Endogenous formation of N'-nitrosonornicotine in F344 rats in the presence of some antioxidants and grape seed extract

N'-Nitrosonornicotine (NNN) is one of the most abundant strong carcinogens in unburned tobacco and cigarette smoke and is classified by the International Agency for Research on Cancer as carcinogenic to humans. Human exposure to NNN mainly occurs upon use of tobacco products. It is also possible that additional amounts of NNN are formed endogenously. The goal of this study was to evaluate the inhibitory effect of some antioxidants, including ascorbic acid and grape seed extract (GSE), on endogenous NNN formation in rats treated with nornicotine and sodium nitrite by gavage twice daily for 3 days. The study included four groups of rats: (1) negative control group A, to which no chemical was administered; (2) negative control group B, treated with nornicotine alone (2.5 micromol per gavage); (3) positive control group, to which both nornicotine (2.5 micromol per gavage) and sodium nitrite (7.5 micromol per gavage) were administered; and (4) rats treated with nornicotine (2.5 micromol per gavage), inhibitor (7.5 or 37.5 micromol per gavage), and sodium nitrite (7.5 micromol per gavage). The mean (+/-SD) total amount of NNN in the 3-day urine of rats treated with both nornicotine and sodium nitrite was 4.78 +/- 2.88 nmol. The order of inhibition of endogenous NNN formation in rats at the molar ratio [nitrite]:[inhibitor] 1:5 was as follows: ascorbic acid (91%) > dihydroxyfumaric acid (86%) approximately catechin (85%) > resveratrol (no inhibition). Treatment of rats with grape seed extract did not produce statistically significant inhibition of endogenous nornicotine nitrosation. This is the first study that demonstrates endogenous NNN formation in rats treated with nornicotine and sodium nitrite and effective inhibition of this process by ascorbic acid, dixydroxyfumaric acid, and catechin.     

New sources of dietary myosmine uptake from cereals,fruits, vegetables,and milk

Myosmine has been regarded as a specific tobacco alkaloid until investigations pointed out that nuts and nut products constitute a significant source of myosmine. In the present study it is shown that the occurrence of myosmine is widespread throughout a large number of plant families. Using a method for extraction practicable for all examined foods, quantitative analysis through internal standard addition showed nanograms per gram amounts. Positively tested edibles were staple foods such as maize, rice, wheat flour, millet, potato, and milk and also cocoa, popcorn, tomato, carrot, pineapple, kiwi, and apples. No myosmine was detectable in other vegetables and fruits such as lettuce, spinach, cucumber, onion, banana, tangerines, and grapes. Myosmine is easily nitrosated giving rise to a DNA adduct identical to the esophageal tobacco carcinogen N-nitrosonornicotine. Therefore, the role of dietary myosmine in esophageal adenocarcinoma should be further investigated.     

Identification of oxidation products of (-)-epigallocatechin gallate and (-)-epigallocatechin with H(2)O(2)

(-)-Epigallocatechin gallate (EGCG) and (-)-epigallocatechin (EGC) are two important antioxidants in tea. They also display some antitumor activities, and these activities are believed to be mainly due to their antioxidative effects. However, the specific mechanisms of antioxidant action of tea catechins remain unclear. In this study are isolated and identified two novel reaction products of EGCG and one product of EGC when they were reacted separately with H(2)O(2). These products are formed by the oxidation and decarboxylation of the A ring in the catechin molecule. This study provides unequivocal proof that the A ring of EGCG and EGC may also be an antioxidant site. This study also indicates an additional reaction pathway for the oxidation chemistry of tea catechins.     

