CN106018635B - The method and detection method of pre-treatment are carried out to tobacco or tobacco product - Google Patents
The method and detection method of pre-treatment are carried out to tobacco or tobacco product Download PDFInfo
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- CN106018635B CN106018635B CN201610658005.2A CN201610658005A CN106018635B CN 106018635 B CN106018635 B CN 106018635B CN 201610658005 A CN201610658005 A CN 201610658005A CN 106018635 B CN106018635 B CN 106018635B
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- dinitrophenylhydrazone
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- 241000208125 Nicotiana Species 0.000 title claims abstract description 73
- 235000002637 Nicotiana tabacum Nutrition 0.000 title claims abstract description 72
- 235000019505 tobacco product Nutrition 0.000 title claims abstract description 70
- 238000000034 method Methods 0.000 title claims abstract description 58
- 238000001514 detection method Methods 0.000 title claims abstract description 51
- 238000002203 pretreatment Methods 0.000 title abstract description 4
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims abstract description 174
- 150000001728 carbonyl compounds Chemical class 0.000 claims abstract description 75
- HORQAOAYAYGIBM-UHFFFAOYSA-N 2,4-dinitrophenylhydrazine Chemical compound NNC1=CC=C([N+]([O-])=O)C=C1[N+]([O-])=O HORQAOAYAYGIBM-UHFFFAOYSA-N 0.000 claims abstract description 46
- 239000000284 extract Substances 0.000 claims abstract description 38
- 239000002253 acid Substances 0.000 claims abstract description 24
- 238000001212 derivatisation Methods 0.000 claims abstract description 19
- 150000002500 ions Chemical class 0.000 claims description 45
- JUJWROOIHBZHMG-UHFFFAOYSA-N Pyridine Chemical compound C1=CC=NC=C1 JUJWROOIHBZHMG-UHFFFAOYSA-N 0.000 claims description 42
- 239000012071 phase Substances 0.000 claims description 33
- VLTRZXGMWDSKGL-UHFFFAOYSA-N perchloric acid Chemical compound OCl(=O)(=O)=O VLTRZXGMWDSKGL-UHFFFAOYSA-N 0.000 claims description 28
- ZTQSAGDEMFDKMZ-UHFFFAOYSA-N butyric aldehyde Natural products CCCC=O ZTQSAGDEMFDKMZ-UHFFFAOYSA-N 0.000 claims description 21
- UMJSCPRVCHMLSP-UHFFFAOYSA-N pyridine Natural products COC1=CC=CN=C1 UMJSCPRVCHMLSP-UHFFFAOYSA-N 0.000 claims description 21
- 238000002386 leaching Methods 0.000 claims description 20
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims description 19
- HGINCPLSRVDWNT-UHFFFAOYSA-N Acrolein Chemical compound C=CC=O HGINCPLSRVDWNT-UHFFFAOYSA-N 0.000 claims description 18
- 230000000391 smoking effect Effects 0.000 claims description 18
- NBBJYMSMWIIQGU-UHFFFAOYSA-N Propionic aldehyde Chemical compound CCC=O NBBJYMSMWIIQGU-UHFFFAOYSA-N 0.000 claims description 14
- 238000003556 assay Methods 0.000 claims description 13
- UEQLSLWCHGLSML-UHFFFAOYSA-N n-(methylideneamino)-2,4-dinitroaniline Chemical compound [O-][N+](=O)C1=CC=C(NN=C)C([N+]([O-])=O)=C1 UEQLSLWCHGLSML-UHFFFAOYSA-N 0.000 claims description 13
- YGIXYAIGWMAGIB-UHFFFAOYSA-N 2,4-dinitro-n-(propan-2-ylideneamino)aniline Chemical compound CC(C)=NNC1=CC=C([N+]([O-])=O)C=C1[N+]([O-])=O YGIXYAIGWMAGIB-UHFFFAOYSA-N 0.000 claims description 10
- JJPZHGIYUVFTGG-BJMVGYQFSA-N 2,4-dinitro-n-[(e)-prop-2-enylideneamino]aniline Chemical compound [O-][N+](=O)C1=CC=C(N\N=C\C=C)C([N+]([O-])=O)=C1 JJPZHGIYUVFTGG-BJMVGYQFSA-N 0.000 claims description 10
- WPWSANGSIWAACK-UHFFFAOYSA-N n-(butan-2-ylideneamino)-2,4-dinitroaniline Chemical compound CCC(C)=NNC1=CC=C([N+]([O-])=O)C=C1[N+]([O-])=O WPWSANGSIWAACK-UHFFFAOYSA-N 0.000 claims description 10
- GFUVNGJSSAEZHW-PDGXAJGZSA-N n-[(e)-[(e)-but-2-enylidene]amino]-2,4-dinitroaniline Chemical compound C\C=C\C=N\NC1=CC=C([N+]([O-])=O)C=C1[N+]([O-])=O GFUVNGJSSAEZHW-PDGXAJGZSA-N 0.000 claims description 10
- IKGRHEWIFBFXPP-IZZDOVSWSA-N n-[(e)-butylideneamino]-2,4-dinitroaniline Chemical compound CCC\C=N\NC1=CC=C([N+]([O-])=O)C=C1[N+]([O-])=O IKGRHEWIFBFXPP-IZZDOVSWSA-N 0.000 claims description 10
- ONBOQRNOMHHDFB-XNWCZRBMSA-N n-[(e)-ethylideneamino]-2,4-dinitroaniline Chemical compound C\C=N\NC1=CC=C([N+]([O-])=O)C=C1[N+]([O-])=O ONBOQRNOMHHDFB-XNWCZRBMSA-N 0.000 claims description 10
- NFQHZOZOFGDSIN-BJMVGYQFSA-N 2,4-dinitro-n-[(e)-propylideneamino]aniline Chemical compound CC\C=N\NC1=CC=C([N+]([O-])=O)C=C1[N+]([O-])=O NFQHZOZOFGDSIN-BJMVGYQFSA-N 0.000 claims description 9
- 238000006243 chemical reaction Methods 0.000 claims description 9
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 9
- SMWDFEZZVXVKRB-UHFFFAOYSA-N Quinoline Chemical compound N1=CC=CC2=CC=CC=C21 SMWDFEZZVXVKRB-UHFFFAOYSA-N 0.000 claims description 8
- MLUCVPSAIODCQM-NSCUHMNNSA-N crotonaldehyde Chemical compound C\C=C\C=O MLUCVPSAIODCQM-NSCUHMNNSA-N 0.000 claims description 8
- MLUCVPSAIODCQM-UHFFFAOYSA-N crotonaldehyde Natural products CC=CC=O MLUCVPSAIODCQM-UHFFFAOYSA-N 0.000 claims description 8
- 239000012074 organic phase Substances 0.000 claims description 8
- ZWEHNKRNPOVVGH-UHFFFAOYSA-N 2-Butanone Chemical compound CCC(C)=O ZWEHNKRNPOVVGH-UHFFFAOYSA-N 0.000 claims description 7
- 235000019504 cigarettes Nutrition 0.000 claims description 7
- 238000001914 filtration Methods 0.000 claims description 7
- 239000007788 liquid Substances 0.000 claims description 7
- WSFSSNUMVMOOMR-NJFSPNSNSA-N methanone Chemical compound O=[14CH2] WSFSSNUMVMOOMR-NJFSPNSNSA-N 0.000 claims description 7
- HGBOYTHUEUWSSQ-UHFFFAOYSA-N valeric aldehyde Natural products CCCCC=O HGBOYTHUEUWSSQ-UHFFFAOYSA-N 0.000 claims description 7
- IKHGUXGNUITLKF-XPULMUKRSA-N acetaldehyde Chemical compound [14CH]([14CH3])=O IKHGUXGNUITLKF-XPULMUKRSA-N 0.000 claims description 6
- 238000010828 elution Methods 0.000 claims description 6
- XVMSFILGAMDHEY-UHFFFAOYSA-N 6-(4-aminophenyl)sulfonylpyridin-3-amine Chemical compound C1=CC(N)=CC=C1S(=O)(=O)C1=CC=C(N)C=N1 XVMSFILGAMDHEY-UHFFFAOYSA-N 0.000 claims description 5
- 101100447665 Mus musculus Gas2 gene Proteins 0.000 claims description 5
- 230000001055 chewing effect Effects 0.000 claims description 5
- 238000001294 liquid chromatography-tandem mass spectrometry Methods 0.000 claims description 5
- 239000002994 raw material Substances 0.000 claims description 5
- 101100348341 Caenorhabditis elegans gas-1 gene Proteins 0.000 claims description 4
- KRHYYFGTRYWZRS-UHFFFAOYSA-N Fluorane Chemical compound F KRHYYFGTRYWZRS-UHFFFAOYSA-N 0.000 claims description 4
- 101100447658 Mus musculus Gas1 gene Proteins 0.000 claims description 4
- 238000012207 quantitative assay Methods 0.000 claims description 4
- 238000000132 electrospray ionisation Methods 0.000 claims description 3
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- 230000010355 oscillation Effects 0.000 claims description 2
- 238000010998 test method Methods 0.000 claims 2
- 125000002915 carbonyl group Chemical group [*:2]C([*:1])=O 0.000 abstract description 10
