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To develop maltol latent fragrant compounds suitable for heated cigarettes, seven maltol carbonates were synthesized by condensation reaction of maltol with different alcohols. The structures of the products were confirmed by nuclear magnetic resonance and high-resolution mass spectrometry techniques. The pyrolysis behavior of seven maltol carbonates was investigated by thermogravimetric analysis and online pyrolysis gas chromatography-mass spectrometry(Py-GC-MS). The results showed that under the optimized conditions, the yields of seven maltol carbonates ranged from 83. 0% and 94. 3%; The synthesized maltol carbonate had mainly two weight loss modes: volatilization and pyrolysis; Upon heating, maltol carbonate could be pyrolyzed to produce maltol, and its pyrolysis ability was closely related to the type of raw material alcohol. Maltol tertiary alcohol and benzyl alcohol carbonate were the most easily pyrolyzed, which could rapidly pyrolyze and release maltol at a relatively low temperature, adapted to the heating temperature of the heated cigarettes. In contrast, maltol secondary alcohol carbonate had weaker pyrolysis ability, while maltol primary alcohol carbonate was difficult to pyrolyze at low temperatures. [ABSTRACT FROM AUTHOR]

In order to develop tobacco flavors with good aroma release stability, a phase transfer catalytic method was used to synthesize carvacrol-β-D-glucoside. The pyrolysis of the synthesized carvacrol-β-D-glucoside was analyzed. It was also added to cigarettes to investigate its pyrolysis release behavior in the particulate phase of mainstream cigarette smoke and its application effect on flavoring. The results showed that: 1) Carvacrol-β-D-glucoside could be obtained by the phase transfer catalytic method, and its structure was confirmed by ¹H NMR, 13C NMR, FTIR, and HRMS. 2) At different heating temperatures, the main cleavage product of carvacrol-β-D-glucoside was carvacrol. 3) By adding carvacrol-β-D-glucoside to cigarettes in different flavoring ways, different flavoring effects could be achieved. The aroma release of cigarettes with carvacrol-β-D-glucoside in cut tobacco was more uniform and stable, and the flavoring effect was better than that of cigarettes with adding carvacrol directly. 4) Cigarettes with added carvacrol-β-D-glucoside could release carvacrol to the particulate phase of mainstream smoke during smoking, and the release of carvacrol increased with the increase of flavoring amount, while the transfer rate of carvacrol to the particulate phase of mainstream smoke was relatively stable. [ABSTRACT FROM AUTHOR]

To study the genes regulating the synthesis of sugar esters in tobacco, transcriptome sequencing analysis was performed on tobacco glandular hair cells and bundle sheath cells, and RT-qPCR was used for verification. The results showed that 11 962 differentially expressed genes were obtained, of which 5145 were up-regulated and 6817 were down-regulated. GO annotation of differentially expressed genes showed that the up-regulated genes mainly included biological processes (2265), molecular functions (1169), and cellular components (363), with a total of 3793 GO terms. Based on the assignment of KEGG pathway, the sequence was mapped to 122 metabolic pathways. Glycolysis/gluconeogenesis, starch and sucrose metabolism, phenylpropanoid biosynthesis related to glycoester synthesis were highly enriched, and acyl sugar pathway in glandular hairs was significantly up-regulated. By comparing with the tobacco reference genome, 12 up-regulated acyltransferase genes were identified in glandular hair cells. RT-qPCR results showed that the expression of 11 genes was higher than that of the control. The expression of gene 63826, gene 16194 and gene 37511 increased by 195.79 times, 166.23 times and 132.65 times, respectively. The above 11 genes may be involved in regulating the synthesis of sugar esters in glandular hair cells, providing a basis for further verification of gene function. [ABSTRACT FROM AUTHOR]

