WO2019010627A1 - Reconstituted tobacco flake and preparation method therefor - Google Patents

Abstract

A reconstituted tobacco flake and a preparation method therefor. The preparation method comprises: mixing a tobacco waste material with a solvent in a mass ratio of 1 : 6 to 10 to obtain a tobacco liquid; adding a complex microbial preparation to the tobacco liquid, fermenting the mixture at 30 °C to 37 °C for 24 h to 48 h, and performing solid-liquid separation so as to obtain a fermentation liquid and a solid, the complex microbial preparation comprising lactobacillus acidophilus, saccharomyces cerevisiae and bacillus subtilis, the amount ratio of lactobacillus acidophilus, saccharomyces cerevisiae and bacillus subtilis per unit volume being 1 : 0.4 to 1.2 : 0.3 to 0.6; processing the solid to tobacco flakes; and wetting the tobacco flakes using the fermentation liquid, so as to obtain reconstituted tobacco flakes.

Description

Recombinant tobacco sheet and preparation method thereof Technical field

The invention relates to the field of tobacco products, in particular to a recombinant tobacco sheet and a preparation method thereof.

Background technique

Tobacco is a widely grown cash crop. Tobacco waste generated during the picking or processing of tobacco, such as tobacco stems, tobacco leaf fragments or cigarettes, can cause a lot of damage. In order to further process the reuse of tobacco waste, the technology of reconstituting tobacco sheets has been extensively developed. However, the application range of recombinant tobacco sheets in cigarettes is very small, mainly because the reconstituted tobacco sheets processed by tobacco waste lack many tobacco aroma substances and are highly irritating.

In summary, traditional recombinant tobacco sheets have a poor aroma, sweetness and softness.

Summary of the invention

Based on this, it is necessary to provide a recombinant tobacco sheet which has good aroma, sweetness and softness and a preparation method thereof.

A method for preparing a recombinant tobacco sheet, comprising the steps of:

Mixing the tobacco waste with the solvent at a mass ratio of 1:6-10 to obtain a tobacco liquid;

Adding a composite microbial preparation to the tobacco liquid, and fermenting at 30 ° C to 37 ° C for 24 h to 48 h, and obtaining a fermentation liquid and a solid material after solid-liquid separation; wherein the composite microbial preparation contains Lactobacillus acidophilus and wine The ratio of the yeast and the Bacillus subtilis per unit volume of the Lactobacillus acidophilus, the Saccharomyces cerevisiae and the Bacillus subtilis is 1:0.4 to 1.2:0.3-0.6;

Processing the solid to form a tobacco sheet;

The tobacco sheet is wetted using the fermentation broth to obtain the recombinant tobacco sheet.

A recombinant tobacco sheet prepared by the above-described method for preparing a recombinant tobacco sheet.

In the above method for preparing a recombinant tobacco sheet, the tobacco waste material is fermented by using a composite microbial preparation containing Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis to obtain a fermentation liquid and a solid matter. The inventors conducted a large number of experiments on the selection of complex microorganisms, the ratio of complex microorganisms, and fermentation conditions. Unexpected discovery of a composite microbial preparation formed by Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis. After fermentation of tobacco liquid under suitable conditions for a period of time, the composite microorganism can decompose some cellulose, protein, polysaccharide and lipid on tobacco waste. Such as macromolecular substances, protein content is reduced, pH value is reduced, and the quality of solid matter formed after fermentation of tobacco waste is improved. At the same time, aroma components such as lactic acid, acetic acid and alcohol are also dissolved in the fermentation liquid to improve the smoke smell of the fermentation liquid. The solid matter is then processed to form a tobacco sheet, and then the fermentation liquid is applied to the tobacco sheet to obtain a recombinant tobacco sheet. The cigarette formed by the processing of the reconstituted tobacco sheet is effectively improved in terms of aroma, sweetness and softness.

DRAWINGS

1 is a flow chart of a method of preparing a recombinant tobacco sheet of an embodiment.

Detailed ways

The above described objects, features and advantages of the present invention will become more apparent from the detailed description. Numerous specific details are set forth in the description below in order to provide a thorough understanding of the invention. However, the present invention can be implemented in many other ways than those described herein, and those skilled in the art can make similar modifications without departing from the spirit of the invention, and thus the invention is not limited by the specific embodiments disclosed below.

As shown in FIG. 1, a method for preparing a recombinant tobacco sheet according to an embodiment includes the following steps S110 to S140.

In step S110, the tobacco waste and the solvent are mixed at a mass ratio of 1:6 to 10 to obtain a tobacco liquid.

Tobacco waste is generally produced during harvesting or processing, such as tobacco stems, tobacco leaf fragments or smoke.

Specifically, the tobacco waste is selected from at least one of a tobacco stem, a tobacco leaf fragment, and a smoke.

In one embodiment, the tobacco waste is a separate tobacco stem, tobacco leaf fragments or tobacco.

In another embodiment, the tobacco waste is a combination of any two or three of tobacco stems, tobacco leaf fragments, and tobacco.

Generally, tobacco waste is difficult to apply to cigarette products. The main reason is that tobacco waste lacks many tobacco aroma substances and is highly irritating. In the present embodiment, by processing and fermenting tobacco waste using a composite microbial preparation, the taste of the cigarette made of tobacco waste can be improved, and waste utilization can be realized.

