CN102221599A - Inspection method capable of inspecting traditional Chinese medicine injection tannins - Google Patents
Inspection method capable of inspecting traditional Chinese medicine injection tannins Download PDFInfo
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- CN102221599A CN102221599A CN2011100464939A CN201110046493A CN102221599A CN 102221599 A CN102221599 A CN 102221599A CN 2011100464939 A CN2011100464939 A CN 2011100464939A CN 201110046493 A CN201110046493 A CN 201110046493A CN 102221599 A CN102221599 A CN 102221599A
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Abstract
The invention relates to an inspection method capable of inspecting traditional Chinese medicine injection tannins, and is used for inspecting the tannin impurities of a traditional Chinese medicine injection to control the quality of the traditional Chinese medicine injection. The inspection method provided by the invention is characterized by comprising the following steps: 1) coating a protein absorption membrane with proteins; 2) enabling a traditional Chinese medicine injection solution to be in contact with the protein coated membrane; 3) washing protein coated membrane by utilizing the solution containing organic solvents; and 4) observing the color of sample application speckles and judging the tannins in the traditional Chinese medicine injection. In the inspection method provided by the invention, the concentration of the tannins in the traditional Chinese medicine injection which are inspected by adopting a sample application mode and can reach 0.75-0.25mg/ml, can reach 0.07mg/ml by adopting a filtering mode; the inspection limit quantity is lower than the quantity in the Pharmacopeia (first chapter) of the People's Republic of China (PRC) in 2010 edition; and simultaneously, the interference resistance is strong and is suitable for the inspection of the traditional Chinese medicine injection tannins.
Description
Affiliated technical field
The present invention relates to traditional Chinese medicine determination of foreign matter field, be used for traditional Chinese medicine tannin class impurity, with the quality of control traditional Chinese medicine.
Technical background
Traditional Chinese medicine is created in the forties in last century the earliest, is started by scholars such as Qian Xinzhong.In China, traditional Chinese medicine has been widely used in clinical and has obtained the approval of industry.One one of the version Pharmacopoeia of the People's Republic of China in 2010 (be called for short " Chinese pharmacopoeia) has just been included folk prescription and compound injections such as erigeron breviscapus, QINGKAILING, 'Zhichuanling.Traditional Chinese medicine is advancing the form of Chinese drug development, expands the range of application of Chinese medicine, performs meritorious deeds never to be obliterated really in aspects such as service numerous people health.Safe, effective, quality controllable still is the gross requirement of traditional Chinese medicine.Technologically speaking, industry has solved the validity problem (although mechanism of action is still not exclusively removed) of traditional Chinese medicine substantially, and safety issue still is an industry difficult problem.
Yet, along with traditional Chinese medicine be widely used in clinically, its bad reaction exposed day by day, particularly lethal bad reaction also have report repeatly.According to present disclosed bad reaction data, the bad reaction of traditional Chinese medicine can be involved each system and internal organs, even causes death.The bad reaction of traditional Chinese medicine and specific impurity are undivided.
Traditional Chinese medicine mostly is plant extracts greatly.As bio-extract, in the raw material leaching process, can bring the big molecular impurity of various plants, as protein, nucleic acid etc.Tannin energy and protein bound are common traditional Chinese medicine algogenic substances.The kind of tannin is more, and the monomer of tannin such as micromolecule such as tannic acid, gallic acid also have the tannin macromolecule polyalcohol of different condensation levels.The tannin instability, oxidation easily, polymerization, variable color, even produce precipitation, be one of main matter that influences the traditional Chinese medicine quality.It is generally acknowledged the pharmacological action of the unlikely participation traditional Chinese medicine of condensed tannin.Because the molecular weight of condensed tannin is bigger, orally do not absorb, if traditional Chinese medicine from the oral formulations exploitation, condensed tannin is impossible the participation curative effect so, also can cause bad reaction on the contrary.Therefore 2010 version " People's Republic of China (PRC) state pharmacopeia (one one) " (to call " pharmacopeia " in the following text) the traditional Chinese medicine determination of foreign matter of special control tannin is also arranged.
Exist the tannin in the traditional Chinese medicine to belong to potpourri, the molecular weight of tannin is a uncertain value, so molecular weight and physicochemical property have bigger difference, is difficult to check with instrument analytical methods such as HPLC and/or mass spectrums.
Because tannin, particularly having the condensed tannin in the traditional Chinese medicine is the important impurity that influences traditional Chinese medicine, therefore is necessary to control as strict as possible.The characteristics that The present invention be directed to traditional Chinese medicine are set up, and tannin class determination of foreign matter is had the characteristics highly sensitive, that antijamming capability is strong.