Based on the significant effects of enzyme treatment on tobacco and its products, the research achievements of enzyme treatment in tobacco quality improvement, aroma production, and harm reduction in recent years were reviewed as follows. The weak links and problems in the research of biological enzymes were analyzed. Significant progress had been made in the research of enzymatic degradation of starch and pectin in tobacco leaves, with composite enzymes being more effective than single enzymes. The combination of laccase and protease had a better effect on treating tobacco stems than single. At the same time, treatments with acid, alkali, and steam explosion could significantly improve the enzymatic hydrolysis effect. In the process of reconstituted tobacco leaf concentrate using raw materials for tobacco thin slice preparation, biological enzymes were more effective, with advantages such as short processing time, high efficiency, and significant effects. In terms of aroma production, it was mainly achieved by producing substances that could participate in the Maillard reaction or catalyze the production of aroma precursors. In terms of harm reduction of cigarettes, it was mainly achieved by reducing the content of tobacco specific Nitrosamine and smoke free radicals and other harmful substances. In the future, in-depth research can be carried out from the aspects of structural characterization and characteristics of tobacco biomacromolecules, creation of tobacco enzyme preparations and analysis of catalytic mechanism, so as to further promote the research and application of biological enzymes in tobacco aroma enhancement, quality improvement and harm reduction. [ABSTRACT FROM AUTHOR]

Investigating the quantitative analysis of two stereoisomeric compounds, specifically (6R, 9R)-3-oxo-α- ionol-β-D-glucopyranoside (rrOIPG) and (6R, 9S)-3-oxo-α-ionol-β-D-pyranoside (rsOIPG), megastigmatrienone precursors of the prominent aroma components in tobacco leaves, represented the core focus of this study. A meticulously- devised methodology combining ultrasound-assisted extraction with ultra-high-performance liquid chromatography (UHPLC) was deployed to ascertain the precise quantification of rrOIPG and rsOIPG. This robust analytical approach was judiciously employed to elucidate the geographical distribution patterns of these stereoisomers within tobacco leaves sourced from seven diverse global regions. The results revealed the compelling linear response characteristics of rrOIPG and rsOIPG, scanninged concentration ranges of 25. 85 to 258. 50 μg/ mL (R2 =0. 999 1) and 6. 28 to 62. 75 μg/ mL (R2 =0. 999 0), respectively. Exceptionally low detection limits of 2. 5 μg/ mL for rrOIPG and 2. 9 μg/ mL for rsOIPG underscore the methods inherent precision. The analysis exhibited superb stability and reproducibility, epitomized by recovery rates of 83. 94% for rrOIPG and 105. 90% for rsOIPG, and RSDs were 1. 22% and 1. 93%, respectively. According to the content distribution of rrOIPG and rsOIPG, the tobacco leaves from domestic and foreign origins could be well distinguished, and the content of rsOIPG configuration in foreign origins was significantly higher than that in domestic origins. In terms of the distribution of the two configurations, the content of rrOIPG configuration was higher than that of rsOIPG configuration. The developed technique exhibits high feasibility and streamlined pretreatment processes, making it well-suitable for the quantitative analysis of rrOIPG and rsOIPG in tobacco leaves. [ABSTRACT FROM AUTHOR]

Shandong tobacco leaves were fermented by Enterobacter cloacae F8-2 and Acinetobacter nosocomialis 7S-2 mixed bacteria. The sensory quality, chemical composition and chemical composition of tobacco smoke and microbial community on tobacco leaf surface before and after fermentation were studied. The results showed that when Enterobacter and Acinetobacter were mixed in 1:1 fermentation for 24 h, the aroma of the tobacco leaf increased and the impurity of the tobacco leaf decreased most significantly. After fermentation, methylheptenone, dihydrotomarone and 5-methylfurfural increased by 89.22%, 35.67% and 33.93%, respectively. Nerolol, solanone and acyl alcohol in tobacco smoke increased by 78.01%, 46.83% and 45.95%, respectively. Mixed fermentation had a great impact on the bacterial community on the surface of tobacco leaves, and the microorganisms cooperated with each other in fermentation and symbiosis. At the same time, Enterobacter and Acinetobacter may be related to the production of aromatic substances such as benzaldehyde and methyl heptene ketone. [ABSTRACT FROM AUTHOR]