In one embodiment, in the step of mixing the tobacco waste material and the solvent in a mass ratio of 1:6 to 10, the mixing operation is agitation treatment at a temperature of 50 ° C to 70 ° C for 1 h to 2 h. Stirring at a relatively high temperature of 50 ° C ~ 70 ° C, accelerate the decomposition of tobacco waste, and promote the release of tobacco components into the tobacco liquid.

Specifically, the stirring speed of the stirring treatment is from 150 rpm/min to 250 rpm/min. For example, 160 rpm/min, 200 rpm/min, 220 rpm/min or 230 rpm/min, and the like. On the one hand, the tobacco waste can be broken up by stirring, and on the other hand, the release of the tobacco component into the tobacco liquid can be accelerated.

Specifically, the solvent mixed with the tobacco waste is water, and the solvent is used in an amount of 6 to 10 times the mass of the tobacco waste.

Step S120, adding a composite microbial preparation to the tobacco liquid obtained in S110, and fermenting at 30 ° C to 37 ° C for 24 h to 48 h, and obtaining a fermentation liquid and a solid matter after solid-liquid separation; wherein the composite microbial preparation contains eosinophil Bacillus, Saccharomyces cerevisiae and Bacillus subtilis, the ratio of the number of Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis per unit volume is 1:0.4 to 1.2:0.3 to 0.6.

The use of a composite microbial preparation to treat tobacco not only reduces the effects of harmful chemical substances (such as acrylamide, nitrosamines, nitrates, etc.) but also reduces polysaccharides, proteins, amino acids or lipids in tobacco waste. In addition, Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis in the composite microbial preparation produce a variety of tobacco flavor substances (such as acetoin, vanillin, phenylethyl alcohol) during fermentation. Different microbial metabolism methods are different, and the produced metabolites are different. The ratio of various bacteria in the composite microbial preparation will affect the overall evaluation of the aroma, sweetness and softness of the final prepared recombinant tobacco sheet. For example, the concentration of Lactobacillus acidophilus is too low, the amount of acidic substances produced is small, and the softness of tobacco waste is less improved. It does not improve the irritancy and enhance the softness. If the concentration of Lactobacillus acidophilus is too high, excessive lactic acid and other by-products are produced, resulting in a pH of the fermentation broth being too low. For example, the concentration of the Saccharomyces cerevisiae is too low, the concentration of the alcohol produced by the decomposition is too low, the amount of vanillin produced is small, the aroma of the tobacco waste is less improved, and the effect of improving the aroma is not obtained. If the concentration of Saccharomyces cerevisiae is too high, too much acid will be produced, causing excessive fermentation of the tobacco waste and producing a sour taste. Further, for example, the concentration of Bacillus subtilis is too low, and the sweetness of the tobacco waste is less improved, and the effect of improving the sweetness is not obtained. If the concentration of Bacillus subtilis is too high, excessive by-products will be produced. The ratio of the number of Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis per unit volume in the composite microbial preparation is 1:0.4 to 1.2:0.3-0.6, and the ratio of various microorganisms is appropriate, and the harmony produces a good aroma and sweetness. And gentle reconstituted tobacco sheets.

In one embodiment, the activation of Lactobacillus acidophilus, Saccharomyces cerevisiae, and Bacillus subtilis is first beneficial to improve fermentation efficiency.

Specifically, the preparation of the composite microbial preparation is obtained by the following preparation method: inoculation of Lactobacillus acidophilus into a medium containing a saccharide compound having a mass fraction of 5% to 15%, and culturing at 30 ° C to 37 ° C for 24 h to 36 h , to obtain a Lactobacillus acidophilus activating solution. Saccharomyces cerevisiae is inoculated into a medium containing a saccharide compound having a mass fraction of 5% to 15%, and cultured at 30 ° C to 40 ° C for 12 h to 24 h to obtain a Saccharomyces cerevisiae activating solution. Bacillus subtilis is inoculated into a medium containing a saccharide compound having a mass fraction of 5% to 15%, and cultured at 30 ° C to 37 ° C for 24 to 36 hours to obtain a Bacillus subtilis activating solution. And the ratio of the ratio of Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis in the combined microbial preparation per unit volume in a ratio of 1:0.4 to 1.2:0.3-0.6, the Lactobacillus acidophilus activating solution, the Saccharomyces cerevisiae activating solution and the grass The Bacillus activation solution is mixed to obtain a composite microbial preparation.

Specifically, after activation, the volume ratio of the Lactobacillus acidophilus activating solution, the Saccharomyces cerevisiae activating solution, and the Bacillus subtilis activating solution is about the ratio of the number of Lactobacillus acidophilus, Saccharomyces cerevisiae, and Bacillus subtilis.

Specifically, the saccharide compound is dissolved in water to obtain a medium containing a saccharide compound having a mass fraction of 5% to 15%, wherein the mass fraction of the saccharide compound in the medium is, for example, 8%, 10%, 12%, etc. . The saccharide compound facilitates the activation of microorganisms and forms a complex microbial preparation with good microbial activity.

Specifically, the saccharide compound may be brown sugar or white sugar or the like.

In the present embodiment, the saccharide compound is brown sugar, and the main component of the brown sugar is sucrose. In addition, the brown sugar also contains vitamins and some trace elements, which is beneficial to the activation of microorganisms and is relatively inexpensive.

Specifically, after a certain period of culture of Lactobacillus acidophilus, Saccharomyces cerevisiae, and Bacillus subtilis, the Lactobacillus acidophilus in the Lactobacillus acidophilus activating solution is in a logarithmic growth phase or a stable growth phase. Saccharomyces cerevisiae in the Saccharomyces cerevisiae activation solution is in logarithmic growth phase or stable growth phase The B. subtilis in the B. subtilis activating solution is in a logarithmic growth phase or a stable growth phase. Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis have better activity and are beneficial for rapid fermentation of tobacco waste.

