Method for fermenting cigar tobacco leaves by using mixtures of saprophytic staphylococcus and ganoderma lucidum extract and application thereof

Abstract

This invention discloses a method for fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract, and its application, belonging to the field of tobacco technology. The method includes the following steps: providing wrapper tobacco leaves; adding wet Staphylococcus saprophyticus cells and Ganoderma lucidum extract to the wrapper tobacco leaves and then fermenting them, turning the leaves during fermentation to obtain fermented tobacco leaves. This invention, by adding Staphylococcus saprophyticus and Ganoderma lucidum extract for synergistic fermentation during cigar wrapper fermentation, significantly increases chlorophyll and carotenoid degradation products in the middle stage of fermentation, and significantly accumulates phenylalanine conversion products and Maillard reaction products in the late stage of fermentation, thereby significantly promoting the production of aroma substances and enhancing the accumulation and harmony of aromatic substances. Furthermore, the fermentation method provided by this invention also effectively improves the color and appearance of the wrapper, providing a feasible path and theoretical support for optimizing the quality of cigar wrappers and developing high-end cigar products.

Description

Technical Field

[0001] This invention belongs to the field of tobacco technology, specifically relating to a method for fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract, and its application. Background Technology

[0002] As a high-end tobacco product, the quality of cigar tobacco leaves not only determines its economic value and market competitiveness but also profoundly influences consumers' acceptance and preference for cigars. This quality is reflected in multi-dimensional sensory experiences such as aroma and taste, and also stems from the complex and harmonious proportions of its chemical components. The wrapper, as the outer surface of the cigar, directly affects its overall color and smoking style; its color uniformity and depth are often considered primary standards for evaluating the quality of high-end cigars. Cigar tobacco leaves require fermentation to improve their quality. This process is subject to the synergistic effects of environmental factors and microbial communities, driving a profound transformation in the appearance, biochemical characteristics, and overall quality of the tobacco leaves, including deepening color, accumulating aroma substances, and reducing astringency. However, current domestic cigar wrapper production is still constrained by multiple factors, including varietal base, technical means, and fermentation conditions. Compared with imported wrappers, there is still a gap in aroma harmony and chemical balance, making it difficult to fully meet the market demand for high-end cigars.

[0003] In recent years, bio-fermentation has been regarded as an important means to overcome the bottleneck of cigar wrapper quality. Studies have shown that polyphenols in tobacco leaves during fermentation are not only important precursors to the formation of aromatic substances, but also key factors determining color changes. On the one hand, microorganisms and their secreted enzymes can promote the degradation and transformation of polyphenols, driving them to transform into phenolic acids, aldehydes, ketones, and heterocyclic aromatic compounds, thereby giving cigars a more mellow and complex aroma. On the other hand, these polyphenols are gradually oxidized to quinone compounds under enzymatic or non-enzymatic conditions, and then undergo polymerization reactions to generate brown substances, gradually deepening the color of the tobacco leaves. The dual mechanism of polyphenols establishes an intrinsic link between aroma enhancement and color deepening, becoming the core pathway for fermentation quality transformation. However, traditional natural fermentation often lacks stable and efficient functional microbial communities, resulting in limited improvement in aroma and color, which has also driven the exploration of the application of exogenous functional microorganisms in cigar fermentation.

[0004] Currently, no use of *Staphylococcus saprophyticus* (S. saprophyticus) has been observed. Staphylococcus saprophyticus ) and Ganoderma lucidum ( Ganoderma lucidum A research report on the mixed fermentation of cigar tobacco leaves with extracts. Summary of the Invention

[0005] The purpose of this invention is to provide a method and its application for fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract. This addresses the problem in existing technologies that lack the use of this mixture to improve the quality of cigar tobacco leaves.

[0006] In a first aspect, the present invention provides a method for fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract, comprising the following steps: providing cigar wrapper tobacco leaves; adding Staphylococcus saprophyticus wet cells and Ganoderma lucidum extract to the cigar wrapper tobacco leaves and then performing fermentation treatment, and turning the leaves over during fermentation to obtain fermented tobacco leaves; wherein, in the step of adding Staphylococcus saprophyticus wet cells and Ganoderma lucidum extract to the cigar wrapper tobacco leaves and then performing fermentation treatment, the amount of Staphylococcus saprophyticus wet cells added is 1-1.5% of the mass of the cigar wrapper tobacco leaves, and the amount of Ganoderma lucidum extract added is 0.5-1.5% of the mass of the cigar wrapper tobacco leaves.

