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The Regulation of Microorganism on the Formation of TSNAs in Tobacco
Congming Zou
Yunnan Academy of Tobacco Agricultural SciencesKunming, China
Zhenyuan Xia, Liping Lei, Wei Huang, Gaokun. Zhao, Yan Jin, Qun Wu
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Yunnan TobaccoProduces around 40% tobacco in China, and 20% flue-cured tobacco in this world !
800000 tons/year
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Oriental Tobacco#1 China
Burley Tobaccowas # 1 China
Sun-cured Yellow Tobacco# 1 China
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Introduction We care about TSNAs…….
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Introduction 1、Tobacco TSNAs mainly form in curing and storage process
2、Tobacco TSNAs formation have a relationship with Microbes
3、Can we use tobacco endophytic bacteria and fungi to control TSNAs formation?
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1、Select microbes (tobacco endophytic bacteria and fungi) which could feed on nicotine
2、TSNAs substrate solution incubation study
3、Tobacco short air-curing study
4、Cigarette leaves incubation study
Experiment Design
5、Tobacco long air-curing study
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1、 Select microbes which could feed on nicotine
122/869 tobacco endophytic bacteria and fungi can successfully feed on nicotine and TSNAs
There microbes mainly belong to Firmicutes,Actinobacteria, Proteobacteria and Bacteroidetes.
16S rDNA sequence method
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2、TSNAs substrate solution incubation study
Treatment NNN(ng/g)
NAT(ng/g)
NAB(ng/g)
NNK(ng/g)
TSNAs(ng/g)
Compared with CK
(%)Ck 344 226 42 21 629 0J45 866 199 32 76 1174 +86.6 J54 1391 315 36 37 1779 +182.8 J70 729 458 41 12 1241 +97.3
05-5402 341 100 37 11 489 -22.3 05-101 381 69 33 13 496 -21.1
Solution from Burley tobacco
Inoculate different microbes
Incubate 3-5 days
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3、Tobacco short air-curing study
Burley tobacco
Half leaves method
Air-cured 7 days
Spray and not spray
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3、Burley tobacco curing study
Treatment NNN(ng/g)
NAT(ng/g)
NAB(ng/g)
NNK(ng/g)
TSNAs(ng/g)
Compared with CK
(%)J45ck 19.25 29.66 2.39 5.31 56.61 0J45 18.84 29.31 3.07 4.44 55.66 -1.7
J54ck 27.85 41.54 2.28 5.44 77.11 0J54 37.14 58.96 2.83 16.12 115.05 +49.2
J70ck 24.43 32.55 2.69 6.01 65.68 0J70 27.09 31.27 2.61 5.21 66.18 +0.8
05-5402ck 26.41 49.69 2.28 6.09 84.47 0 05-5402 21.19 43.38 1.09 4.18 69.84 -30.2 05-101ck 22.39 15.95 1.5 4.41 44.25 0 05-101 20.97 17.54 1.41 3.69 43.61 -1.4
2017
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4、Cigarette leaves incubation study
Cigarette leave from Zhongnanhai
Inoculate different microbes
Incubate 5 days
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4、Cigarette leaves incubation study
Treatment NNN(ng/g)
NAT(ng/g)
NAB(ng/g)
NNK(ng/g)
TSNAs(ng/g)
Compared with CK(%)
CK 5148.31 1404.29 44.55 196.5 6793.65 0.0
J45 5361.76 1414.24 47.33 207.43 7030.82 +3.5
J54 5640.51 1403.84 52.04 209.9 7305.93 +7.5
J70 4824.2 1361.41 36.66 177.64 6399.76 -5.8
05-5402 4225.38 1268.09 39.37 185.63 5706.81 -16.0
05-101 5002.29 1436.27 45.08 211.89 6695.53 -1.4
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Summary
Class Study 1 Study 2 Study 3
J45 Pseudomonas +86.6 -1.7 +3.5
J54 Pseudomonas +182.8 +49.2 +7.5
J70 Pseudomonas +97.3 +0.8 -5.8
05-5402 Bacillus -22.3 -30.2 -16.0
05-101 Paenibacillus -21.1 -1.4 -1.4
Relatively stable microorganisms
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5、Tobacco long air-curing study
Air-cured 50 days
Spray and not spray
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5、Tobacco long air-curing study
Treatment NNN (ng/g)
NAT(ng/g)
NAB (ng/g)
NNK (ng/g)
TSNAs (ng/g)
Compared with CK(%)
CK 567.4 783.5 21.9 162.5 1535.3 0
J45 270.2 501.1 13.9 117.6 902.8 -41.2*J54 222.5 462.7 12.4 122.2 819.8 -46.6*
J70 393.8 654.2 18.8 132.6 1199.3 -21.9*
05-5402 294.1 584.0 16.5 138.9 1033.5 -32.7*
05-101 435.2 838.5 25.2 160.6 1459.4 -4.9
2017
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Question?
Microbes which can successfully feed on nicotine and TSNAs
could potentially degrade the formation of TNSAs. However,
it might depend on microbe’s incubation time, substitute
substrate availability and environment conditions.
2017
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123_
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6、Future action ( for flue-cured tobacco flue-curing)
Pool with selected bacteria Ready for flue-curing
2017
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123_
Zou
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gmin
g.pd
fT
SR
C20
17(7
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Doc
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r-re
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