Specifically, after inoculating Bacillus subtilis into the culture medium, the mixture is cultured at a certain stirring rate at a stirring speed of 100 rpm/min to 200 rpm/min to promote the growth of Bacillus subtilis, and a Bacillus subtilis activating solution is obtained.

In one embodiment, the Lactobacillus acidophilus, Saccharomyces cerevisiae, and Bacillus subtilis in the composite microbial preparation are all in a stationary phase.

In one embodiment, the concentration of Lactobacillus acidophilus in the Lactobacillus acidophilus activating solution is about 1 x 10 ⁷ /mL to 1 x 10 ⁸ /mL. The concentration of Saccharomyces cerevisiae in the Saccharomyces cerevisiae activating solution is about 1 × 10 ⁷ /mL to 1 × 10 ⁸ /mL. The concentration of Bacillus subtilis in the Bacillus subtilis activating solution is about 1 × 10 ⁷ /mL to 1 × 10 ⁸ /mL.

Specifically, the mass ratio of the composite microbial preparation to the tobacco liquid is 4-6:100, and the ratio is suitable, which is suitable for the growth of the composite microorganism.

Specifically, after the composite microbial preparation is added to the tobacco liquid and fermented at 30 ° C to 37 ° C for 24 h to 48 h, the fermentation product is passed through a sieve having a pore diameter of 0.5 μm to 5 μm, and the fermentation liquid and the solid matter are separately obtained. The filter screen is, for example, gauze or the like.

Step S130, processing the solid matter obtained in S120 to form a tobacco sheet.

Specifically, the separated solid matter is dried and pressed to form a tobacco sheet to prepare a cigarette.

Specifically, the pressed tobacco sheets can be cut into various sizes as needed.

Step S140, the tobacco sheet obtained in S130 is wetted using the fermentation liquid obtained in S120 to obtain a recombinant tobacco sheet.

In one embodiment, before the step of using the fermentation liquid to wet the tobacco sheet, the method further comprises the step of concentrating the fermentation liquid: concentrating the fermentation liquid under vacuum, the concentration temperature is 50 ° C to 70 ° C, and concentrating the fermentation liquid before The volume is 3 to 5 times the volume of the fermentation broth after concentration. The concentrated fermentation broth has a higher concentration, and the tobacco flakes obtained after being added to the tobacco flakes have a stronger aroma.

Specifically, the step of moisturizing the tobacco sheet using the fermentation liquid is specifically: adding the fermentation liquid to the tobacco sheet by coating, spraying or dipping, the fermentation liquid being sufficient to wet the tobacco sheet.

The inventors conducted a large number of experiments on the selection of complex microorganisms, the ratio of complex microorganisms, and fermentation conditions. Unexpected discovery of a composite microbial preparation formed by Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis. After fermentation of tobacco liquid under suitable conditions for a period of time, the composite microorganism can decompose some cellulose, protein, polysaccharide and lipid on tobacco waste. Such as macromolecular substances, protein content is reduced, pH value is reduced, and the quality of solid matter formed after fermentation of tobacco waste is improved. At the same time, aroma components such as lactic acid, acetic acid and alcohol are also dissolved in the fermentation liquid during the fermentation process. The recombinant tobacco sheet obtained by adding the fermentation broth to the tobacco sheet formed of the solid material, the cigarette formed by the processing of the recombinant tobacco sheet is effectively improved in aroma, sweetness and softness.

Moreover, the preparation method of the above-mentioned recombinant tobacco sheet is simple in operation, easy to industrialize, and can effectively expand the application range of tobacco waste, thereby realizing efficient waste utilization. At the same time, the preparation method of the above-mentioned recombinant tobacco sheet does not use any artificially synthesized additive in the preparation process, and is more safe to apply.

The recombinant tobacco sheet of one embodiment is prepared by the method for preparing the above-mentioned recombinant tobacco sheet, and the recombinant tobacco sheet has good aroma, sweetness and softness.

The experimental results show that the cigarettes processed by the above-mentioned recombinant tobacco sheets have better taste and finer taste, lower smoke irritability, lower oral residue, lower gas and increased aroma.

The following are examples of the examples (the following examples, unless otherwise specified, do not contain components other than the unavoidable impurities, which are not explicitly indicated.)

Example 1

The preparation process of the recombinant tobacco sheet of this embodiment is as follows:

(1) Preparation of a composite microbial preparation

Medium preparation: Take 10g of brown sugar, add 100g of water, mix and mix, sterilize at 100 °C for 30min, The medium was obtained and the medium was divided into three portions.

Preparation of Lactobacillus acidophilus activating solution: Lactobacillus acidophilus was inoculated into one of the above mediums, and cultured at 37 ° C for 24 hours.

Preparation of Saccharomyces Cerevisiae Activating Solution: Saccharomyces cerevisiae was inoculated into one of the above mediums and cultured at 30 ° C for 24 h.

Preparation of Bacillus subtilis activating solution: Bacillus subtilis was inoculated into one of the above-mentioned mediums, and cultured at 37 ° C, 121 rpm / min for 24 hours.

The Lactobacillus acidophilus activating solution, the Saccharomyces cerevisiae activation solution, and the Bacillus subtilis activating solution are mixed in a ratio of the ratio of Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis per unit volume in a ratio of 1:0.6:0.3 to obtain a composite microorganism. preparation.

