CN104792915B - Method for determining nystose content of morinda officinalis roots - Google Patents
Method for determining nystose content of morinda officinalis roots Download PDFInfo
- Publication number
- CN104792915B CN104792915B CN201510163742.0A CN201510163742A CN104792915B CN 104792915 B CN104792915 B CN 104792915B CN 201510163742 A CN201510163742 A CN 201510163742A CN 104792915 B CN104792915 B CN 104792915B
- Authority
- CN
- China
- Prior art keywords
- solution
- reference substance
- water
- sugar
- lamellae
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- 238000000034 method Methods 0.000 title claims abstract description 35
- 241000096284 Gynochthodes officinalis Species 0.000 title abstract description 5
- FLDFNEBHEXLZRX-DLQNOBSRSA-N Nystose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)OC[C@]1(OC[C@]2(O[C@@H]3[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O3)O)[C@H]([C@H](O)[C@@H](CO)O2)O)[C@@H](O)[C@H](O)[C@@H](CO)O1 FLDFNEBHEXLZRX-DLQNOBSRSA-N 0.000 title abstract 2
- FLDFNEBHEXLZRX-UHFFFAOYSA-N nystose Natural products OC1C(O)C(CO)OC1(CO)OCC1(OCC2(OC3C(C(O)C(O)C(CO)O3)O)C(C(O)C(CO)O2)O)C(O)C(O)C(CO)O1 FLDFNEBHEXLZRX-UHFFFAOYSA-N 0.000 title abstract 2
- 238000004445 quantitative analysis Methods 0.000 claims abstract description 9
- 230000010354 integration Effects 0.000 claims abstract description 8
- 239000000243 solution Substances 0.000 claims description 75
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 57
- 239000013558 reference substance Substances 0.000 claims description 47
- 238000012360 testing method Methods 0.000 claims description 44
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 42
- 239000012567 medical material Substances 0.000 claims description 27
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 claims description 26
- 239000000523 sample Substances 0.000 claims description 24
- 239000000706 filtrate Substances 0.000 claims description 21
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 claims description 20
- 238000010438 heat treatment Methods 0.000 claims description 17
- 229960000583 acetic acid Drugs 0.000 claims description 16
- 238000001914 filtration Methods 0.000 claims description 16
- 239000002904 solvent Substances 0.000 claims description 16
- 239000012362 glacial acetic acid Substances 0.000 claims description 14
- 238000002360 preparation method Methods 0.000 claims description 14
- 239000001117 sulphuric acid Substances 0.000 claims description 14
- 235000011149 sulphuric acid Nutrition 0.000 claims description 14
- 238000010521 absorption reaction Methods 0.000 claims description 11
- 238000001514 detection method Methods 0.000 claims description 11
- 238000001035 drying Methods 0.000 claims description 11
- 239000003208 petroleum Substances 0.000 claims description 11
- 239000000284 extract Substances 0.000 claims description 10
- 238000000605 extraction Methods 0.000 claims description 9
- 239000012528 membrane Substances 0.000 claims description 9
- 238000009825 accumulation Methods 0.000 claims description 8
- 239000000463 material Substances 0.000 claims description 6
- 239000007788 liquid Substances 0.000 claims description 5
- 239000011259 mixed solution Substances 0.000 claims description 5
- 238000003384 imaging method Methods 0.000 claims description 4
- 238000002156 mixing Methods 0.000 claims description 3
- 239000000843 powder Substances 0.000 claims description 3
- 238000000079 presaturation Methods 0.000 claims description 3
- 239000012488 sample solution Substances 0.000 claims description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 33
- 238000004809 thin layer chromatography Methods 0.000 description 19
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 16
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 13
- 238000004458 analytical method Methods 0.000 description 12
- DLYUQMMRRRQYAE-UHFFFAOYSA-N tetraphosphorus decaoxide Chemical compound O1P(O2)(=O)OP3(=O)OP1(=O)OP2(=O)O3 DLYUQMMRRRQYAE-UHFFFAOYSA-N 0.000 description 10
- 239000003814 drug Substances 0.000 description 7
- 229920001542 oligosaccharide Polymers 0.000 description 6
- 150000002482 oligosaccharides Chemical class 0.000 description 6
- 239000012224 working solution Substances 0.000 description 6
- 238000011049 filling Methods 0.000 description 5
- 238000001291 vacuum drying Methods 0.000 description 5
- 238000009792 diffusion process Methods 0.000 description 4
- 229940079593 drug Drugs 0.000 description 4
- 235000013305 food Nutrition 0.000 description 4
- 239000000047 product Substances 0.000 description 4
- 238000011084 recovery Methods 0.000 description 4
- 239000003153 chemical reaction reagent Substances 0.000 description 3
- 238000010790 dilution Methods 0.000 description 3
- 239000012895 dilution Substances 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 150000002772 monosaccharides Chemical class 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 2
- 229930006000 Sucrose Natural products 0.000 description 2
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 2
- 238000013459 approach Methods 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 230000033228 biological regulation Effects 0.000 description 2
- 210000000988 bone and bone Anatomy 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 239000008103 glucose Substances 0.000 description 2
- 239000000413 hydrolysate Substances 0.000 description 2
- 238000002386 leaching Methods 0.000 description 2
- 238000005259 measurement Methods 0.000 description 2
- 210000003205 muscle Anatomy 0.000 description 2
- 125000004108 n-butyl group Chemical group [H]C([H])([H])C([H])([H])C([H])([H])C([H])([H])* 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 238000013112 stability test Methods 0.000 description 2
- 239000005720 sucrose Substances 0.000 description 2
- 238000010200 validation analysis Methods 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- 208000006820 Arthralgia Diseases 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 208000035126 Facies Diseases 0.000 description 1
- 229930091371 Fructose Natural products 0.000 description 1
- 239000005715 Fructose Substances 0.000 description 1
- RFSUNEUAIZKAJO-ARQDHWQXSA-N Fructose Chemical compound OC[C@H]1O[C@](O)(CO)[C@@H](O)[C@@H]1O RFSUNEUAIZKAJO-ARQDHWQXSA-N 0.000 description 1
- 206010021118 Hypotonia Diseases 0.000 description 1
- 208000019255 Menstrual disease Diseases 0.000 description 1
- 208000002193 Pain Diseases 0.000 description 1
- 208000025747 Rheumatic disease Diseases 0.000 description 1
- 241001107098 Rubiaceae Species 0.000 description 1
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 1
- 210000001015 abdomen Anatomy 0.000 description 1
- 239000000443 aerosol Substances 0.000 description 1
- LKDRXBCSQODPBY-ZXXMMSQZSA-N alpha-D-fructopyranose Chemical compound OC[C@]1(O)OC[C@@H](O)[C@@H](O)[C@@H]1O LKDRXBCSQODPBY-ZXXMMSQZSA-N 0.000 description 1
- PYKYMHQGRFAEBM-UHFFFAOYSA-N anthraquinone Natural products CCC(=O)c1c(O)c2C(=O)C3C(C=CC=C3O)C(=O)c2cc1CC(=O)OC PYKYMHQGRFAEBM-UHFFFAOYSA-N 0.000 description 1
- 150000004056 anthraquinones Chemical class 0.000 description 1
- 230000001430 anti-depressive effect Effects 0.000 description 1
