CN1544935A - Multiple wavelength Chinese traditional medicine chromatogram fingerprint, fabricating method and application thereof - Google Patents
Multiple wavelength Chinese traditional medicine chromatogram fingerprint, fabricating method and application thereof Download PDFInfo
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Abstract
The invention relates to a multi-wavelength Chinese medicinal chromatographic fingerprint pattern and its constructing method and application, by investigation and optimization of chromatographic conditions and UV response, determining the gradient curve and many UV detected wavelengths within the eluting time of 1-1.5 hours to measure the fingerprint pattern and make chemical verification; adopting relative response valve and quantifiable peak as fixed-point indexes to make relativized processing and matrixing, so as to establish quantized multi-wavelength fingerprint pattern shared-mode data matrix; making similarity identification and component difference comparison on the unknown sample by the matrix, so as to implement quality identification and evaluation, qualitatively and quantitatively. Its advantages: it can not only grasp varieties and quality conditions of Chinese medicines by the whole chromatographic characteristic, but also better characterize the peculiarity of Chinese medicinal components by multi-wavelength selection and fingerprint peak quantitative identification, and strengthens the expression ability and standard degree of the fingerprint pattern for varieties and quality of Chinese medicines by quantizing main components. The established fingerprint pattern can be widely applied to Chinese medicinal quality detection and evaluating technique fields.
Description
Technical field
The present invention relates to a kind of multi-wavelength chromatographic fingerprints of Chinese materia medica and construction method and application.
Background technology
Traditional Chinese medicine fingerprint is a kind of effective means of the characteristic of Chinese medicine analytic sample being expressed by chromatogram or spectrum means.In the directive document of related drugs research, traditional Chinese medicine is done corresponding requirement, and be used for the discriminating of traditional Chinese medicine.In recent years, putting into practice in the evolution of traditional Chinese medicine quality control and evaluation, because the raising of quality requirements and the development of relevant new drug research, more and more require the Chinese medicine sample is carried out multianalysis and whole the expression, but, because a large amount of not existence of principal component in the Chinese medicine sample, illustrate pointedly and comprise composition in the Chinese medicine sample and relative content has bigger difficulty, therefore in modern study, the modes of a large amount of utilization finger-prints are carried out sample characteristics of for example and are expressed, simultaneously at Chinese crude drug, medicine materical crude slice, also utilized chromatogram or spectrum means to carry out the research of finger-print in the analysis of Chinese medical extract and Chinese medicine preparation widely.
In the research of numerous chromatographic fingerprintings, mainly be that utilization ultraviolet detection, evaporation light detect and the Mass Spectrometer Method mode, wherein in view of practical purpose, ultraviolet detection is the main detection means of determining fingerprint pattern, but comparatively speaking, there is following problem in general ultraviolet detection mode: 1, detect the uniqueness of wavelength, make the material information of a large amount of other non-characteristic absorption is not expressed preferably; 2, collection of illustrative plates is not controlled the relative quantity of composition wherein under the situation of overall similarity comparison; 3, in the process that fingerprint peaks is investigated, only take the relative retention value mode to differentiate, do not set up the fingerprint peaks data sequence that contains characteristic information, be used for the specificity evaluation; 4, in the finger-print evaluation method, result's qualitative and quantitative information is not used in combination, and the judged result specific aim is not strong.
Summary of the invention
At the problems referred to above, the purpose of this invention is to provide a kind of adopt multi-wavelength chromatographic fingerprints of Chinese materia medica and construction method and application qualitative and that quantitative target combines.
