, and in conjunction with each boiling point substance, determines gas chromatogram optimum analysis condition such as
Under：
Gas chromatographicanalyzer analysis condition
Chromatographic column：30m × 0.25mm DB-17 chromatographic column；Carrier gas：N2；Flow rate of carrier gas：1mL/min；Chromatogram column temperature：
120℃；Sample size：1μL；Gasification room temperature：260℃；Detector temperature：280℃；Detector is fid detector；
1.2) preparation of external standard solution：According to the mass fraction of external standard, weigh respectively different quality osthole,
α-linoleic acid, Palmic acid and (+)-Nootkatone, are added in the volumetric flask of 10mL with acetonitrile respectively, are settled to scale with acetonitrile, are made into
Concentration is 1000mg/L external standard solution, and is made into the osthole that concentration is 100mg/L, α-linoleic acid, Palmic acid and Nuo Ka
The hybrid standard External standards solutions of ketone；
1.3) preparation of detection sample：Accurately weigh 5g sample (fresh weight) in 100mL centrifuge tube, add 25mL acetonitrile,
Vortex 2min, ultrasonic 30min, 4500r/min are centrifuged 3min, extract supernatant 1.5mL, filter through 0.22 μm of mocromembrane, treat test sample
The osthole of product, α-linoleic acid, Palmic acid and (+)-Nootkatone；
2) adopt the method that liquid chromatograph-UV-detector identifies Fujian small stream honey Fructus Citri grandiss, operating procedure is as follows：
2.1), with 254nm for uv absorption wavelength, gradient elution, for meeting the peak of all the components in sample for chromatograph of liquid
It is kept completely separate, in conjunction with each material polarity, determine that liquid chromatograph optimum analysis condition is as follows：
Chromatographic analyzer of liquid phase analysis condition
Chromatographic column：Zorbax Eclipse XDB C18 250mm × 4.6mm × 5 μm chromatographic column；Mobile phase：A is aqueous phase,
B is acetonitrile；Elution flow rate：1mL/min；Chromatogram column temperature：30℃；Sample size：10μL；Gradient elution program：Water A%:Acetonitrile
B% is 70:30, eluting 0～15min, water A%:Acetonitrile B% is 40:60, eluting 15～20min, water A%:Acetonitrile B% is 40:
60, eluting 20～40min, water A%:Acetonitrile B% is 20:80, eluting 40～45min, water A%:Acetonitrile B% is 20:80, eluting
45～65min, water A%:Acetonitrile B% is 70:30, eluting 65～70min；Detector is UV-detector；
2.2) preparation of external standard solution：According to the mass fraction of external standard, weigh limonene, the paddy of different quality respectively
Sterol and oleamide, are added in the volumetric flask of 10mL with acetonitrile dissolving respectively, are settled to scale with acetonitrile, being made into concentration is
1000mg/L external standard solution, and it is made into the mixed of the osthole that concentration is 100mg/L, α-linoleic acid, Palmic acid and (+)-Nootkatone
Standardization External standards solutions；
2.3) preparation of detection sample：Accurately weigh the fresh sample of 5g in 100mL centrifuge tube, add 25mL acetonitrile, be vortexed
2min, ultrasonic 30min, 4500r/min are centrifuged 3min, extract supernatant 1.5mL, filter through 0.22 μm of mocromembrane, testing sample
Osthole, α-linoleic acid, Palmic acid and (+)-Nootkatone.
The gas chromatograph of the present invention is Agilent7890A gas chromatograph, and chromatographic column is that separation efficiency is high, separates speed
Spend fast quartzy capillary column, detected with hydrogen flame detector, each component in sample and external standard are carried out one a pair
Ratio accurately detects the chemical constituent in the small stream honey Fructus Citri grandiss of Fujian.Chromatograph of liquid is Water-e2695 high performance liquid chromatography
Instrument, chromatographic column is Zorbax Eclipse XDB C18 reversed-phase column, with UV-detector, to each component in sample and external standard
Thing is contrasted one by one, accurately detects the chemical constituent in the small stream honey Fructus Citri grandiss of Fujian.
The present invention compared with prior art, has the following advantages that：
1. change the deficiency in the past judging the Fructus Citri grandiss place of production and quality technically science with the method for sensory evaluation.
2. measure from chromatographic technique, inclusion diversity, being identified, result is more objective to the place of production of sweet Fructus Citri grandiss, quality, more
Accurately more scientific.
3. adopt gas chromatogram and liquid chromatography technology simultaneously, both can detect volatile material, and can detect again
Polar substancess, the more scientific reliability of result.
4., in the non-overloading scope of chromatographic column, qualitative results are unrelated with sample size repeatability.