- 230000035945 sensitivity Effects 0.000 abstract description 6
- 150000001875 compounds Chemical class 0.000 abstract description 4
- 238000004445 quantitative analysis Methods 0.000 abstract description 2
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- 239000012224 working solution Substances 0.000 description 13
- 239000000523 sample Substances 0.000 description 10
- 239000012488 sample solution Substances 0.000 description 10
- WSFSSNUMVMOOMR-UHFFFAOYSA-N Formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 description 9
- IKHGUXGNUITLKF-UHFFFAOYSA-N Acetaldehyde Chemical compound CC=O IKHGUXGNUITLKF-UHFFFAOYSA-N 0.000 description 8
- 239000000779 smoke Substances 0.000 description 8
- 229940125782 compound 2 Drugs 0.000 description 6
- 238000000605 extraction Methods 0.000 description 6
- 238000004458 analytical method Methods 0.000 description 5
- KEUXLNUAJRPQNF-UHFFFAOYSA-N acetaldehyde;propan-2-one Chemical compound CC=O.CC(C)=O KEUXLNUAJRPQNF-UHFFFAOYSA-N 0.000 description 4
- 239000002904 solvent Substances 0.000 description 4
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- 238000011084 recovery Methods 0.000 description 3
- 241000894007 species Species 0.000 description 3
- 239000011550 stock solution Substances 0.000 description 3
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 description 2
- 235000003255 Carthamus tinctorius Nutrition 0.000 description 2
- 244000020518 Carthamus tinctorius Species 0.000 description 2
- -1 aldehyde compounds Chemical class 0.000 description 2
- 239000007864 aqueous solution Substances 0.000 description 2
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- 238000005516 engineering process Methods 0.000 description 2
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- 238000002552 multiple reaction monitoring Methods 0.000 description 2
- 239000003960 organic solvent Substances 0.000 description 2
- 150000002989 phenols Chemical class 0.000 description 2
- 238000000926 separation method Methods 0.000 description 2
- 231100000419 toxicity Toxicity 0.000 description 2
- 230000001988 toxicity Effects 0.000 description 2
- 238000005303 weighing Methods 0.000 description 2
- NFQHZOZOFGDSIN-UHFFFAOYSA-N 2,4-dinitro-n-(propylideneamino)aniline Chemical compound CCC=NNC1=CC=C([N+]([O-])=O)C=C1[N+]([O-])=O NFQHZOZOFGDSIN-UHFFFAOYSA-N 0.000 description 1
- 244000061322 Nicotiana alata Species 0.000 description 1
- 244000006449 Nicotiana forgetiana Species 0.000 description 1
- 241000208128 Nicotiana glauca Species 0.000 description 1
- 244000061176 Nicotiana tabacum Species 0.000 description 1
- GRYLNZFGIOXLOG-UHFFFAOYSA-N Nitric acid Chemical compound O[N+]([O-])=O GRYLNZFGIOXLOG-UHFFFAOYSA-N 0.000 description 1
- 241000208292 Solanaceae Species 0.000 description 1
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 150000001299 aldehydes Chemical class 0.000 description 1
- WNROFYMDJYEPJX-UHFFFAOYSA-K aluminium hydroxide Chemical compound [OH-].[OH-].[OH-].[Al+3] WNROFYMDJYEPJX-UHFFFAOYSA-K 0.000 description 1
- 239000000908 ammonium hydroxide Substances 0.000 description 1
- 125000003118 aryl group Chemical group 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- DUEPRVBVGDRKAG-UHFFFAOYSA-N carbofuran Chemical compound CNC(=O)OC1=CC=CC2=C1OC(C)(C)C2 DUEPRVBVGDRKAG-UHFFFAOYSA-N 0.000 description 1
- 150000001735 carboxylic acids Chemical class 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 235000019506 cigar Nutrition 0.000 description 1
- 210000004081 cilia Anatomy 0.000 description 1
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- 238000010438 heat treatment Methods 0.000 description 1
- 235000008216 herbs Nutrition 0.000 description 1
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- 238000010829 isocratic elution Methods 0.000 description 1
- 150000002576 ketones Chemical class 0.000 description 1
- 238000004949 mass spectrometry Methods 0.000 description 1
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- 239000000463 material Substances 0.000 description 1
- 239000013618 particulate matter Substances 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 238000000197 pyrolysis Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000000638 solvent extraction Methods 0.000 description 1
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- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 239000011701 zinc Substances 0.000 description 1
- UGZADUVQMDAIAO-UHFFFAOYSA-L zinc hydroxide Chemical compound [OH-].[OH-].[Zn+2] UGZADUVQMDAIAO-UHFFFAOYSA-L 0.000 description 1
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Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
- G01N2030/067—Preparation by reaction, e.g. derivatising the sample
Landscapes
- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Other Investigation Or Analysis Of Materials By Electrical Means (AREA)
Abstract
The invention belongs to test and analyze field, the method for being related to a kind of pair of tobacco or tobacco product progress pre-treatment includes the following steps, (1) extracts tobacco sample or tobacco product sample with acetonitrile, obtains the first extract of carbonyl containing compound；(2) 2,4-dinitrophenylhydrazine and/or 2,4-dinitrophenylhydrazine solution is added into first extract and strong acid solution performs the derivatization, the extracting solution I of isolated carbonyl containing compound derivatization product.The invention further relates to method, kit and the purposes of carbonyls in detection tobacco or tobacco product.The present invention confirms for the first time contains carbonyls in tobacco and tobacco product, and the method for the present invention accurate quantitative analysis goes out carbonyl compound content in tobacco or tobacco product, and detection limit is low, high sensitivity, reproducible.
Description
Technical Field
The invention belongs to the field of detection and analysis, and particularly relates to a method for pretreating tobacco or tobacco products, and a method for detecting carbonyl compounds in the tobacco or tobacco products.
Background
Carbonyl compounds are an important class of harmful constituents in the smoke produced by smoking tobacco products and are included in the "hoffman list" of 44 harmful constituents in smoke. Such compounds include formaldehyde, acetaldehyde, acrolein, and the like, with acetaldehyde being the largest. The low molecular weight carbonyl compound has stronger cilium toxicity, and the most toxic is low molecular weight aldehyde compounds such as acrolein, formaldehyde, acetaldehyde and the like, and the toxicity of the low molecular weight carbonyl compound is gradually reduced along with the increase of the molecular weight. Carbonyl compounds in the smoke mainly come from pyrolysis of phenols in the tobacco products, and the phenols in the tobacco products are formed by non-enzymatic browning reactions.
