CN108508223A - A kind of high-throughput Continuous Flow Analysis method for chlorinity in tobacco gene editor's material - Google Patents
A kind of high-throughput Continuous Flow Analysis method for chlorinity in tobacco gene editor's material Download PDFInfo
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- CN108508223A CN108508223A CN201810147250.6A CN201810147250A CN108508223A CN 108508223 A CN108508223 A CN 108508223A CN 201810147250 A CN201810147250 A CN 201810147250A CN 108508223 A CN108508223 A CN 108508223A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
- G01N35/10—Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/34—Purifying; Cleaning
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Abstract
The invention discloses a kind of high-throughput Continuous Flow Analysis methods for chlorinity in tobacco gene editor's material, including step：A, it accurately weighs the sample that 0.25g contains chlorine to be placed in 25mL sample cells, excellent sample feeding pipe will be claimed to be put into successively on the sample injection disc of Continuous Flow Analysis instrument autosampler, until all piling；B, deionized water 25mL is separately added into each sample cell with liquid-transfering gun, then it is sequentially inserted into the inner sleeve of filtering sieve plate of the bottom with 0.45 μm of aperture, solution splashes out during preventing oscillation extraction, and sample disc is put on oscillator after being plugged inner sleeve, carries out oscillation extraction；C, inner sleeve is pushed after having extracted, the sample solution extracted is allowed to be entered in inner sleeve by the filtering sieve plate, sample disc is placed on the autosampler of Continuous Flow Analysis instrument again, the sample introduction needle of autosampler is only insertd at the supernatant in inner sleeve every time to draw filtered sample, is transferred in Continuous Flow Analysis instrument and is analyzed.
Description
Technical field
The invention belongs to tobacco chemistry research fields, and in particular to chlorinity is examined in a kind of suitable tobacco gene editor material
The method for high-flux analysis of survey.
Background technology
With tobacco business " batch production breeding side of the tobacco gene group plan key special subjects based on genome editing technique
The propulsion of case " screens material by chemical composition analysis, and pass through tobacco leaf using all gene editing materials as object
Compatibility test and cigarette composition verification, determine the strong kind of industrial applicability, become the pass of tobacco business active demand
Key technology content.
Chlorine is the important indicator of routine chemical components and tobacco quality evaluation important in tobacco.Chlorine in tobacco at present
Detection commonly use connected neighbor set.Standard working solution, sample, reagent are pumped into chemistry by Continuous Flow Analysis technology in proportion
In analyzer, and gas is sent into timing simultaneously, makes to form continuous fluid separating system in pipeline, makes the substance that participation is reacted even
It is sufficiently mixed during afterflow is dynamic, then such as mixes, heats, filters by series reaction, examined finally by detector
Corresponding peak electrical is measured, then corresponding concentration is calculated automatically from by standard curve.Continuous Flow Analysis technology
Have many advantages, such as that fast detection speed, sample and reagent dosage are few, at home and abroad tobacco business is widely applied.
In the standard method (YC/T 162-2011) of current tobacco routine chemical components analysis, the measurement of chlorine uses water
Sample is extracted, is then measured again with Continuous Flow Analysis instrument.In sample pretreatment process, sample need to be weighed to ground
In triangular flask, extractant oscillation extraction is added, is filtered with qualitative filter paper, filtered solution is transferred to Continuous Flow Analysis instrument
In sample feeding pipe, subsequent measurements then could be carried out.It during the filtration process, need to be by solid sample residue and extracting solution in triangular flask
Mixture be poured into bottleneck be placed with qualitative filter paper folding funnel in, by after filter extracting solution collect in receiving flask.Sample
It needs to shift sample several times in product pretreatment process, it is not easy enough, influence the working efficiency of detection；And it may in transfer process
The reliability of testing result is influenced because introducing accidentally splashing etc. for impurity and extracting solution.
Gene editing batch production breeding at least will produce tens thousand of editor's materials, at present the day sample treatment of chlorine sample analysis
Amount only 60 or so, the routine chemical components analysis for completing tens thousand of a materials need very long period, and the lag of analysis result will
It influences whether the progress of entire batch production breeding, therefore quickly divides accurately and reliably high-throughput in gene editing breeding process
The demand of analysis method is very urgent.