C4F tobacco leaves in Hechi area, Guangxi were treated by solid-state fermentation with Bacillus sp. ZZ-8. Single factor test and response surface methodology experiment were used to obtain the best processing conditions of Guangxi tobacco leaves treated by Bacillus sp. ZZ-8. The smoking quality of treated tobacco leaves was evaluated by analyzing the content changes of conventional chemical components and important aroma components, combined with sensory evaluation technology. Research results demonstrated that the optimum extracting condition was as follow: inoculum 3 mL, fermentation temperature 28 T, fermentation time 36 h, and tobacco leaf relative humidity 60%. Under this condition, the highest content of petroleum ether extract was 9.01%. The total sugar content of tobacco leaves decreased from 23. 10% to 22. 20%, the reducing sugar content increased from 22. 19% to 22.40%, and the sugar alkali ratio decreased from 10.23 to 9.45. Benzaldehyde and β-cyclocitral were produced in tobacco leaves after fermentation, the overall content of aroma components in C4F tobacco increased from 170. 444 µg/g to 186. 267 µg/g in which the content of aroma substances such as damascene, solanone, megastigmatrienone, dihydrodamascone, geranyl acetone and dihydroactinidide changed greatly. After sensory evaluation, the aroma quality of treated tobacco leaves increased, the irritation and miscellaneous gas decreased, the flue gas was fine and soft, the aftertaste was comfortable, and the smoking quality was improved. [ABSTRACT FROM AUTHOR]

The strains that could degrade β-carotene with high efficiency were screened from fresh Yuyan 13 tobacco leaves, and the main degradation products were detected by GC-MS, the species of the strain were identified by morphology and 16S rDNA phylogenetic tree, the fermentation conditions of the strain were optimized by single factor test and orthogonal test. The results showed that YT-3 strain was Enterobacter hormanii subsp. which could degrade β-carotene with high efficiency. The main degradation products were β-ionone, dihydrokiwifruit lactone, β-cyclic citral (3.21%) and other flavor substances. The optimum fermentation conditions of YT-3 strain were as follows: NaNO3 concentration 4 g/L, sucrose concentration 30 g/L, yeast powder concentration 3 g/L, initial pH value 7.0. Under these fermentation conditions, the degradation rate of β-carotene reached 97.05%. [ABSTRACT FROM AUTHOR]

Based on the fermentation pretreatment of B3F tobacco leaves in Baise, Guangxi by Saccharomyces cerevisiae, the tobacco leaves were further fermented by oligotrophic maltophilia. The optimum fermentation conditions were determined by single factor test and orthogonal test; The changes of conventional chemical components and aroma components of B3F tobacco leaves in Baise, Guangxi before and after mixed microbial fermentation were compared and analyzed, and the improvement level of sensory evaluation quality of B3F tobacco leaves after fermentation was studied. The results showed that the optimum fermentation conditions of oligotrophic maltophilia were inoculation amount of 8 ml, fermentation temperature of 34 °C, fermentation time of 60 h and humidity of 70%. Under these conditions, the mass fraction of petroleum ether extract of fermented tobacco leaves reached 16.50%. After mixed microbial fermentation, the contents of total sugar and reducing sugar in conventional chemical components of tobacco leaves increased, arid the sugar alkali ratio of tobacco leaves increased from 6.77 to 7.55. The mass concentration of six important aroma components in tobacco leaves (megaketene, damascus, eggplant, dihydrodamadone, vanillin and dihydrokiwifruit lactone) increased by 6.65% compared with that before fermentation. After fermentation, the aroma quality and aroma plumpness of tobacco leaves increased, and the irritation and miscellaneous gas of tobacco leaves decreased. [ABSTRACT FROM AUTHOR]

Taking the strong-flavor, intermediate-flavor and clear-flavor reconstituted tobacco leaf concentrates as the research object, the contents of conventional chemical components was compared by continuous flow method, the differences of neutral aroma components were detected by GC-MS, and the differences of microbial community structure and diversity in different flavor concentrates were compared by high-throughput sequencing. The results showed that the content of water-soluble total sugar in strong-flavor concentrate was lower, and the content of other conventional chemical components was higher. The contents of α-damascenone and megastigmatrienone were higher in the clear-flavor, the contents of nicotinine and acetylpyrrole were higher in the strong-flavor, and the contents of most flavor components in intermediate-flavor were in the middle. The species diversity of bacteria in clear-flavor concentrate was the largest, and the species diversity of fungi in strong-flavor concentrate was the largest. The dominant bacterial genera of the three concentrates were Bacillus, the dominant fungal genera of the strong-flavor and intermediate-flavor concentrates were Debaryomyces, and the strong-flavor concentrated fragrance was Ogataea. [ABSTRACT FROM AUTHOR]