Comparative analysis of tobacco-specific nitrosamines and total N-nitroso compounds in moldovan cigarette tobacco

While previous studies have evaluated levels of tobacco-specific nitrosamines (TSNA) and total N-nitroso compounds (NOC) in tobacco, there are no reports in the literature on TSNA and total NOC in the same tobacco products. We compared levels of TSNA, total NOC, and NOC precursors (NOCP) in tobacco of cigarettes purchased in Moldova and in some tobacco types commonly used for the manufacturing of Moldovan cigarettes. Cigarette tobaccos included those from non-Moldovan, traditional Moldovan, and blended Moldovan cigarettes. The results demonstrate that tobacco of non-Moldovan cigarettes contains higher TSNA and NOC levels (mean, 16 and 63 nmol/g tobacco, n = 6) than that of Moldovan cigarettes (mean, 5 and 23 nmol/g tobacco, n = 25). TSNA and NOC levels were also generally higher in tobacco of blended than in traditional Moldovan cigarettes. NOCP levels in Moldovan and non-Moldovan cigarette tobacco were similar as follows: 29000 +/- 30000 and 33000 +/- 28000 nmol/g tobacco (mean +/- SD). Total NOC were strongly correlated with total TSNA levels (r = 0.66; P < 0.0001). These findings demonstrate that current technologies involved in the manufacture of some blended cigarettes create conditions that favor N-nitrosation of alkaloids and other tobacco constituents.     

Relative quantification of N(epsilon)-(Carboxymethyl)lysine,imidazolone A,and the Amadori product in glycated lysozyme by MALDI-TOF mass spectrometry

The nonenzymatic glycation of proteins by reducing sugars, also known as the Maillard reaction, has received increasing recognition from nutritional science and medical research. In this study, we applied matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) to perform relative and simultaneous quantification of the Amadori product, which is an early glycation product, and of N(epsilon)-(carboxymethyl)lysine and imidazolone A, two important advanced glycation end products. Therefore, native lysozyme was incubated with d-glucose for increasing periods of time (1, 4, 8, and 16 weeks) in phosphate-buffered saline pH 7.8 at 50 degrees C. After enzymatic digestion with endoproteinase Glu-C, the N-terminal peptide fragment (m/z 838; amino acid sequence KVFGRCE) and the C-terminal peptide fragment (m/z 1202; amino acid sequence VQAWIRGCRL) were used for relative quantification of the three Maillard products. Amadori product, N(epsilon)-(carboxymethyl)lysine, and imidazolone A were the main glycation products formed under these conditions. Their formation was dependent on glucose concentration and reaction time. The kinetics were similar to those obtained by competitive ELISA, an established method for quantification of N(epsilon)-(carboxymethyl)lysine and imidazolone A. Inhibition experiments showed that coincubation with N(alpha)-acetylargine suppressed formation of imidazolone A but not of the Amadori product or N(epsilon)-(carboxymethyl)lysine. The presence of N(alpha)-acetyllysine resulted in the inhibition of lysine modifications but in higher concentrations of imidazolone A. o-Phenylenediamine decreased the yield of the Amadori product and completely inhibited the formation of N(epsilon)-(carboxymethyl)lysine and imidazolone A. MALDI-TOF-MS proved to be a new analytical tool for the simultaneous, relative quantification of specific products of the Maillard reaction. For the first time, kinetic data of defined products on specific sites of glycated protein could be measured. This characterizes MALDI-TOF-MS as a valuable method for monitoring the Maillard reaction in the course of food processing.     

Potential nitrite scavengers as inhibitors of the formation of N-nitrosamines in solution and tobacco matrix systems

