The formation of tobacco-specific nitrosamines from the major tobacco alkaloid nicotine was examined. Detached leaf tobacco was fed either [2'-14C]nicotine or [2'-14C]nornicotine and air cured. The cured leaf was then analyzed for [2'-14C]N'-nitrosonornicotine ([2'-14C]NNN). The yield of [2'-14C]NNN was 0.007% from nornicotine and 0.009% from nicotine. Because the ratio of nicotine to nornicotine in conventional nicotine-type tobacco is 20-100:1, nicotine is considered to be the major precursor for the carcinogen NNN in tobacco. The formation of other nitrosamines from nicotine in vitro was then studied. Reaction of nicotine with NaNO2 gave rise to NNN, as well as to two other nitrosamines, 4-(N-methyl-N-nitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and 4-(N-methyl-N-nitrosamino)-4-(3-pyridyl)butanal (NNA). Analysis of market products revealed the presence of NNK (0.6-24 microgram/g) in chewing tobacco and snuff. The tumorigenic activity of NNN, NNK, and NNA in strain A mice was studied. NNK induced more lung adenomas per mouse than did NNN, whereas NNA was less active than NNN. In addition, two cases of undifferentiated carcinoma of the salivary glands occurred in the NNN experimental groups.

This research was partially supported by a grant from the Brown Hazen Fund of the Research Corporation. Names of companies or commercial products are given solely for the purpose of providing specific information; their mention does not imply recommendation or endorsement by the U.S. Department of Agriculture over others not mentioned.

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Various factors which might contribute to the presence of or reflect on the formation of N'-nitrosonornicotine (NNN) or related nitrosamines in tobacco were studied. Using high-performance liquid chromatography (HPLC), the E- and Z-isomers of NNN were clearly separated and the rates of interconversion were determined. The E/Z ratio in tobacco approximated that observed in solution at similar pH and temperature. The influence of curing and stalk position on NNN levels in tobacco was determined. NNN was not detected in green tobacco but was detected in air-cured leaves from the same crop. No significant correlation was observed among stalk position and NNN levels in one variety of bright tobacco. Since NNN may derive from nicotine, two new nitrosamines, a nitrosamino ketone and a nitrosamino aldehyde, which could theoretically arise from nicotine, were synthesized. Analysis of tobacco for these components is currently in progress

Test plants were grown within a chamber enriched with radon-222 in the atmosphere, in tobacco fields with different sources of phosphate-containing fertilizer, and in culture containing lead-210 in the nutrient solution. Harvested leaves were subjected to three curing conditions. The major portion of the lead-210 in the plant was probably absorbed through the roots. Airborne radon 222 and its daughters contributed much less to the plant's content of lead-210 and of polonium-210. The stage of leaf development and the methods used to cure the leaf affected the final amount of polonium-210 in tobacco leaf.

The preceding paper (11) Craig counterculrrent apparatus (2) was used. In the course of this development the separation of a mixture prepared from nicotine, nornicotine, and anabasine was investigated by means of this apparatus,

Tobacco plants (Nicotiana tabacum L.) were grown on long or short photoperiods followed by 5 minutes of red or far red radiation each day. Plants that received 16-hour photoperiods had a significantly higher concentration of total alkaloids and total phenolics than those that received 8-hour photoperiods. Significantly higher total alkaloid content was found in plants that received red rather than far red radiation last each day. Within each photoperiod, plants that received far red had higher concentrations of soluble phenols, particularly of chlorogenic acid. The interactions among these variables upon alkaloid and phenolic contents are discussed.were trimmed to a 2-cm "brush," and leaves at lower stalk positions were removed so that the characteristics of new growth were a function of the assigned treatment. The "conditioned" test plants were then set in aerated nutrient solution (4) and placed under either 8-or 16-hr photoperiods at 25 C under 2000 ft-c of illumination from cool white VHO fluorescent lamps. Roots of test plants were protected from light exposure by placing aluminum foil over the nutrient solution containers.Thirty plants were used for each photoperiod treatment. At the end of their respective photoperiods, each day, half of the test plants were irradiated with 5 min of R, while the other half received 5 min of FR. The intensity of R and FR radiation was adjusted to approximately 360 j,W-2 over the wave length bands of 600 to 700 and 700 to 770 nm, respectively, as described in an earlier report (8).Plants on 16-hr photoperiods were treated for 18 days. Those on 8-hr photoperiods received 20 days of treatment. This 2-day Photocontrol of plant growth and development has been widely studied. Pronounced developmental differences associated with red and far red light, acting through the phytochrome system, occur when such irradiations follow relatively short photoperiods and are followed by long uninterrupted nights (8). Tobacco plants irradiated briefly with FR2 at the end of short days develop long stems with light green leaves relative to those on plants irradiated with R at the end of the day (8). Light of defined spectra was observed to affect levels of phenolic components of many plants (15,16). However, the exact relationships between photoinduced biosynthesis of the compounds and photoinduced growth and development are yet to be established (3). The objective of the research reported herein was to determine whether levels of alkaloids and phenolic compounds of tobacco were influenced by length of photoperiod and/or by manipulation of the phytochrome system with R and FR at the end of the day. MATERIALS AND METHODSTobacco (Nicotiana tabacum L. cv. TI-204-E) seedlings were started and grown for about 6 weeks at 28 C under 14-hr, 1600 ft-c photoperiods from cool white fluorescent lamps. The seedlings were started in vermiculite and subirrigated with halfstrength Hoagland's nutrient solution (4).Photoperiod

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