• 한국연초학회지
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• 제27권1호
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• pp.51-58
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• 2005

The objective of this study was to evaluate the effect of cigarette filter on in vitro cytotoxicity of cigarette mainstream smoke from the cigarette. In this work, we used 3 types of cigarettes included non-filtered 2R4F cigarette, cellulose acetate-filtered 2R4F cigarette, and carbon dual-filtered 2R4F cigarette which was made from original 2R4F by replacing with an acetate filter containing carbon. The cytotoxicity of both the cigarette smoke condensate (CSC), which was collected in Cambridge filter pad, and the gas/vapor phase (GVP), which was bubbled through in phosphate-buffered saline in a gas-washing bottle, was determined using a neutral red uptake assay with CHO-K1 cells. With regard to cytotoxicity when calculated on an equal puff basis, the cytotoxicity of CSC from the filtered cigarettes was lower than that of the non filtered cigarette. Also, $EC_{50}$ vlaue of GVP from carbon filter cigarette was 40.9 puff/L, indicating the cytotoxicity to be $20\%$ lower than that of the CA filter cigarette. The cytotoxicity of the GVP was correlated to the several vapor phase components (formaldehyde, acetaldehyde, acetone, acrolein, crotonaldehyde and MEK). In conclusion, carbon filter, which significantly reduced the amount of carbonyl compounds in mainstream cigarette smoke, results in significant reductions in the cytotoxicity potential of the smoke.

• 한국연초학회지
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• 제29권2호
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• pp.110-117
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• 2007

In vitro toxicity tests such as cytotoxicity, mutagenicity and genotoxicity assay are useful for evaluating the relative toxicity of smoke or smoke condensates obtained from different cigarette configurations. A major disadvantage of these tests is relatively time-consuming, complicated and expensive. Recently, a cell-free glutathione consumption assay (GCA) as a rapid and simple screening method for the toxicity assessment of smoke has been reported by Cahours et al. (CORESTA, 2006). This study was carried out to assess the GCA application capable of predicting the toxicity of gas/vapor phase (GVP) of cigarette smoke and to identify individual compounds responsible for the glutathione (GSH) consumption in smoke. Each GVPs from 2R4F, standard cigarette, carbon filter cigarette (ExC) and new carbon filter cigarette (ExN), test cigarettes were collected by automatic smoking machine and evaluated the relative toxicity by GCA and neutral red uptake (NRU) assay. Toxic compounds existed in smoke were also chosen, relative toxicities of these compounds were screened by using two methods and compared individually. The overall order of toxicity by GCA was 2R4F > ExC > ExN, which was consistent with the result of Neutral Red Uptake assay. The levels of carbonyl compounds of ExN were lower than those of 2R4F and ExC, indicating that GSH consumption was associated with carbonyl compound yields. A major toxicant under current study is acrolein, which contributed to more than half of the GSH consumption. Collectively, the toxicity of GVP determined by GCA method may be mainly attributed to acrolein.

• 한국연초학회지
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• 제29권1호
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• pp.30-40
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• 2007

The objective of this study was to investigate the contribution of various smoke constituents to the toxicological activity of total particulate matter(TPM) or the gas/vapor phase(GVP). These components included phenol compounds, aromatic amines, polycyclic aromatic hydrocarbons, heterocyclic amines, and carbonyl compounds. The mutagenic and cytotoxic potencies were assessed using the Salmonella mutagenicity assay with S. typimurium TA98 strain and the neutral red uptake cytotoxicity assay(NRU) with BALB/c 3T3 fibroblast cells, respectively. The Salmonella mutagenicity test showed that heterocyclic amines exhibited significantly higher levels of toxicity compared to other smoke constituents. Among them, 2-amino-3,4-dimethylimidazo[4,5-f]quinoline(MeIQ) was shown the most mutagenic compound with a specific mutagenicity of $7.9{\times}10^5\;revertants/{\mu}g$. An analysis of the possible contribution revealed that MeIQ account for only 0.85% of the 2R4F-TPM mutagenicity in TA98. NRU data demonstrated that high cytotoxic activity was obtained for hydroquinone, formaldehyde, and acrolein. Based on the results of the present study, the contribution of acrolein to the cytotoxicity of the GVP fraction was calculated as 61%. Thus, a large proportion of the cytotoxic activity of this complex mixture, cigarette smoke gas phase, can be attributed to the acrolein.