The ability of 20 compounds, all but one tobacco constituents, to inhibit the formation of tobacco-specific N-nitrosamines (TSNA) was investigated in buffer and detergent solution and in tobacco midrib and lamina systems. In solution at pH 5.5, ascorbic acid and the phenolic acids caffeic and ferulic acid were the most potent inhibitors of the reaction between nornicotine and nitrite, with nearly complete inhibition at molar ratios test compound/nitrite > 1:1. Also, cysteine > dihydrocaffeic acid > protocatechuic acid approximately catechin acted as strong inhibitors with >90% inhibition at a ratio of 3:1. Lower inhibitions were observed with chlorogenic acid > p-coumaric acid > sclareol > serine. Rutin showed an inhibition of 34% at a ratio of 0.1:1. Sclareol, alanine, proline, and serine did not significantly affect the N-nitrosonornicotine (NNN) formation. alpha-Tocopherol and glutathione enhanced NNN formation at pH 5.5 but were inhibitors at pH 3. Cinnamic acid, vanillic acid, eugenol, and esculin enhanced NNN formation. Increased NNN formation was also observed for dihydrocaffeic acid, chlorogenic acid, protocatechuic acid, and catechin at a less-than-equimolar ratio of test compound to nitrite. The tobacco matrix experiments were performed with air-cured, ground tobacco midrib and lamina. Caffeic acid, ferulic acid, dihydrocaffeic acid and catechin were potent inhibitors of the formation of TSNA in the midrib as well as in the lamina. Also protocatechuic acid, glutathione, ascorbic acid, p-coumaric acid, chlorogenic acid and cysteine were inhibitors, while alpha-tocopherol and rutin inhibited the reaction in the midrib but not in the lamina. Cinnamic acid, vanillic acid, eugenol, alanine, proline and serine showed small effects only. The molar ratio of secondary alkaloid(s)/nitrite in the test systems were 0.1:1 (solution), approximately 0.25:1 (midrib), and approximately 1:1 (lamina) and is most likely the major contributor to the observed order of inhibition potency (solution > midrib > lamina) of the test compounds. The vicinal phenolic hydroxyl groups of polyphenols and the simultaneous presence of a phenol group and an olefinic bond in hydroxycinnamic acids were the most characteristic structural elements of the potent inhibitors.     

L-(-)-malic acid production by Saccharomyces spp. during the alcoholic fermentation of wine (1)

In an attempt to increase the acidity of wine by biological means, malate-producing yeasts were selected from a collection of 282 strains isolated in different parts of Spain. Only 4% of these strains (all of which belonged to Saccharomyces cerevisiae) produced l-(-)-malic acid in the range of 0.5-1 g/L. This was formed between days 2 and 6 of alcoholic fermentation, reaching a maximum on days 3 and 4; the concentration remained stable from day 7. Malic acid production was favored by temperatures in the 18-25 degrees C range and by musts with a high pH and low concentrations of sugar, initial malic acid, and yeast-assimilable nitrogen. Oxaloacetic acid, a precursor of malic acid, had no influence on malate production. The precursors pyruvic and fumaric acid did, however, have a significant effect on the production of this acid in some strains. No direct relation between pyruvate and malate metabolism was observed.     

Synthesis of [3-(3-chloro-4-fluorophenyl)-2-oxo-3,4-dihydro-2H-2lambda5- benzo[e][1,3,2]oxazaphosphinin-2-yl]-(aryl/alkyl) methanols and their bioactivity on sugarcane smut

Synthesis of some new substituted [3-(3-chloro-4-fluorophenyl)-2-oxo-3,4-dihydro-2H-2lambda(5)-benzo[e][1,3,2]oxazaphosphinin-2-yl]-(aryl/alkyl)methanols (7a-k) based on the Pudovick reaction was accomplished in the presence of niobium pentoxide (Nb(2)O(5)) without using an external chiral ligand. Nb(2)O(5) appears to form the metal complex intermediate catalyst system (6) by reacting with 3-(3-chloro-4-fluoro-phenyl)-3,4-dihydrobenzo[e][1,3,2]oxazaphosphinine-2-oxide (4), which not only directs the Pudovick addition reactions of aldehyde but also increases the yields and purity of the products. These compounds exhibited a lethal effect on whip smut of sugarcane and were degraded in the environment in the presence of bacteria and fungi to nontoxic phosphate residues that act as possible plant nutrients. Thus, a new class of benzooxazaphosphininyl methanol derivatives that act in synergy both as antipathogens and as plant nutrients in the environment have been discovered.     