Currently, methods for detecting carbonyl compounds in mainstream smoke include: the smoke particulate matter of the main stream smoke is collected by a glass fiber filter disc, and is subjected to solvent extraction and quantitative analysis by a high performance liquid chromatography-ultraviolet detector. The solvent used for the extraction is usually a mixture of an organic solvent and water. Moreover, it has been found that the detection accuracy of a part of carbonyl compounds is reduced and some carbonyl compounds cannot be detected even if the proportion of the organic solvent in the solvent is too high.
So far, researchers have not found that the tobacco or the tobacco products contain carbonyl compounds, and have not carried out detection analysis on the carbonyl compounds.
Disclosure of Invention
The inventor proves that the tobacco and/or the tobacco products contain carbonyl compounds through a great deal of experimental research. The present inventors have also obtained a method for pre-treating tobacco or tobacco products which is capable of extracting carbonyl compounds from the tobacco or tobacco products and converting them into derivatised products. On the basis, the inventor also obtains a method for detecting carbonyl compounds in tobacco or tobacco products, and the carbonyl compounds in the tobacco or tobacco products are quantitatively detected by detecting the derivative products of the carbonyl compounds. The method has the advantages of high accuracy, high sensitivity, low detection limit and good repeatability.
The invention relates in a first aspect to a method for pretreating tobacco or tobacco products, comprising the following steps:
(1) leaching a tobacco sample or a tobacco product sample by using acetonitrile to obtain primary extract containing carbonyl compounds;
(2) and adding a 2, 4-dinitrophenylhydrazine and/or a 2, 4-dinitrophenylhydrazine solution and a strong acid solution into the primary extract for derivatization, and separating to obtain an extract I containing a carbonyl compound derivatization product.
In one embodiment of the first aspect of the present invention, the method further comprises the following steps:
(3) and adding weak base or weak base solution into the extracting solution I, and mixing to obtain an extracting solution II.
In one embodiment, the weak base is selected from one or more of pyridine and quinoline, preferably pyridine.
In one embodiment, the weak base solution is selected from one or more of an acetonitrile solution of pyridine and an acetonitrile solution of quinoline, preferably an acetonitrile solution of pyridine.
In one embodiment, the pyridine in acetonitrile (as pyridine) or pyridine is added in an amount of 300-.
In one embodiment of the first aspect of the present invention, any one or more of the following a to O are included:
A. the tobacco product is selected from one or more of a combustible smoking tobacco product and a non-combustible smoking tobacco product; preferably, the tobacco product is a combustible smoking tobacco product, more preferably a cigarette;
B. the carbonyl compound is selected from one or more of formaldehyde, acetaldehyde, acetone, propionaldehyde, acrolein, butyraldehyde, 2-butanone and crotonaldehyde;
C. in the step (1), the leaching feed-liquid ratio is 1 (10-50) (g/mL), preferably 1 (10-30) (g/mL), and more preferably 1:20 (g/mL);
D. in the step (1), the leaching is carried out under oscillation conditions;
E. in the step (1), the leaching time is 10 to 60 minutes, preferably 20 to 50 minutes, and more preferably 20 minutes, 30 minutes or 50 minutes;
F. in the step (1), the leaching temperature is normal temperature, preferably 5-40 ℃, more preferably 10-35 ℃, and further preferably 10 ℃, 25 ℃ or 30 ℃;
G. in the step (2), the addition amount of the 2, 4-dinitrophenylhydrazine solution (calculated by 2, 4-dinitrophenylhydrazine) or the 2, 4-dinitrophenylhydrazine is 0.2-5mg/(mL of the primary extract), preferably 1-3mg/(mL of the primary extract), and more preferably 2mg/(mL of the primary extract);
H. in the step (2), the dosage of the strong acid solution is 300-;
I. in the step (2), the 2, 4-dinitrophenylhydrazine solution is acetonitrile solution of 2, 4-dinitrophenylhydrazine; preferably, the concentration of the 2, 4-dinitrophenylhydrazine in the acetonitrile solution of the 2, 4-dinitrophenylhydrazine is 1-40mg/mL, more preferably 5-20mg/mL, and further preferably 10 mg/mL;
J. in the step (2), the strong acid solution is acetonitrile solution of strong acid; preferably, the concentration of the strong acid in the acetonitrile solution of the strong acid is 0.05% to 2% (W/W), more preferably 0.1% to 1% (W/W), and still more preferably 0.2% (W/W);
K. in the step (2), the strong acid solution is selected from one or more of perchloric acid solution and hydrofluoric acid solution;
l. in step (2), the time of derivatization is 20 to 60 minutes, preferably 30 to 50 minutes, more preferably 40 minutes;
m, in the step (2), the derivatization temperature is normal temperature, preferably 10-35 ℃;
n, in the step (2), the derivative product of the carbonyl compound is selected from one or more of formaldehyde-2, 4-dinitrophenylhydrazone, acetaldehyde-2, 4-dinitrophenylhydrazone, acetone-2, 4-dinitrophenylhydrazone, propionaldehyde-2, 4-dinitrophenylhydrazone, acrolein-2, 4-dinitrophenylhydrazone, butyraldehyde-2, 4-dinitrophenylhydrazone, 2-butanone-2, 4-dinitrophenylhydrazone and crotonaldehyde-2, 4-dinitrophenylhydrazone;
o. step (1), before leaching, further comprising the step of crushing the tobacco sample or tobacco product sample.
The method for pretreating tobacco or a tobacco product according to the first aspect of the present invention is a method for separating carbonyl compounds from tobacco or a tobacco product.
The second aspect of the invention relates to a method for detecting carbonyl compounds in tobacco or tobacco products, which comprises the steps of obtaining an extract I or an extract II according to the pretreatment method of any one of the first aspect of the invention; the method also comprises the step of measuring the derivative products of the carbonyl compounds in the extracting solution I or the extracting solution II by using a liquid chromatography-tandem mass spectrometry method.
In one embodiment of the second aspect of the present invention, further comprising the step of filtering extract I or extract II before the liquid chromatography tandem mass spectrometry measurement; preferably, filtration is performed with an organic phase filtration membrane; preferably, the pore size of the organic phase filter is 0.2-0.6. mu.m, more preferably 0.45. mu.m.
In one embodiment of the second aspect of the present invention, the assay is a quantitative assay, preferably an internal standard method quantitative assay; more preferably, the internal standard is d3Formaldehyde-2, 4 dinitrophenylhydrazone.
In one embodiment of the second aspect of the present invention, the operating conditions of the liquid chromatography comprise any one or more of the following a to d:
a. the chromatographic column is a liquid chromatographic column of the Danan Acclaim Explosive E2; preferably, the column has a size of 250mm x 4.6mm, 5 μm,
b. the temperature of the chromatographic column is 36-50 ℃, and preferably 40 ℃;
c. the sample amount is 1-10 μ L, preferably 3-7 μ L, more preferably 5 μ L;
d. the mobile phase consists of a mobile phase A and a mobile phase B, wherein the mobile phase A is acetonitrile, and the mobile phase B is water;
preferably, the elution procedure for the mobile phase is: 0 min, 60% (V/V) mobile phase a; 26 minutes, 60% (V/V) mobile phase a; 28 min, 70% (V/V) mobile phase a; 43 minutes, 70% (V/V) mobile phase a; 45 minutes, 60% (V/V) mobile phase a, hold until 50 minutes; the flow rate was always 0.6 mL/min.