For achieving the above object, the present invention takes following technical scheme: a kind of construction method of multi-wavelength chromatographic fingerprints of Chinese materia medica, and it may further comprise the steps:
(1) the Chinese medicine sample is prepared, in chromatogram preliminary election moving phase system, carry out rough segmentation, wherein stronger UV response is discerned and assert, result according to identification and identification, determine a plurality of detection wavelength and the tracking response of sample, measure capacity factor measure and corresponding moving phase proportionate relationship under the multistage wash-out ratio;
(2) concern according to aforementioned proportion, determine in the gradient elution method of setting under the moving phase system, carrying out actual sample measures, detect chromatography eluant behavior under the wavelength according to separation case and each, carry out global optimization, adjust the gradient elution curve in real time, finally identification situation and the capacity factor measure according to tracking response distributes, determine the gradient curve under 1 hour～1.5 hours elution times, thereby determine the finger print measuring method of this sample;
(3) above-mentioned definite assay method is carried out efficiency evaluation, comprising specificity, accuracy, precision, repeatability, stability, measurement range and durability;
(4) actual sample that is no less than 10 batches is measured, measurement result comprises collection of illustrative plates and the data under a plurality of detection wavelength, described data are vertically launched with retention time, gained relative peak area value under each wavelength or peak height value are carried out the consistance processing, set up the relative peak area value of corresponding relative retention time of working sample or the finger-print data sequence of peak height value, corresponding sequence is averaged or gets median, form finger-print common pattern sequence under a plurality of single wavelength, carry out the data integration under a plurality of wavelength then, detecting the relative retention time that can respond the fingerprint peaks correspondence in wavelength coverages entirely with all is row, each detects wavelength data carry out sequence matrixization for row, corresponding time point does not have the position of response fills with 0, forms multi-wavelength finger-print common pattern data matrix;
(5) identical Chinese medicine sample is taked multiple preparation method preparation or taked multiple detection method that same sample is measured, and respectively repeat steps (4), the a plurality of data matrixes that obtain are carried out diversity ratio, on this basis the data of step (4) gained are carried out stratification;
(6) it is qualitative the main chromatographic peak in the finger-print to be carried out finger-print;
(7) it is quantitative the main chromatographic peak in the finger-print to be carried out finger-print.
In the above-mentioned steps (6), finger-print is qualitative to be comprised three and infers levels, promptly 1. adopts the reference substance contrast qualitative; 2. adopting the known structure tester is reference, utilize the ultra-violet absorption spectrum feature of the middle quasi-molecular ion of liquid chromatography/mass spectrometry coupling (LC/MS) or molion and corresponding fragment cracking rule and liquid chromatography/diode array detector (LC/DAD) to infer that fingerprint peaks contains the structure type of composition, is used for the supposition to the fingerprint peaks composition; 3. adopt the material composition of LC/MS and LC/DAD information and retention sign fingerprint peaks representative, content comprises that the retention of fingerprint peaks, real-time ultra-violet absorption spectrum and possible molecular weight and ms fragment information are used to identify this fingerprint peaks.
In the above-mentioned steps (7), finger-print quantitatively comprises three levels, 1. adopts reference substance to carry out accurately quantitatively; 2. be reference with accurately quantitative fingerprint peaks, the fingerprint peaks that supposition is had the same structure type compares with it, adopts relative response to carry out quantitatively; 3. adopt the mode of percentage response, to carrying out quantified controlling with the message identification fingerprint peaks.
In the above-mentioned steps (5), stratification is meant according to response magnitude or according to material composition differentiates situation, carry out data qualification, the aggregation of data that similar difference is changed is a class, identical category fingerprint peaks data are compared, choose wherein quantitatively composition or substantially qualitatively the stable elements peak be reference, other data in the class are carried out relativization to be handled, carry out the quantification treatment result of finger print data as relative content or relative characteristic ratio, the result inserts in the data matrix with the different levels quantification treatment, promptly forms to quantize finger-print common pattern data matrix.
In the above-mentioned steps (5), multiple preparation method comprises that cold-maceration, temperature are soaked method, percolation, decocting method, circumfluence method, pressurization leaching, leaching, ultrasonic extraction, Microwave Extraction method, super critical extraction reduce pressure.
In the above-mentioned steps (4), collection of illustrative plates under described a plurality of detection wavelength and data are to detect to measure simultaneously under the wavelength to obtain or measure respectively under the same terms different wave length obtaining at each.
The multi-wavelength chromatographic fingerprints of Chinese materia medica that construction method by above-mentioned multi-wavelength chromatographic fingerprints of Chinese materia medica obtains.
The application of above-mentioned multi-wavelength chromatographic fingerprints of Chinese materia medica in quality discrimination.
The application of above-mentioned multi-wavelength chromatographic fingerprints of Chinese materia medica in quality assessment is analyzed.