5. simple to operate, qualitative accurately, detection quick it is adaptable to qualitative point of chemical constituent in sweet Fructus Citri grandiss and other Fructus Citri grandiss
Analysis.
Brief description
Fig. 1 is the gas chromatogram of external standard osthole.
Fig. 2 is the gas chromatogram of external standard (+)-Nootkatone.
Fig. 3 is the gas chromatogram of external standard Palmic acid.
Fig. 4 is external standard α-linoleic gas chromatogram.
The gas chromatogram of Fig. 5 hybrid standard external standard.
Fig. 6 is the liquid chromatogram of external standard sitosterol.
Fig. 7 is the liquid chromatogram of external standard oleamide.
Fig. 8 is the liquid chromatogram of external standard limonene.
The liquid chromatogram of Fig. 9 hybrid standard external standard.
Figure 10 is the gas chromatogram of Fujian small stream honey Fructus Citri grandiss exocuticle sample.
Figure 11 is the gas chromatogram of Fujian small stream honey Fructus Citri grandiss endepidermis sample.
Figure 12 is the gas chromatogram of Fujian small stream honey Fructus Citri grandiss endocarp sample.
Figure 13 is the gas chromatogram of Fujian small stream honey Fructus Citri grandiss Meat Sample.
Figure 14 is the liquid chromatogram of Fujian small stream honey Fructus Citri grandiss exocuticle sample.
Figure 15 is the liquid chromatogram of Fujian small stream honey Fructus Citri grandiss endepidermis sample
Figure 16 is the liquid chromatogram of Fujian small stream honey Fructus Citri grandiss endocarp sample
Figure 17 is the liquid chromatogram of Fujian small stream honey Fructus Citri grandiss Meat Sample
Specific embodiment
In following examples, the instrument of application is included with reagent：
Instrument and material：Agilent 7890A gas chromatograph, band hydrogen flame ionization detector (FID)；DB-17 type stone
English capillary chromatographic column, 30m × 0.25mm；Carrier gas is more than 99.999% nitrogen for purity, and combustion gas is more than for purity
99.999% hydrogen, combustion-supporting gas is the compressed air of 0.5MPa；Water-e2695 high performance liquid chromatograph, carries ultraviolet detection
Device and chromatographic work station；Zorbax Eclipse XDB C18 chromatographic column, 250mm × 4.6mm × 5 μm.
Reagent：Acetonitrile (analyzing pure, chromatographically pure), ultra-pure water.
External standard：Osthole, limonene, α-linoleic acid, Palmic acid, sitosterol, oleamide and (+)-Nootkatone.
Sample：Small stream honey Fructus Citri grandiss are picked up under Fujian Zhang Zhou Pinghe County Wen Feng town Chai Chuan village bank in Fujian.
Embodiment 1
The method identifying Fujian small stream honey Fructus Citri grandiss using gas chromatograph, hydrogen flame ionization detector, concrete operation step is such as
Under：
1. the chromatographic column initial temperature of gas chromatograph is room temperature, and 10 DEG C/min of heating rate rises to 50～280 DEG C of temperature,
Peak for meeting all the components in sample is kept completely separate, and in conjunction with each boiling point substance, determines that gas chromatogram optimum analysis condition is as follows：
Gas chromatographicanalyzer analysis condition
Chromatographic column：30m × 0.25mm DB-17 chromatographic column；Carrier gas：N2；Flow rate of carrier gas：1mL/min；Chromatogram column temperature：
120℃；Sample size：1μL；Gasification room temperature：260℃；Detector temperature：280℃；Detector is fid detector.
2. the preparation of external standard solution：According to the mass fraction of external standard, weigh respectively the osthole of different quality, α-
Linoleic acid, Palmic acid and (+)-Nootkatone, are added in the volumetric flask of 10mL with acetonitrile dissolving respectively, are settled to scale with acetonitrile, join
Become 1000mg/L external standard solution, and be made into the hybrid standard external standard solution that concentration is 100mg/L.Through gas chromatographic detection,
As shown in accompanying drawing 1, Fig. 2, Fig. 3, Fig. 4 and Fig. 5, Fig. 1 is osthole gas phase spectrogram to its testing result；Fig. 2 is (+)-Nootkatone gas phase
Chromatogram；Fig. 3 is Palmic acid gas chromatogram；Fig. 4 is α-linoleic acid gas chromatogram；Fig. 5 is hybrid standard external standard gas phase
Chromatogram.