- 230000003064 anti-oxidating effect Effects 0.000 description 1
- 230000002180 anti-stress Effects 0.000 description 1
- 239000000935 antidepressant agent Substances 0.000 description 1
- 229940005513 antidepressants Drugs 0.000 description 1
- 230000006907 apoptotic process Effects 0.000 description 1
- 238000000149 argon plasma sintering Methods 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 150000001720 carbohydrates Chemical class 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 239000003086 colorant Substances 0.000 description 1
- 235000009508 confectionery Nutrition 0.000 description 1
- 230000018044 dehydration Effects 0.000 description 1
- 238000006297 dehydration reaction Methods 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 238000004821 distillation Methods 0.000 description 1
- GUTPQYNJVRSYHY-UHFFFAOYSA-N ethyl acetate;sulfuric acid Chemical compound OS(O)(=O)=O.CCOC(C)=O GUTPQYNJVRSYHY-UHFFFAOYSA-N 0.000 description 1
- 239000012467 final product Substances 0.000 description 1
- 208000017561 flaccidity Diseases 0.000 description 1
- 150000004676 glycans Chemical class 0.000 description 1
- 229930182470 glycoside Natural products 0.000 description 1
- 150000002338 glycosides Chemical class 0.000 description 1
- 230000008676 import Effects 0.000 description 1
- 201000001881 impotence Diseases 0.000 description 1
- 208000021267 infertility disease Diseases 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 210000003734 kidney Anatomy 0.000 description 1
- 238000012417 linear regression Methods 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 238000001819 mass spectrum Methods 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 239000010413 mother solution Substances 0.000 description 1
- 238000005457 optimization Methods 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 238000001121 post-column derivatisation Methods 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 238000010992 reflux Methods 0.000 description 1
- 230000003252 repetitive effect Effects 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 230000000552 rheumatic effect Effects 0.000 description 1
- 238000007790 scraping Methods 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 229930000044 secondary metabolite Natural products 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 239000000741 silica gel Substances 0.000 description 1
- 229910002027 silica gel Inorganic materials 0.000 description 1
- 230000003595 spectral effect Effects 0.000 description 1
- 239000004575 stone Substances 0.000 description 1
- 238000005728 strengthening Methods 0.000 description 1
- 230000001629 suppression Effects 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
- 238000002604 ultrasonography Methods 0.000 description 1
Abstract
The invention provides a method for determining the nystose content of morinda officinalis roots by scanning a thin-layer image through software. According to the method, a trapezoidal integral method is adopted for integration of chromatographic peaks, and quantitative analysis is performed according to the relation between the peak area and the sample concentration. The method is simple to operate and can substitute a traditional thin layer scanner.
Description
Technical field
The present invention relates to field of medicaments, especially relate to a kind of Radix Morindae Officinalis this sugar thin layer quantitative detecting method resistance to.
Background technology
Morinda officinalis How is the dry root of dicotyledon Rubiaceae Radix Morindae Officinalis Morinda officinalis How.Radix Morindae Officinalis medicine is eaten
Homology, Shennong's Herbal is classified as top grade, is also one of Chinese Famous " four great Nan medicines ".Sweet, pungent, the tepor of property,
Returning kidney, Liver Channel, have kidney-replenishing, bone and muscle strengthening, the function of wind-damp dispelling, for impotence and seminal emission, cold womb is infertile, menoxenia,
Few abdomen cold type of pain, rheumatic arthralgia, muscles and bones flaccidity is soft.
Chemical research to Radix Morindae Officinalis shows wherein to contain anthraquinone, iridoid and glycoside thereof, organic acid, saccharide etc. in recent years
Point.The oligosaccharide wherein occupying absolute content advantage is its main active component, and other component content is less.Research report bar halberd
The oligosaccharide separated in it has the pharmacologically actives such as antioxidation, anti-stress, antidepressant, suppression apoptosis.Therefore, oligosaccharide content becomes
The important indicator that quality of medicinal material is good and bad.Owing to oligosaccharide kind composition does not has strong uv absorption, conventional UV-detector to be not suitable for
Measure and analyze.Detection for oligosaccharide kind uses differential refraction, evaporat light scattering, pulsed amperometric, charged aerosol, mass spectrum more
Etc. method, in addition, the method taking post-column derivatization detection fluorescence can also be used for it and measures analysis.Assay method is many at present
Need expensive accurate instrument and the sample handling processes of complexity.
TLC technique with it quickly, low cost, high information quantity, low technical threshold be at modern analysis such as medicine, food, change
Work field still occupy critical role.It is an object of the invention to provide a kind of content assaying method based on thin layer chromatography, new method
Software scans thin layer image is utilized to replace tradition thin-layer chromatogram scanner.
Summary of the invention
The present invention provides one to utilize software scans thin layer determining image Radix Morindae Officinalis this sugar thin layer quantitative detecting method resistance to.The method is grasped
Make simple, tradition thin-layer chromatogram scanner can be acted on behalf of.
The present invention is achieved by the following technical solutions:
A kind of Radix Morindae Officinalis this sugar thin layer quantitative detecting method resistance to, it is characterised in that comprise the steps:
(1) prepared by solution: include Radix Morindae Officinalis need testing solution and Nai Si sugar reference substance solution；
(2) launch and develop the color: by gained need testing solution and reference substance solution ribbon point sample to High Performance Thin precoated plate, with
Acetate-methanol-water-glacial acetic acid is that developing solvent launches and dries, and lamellae is immersed the ethyl acetate stone containing concentrated sulphuric acid
Several seconds in the developer of the mixed solution of oil ether, taking-up is dried, and is placed in thin layer heating plate up to clear spot；
(3) imaging and image procossing: the lamellae developed the color through step 2 is placed under uviol lamp and takes pictures, photo is imported at image
Reason software, sample strip is scanned by software, separates rgb value and adds up line by line, then utilizing trapezoidal integration to check colors
Spectral peak is integrated, and carries out quantitative analysis according to the relation of peak area and sample concentration.