N-(1-deoxy-D-fructos-1-yl) fumonisin B(1), the initial reaction product of fumonisin B(1) and D-glucose

Incubation of fumonisin B(1) and D-glucose in aqueous solutions resulted in the formation of N-(1-deoxy-D-fructos-1-yl) fumonisin B(1) in addition to the previously reported N-(carboxymethyl) fumonisin B(1). N-(1-Deoxy-D-fructos-1-yl) fumonisin B(1) is the first stable product formed after the Amadori rearrangement of the Schiff base formed by the reaction of the primary amine of fumonisin B(1) and the aldehyde group of D-glucose. N-(1-Deoxy-D-fructos-1-yl) fumonisin B(1) was synthesized by reacting fumonisin B(1) with an excess of D-glucose in methanol and heating for 6 h at 64 degrees C. It was purified using C(18) and strong cation exchange solid-phase extraction cartridges and characterized by nuclear magnetic resonance and liquid chromatography-mass spectrometry. Subsequently, N,N-dimethylformamide was found to be a better reaction solvent, requiring reaction for only 2-3 h at 64 degrees C and eliminating the formation of methyl esters. Alkaline hydrolysis of N-(1-deoxy-D-fructos-1-yl) fumonisin B(1) gave a mixture of hydrolyzed fumonisin B(1) and hydrolyzed N-(carboxymethyl) fumonisin B(1).     

Analysis of tobacco-specific nitrosamines in Moldovan cigarette tobacco

Tobacco-specific nitrosamines (TSNA) are among the most important and abundant strongly carcinogenic agents in unburned tobacco. It has been established that 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) induces lung tumors in rodents independent of the route of administration. N'-Nitrosonornicotine (NNN) causes tumors of the esophagus and nasal cavity in rats, lung in mice, and respiratory tract in hamsters. Although the manufacturing of cigarettes is an important domain of Moldovan industry, there are no reports in the literature on TSNA analysis in Moldovan tobacco. The main purpose of the present study was an initial evaluation of TSNA levels in Moldovan cigarette tobacco. Eighteen brands of Moldovan cigarettes, representing 78% of all brands produced in Moldova, were analyzed. Four TSNA-NNN, NNK, N'-nitrosoanatabine (NAT), and N'-nitrosoanabasine (NAB)-were analyzed by gas chromatography with nitrosamine selective detection (GC-TEA). Levels of TSNA in most Moldovan cigarettes were substantially lower than in American brands. Mean levels of NNN in three commercial American brands were 3.32 +/- 0.88 (SD) microg/g as compared to 0.579 +/- 0.548 microg/g, range 0.093-2.09 microg/g (N = 18), in the cigarettes produced in Moldova. For NNK and NAT, mean levels in the American brands were 1.57 +/- 0.178 and 1.99 +/- 0.579 microg/g, respectively, while the corresponding values for Moldovan cigarettes were 0.193 +/- 0.089, range 0.104-0.484 microg/g, and 0.160 +/- 0.114 microg/g, range 0.055-0.481 microg/g. The highest levels of NNN-1.10-2.09 microg/g-were observed in "American type" cigarettes manufactured from high-quality tobacco. The results of this study should be useful in heightening the awareness of the dangers of smoking in Moldova and can be envisioned as the initial step in the control of tobacco-related cancer in this republic.     

Enzymatic synthesis of a selective inhibitor for alpha-glucosidases: alpha-acarviosinyl-(1-->9)-3-alpha-D-glucopyranosylpropen