In one embodiment of the second aspect of the invention, the operating conditions or parameters of the mass spectrometer include any one or more of the following 1) to 30):
1) the ion source is an electrospray ionization source;
2) the scanning mode is negative ion scanning;
3) the detection mode is multi-reaction monitoring;
4) the electrospray voltage is 3000-6000V, preferably 3500-5000V, and more preferably 4500V;
5) the ion source temperature is 400-650 ℃, preferably 450-550 ℃, and more preferably 500 ℃;
6) the pressure of the auxiliary Gas Gas1 and/or Gas2 is 40-70psi, preferably 50-60psi, more preferably 50psi or 60 psi;
7) the quantitative ion pair of the formaldehyde-2, 4-dinitrophenylhydrazone is 209.0/163.0;
8) the qualitative ion pair of the formaldehyde-2, 4-dinitrophenylhydrazone is 209.0/151.0;
9) the collision energy of the formaldehyde-2, 4-dinitrophenylhydrazone is-12V;
10) the quantitative ion pair of the acetaldehyde-2, 4-dinitrophenylhydrazone is 223.0/163.0;
11) the qualitative ion pair of the acetaldehyde-2, 4-dinitrophenylhydrazone is 223.0/181.0;
12) the collision energy of the acetaldehyde-2, 4-dinitrophenylhydrazone is-16V;
13) the quantitative ion pair of the acetone-2, 4-dinitrophenylhydrazone is 237.0/207.0;
14) the qualitative ion pair of the acetone-2, 4-dinitrophenylhydrazone is 237.0/178.0;
15) the collision energy of the acetone-2, 4-dinitrophenylhydrazone is-14V;
16) the quantitative ion pair of propionaldehyde-2, 4-dinitrophenylhydrazone is 237.0/163.0;
17) the qualitative ion pair of propionaldehyde-2, 4-dinitrophenylhydrazone is 237.0/152.0;
18) the collision energy of the propionaldehyde-2, 4-dinitrophenylhydrazone is-15V;
19) the quantitative ion pair of the acrolein-2, 4-dinitrophenylhydrazone is 235.0/158.0;
20) the qualitative ion pair of the acrolein-2, 4-dinitrophenylhydrazone is 235.0/163.0;
21) the collision energy of the acrolein-2, 4-dinitrophenylhydrazone is-16V;
22) the quantitative ion pair of the butyraldehyde-2, 4-dinitrophenylhydrazone is 251.0/163.0;
23) the qualitative ion pair of the butyraldehyde-2, 4-dinitrophenylhydrazone is 251.0/152.0;
24) the collision energy of the butyraldehyde-2, 4-dinitrophenylhydrazone is-14V;
25) the quantitative ion pair of the 2-butanone-2, 4-dinitrophenylhydrazone is 251.0/152.0;
26) the qualitative ion pair of the 2-butanone-2, 4-dinitrophenylhydrazone is 251.0/163.0;
27) the collision energy of the 2-butanone-2, 4-dinitrophenylhydrazone is-22V;
28) the quantitative ion pair of the crotonaldehyde-2, 4-dinitrophenylhydrazone is 249.0/181.0;
29) the qualitative ion pair of the crotonaldehyde-2, 4-dinitrophenylhydrazone is 249.0/172.0;
30) the collision energy of crotonaldehyde-2, 4-dinitrophenylhydrazone is-18V.
In a third aspect, the invention relates to a kit comprising,
I. acetonitrile;
II, acetonitrile solution of 2, 4-dinitrophenylhydrazine and/or 2, 4-dinitrophenylhydrazine; and
III. perchloric acid in acetonitrile.
In one embodiment of the third aspect of the invention, the kit further comprises pyridine and/or an acetonitrile solution of pyridine.
In one embodiment of the third aspect of the present invention, the kit further comprises an organic phase filter and d3Formaldehyde-2, 4 dinitrophenylhydrazone.
In one embodiment of the third aspect of the present invention, the concentration of 2, 4-dinitrophenylhydrazine in the acetonitrile solution of 2, 4-dinitrophenylhydrazine is 1 to 40mg/mL, preferably 5 to 20mg/mL, more preferably 10 mg/mL.
In one embodiment of the third aspect of the present invention, the concentration of perchloric acid in an acetonitrile solution of perchloric acid is between 0.05% and 2% (W/W), preferably between 0.1% and 1% (W/W), more preferably 0.2% (W/W).
The fourth aspect of the present invention relates to the use of the kit according to any one of the third aspects of the present invention for the pretreatment of tobacco or a tobacco product or for the detection of carbonyl compounds in tobacco or a tobacco product; preferably, the smoking article is selected from one or more of a combustible smoking article and a non-combustible smoking article, more preferably a combustible smoking article, and even more preferably a cigarette.
In one embodiment of the fourth aspect of the present invention, the carbonyl compound is selected from any one or more of formaldehyde, acetaldehyde, acetone, propionaldehyde, acrolein, butyraldehyde, 2-butanone, and crotonaldehyde.
In the present invention,
unless otherwise specified, "leaching" refers to the process of leaching soluble active ingredients from a raw material using a suitable solvent and method.
Unless otherwise specified, "tobacco" refers to solanaceae, genus Nicotiana (Nicotiana L.), annual or perennial herbs, and hobby crops. There are more than 60 species of this genus, most of which are wild species. The main cultivated species are safflower tobacco and yellow flower tobacco. Both are native to south america and are naturally occurring amphidiploids. The safflower tobacco is also called common tobacco and is a tobacco seed for producing worldwide commodities. And various types are formed due to different modulation methods, seed sources, regions and cultivation measures: flue-cured tobacco, burley tobacco, maryland tobacco, cigar tobacco, smoking, aromatic tobacco, sun-cured red tobacco, sun-cured yellow tobacco, etc. There are many cultivars in each type. The yellow tobacco is cultivated in more than the former Soviet Union and India, and is cultivated in a small amount in Xinjiang, Gansu and other places in China. The flower tobacco (N.alata Link and Otta) and the Pink blue tobacco (N.glauca Graham) in the genus are planted for appreciation.
Unless otherwise specified, "tobacco product" means a product for smoking, sucking, chewing or snuffing, produced in whole or in part from tobacco leaves as a raw material. Such tobacco products include, but are not limited to, cigarettes and chewing tobacco.
Unless otherwise specified, "smokable tobacco products" means tobacco products that require ignition and inhalation of smoke either directly, or filtered through a filter or water, most commonly a traditional cigarette that is popular worldwide.
If not specifically stated, the term "non-combustible smoking tobacco product" means that the tobacco product does not need to be burnt, reduces harmful ingredients such as tar generated by burning, does not generate second-hand smoke, and meets the requirements of consumers on tobacco products to a certain extent, and mainly comprises smokeless tobacco products, heating non-combustible cigarettes and electronic cigarettes. The preparation process of the non-smoking tobacco product is obviously different from the smoking tobacco product.
Unless otherwise specified, "carbonyl compound" refers to compounds containing carbonyl groups in the molecule and are collectively referred to as carbonyl compounds, representative two types of carbonyl compounds are ① aldehydes and ketones, ② carboxylic acids and derivatives thereof.
Unless otherwise specified, "strong acid" refers to an acid capable of almost complete ionization in aqueous solution, having a strong acidic reaction, such as HCl, H, sulfate2SO4HNO, nitric acid3Perchloric acid HCl4And the like.
Unless otherwise specified, "weak base" refers to a base that is only partially ionized in an aqueous solution, and has a weak base reaction. For example, ammonium hydroxide NH4OH, Zinc hydroxide Zn (OH)2Aluminum hydroxide Al (OH)3。
Unless otherwise specified, "internal standard method" refers to a method of plotting a standard curve in instrumental analysis. The principle is as follows: adding a certain amount of internal standard substance into standard solutions with different concentration levels, and then calculating the signal intensity ratio of the component to be detected and the internal standard substance as a longitudinal axis; and then, the quantity of the component to be measured or the ratio of the component to be measured to the internal standard quantity is calculated and used as a horizontal axis to draw a relation curve. Generally, the internal standard substance has physical and chemical properties similar to those of the component to be measured, and can be separated from the component to be measured. The internal standard method can eliminate the influence of measuring instrument, measuring condition change, sample amount change, etc., thus improving the analysis precision and being widely used for chromatographic analysis and spectral analysis.
The invention has the following beneficial effects:
1. the invention discovers that the tobacco contains carbonyl compounds for the first time, and simultaneously discovers that the tobacco products contain the carbonyl compounds for the first time.