Need testing solution preparation method described in step (1): by Radix Morindae Officinalis fine powder aqueous methanol or water supersound extraction 15-30
Minute, filter, filtrate constant volume and get final product.Described aqueous methanol is 50-80% methanol, preferably 60% methanol.
Preferably Radix Morindae Officinalis medicinal material drying is ground into fine powder, by water or 60% methanol supersound extraction 20 minutes, filters, filtering residue water
Washing, merging filtrate constant volume.
Preferably take Radix Morindae Officinalis medical material use preposition in 60 DEG C of drying baker overnight, pulverize and cross 20 mesh sieves, accurately weighed, be placed in
In conical flask, the ultrasonic 80kHz that adds water extracts 20min, filters, and filtering residue washs with little water, and merging filtrate constant volume shake up,
By 0.45 μm microporous filter membrane, take subsequent filtrate and prepare need testing solution 20mg/ml.
Prepared by described reference substance: take resistance to this sugar reference substance, accurately weighed, be dissolved in water and be diluted to scale, shake up prepare often contain
The reference substance storing solution of 1.586mg/mL this sugar resistance to.The appropriate dilute of accurate absorption prepares series concentration reference substance solution.Institute
The series concentration stated is respectively 1.190,0.893,0.670 and 0.446mg/mL
Developing solvent acetate-methanol-water-glacial acetic acid=8:3:(2-2.5 described in step of the present invention (2)): 3, preferably acetic acid second
Ester-methanol-water-glacial acetic acid=8:3:2.3:3.The ratio of the water R to this sugar resistance in developing solventfValue impact is relatively big, but too low having
Machine Phase Proportion can cause the excess diffusion of band to affect separating degree.
The uniformity coefficient of developer directly influences analysis and the accuracy of quantitative result because the selection of developer and proportioning for
It is very important for thin layer chromatography, in order to ensure the diffusion uniformly and not causing speckle of developer, step of the present invention (2)
In preferred developer: the mixed solution of the ethyl acetate petroleum ether 5-7:5-3 containing 1-2.5% concentrated sulphuric acid, heating colour developing 5-12min.
Further preferred developer is the mixed solution of the ethyl acetate petroleum ether 7:3 of 2% concentrated sulphuric acid, and uses the mode of leaching plate, heating
Colour developing in 10 minutes.Chromatographic peak signal is had an impact by the concentration of the concentrated sulphuric acid in developer, and when concentration is 2%, chromatographic peak signal is relatively strong,
And speckle gelatinizing will not be caused.
Lamellae of the present invention is precoated silica gel plate GF254, the most prefabricated SIL G-25UV254Lamellae.
The described spotting methods of step of the present invention (2), ribbon point sample, strip width: 4mm；Striation widths: 10mm；
Away from base distance: 8mm；Point sample amount: 2 μ L.
Lamellae after step of the present invention (2) preferably point sample, needs to be dried more than 4h in the pre-deployed.
Taking pictures described in step of the present invention (3) refers to take pictures parameter under uviol lamp: time of exposure 2000-6000ms, gain
1.0, accumulation is closed.Preferably expose 5000ms.
Thin layer image described in step of the present invention (3) is directed in PaintShop and each band is carried out digital scanning, separates
Rgb value the integrated value utilizing trapezoidal integration to obtain corresponding chromatographic peak that adds up line by line.
Described PaintShop refers to, preferably TLC Image Processor software, more preferably TLC Image Processor3.0
Version software
Described rgb value refers to constitute the value of the Red Green Blue of image pixel, and three kinds of values are overlapped mutually available various
The color of various kinds.
Described trapezoidal integration refers to: according to certain step-length, chromatographic peak is divided into interval as much as possible, each interval
What area can approximate tries to achieve by trapezoidal area computing formula, and the area of chromatographic peak is equal to each intervening areas sum.The interval divided
Number is the most, and error is the least, and formula is: on interval [a, b],
Beneficial effects of the present invention
1, the analysis of quantitative result is directly influenced due to the uniformity coefficient of colour developing, in order to ensure the uniform of developer and not cause speckle
The diffusion of point, first important additional technical feature of the present invention is with the ethyl acetate petroleum ether of 1-2.5% concentrated sulphuric acid
The mode of the mixed solution leaching plate of 5-7:5-3 develops the color.
2, in order to reduce the cost of detection, not relying on accurate expensive detector, second important supplementary technology of the present invention is special
Levy the mode being to utilize software scans thin layer image and replace traditional thin-layer chromatogram scanner: after image imports software, to choosing band
Carrying out picture element scan line by line, add up respective rgb value and total value respectively, and output comprises the curve of chromatographic peak in window, then
Trapezoidal integration is utilized to be integrated chromatographic peak obtaining peak area.
3, in order to reduce the interference of lamellae background, the 3rd the important additional technical feature of the present invention is to utilize isolated red
Chrominance channel (R) numerical value is integrated calculating, and this is owing to lamellae background is the bluest, uses isolated red value can avoid the back of the body
Scape disturbs, and effectively prevent the drift of baseline.
In order to preferably embody above-mentioned beneficial effect, illustrated by following test example
The present invention establishes a kind of thin layer quantitative approach not relying on thin-layer chromatogram scanner, replaces entity scanner with software, to bar
In halberd sky, this sugar content resistance to is determined, Method validation show this method have good specificity, precision, stability and
Accuracy, can be used for Radix Morindae Officinalis quality of medicinal material control as a kind of quick, quantitative detecting method of low cost, low threshold.
2010 editions Chinese Pharmacopoeia regulations, this sugar content resistance to of Radix Morindae Officinalis medical material must not be less than 2%.In the actual sample collected, only lot number
Be 130501 a collection of medical material against regulation.Finding through analyzing with other medical material profile comparisons, this medical material is thinner, thus it is speculated that former
Because Growing years is shorter, secondary metabolite accumulation is not.
Test example one, methodological study
1. material, reagent and instrument
1.1. standard substance
This sugar (lot number: AWG0714, purity 99.0%) resistance to, Wako Pure Chemical Industries, Ltd. of Japan
Sucrose (lot number 111507-201303:, purity 99.8%), National Institute for Food and Drugs Control
Glucose (lot number 110833-201303:, purity 100%), National Institute for Food and Drugs Control
Fructose (lot number 100231-201305:, purity 99.4%), National Institute for Food and Drugs Control
1.2. reagent
Ethyl acetate, methanol, glacial acetic acid analytical pure are purchased from Guangzhou Chemical Reagent Factory, and water is obtained by triple distillation water system controlling.