(2) Preparation of tobacco liquid

Tobacco waste pretreatment: tobacco processing waste containing tobacco leaf fragments, stems and cigarettes and water at a mass ratio of 1:8, water temperature of 60 ° C, placed for 1.5 h, 200 rpm / min, to obtain tobacco liquid.

(3) Fermented tobacco liquid

The composite microbial preparation was added to the tobacco liquid at a mass ratio of 5:100, and fermented at 37 ° C for 24 h. The fermented tobacco liquid was filtered with a layer of gauze to collect the fermentation broth and solid matter. The fermentation broth was concentrated 3 times under vacuum and the concentration was 50 °C.

The solid is dried and pressed to form a tobacco sheet, and then the fermentation liquid is coated on the tobacco sheet. When coated, the fermentation liquid is sufficient to wet the tobacco sheet to obtain a recombinant tobacco sheet.

Protein content test: The protein content in the tobacco liquid and the fermentation broth obtained after fermentation was tested by Coomassie brilliant blue staining. The protein content of the fermentation broth decreased by 7.16% after fermentation, indicating that the composite microorganism can effectively reduce the protein content in the tobacco waste. .

pH test: The pH value of the tobacco liquid and the fermentation liquid obtained after fermentation was tested by a pH meter. The pH value of the tobacco liquid was 6.32, and the pH value of the fermentation liquid was 4.63, indicating that the composite microorganism can effectively reduce the pH of the tobacco waste. value.

The prepared recombinant tobacco sheet was cut into a cigarette, and tested for aroma, sweetness, and softness.

Aroma test: Inviting 20 men with smoking habits (between 21 and 35 years old), taking 10 cigarettes, each at least 1 hour apart. In the process of sucking, the smell and taste of the smoke smell is pleasant and pleasant. The smoke aroma is purely 10 points, the smoke aroma is generally 5 points, and the completely smokeless aroma is 0 points. The scores of 20 people were counted, and the average value was calculated and recorded in Table 1.

Sweetness test: Inviting 20 men with smoking habits (between 21 and 35 years old) to smoke 10 cigarettes, each at least 1 hour apart. In the process of judging, the degree of the sweetness of the smoke is perceived by the sense of smell and taste. The sweetness of the smoke is 10 points, the sweetness of the smoke is generally 5 points, and the taste of sourness is 0 points. The scores of 20 people were counted, and the average value was calculated and recorded in Table 1.

Melting test: Inviting 20 men with smoking habits (between 21 and 35 years old), smoking 10 cigarettes, each at least 1 hour apart. In the process of sucking, the degree of softness of the smoke is perceived by the sense of smell and taste. The softness of the smoke is 10 points, the softness of the smoke is generally 5 points, and the taste is very spicy. The scores of 20 people were counted, and the average value was calculated and recorded in Table 1.

Example 2

The preparation process of the recombinant tobacco sheet of this embodiment is as follows:

(1) Preparation of a composite microbial preparation

Medium preparation: 10 g of brown sugar was added, 100 g of water was added, mixed, and sterilized at 100 ° C for 30 min to obtain a medium, and the medium was divided into three portions.

Preparation of Lactobacillus acidophilus activating solution: Lactobacillus acidophilus was inoculated into one of the above mediums, and cultured at 30 ° C for 36 hours.

Preparation of Saccharomyces Cerevisiae Activating Solution: Saccharomyces cerevisiae was inoculated into one of the above mediums and incubated at 40 ° C for 12 h.

Preparation of Bacillus subtilis activating solution: Bacillus subtilis was inoculated into one of the above mediums, and cultured at 30 ° C, 121 rpm / min for 36 h.

The Lactobacillus acidophilus activating solution, the Saccharomyces cerevisiae activating solution and the Bacillus subtilis activating solution are mixed in a ratio of the ratio of Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis per unit volume in a ratio of 1:0.8:0.4 to obtain a composite microbial preparation. .

(2) Preparation of tobacco liquid

Tobacco waste pretreatment: Tobacco liquid was obtained by mixing tobacco processing waste containing tobacco leaf fragments, tobacco stems and tobacco with water at a mass ratio of 1:10, water temperature of 70 ° C, and standing for 1 h at 150 rpm/min.

(3) Fermented tobacco liquid

The composite microbial preparation was added to the tobacco liquid at a mass ratio of 4:100, and fermented at 30 ° C for 48 h. The fermented tobacco liquid was filtered with a layer of gauze to collect the fermentation broth and solid matter. The fermentation broth was concentrated 5 times under vacuum and the concentration was 70 °C.

The solid is dried and pressed to form a tobacco sheet, and then the fermentation liquid is coated on the tobacco sheet. When coated, the fermentation liquid is sufficient to wet the tobacco sheet to obtain a recombinant tobacco sheet.

Protein content test: The protein content in the tobacco liquid and the fermentation broth obtained after fermentation was tested by Coomassie brilliant blue staining. The protein content of the fermentation broth decreased by 8.16% after fermentation, indicating that the composite microorganism can effectively reduce the protein content in the tobacco waste. .

pH test: The pH value of the tobacco liquid and the fermentation liquid obtained after fermentation were respectively measured by a pH meter. The pH value of the tobacco liquid was 6.32, and the pH value of the fermentation liquid was 4.33, indicating that the composite microorganism can effectively reduce the pH of the tobacco waste. value.

The prepared recombinant tobacco sheets were cut into cigarettes, and the aroma, sweetness and softness were tested in the same manner as in Example 1. The scores are shown in Table 1.