2. The pretreatment method of the invention can extract and derivatize carbonyl compounds in tobacco or tobacco products.
3. The detection method can accurately determine and measure the carbonyl compound in the tobacco or the tobacco product, and has the advantages of low detection limit, high sensitivity and good repeatability.
4. In the detection method, pure acetonitrile is used as an extraction solvent, so that more carbonyl compounds can be detected, the accuracy is higher, the detection limit is lower, and the sensitivity is higher.
5. In the detection method, the liquid chromatogram adopts gradient elution, so that the chromatographic peak separation degree of each carbonyl compound is higher.
Drawings
In order that the present disclosure may be more readily and clearly understood, reference is now made to the following detailed description of the embodiments of the present disclosure taken in conjunction with the accompanying drawings, in which
FIG. 1 is a flow diagram of the present invention for extracting carbonyl compounds from tobacco or tobacco products;
FIG. 2 is a reaction equation of derivatization reaction of a carbonyl compound with 2, 4-dinitrophenylhydrazine according to the present invention;
FIG. 3 is a chromatogram of a standard working solution grade S3 in example 1 of the present invention;
FIG. 4 is a chromatogram of a sample solution in example 1 of the present invention;
FIG. 5 is a chromatogram of a sample solution in comparative example 2 of the present invention.
Detailed Description
Example 1
1. Materials and reagents
(1)8 carbonyl compounds, 2, 4-dinitrophenylhydrazone (purchased from carbofuran technologies, ltd): formaldehyde-2, 4-dinitrophenylhydrazone, acetaldehyde-2, 4-dinitrophenylhydrazone, propionaldehyde-2, 4-dinitrophenylhydrazone, acetone-2, 4-dinitrophenylhydrazone, acrolein-2, 4-dinitrophenylhydrazone, butyraldehyde-2, 4-dinitrophenylhydrazone, 2-butanone-2, 4-dinitrophenylhydrazone, and crotonaldehyde-2, 4-dinitrophenylhydrazone.
(2) Internal standard d 3-Formaldehyde-2, 4-dinitrophenylhydrazone, purchased from Bailingwei science and technology, Inc.
(3) Internal standard stock solution: the internal standard d 3-formaldehyde-2, 4-dinitrophenylhydrazone was prepared into an internal standard stock solution with a concentration of 50ng/mL with acetonitrile.
(4) Standard working solution: weighing 8 carbonyl compounds, namely 2, 4-dinitrophenylhydrazone standard substances and internal standard substances, adding acetonitrile to dissolve the carbonyl compounds, and preparing 5-grade standard working solution with concentration gradient. The concentration of each carbonyl compound, namely-2, 4-dinitrophenylhydrazone, in the 5-grade standard working solution is shown in Table 1, and the concentration of the internal standard substance is 50 ng/mL.
TABLE 15 concentration of 2, 4-dinitrophenylhydrazone as each carbonyl compound in the Standard working solution (ng/mL)
| Serial number | Substance(s) | S1 | S2 | S3 | S4 | S5 |
| 1 | Formaldehyde-2, 4-dinitrophenylhydrazone | 10.01 | 30.03 | 50.05 | 100.10 | 300.30 |
| 2 | Acetaldehyde-2, 4-dinitrophenylhydrazone | 7.43 | 22.30 | 37.16 | 74.33 | 222.98 |
| 3 | Acetone-2, 4-dinitrophenylhydrazone | 10.48 | 31.44 | 52.40 | 104.80 | 314.41 |
| 4 | Propanal-2, 4-dinitrophenylhydrazone | 9.80 | 29.40 | 49.00 | 97.99 | 293.97 |
| 5 | Acrolein-2, 4-dinitrophenylhydrazone | 4.86 | 14.59 | 24.32 | 48.63 | 145.90 |
| 6 | Butyraldehyde-2, 4-dinitrophenylhydrazone | 9.50 | 28.50 | 47.50 | 94.99 | 284.97 |
| 7 | 2-butanone-2, 4-dinitrophenylhydrazone | 8.42 | 25.26 | 42.11 | 84.21 | 252.63 |
| 8 | Crotonaldehyde-2, 4-dinitrophenylhydrazone | 9.46 | 28.39 | 47.32 | 94.64 | 283.93 |
The concentrations of the substances in the standard working solutions of grade 5, calculated as carbonyl compounds, are shown in Table 2. The internal standard concentration was still 50 ng/mL. If the sample concentration exceeds the standard working solution coverage, the standard working solution coverage can be properly expanded.
TABLE 2 concentration of substance in standard working solution (ng/mL) in terms of carbonyl compound
| Serial number | Substance(s) | S1 | S2 | S3 | S4 | S5 |
| 1 | Formaldehyde (I) | 1.43 | 4.29 | 7.15 | 14.30 | 42.90 |
| 2 | Acetaldehyde | 1.46 | 4.38 | 7.30 | 14.60 | 43.80 |
| 3 | Acetone (II) | 2.55 | 7.66 | 12.77 | 25.54 | 76.62 |
| 4 | Propionaldehyde | 2.39 | 7.16 | 11.94 | 23.88 | 71.64 |
| 5 | Acrolein | 1.15 | 3.46 | 5.77 | 11.54 | 34.62 |
| 6 | Butyraldehyde | 2.71 | 8.14 | 13.57 | 27.14 | 81.42 |
| 7 | 2-butanone | 2.41 | 7.22 | 12.03 | 24.06 | 72.18 |
| 8 | Crotonaldehyde | 2.65 | 7.95 | 13.25 | 26.50 | 79.50 |
2. Extraction of
The extraction of carbonyl compounds from tobacco was carried out according to the scheme of FIG. 1.
Grinding tobacco leaves 1 (Yunnan tobacco leaves) into tobacco powder, accurately weighing 1g of tobacco powder sample, placing the tobacco powder sample into a 50mL centrifuge tube, accurately adding 20mL of acetonitrile, and carrying out oscillating extraction at the extraction temperature of 25 ℃ for 30 minutes; 1mL of the primary extract after leaching was taken out and put into a 10mL volumetric flask, 200. mu.L of 10mg/mL acetonitrile solution of 2, 4-dinitrophenylhydrazine and 500. mu.L of 0.2% (W/W) acetonitrile solution of perchloric acid were added, and the mixture was mixed well and left at room temperature for 40 minutes to perform derivatization reaction (the reaction equation is shown in FIG. 2). Then, 500. mu.L of pyridine and 20. mu.L of the stock solution of the internal standard were added, the volume was adjusted to 10mL with acetonitrile, and the mixture was filtered through a 0.45 μm organic phase filter to obtain a sample solution.
3. Detection of
The 5-grade standard working solution and the sample solution were subjected to detection analysis by a high performance liquid chromatography tandem mass spectrometer (ABCIEX Qtrap 5500). The chromatogram of the standard working solution of grade S3 is shown in FIG. 3. The chromatogram of the sample solution is shown in FIG. 4.
The chromatographic operation conditions are as follows: the chromatographic column adopts a Dyan Acclaim Explosive E2 liquid chromatographic column with the specification of 250mm multiplied by 4.6mm and 5 mu m,the temperature of the chromatographic column is 40 ℃; the sample injection amount is 5 mu L; mobile phase: the mobile phase A is acetonitrile, and the mobile phase B is water; gradient elution procedure of mobile phase: 0 min, 60% (V/V) mobile phase a; 26 minutes, 60% (V/V) mobile phase a; 28 min, 70% (V/V) mobile phase a; 43 minutes, 70% (V/V) mobile phase a; 45 minutes, 60% (V/V) mobile phase a; hold until 50 minutes; the flow rate was always 0.6 mL/min.
Mass spectrum operating conditions: an ion source: electrospray ionization source (ESI); the scanning mode is as follows: scanning negative ions; the detection mode is as follows: multiple Reaction Monitoring (MRM); electrospray voltage: 4500V; ion source temperature: 500 ℃; pressure of assist Gas 1: 60 psi; pressure of assist Gas 2: 50 psi.
