The pooled odds ratio (OR) from four case-control studies of lung cancer associated with menthol vs. nonmenthol cigarettes was 0.93 (95% confidence interval 0.82–1.05) (4
). Menthol was not associated with an increased risk for other cancers (30
). Menthol is not pyrolized during tobacco combustion (http://tobaccodocuments.org/product_design/504331475-1477.html
), and does not induce tumors in animals via intraperitoneal injection (32
) or drinking water supplementation (24
). Menthol may not be a direct carcinogen or co-carcinogen, but it causes several pharmacologic and physiological effects that may affect cancer risk (33
). Despite the lack of increased risk in the case-control studies, the effect size and statistical power to detect differences of one type of cigarette relative to another is likely to be small. However even a small relative effect of menthol could cause substantial numbers of cancers since millions of Americans smoke menthol cigarettes.
Consequently, biomarker and mechanistic studies are seemingly important tools for indirectly assessing possible risks. One of the limitations of studying the effects of menthol has been separating its effects from race. The majority of blacks who smoke prefer menthol, and studies of smoking topography or cotinine levels may have controlled for race but did not examine the race-specific effects of menthol (34
). The current study examined the race-specific levels of several tobacco smoke biomarkers between menthol and nonmenthol smokers. We found no significant differences in mean cotinine and thiocyanate levels.
A limitation of the study is that it included subjects who smoked at least five cigarettes per day. The mean cotinine levels were higher than that reported for NHANES data (37
), where about 25% of smokers reported smoking 7 or fewer cigarettes daily. The current study therefore does not address possible differences in biomarker levels by menthol status among very light smokers. We observed no significant differences in the urinary levels of the lung carcinogen NNAL between menthol and nonmenthol smokers in race-specific analyses, although the number of blacks who smoked nonmenthol cigarettes was relatively small and the differences in NNAL by menthol status was not significant. Assuming the same effect size and variances that were observed, the study would have required about 6 times the number of nonmenthol black smokers to detect significant differences. However, if the effect size is similar to that what was observed in whites, much smaller numbers would be needed.
Menthol has been studied in relation to increased addiction to nicotine. In a two-week randomized pharmacokinetic crossover study of 14 smokers who switched from menthol to nonmenthol or nonmenthol to menthol, menthol was on average unrelated to blood levels of unlabelled nicotine (14
). The effect of menthol on behavioral measures of nicotine dependence such as the FTND was negligible in adolescent smokers (38
). The current study showed similar findings in adult smokers. It has been suggested that the amount of time between waking up and smoking the first cigarette is possibly a better measure of nicotine dependence than smoking amount or the FTND since it was shown to be a better predictor of plasma cotinine concentrations than daily smoking amount in adults (39
). However, in our data, the correlations of cotinine (both plasma and urinary) were very similar between cigarettes per day, time since waking and FTND. We did observe that menthol smokers were more likely to smoke cigarettes within 30 minutes after waking up than nonmenthol smokers, which is similar to findings in the adolescent smoker study, but differs from the Community Intervention Trial for Smoking Cessation (COMMIT) which found a longer time to first cigarette after waking for menthol smokers (16
). The literature is inconsistent in this area and more studies might be helpful to understand this relationship. Menthol may inhibit quitting (19
), although other large scale studies show similar quit rates between menthol and nonmenthol smokers (17
Another limitation is the interpretation of these analyses is that menthol content varies by cigarette brand, and younger smokers have been reported to smoke menthol brands with lower menthol content than older menthol smokers (40
). In the present study, cigarettes were classified as menthol or nonmenthol, and therefore the magnitude of the effects (or lack thereof) associated with menthol reported here may not necessarily be generalizeable to all smokers.
We previously reported no racial differences in the ratio of NNAL-Gluc/NNAL levels in men, although in a subgroup of nine women who had a very high NNAL-Gluc/NNAL ratio (≥6), eight were white (29
). We examined the cigarette brands of these women and found that only one subject was a menthol smoker so it is possible that the racial differences in NNAL glucuronidation in women could be attributed to menthol.
In the current study, menthol smokers had lower urinary ratios of NNAL-Gluc/NNAL than nonmenthol smokers, and menthol inhibited the glucuronidation of both NNAL-N-Gluc and NNAL-O-Gluc formation in vitro
. These effects might be mediated by UGT2B10, which is the major enzyme involved in the N
-glucuronidation of NNAL (41
), and by UGT2B17 and UGT2B7 which are the major enzymes involved in NNAL O
). While UGT2B7 genotype was not associated with human liver microsomal glucuronidation activity against menthol (44
), in vitro studies have demonstrated that UGT2B7 is active against menthol and could be potentially inhibited by this substrate (45
). The potential exists that menthol could therefore act as a substrate and inhibitor of other UGTs as well. Studies by Benowitz et al have demonstrated that menthol significantly inhibits nicotine-glucuronide formation in vivo
). Since UGT2B10 is the major enzyme responsible for the N
-glucuronidation of both nicotine and NNAL, it is likely that UGT2B10 may be the target for the inhibitory effects of menthol on nicotine and NNAL glucuronidation activity in vivo
. Studies examining the interaction between menthol and NNAL/nicotine glucuronidation in cells specifically over-expressing UGTs 2B7, 2B10 and 2B17 are currently planned. The major limitation of in vitro studies that attempt to identify the underlying mechanisms in epidemiologic data is that it is difficult to mimic in vivo conditions experimentally. In particular the human liver microsomes are not equivalent to ER in intact cells and do not interact similarly with other cellular components and vascular supply, and nutrients. The levels of exposure in the in vitro model may not reflect in vivo exposure. Despite these limitations, cellular pathways are often activated similarly in both in vitro and in vivo systems and human liver microsomes are commonly used to screen for drug interactions in pharmacologic research (47
In summary, these data indicate that menthol is not associated with a higher exposure to tobacco smoke carcinogens but the findings on nicotine dependence are inconclusive. Menthol may not be more hazardous than other cigarette formulations for most smokers, although it cannot be ruled out at this time that some menthol smokers are possibly at increased risk of lung cancer due to selective inhibition of UGT enzymes.