Example 3

The preparation process of the recombinant tobacco sheet of this embodiment is as follows:

(1) Preparation of a composite microbial preparation

Medium preparation: 10 g of brown sugar was added, 100 g of water was added, mixed, and sterilized at 100 ° C for 30 min to obtain a medium, and the medium was divided into three portions.

Preparation of Lactobacillus acidophilus activating solution: Lactobacillus acidophilus was inoculated into one of the above mediums, and cultured at 35 ° C for 30 hours.

Preparation of Saccharomyces Cerevisiae Activating Solution: Saccharomyces cerevisiae was inoculated into one of the above mediums and incubated at 37 ° C for 20 h.

Preparation of Bacillus subtilis activating solution: Bacillus subtilis was inoculated into one of the above-mentioned mediums, and cultured at 35 ° C, 121 rpm / min for 30 hours.

The Lactobacillus acidophilus activating solution, the Saccharomyces cerevisiae activating solution and the Bacillus subtilis activating solution are mixed in a ratio of the ratio of Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis per unit volume in a ratio of 1:1.0:0.5 to obtain a composite microbial preparation. .

(2) Preparation of tobacco liquid

Tobacco waste pretreatment: tobacco processing waste containing tobacco leaf fragments, stems and cigarettes and water at a mass ratio of 1:8, water temperature of 60 ° C, placed for 1.5 h, 200 rpm / min, to obtain tobacco liquid.

(3) Fermented tobacco liquid

The composite microbial preparation was added to the tobacco liquid at a mass ratio of 5:100, and fermented at 35 ° C for 36 h. The fermented tobacco liquid was filtered with a layer of gauze to collect the fermentation broth and solid matter. The fermentation broth was concentrated 4 times under vacuum and the concentration was 60 °C.

The solid is dried and pressed to form a tobacco sheet, and then the fermentation liquid is coated on the tobacco sheet. When coated, the fermentation liquid is sufficient to wet the tobacco sheet to obtain a recombinant tobacco sheet.

Protein content test: The protein content in the tobacco liquid and the fermentation broth obtained after fermentation was tested by Coomassie brilliant blue staining. The protein content of the fermentation broth decreased by 10.36% after fermentation, indicating that the composite microorganism can effectively reduce the protein content in the tobacco waste. .

pH test: The pH value of the tobacco liquid and the fermentation liquid obtained after fermentation were respectively measured by a pH meter. The pH value of the tobacco liquid was 6.32, and the pH value of the fermentation liquid was 4.13, indicating that the composite microorganism can effectively reduce the pH of the tobacco waste. value.

The prepared recombinant tobacco sheets were cut into cigarettes, and the aroma, sweetness and softness were tested in the same manner as in Example 1. The scores are shown in Table 1.

Example 4

The preparation process of the recombinant tobacco sheet of this embodiment is as follows:

(1) Preparation of a composite microbial preparation

Medium preparation: 10 g of brown sugar was added, 100 g of water was added, mixed, and sterilized at 100 ° C for 30 min to obtain a medium, and the medium was divided into three portions.

Preparation of Lactobacillus acidophilus activating solution: Lactobacillus acidophilus was inoculated into one of the above mediums, and cultured at 37 ° C for 28 hours.

Preparation of Saccharomyces Cerevisiae Activating Solution: Saccharomyces cerevisiae was inoculated into one of the above mediums and incubated at 37 ° C for 12 h.

Preparation of Bacillus subtilis activating solution: Bacillus subtilis was inoculated into one of the above mediums, and cultured at 30 ° C, 121 rpm / min for 36 h.

The Lactobacillus acidophilus activating solution, the Saccharomyces cerevisiae activating solution and the Bacillus subtilis activating solution are mixed in a ratio of the ratio of Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis per unit volume in a ratio of 1:1.2:0.6 to obtain a composite microbial preparation. .

(2) Preparation of tobacco liquid

Tobacco waste pretreatment: tobacco processing waste containing tobacco leaf fragments, stems and cigarettes and water at a mass ratio of 1:8, water temperature of 60 ° C, placed for 1.5 h, 200 rpm / min, to obtain tobacco liquid.

(3) Fermented tobacco liquid

The composite microbial preparation was added to the tobacco liquid at a mass ratio of 6:100, and fermented at 35 ° C for 36 h. The fermented tobacco liquid was filtered with a layer of gauze to collect the fermentation broth and solid matter. The fermentation broth was concentrated 3 times under vacuum and the concentration was 50 °C.

The solid is dried and pressed to form a tobacco sheet, and then the fermentation liquid is coated on the tobacco sheet. When coated, the fermentation liquid is sufficient to wet the tobacco sheet to obtain a recombinant tobacco sheet.

Protein content test: The protein content in the tobacco liquid and the fermentation broth obtained after fermentation was tested by Coomassie brilliant blue staining. The protein content of the fermentation broth decreased by 16.36% after fermentation, indicating that the composite microorganism can effectively reduce the protein content in the tobacco waste. .

pH test: the pH value of the tobacco liquid and the fermentation liquid obtained after fermentation were respectively measured by a pH meter. The pH value of the tobacco liquid was 6.32, and the pH value of the fermentation liquid was 4.10, indicating that the composite microorganism can effectively reduce the pH of the tobacco waste. value.

The prepared recombinant tobacco sheets were cut into cigarettes, and the aroma, sweetness and softness were tested in the same manner as in Example 1. The scores are shown in Table 1.