The present invention is owing to take above design, it has the following advantages: 1, because the present invention adopts the multi-wavelength detection mode, the figure that records under a plurality of wavelength is made up arrangement, made up the finger-print of comparatively complete reflection sample feature under each wavelength, therefore can characterize various traditional Chinese medicine ingredients specificity informations more fully, and in discriminating of Chinese medicine sample quality and evaluation analysis, easily the Pre-testing article are analyzed comparison qualitatively, and draw and judge conclusion comparatively accurately.2, the present invention is owing to carried out consistance and layering processing with the fingerprint peaks data that record under a plurality of wavelength, and be integrated into the data matrix of common pattern, therefore the principal ingredient in the traditional Chinese medicine ingredients can be quantized, strengthened ability to express and the standardized degree of finger-print to herbal species and quality thereof, make in discriminating of Chinese medicine sample quality and evaluation analysis, not only can obtain being verified article analysis result qualitatively, but also can obtain pointed quantitative judgement easily.3, the present invention is owing to provide the construction method of the multi-wavelength chromatographic fingerprinting that the qualitative and quantitative of a whole set of standard combines, therefore when the chromatographic fingerprints of Chinese materia medica that uses this method to obtain carries out the traditional Chinese medicine ingredients evaluation, analyzes and estimates, not only method is easy, reliable and stable, and is easy to grasp and use.The present invention can be widely used in the complex system that Chinese crude drug, medicine materical crude slice, extract, Chinese medicine preparation and health products, food etc. contain the multiclass chemical constitution, carry out qualitative and quantitative analysis by making up the multi-wavelength chromatographic fingerprinting, reach the purpose of discriminating, quality assessment and quality control.
Description of drawings
Fig. 1 is a cape jasmine medicinal material full wavelength scanner collection of illustrative plates
Fig. 2 a～c be the cape jasmine medicinal material respectively 240,330 and the 440nm wavelength under finger-print
Fig. 3 is the finger-print (combination collection of illustrative plates) of cape jasmine medicinal material multi-wavelength combination
Fig. 4 a～c is a cape jasmine medicinal material multi-wavelength HPLC qualitative and quantitative analysis reference substance collection of illustrative plates
Fig. 5 a～f is the multi-wavelength finger-print contrast of cultivation (a) and wild (b) cape jasmine medicinal material
Fig. 6 is a qingkailing injections full wavelength scanner collection of illustrative plates
Fig. 7 a～c is the uv absorption spectra of principal ingredient scutelloside, Gardenoside and chlorogenic acid in the qingkailing injections
Fig. 8 a～c be qingkailing injections respectively 240,280 and the 330nm wavelength under finger-print
Fig. 9 a～c is three kinds of principal ingredient reference substance HPLC chromatograms in the qingkailing injections
Figure 10 is that qingkailing injections is by the retention classification results
Figure 11 a～d is that similarity compared before the different manufacturers qingkailing injections adopted this fingerprint spectrum method to carry out Quality Control
Figure 12 a～d is that similarity compared after the different manufacturers qingkailing injections adopted this fingerprint spectrum method to carry out Quality Control
Embodiment
The construction method of multi-wavelength chromatographic fingerprints of Chinese materia medica of the present invention may further comprise the steps:
1, the Chinese medicine sample is prepared, in chromatogram preliminary election moving phase system, carry out rough segmentation, wherein stronger UV response is discerned and assert, result according to identification and identification, determine a plurality of detection wavelength and the tracking response of sample, measure capacity factor measure and corresponding moving phase proportionate relationship under the multistage wash-out ratio;
2, concern according to aforementioned proportion, determine in the gradient elution method of setting under the moving phase system, carrying out actual sample measures, detect chromatography eluant behavior under the wavelength according to separation case and each, carry out global optimization, adjust the gradient elution curve in real time, finally identification situation and the capacity factor measure according to tracking response distributes, determine the gradient curve under 1 hour～1.5 hours elution times, thereby determine the finger print measuring method of this sample;
3, above-mentioned definite assay method is carried out efficiency evaluation, comprising specificity, accuracy, precision, repeatability, stability, measurement range and durability;