Summary of the invention
The present invention utilizes the characteristics of albumen mass-energy high strength adsorption precipitation tannin to set up this method, to improve sensitivity and the specificity that the traditional Chinese medicine tannin is checked.This method may further comprise the steps:
1, with the protein adsorption film with protein bag quilt;
2, the film of Chinese medicine injection agent solution with protein bag quilt contacted;
3, with the solution washing protein coated film that contains organic solvent;
4, observe the shade of point sample spot, judge the tannin in the traditional Chinese medicine.
In above step 1, include but not limited to nitrocellulose filter, nylon membrane, pvdf membrane (Polyvinylidene fluoride, PVDF membrane) with the film of protein bag quilt, used protein includes but not limited to egg white, bovine serum albumin(BSA).
In above step 2, the mode that the Chinese medicine injection agent solution contacts with the protein coated film includes but not limited to point sample mode and filter type, can carry out moistening processing with organic solvents such as methyl alcohol earlier before contacting with the protein coated film.
In above step 3, the organic solution concentration of washed protein coated film is 0-100%, and organic solvent includes but not limited to methyl alcohol and DMSO (dimethyl sulfoxide (DMSO)).Adopt the amount of organic solution washing and the number of times of washing not to limit.
The effect that this method is implemented is, checks the sensitivity and the specificity height of traditional Chinese medicine tannin, and antijamming capability is strong.
The recommendation condition of the inventive method is: the film of protein bag quilt adopts pvdf membrane, and the way of contact of parenteral solution and protein coated film is a filter type, and the organic solution of washed protein coated film is DMSO, and washing is 3 times behind the point sample, each 10 minutes.
Description of drawings
Fig. 1 sets up the tannin inspection method for the pvdf membrane that adopts protein bag quilt in the point sample mode, determines that the present invention checks that the figure as a result of limit: A is the result after some variable concentrations tannin solution on the common pvdf membrane is by methanol wash; B is for adopting the picture after 10% egg white solution bag is by the washing of pvdf membrane point sample; 20.00,6.67,2.22,0.74,0.25,0.08,0.03,0mg/ml point 1 to 8 pyrogallic acid content is respectively:.Point sample amount 1 μ l.
Fig. 2 sets up the tannin inspection method for the nitrocellulose filter (NC film) that adopts protein bag quilt in the point sample mode, determines that the present invention checks that the figure as a result of limit: A is the result after some variable concentrations tannin solution on the common NC film is by methanol wash; B is for adopting the picture after 10% egg white solution bag is by the washing of NC film spot sample; 20.00,6.67,2.22,0.74,0.25,0.08,0.03,0mg/ml point 1 to 8 pyrogallic acid content is respectively:.Point sample amount 1 μ l.
Fig. 3 is checked that with filter type the tannin in the solution limits the quantity of by pvdf membrane for the albumen bag: A be the albumen bag by pvdf membrane, B is common pvdf membrane.The tannin concentration of point 1-5 is 2.00,0.67,0.22,0.07,0mg/ml (by pyrogallic acid), and the filtration volume is 0.5ml.
Fig. 4 for the protein bag by the pvdf membrane point sample after by the figure as a result of methyl alcohol DMSO mixed solution washing: the tannin concentration of point sample is: 20.00mg/ml (by pyrogallic acid); Point sample amount 1 μ l.
Fig. 5 for the present invention check in the point sample mode figure as a result of phenol traditional Chinese medicine ingredients scutellarin: A for the scutellarin aqueous solution point of saturation concentration on common pvdf membrane, the spot figure through the methanol wash front and back; B be with the scutellarin aqueous solution point of saturation concentration the protein bag by pvdf membrane on, the spot figure before and after methanol wash; Point sample amount 1 μ l.
Fig. 6 is that the tannin of gradient concentration is according to " after the Chinese pharmacopoeia method precipitation tannin, getting the figure as a result that supernatant is checked with this law in the point sample mode again: tannin theoretical concentration＞1000,333,111,37.0,12.3,4.11, the 0 μ g/ml of some 1-7.Point sample amount 1 μ l.
Fig. 7 mixes the figure as a result of centrifuged supernatant with point sample mode check criteria tannin solution and 1% egg white solution for the present invention: the content of each point tannin is respectively from left to right: 20,000,6,667,2,222,741,246,82,27,9,3,1.5,0 μ g/ml (by pyrogallic acid), point sample amount 1 μ l.