1.3. sample collection
Radix Morindae Officinalis medical material is collected from medical material market, Deqing city of Guangdong Province, and kind is reflected through positive pharmaceutcal corporation, Ltd professor Xie Peishan in Guangzhou ten thousand
It is set to Morinda officinalis How.Lot number is respectively 100201,100401,100701,110401,120201,
130501,130801,130901,131101,140401。
1.4. key instrument
BT25S type electronic balance (100,000/), Sai Duolisi group of Germany
The automatic triple pure water distillators of SZ-97A type, Shanghai Yarong Biochemical Instrument Plant
The high-frequency digitally controlled Ultrasound Instrument of KQ-500TDE type, Kunshan Ultrasonic Instruments Co., Ltd.
The automatic sample applicator of ATS-4 thin layer, CAMAG company of Switzerland
TLC lamellae heater III, CAMAG company of Switzerland
TLC scope, CAMAG company of Switzerland
2. test method
2.1. the preparation of working solution
2.1.1. the preparation of series concentration reference substance solution
Take resistance to this sugar reference substance about 15mg, accurately weighed, it is placed in 10mL volumetric flask, is dissolved in water and is diluted to scale,
Shake up the prepared every mL reference substance storing solution containing 1.586mg this sugar resistance to.The appropriate dilute of accurate absorption prepares series concentration pair
According to product solution.
2.1.2. the preparation of need testing solution
Radix Morindae Officinalis medical material use preposition in 60 DEG C of drying baker overnight, pulverize and cross 20 mesh sieves, take about 0.5g, accurately weighed,
Being placed in conical flask, add 20mL water, ultrasonic (80kHz) extracts 20min, filters, and filtering residue washs with little water, merges
Filtrate is also settled to 25mL, shakes up, and by 0.45 μm microporous filter membrane, takes subsequent filtrate and prepares need testing solution.
2.2. method is analyzed
2.2.1. thin layer chromatography condition
Lamellae: prefabricated SIL G-25UV254Lamellae, 10 × 15cm；Strip width: 4mm；Striation widths: 10mm；
Away from base distance: 8mm；Point sample amount: 2 μ L；Presaturation time: 15min；Developing solvent: acetate-methanol-water-ice vinegar
Acid (8:3:2.3:3, v/v/v/v)；Open up away from: exhibition to upper edge；Developer: the ethyl acetate oil containing 2% concentrated sulphuric acid
Ether mixed liquor (7:3, v/v), contaminates formula；Colour developing heat time heating time: 10min；Scope is taken pictures parameter: time of exposure-5000ms,
Gain-1.0, accumulation-close.
2.2.2. image procossing
Pixel be 715 × 1087 BMP thin layer image be directed in self-editing 3.0 editions softwares of TLC Image Processor respectively
Band carries out digital scanning, it is thus achieved that the integrated value of corresponding chromatographic peak.
2.3. methodological study
2.3.1. specificity test
Being launched on lamellae by need testing solution linear spotting, lamellae carries out developing portions, scraping has phase with this sugar resistance to
Same RfThe non-coloured moiety of value, with water dissolution, filters, point sample, with n-butyl alcohol-isopropanol-water-glacial acetic acid (7:5:2:1) is
Developing solvent carries out second outspread, is expanded at 70mm for the first time, and at second time to 100mm, checking speckle is the most single.
Separately take reference substance storing solution appropriate, add sulphuric acid to 0.1M, hydrolyze 4h, detection hydrolyzate whether interference measurement.
2.3.2. the drafting of limit, quantitative limit and standard curve is detected
Taking this sugar reference substance solution appropriateness dilution resistance to, method measures according to the above analysis, and the peak height of scanning signal is equivalent to noise
The detection that solute amount is method limit when 3 times, is the quantitative limit of method when 10 times.
Take the appropriate mother solution stepwise dilution that adds water to prepare series concentration and be respectively 1.190,0.893,0.670 and 0.446mg/mL
Other 4 concentration point.With the concentration of this sugar resistance to as abscissa (x), this sugar spot pixels red channel curvilinear integral area resistance to is
Vertical coordinate (y) is drawn standard curve and investigates correlation coefficient.
2.3.3. precision test
Take the reference substance solution that concentration is 0.446,0.893 and 1.586mg/mL, continuous point sample 6 on same thin laminate
Secondary, precision in investigating plate with integrated peak areas for index.Between plate precision for by three concentrations control product solution respectively o'clock in 5
On block difference lamellae, equally with peak area as inspection target.Relative standard in the plate of precision reference substance solution and between plate
Deviation (RSD) represents, relative standard deviation requires to be not more than 5%.
2.3.4. replica test
Take with 6 parts of a collection of Radix Morindae Officinalis sample, according to need testing solution preparation method operation repetitive, and measure by above-mentioned analysis method
Analyze, calculate content results and RSD value according to retinue standard curve.
2.3.5. stability test
Taking above-mentioned need testing solution, at room temperature (25 DEG C) are transferred and are set to 0, by above-mentioned point after 8,16,24,32,40,48h
Analysis method measures to be analyzed, and calculates content results and RSD value according to retinue standard curve.Single batch sample should be exceeded standing time
The required time to be analyzed such as product.
2.3.6. recovery test it is loaded
Take the Radix Morindae Officinalis medical material 9 parts of known this sugar content resistance to, be equivalent to the 50% of medical material content accurate addition respectively, 100%, 150%
Resistance to this sugar reference substance, each concentration point 3 parts, prepare according to need testing solution preparation method, and press said determination analysis method
The measure and calculation response rate and RSD value.
3. result of the test
3.1. specificity test
Being after developing solvent launches with n-butyl alcohol-isopropanol-water-glacial acetic acid (7:5:2:1), the sample that scraper plate obtains is still for single
The speckle of rule, RfIt is about 0.45.This sugar resistance to hydrolyzes 4h in 0.1M sulfuric acid solution, obtains final hydrolyzate monosaccharide,
Result is shown in accompanying drawing 2, differs 1 glycan molecule to R in molecular structurefThe impact of value is relatively big, such as monosaccharide and the R of disaccharidasefValue difference is near
0.1.Therefore, method uses developing solvent this sugar resistance to can be played preferable separating effect, and its hydrolyzate can't be to mensuration
Producing interference, method specificity is good.
3.2. limit, quantitative limit and standard curve are detected
When signal to noise ratio (Signal to noise ratio, S/N) is 3, the solute amount on band, i.e. detection are limited to 224ng,
When S/N is 10, the solute amount on band, the most quantitatively it is limited to 747ng.Take this sugar reference substance solution resistance to of 5 gradient concentrations,
Method is measured according to the above analysis, with the peak area of digital scanning red channel curve as vertical coordinate (y), right with this sugar resistance to
Concentration (mg/mL) according to product solution is abscissa (x), and the standard curve equation obtained by linear regression is: y=16455.6
* x 1095, R2=0.9945.This sugar resistance to is in the range of 0.446-1.586mg/mL concentration, and linear relationship is good.