4, the actual sample that is no less than 10 batches is measured, employing respectively detects the mode of measuring simultaneously under the wavelength or measuring respectively and measures under the same terms different wave length, the measurement result that obtains comprises collection of illustrative plates and the data under a plurality of detection wavelength, described data are vertically launched with retention time, gained relative peak area value under each wavelength or peak height value are carried out the consistance processing, be about to the data relativization, generally can adopt relative peak area value or peak height value, and the material composition peak by can be quantitative is as the fixed point index, carrying out the relativization of retention time handles, the data of mating corresponding mensuration batch, the total fingerprint peaks of identification, set up the relative peak area value of corresponding relative retention time of working sample or the finger-print data sequence of peak height value, corresponding sequence is averaged or gets median, form finger-print common pattern sequence under a plurality of single wavelength, carry out the data integration under a plurality of wavelength then, detecting the relative retention time that can respond the fingerprint peaks correspondence in wavelength coverages entirely with all is row, each detects wavelength data carry out sequence matrixization for row, corresponding time point does not have the position of response fills with 0, forms multi-wavelength finger-print common pattern data matrix;
5, identical Chinese medicine sample is taked that cold-maceration, temperature soak that method, percolation, decocting method, circumfluence method, pressurization leach, after one or more preparation methods in the decompression leaching, ultrasonic extraction, Microwave Extraction method, super critical extraction are prepared same sample, respectively repeated steps 4; Perhaps take one or more detection methods in uv detection method, evaporation light detection method, the Mass Spectrometer Method method that same sample is measured, respectively repeat steps 4.The a plurality of data matrixes that obtain are carried out diversity ratio, on this basis the data of step 4 gained are carried out stratification.Stratification is meant according to response magnitude or according to material composition differentiates situation, carry out data qualification, the aggregation of data that similar difference is changed is a class, identical category fingerprint peaks data are compared, choose wherein quantitatively composition or substantially qualitatively the stable elements peak be reference, other data in the class are carried out relativization to be handled, carry out the quantification treatment result of finger print data as relative content or relative characteristic ratio, the result inserts in the data matrix with the different levels quantification treatment, promptly forms to quantize finger-print common pattern data matrix.
6, it is qualitative the main chromatographic peak in the finger-print to be carried out finger-print, qualitatively can comprise three and infers levels: (1) adopts the reference substance contrast qualitative; (2) adopting the known structure tester is reference, utilizes among the LC/MS quasi-molecular ion or molion and corresponding fragment cracking rule and LC/DAD ultra-violet absorption spectrum feature to infer that fingerprint peaks contains the structure type of composition, is used for the supposition to the fingerprint peaks composition; (3) material composition that adopts LC/MS and LC/DAD information and retention sign fingerprint peaks to represent, content comprises that the retention of fingerprint peaks, real-time ultra-violet absorption spectrum and possible molecular weight and ms fragment information are used to identify this fingerprint peaks.
7, it is quantitative the main chromatographic peak in the finger-print to be carried out finger-print, quantitatively can comprise three levels, and (1) adopts reference substance to carry out accurately quantitatively; (2) be reference with accurately quantitative fingerprint peaks, the fingerprint peaks that supposition is had the same structure type compares with it; (3) adopt relative response to carry out quantitatively, adopt the mode of percentage response, carrying out quantified controlling with the message identification fingerprint peaks.
The multi-wavelength chromatographic fingerprints of Chinese materia medica that obtains through above-mentioned building mode, can measure for test agent the unknown, the similarity of utilizing above-mentioned matrix to carry out sample is differentiated and component difference compares, thereby realizes quality discrimination and the quality assessment analysis that qualitative, quantitative combines.
Below in conjunction with embodiment, the present invention further is described.
Embodiment one:
1, the structure of cape jasmine medicinal material multi-wavelength finger-print
(a) Agilent 1100 type high performance liquid chromatographs (DAD diode array detector), Phenomenex Luna C18 chromatographic column (250 * 4.6mm, 5 μ m) are adopted in experiment.
(b) after cape jasmine fruit is pulverized, cross 40 mesh sieves, add an amount of methyl alcohol and soaked 1 hour, sonicated 20 minutes is put coldly, and 3000rpm gets supernatant liquid filtering promptly after centrifugal 10 minutes.
(c) chromatographic condition: moving phase is the first alcohol and water, flow velocity 0.8ml/min, and 35 ℃ of column temperatures adopt linear gradient elution.
(d) according to the DAD detecting device at 200～600nm full wavelength scanner figure (as shown in Figure 1), for being reached, each component absorbs purpose strong, that interference is little, be chosen in 240nm and measure iridoid glycosides material (shown in Fig. 2-a), 330nm measures chlorogenic acid (shown in Fig. 2-b), 440nm measures crocin class material (shown in Fig. 2-c), obtains the independent finger-print of cape jasmine under three wavelength; Independent finger-print under these three wavelength is organically combined the combination finger-print (as shown in Figure 3) that just constitutes cape jasmine, and the spectrogram of heterogeneity adopts difference after the optimization to detect wavelength respectively to record among the figure, and it is respectively: 1, gardenia acid; 2, different Gardenoside; 4, Geniposide gentiobiose glycosides; 8, Gardenoside; 7, chlorogenic acid; 14, crocin 1; 16, crocin 2; 18, crocin 3; 19, crocetin.