• 한국연초학회지
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• 제30권1호
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• pp.14-24
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• 2008

The objective of this study was to determine the effect of smoking conditions on the in vitro toxicological activity of mainstream smoke. The 2R4F reference cigarette was machine-smoked by International Organization for Standardization(ISO) and Canadian Intense(CI) conditions. Smoke was analysed for chemical composition and in vitro toxicity. The cytotoxic potencies of both the total particulate matter(TPM), which were collected in Cambridge filter pad, and gas/vapor phase(GVP), which was bubbled through in phosphate-buffer saline in a gas-washing bottle, were assessed neutral red up take assay with chinese hamster ovary(CHO) cells. The assessment for genotoxicity of TPMs generated under ISO and CI conditions was determined using Salmonella mutagenicity assay and in vitro micronucleus assay. When calculated on an equal TPM basis, in vitro toxicity of TPM obtained under CI condition was decreased compared to TPM generated under ISO condition. The results of chemical composition analyses revealed that the lower toxicological activity under CI condition than that of ISO condition could be explained by the decreased in the contents of phenols, N-nitrosoamines and aromatic amines of TPM on an equal TPM basis.

• 한국연초학회지
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• 제32권1호
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• pp.19-27
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• 2010

The aim of this study is to compare the sensitivity of both two NCI-H292 and A549 cell types to acute inflammatory responses induced by cigarette smoke. For this, we treated two kinds of smoke fractions derived from 2R4F reference cigarettes: total particulate matter(TPM) collected onto a Cambridge filter pad and gas/vapor phase(GVP) prepared by bubbling through in buffer solution. When we measured cellular cytotoxicity by neutral red uptake assay after treatment for 24 hours, TPM and GVP induced cytotoxic effect in a dose-dependent manner in the range of 10-$100{\mu}g$/mL and 60-$300 {\mu}g$/mL., respectively, in both cell types without any cellular difference. Additionally, when we examined acute inflammatory responses by analyzing cytokines secreted into culture media including tumor necrosis factor-$\alpha$ (TNF-$\alpha$), interleukin-8(IL-8), and transforming growth factor-$\alpha$(TGF-$\alpha$) as well as matrix metalloproteinase-1(MMP-1), the treatment with smoke fractions increased those marker proteins in a dose-dependent manner in NCI-H292. Meanwhile, in A549 cells only MMP-1 was observed to be increased in a dose-dependent fashion. Collectively, our data indicate that NCI-H292 cell type is more sensitive to cigarette smoke-induced inflammatory response than A549 cells. This suggests that NCI-H292 could be useful as an in vitro evaluation tool to assess harmful effects of cigarette smoke.

• 한국연초학회지
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• 제31권2호
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• pp.75-84
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• 2009

The objective of this study was to investigate the effect of nitrogen compounds such as protein on the chemical composition and toxicity of cigarette mainstream smoke. BSA protein was treated into the tobacco leaf of original 2R4F cigarette at 1~4 % level. The studies were performed which included a bacterial mutagenicity assay and a mammalian cell cytotoxicity assay for total particulate matter(TPM), and glutathione(GSH) consumption assay for gas/vapor phase(GVP) and determination of smoke chemical constitute. Cigarettes treated with protein were observed dose-dependent increase in yield of volatiles, semi-volatiles and aromatic amines compared with control cigarette. However, carbonyl compounds such as acrolein was lower than that of control cigarette when calculated on an equal TPM basis. The cytotoxicity of TPM obtained from the protein-added cigarettes was not different from that of control cigarette. However, the mutagenicity of the TPM from protein-treated cigarettes(1~4 %) was up to 10-27 % higher than that of control. On the other hand, toxicity of GVP from protein-treated cigarette(4 %) was significantly decreased compared with control cigarette. An overall assessment of our data suggests that nitrogen compounds such as protein should be important for the chemical composition and biological activity of cigarette mainstream smoke.