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A Comparative In Vitro Toxicological Screening of a Closed-End Heated Tobacco Product * Cover

A Comparative In Vitro Toxicological Screening of a Closed-End Heated Tobacco Product *

Contributions to Tobacco & Nicotine Research
Volume 32 (2023): Issue 4 (December 2023)
By: Zhenyu Niu,  Qi Zhang,  Meng Nie,  Lili Fu,  Junjun Luo,  Xinduo Li,  Jie Zhou,  Jiakai Liu,  Zheng Song,  Chuan Liu,  Ke Zhang,  Jingjing Liang,  Xingtao Jiang,  Zehong Wu,  Bin Li and  Te Xu  
Open Access
|Dec 2023

Figures & Tables

Figure 1.

Schematic diagram comparing air flow path for the closed-end HTP-A (a: the test product) and an open-end HTB-B (b: see Ref. 8, 9 )
Schematic diagram comparing air flow path for the closed-end HTP-A (a: the test product) and an open-end HTB-B (b: see Ref. 8, 9 )

Figure 2.

Cell viability in NRU assay of HTP-A and HTP-B aerosol, and 3R4F smoke. Cell viability of CHO cells was determined by NRU assay following 24 h exposures of DMSO (a) or HTP-A (b), HTP-B (c) and 3R4F (d). The data represent the mean ± SD (n = 3) (###P < 0.01).
Cell viability in NRU assay of HTP-A and HTP-B aerosol, and 3R4F smoke. Cell viability of CHO cells was determined by NRU assay following 24 h exposures of DMSO (a) or HTP-A (b), HTP-B (c) and 3R4F (d). The data represent the mean ± SD (n = 3) (###P < 0.01).

Salmonella typhimurium strains characteristics, source and positive controls_

StrainsMutationAntibiotic resistancePositive controls
Without S9 (μg/plate)With S9 (μg/plate)
TA98His D3052Ampicillin2-Nitrofluorene (4 μg/plate)2-Amlinofluorene (10 μg/plate)
TA100His G46Ampicillin4-Nitroquinoline N-oxide (0.5 μg/plate)2-Amlinofluorene (10 μg/plate)

In vitro mammalian cell micronucleus test_

Positive controlsUltimate density
Cyclophosphamide A (short-term + S9)0.2 μg/mL
Mitomycin C (short-term - S9)1.0 μg/mL
Mitomycin C (long-term - S9)0.5 μg/mL

Revertant colonies of Salmonella typhimurium obtained following exposure to different doses of TPM from HTP-A and HTP-B_

Dose (μg/plate)TA98TA100
HTP-AHTP-BHTP-AHTP-B
−S9
NC38.0 ± 2.438.0 ± 2.4115.3 ± 3.8115.3 ± 3.8
025.7 ± 4.025.7 ± 4.0121.7 ± 6.0121.7 ± 6.0
5024.0 ± 3.730.3 ± 4.0124.0 ± 4.1108.0 ± 9.9
50035.0 ± 3.728.7 ± 4.1120.3 ± 8.7105.0 ± 8.3
100030.7 ± 2.433.3 ± 5.7114.3 ± 6.2129.3 ± 5.3
200030.7 ± 3.832.7 ± 4.5120.7 ± 7.9125.3 ± 3.1
PC899.0 ± 24.0899.0 ± 24.02719.7 ± 55.52719.7 ± 55.5
     
+S9
NC60.0 ± 5.960.0 ± 5.9156.3 ± 2.9156.3 ± 2.9
051.0 ± 5.151.0 ± 5.1107.0 ± 6.4107.0 ± 6.4
5048.3 ± 7.648.3 ± 3.1116.0 ± 19.0100.0 ± 4.3
50048.7 ± 7.456.0 ± 4.3112.0 ± 10.7100.7 ± 5.4
100049.0 ± 7.140.3 ± 5.2113.7 ± 12.4100.7 ± 9.2
200051.0 ± 2.841.7 ± 7.094.0 ± 16.8105.3 ± 1.2
PC2437.3 ± 106.02437.3 ± 106.0781.7 ± 95.7781.7 ± 95.7

Selected toxicant yields from two HTPs and 3R4F_

AnalyteUnitHTP-AHTP-B3R4F
Nicotinemg/stick0.47 ± 0.010.981.05
Total particulate mattermg/stick29.06 ± 1.3630.1524.23
 mg/mg nicotine61.83 ± 2.9030.7723.08
Carbon monoxidemg/stick0.19 ± 0.010.2524.69
 mg/mg nicotine0.40 ± 0.020.2623.52
Formaldehydeμg/stick1.08 ± 0.184.46 ± 0.5626.60 ± 2.14
 μg/mg nicotine2.30 ± 0.384.55 ± 0.5725.33 ± 2.04
Acetaldehydeμg/stick123.5 ± 19.53172.42 ± 6.10722.47 ± 36.11
 μg/mg nicotine262.77 ± 41.55175.94 ± 6.22688.07 ± 34.39
Acroleinμg/stick4.78 ± 0.9110.90 ± 0.1871.92 ± 2.84
 μg/mg nicotine1.02 ± 1.9411.12 ± 0.1868.50 ± 2.70
1,3-Butadieneμg/stickNQNQ8.13 ± 0.17
 μg/mg nicotineNQNQ7.74 ± 0.16
Benzeneμg/stick0.17 ± 0.000.36 ± 0.0549.95 ± 1.54
 μg/mg nicotine0.36 ± 0.000.37± 0.0547.57 ± 1.47
Benzo(a)pyreneng/stickNQNQ24.00 ± 0.64
 ng/mg nicotineNQNQ22.86 ± 0.61
NNKng/stick6.61 ± 0.865.32 ± 0.89281.00 ± 16
 ng/mg nicotine14.06 ± 0.835.43 ± 0.91267.62 ± 15
NNNng/stick1.79 ± 0.268.37 ± 0.55163.43 ± 11.19
 ng/mg nicotine3.81 ± 0.558.54 ± 0.56155.65 ± 10.66