(e) it is qualitative to adopt reference substance that the sample material composition is carried out, and shown in Fig. 4 a～c, is the multi-wavelength HPLC quantitative test reference substance collection of illustrative plates of cape jasmine medicinal material, has provided the qualitative qualification result of 9 kinds of compositions in the cape jasmine: 1, gardenia acid; 2, different Gardenoside; 3, Geniposide gentiobiose glycosides; 4, Gardenoside; 5, chlorogenic acid; 6, crocin 1; 7, crocin 2; 8, crocin 3; 9, crocetin takes the consistency analysis of online scanning spectra to carry out qualitative in conjunction with LC/MS to its homologue and the composition that do not have a reference substance.
(f) adopt the reference substance external standard method that the sample material composition is carried out quantitative measurement, in multi-wavelength HPLC finger-print (shown in Fig. 4 a～c), adopt 9 kinds of compositions in the reference substance quantitative measurement cape jasmine medicinal material, adopt relative peak area that its homologue and the composition that do not have a reference substance are carried out quantitative measurement, its measurement result is as shown in table 1:
Table 1 cape jasmine multi-wavelength finger-print common pattern data matrix tables of data
Peak area
Relative retention time
240nm 330nm 440nm
3.851 0 0 1706
12.064 2554908 0 0
14.037 909293 0 0
14.784 0 146862 0
15.691 506727 0 0
17.685 5684359 0 0
18.315 805815 1188839 2885
20.139 0 361535 0
20.587 26273922 0 0
22.933 558051 0 0
23.072 0 94150 0
23.413 403608 0 0
23.435 0 80959 0
30.080 0 114335 0
30.528 0 0 8621
31.040 0 138811 0
31.477 0 0 4722
31.659 0 231528 0
31.861 2974654 2652173 0
32.416 0 508344 0
32.427 120161 0 0
32.448 0 0 4189
32.747 756831 1450702 0
32.768 0 0 14450
33.28 362344 482370 0
33.792 0 0 4112
33.824 297857 436311 0
34.421 0 0 8598
34.432 848036 2333891 0
35.68 129105 0 0
35.883 156509 189857 0
36.053 0 0 4891
36.192 0 0 2156
37.493 63805488 76523788 343283
39.104 7243344 0 0
39.808 2541040 0 0
46.763 0 1598107 0
47.765 829225 903816 0
48.373 0 227272 0
49.749 0 1791878 0
(g) finger-print common pattern data matrix (as shown in table 1) be will quantize, the similarity differentiation and evaluation of sample will be used to carry out.
2, cape jasmine medicinal material multi-wavelength Application of Fingerprint
Shown in Fig. 5 a～f, be the multi-wavelength finger-print contrast of the cape jasmine medicinal material (a) (shown in Fig. 5-a, c, e) and the wild cape jasmine medicinal material (b) (shown in Fig. 5-b, d, f) of cultivation, this fingerprint spectrum method can clearly characterize the similarities and differences between the two.The result shows that Gardenoside in the wild cape jasmine and crocin constituents content generally a little more than the content of cultivation product, show that the cultivation quality amount is lower than wild kind.
Embodiment two:
1, the structure of QINGKAILING multi-wavelength finger-print
(a) adopt Tianjin, island LC-2010A type high performance liquid chromatograph (having SPD-M10 A type diode array detector, automatic sampling apparatus), Kromasil C18 stratographic analysis post (250 * 4.6mm, 5 μ m).
(b) qingkailing injections with 0.45 μ m membrane filtration after direct injected.
(c) chromatographic condition: moving phase is the A phase: 0.3% aqueous formic acid, B phase: methyl alcohol, column temperature: 30 ℃; Flow velocity: 1.0ml/min adopts linear gradient elution.