Invention content
The purpose of the present invention is to provide a kind of high-throughput Continuous Flow Analysis method of gene editing material chlorinity, packets
Include following steps：
A, it accurately weighs the sample that 0.25g contains chlorine to be placed in 25mL sample cells, from Continuous Flow Analysis instrument automatic sampling
Sample injection disc is removed on device, excellent sample feeding pipe will be claimed to be put into successively on Continuous Flow Analysis instrument sample injection disc, until by the sample injection disc
In 100 sample cell positions all pile sample cell；
B, it is separately added into deionized water 25mL into each sample cell with the liquid-transfering gun of 25mL, is then sequentially inserted into bottom belt
There is the inner sleeve of the filtering sieve plate in 0.45 μm of aperture, solution splashes out during preventing oscillation extraction, is plugged after inner sleeve sample
Disk is put on oscillator, and oscillation extraction is carried out by the oscillation extraction condition that standard method is selected；
C, the inner sleeve of sample cell is pushed after having extracted, the sample solution extracted is allowed to enter by the filtering sieve plate
It is placed on the autosampler of Continuous Flow Analysis instrument into inner sleeve, then sample disc, the sample introduction needle of autosampler is every
To draw filtered sample at the secondary supernatant only insertd into inner sleeve, it is transferred in Continuous Flow Analysis instrument and is analyzed.
Preferably, sealing ring is carried at the top of the sample cell, the inner sleeve is inserted into the sealing before oscillation extraction
In circle, extract liquor is then added, inner sleeve is stuck in the oral area of sample cell prevents extract liquor in oscillatory process from splashing out, but inner sleeve
The filtering sieve plate of bottom is not contacted with sample；Inner sleeve is pushed after sample extraction is complete, extract liquor, which is just filtered by sieve plate, to be entered
Into inner sleeve, autosampler can extract sample directly from inner sleeve and be analyzed.
In order to reduce transfer workload, inventor have also been devised with the matched oscillator of the sample disc of autosampler, can
Sample disc is removed from autosampler, fills direct oscillation extraction after a disk sample, inner sleeve filtering, entire sample after having extracted
Product disk is put on autosampler and is analyzed again.Whole operation flow need not shift specific sample, before sample
Processing procedure is greatly simplified, and sample analysis efficiency is compared with current tobacco business standard method and several times can be improved.And
The reduction of transfer step can effectively prevent the error in sample pre-treatments operating process, greatly reduce the introducing of human error.
In addition, in conventional tobacco chlorine flow analysis continuous mode, tobacco need to be broken into fine powder and be extracted, in conventional method
Fine powder meeting partial gelatinization blocks filter paper in sample pretreatment process, and the rate of filtration is slower, and in improved processing method, oscillation
It is slightly placed after the completion of extraction, under tobacco sample powder just sinks to, sieve plate when filtering in inner sleeve only connects with supernatant
It touches, can effectively avoid the sieve plate in filter process from blocking in this way, faster, filter operation is easier to realize the rate of filtration.
When sample analysis, by tobacco business standard YC/T 269-2008, (measurement of tobacco and tobacco product chlorine continuously flows
Method) selected condition is measured.It is first worked curve with sulfuric acid sodium standard solution when measurement, sample is placed in automatically in order
On injector, the titer of an intermediate concentration is put after 10 samples, and as quality control, (sample introduction frequency is every two minutes 1
A sample)；Analysis after hours, according to peak height from working curve calculate sample in chlorinity.
It is higher than conventional method to the uniformity requirement of sample since connected neighbor set sample volume is small, the present invention
In test influence of the sample comminution grain size to measurement result, the results showed that：Sample is not sieved, the precision of measurement result compared with
Difference, with the increase for crossing grit number, the precision of sample measurement result increases, and when mesh number reaches 60 mesh or more, sample measures knot
The precision of fruit tends towards stability, and the relative standard deviation (RSD) of measurement is 1% hereinafter, therefore selection crushes simultaneously in the present invention
Cross 60 sieves.
Choose flue-cured tobacco, suncured tabacco, Turkish tobaccos and finished cigarettes sample, respectively in the present inventive method with tobacco business standard
YC/T 162-2011 (the measurement continuous flow method of tobacco and tobacco product chlorine) are measured, and compare (table to result
1).The method of the present invention measurement result and at present without significant difference, analysis result is accurate with tobacco business standard method measurement result
It is really reliable.