3.3. precision test
In plate, between precision and plate, the result of precision is as shown in table 1, the RSD of this sugar reference substance solution resistance to of low middle high three concentration
All within the scope of 5%, method precision is good.
Table 1. this sugar Precision test result resistance to
3.4. replica test
Take 6 parts of the Radix Morindae Officinalis sample that lot number is 100201, according to the parallel sample solution of preparing of need testing solution preparation method, and depend on
Above-mentioned analytical measures the content of this sugar resistance to, and the RSD of 6 results is 2.98%, and method repeatability is good.
3.5. stability test
Take the Radix Morindae Officinalis sample that lot number is 100201, prepare sample solution according to need testing solution preparation method, 0-48 after preparation
Measuring once every 8h between h, the RSD of 7 measurement results is 2.66%, and sample is good at 48 hours internal stabilities.
3.6. recovery test it is loaded
Take the Radix Morindae Officinalis medical material 0.5g that lot number is 130501, be separately added into and be equivalent to the 50% of medical material this sugar content resistance to, 100% He
This sugar reference substance resistance to of 150% amount, prepares solution according to the method described above and analyzes mensuration, low middle high three sample-adding hydraulic test gained
Average recovery is all between 95%-105%, and as shown in table 2, RSD is 2.77%, and the accuracy of method is good.
Table 2. average recovery is tested
The most multiple batches of sample determination analysis
Being prepared according to need testing solution preparation method by 10 batches of Radix Morindae Officinalis medical materials, method measures and analyzes according to the above analysis, this sugar resistance to
Content results is shown in Table 3, and digital scanning figure is as shown in Figure 4.
Table 3. multiple batches of medical material quantitative analysis results
The present invention establishes a kind of thin layer quantitative approach not relying on thin-layer chromatogram scanner, replaces entity scanner with software, to bar
In halberd sky, this sugar content resistance to is determined, Method validation show this method have good specificity, precision, stability and
Accuracy, can be used for Radix Morindae Officinalis quality of medicinal material control as a kind of quick, quantitative detecting method of low cost, low threshold.
Test example two, experimental condition preferred
The present invention takes single_factor method that experimental condition includes the preparation of sample with the scanning peak area of this sugar resistance to for optimizing index, aobvious
Toner concentration, picture collection parameter all made screening and optimize, data are shown in Table 4.Experiment compares the extraction of 60% ethanol and pure water
Effect, the extraction efficiency of result display pure water is higher.Extracting mode has investigated supersound extraction and hot reflux, and supersound extraction is convenient fast
Prompt and extraction ratio will not be reduced.Extraction time experiment display, extracts 20 minutes content peakings, and extracting solution for the second time is resistance to
This sugar content is less than the detection limit of method.The ratio of the water R to this sugar resistance in developing solventfValue impact is relatively big, but too low
Organic facies causes the excess diffusion two of band to affect separating degree than regular meeting.Sulfuric acid concentration in developer is investigated by experiment, dense
When degree is 2%, chromatographic peak signal is relatively strong, and will not cause speckle gelatinizing.When the time of exposure of image acquisition is 5000ms, chromatograph
Peak-to-peak signal is relatively strong, and peak shape is preferable.
Table 4 experimental condition optimization result
Accompanying drawing explanation
Fig. 1 is the principle of the invention and overhaul flow chart: oligosaccharide is hydrolyzed to monosaccharide, and under effect of sulfuric acid, dehydration forms conjugated system and obtains
Obtain uv absorption, utilize the fluorescence intensity produced after exciting to be measured analyzing.
Fig. 2 is this sugar hydrolysate identification result resistance to: 1. glucose, 2. fructose, 3. sucrose, this sugar the most resistance to, S. Radix Morindae Officinalis pair
According to medical material, this sugar hydrolysate the most resistance to.
Fig. 3 is software separation pixel RGB code: first half section code is for the image of analysis of fluorescence, and second half section code is used for analyzing visible
The image gathered under light.
Fig. 4 is software scans figure based on thin layer image: this sugar resistance to all can reach baseline separation, R with chromatographic peak front and backfIt is 0.35.
Detailed description of the invention
Further illustrate the present invention by the following examples, but scope of the presently claimed invention is not limited.
Embodiment 1
(1) prepared by reference substance solution: take resistance to this sugar reference substance about 15mg, accurately weighed, is placed in 10mL volumetric flask, adds 60%
Ethanol dissolves and is diluted to scale, shakes up the prepared every mL reference substance storing solution containing 1.586mg this sugar resistance to.Accurate absorption is appropriate
Add 60% ethanol dilution and prepare other concentration point reference substance solution.
Prepared by need testing solution: take Radix Morindae Officinalis medical material (lot number: 100201) and be placed in 60 DEG C of drying baker overnight, pulverize and mistake
20 mesh sieves, take about 0.5g, accurately weighed, are placed in conical flask, add 20mL 60% ethanol, and ultrasonic (80kHz) extracts 20
Min, filters, and filtering residue is also settled to 25mL with a little 60% washing with alcohol, merging filtrate, shakes up, by 0.45 μm micropore
Filter membrane, takes subsequent filtrate and prepares need testing solution.
(2) by 2 μ L working solution ribbon point samples in prefabricated SIL G-25UV254On lamellae (10 × 15cm), base away from 8mm,
Strip width 4mm, striation widths 10mm.Put into dry more than the 4h of vacuum drying apparatus filling phosphorus pentoxide, take out,
It is placed in double flute expansion cylinder, adds developing solvent (ethyl acetate: methanol: water: glacial acetic acid=8:3:2.3:3, v/v/v/v) forecast and 15
Min, is expanded to plate top edge, takes out, and cold air dries up, and plate immerses the ethyl acetate petroleum ether mixing containing 2% concentrated sulphuric acid
In liquid (7:3, v/v) about 3 seconds, taking-up was dried, and was placed in 105 DEG C of heating 10min in thin layer heating plate.
(3) lamellae is placed in TLC scope, collection image under 366nm uviol lamp, time of exposure 5000ms,
Gain: 1.0, accumulation: close.The BMP thin layer image that the pixel of collection is 715 × 1087 is directed into self-editing TLC Image
3.0 editions softwares of Processor carry out digital scanning to each band, it is thus achieved that the integrated value of corresponding chromatographic peak.
Linear relationship according to peak area with this sugar concentration resistance to carries out quantitative analysis, records in the Radix Morindae Officinalis medical material that lot number is 100201
The content of this sugar resistance to is 4.09%.