(d) pass through the DAD detecting device at 200～600nm full wavelength scanner (as shown in Figure 6), scanning result (solvent: methyl alcohol) show that Gardenoside has absorption maximum at the 238nm place; Scutelloside has absorption maximum at the 280nm place; Chlorogenic acid has absorption maximum at the 329nm place.Fig. 7 a～c is the uv absorption spectra of principal ingredient scutelloside, Gardenoside and chlorogenic acid in the qingkailing injections, among the figure: 1, Gardenoside, 2, scutelloside, 3, chlorogenic acid, solvent is a methyl alcohol.For being reached, each component absorbs purpose strong, that interference is little, finally be chosen in 238nm and measure Gardenoside (shown in Fig. 8-a), 280nm measures scutelloside (shown in Fig. 8-b), 330nm measures chlorogenic acid (shown in Fig. 8-c), obtain three finger-prints under the wavelength simultaneously, among the figure: 1, Gardenoside, 2, scutelloside, 3, chlorogenic acid.
(e) adopt reference substance to carry out qualitative to the sample material composition, take online LC/UV to carry out qualitative (shown in Fig. 9 a～c) to its homologue and the composition that do not have a reference substance in conjunction with LC/MS, among the figure three kinds of principal ingredients 1 in the QINGKAILING ZHUSHEJI, Gardenoside, 2, scutelloside, 3 chlorogenic acids qualitative.
(f) adopt the reference substance external standard method that the sample material composition is carried out quantitative measurement, adopts relative peak area that its homologue and the composition that do not have a reference substance are carried out quantitative measurement.
(g) the QINGKAILING fingerprint peaks is pressed retention and classified (as shown in figure 10), classification results adopts and quantizes finger-print common pattern data matrix, carries out the similarity of sample and differentiates and evaluation.Common pattern data matrix data are as shown in table 2:
Table 2 QINGKAILING multi-wavelength finger-print common pattern data matrix data
Peak area
Relative retention time
238nm 280nm 330nm
4.768 453610 0 0
4.779 0 389856 32210
6.293 0 882817 0
7.893 0 340205 0
10.272 0 536179 0
11.243 0 107203 0
11.659 844011 0 0
11.669 0 2023175 0
13.195 0 427316 0
13.205 557140 0 0
14.101 926108 324157 0
15.467 669109 595233 0
18.347 384221 0 0
18.389 0 0 103891
20.459 898142 0 0
20.469 0 449928 0
21.141 1998911 0 0
21.333 1357095 0 0
21.376 0 2935551 0
22.027 554408 0 0
23.691 0 163450 540045
28.363 0 0 426801
29.067 775892 0 0
29.728 0 0 428852
30.933 0 0 313032
32.395 5410397 0 0
42.24 0 0 152117
42.528 0 0 215112
43.371 1015044 2445299 3328821
44.928 479956 0 0
46.304 0 276533 172976
49.493 1.09E+08 0 0
49.504 0 0 1.15E+08
51.392 0 0 167862
52.171 0 0 332877
52.971 0 334864 0
53.611 1010591 2181996 0
53.621 0 0 1192408
54.731 3135701 7076680 3226072
75.328 4676526 625855 0
2, QINGKAILING multi-wavelength Application of Fingerprint
Shown in Figure 11 a～d, be that the different manufacturers qingkailing injections adopts finger-print of the present invention to carry out carrying out similarity relatively with standard diagram before the Quality Control.The finger-print of each factory's product reflects original situation of product substantially, and finger-print differs greatly between each producer, and similarity is respectively 0.958,0.780,0.650.
Shown in Figure 12 a～d, be that similarity compared after the different manufacturers qingkailing injections adopted finger-print of the present invention to carry out Quality Control.Through carrying out process modification under the finger-print guidance, finger-print reaches unanimity between each producer's product, and similarity is respectively 0.974,0.941,0.925.
From the above description as can be seen, multi-wavelength finger-print obtained by the method for the present invention can effectively, comprehensively, synthetically reflect the qualitative variability of tcm product, can carry out quality control effectively, make different manufacturers can produce product unified, stable, equal in quality and that reach quality standards from medicinal material, semi-manufacture to the final products overall process.