Tobacco-derived microorganisms refer to a class of microorganisms that adapt to and come from the chemical environment of tobacco. The screening, physiological and biochemical function mining and application technology research of these microorganisms are of great value for cigarette quality improvement, aroma enhancement and harm reduction. This paper reviews the sources and types of tobacco-derived microorganisms, as well as the application in tobacco leaf fermentation treatment, reconstituted tobacco leaf concentrate fermentation treatment and tobacco flavor preparation. The weak links and problems of tobacco-derived microorganisms were analyzed in the construction of resource system, function mining and the study of tobacco treatment mechanism. The development prospect of tobacco-derived microorganisms was prospected from the aspects of strengthening basic theoretical research and deepening applied technology research. [ABSTRACT FROM AUTHOR]

The medium of Stenotrophomonas sp. H3-1 was optimized by one single factor experiment. The strain was prepared into bacteria-preparation and preliminarily used in tobacco fermentation, and the quality changes of tobacco leaves before and after fermentation were compared. It showed that the optimized culture medium was 2.0 g/L of maltose as the carbon source and the nitrogen source is ammonium sulfate of 2.0 g/L, initial pH value 8, temperature 40 °C. After tobacco fermentation, the mass concentrations of five flavor substances changed obviously, the mass concentrations of solanone and 9-hydroxy-4,7-megadiene-3-one, which had greater effect on aroma composition, all increased to different degrees. The evaluation results showed that the aroma quality, aroma quantity and aroma concentration of tobacco leaves were improved after fermented with bacterial preparations. [ABSTRACT FROM AUTHOR]

The volatile chemical components of tobacco flavor were prepared by simultaneous distillation extraction and analyzed using gas chromatography-mass spectrometry. The aroma-active violates were determined by means of gas chromatography-mass spectrometry-olfactometry combined with aroma extraction dilution analysis, ad the important odorants were further screened out according to quantitative study and odor activity values. Aroma recombination and omission experiments identified the key odorants of the tobacco flavor. The results exhibited that simultaneous distillation extraction of tobacco flavor included 54 kinds of volatile compounds, including 22 aroma-active compounds, and 17 odorants were screened out as the predominant odorants according to flavor dilution factors and odor activity values, β-damascarone, linalool, ethyl maltol, linalyl acetate, neryl acetate, myrcene, nerol, geranyl acetate, D-limonene, and benzyl alcohol were confirmed as the key odorants of tobacco flavor. [ABSTRACT FROM AUTHOR]

The structure of tobacco stems from He'nan province was characterized by chemical composition analysis, infrared spectrum analysis and crystallinity analysis. The thermal decomposition characteristics of tobacco stems were studied by thermogravimetric analysis, and the pyrolysis kinetic parameters were also calcu-lated. Results showed that there were typical biomass components in tobacco stems, and the contents of cellulose, hemicellulose, lignin and pectin were 28.15%, 20.01%, 5.67% and 5.32%, respectively. There were characteristic structural groups of lignocellulose in tobacco stem. The crystallinity of tobacco stem was 25.37%, which indicated that the pyrolysis reaction of tobacco stem was easier to be carried out. During the pyrolysis of tobacco stems, the weight loss of cellulose, hemicellulose and lignin were 18.83%, 17.89% and 15.06%, respectively. Corresponding activation energies of pyrolysis reactionswere 31.72 kJ/mol, 27.82 kJ/mol and 58.99 kJ/mol, respectively. The pyrolysis of cellulose and hemicellulose were dominated by a first-order chemical reaction, while lignin pyrolysis was controlled by a third-order chemical reaction. [ABSTRACT FROM AUTHOR]