The quantitative ion pair, the qualitative ion pair and the Collision Energy (CE) of each carbonyl compound, 2, 4-dinitrophenylhydrazone are shown in Table 3.
TABLE 3 Mass Spectrometry parameters for carbonyl Compound 2, 4-dinitrophenylhydrazone
4. Computing
(1) According to the detection result of the medium-grade 5 standard working solution in the above 3, the concentration ratio of the carbonyl compound-2, 4-dinitrophenylhydrazone to the internal standard substance is subjected to linear regression by using the peak area ratio of the carbonyl compound-2, 4-dinitrophenylhydrazone to the internal standard substance, so as to obtain a standard curve equation, which is shown in table 4. Wherein the concentration of the carbonyl compound, 2, 4-dinitrophenylhydrazone, is given in Table 2 as the concentration of the carbonyl compound.
In the standard curve equation, the abscissa represents the ratio of the concentration of the carbonyl compound-2, 4-dinitrophenylhydrazone (in terms of the concentration of the carbonyl compound) to the concentration of the internal standard substance, and the ordinate represents the ratio of the peak area of the carbonyl compound-2, 4-dinitrophenylhydrazone to the peak area of the internal standard substance. The lowest concentration standard working solution was taken and subjected to 10 replicates to calculate the standard deviation, 3 times the standard deviation being the detection limit and 10 times the standard deviation being the quantitation limit, as shown in table 4.
TABLE 4 Standard Curve equations, detection limits and quantitation limits
| Target object | Linear regression equation | r | Detection limit (μ g/g) | Limit of quantitation (ug/g) |
| Formaldehyde (I) | y＝0.16x+0.012 | 0.9994 | 0.14 | 0.5 |
| Acetaldehyde | y＝0.18x-0.039 | 0.9998 | 0.14 | 0.5 |
| Acetone (II) | y＝0.23x-0.042 | 0.9995 | 0.26 | 0.9 |
| Propionaldehyde | y＝0.27x+0.033 | 0.9994 | 0.30 | 1.0 |
| Acrolein | y＝0.25x-0.07 | 0.9997 | 0.15 | 0.5 |
| Butyraldehyde | y＝0.29x-0.016 | 0.9996 | 0.30 | 1.0 |
| 2-butanone | y＝0.19x-0.026 | 0.9998 | 0.25 | 0.9 |
| Crotonaldehyde | y＝0.24x-0.061 | 0.9997 | 0.26 | 0.9 |
(2) According to the peak area detection results of the carbonyl compound-2, 4-dinitrophenylhydrazone and the internal standard substance in the sample solution, the concentration of the carbonyl compound in the sample solution is calculated by combining the standard curve equation of the table 4, and then the content of the carbonyl compound in the tobacco leaf is calculated according to the following formula, and the result is shown in the table 5.
Wherein,
y-carbonyl compound content in tobacco leaf, mu g/g;
c-the concentration of the carbonyl compound in the sample solution, μ g/mL;
v is the volume of the sample solution, mL;
m is the weight of tobacco leaves, g.
Example 2
The contents of carbonyl compounds in tobacco leaves 2, 3, 4 and 5 were measured by the method of example 1, and the results are shown in Table 5.
TABLE 5 content of carbonyl Compounds in different tobacco leaves (. mu.g/g)
| Formaldehyde (I) | Acetaldehyde | Acetone (II) | Propionaldehyde | Acrolein | Butyraldehyde | 2-butanone | Crotonaldehyde | |
| Tobacco leaf 1 | 2.33 | 1.46 | 2.26 | 1.03 | LOQ | LOQ | LOQ | LOQ |
| Tobacco leaf 2 | 2.46 | 1.72 | 1.93 | LOQ | LOQ | LOQ | LOQ | LOQ |
| Tobacco leaf 3 | 2.98 | 2.06 | 2.85 | 1.24 | LOQ | LOQ | LOQ | LOQ |
| Tobacco leaf 4 | 3.04 | 2.13 | 2.97 | LOQ | LOQ | LOQ | LOQ | LOQ |
| Tobacco leaf 5 | 1.96 | 1.43 | 2.31 | LOQ | LOQ | LOQ | LOQ | LOQ |
Note: LOQ indicates no detection.
As is clear from Table 5, the method of the present invention has a low detection limit for carbonyl compounds and a high sensitivity.
Example 3
Based on example 1, 8 carbonyl compounds were spiked at three levels, high, medium and low, respectively, and the spiked samples were averaged in triplicate to determine recovery and Reproducibility (RSD), with the results shown in table 6.
TABLE 6 recovery and reproducibility of the method
As can be seen from Table 6, the recovery rates of the 8 carbonyl compounds by the method of the present invention are all over 92%, the detection accuracy is high, and the repeatability is good.
Comparative example 1
The procedure of example 1 was repeated except that the pure acetonitrile was replaced with mixed solvents of acetonitrile and water at volume ratios of 1:1 and 2:1, respectively, to extract tobacco leaf 1.
The measured content of each carbonyl compound in the tobacco leaves 1 is shown in table 7.
TABLE 7
Note: LOQ indicates no detection.
Comparing the detection results of the tobacco leaves 1 in the table 7 and the table 5, the method has higher detection accuracy, more measurable carbonyl compounds, lower detection limit and higher sensitivity when the method is leached by pure acetonitrile compared with the method of leaching by using the mixed solvent of acetonitrile and water.
Comparative example 2
Isocratic elution was carried out using 60% (V/V) acetonitrile + 40% (V/V) water as a mobile phase, and the rest of the operation was carried out with reference to example 1, to obtain a chromatogram of the sample solution as shown in FIG. 5.
As can be seen from a comparison of FIGS. 4-5, the gradient elution used in the process of the present invention provides a better separation of the chromatographic peaks for each carbonyl compound than an equivalent elution with 60% (V/V) acetonitrile + 40% (V/V) water as the mobile phase.
It should be understood that the above examples are only for clarity of illustration and are not intended to limit the embodiments. Other variations and modifications will be apparent to persons skilled in the art in light of the above description. And are neither required nor exhaustive of all embodiments. And obvious variations or modifications therefrom are within the scope of the invention.
Claims (64)
1. A method for detecting carbonyl compounds in tobacco or tobacco products comprises the steps of carrying out pretreatment on the tobacco or the tobacco products to obtain an extracting solution I containing carbonyl compound derivatization products; the method also comprises the step of determining the derivative product of the carbonyl compound in the extracting solution I by using a liquid chromatography-tandem mass spectrometry method;
wherein, the tobacco or tobacco products are pretreated by the following steps:
(1) leaching a tobacco sample or a tobacco product sample by using acetonitrile to obtain primary extract containing carbonyl compounds;
(2) adding 2, 4-dinitrophenylhydrazine and/or 2, 4-dinitrophenylhydrazine solution and strong acid solution into the primary extract for derivatization, and separating to obtain an extract I containing a carbonyl compound derivatization product;
and, the tobacco product is a product for smoking, sucking, chewing or snuffing, produced wholly or partly from tobacco leaves as raw material; the carbonyl compound is selected from one or more of formaldehyde, acetaldehyde, acetone, propionaldehyde, acrolein, butyraldehyde, 2-butanone and crotonaldehyde.
2. A method for detecting carbonyl compounds in tobacco or tobacco products comprises the steps of carrying out pretreatment on the tobacco or the tobacco products to obtain an extracting solution II containing carbonyl compound derivatization products; the method also comprises the step of determining the derivative product of the carbonyl compound in the extracting solution II by using a liquid chromatography-tandem mass spectrometry method;
wherein, the tobacco or tobacco products are pretreated by the following steps:
(1) leaching a tobacco sample or a tobacco product sample by using acetonitrile to obtain primary extract containing carbonyl compounds;
(2) adding 2, 4-dinitrophenylhydrazine and/or 2, 4-dinitrophenylhydrazine solution and strong acid solution into the primary extract for derivatization, and separating to obtain an extract I containing a carbonyl compound derivatization product;
(3) adding weak base or weak base solution into the extracting solution I, and mixing to obtain an extracting solution II;
and, the tobacco product is a product for smoking, sucking, chewing or snuffing, produced wholly or partly from tobacco leaves as raw material; the carbonyl compound is selected from one or more of formaldehyde, acetaldehyde, acetone, propionaldehyde, acrolein, butyraldehyde, 2-butanone and crotonaldehyde.