Claims (10)
1, a kind of construction method of multi-wavelength chromatographic fingerprints of Chinese materia medica, it may further comprise the steps:
(1) the Chinese medicine sample is prepared, in chromatogram preliminary election moving phase system, carry out rough segmentation, wherein stronger UV response is discerned and assert, result according to identification and identification, determine a plurality of detection wavelength and the tracking response of sample, measure capacity factor measure and corresponding moving phase proportionate relationship under the multistage wash-out ratio;
(2) concern according to aforementioned proportion, determine in the gradient elution method of setting under the moving phase system, carrying out actual sample measures, detect chromatography eluant behavior under the wavelength according to separation case and each, carry out global optimization, adjust the gradient elution curve in real time, finally identification situation and the capacity factor measure according to tracking response distributes, determine the gradient curve under 1 hour～1.5 hours elution times, thereby determine the finger print measuring method of this sample;
(3) above-mentioned definite assay method is carried out efficiency evaluation, comprising specificity, accuracy, precision, repeatability, stability, measurement range and durability;
(4) actual sample that is no less than 10 batches is measured, measurement result comprises collection of illustrative plates and the data under a plurality of detection wavelength, described data are vertically launched with retention time, gained relative peak area value under each wavelength or peak height value are carried out the consistance processing, set up the relative peak area value of corresponding relative retention time of working sample or the finger-print data sequence of peak height value, corresponding sequence is averaged or gets median, form finger-print common pattern sequence under a plurality of single wavelength, carry out the data integration under a plurality of wavelength then, detecting the relative retention time that can respond the fingerprint peaks correspondence in wavelength coverages entirely with all is row, each detects wavelength data carry out sequence matrixization for row, corresponding time point does not have the position of response fills with 0, forms multi-wavelength finger-print common pattern data matrix;
(5) identical Chinese medicine sample is taked multiple preparation method preparation or taked multiple detection method that same sample is measured, and respectively repeat steps (4), the a plurality of data matrixes that obtain are carried out diversity ratio, on this basis the data of step (4) gained are carried out stratification;
(6) it is qualitative the main chromatographic peak in the finger-print to be carried out finger-print;
(7) it is quantitative the main chromatographic peak in the finger-print to be carried out finger-print.
2, the construction method of multi-wavelength chromatographic fingerprints of Chinese materia medica as claimed in claim 1 is characterized in that: in the step (6), described finger-print is qualitative to be comprised three and infers levels, promptly 1. adopts the reference substance contrast qualitative; 2. adopting the known structure tester is reference, utilize the ultra-violet absorption spectrum feature of quasi-molecular ion in the liquid chromatography/mass spectrometry coupling or molion and corresponding fragment cracking rule and liquid chromatography/diode array detector to infer that fingerprint peaks contains the structure type of composition, is used for the supposition to the fingerprint peaks composition; 3. adopt the material composition of liquid chromatography/mass spectrometry coupling and liquid chromatography/diode array detector information and retention sign fingerprint peaks representative, content comprises that the retention of fingerprint peaks, real-time ultra-violet absorption spectrum and possible molecular weight and ms fragment information are used to identify this fingerprint peaks.
3, the construction method of multi-wavelength chromatographic fingerprints of Chinese materia medica as claimed in claim 1 is characterized in that: in the step (7), described finger-print quantitatively comprises three levels, 1. adopts reference substance to carry out accurately quantitatively; 2. be reference with accurately quantitative fingerprint peaks, the fingerprint peaks that supposition is had the same structure type compares with it, adopts relative response to carry out quantitatively; 3. adopt the mode of percentage response, to carrying out quantified controlling with the message identification fingerprint peaks.
4, the construction method of multi-wavelength chromatographic fingerprints of Chinese materia medica as claimed in claim 1, it is characterized in that: in the step (5), described stratification is meant according to response magnitude or according to material composition differentiates situation, carry out data qualification, the aggregation of data that similar difference is changed is a class, identical category fingerprint peaks data are compared, choose wherein quantitatively composition or substantially qualitatively the stable elements peak be reference, other data in the class are carried out relativization to be handled, carry out the quantification treatment result of finger print data as relative content or relative characteristic ratio, the result inserts in the data matrix with the different levels quantification treatment, promptly forms to quantize finger-print common pattern data matrix.
5, the construction method of multi-wavelength chromatographic fingerprints of Chinese materia medica as claimed in claim 1, it is characterized in that: in the step (5), described multiple preparation method comprises that cold-maceration, temperature are soaked method, percolation, decocting method, circumfluence method, pressurization leaching, leaching, ultrasonic extraction, Microwave Extraction method, super critical extraction reduce pressure.
6, the construction method of multi-wavelength chromatographic fingerprints of Chinese materia medica as claimed in claim 1, it is characterized in that: in the step (4), described consistance is handled and is meant employing relative peak area value or peak height value, carrying out the relativization of retention time handles, the data of mating corresponding mensuration batch, and the material composition peak by can be quantitative is as the fixed point index, the total fingerprint peaks of identification.