Here, we describe the enzymatic synthesis of novel inhibitors using acarviosine-glucose as a donor and 3-alpha-D-glucopyranosylpropen (alphaGP) as an acceptor. Maltogenic amylase from Thermus sp. (ThMA) catalyzed the transglycosylation of the acarviosine moiety to alphaGP. The two major reaction products were isolated using chromatographies. Structural analyses revealed that acarviosine was transferred to either C-7 or C-9 of the alphaGP, which correspond to C-4 and C-6 of glucose. Both inhibited rat intestine alpha-glucosidase competitively but displayed a mixed-type inhibition mode against human pancreatic alpha-amylase. The alpha-acarviosinyl-(1-->7)-3-alpha-D-glucopyranosylpropen showed weaker inhibition potency than acarbose against both alpha-glycosidases. In contrast, the alpha-acarviosinyl-(1-->9)-3-alpha-D-glucopyranosylpropen exhibited a 3.0-fold improved inhibition potency against rat intestine alpha-glucosidase with 0.3-fold inhibition potency against human pancreatic alpha-amylase relative to acarbose. In conclusion, alpha-acarviosinyl-(1-->9)-3-alpha-D-glucopyranosylpropen is a novel alpha-glucosidase-selective inhibitor with 10-fold enhanced selectivity toward alpha-glucosidase over alpha-amylase relative to acarbose, and it could be applied as a potent hypoglycemic agent.     

Defektverbindung und fremdionen-dotierung bei der ausfällung von eisen(II)- und eisen(III)- phosphaten

The products obtained by precipitation of iron(III)-phosphates in the pH-range from 2 to 13.35 differ in their composition from the ones reported in the literature, e.g. FePO₄2H₂O or FeH₂(OH)₂PO₄, except at low pH values.The molar ratio of 1 : 1 for P and Fe is disturbed to a significant extent and foreign ions are also incorporated as integral parts of the precipitation products.The change of the P/Fe ratio within the pH-range from 2 to 11 is the result of continuous substitution of OH^? for PO₄^? ions. This exchange may be explained as a transition between phosphate- and hydroxide-“Defektserie” (reference is to sets of compounds which form partial rather than complete isomorphous series), whereby the critical composition and the critical precipitation product are defined by the inflection point of the exchange curve.During the hydrothermal recrystallization of amorphous precipitation products, crystalline iron phosphate (strengite) is formed at low pH values, and crystalline iron hydroxide is formed at high pH values. The precipitation products formed at intermediate pH values have a heterogeneous composition and fail to crystallize under these conditions.     

Thermal decomposition of 5-(hydroxymethyl)-2-furaldehyde (HMF) and its further transformations in the presence of glycine

Thermal decomposition of HMF has been so far studied indirectly through carbohydrate degradation reactions assuming HMF as the main product. Such studies, however, do not necessarily generate relevant information on HMF decomposition because many other products are generated simultaneously. Direct thermal decomposition using different concentrations of HMF in silica gel was studied using pyrolysis-GC-MS. Undiluted HMF generated four peaks corresponding to 5-methylfurfural, 2,5-furandicarboxaldehdye, HMF, and a major unknown peak at retention time of 20.73 min. The diluted HMF in silica gel (15-fold) generated only the first three peaks. The generation of the unknown peak was dependent on the concentration of HMF, indicating the possibility of a dimeric structure; furthermore, when HMF was generated from [U-13C6]glucose in the reaction mixture, the highest mass in the spectrum of the unknown peak showed the incorporation of 11 carbon atoms from the glucose. Thermal decomposition studies of HMF have also indicated that in the absence of amino acids it can mainly dimerize and the initially formed dimer can degrade to generate 5-methylfurfural and 2,5-furandicarboxaldehyde. On the other hand, thermal degradation of HMF in the presence of glycine generated Schiff base adducts of HMF, 5-methylfurfural, and 2,5-furandicarboxaldehdye in addition to 2-acetyl-5-methylfuran and a newly discovered adduct, 5-[(dimethylamino)methyl]-2-furanmethanol.     