The comparison (%) of 1 the method for the present invention of table and current standard
Beneficial effects of the present invention
Project team is directed in current tobacco deficiency present in chlorine Continuous Flow Analysis quadrat method, targetedly devises
Quick sample pretreatment method.In improving analysis method, whole operation flow need not shift sample, sample pretreatment process
It is greatly simplified.Using more than two sample disc alternate treatments (a disk sample introduction, a disk pre-treatment), can it is continual into
Row analysis, daily 2 people can handle 8-10 disk samples, and routinely 60 samples of sample introduction speed/h is calculated, and can analyze 400 or more daily
Sample, sample analysis efficiency are compared with current tobacco business standard method and several times can be improved, and actual sample analysis result and
Industry standard methods are consistent at present.The present invention is that the high flux screening for magnanimity editor's material that gene batch production breeding generates carries
High-throughput rapid analysis method has been supplied, the process that chlorine is analyzed in gene editing material screening process is greatly accelerated.
In conventional tobacco chlorine flow analysis continuous mode, tobacco need to be broken into fine powder and be extracted, in conventional method sample
Fine powder meeting partial gelatinization blocks filter paper in pretreatment process, and the rate of filtration is slower, and in improved processing method, oscillation extraction
It slightly places after the completion, under tobacco sample powder just sinks to, sieve plate when filtering in inner sleeve is only contacted with supernatant, this
Sample can effectively avoid the sieve plate in filter process from blocking, and faster, filter operation is easier to realize the rate of filtration.
Description of the drawings
Fig. 1 is the sample injection disc used in the present invention.
Fig. 2 is the sample feeding pipe used in the present invention and the schematic diagram of inner sleeve.Wherein a figures are sample feeding pipes；B figures are inner sleeves
Pipe；C figures are schematic diagram when inner sleeve is inserted at the top of sample feeding pipe；D figures are the schematic diagram that inner sleeve presses downwards；E figures are inner sleeve
Pipe is pressed onto bottom and completes the schematic diagram for being inserted into sampling probe Aspirate supernatant after original position is filtered again.
Specific implementation mode
The present invention is further described by following specific examples, but does not limit the present invention.
Embodiment 1：The measurement of chlorine in cured tobacco leaf sample A
1) sample pre-treatments：Tobacco leaf (or pipe tobacco) is dried first, tobacco and tobacco product are carried out with high speed whirlwind mill
Then cell wall broken wall crosses 60 mesh screens, be sealed as standby sample.By YC/T31-1996 standardized test specimen moisture content.By with
Lower processing：
A, it accurately weighs the sample that 0.25g contains chlorine to be placed in 25mL sample cells, from Continuous Flow Analysis instrument automatic sampling
Sample injection disc is removed on device, excellent sample feeding pipe will be claimed to be put into successively on Continuous Flow Analysis instrument sample injection disc, until by the sample injection disc
In 100 sample cell positions all pile sample cell；
B, it is separately added into deionized water 25mL into each sample cell with the liquid-transfering gun of 25mL, is then sequentially inserted into bottom belt
There is the inner sleeve of the filtering sieve plate in 0.45 μm of aperture, solution splashes out during preventing oscillation extraction, is plugged after inner sleeve sample
Disk is put on oscillator, and oscillation extraction is carried out by the oscillation extraction condition that standard method is selected；
C, the inner sleeve of sample cell is pushed after having extracted, the sample solution extracted is allowed to enter by the filtering sieve plate
It is placed on the autosampler of Continuous Flow Analysis instrument into inner sleeve, then sample disc, the sample introduction needle of autosampler is every
To draw filtered sample at the secondary supernatant only insertd into inner sleeve, it is transferred in Continuous Flow Analysis instrument and is analyzed.
2) according to standard method YC/T 162-2011 (the measurement continuous flow method of tobacco and tobacco product chlorine) relevant regulations
Complete the subsequent measurements work of chlorine in sample.
It is 0.62% to measure chlorinity in cured tobacco leaf sample A.
Embodiment 2：The measurement of chlorine in finished cigarettes sample B
The process in embodiment 1 is repeated, selects finished cigarettes B to carry out the measurement of chlorine in its pipe tobacco, measuring chlorinity is
0.81%.