Embodiment 2
(1) prepared by reference substance solution: take resistance to this sugar reference substance about 15mg, accurately weighed, is placed in 10mL volumetric flask, adds water-soluble
Solve and be diluted to scale, shaking up the prepared every mL reference substance storing solution containing 1.586mg this sugar resistance to.Accurate absorption adds water dilute in right amount
Release other concentration point reference substance solution prepared.
Prepared by need testing solution: take Radix Morindae Officinalis medical material (lot number: 100201) and be placed in 60 DEG C of drying baker overnight, pulverize and mistake
20 mesh sieves, take about 0.5g, accurately weighed, are placed in conical flask, add 20mL water, and ultrasonic (80kHz) extracts 20min,
Filtering, filtering residue washs with little water, and merging filtrate is also settled to 25mL, shakes up, and by 0.45 μm microporous filter membrane, takes continuous
Filtrate prepares need testing solution.
(2) by 2 μ L working solution ribbon point samples in prefabricated SIL G-25UV254On lamellae (10 × 15cm), base away from 8mm,
Strip width 4mm, striation widths 10mm.Put into dry more than the 4h of vacuum drying apparatus filling phosphorus pentoxide, take out,
It is placed in double flute expansion cylinder, adds developing solvent (ethyl acetate: methanol: water: glacial acetic acid=8:3:2:3, v/v/v/v) forecast and 15min,
Be expanded to plate top edge, take out, cold air dries up, by plate immerse containing 1% concentrated sulphuric acid ethyl acetate petroleum ether mixed liquor (5:5,
V/v) in about 3 seconds, taking-up was dried, and was placed in 105 DEG C of heating 5min in thin layer heating plate.
(3) lamellae is placed in TLC scope, collection image under 366nm uviol lamp, time of exposure 2000ms,
Gain: 1.0, accumulation: close.The BMP thin layer image that the pixel of collection is 715 × 1087 is directed into self-editing TLC Image
3.0 editions softwares of Processor carry out digital scanning to each band, it is thus achieved that the integrated value of corresponding chromatographic peak.
Linear relationship according to peak area with this sugar concentration resistance to carries out quantitative analysis, records in the Radix Morindae Officinalis medical material that lot number is 100201
The content of this sugar resistance to is 4.17%.
Embodiment 3
(1) prepared by reference substance solution: take resistance to this sugar reference substance about 15mg, accurately weighed, is placed in 10mL volumetric flask, adds water-soluble
Solve and be diluted to scale, shaking up the prepared every mL reference substance storing solution containing 1.586mg this sugar resistance to.Accurate absorption adds water dilute in right amount
Release other concentration point reference substance solution prepared.
Prepared by need testing solution: take Radix Morindae Officinalis medical material (lot number: 100201) and be placed in 60 DEG C of drying baker overnight, pulverize and mistake
20 mesh sieves, take about 0.5g, accurately weighed, are placed in conical flask, add 20mL water, and ultrasonic (80kHz) extracts 20min,
Filtering, filtering residue washs with little water, and merging filtrate is also settled to 25mL, shakes up, and by 0.45 μm microporous filter membrane, takes continuous
Filtrate prepares need testing solution.
(2) by 2 μ L working solution ribbon point samples in prefabricated SIL G-25UV254On lamellae (10 × 15cm), base away from 8mm,
Strip width 4mm, striation widths 10mm.Put into dry more than the 4h of vacuum drying apparatus filling phosphorus pentoxide, take out,
It is placed in double flute expansion cylinder, adds developing solvent (ethyl acetate: methanol: water: glacial acetic acid=8:3:2.5:3, v/v/v/v) forecast and 15
Min, is expanded to plate top edge, takes out, and cold air dries up, and is mixed by the ethyl acetate petroleum ether that plate immerses containing 2.5% concentrated sulphuric acid
Closing in liquid (6:4, v/v) about 3 seconds, taking-up is dried, and is placed in 105 DEG C of heating 8min in thin layer heating plate.
(3) lamellae is placed in TLC scope, collection image under 366nm uviol lamp, time of exposure 6000ms,
Gain: 1.0, accumulation: close.The BMP thin layer image that the pixel of collection is 715 × 1087 is directed into self-editing TLC Image
3.0 editions softwares of Processor carry out digital scanning to each band, it is thus achieved that the integrated value of corresponding chromatographic peak.
Linear relationship according to peak area with this sugar concentration resistance to carries out quantitative analysis, records in the Radix Morindae Officinalis medical material that lot number is 100201
The content of this sugar resistance to is 4.23%.
Embodiment 4
(1) prepared by reference substance solution: take resistance to this sugar reference substance about 15mg, accurately weighed, is placed in 10mL volumetric flask, adds water-soluble
Solve and be diluted to scale, shaking up the prepared every mL reference substance storing solution containing 1.586mg this sugar resistance to.Accurate absorption adds water dilute in right amount
Release other concentration point reference substance solution prepared.
Prepared by need testing solution: take Radix Morindae Officinalis medical material (lot number: 100201) and be placed in 60 DEG C of drying baker overnight, pulverize and mistake
20 mesh sieves, take about 0.5g, accurately weighed, are placed in conical flask, add 20mL water, and ultrasonic (80kHz) extracts 20min,
Filtering, filtering residue washs with little water, and merging filtrate is also settled to 25mL, shakes up, and by 0.45 μm microporous filter membrane, takes continuous
Filtrate prepares need testing solution.
(2) by 2 μ L working solution ribbon point samples in prefabricated SIL G-25UV254On lamellae (10 × 15cm), base away from 8mm,
Strip width 4mm, striation widths 10mm.Put into dry more than the 4h of vacuum drying apparatus filling phosphorus pentoxide, take out,
It is placed in double flute expansion cylinder, adds developing solvent (ethyl acetate: methanol: water: glacial acetic acid=8:3:2.5:3, v/v/v/v) forecast and 15
Min, is expanded to plate top edge, takes out, and cold air dries up, and is mixed by the ethyl acetate petroleum ether that plate immerses containing 2.5% concentrated sulphuric acid
Closing in liquid (6:4, v/v) about 3 seconds, taking-up is dried, and is placed in 105 DEG C of heating 8min in thin layer heating plate.
(3) lamellae is placed in TLC scope, collection image under 366nm uviol lamp, time of exposure 6000ms,
Gain: 1.0, accumulation: close.The BMP thin layer image that the pixel of collection is 715 × 1087 is directed into self-editing TLC Image
3.0 editions softwares of Processor carry out digital scanning to each band, it is thus achieved that the integrated value of corresponding chromatographic peak.
Linear relationship according to peak area with this sugar concentration resistance to carries out quantitative analysis, records in the Radix Morindae Officinalis medical material that lot number is 100201
The content of this sugar resistance to is 4.27%.