Revertant colonies of Salmonella typhimurium obtained following exposure to different doses of TPM from 3R4F_

Dose (μg/plate)TA98TA100
−S9+S9−S9+S9
NC38.0 ± 2.460.0 ± 5.9115.3 ± 3.8156.3 ± 2.9
025.7 ± 4.051.0 ± 5.1121.7 ± 6.0107.0 ± 6.4
5038.3 ± 8.287.7 ± 5.2125.7 ± 8.5105.0 ± 3.7
10028.3 ± 7.5124.3 ± 5.4*127.3 ± 7.6115.3 ± 7.5
25032.7 ± 6.1237.7 ± 11.8*143.3 ± 8.3133.0 ± 4.1
50034.7 ± 2.1233.7 ± 33.0*137.7 ± 11.1171.7 ± 13.0
PC899.0 ± 24.02437.3 ± 106.02719.7 ± 55.5781.7 ± 95.7

Analytical procedures used for HTP aerosol and cigarette smoke chemical analyses_

AnalyteBrief descriptionReference
NicotineBy gas chromatography with flame ionization detectionISO 10315 (12)
COBy nondispersive infrared photometryISO 8454 (13)
Polycyclic aromatic hydrocarbonsExtracted by hexane and analyzed by GC-MSCRM 91 (14)
N-nitrosaminesExtracted by acetic acid and analyzed by HPLC-MS/MSCRM 75 (15)
Volatile organic compoundsBy GC-MS using simultaneous trapping of adsorbent cartridges in a Cambridge filter without cryogenic impingerWHO SOP 09 (16)
Carbonyl compoundsBy HPLC, using simultaneous trapping of adsorbent cartridges in a Cambridge filter without cryogenic impingeWHO SOP 08 (17)

Cell genotoxicity of different TPM fractions of HTP-A and HTP-B, and 3R4F_

SampleConcentration (μg/mL)MN frequencies (%)
3h + 21h -S93h + 21h +S924h + 0h -S9
Vehicle111.49 ± 0.231.52 ± 0.541.08 ± 0.08
HTP-A
751.37 ± 0.331.28 ± 0.041.29 ± 0.08
1501.08 ± 0.371.20 ± 0.071.19 ± 0.04
3001.05 ± 0.071.21 ± 0.011.40 ± 0.08
6001.15 ± 0.171.10 ± 0.101.30 ± 0.01
HTP-B
751.11 ± 0.021.20 ± 0.141.26 ± 0.16
1501.33 ± 0.221.07 ± 0.181.28 ± 0.11
3001.36 ± 0.301.28 ± 0.011.33 ± 0.13
6001.69 ± 0.301.29 ± 0.211.32 ± 0.11
3R4F
14.32.03 ± 0.212.16 ± 0.922.53 ± 0.05
28.53.21 ± 1.022.46 ± 0.982.85 ± 0.20
57.03.93 ± 1.342.31 ± 1.292.65 ± 0.05
114.04.49 ± 1.582.67 ± 1.493.03 ± 0.30
PC-5.68 ± 0.295.08 ± 0.556.09 ± 0.37
NC-1.40 ± 0.301.32 ± 0.301.20 ± 0.11

Overview of test product specifications_ The heating device of HTP-A is branded as Mr_ Yeah and the tobacco sticks carry the brand FARSTAR_ HTP-B consists of an IQOS™ branded heating device and TEREA branded tobacco sticks_

ParameterHTP-AHTP-BCombustible cigarette
Commercial product (name)Mr. YeahIQOS™3R4F
Puffing air flow designClosed-endOpen-endOpen
Heater designExternal heatingCentral heatingCombustion
Maximum heating temp. (°C)260350/
Puff n° under HCI regime8128
Tobacco stick-length (mm)454584
Tobacco stick-weight (mg)721.70664.301035.00
Tobacco stick-diameter (mm)7.367.187.98
DOI: https://doi.org/10.2478/cttr-2023-0018 | Journal eISSN: 2719-9509
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Language: English
Page range: 146 - 156
Submitted on: Jun 5, 2023
Accepted on: Nov 23, 2023
Published on: Dec 16, 2023
Published by: Institut für Tabakforschung GmbH
In partnership with: Paradigm Publishing Services
Publication frequency: 4 issues per year
Keywords:
Heated tobacco product,
aerosol,
chemistry,
in vitro,
toxicological assessment
Related subjects:
General interest,
Life sciences,
Life sciences, other,
Physics,
Physics, other

© 2023 Zhenyu Niu, Qi Zhang, Meng Nie, Lili Fu, Junjun Luo, Xinduo Li, Jie Zhou, Jiakai Liu, Zheng Song, Chuan Liu, Ke Zhang, Jingjing Liang, Xingtao Jiang, Zehong Wu, Bin Li, Te Xu, published by Institut für Tabakforschung GmbH
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