Fig. 8 is for adopting absorption curve method check criteria tannin solution abosrption spectrogram: the tannin concentration of R1-10 is respectively 20,000,6,667,2,222,741,246,82,27,9,3,1.5 μ g/ml (by pyrogallic acid), and R0 does not contain tannin.
Fig. 9 mixes the abosrption spectrogram of centrifuged supernatant mixing centrifuged supernatant for adopting the molten and 1% egg white solution of absorption curve method check criteria tannin: the tannin concentration of R1-10 is respectively 20,000,6,667,2,222,741,246,82,27,9,3,1.5 μ g/ml (by pyrogallic acid), R0 does not contain tannin.
Figure 10 checks the figure as a result of four kinds of traditional Chinese medicine tannins for the present invention in the point sample mode: 1,2,3,4 be respectively qingkailing injections, Shuanhuanglian injection, danshen injections and be the spot of erigeron breviscapus parenteral solution.
Figure 11 checks the figure as a result of four kinds of traditional Chinese medicine tannins with filter type for the present invention: 1,2,3,4 be respectively qingkailing injections, Shuanhuanglian injection, danshen injections and be the spot of erigeron breviscapus parenteral solution.Filtrable volume 0.5ml.
Embodiment
Exemplifying embodiment 1
Adopt the point sample mode to set up traditional Chinese medicine tannin inspection method, determine to check limit with the pvdf membrane of protein bag quilt
Get a pvdf membrane, be immersed in after moistening with methyl alcohol in 10% the egg white solution, the room temperature jolting was taken out air-dry standby after 3 hours.
Make the pyrogallic acid solution 5ml that concentration is 20mg/ml with distilled water, room temperature is placed naturally, and (410nm d=1cm) reaches more than 3 as tannin standard inventory solution etc. solution absorbency.Get 8 1.5ml test tube numbering 1-8.No. 1 pipe adding standard tannin stock solution 1.5ml.Respectively add distilled water 1ml in 2-8 number pipe subsequently, solution 0.5ml joins pipe No. 2 from No. 1 pipe, gets 0.5ml behind the mixing again and joins pipe No. 3, is formulated into pipe successively No. 7.Promptly obtain concentration gradient and be one group of tannin standard solution of 3 times, the concentration of tannin from No. 1 pipe to 8 pipe be followed successively by 20.00,6.67,2.22,0.74,0.25,0.08,0.03,0mg/ml.
Will by the pvdf membrane of protein bag quilt with wetted with methanol after, put gradient tannin standard solution 1 μ l respectively.Then this film is immersed in the methanol solution and washs each 10 minutes 3 times.Take pictures this film is air-dry subsequently, the result as shown in Figure 1.
From Fig. 1 as seen, sample spot on not by the pvdf membrane of protein bag quilt, this film colourless basically (Figure 1A) after methanol wash, and on the pvdf membrane of protein bag quilt, after methanol wash, still can demonstrate tannin standard solution point the color (Figure 1B) of tannin.Adopt the pvdf membrane of protein bag quilt, this law checks that limiting the quantity of of tannin is 0.75-0.25mg/ml.
Exemplifying embodiment 2
Adopt the point sample mode to set up traditional Chinese medicine tannin inspection method, determine to check limit with the NC film of protein bag quilt
Get a NC film and be immersed in 10% the egg white solution, the room temperature jolting was taken out air-dry standby after 3 hours.
Make the pyrogallic acid solution 5ml that concentration is 20mg/ml with distilled water, room temperature is placed naturally, and (410nm d=1cm) reaches more than 3 as tannin standard inventory solution etc. solution absorbency.Get 8 1.5ml test tube numbering 1-8.No. 1 pipe adding standard tannin stock solution 1.5ml.Respectively add distilled water 1ml in 2-8 number pipe subsequently, solution 0.5ml joins pipe No. 2 from No. 1 pipe, gets 0.5ml behind the mixing again and joins pipe No. 3, is formulated into pipe successively No. 7.Promptly obtain concentration gradient and be one group of tannin standard solution of 3 times, the concentration of tannin from No. 1 pipe to 8 pipe be followed successively by 20.00,6.67,2.22,0.74,0.25,0.08,0.03,0mg/ml.
On by the NC film of protein bag quilt, put gradient tannin standard solution 1 μ l respectively.Then this film is immersed in 50% methanol solution and washs each 10 minutes 3 times.Take pictures this film is air-dry subsequently, the result as shown in Figure 1.