Embodiment 5
(1) prepared by reference substance solution: take resistance to this sugar reference substance about 15mg, accurately weighed, is placed in 10mL volumetric flask, adds water-soluble
Solve and be diluted to scale, shaking up the prepared every mL reference substance storing solution containing 1.586mg this sugar resistance to.Accurate absorption adds water dilute in right amount
Release other concentration point reference substance solution prepared.
Prepared by need testing solution: take Radix Morindae Officinalis medical material (lot number: 100201) and be placed in 60 DEG C of drying baker overnight, pulverize and mistake
20 mesh sieves, take about 0.5g, accurately weighed, are placed in conical flask, add 20mL water, and ultrasonic (80kHz) extracts 20min,
Filtering, filtering residue washs with little water, and merging filtrate is also settled to 25mL, shakes up, and by 0.45 μm microporous filter membrane, takes continuous
Filtrate prepares need testing solution.
(2) by 2 μ L working solution ribbon point samples in prefabricated SIL G-25UV254On lamellae (10 × 20cm), base away from 8mm,
Strip width 4mm, striation widths 10mm.Put into dry more than the 4h of vacuum drying apparatus filling phosphorus pentoxide, take out,
It is placed in double flute expansion cylinder, adds developing solvent (ethyl acetate: methanol: water: glacial acetic acid=8:3:2.5:3, v/v/v/v) forecast and 15
Min, is expanded to plate top edge, takes out, and cold air dries up, and plate immerses the ethyl acetate petroleum ether mixing containing 2% concentrated sulphuric acid
In liquid (7:3, v/v) about 3 seconds, taking-up was dried, and was placed in 105 DEG C of heating 12min in thin layer heating plate.
(3) lamellae is placed in TLC scope, collection image under 366nm uviol lamp, time of exposure 4000ms,
Gain: 1.0, accumulation: close.The BMP thin layer image that the pixel of collection is 715 × 1087 is directed into self-editing TLC Image
3.0 editions softwares of Processor carry out digital scanning to each band, it is thus achieved that the integrated value of corresponding chromatographic peak.
Linear relationship according to peak area with this sugar concentration resistance to carries out quantitative analysis, records in the Radix Morindae Officinalis medical material that lot number is 100201
The content of this sugar resistance to is 4.33%.
Claims (5)
1. Radix Morindae Officinalis this sugar thin layer quantitative detecting method resistance to, it is characterised in that comprise the steps:
(1) prepared by solution: include Radix Morindae Officinalis need testing solution and Nai Si sugar reference substance solution；
Described need testing solution preparation method: Radix Morindae Officinalis medicinal material drying is ground into fine powder, by water supersound extraction 15-30 minute,
Filtering, filtering residue washes with water, merging filtrate constant volume；
(2) launch and develop the color: by gained need testing solution and reference substance solution ribbon point sample to thin layer precoated plate, with ethyl acetate
-methanol-water-glacial acetic acid=8:3:2.3:3 is that developing solvent launches and dries, and immerses lamellae containing 1-2.5% concentrated sulphuric acid
Several seconds in the developer of the mixed solution of ethyl acetate petroleum ether 5-7:5-3, taking-up is dried, and is placed in thin layer heating plate colour developing and adds
Hot 5-12min；
(3) imaging and image procossing: the lamellae developed the color through step 2 is placed under uviol lamp and takes pictures, photo is imported image procossing soft
Part, sample strip is scanned by software, separates rgb value and adds up line by line, then utilizing trapezoidal integration to chromatographic peak
It is integrated, carries out quantitative analysis according to the relation of peak area and sample concentration.
2. detection method as claimed in claim 1, it is characterised in that parameter of taking pictures under the uviol lamp described in step (3): expose
Light time 2000-6000ms, gain 1.0, accumulation is closed.
3. detection method as claimed in claim 1, it is characterised in that the thin layer image described in step (3) is directed into TLC Image
Each band is carried out digital scanning by 3.0 editions graphics softwares of Processor, separates rgb value and the acquisition that adds up line by line is corresponding
The integrated value of chromatographic peak.
4. detection method as claimed in claim 1, it is characterised in that comprise the steps:
(1) prepared by solution:
Reference substance solution is prepared: take this sugar reference substance resistance to, accurately weighed, is dissolved in water and is diluted to scale, shaking up and prepare often containing 1.586
The reference substance storing solution of mg/mL this sugar resistance to, the appropriate dilute of accurate absorption prepares series concentration reference substance solution；
The preparation of need testing solution: Radix Morindae Officinalis medical material use preposition in 60 DEG C of drying baker overnight, pulverize and cross 20 mesh sieves, accurate claim
Fixed, it is placed in conical flask, the ultrasonic 80kHz that adds water extracts 20min, filters, and filtering residue washs with little water, and merging filtrate is the most fixed
Hold, shake up, by 0.45 μm microporous filter membrane, take subsequent filtrate and prepare need testing solution 20mg/ml；
(2) launch and develop the color:
Lamellae: prefabricated U V254Lamellae, presaturation time: 15min；Developing solvent: acetate-methanol-water-glacial acetic acid
8:3:2.3:3,v/v/v/v；Open up away from: exhibition to upper edge；Developer: containing the ethyl acetate petroleum ether of 1-2.5% concentrated sulphuric acid
Mixed liquor 7:3, v/v, contaminate formula；Colour developing heat time heating time: 5-12min；
(3) imaging and image procossing
By lamellae as in TCL scope, under 366nm uviol lamp, gather image, time of exposure 5000ms, gain 1.0, tire out
Long-pending-to close, the BMP thin layer image that the pixel of collection is 715 × 1087 is directed into TLC Image Processor 3.0 domain
In shape software, each band being carried out digital scanning, add up respective rgb value and total value respectively, and output comprises the curve of chromatographic peak extremely
In window, trapezoidal integration is then utilized to be integrated chromatographic peak obtaining peak area.