Claims (2)
1. a kind of high-throughput Continuous Flow Analysis method for chlorinity in tobacco gene editor's material, which is characterized in that packet
Include following steps：
A, it accurately weighs the sample that 0.25g contains chlorine to be placed in 25mL sample cells, from Continuous Flow Analysis instrument autosampler
Sample injection disc is removed, excellent sample feeding pipe will be claimed to be put into successively on Continuous Flow Analysis instrument sample injection disc, until will be in the sample injection disc
All pile sample cell in 100 sample cell positions；
B, it is separately added into deionized water 25mL into each sample cell with the liquid-transfering gun of 25mL, is then sequentially inserted into bottom and carries
The inner sleeve of the filtering sieve plate in 0.45 μm of aperture, solution splashes out during preventing oscillation extraction, is plugged after inner sleeve sample disc
It is put on oscillator, oscillation extraction is carried out by the oscillation extraction condition that standard method is selected；
C, the inner sleeve of sample cell is pushed after having extracted, allows the sample solution extracted to be entered by the filtering sieve plate interior
In casing, then sample disc is placed on the autosampler of Continuous Flow Analysis instrument, the sample introduction needle of autosampler is every time only
To draw filtered sample at the supernatant being inserted into inner sleeve, it is transferred in Continuous Flow Analysis instrument and is analyzed.
2. high throughput Continuous Flow Analysis method according to claim 1, which is characterized in that carried at the top of the sample cell
Sealing ring, the inner sleeve are inserted into before oscillation extraction in the sealing ring, and extract liquor is then added, and inner sleeve is stuck in sample
The oral area of quality control prevents extract liquor in oscillatory process from splashing out, but the filtering sieve plate of inner sleeve bottom of the tube is not contacted with sample；Sample extracts
Inner sleeve is pushed after taking, extract liquor is just entered by sieve plate filtering in inner sleeve, and autosampler can be directly from inner sleeve
Sample is extracted in pipe to be analyzed.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810147250.6A CN108508223A (en) | 2018-02-12 | 2018-02-12 | A kind of high-throughput Continuous Flow Analysis method for chlorinity in tobacco gene editor's material |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810147250.6A CN108508223A (en) | 2018-02-12 | 2018-02-12 | A kind of high-throughput Continuous Flow Analysis method for chlorinity in tobacco gene editor's material |
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| Publication Number | Publication Date |
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| CN108508223A true CN108508223A (en) | 2018-09-07 |
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| Application Number | Title | Priority Date | Filing Date |
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| CN201810147250.6A Pending CN108508223A (en) | 2018-02-12 | 2018-02-12 | A kind of high-throughput Continuous Flow Analysis method for chlorinity in tobacco gene editor's material |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109142600A (en) * | 2018-10-18 | 2019-01-04 | 云南中烟工业有限责任公司 | A kind of method of 4 kinds of mercapturic acids in measurement smoker urine |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101071104A (en) * | 2007-04-17 | 2007-11-14 | 广东中烟工业公司 | Method for determining chloride content in tobacco |
| CN203264748U (en) * | 2013-05-24 | 2013-11-06 | 中国热带农业科学院分析测试中心 | Filter cover of soil sample effective state analysis centrifugal tube |
| CN107199063A (en) * | 2017-05-19 | 2017-09-26 | 黑龙江谱尼测试科技有限公司 | A kind of novel filter-type colorimetric cylinder |
-
2018
- 2018-02-12 CN CN201810147250.6A patent/CN108508223A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101071104A (en) * | 2007-04-17 | 2007-11-14 | 广东中烟工业公司 | Method for determining chloride content in tobacco |
| CN203264748U (en) * | 2013-05-24 | 2013-11-06 | 中国热带农业科学院分析测试中心 | Filter cover of soil sample effective state analysis centrifugal tube |
| CN107199063A (en) * | 2017-05-19 | 2017-09-26 | 黑龙江谱尼测试科技有限公司 | A kind of novel filter-type colorimetric cylinder |
Non-Patent Citations (1)
| Title |
|---|
| 国家烟草专卖局: "《中华人民共和国烟草行业标准 YC/T 162-2011 烟草及烟草制品 氯的测定 连续流动法》", 31 December 2011, 中国标准出版社 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109142600A (en) * | 2018-10-18 | 2019-01-04 | 云南中烟工业有限责任公司 | A kind of method of 4 kinds of mercapturic acids in measurement smoker urine |
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Application publication date: 20180907 |