From Fig. 2 as seen, sample spot is not on by the NC film of protein bag quilt, this film colourless basically (Fig. 2 A) after 50% methanol wash, and the tannin standard solution point color (Fig. 2 B) that point 1 still can demonstrate tannin after 50% methanol wash on the NC film of protein bag quilt.Adopt the NC film of protein bag quilt, this law checks that limiting the quantity of of tannin is 20mg/ml, and the pvdf membrane of protein bag quilt is adopted in contrast, and the NC film of protein bag quilt checks that effect is more bad.
Exemplifying embodiment 3
Adopt the filtration mode to set up the protein bag, determine to check and limit the quantity of by pvdf membrane tannin inspection method
With the pyrogallic acid solution (pH4-5) of 6mg/ml, in 80 ℃ of constant temperature 3 days, after the 410nm absorption value reaches more than 3 as the tannin reference material, and according to 3 times of gradients preparation tannin dilutions; Get a usefulness 10% egg white bag by pvdf membrane, be installed on the aperture filter connection vacuum pump; With the moistening filter membrane of 0.1ml methyl alcohol, add the tannin aqueous solution 0.5ml of variable concentrations subsequently earlier, negative pressure is passed through filter membrane; Filter membrane is taken out, add methyl alcohol (DMSO) solution and place jolting 30min on the shaking table; So with not operated in contrast by the pvdf membrane of protein bag quilt; Wash to the colourless substantially termination of control film washing, and take pictures on the gel imaging instrument, the result as shown in Figure 3.
The result shows that filter type is checked limiting the quantity of at least below 0.07mg/ml of tannin.Therefore the filtration mode checks that than point sample mode limit is low, if increase detection volume, limiting the quantity of of detection will further descend.
Exemplifying embodiment 4
The protein bag is by the study on the stability of pvdf membrane tannin spot
Get a pvdf membrane, be immersed in after moistening with methyl alcohol in 10% the egg white solution, the room temperature jolting was taken out air-dry standby after 3 hours.
Make the pyrogallic acid solution 5ml that concentration is 20mg/ml with distilled water, room temperature is placed naturally, and (410nm d=1cm) reaches more than 3 as tannin standard inventory solution etc. solution absorbency.
With the pvdf membrane of protein bag quilt with wetted with methanol after, the tannin standard solution 1 μ l of many places point 20mg/ml.Then this film is immersed in the methanol solution and washs each 10 minutes 3 times.Subsequently this film is cut off to be immersed in again in the mixed solution that contains different methyl alcohol and DMSO and washed 3 times, each 10 minutes, take out subsequently and take pictures.Use not by parallel test of pvdf membrane of protein bag quilt in addition.The result as shown in Figure 4.
From Fig. 4 as seen, tannin point is on the pvdf membrane of protein bag quilt, after methanol wash, wash with methyl alcohol DMSO mixed solution once more, still there be (Fig. 3 A) in positive spots, and tannin standard solution point is handled back positive spots disappearance (Fig. 3 B) through same procedure on not by the pvdf membrane of protein bag quilt.
Exemplifying embodiment 5
Check Chinese medicine phenolic constituent scutellarin with this law point sample mode
Get a pvdf membrane, be immersed in after moistening with methyl alcohol in 10% the egg white solution, the room temperature jolting was taken out air-dry standby after 3 hours.
Get a sterile injection powder that contains scutellarin, standby with the saturated aqueous solution of distilled water preparation scutellarin, solution is golden yellow.
With the pvdf membrane of protein bag quilt with wetted with methanol after, some scutellarin saturated aqueous solution 1 μ l.Then this film is immersed in the methanol solution and washs each 10 minutes 3 times.Take pictures before this film washing, also take pictures behind the laundry air-dry.The result as shown in Figure 5.
From Fig. 5 as seen, scutellarin tannin point all do not had some idea of a little before the methanol wash, but spot all disappears after methanol wash on the common pvdf membrane and on the pvdf membrane of protein bag quilt.Show that the coloured phenolic substance of micromolecule in the Chinese medicine is less to the interference of this method.
Exemplifying embodiment 6
This law point sample mode is checked the comparison of tannin and official method
Get a pvdf membrane, be immersed in after moistening with methyl alcohol in 10% the egg white solution, the room temperature jolting was taken out air-dry standby after 3 hours.Other prepares 1% egg white normal saline solution.