3. The detection method according to claim 2, wherein in the step (3), the weak base is selected from one or more of pyridine and quinoline.
4. The detection method according to claim 2, wherein in the step (3), the weak base is pyridine.
5. The detection method according to claim 2, wherein in the step (3), the weak base solution is one or more selected from an acetonitrile solution of pyridine and an acetonitrile solution of quinoline.
6. The detection method according to claim 2, wherein in the step (3), the weak base solution is an acetonitrile solution of pyridine.
7. The detecting method according to claim 4 or 6, wherein in the step (3), the amount of the acetonitrile solution corresponding to pyridine in terms of pyridine per ml of the initial extract is 300-700. mu.L, or the amount of pyridine in terms of pyridine per ml of the initial extract is 300-700. mu.L.
8. The detecting method according to claim 4 or 6, wherein in the step (3), the amount of the acetonitrile solution corresponding to pyridine in terms of pyridine per ml of the initial extract is 400-600. mu.L, or the amount of pyridine in terms of pyridine per ml of the initial extract is 400-600. mu.L.
9. The detection method according to claim 4 or 6, wherein in the step (3), the amount of the acetonitrile solution containing pyridine per ml of the initial extract solution is 500. mu.L, or the amount of pyridine per ml of the initial extract solution is 500. mu.L.
10. The detection method according to claim 1 or 2, characterized by any one or more of the following a to N:
A. the tobacco product is selected from one or more of a combustible smoking tobacco product and a non-combustible smoking tobacco product;
B. in the step (1), the leaching material-liquid ratio is 1 (10-50) (g/mL);
C. in the step (1), the leaching is carried out under oscillation conditions;
D. in the step (1), the leaching time is 10-60 minutes;
E. in the step (1), the leaching temperature is normal temperature;
F. in the step (2), the addition amount of 2, 4-dinitrophenylhydrazine solution calculated by 2, 4-dinitrophenylhydrazine in each milliliter of primary extract is 0.2-5mg, or the addition amount of 2, 4-dinitrophenylhydrazine in each milliliter of primary extract is 0.2-5 mg;
G. in the step (2), the dosage of the strong acid solution in each milliliter of the primary extract is 300-;
H. in the step (2), the 2, 4-dinitrophenylhydrazine solution is acetonitrile solution of 2, 4-dinitrophenylhydrazine;
I. in the step (2), the strong acid solution is acetonitrile solution of strong acid;
J. in the step (2), the strong acid solution is selected from one or more of perchloric acid solution and hydrofluoric acid solution;
K. in the step (2), the derivatization time is 20-60 minutes;
l, in the step (2), the derivatization temperature is normal temperature;
m, in the step (2), the derivative product of the carbonyl compound is selected from one or more of formaldehyde-2, 4-dinitrophenylhydrazone, acetaldehyde-2, 4-dinitrophenylhydrazone, acetone-2, 4-dinitrophenylhydrazone, propionaldehyde-2, 4-dinitrophenylhydrazone, acrolein-2, 4-dinitrophenylhydrazone, butyraldehyde-2, 4-dinitrophenylhydrazone, 2-butanone-2, 4-dinitrophenylhydrazone and crotonaldehyde-2, 4-dinitrophenylhydrazone;
and N, before leaching, the step (1) further comprises the step of crushing the tobacco sample or the tobacco product sample.
11. The method of detecting according to claim 10, wherein in item A, the tobacco product is a smokable tobacco product.
12. The method according to claim 10, wherein in item A, the tobacco product is a cigarette.
13. The detection method according to claim 10, wherein in item B, in the step (1), the leaching feed-liquor ratio is 1 (10-30) (g/mL).
14. The assay of claim 10, wherein in step (1) in item B, the feed to liquor ratio of the leach is 1:20 (g/mL).
15. The assay of claim 10 wherein in step (D), the leaching time is from 20 to 50 minutes in step (1).
16. The assay of claim 10 wherein in step (1) the leaching time is 20 minutes, 30 minutes or 50 minutes.
17. The assay of claim 10 wherein in step (E), the temperature of the leach is 5-40 ℃ in step (1).
18. The assay of claim 10 wherein in step (E), the temperature of the leach is 10-35 ℃ in step (1).
19. The assay of claim 10 wherein in step (E), the temperature of the leach is 10 ℃, 25 ℃ or 30 ℃ in step (1).
20. The detecting method according to claim 10, wherein in the item F, in the step (2), the amount of the 2, 4-dinitrophenylhydrazine solution added per ml of the primary extract solution in terms of 2, 4-dinitrophenylhydrazine is 1 to 3mg, or the amount of the 2, 4-dinitrophenylhydrazine added per ml of the primary extract solution is 1 to 3 mg.
21. The method according to claim 10, wherein in the item F, in the step (2), the amount of the 2, 4-dinitrophenylhydrazine solution per ml of the primary extract solution is 2mg in terms of 2, 4-dinitrophenylhydrazine, or the amount of the 2, 4-dinitrophenylhydrazine solution per ml of the primary extract solution is 2 mg.
22. The detecting method according to claim 10, wherein in the item G, the amount of the strong acid solution used in step (2) is 400-600. mu.L per ml of the initial extract.
23. The method according to claim 10, wherein the amount of the strong acid solution used in the step (2) is 500. mu.L/ml of the primary extract in the item G.
24. The method according to claim 10, wherein in the item H, the concentration of 2, 4-dinitrophenylhydrazine in the acetonitrile solution of 2, 4-dinitrophenylhydrazine in the step (2) is 1 to 40 mg/mL.
25. The method according to claim 10, wherein in the item H, the concentration of 2, 4-dinitrophenylhydrazine in the acetonitrile solution of 2, 4-dinitrophenylhydrazine in the step (2) is 5 to 20 mg/mL.
26. The method according to claim 10, wherein in the step (2), the concentration of 2, 4-dinitrophenylhydrazine in the acetonitrile solution of 2, 4-dinitrophenylhydrazine is 10 mg/mL.
27. The method according to claim 10, wherein in the step (2), the concentration of the strong acid in the acetonitrile solution of the strong acid is 0.05% to 2% (W/W).
28. The method according to claim 10, wherein in the step (2), the concentration of the strong acid in the acetonitrile solution of the strong acid is 0.1 to 1% (W/W).
29. The method according to claim 10, wherein in the step (2), the concentration of the strong acid in the acetonitrile solution of the strong acid is 0.2% (W/W).
30. The assay of claim 10 wherein in item K, the derivatization time is from 30 to 50 minutes.
31. The assay of claim 10 wherein in item K, the derivatization time is 40 minutes.
32. The method of claim 10, wherein the temperature of derivatization is 10 ℃ to 35 ℃.
33. The detection method according to claim 1 or 2, further comprising a step of filtering the extract I or the extract II before the measurement by liquid chromatography tandem mass spectrometry.
34. The detection method according to claim 33, wherein the filtration is performed using an organic phase filtration membrane.
35. The detection method according to claim 34, wherein the pore size of the organic phase filtration membrane is 0.2 to 0.6 μm.
36. The detection method according to claim 34, wherein the pore size of the organic phase filter is 0.45 μm.
37. The assay of claim 1 or 2 wherein the assay is a quantitative assay.
38. The assay of claim 37 wherein the assay is an internal standard quantitative assay.
39. The detection method according to claim 38, wherein the internal standard is d 3-formaldehyde-2, 4-dinitrophenylhydrazone.
