Study on the effect of enzymatic treatment of tobacco on HnB cigarettes and microbial succession during fermentation.

Starch and cellulose are the fundamental components of tobacco, while their excessive content will affect the quality of tobacco. Enzymatic treatment with different enzymes is a promising method to modulate the chemical composition and improve the sensory quality of tobacco leaves. In this study, enzymatic treatments, such as amylase, cellulase, and their mixed enzymes, were used to improve tobacco quality, which could alter the content of total sugar, reducing sugar, starch, and cellulose in tobacco leaves. The amylase treatment changed surface structure of tobacco leaves, increased the content of neophytadiene in tobacco by 16.48%, and improved the total smoking score of heat-not-burn (HnB) cigarette products by 5.0 points compared with the control. The Bacillus, Rubrobacter, Brevundimonas, Methylobacterium, Stenotrophomonas, Acinetobacter, Pseudosagedia-chlorotica, and Sclerophora-peronella were found to be significant biomarkers in the fermentation process by LEfSe analysis. The Basidiomycota and Agaricomycetes were significantly correlated with aroma and flavor, taste, and total score of HnB. The results showed that microbial community succession occurred due to amylase treatment, which promoted the formation of aroma compounds, and regulated the chemical composition of tobacco, and improved tobacco quality during tobacco fermentation. This study provides a method for enzymatic treatment to upgrade the quality of tobacco raw materials, thereby improving the quality of HnB cigarettes, and the potential mechanism is also revealed by chemical composition and microbial community analysis. KEY POINTS: Enzymatic treatment can change the chemical composition of tobacco leaves. The microbial community was significantly affected by enzymatic treatment. The quality of HnB cigarettes was significantly improved by amylase treatment.

Tobacco microbial screening and application in improving the quality of tobacco in different physical states.

The first-cured tobacco contains macromolecular substances with negative impacts on tobacco products quality, and must be aged and fermented to mitigate their effects on the tobacco products quality. However, the natural fermentation takes a longer cycle with large coverage area and low economic efficiency. Microbial fermentation is a method to improve tobacco quality. The change of chemical composition of tobacco during the fermentation is often correlated with shapes of tobacco. This study aimed to investigate the effects of tobacco microorganisms on the quality of different shapes of tobacco. Specifically, Bacillus subtilis B1 and Cytobacillus oceanisediminis C4 with high protease, amylase, and cellulase were isolated from the first-cured tobacco, followed by using them for solid-state fermentation of tobacco powder (TP) and tobacco leaves (TL). Results showed that strains B1 and C4 could significantly improve the sensory quality of TP, enabling it to outperform TL in overall texture and skeleton of tobacco products during cigarette smoking. Compared with the control, microbial fermentation could increase reducing sugar; regulate protein, starch, and cellulose, reduce nicotine, improve total aroma substances, and enable the surface of fermented TP and TL to be more loose, wrinkled, and porous. Microbial community analysis indicated that strains B1 and C4 could change the native structure of microbial community in TP and TL. LEfSe analysis revealed that the potential key biomarkers in TP and TL were Bacilli, Pseudonocardia, Pantoea, and Jeotgalicoccus, which may have cooperative effects with other microbial taxa in improving tobacco quality. This study provides a theoretical basis for improving tobacco fermentation process for better cigarettes quality.