Characterization of fumonisin B(1)-glucose reaction kinetics and products

The reaction of fumonisin B(1) with the reducing sugar D-glucose can block the primary amine group of fumonisin B(1) and may detoxify this mycotoxin. A method to separate hundred milligram quantities of fumonisin B(1)-glucose reaction products from the excess D-glucose with a reversed-phase C(18) cartridge was developed. Mass spectrometry revealed that there were four primary products in this chain reaction when fumonisin B(1) was heated with D-glucose at 65 degrees C for 48 h: N-methyl-fumonisin B(1), N-carboxymethyl-fumonisin B(1), N-(3-hydroxyacetonyl)-fumonisin B(1), and N-(2-hydroxy, 2-carboxyethyl)-fumonisin B(1). The N-(1-deoxy-D-fructos-1-yl) fumonisin B(1) (fumonisin B(1)-glucose Schiff's base) was detected by mass spectrometry when fumonisin B(1) was heated with D-glucose at 60 degrees C. The nonenzymatic browning reaction of fumonisin B(1) with excess D-glucose followed apparent first-order kinetics. The activation energy, E(a), was 105.7 kJ/mol. Fumonisin B(1) in contaminated corn could precipitate the nonenzymatic browning reaction with 0.1 M D-glucose at 60 and 80 degrees C.     

Analysis of a model reaction system containing cysteine and (E)-2-methyl-2-butenal, (E)-2-hexenal, or mesityl oxide

Cysteine conjugates, resulting from the addition of cysteine to alpha,beta-unsaturated carbonyl compounds, are important precursors of odorant sulfur compounds in food flavors. The aim of this work was to better understand this chemistry in the light of the unexpected double addition of cysteine to two unsaturated aldehydes. These reactions were studied as a function of pH. When (E)-2-methyl-2-butenal (tiglic aldehyde, 4) was treated with cysteine in water at pH 8, the major product formed was the new compound (4R)-2-(2-[[(2R)-2-amino-2-carboxyethyl]thio]methylpropyl)-1,3-thiazolidine-4-carboxylic acid (6). Under acidic conditions (pH 1), we also observed a double addition, but the second cysteine was linked by a vinylic sulfide bond to form the previously unreported major product, (2R,2'R,E)-S,S'-(2,3-dimethyl-1-propene-1,3-diyl)bis-cysteine (7). When (E)-2-hexenal (12) was treated with cysteine under acidic conditions, the major product was the novel (4R,2' 'R)-2-[2'-(2' '-amino-2' '-carboxyethylthio)pentyl]-1,3-thiazolidine-4-carboxylic acid (13), and the formation of an vinylic sulfide compound analogous to 7 was not observed. Reduction of the acidic crude reaction mixture with NaBH(4) afforded 13 and the cysteine derivative (R)-S-[1-(2-hydroxyethyl)butyl]cysteine (14) in 14% yield. Treating (E)-2-hexenal with cysteine at pH 8 followed by NaBH(4) reduction yielded the new product (3R)-7-propylhexahydro-1,4-thiazepine-3-carboxylic acid (15). Addition of cysteine to mesityl oxide (16), at pH 8, followed by reduction with NaBH(4) furnished (R)-S-(3-hydroxy-1,1-dimethylbutyl)cysteine (3) and the new compound (3R)-hexahydro-5,7,7-trimethyl-1,4-thiazepine-3-carboxylic acid (18).     

The formation of 2-chlorobenzamide upon hydrolysis of the benzoylphenylurea insecticide 1-(2-chlorobenzoyl)-3-(4-chlorophenyl) urea in different water systems

It has been reported that 1-(2-chlorobenzoyl)-3-(4-chlorophenyl) urea (CCU), an insect growth regulator, has no measurable toxicity to nontarget organisms and is essentially harmless to humans. However, one of its degradation products, 2-chlorobenzamide (CBA), is suspected of being a carcinogen. Therefore, the maximum concentration of CBA formed and the dynamics of its formation need to be given careful attention after CCU is used in the field. This paper describes the degradation of CCU to form CBA in three different water systems (distilled water, spring water, and simulated seawater) and the effects of temperature on the dynamics of CBA formation. The results indicate that the maximum level of CBA concentration is different in the different systems (highest in spring water) and that the temperature has a significant impact on the process (higher temperature leads to higher and earlier peak of CBA concentration). The maximum concentration of CBA after application of CCU was approximately 3.8% of the initial concentration of CCU at 35 degrees C in distilled water, and 2.4% in spring water at 25 degrees C.     