Make the pyrogallic acid solution 5ml that concentration is 6mg/ml with distilled water, room temperature is placed naturally, and (410nm d=1cm) reaches more than 3 as tannin standard inventory solution etc. solution absorbency.Get 8 1.5ml test tube numbering 1-7.No. 1 pipe adding standard tannin stock solution 1.5ml.Respectively add distilled water 1ml in 2-8 number pipe subsequently, solution 0.5ml joins pipe No. 2 from No. 1 pipe, gets 0.5ml behind the mixing again and joins pipe No. 3, is formulated into pipe successively No. 6.Promptly obtain concentration gradient and be one group of tannin standard solution of 3 times, the concentration of tannin from No. 1 pipe to 8 pipe be followed successively by 6.00,2.00,0.67,0.22,0.07,0.02,0mg/ml.
Get each standard tannin liquid 1ml and join in the color comparison tube, add 5ml 1% egg white normal saline solution again, placed observations 10 minutes.Get liquid 1ml from color comparison tube subsequently and join the 1.5ml centrifuge tube, through 15, centrifugal 5 minutes of 000g, observation has or not precipitation.The result is as shown in table 1.Table 1 result shows that " Chinese pharmacopoeia is checked limiting the quantity of＞6mg/ml of tannin, and limiting the quantity of of centrifuge method is about 6mg/ml.
The protein precipitation of each standard tannin pipe of table 1 changes
+ expression visible precipitate or muddiness ,-expression are not seen precipitation or muddy.
*Be theoretical concentration,, cause the part tannin no longer in solution because No. 1 pipe has precipitation after centrifugal.
With the pvdf membrane of protein bag quilt with wetted with methanol after, get above-mentioned centrifuged supernatant and put gradient tannin standard solution 1 μ l respectively.Then this film is immersed in the methanol solution and washs each 10 minutes 3 times.Take pictures this film is air-dry subsequently, the result as shown in Figure 6.
From Fig. 6 visible point 1 and point 2 significantly positive spots is arranged, tannin is by " Chinese pharmacopoeia method post precipitation, still visible tannin, the inspection in check result and the exemplifying embodiment 1 limit the quantity of (0.75-0.25mg/ml) coincide, so the inspection of this law limit is lower than " Chinese pharmacopoeia method.
Check and tannin and the residual tannin of the mixed centrifugal supernatant in back of 10% egg white solution with this law point sample mode
Get a pvdf membrane and be immersed in 10% the egg white solution, the room temperature jolting was taken out air-dry standby after 3 hours.
Make the pyrogallic acid solution 5ml that concentration is 20mg/ml with distilled water, room temperature is placed naturally, and (410nm d=1cm) reaches more than 3 as tannin standard inventory solution etc. solution absorbency.Get 11 1.5ml test tube numbering 1-10,0.No. 1 pipe adding standard tannin stock solution 1.5ml.Respectively add distilled water 1ml in 2-8 number pipe subsequently, solution 0.5ml joins pipe No. 2 from No. 1 pipe, gets 0.5ml behind the mixing again and joins pipe No. 3, is formulated into pipe successively No. 10.Promptly obtain concentration gradient and be one group of tannin standard solution of 3 times, the concentration of tannin is followed successively by 20,000,6,667,2,222,741,246,82,27,9,3,1.5 μ g/ml from No. 1 pipe to 10 pipe, and No. 0 pipe does not contain tannin.
Get each standard tannin solution 0.5ml of preparation, add 0.5ml 10% egg white solution and fully mix, through 15,000g is after centrifugal 5 minutes, with supernatant 1 μ l point sample to the pvdf membrane of protein bag quilt.Then this film is immersed in the methanol solution and washs each 10 minutes 3 times.Take pictures this film is air-dry subsequently, the result as shown in Figure 7.The result shows that by the egg white post precipitation, this law still can be checked out the tannin in the supernatant solution.
Other gets each the tannin standard solution 50 μ l that does not mix with egg white solution, joins in 384 orifice plates, and each hole sample carries out absorption curve scanning, the results are shown in Figure 8.Also get simultaneously with egg white solution and mix centrifugal back supernatant solution 100 μ l, also join in 384 orifice plates.Each hole sample also carries out absorption spectrum scanning, the results are shown in Figure 9.Comparison diagram 8 and Fig. 9, tannin is behind protein precipitation, and absorption curve moves to left, but is leaning on the 400nm place, and absorption value is still higher, shows that protein is difficult to coloured tannin precipitation fully.
This law point sample mode detects four kinds of traditional Chinese medicine tannins
Get a pvdf membrane, be immersed in after moistening with methyl alcohol in 10% the egg white solution, the room temperature jolting was taken out air-dry standby after 3 hours.