5. detection method as claimed in claim 4, it is characterised in that comprise the steps:
(1) prepared by solution:
Reference substance solution is prepared: take this sugar reference substance resistance to, accurately weighed, is dissolved in water and is diluted to scale, shaking up and prepare often containing 1.586
The reference substance storing solution of mg/mL this sugar resistance to, the appropriate dilute of accurate absorption prepares series concentration reference substance solution, and concentration is respectively
It is 1.190,0.893,0.670 and 0.446mg/mL；
The preparation of need testing solution: Radix Morindae Officinalis medical material use preposition in 60 DEG C of drying baker overnight, pulverize and cross 20 mesh sieves, take about
0.5g, accurately weighed, it is placed in conical flask, adds 20mL water, ultrasonic 80kHz extracts 20min, filters, and filtering residue is with few
Xu Shui washs, and merging filtrate is also settled to 25mL, shakes up, and by 0.45 μm microporous filter membrane, takes subsequent filtrate and prepares test sample
Solution；
(2) launch and develop the color:
Lamellae: prefabricated U V254Lamellae,；Presaturation time: 15min；Developing solvent: acetate-methanol-water-glacial acetic acid
8:3:2.3:3,v/v/v/v；Open up away from: exhibition to upper edge；Developer: the ethyl acetate petroleum ether mixing containing 2% concentrated sulphuric acid
Liquid 7:3, v/v, contaminate formula；Colour developing heat time heating time: 10min；
(3) imaging and image procossing
By lamellae as in TCL scope, under 366nm uviol lamp, gather image, time of exposure 5000ms, gain 1.0, tire out
Long-pending-to close, the BMP thin layer image that the pixel of collection is 715 × 1087 is directed into TLC Image Processor 3.0 domain
In shape software, each band being carried out digital scanning, add up respective rgb value and total value respectively, and output comprises the curve of chromatographic peak extremely
In window, trapezoidal integration is then utilized to be integrated chromatographic peak obtaining peak area.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510163742.0A CN104792915B (en) | 2015-04-08 | 2015-04-08 | Method for determining nystose content of morinda officinalis roots |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510163742.0A CN104792915B (en) | 2015-04-08 | 2015-04-08 | Method for determining nystose content of morinda officinalis roots |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN104792915A CN104792915A (en) | 2015-07-22 |
| CN104792915B true CN104792915B (en) | 2017-01-11 |
Family
ID=53557894
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201510163742.0A Active CN104792915B (en) | 2015-04-08 | 2015-04-08 | Method for determining nystose content of morinda officinalis roots |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN104792915B (en) |
Families Citing this family (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108776121B (en) * | 2018-04-08 | 2021-07-30 | 广州卡马生物科技有限公司 | High-throughput screening method for antioxidant in medicinal materials based on fluorescence quenching principle |
| CN109959750A (en) * | 2019-04-09 | 2019-07-02 | 浙江大学 | Fat-soluble natural products thin layer quantitative image recognition detection method |
| CN110824095A (en) * | 2019-11-29 | 2020-02-21 | 北京康仁堂药业有限公司 | Method for identifying morinda officinalis formula particles and prepared morinda officinalis formula particles |
Family Cites Families (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100347541C (en) * | 2005-11-11 | 2007-11-07 | 清华大学 | Thin layer chromatography quantitative analysis method based on image processing technology |
| US8476075B2 (en) * | 2009-04-06 | 2013-07-02 | Board Of Regents, The University Of Texas System | Analytical technique for measuring bound glycerides in a biodiesel composition |
| CN101900715A (en) * | 2009-05-25 | 2010-12-01 | 姚志湘 | Thin layer chromatogram scanner based on postpositive light splitting |
| CN202171586U (en) * | 2011-06-15 | 2012-03-21 | 李传润 | Integrated type thin-layer chromatography analyzer |
| CN103983733A (en) * | 2014-05-21 | 2014-08-13 | 江南大学 | Method for detecting biogenic amine in yellow wine by using chromatographic sheet |
-
2015
- 2015-04-08 CN CN201510163742.0A patent/CN104792915B/en active Active
Also Published As
| Publication number | Publication date |
|---|---|
| CN104792915A (en) | 2015-07-22 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN107843677A (en) | Radix paeoniae rubrathe reference extract and its preparation method and application | |
| CN104487823B (en) | Method for determining a concentration of a polysorbate species in a mixture | |
| CN104007221B (en) | The detection method for the treatment of functional uterine bleeding Chinese medicine composition | |
| CN105259295A (en) | Quality detection method for ginseng, cassia twig and poria cocos oral solution | |
| CN104792915B (en) | Method for determining nystose content of morinda officinalis roots | |
| CN113325125A (en) | Method for thin-layer identification and total flavone content determination of ancient classic famous prescription compound preparation Juanbi granules | |
| CN109632978A (en) | A kind of Poria cocos reference extract and its preparation method and application | |
| CN103940972B (en) | A kind of detection method for the treatment of hypertensive Chinese medicine preparation | |
| CN106501393B (en) | A method of detection hickory chick Nucleosides | |
| CN101264226B (en) | Quality detecting method of diabetes treating medicine | |
| CN106770719A (en) | The fingerprint atlas detection method of low-sugar type intensified loquet distillate | |
| CN112730674B (en) | Quality detection method of momordica grosvenori tea | |
| CN102266478A (en) | Method for controlling quality of drug combination for liver benefiting and eyesight improving, and liver and kidney nourishing | |
| CN111007190B (en) | Method for constructing UPLC (ultra performance liquid chromatography) characteristic spectrum of rhizoma bolbostemmae medicinal material and method for measuring component content of rhizoma bolbostemmae medicinal material | |
| CN107655849A (en) | A kind of herbal tea near infrared online detection method | |
| CN107271396A (en) | The quick determination method of general flavone content in a kind of tealeaves | |
| CN106770764A (en) | A kind of content assaying method of Xue bijing intermediate multi-target ingredient | |
| CN105628853A (en) | Quality detection method for medicinal herb herba cymbopogonis | |
| CN114216980B (en) | Method for establishing HPLC-ELSD fingerprint of starwort root | |
| CN108037234B (en) | Quality detection method of abrus herb hepatitis granules | |
| CN115575541A (en) | Method for simultaneously determining multiple active ingredients in Yinhuang Erchen mixture | |
| CN103675192A (en) | Detection method of inflammation-diminishing compound pierasma quassioides benn capsule | |
| CN109557233B (en) | Method for determining content of multi-index components in white paeony root extracting solution | |
| CN110308213B (en) | Method for constructing fingerprint spectrum of kidney-nourishing and fetus-growing pill and application of fingerprint spectrum in quality detection | |
| CN102475728B (en) | Detection method of Pianzaihuang |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| EXSB | Decision made by sipo to initiate substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20231203 Address after: No. 2 Fangcaodian Road, Guangzhou Science City, Guangzhou High tech Industrial Development Zone, Guangzhou, Guangdong Province, 510663 Patentee after: GUANGZHOU WANZHENG PHARMACEUTICAL CO.,LTD. Address before: 510663 Guangzhou High tech Industrial Development Zone, Guangdong Province Science City, No. 2 Fangcaodian Road, Self Compiled 208 Patentee before: GUANGZHOU HAOYIWAN PHARMACEUTICAL TECHNOLOGY Co.,Ltd. |
|
| TR01 | Transfer of patent right |