Example 5

The preparation process of the recombinant tobacco sheet of this embodiment is as follows:

(1) Preparation of a composite microbial preparation

Medium preparation: 10 g of brown sugar was added, 100 g of water was added, mixed, and sterilized at 100 ° C for 30 min to obtain a medium, and the medium was divided into three portions.

Preparation of Lactobacillus acidophilus activating solution: Lactobacillus acidophilus was inoculated into one of the above mediums, and cultured at 37 ° C for 24 hours.

Preparation of Saccharomyces Cerevisiae Activating Solution: Saccharomyces cerevisiae was inoculated into one of the above mediums and incubated at 37 ° C for 24 h.

Preparation of Bacillus subtilis activating solution: Bacillus subtilis was inoculated into one of the above mediums, and cultured at 30 ° C, 121 rpm / min for 28 h.

The Lactobacillus acidophilus activating solution, the Saccharomyces cerevisiae activating solution and the Bacillus subtilis activating solution are mixed in a ratio of the ratio of Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis per unit volume in a ratio of 1:0.4:0.3 to obtain a composite microbial preparation. .

(2) Preparation of tobacco liquid

Tobacco waste pretreatment: tobacco processing waste containing tobacco leaf fragments, stems and cigarettes and water at a mass ratio of 1:8, water temperature of 60 ° C, placed for 1.5 h, 200 rpm / min, to obtain tobacco liquid.

(3) Fermented tobacco liquid

The composite microbial preparation was added to the tobacco liquid at a mass ratio of 5:100, and fermented at 35 ° C for 36 h. The fermented tobacco liquid was filtered with a layer of gauze to collect the fermentation broth and solid matter. The fermentation broth was concentrated 3 times under vacuum and the concentration was 50 °C.

The solid is dried and pressed to form a tobacco sheet, and then the fermentation liquid is coated on the tobacco sheet. When coated, the fermentation liquid is sufficient to wet the tobacco sheet to obtain a recombinant tobacco sheet.

Protein content test: The protein content in the tobacco liquid and the fermentation broth obtained after fermentation was tested by Coomassie brilliant blue staining. The protein content of the fermentation broth decreased by 6.53% after fermentation, indicating that the composite microorganism can effectively reduce the protein content in the tobacco waste. .

pH test: The pH value of the tobacco liquid and the fermentation liquid obtained after fermentation were respectively measured by a pH meter. The pH value of the tobacco liquid was 6.32, and the pH value of the fermentation liquid was 4.50, indicating that the composite microorganism can effectively reduce the pH of the tobacco waste. value.

The prepared recombinant tobacco sheet was cut into a cigarette, and the same method as in Example 1 was carried out. Tests for aroma, sweetness and mildness are given in Table 1.

Comparative example 1

The preparation process of the recombinant tobacco sheet of Comparative Example 1 is substantially the same as that of the recombinant tobacco sheet of Example 3, except that the microorganism used in the step (3) fermentation of the tobacco liquid is the Lactobacillus acidophilus activating solution, and the The preparation method of the Lactobacillus acidophilus activating solution was the same as in Example 3, and the mass ratio of the Lactobacillus acidophilus activating solution to the tobacco liquid was 5:100.

The prepared recombinant tobacco sheet shreds were also processed into cigarettes, and the aroma, sweetness and softness were tested in the same manner as in Example 1, and the scores are shown in Table 1.

Comparative example 2

The preparation process of the recombinant tobacco sheet of Comparative Example 2 was substantially the same as that of the recombinant tobacco sheet of Example 3, except that the microorganism used in the step (3) fermentation of the tobacco liquid was Saccharomyces cerevisiae activation liquid, and the Saccharomyces cerevisiae was activated. The liquid was prepared in the same manner as in Example 3, and the mass ratio of the Saccharomyces cerevisiae activation liquid to the tobacco liquid was 5:100.

The prepared recombinant tobacco sheet shreds were also processed into cigarettes, and the test scores of the aroma, sweetness and softness in the same manner as in Example 1 are shown in Table 1.

Comparative example 3

The preparation process of the recombinant tobacco sheet of Comparative Example 3 was substantially the same as that of the recombinant tobacco sheet of Example 3, except that the microorganism used in the step (3) fermentation of the tobacco liquid was Bacillus subtilis activating liquid, Bacillus subtilis. The preparation method of the bacillus activating solution was the same as in Example 3, and the mass ratio of the Bacillus subtilis activating solution to the tobacco liquid was 5:100.

The prepared recombinant tobacco sheet shreds were also processed into cigarettes, and the test scores of the aroma, sweetness and softness in the same manner as in Example 1 are shown in Table 1.

Comparative example 4

The preparation process of the recombinant tobacco sheet of Comparative Example 4 was substantially the same as that of the recombinant tobacco sheet of Example 3, except that the microorganism used in the step (3) fermentation of the tobacco liquid was Saccharomyces cerevisiae activation liquid and Bacillus subtilis. A mixture of activating fluids. The preparation method of the Saccharomyces cerevisiae activating liquid and the Bacillus subtilis activating solution is the same as that of the third embodiment, and the brewer's yeast and the herbaceous buds are in a unit volume. The S. cerevisiae activation solution and the Bacillus subtilis activating solution were mixed at a ratio of the ratio of the number of the bacillus. The mass ratio of the mixture of the yeast activating solution and the Bacillus subtilis activating solution to the tobacco liquid was 5:100.