40. The detection method according to claim 1 or 2, wherein the operating conditions of the liquid chromatography include any one or more of the following a to d:
a. the chromatographic column is a liquid chromatographic column of the Danan Acclaim Explosive E2;
b. the temperature of the chromatographic column is 36-50 ℃;
c. the sample amount is 1-10 mu L;
d. the mobile phase consists of a mobile phase A and a mobile phase B, wherein the mobile phase A is acetonitrile, and the mobile phase B is water.
41. The detection method according to claim 40, wherein in item a, the size of the chromatographic column is 250mm x 4.6mm, 5 μm,
42. a test method according to claim 40 wherein in item b the temperature of the column is 40 ℃.
43. The test method according to claim 40, wherein the amount of the sample is 3 to 7. mu.L in the item c.
44. The method according to claim 40, wherein the amount of the sample is 5. mu.L in item c.
45. The detection method according to claim 40, wherein in item d, the elution procedure of the mobile phase is as follows: 0 min, 60% (V/V) mobile phase a; 26 minutes, 60% (V/V) mobile phase a; 28 min, 70% (V/V) mobile phase a; 43 minutes, 70% (V/V) mobile phase a; 45 minutes, 60% (V/V) mobile phase a, hold until 50 minutes; the flow rate was always 0.6 mL/min.
46. The detection method of claim 1 or 2, wherein the operating conditions or parameters of the mass spectrometer comprise any one or more of the following 1) to 30):
1) the ion source is an electrospray ionization source;
2) the scanning mode is negative ion scanning;
3) the detection mode is multi-reaction monitoring;
4) the electrospray voltage is 3000-6000V;
5) the temperature of the ion source is 400-650 ℃;
6) the pressure of the auxiliary Gas Gas1 and/or Gas2 is 40-70 psi;
7) the quantitative ion pair of the formaldehyde-2, 4-dinitrophenylhydrazone is 209.0/163.0;
8) the qualitative ion pair of the formaldehyde-2, 4-dinitrophenylhydrazone is 209.0/151.0;
9) the collision energy of the formaldehyde-2, 4-dinitrophenylhydrazone is-12V;
10) the quantitative ion pair of the acetaldehyde-2, 4-dinitrophenylhydrazone is 223.0/163.0;
11) the qualitative ion pair of the acetaldehyde-2, 4-dinitrophenylhydrazone is 223.0/181.0;
12) the collision energy of the acetaldehyde-2, 4-dinitrophenylhydrazone is-16V;
13) the quantitative ion pair of the acetone-2, 4-dinitrophenylhydrazone is 237.0/207.0;
14) the qualitative ion pair of the acetone-2, 4-dinitrophenylhydrazone is 237.0/178.0;
15) the collision energy of the acetone-2, 4-dinitrophenylhydrazone is-14V;
16) the quantitative ion pair of propionaldehyde-2, 4-dinitrophenylhydrazone is 237.0/163.0;
17) the qualitative ion pair of propionaldehyde-2, 4-dinitrophenylhydrazone is 237.0/152.0;
18) the collision energy of the propionaldehyde-2, 4-dinitrophenylhydrazone is-15V;
19) the quantitative ion pair of the acrolein-2, 4-dinitrophenylhydrazone is 235.0/158.0;
20) the qualitative ion pair of the acrolein-2, 4-dinitrophenylhydrazone is 235.0/163.0;
21) the collision energy of the acrolein-2, 4-dinitrophenylhydrazone is-16V;
22) the quantitative ion pair of the butyraldehyde-2, 4-dinitrophenylhydrazone is 251.0/163.0;
23) the qualitative ion pair of the butyraldehyde-2, 4-dinitrophenylhydrazone is 251.0/152.0;
24) the collision energy of the butyraldehyde-2, 4-dinitrophenylhydrazone is-14V;
25) the quantitative ion pair of the 2-butanone-2, 4-dinitrophenylhydrazone is 251.0/152.0;
26) the qualitative ion pair of the 2-butanone-2, 4-dinitrophenylhydrazone is 251.0/163.0;
27) the collision energy of the 2-butanone-2, 4-dinitrophenylhydrazone is-22V;
28) the quantitative ion pair of the crotonaldehyde-2, 4-dinitrophenylhydrazone is 249.0/181.0;
29) the qualitative ion pair of the crotonaldehyde-2, 4-dinitrophenylhydrazone is 249.0/172.0;
30) the collision energy of crotonaldehyde-2, 4-dinitrophenylhydrazone is-18V.
47. The detection method according to claim 46, wherein the electrospray voltage is 3500-5000V in item 4).
48. The detection method according to claim 46, item 4), wherein the electrospray voltage is 4500V.
49. The detection method according to claim 46, wherein in item 5), the ion source temperature is 450 ℃ -550 ℃.
50. The detection method according to claim 46, wherein in item 5), the ion source temperature is 500 ℃.
51. The method according to claim 46, wherein in item 6), the pressure of the auxiliary Gas Gas1 and/or Gas2 is 50-60 psi.
52. The method according to claim 46, wherein in item 6), the pressure of the auxiliary Gas Gas1 and/or Gas2 is 50psi or 60 psi.
53. The use of a kit for detecting carbonyl compounds in tobacco or tobacco products;
wherein the kit comprises:
I. acetonitrile;
II, acetonitrile solution of 2, 4-dinitrophenylhydrazine and/or 2, 4-dinitrophenylhydrazine; and
III, acetonitrile solution of perchloric acid;
and, the tobacco product is a product for smoking, sucking, chewing or snuffing, produced wholly or partly from tobacco leaves as raw material; the carbonyl compound is selected from one or more of formaldehyde, acetaldehyde, acetone, propionaldehyde, acrolein, butyraldehyde, 2-butanone and crotonaldehyde.
54. The use of claim 53, wherein the kit further comprises pyridine and/or an acetonitrile solution of pyridine.
55. The use of claim 53, wherein the kit further comprises an organic phase filter and d3Formaldehyde-2, 4 dinitrophenylhydrazone.
56. The use according to claim 53, wherein in the kit the concentration of 2, 4-dinitrophenylhydrazine in acetonitrile solution of 2, 4-dinitrophenylhydrazine is 1-40 mg/mL.
57. The use according to claim 53, wherein in the kit the concentration of 2, 4-dinitrophenylhydrazine in acetonitrile solution of 2, 4-dinitrophenylhydrazine is 5-20 mg/mL.
58. The use according to claim 53, wherein the concentration of 2, 4-dinitrophenylhydrazine in acetonitrile solution of 2, 4-dinitrophenylhydrazine in the kit is 10 mg/mL.
59. The use according to claim 53, wherein the concentration of perchloric acid in an acetonitrile solution of perchloric acid in the kit is between 0.05% and 2% (W/W).
60. The use according to claim 53, wherein the concentration of perchloric acid in an acetonitrile solution of perchloric acid in the kit is between 0.1% and 1% (W/W).
61. The use according to claim 53, wherein the concentration of perchloric acid in an acetonitrile solution of perchloric acid in the kit is 0.2% (W/W).
62. The use of claim 53, wherein the tobacco product is selected from one or more of a smokable tobacco product and a non-smokable tobacco product.
63. The use of claim 53, wherein the tobacco product is a smokable tobacco product.
64. The use according to claim 53, wherein the tobacco product is a cigarette.
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| CN107966518B (en) * | 2017-12-07 | 2020-08-11 | 中国烟草总公司郑州烟草研究院 | Treatment method of smokeless tobacco product and determination method of small molecular aldehyde in smokeless tobacco product |
| CN109142566A (en) * | 2018-07-30 | 2019-01-04 | 苏州中汽检测技术服务有限公司 | The separation method and measuring method of automotive upholstery volatile matter methacrylaldehyde and acetone |
| KR102695238B1 (en) | 2023-06-25 | 2024-08-14 | 차이나 타바코 윈난 인더스트리얼 컴퍼니 리미티드 | Identification method of tobacco carbonyl components by non-targeted mass spectrometry |
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