3. the preparation of sample：Accurately weigh 5g sample (fresh weight) in 100mL centrifuge tube, add 25mL acetonitrile, be vortexed
2min, ultrasonic 30min, 4500r/min are centrifuged 3min, extract supernatant 1.5mL, filter through 0.22 μm of mocromembrane, through gas chromatogram
Detection, as shown in accompanying drawing 10, Figure 11, Figure 12 and Figure 13, Figure 10 is exocuticle sample spectrogram to its testing result；Figure 11 is endepidermis
Sample spectrogram；Figure 12 is endocarp sample spectrogram；Figure 13 is Meat Sample spectrogram.
Embodiment 2
The method identifying Fujian small stream honey Fructus Citri grandiss using liquid chromatograph-UV-detector, concrete operation step is as follows：
1. chromatograph of liquid is with 254nm for uv absorption wavelength, gradient elution, and the peak for meeting all the components in sample is complete
Fully separating, in conjunction with each material polarity, determine that liquid chromatograph optimum analysis condition is as follows：
Chromatographic analyzer of liquid phase analysis condition
Chromatographic column：Zorbax Eclipse XDB C18 250mm × 4.6mm × 5 μm chromatographic column；Mobile phase：A is aqueous phase,
B is acetonitrile；Elution flow rate：1mL/min；Chromatogram column temperature：30℃；Sample size：10μL；Gradient elution program：Water A%:Acetonitrile
B% is 70:30, eluting 0～15min, water A%:Acetonitrile B% is 40:60, eluting 15～20min, water A%:Acetonitrile B% is 40:
60, eluting 20～40min, water A%:Acetonitrile B% is 20:80, eluting 40～45min, water A%:Acetonitrile B% is 20:80, eluting
45～65min, water A%:Acetonitrile B% is 70:30, eluting 65～70min；Detector is UV-detector；
2. the preparation of external standard solution：According to the mass fraction of external standard, weigh limonene, the paddy steroid of different quality respectively
Alcohol and oleamide, are added in the volumetric flask of 10mL with acetonitrile dissolving respectively, are settled to scale with acetonitrile, are made into 1000mg/L
External standard solution, and it is made into the hybrid standard External standards solutions that concentration is 100mg/L.Through liquid chromatographic detection, its testing result is such as
Shown in accompanying drawing 6, Fig. 7, Fig. 8 and Fig. 9, Fig. 6 is the liquid chromatogram of sitosterol；Fig. 7 is the liquid chromatogram of oleamide；Fig. 8
Liquid chromatogram for limonene；Fig. 9 is the liquid chromatogram of hybrid standard external standard.
3. the preparation of sample：Accurately weigh 5g sample (fresh weight) in 100mL centrifuge tube, add 25mL acetonitrile, be vortexed
2min, ultrasonic 30min, 4500r/min are centrifuged 3min, extract supernatant 1.5mL, filter through 0.22 μm of mocromembrane, through liquid chromatograph
Detection, as shown in accompanying drawing 14, Figure 15, Figure 16 and Figure 17, Figure 14 is the liquid chromatogram of exocuticle sample to its testing result；Figure 15
Liquid chromatogram for endepidermis sample；Figure 16 is the liquid chromatogram of endocarp sample；Figure 17 is the liquid phase color of Meat Sample
Spectrogram.
Claims (1)
1. a kind of employing chromatography identifies the method for Fujian small stream honey Fructus Citri grandiss it is characterised in that initially with acetonitrile to sweet Fructus Citri grandiss appearance
Organic substance in each position such as skin, endepidermis, endocarp and sarcocarp is extracted, and then adopts gas chromatograph, hydrogen fire simultaneously
Flame ion detector and liquid chromatograph-UV-detector are detected, carry out color to Organic substance in the Fujian small stream honey each position of Fructus Citri grandiss
Spectrum qualitative analyses, detect osthole in Fujian small stream honey Fructus Citri grandiss, limonene, α-linoleic acid, Palmic acid, paddy steroid exactly
The specificity substances such as alcohol, oleamide and (+)-Nootkatone, are successfully established sweet Fructus Citri grandiss specificity substance chromatographic fingerprinting；
1) adopt the method that Fujian small stream honey Fructus Citri grandiss identified by gas chromatograph, hydrogen flame ionization detector, operating procedure is as follows：
1.1) the chromatographic column initial temperature of gas chromatograph is room temperature, and 10 DEG C/min of heating rate rises to 50～280 DEG C of temperature, is
The peak meeting all the components in sample is kept completely separate, and in conjunction with each boiling point substance, determines that gas chromatogram optimum analysis condition is as follows：