The prepared recombinant tobacco sheet shreds were also processed into cigarettes, and the test scores of the aroma, sweetness and softness in the same manner as in Example 1 are shown in Table 1.

Comparative example 5

The preparation process of the recombinant tobacco sheet of Comparative Example 5 was substantially the same as that of the recombinant tobacco sheet of Example 3, except that the microorganism used in the step (3) fermentation of the tobacco liquid was Lactobacillus acidophilus active liquid and dry grass. A mixture of Bacillus activation fluids. The preparation method of the Lactobacillus acidophilus activating solution and the Bacillus subtilis activating solution is the same as that of the third embodiment, and the Lactobacillus acidophilus activating solution and the ratio of the number of Lactobacillus acidophilus and Bacillus subtilis per unit volume are 1:0.5. Bacillus subtilis activating solution is mixed. The mass ratio of the mixture of the Lactobacillus acidophilus activating solution and the Bacillus subtilis activating solution to the tobacco liquid was 5:100.

The prepared recombinant tobacco sheet shreds were also processed into cigarettes, and the test scores of the aroma, sweetness and softness in the same manner as in Example 1 are shown in Table 1.

Comparative example 6

The preparation process of the recombinant tobacco sheet of Comparative Example 6 was substantially the same as that of the recombinant tobacco sheet of Example 3, except that: Comparative Example 6 Step (3) The microorganism used for fermenting the tobacco liquid was Lactobacillus acidophilus activation liquid and wine making. A mixture of yeast activating fluids. The Lactobacillus acidophilus activating solution and the Saccharomyces cerevisiae activating solution are prepared in the same manner as in Example 3, and the Lactobacillus acidophilus activating solution and the Saccharomyces cerevisiae are ratios in a ratio of 1:1 by the ratio of Lactobacillus acidophilus to Saccharomyces cerevisiae per unit volume. The activation solution is mixed. The mass ratio of the mixture of the Lactobacillus acidophilus activating solution and the yeast activating solution to the tobacco liquid was 5:100.

The prepared recombinant tobacco sheet shreds were also processed into cigarettes, and the test scores of the aroma, sweetness and softness in the same manner as in Example 1 are shown in Table 1.

Comparative example 7

The same tobacco waste of Example 3 was directly processed to form a cigarette, and the test scores of the aroma, sweetness and softness in the same manner as in Example 1 are shown in Table 1.

Comparative example 8

The test scores for the aroma, sweetness and softness of the same method as in Example 1 were obtained by cutting from a normal tobacco leaf into a cigarette.

Table 1 shows the scoring data of the aroma, sweetness and softness of the cigarettes of the recombinant tobacco sheet processed cigarettes of Examples 1 to 5 and Comparative Examples 1 to 8. The overall score is the average of aroma, sweetness and softness.

Table 1: Scoring data for aroma, sweetness and softness of cigarettes

	Aroma (minutes)	Sweet feeling (minutes)	Softness (minutes)	Comprehensive score (minutes)
Example 1	8.5	8.4	9.5	8.8
Example 2	8.6	8.8	9.6	9.0
Example 3	9.5	9.0	9.8	9.4
Example 4	9.1	8.5	8.5	8.7
Example 5	8.5	9.2	8.8	8.8
Comparative example 1	1.0	2.0	7.5	3.5
Comparative example 2	6.5	2.5	2.0	3.7
Comparative example 3	1.5	6.4	3.5	3.8
Comparative example 4	6.2	5.1	2.0	4.4
Comparative example 5	1.0	5.0	6.0	4.0
Comparative example 6	5.5	2.0	4.5	4.0
Comparative example 7	0.2	1.0	0.3	0.5
Comparative example 8	9.8	9.6	9.5	9.6

As can be seen from Table 1, the recombinant tobacco sheet processing cigarettes of Examples 1 to 5 have a score of at least 8.5, a sweetness score of at least 8.4, a softness score of at least 8.5, and a comprehensive score of at least 8.7. . Moreover, the aroma, sweetness and softness scores of the processed tobacco sheet processing cigarettes of Example 3 were 9.5, 9.0 and 9.8, respectively, and the overall score was 9.4, and the comparison ratio was normal. The aroma of cigarettes formed by tobacco processing is basically the same, and even the score of softness is better than the ratio of 8. For the cigarettes processed from untreated tobacco waste, the aroma, sweetness and softness scores were only 0.2, 1.0 and 0.3, with a composite score of 0.5. The aroma, sweetness, and softness of the processed tobacco sheet processing of Examples 1 to 5 were apparently due to Comparative Example 7.

In addition, the addition of only Lactobacillus acidophilus to the fermentation of tobacco liquid was compared with a softness score of 7.5, which was higher than that of Comparative Example 2 and Comparative Example 3 without the addition of Lactobacillus acidophilus, indicating the addition of Lactobacillus acidophilus to the smell of tobacco. The softness has a certain improvement effect.

Only the proportion 2 of the fermentation of tobacco liquid by Saccharomyces cerevisiae was added, and the aroma score was 6.5 points, which was higher than that of Comparative Example 1 and Comparative Example 3 without adding S. cerevisiae, indicating that the addition of Saccharomyces cerevisiae had a certain improvement effect on the aroma of smoke.

Only the proportion 3 of the fermentation of tobacco liquid by Bacillus subtilis was added, and the sweetness score was 6.4, which was higher than that of Comparative Example 1 and Comparative Example 2 without added Bacillus subtilis, indicating that the added sweetness of B. subtilis had a certain sweetness to smoke. Improvement.