[0007] In this invention, the inventors discovered that by adding Staphylococcus saprophyticus and Ganoderma lucidum extract for mixed synergistic fermentation during the cigar wrapper fermentation process, the degradation products of chlorophyll and carotenoids are significantly increased in the middle stage of fermentation, and phenylalanine conversion products and Maillard reaction products are also significantly accumulated in the late stage of fermentation. This significantly promotes the production of aroma substances and enhances the accumulation and harmony of aromatic substances. In addition, the fermentation method provided by this invention also effectively improves the color and appearance of the wrapper, providing a feasible path and theoretical support for the quality optimization of cigar wrappers and the development of high-end cigar products.

[0008] In some implementations, the moisture content of the shroud tobacco leaves is 29-31% during the step of providing the shroud tobacco leaves.

[0009] In some implementations, the fermentation process, which involves adding wet Staphylococcus saprophyticus cells and Ganoderma lucidum extract to the tobacco leaves and then turning them over during fermentation, specifically includes fermenting for 23-27 days at a temperature of 36-38°C.

[0010] In some implementation schemes, the turning process specifically includes: first turning the fermentation chamber on day 9-11, then turning it on day 16-18, and finally turning it on day 24-26.

[0011] In a second aspect, the present invention provides the application of the method of fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract as described above in any of the following: A1) increasing the content of aroma precursors in tobacco leaves; A2) increasing the content of aroma substances in tobacco leaves.

[0012] In some embodiments, A1), the tobacco aroma precursors include at least one of chlorophyll degradation products, carotenoid degradation products, phenylalanine conversion products, and Maillard reaction products.

[0013] In some embodiments, the carotenoid degradation products include at least one of dihydroactinidone and farnesylacetone; and / or, the phenylalanine conversion products include at least one of phenylacetaldehyde and phenylethanol; and / or, the Maillard reaction products include at least one of furfural and furfuryl alcohol.

[0014] In some embodiments, A2), the aroma compounds of tobacco leaves include at least one of neophytadiene, phytone, phenylacetaldehyde, phenylethanol, and indole.

[0015] In a third aspect, the present invention provides the application of the method of fermenting cigar tobacco leaves with a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract as described above in improving the sensory quality of tobacco leaves; wherein the sensory quality of tobacco leaves includes at least one of aroma quality, aroma intensity, aftertaste, off-flavors, sweetness, irritation, and combustibility.

[0016] In a fourth aspect, the present invention provides the application of the method of fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract as described above in improving the color of tobacco leaves; wherein improving the color of tobacco leaves includes deepening the color of tobacco leaves.

[0017] The beneficial effects of this invention are as follows: Unlike existing technologies, this invention incorporates Staphylococcus saprophyticus and Ganoderma lucidum extract into the cigar wrapper fermentation process for synergistic fermentation. This significantly increases the degradation products of chlorophyll and carotenoids in the middle stage of fermentation, and also significantly accumulates phenylalanine conversion products and Maillard reaction products in the final stage of fermentation. This, in turn, significantly promotes the production of aroma substances and enhances the accumulation and harmony of aromatic compounds. Furthermore, the fermentation method provided by this invention effectively improves the color and appearance of the wrapper, offering a feasible path and theoretical support for optimizing cigar wrapper quality and developing high-end cigar products. Attached Figure Description

[0018] Figure 1 This is a bar chart showing the aroma compounds of cigar wrapper tobacco leaves from different treatment groups during the fermentation process in performance test 1 of this invention. Figure 2 This is a heat map showing the changes in aroma compounds of cigar wrapper tobacco leaves in different treatment groups during the fermentation process in performance test 1 of this invention. Figure 3A The lightness value L of the cigar wrapper tobacco leaves in different treatment groups in performance test 3 of this invention. Result of the change; Figure 3B The redness value 'a' of the cigar wrapper tobacco leaves in different treatment groups in performance test 3 of this invention. Result of the change; Figure 3C The yellowness value b of the cigar wrapper tobacco leaves in different treatment groups in performance test 3 of this invention. Result of the change; Figure 3D The results show the changes in the total color difference ΔE of the cigar wrapper tobacco leaves in different treatment groups during performance test 3 of this invention. Detailed Implementation

[0019] The technical solutions of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0020] Experimental methods not specified in the examples are generally performed under conventional conditions and as described in the manual, or as recommended by the manufacturer. Unless otherwise specified, the general equipment, materials, reagents, etc. used are commercially available.

[0021] Currently, existing technologies lack the ability to improve the quality of cigar tobacco by using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract for fermentation.

[0022] To address the lack of existing technologies that utilize a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract in the fermentation of cigar tobacco leaves to improve tobacco quality, this invention provides a method for fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract, and its application.