Gas chromatographicanalyzer analysis condition
Chromatographic column：30m × 0.25mm DB-17 chromatographic column；Carrier gas：N2；Flow rate of carrier gas：1mL/min；Chromatogram column temperature：120℃；
Sample size：1μL；Gasification room temperature：260℃；Detector temperature：280℃；Detector is fid detector；
1.2) preparation of external standard solution：According to the mass fraction of external standard, weigh osthole, the α-Asia of different quality respectively
Oleic acid, Palmic acid and (+)-Nootkatone, are added in the volumetric flask of 10mL with acetonitrile respectively, are settled to scale with acetonitrile, are made into concentration
For 1000mg/L external standard solution, and it is made into the osthole that concentration is 100mg/L, α-linoleic acid, Palmic acid and (+)-Nootkatone
Hybrid standard External standards solutions；
1.3) preparation of detection sample：Accurately weigh 5g sample (fresh weight) in 100mL centrifuge tube, add 25mL acetonitrile, be vortexed
2min, ultrasonic 30min, 4500r/min are centrifuged 3min, extract supernatant 1.5mL, filter through 0.22 μm of mocromembrane, testing sample
Osthole, α-linoleic acid, Palmic acid and (+)-Nootkatone；
2) adopt the method that liquid chromatograph-UV-detector identifies Fujian small stream honey Fructus Citri grandiss, operating procedure is as follows：
2.1) chromatograph of liquid is with 254nm for uv absorption wavelength, gradient elution, and the peak for meeting all the components in sample is complete
Separate, in conjunction with each material polarity, determine that liquid chromatograph optimum analysis condition is as follows：
Chromatographic analyzer of liquid phase analysis condition
Chromatographic column：Zorbax Eclipse XDB C18 250mm × 4.6mm × 5 μm chromatographic column；Mobile phase：A is aqueous phase, and B is
Acetonitrile；Elution flow rate：1mL/min；Chromatogram column temperature：30℃；Sample size：10μL；Gradient elution program：Water A%:Acetonitrile B%
For 70:30, eluting 0～15min, water A%:Acetonitrile B% is 40:60, eluting 15～20min, water A%:Acetonitrile B% is 40:60,
Eluting 20～40min, water A%:Acetonitrile B% is 20:80, eluting 40～45min, water A%:Acetonitrile B% is 20:80, eluting 45
～65min, water A%:Acetonitrile B% is 70:30, eluting 65～70min；Detector is UV-detector；
2.2) preparation of external standard solution：According to the mass fraction of external standard, weigh limonene, the sitosterol of different quality respectively
And oleamide, it is added in the volumetric flask of 10mL with acetonitrile dissolving respectively, be settled to scale with acetonitrile, being made into concentration is
1000mg/L external standard solution, and it is made into the mixed of the osthole that concentration is 100mg/L, α-linoleic acid, Palmic acid and (+)-Nootkatone
Standardization External standards solutions；
2.3) preparation of detection sample：Accurately weigh the fresh sample of 5g in 100mL centrifuge tube, addition 25mL acetonitrile, vortex 2min,
Ultrasonic 30min, 4500r/min are centrifuged 3min, extract supernatant 1.5mL, filter through 0.22 μm of mocromembrane, the Fructus Cnidii of testing sample
Element, α-linoleic acid, Palmic acid and (+)-Nootkatone.
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| CN201610823343.7A CN106442777B (en) | 2016-09-14 | 2016-09-14 | A method of Fu Jian Guanxi small stream honey shaddock is identified using chromatography |
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1959407A (en) * | 2006-10-20 | 2007-05-09 | 中山大学 | Method for constructing fingerprint HPLC of Shatian pomelo, and normal fingerprint |
| CN102787017A (en) * | 2012-05-15 | 2012-11-21 | 浙江大学 | Extraction process and component analysis of pomelo peel essential oil, and employed response surface optimization method |
-
2016
- 2016-09-14 CN CN201610823343.7A patent/CN106442777B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1959407A (en) * | 2006-10-20 | 2007-05-09 | 中山大学 | Method for constructing fingerprint HPLC of Shatian pomelo, and normal fingerprint |
| CN102787017A (en) * | 2012-05-15 | 2012-11-21 | 浙江大学 | Extraction process and component analysis of pomelo peel essential oil, and employed response surface optimization method |
Non-Patent Citations (3)
| Title |
|---|
| XIAO-QI ZHANG 等: "A new coumarin from Citrus grandis‘Shatianyu’", 《BIOCHEMICAL SYSTEMATICS AND ECOLOGY》 * |
| 嵇海峰 等: "3种福建省柚子汁挥发性成分及香气分析", 《集美大学学报（自然科学版）》 * |
| 陈兆杰 等: "色谱质谱联用技术在我国3种柚子组分鉴定及指纹图谱建立中的应用", 《色谱》 * |
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