Using genetic engineering techniques, the recombinant plasmid pPICZαA-AoFT was linearized, and then electroporated into Pichia pastoris X33 to construct recombinant Pichia pastoris engineering strain pPICZαA-AoFT/X33 and achieve the recombinant fructosyl transferase AoFT in Pichia pastoris. Furthermore, the enzymatic properties of the purified fructosyl transferase AoFT were studied. The results showed that the pPICZαA-AoFT/X33 multiplied rapidly with the increase of fermentation time. The OD600 value of the cell reached a peak at 120 h, and the total protein concentration of the extracellular protein in the fermentation broth was 1.64 mg/mL. The specific activity of the purified fructosyl transferase AoFT enzyme was 71.4 U/mg, and the yield was 70.6%. The peptide N-glycosidase F digestion reaction confirmed that the purified fructosyl transferase AoFT had some N-glycosylation sites. The optimal reaction temperature of fructosyl transferase AoFT was 45 °C, and the optimal buffer system was sodium acetate buffer (pH value was 4.5 ~ 6.0). Metal ions Mn2+ and Ca2+ could activate fructosyl transferase AoFT, while Al3+, Fe2+, Ni2+, Zn2+ and EDTA could inhibit fructosyl transferase AoFT. Surfactants (SDS, Tween-20 and Triton X-100) had little effect on the activity of fructosyl transferase AoFT, while the reductant DTT had obvious inhibition effect on the activity of fructosyl transferase AoFT. The low concentration of PMSF had no significant effect on the activity of fructosyl transferase,but the high concentration could reduce the activity. The activity of fructosyl transferase AoFT showed strong resistance to organic solvents (ethanol, DMSO, dimethylformamide, n-butanol, n-hexane and toluene). [ABSTRACT FROM AUTHOR]

The fermentation conditions of β-carotene degrading strain HC-3 were optimized by single factor experiment and orthogonal experiment,and the fermentation liquid of strain HC-3 obtained under the optimal fermentation conditions was applied to the reconstituted tobacco concentrate to study its aroma enhancing effect. The results showed that the best fermentation conditions for β-carotene degradation were 3 g/L of NaNO3,30 g/L of sucrose,3 g/L of yeast powder and 7.0 of initial pH value. Under this condition,the degradation rate of β-carotene could reach 93.35%. After being treated by strain HC-3,the eight flavor components of dihydroactyl lactone, 4,7,9-Mega three ene -3- keton, 4-hydroxyl -β-two hydrogen damascene, 9-hydroxy - 4,7-Mega three ene ketene, triethyl citrate, 4- (3-hydroxyl butyl) -3,5,5- three methyl -2- cyclohexene -1- ketone, tetradecanoic acid and neophytadiene in the reconstituted tobacco concentrate were significantly increased. Compared with the untreated reconstituted tobacco concentrate,the fragrances made from the reconstituted tobacco concentrate treated by strain HC-3 had better quality,more aroma,higher aroma concentration and lower offensive odor content. [ABSTRACT FROM AUTHOR]

A flavor-enhancing strain HS-1, was screened and isolated from the reconstituted tobacco concentrate by dilute coating plate method. The strain was identified and the culture conditions for fermentation of the strain were optimized. The neutral aroma components of the reconstituted tobacco concentrate fermented under the optimal culture conditions were analyzed. The results showed that HS-1 was Planococcus sp., the optimal culture conditions for fermentation were fermentation temperature of 30 °C, inoculation amount of 5%, and fermentation time of 36 h. Under this culture condition, the total content of neutral aroma components in the reconstituted tobacco concentrate fermented by HS-1 was increased from 56.845% before fermentation to 72.527%, and the types and contents of neutral aroma components had increased. 5-methylfurfural and β-ionone were new ingredients after fermentation, the components such as benzyl alcohol, phenylacetaldehyde, 2-acetylpyrrole, phenethyl alcohol, solanone, S-amascenone, dihydroactinidiolide and megastig-matrienone had increased in varying degrees after fermentation. [ABSTRACT FROM AUTHOR]