Quantitative analysis, occurrence, and stability of (E)-1-(2,3,6-Trimethylphenyl)buta-1,3-diene in wine

A stable isotope dilution assay has been developed for quantification of (E)-1-(2,3,6-trimethylphenyl)buta-1,3-diene (4) in wine using a [(2)H(6)]-analogue. Using this method, 4 was found in 96 out of 97 white wines, but in none of 12 red wines analyzed. 4 was found to be most prevalent in Semillon wines, followed by Chardonnay, with Riesling showing the least amount of 4 among these three varieties. 4, like 1,1,6-trimethyldihydronaphthalene (TDN, 3), appears to be formed during the aging process. 4 was found to be unstable in model wine, and in both white and red wine, with the order of stability being model > white > red. In a PVPP-stripped red wine, the rate of degradation of 4 was substantially lessened, with the final concentrations very close to those observed in model wine. When treated with either grape or wine tannin extracts in model wine, the concentration of 4 was found to decrease to levels very close to those observed with an untreated red wine. When white wine was heated at 45 degrees C, 4 was formed, indicating the presence of precursor forms. The amounts formed were much higher than those found in a commercial white wine. 4 was also observed in red wine heated to 45 degrees C, but the concentration produced was much less than that with white wine.     

Identification of fructo- and malto-oligosaccharides in cured tobacco leaves (Nicotiana tabacum)

Fructo-oligosaccharides (FOSs) and malto-oligosaccharides (MOSs) in cured tobacco leaves ( Nicotiana tabacum ) were detected and quantified using high-performance liquid chromatography (HPLC) coupled with electrospray ionization (ESI) tandem mass spectrometry (MS/MS). These oligosaccharides were present in several tobacco materials including flue-cured tobacco, sun/air-cured tobacco, and cut filler of commercially available tobacco products, but were not detected in air-cured tobacco. The changes in these oligosaccharides during storage were also investigated. The results revealed that MOSs simply decreased under a warm atmosphere and FOSs increased. In addition, the formation of FOSs in cured tobacco leaves occurred in the presence of sucrose. With regard to FOSs, it has been reported that green tobacco leaves do not contain FOSs such as kestose, nystose, and fructosyl-nystose. The results of a model test suggested that the changes in FOS amount were caused by enzymatic reactions.     

Stimulation of nicotine demethylation by NaHCO(3) treatment using greenhouse-grown burley tobacco

Experiments were conducted in tobacco (Nicotiana tabacum L.) to investigate the effect of sodium bicarbonate (NaHCO(3)) on the conversion of nicotine to nornicotine, a secondary alkaloid that can form the tobacco-specific nitrosamine N-nitrosonornicotine (NNN). The results showed that, under optimum conditions, NaHCO(3) stimulated nicotine conversion in converter plants to the maximum level predetermined by the genetic background. The conversion level in NaHCO(3)-treated leaves was 2-3 times higher than that in control leaves. For young seedlings the optimum concentration of sodium bicarbonate was 0.8% aqueous solution, and for adult plants the optimum concentration was 1%. Lower concentrations resulted in partial stimulation, whereas higher concentration damaged leaf tissue and resulted in a lower conversion level. Studies with different temperatures (from 22 to 43 degrees C) showed that 37 degrees C was optimal. This temperature allowed the least amount of time, 2-3 days for mature leaves and 4-6 days for green leaves, for the major converters to reach >95% of nicotine conversion. An examination of leaves from different growth stages and stalk positions showed that the amount of time needed for conversion was longer for young leaves and shorter for mature leaves. Treatment of leaves with NaHCO(3) affords a rapid and convenient means of identifying and removing nornicotine converter plants during growth in the greenhouse or field.