Study On Preparation And Application Of Active Biological Agent Based On Tobacco Extract

The technology of synthesizing tobacco flavors or improving the quality of cigarettes based on biotechnology has become a research hotspot in the tobacco industry in recent years.This study is divided into four parts: Firstly,appropriate tobacco extract was obtained through process optimization as microbial fermentation substrate;Nextly,the aroma-producing microorganisms were screened from the tobacco surface and the microorganisms with obvious aroma-producing characteristics were identified;Then,an active biological agents based on the above tobacco extracts and typical microorganisms were prepared;Finally,the active biological agent was applied to ferment tobacco leaves to improve its quality.This study not only prepared natural tobacco flavor with obvious aroma and style characteristics,but also obtained active biological agents that could improve the internal quality of tobacco.The main research is as follows:1?Study on extraction of tobacco extractTaking sugar content and extraction rate as indicators,the extraction effect of tobacco leaf was studied from the ratio of extraction material to liquid,time and temperature.Firstly,the optimal conditions of tobacco extraction were determined by single factor experiment,and then the response surface analysis was carried out by using the Box-Behnken principle.The results showed that the order of the influence of the three factors on the extraction rate was extraction time > solid-liquid ratio > extraction temperature.The optimization results showed that the optimal conditions for tobacco extraction were solid-liquid ratio of 1:12,constant temperature extraction at 60 ? for 2 h.At this time,the extraction rate and sugar content of tobacco extraction could reach 55.34% and 13.14 g/L,respectively.2?Screening and identification of aroma-producing microorganismsFour kinds of tobacco leaves from different sources were selected,and 18 microorganisms were isolated from the surface of the tobacco leaves.By smelling the flavor changes of tobacco extracts before and after fermentation of 18 strains,combined with the microscopic examination results of the strains,the morphology and smell of the strains on the plate,and the molecular identification results of the strains,comprehensive consideration was made,a strain of Saccharomyces cerevisiae(labeled Ye13 in the laboratory)was selected as the strain used in the subsequent experiments.The growth characteristics of Ye13 were analyzed,and the optimal growth temperature was 30 ?,the optimal PH was 5.0,and the optimal growth speed was 200 rpm.Considering that Ye13 originates from the surface of dry tobacco leaves,it can be suitably applied to tobacco leaf fermentation to improve the quality of tobacco leaves.Therefore,the feasibility of preparing active biological agents from Ye13 is further investigated.3?Research on active biological agentsIn order to better retain the original aroma of tobacco,this study further prepared active biological agents based on tobacco extract after obtaining high-quality tobacco extract and typical aroma-producing microorganisms.The results showed that the inoculation amount and fermentation time had a great influence on the fermentation effect,and the fermentation temperature had a little influence on the fermentation result.The results of orthogonal experiment showed that the optimal fermentation conditions were as follows: the inoculation amount was 5%,the fermentation temperature was 30 ?,and the fermentation time was 24 h.Under the optimal fermentation conditions,the content of macromolecules in the active biological agent decreased after fermentation.Total sugar,reducing sugar and amino acid decreased significantly compared with those before fermentation,which decreased by 51.30%,55.46% and 51.65% respectively.The determination results of aroma components showed that the content of most of the alcohol and ketone aroma substances in the active biological preparations increased,in addition,a variety of acid aroma substances were added.4?Study on the application of active biological agents in the fermentation of tobacco leavesThe active biological agent prepared above were applied to the fermentation of tobacco leaves,and after adding the active biological agent,the content of representative components of burnt sweet flavor in the mainstream smoke of cigarettes prepared from the fermented tobacco leaves was increased.In addition,a variety of ethers,aldehydes,acids,ester carbonyl compounds and some phenols had been added.The sensory evaluation of the prepared cigarettes showed that after adding the active biological agent,the strength of cigarettes became full,and the aroma quality and quantity increased significantly.Active biological agents can significantly improve the smoking and aftertaste of cigarettes and improve its internal quality.GC-MS analysis of aroma components extracted from fermented tobacco leaves by simultaneous distillation showed that: active biological agents can promote the increase of the content and quantity of aroma components in tobacco leaves,and can effectively improve the content of several important tobacco aroma components.The microbial diversity analysis of the fermented tobacco leaves showed that the overall microbial community species diversity of the tobacco leaves after the fermentation with the active biological agent decreased,but the content of Saccharomyces cerevisiae increased significantly,indicating that Ye13 maintained a certain activity during the fermentation process,which is one of the important factors to improve the quality of tobacco leaves.In summary,through the four parts of tobacco extract process optimization,microbial isolation and identification,preparation of active biological agents from tobacco extract and tobacco fermentation,this study not only obtained a microorganism with significant aromaproducing characteristics and adapted to the complex environment of tobacco,but also obtained an active biological agent of tobacco extract that can be used to improve the quality of tobacco.It has a certain application prospect for improving the internal quality of cigarettes and tobacco fermentation.