The only difference from Example 3 was that Comparative Example 4 in which Lactobacillus acidophilus was not added, the scores of aroma, sweetness and softness were 6.2, 5.1 and 2.0, respectively.

The only difference from Example 3 was that Comparative Example 5 without the addition of Saccharomyces cerevisiae, the scores of aroma, sweetness and softness were 1.0, 5.0 and 6.0, respectively.

The only difference from Example 3 was that Comparative Example 6 was not added with Bacillus subtilis, and the scores of aroma, sweetness, and softness were 5.5, 2.0, and 4.5, respectively.

The aroma, sweetness and softness scores of Comparative Example 4 to Comparative Example 6 were all not lower than that of Example 3, indicating that Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis were compounded in a certain ratio, and the individual use could be improved. Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis improve the aroma, sweetness and softness, or the combination of any two combinations of Lactobacillus acidophilus, Saccharomyces cerevisiae and Bacillus subtilis for aroma, sweetness and softness , obtained the effect of 1+1>2.

The above-mentioned embodiments are merely illustrative of one or more embodiments of the present invention, and the description thereof is more specific and detailed, but is not to be construed as limiting the scope of the invention. It should be noted that those skilled in the art can also do without departing from the inventive concept. A number of variations and modifications are within the scope of the invention. Therefore, the scope of the invention should be determined by the appended claims.

Claims (17)

1. A method for preparing a recombinant tobacco sheet, comprising:

   Mixing the tobacco waste with the solvent at a mass ratio of 1:6-10 to obtain a tobacco liquid;

   Adding a composite microbial preparation to the tobacco liquid, and fermenting at 30 ° C to 37 ° C for 24 h to 48 h, and obtaining a fermentation liquid and a solid material after solid-liquid separation; wherein the composite microbial preparation contains Lactobacillus acidophilus and wine The ratio of the yeast and the Bacillus subtilis per unit volume of the Lactobacillus acidophilus, the Saccharomyces cerevisiae and the Bacillus subtilis is 1:0.4 to 1.2:0.3-0.6;

   Processing the solid to form a tobacco sheet;

   The tobacco sheet is wetted using the fermentation broth to obtain the recombinant tobacco sheet.
2. The method according to claim 1, wherein the composite microbial preparation is obtained by the following preparation method:

   Lactobacillus acidophilus is inoculated into a medium containing a saccharide compound having a mass fraction of 5% to 15%, and cultured at 30 ° C to 37 ° C for 24 h to 36 h to obtain a Lactobacillus acidophilus activating solution;

   The Saccharomyces cerevisiae is inoculated into a medium containing a saccharide compound having a mass fraction of 5% to 15%, and cultured at 30 ° C to 40 ° C for 12 h to 24 h to obtain a Saccharomyces cerevisiae activating solution;

   Bacillus subtilis is inoculated into a medium containing a saccharide compound having a mass fraction of 5% to 15%, and cultured at 30 ° C to 37 ° C for 24 h to 36 h to obtain a Bacillus subtilis activating solution;

   The Lactobacillus acidophilus activating solution, the Saccharomyces cerevisiae activating solution, and the Bacillus subtilis activating solution are mixed to obtain the composite microbial preparation.
3. The method according to claim 2, wherein said Lactobacillus acidophilus in said Lactobacillus acidophilus activating solution is in a logarithmic growth phase or a stable growth phase.
4. The method according to claim 2, wherein said Saccharomyces cerevisiae in said Saccharomyces cerevisiae activating solution is in a logarithmic growth phase or a stable growth phase.
5. The method according to claim 2, wherein said Bacillus subtilis in said Bacillus subtilis activating solution is in a logarithmic growth phase or a stable growth phase.
6. The method of claim 2 wherein the saccharide compound is brown sugar.
7. The method of claim 1 wherein said tobacco waste is selected from the group consisting of a tobacco stem, At least one of tobacco leaf fragments and smoke.
8. The method according to claim 1, wherein the mass ratio of the composite microbial preparation to the tobacco liquid is from 4 to 6:100.
9. The method of claim 1 further comprising the step of concentrating said fermentation broth prior to said step of wetting said tobacco sheet using said fermentation broth.
10. The method according to claim 9, wherein the step of concentrating the fermentation broth is specifically: concentrating the fermentation broth under vacuum conditions, the concentration of the concentration being from 50 ° C to 70 ° C.
11. The method according to claim 9, wherein the volume of the fermentation liquid before concentration is 3 to 5 times the volume of the fermentation liquid after concentration.
12. The method according to claim 1, wherein the step of wetting the tobacco sheet with the fermentation liquid is specifically: adding the fermentation liquid to the tobacco sheet by coating, spraying or dipping The fermentation broth is sufficient to wet the tobacco sheet.
13. The method according to claim 1, wherein in the step of mixing the tobacco waste with the solvent in a mass ratio of 1:6 to 10, the mixing is performed at a temperature of 50 ° C to 70 ° C. Stirring treatment for 1 h to 2 h.
14. The method according to claim 13, wherein the stirring speed of the stirring treatment is from 150 rpm/min to 250 rpm/min.
15. The method according to claim 1, wherein in the step of mixing the tobacco waste with the solvent in a mass ratio of 1:6 to 10, the solvent is water.
16. The method according to claim 1, wherein the operation of the solid-liquid separation is specifically: filtering the product after the fermentation through a sieve having a pore diameter of 0.5 μm to 5 μm.
17. A recombinant tobacco sheet produced by the method for producing a recombinant tobacco sheet according to any one of claims 1 to 16.

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