7, the construction method of multi-wavelength chromatographic fingerprints of Chinese materia medica as claimed in claim 1, it is characterized in that: in the step (4), collection of illustrative plates under described a plurality of detection wavelength and data are to detect to measure simultaneously under the wavelength to obtain or measure respectively under the same terms different wave length obtaining at each.
8, a kind of multi-wavelength chromatographic fingerprints of Chinese materia medica that obtains by the construction method of the described multi-wavelength chromatographic fingerprints of Chinese materia medica of claim 1.
9, the application of multi-wavelength chromatographic fingerprints of Chinese materia medica as claimed in claim 8 in quality discrimination.
10, the application of multi-wavelength chromatographic fingerprints of Chinese materia medica as claimed in claim 8 in quality assessment is analyzed.
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| CN1544935A true CN1544935A (en) | 2004-11-10 |
| CN1281948C CN1281948C (en) | 2006-10-25 |
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101788469A (en) * | 2010-03-08 | 2010-07-28 | 辅仁药业集团有限公司 | Method for online detecting near infrared spectrum of active ingredients of compound eucommia bark capsules |
| CN102692460A (en) * | 2012-05-22 | 2012-09-26 | 辽宁中医药大学 | Process for determining content of four components of honeysuckle during full-time-interval three-wavelength fusion |
| CN105920071A (en) * | 2016-04-29 | 2016-09-07 | 中国科学院新疆理化技术研究所 | Applications of safflower extract with definite spectrum-effect relationship |
| CN106568882A (en) * | 2017-02-21 | 2017-04-19 | 广东铭康香精香料有限公司 | Method for analyzing aroma components of shampoo |
| CN108088947A (en) * | 2017-12-14 | 2018-05-29 | 中国日用化学工业研究院 | The assay method of the polysaccharide degree of polymerization in polyglycoside surfactants |
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| CN113075370A (en) * | 2021-03-12 | 2021-07-06 | 药都(本溪)一致科技有限公司 | Method, system, medium and application for measuring quality consistency of traditional Chinese medicine |
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2003
- 2003-11-14 CN CN 200310103862 patent/CN1281948C/en not_active Expired - Fee Related
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101788469A (en) * | 2010-03-08 | 2010-07-28 | 辅仁药业集团有限公司 | Method for online detecting near infrared spectrum of active ingredients of compound eucommia bark capsules |
| CN102692460A (en) * | 2012-05-22 | 2012-09-26 | 辽宁中医药大学 | Process for determining content of four components of honeysuckle during full-time-interval three-wavelength fusion |
| CN105920071A (en) * | 2016-04-29 | 2016-09-07 | 中国科学院新疆理化技术研究所 | Applications of safflower extract with definite spectrum-effect relationship |
| CN105920071B (en) * | 2016-04-29 | 2019-07-23 | 中国科学院新疆理化技术研究所 | A kind of purposes of the safflower extract with clear spectrum effect relationship |
| CN106568882A (en) * | 2017-02-21 | 2017-04-19 | 广东铭康香精香料有限公司 | Method for analyzing aroma components of shampoo |
| CN106568882B (en) * | 2017-02-21 | 2019-02-12 | 广东铭康香精香料有限公司 | A method of analysis shampoo fragrance component |
| CN108088947A (en) * | 2017-12-14 | 2018-05-29 | 中国日用化学工业研究院 | The assay method of the polysaccharide degree of polymerization in polyglycoside surfactants |
| CN109580803A (en) * | 2018-08-24 | 2019-04-05 | 成都中医药大学 | A kind of Russia's color leaf extract and its quality determining method |
| CN109580803B (en) * | 2018-08-24 | 2021-09-17 | 成都中医药大学 | Russian leaf extract and quality detection method thereof |
| CN109307723A (en) * | 2018-11-26 | 2019-02-05 | 成都中医药大学 | A kind of the UPLC detection method and its discrimination method of cape jasmine and water cape jasmine |
| CN113075370A (en) * | 2021-03-12 | 2021-07-06 | 药都(本溪)一致科技有限公司 | Method, system, medium and application for measuring quality consistency of traditional Chinese medicine |
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| Publication number | Publication date |
|---|---|
| CN1281948C (en) | 2006-10-25 |
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