Buy qingkailing injections, Shuanhuanglian injection, danshen injections and erigeron breviscapus parenteral solution from the market, adopt the freeze drying mode that these four kinds of parenteral solutions are concentrated, the preparation sample.
With the pvdf membrane of protein bag quilt with wetted with methanol after, put each sample 1 μ l respectively.Then this film is immersed in the methanol solution and washs each 10 minutes 3 times.Take pictures this film is air-dry subsequently, the result as shown in figure 10.
From Figure 10 as seen, four kinds of traditional Chinese medicine injection sample spot of different cycles of concentration on the pvdf membrane of protein bag quilt, the color of this film showed different after methanol wash.For injection of the same race, concentrating degree is high more, and it is dark more to develop the color.On the whole, the content of tannin of danshen injections is higher.
This law is filtered suction type and is detected four kinds of traditional Chinese medicine tannins
Get a usefulness 10% egg white bag by pvdf membrane, be installed on aperture (diameter 2mm) filter connection vacuum pump; With the moistening filter membrane of 0.1ml methyl alcohol, add different parenteral solutions (qingkailing injections, Shuanhuanglian injection, danshen injections and erigeron breviscapus parenteral solution) aqueous solution 0.5ml subsequently earlier, negative pressure is passed through filter membrane; Filter membrane is taken out, add methyl alcohol (DMSO) solution and place jolting 30min on the shaking table; So with not operated in contrast by the pvdf membrane of protein bag quilt; Wash to the colourless substantially termination of control film washing, and take pictures on the gel imaging instrument, the result as shown in figure 11.
Figure 11 result shows that the content of tannin in the danshen injections is higher, and the content of tannin in qingkailing injections and the erigeron breviscapus parenteral solution is less.
Claims (4)
1. method of checking the traditional Chinese medicine tannin, its feature with carry out according to the following steps:
1) with the protein adsorption film with protein bag quilt;
2) the Chinese medicine injection agent solution is contacted with the protein coated film;
3) with the solution washing protein coated film that contains organic solvent;
4) shade of observation point sample spot is judged the tannin in the traditional Chinese medicine.
2. according to the described traditional Chinese medicine tannin of claim 1 inspection method, it is characterized in that the protein adsorption film includes but not limited to pvdf membrane (PVDF membrane), nitrocellulose filter (NC film), the protein of coated film includes but not limited to egg white, bovine serum albumin(BSA).
3. according to the described traditional Chinese medicine tannin of claim 1 inspection method, it is characterized in that the mode that the Chinese medicine injection agent solution contacts with the protein coated film includes but not limited to point sample mode and filter type.
4. according to the described traditional Chinese medicine tannin of claim 1 inspection method, it is characterized in that the used organic solution concentration of washed protein coated film is 0-100%, organic solvent includes but not limited to methyl alcohol and DMSO (dimethyl sulfoxide (DMSO)), and the mixing of their two or more arbitrary proportions; Adopt the amount of organic solution washing and the number of times of washing not to limit.
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| CN109668886A (en) * | 2019-02-26 | 2019-04-23 | 云南中医学院 | A kind of inspection method of the micro tannin of traditional Chinese medicine |
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| CN101526520A (en) * | 2008-03-07 | 2009-09-09 | 国家纳米科学中心 | Method and device for biological sample detection |
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| CN101865926A (en) * | 2010-06-13 | 2010-10-20 | 云南中医学院 | Detection method for trace protein of Chinese medicinal injection |
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| US6057160A (en) * | 1997-08-06 | 2000-05-02 | Washington State University Research Foundation | Methods for preparation and use of a coomassie brilliant blue/protein complex |
| EP1113270A2 (en) * | 1999-12-28 | 2001-07-04 | Matsushita Electronics Corporation | Reagent and method for measuring a concentration of protein |
| CN101526520A (en) * | 2008-03-07 | 2009-09-09 | 国家纳米科学中心 | Method and device for biological sample detection |
| CN101625313A (en) * | 2009-01-22 | 2010-01-13 | 九芝堂股份有限公司 | Method for measuring content of tannide in nakedflower beautyberry |
| CN101865926A (en) * | 2010-06-13 | 2010-10-20 | 云南中医学院 | Detection method for trace protein of Chinese medicinal injection |
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| CN109668886A (en) * | 2019-02-26 | 2019-04-23 | 云南中医学院 | A kind of inspection method of the micro tannin of traditional Chinese medicine |
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