[0023] In a first aspect, the present invention provides a method for fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract, comprising the following steps: providing cigar wrapper tobacco leaves; adding Staphylococcus saprophyticus wet cells and Ganoderma lucidum extract to the cigar wrapper tobacco leaves and then performing fermentation treatment, and turning the leaves over during fermentation to obtain fermented tobacco leaves; wherein, in the step of adding Staphylococcus saprophyticus wet cells and Ganoderma lucidum extract to the cigar wrapper tobacco leaves and then performing fermentation treatment, the amount of Staphylococcus saprophyticus wet cells added is 1-1.5% of the mass of the cigar wrapper tobacco leaves, preferably 1.3%; the amount of Ganoderma lucidum extract added is 0.5-1.5% of the mass of the cigar wrapper tobacco leaves, preferably 1%.

[0024] The method provided by this invention involves the addition of Staphylococcus saprophyticus and Ganoderma lucidum extract for synergistic fermentation during the cigar wrapper fermentation process. This significantly increases the degradation products of chlorophyll and carotenoids in the middle stage of fermentation, and also significantly accumulates phenylalanine conversion products and Maillard reaction products in the late stage of fermentation. This significantly promotes the production of aroma substances and enhances the accumulation and harmony of aromatic substances. In addition, the fermentation method provided by this invention also effectively improves the color and appearance of the wrapper, thus providing a new process idea for the application of domestically produced wrappers in the production of high-end cigars.

[0025] In some implementations, the moisture content of the shroud tobacco leaves is 29-31% during the step of providing the shroud tobacco leaves.

[0026] In this invention, by controlling the moisture content of the tobacco leaves within a specific range, it is easier to ferment the tobacco leaves in the subsequent process.

[0027] In some implementations, the fermentation process, which involves adding wet Staphylococcus saprophyticus cells and Ganoderma lucidum extract to the tobacco leaves and then turning the leaves over during fermentation, specifically includes fermenting for 23-27 days, preferably 25 days, at a temperature of 36-38°C (preferably 37°C).

[0028] In this invention, by controlling the parameters in the fermentation process within a specific range, Staphylococcus saprophyticus, Ganoderma lucidum extract, and tobacco leaves can be fully fermented, significantly improving the quality of the tobacco leaves.

[0029] In some implementation schemes, the turning process specifically includes: first turning the fermentation chamber on day 9-11, then turning it on day 16-18, and finally turning it on day 24-26.

[0030] In this invention, by employing a turning-over process, the fermentation process can be precisely controlled, thereby significantly improving the quality of tobacco leaves.

[0031] It is understandable that the method of opening the box can be selected according to actual usage needs. For example, in this invention, the opening of the box follows the principle of "from top to bottom and from outside to inside".

[0032] In a second aspect, the present invention provides the application of the method of fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract as described above in any of the following: A1) increasing the content of aroma precursors in tobacco leaves; A2) increasing the content of aroma substances in tobacco leaves.

[0033] In this invention, serotonin and Ganoderma lucidum extract are used to synergistically ferment cigar tobacco leaves, which can significantly promote the production of aroma substances, making the tobacco leaves more balanced in terms of sweetness, floral aroma and roasting aroma, thereby significantly improving the quality of the tobacco leaves.

[0034] In some embodiments, A1), the tobacco aroma precursors include at least one of chlorophyll degradation products, carotenoid degradation products, phenylalanine conversion products, and Maillard reaction products.

[0035] In some embodiments, the carotenoid degradation products include at least one of dihydroactinidone and farnesylacetone; and / or, the phenylalanine conversion products include at least one of phenylacetaldehyde and phenylethanol; and / or, the Maillard reaction products include at least one of furfural and furfuryl alcohol.

[0036] In this invention, dihydroactinol and farnesylacetone in the carotenoid degradation products impart distinct floral and fruity aromas to cigar tobacco leaves; phenylacetaldehyde and phenylethanol in the phenylalanine conversion products enhance the sweet and floral aromas, improving the smoothness and harmony of the tobacco leaves; furfural and furfuryl alcohol in the Maillard reaction products impart nutty and roasted aromas to the tobacco leaves.

[0037] In some embodiments, A2), the aroma compounds of tobacco leaves include at least one of neophytadiene, phytone, phenylacetaldehyde, phenylethanol, and indole.

[0038] In this invention, neophytadiene imparts a woody aroma and mellowness to tobacco leaves, while indole imparts a jasmine aroma to tobacco leaves.

[0039] In a third aspect, the present invention provides the application of the method of fermenting cigar tobacco leaves with a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract as described above in improving the sensory quality of tobacco leaves; wherein the sensory quality of tobacco leaves includes at least one of aroma quality, aroma intensity, aftertaste, off-flavors, sweetness, irritation, and combustibility.

[0040] In this invention, the co-fermentation of Staphylococcus saprophyticus and Ganoderma lucidum extract not only significantly enhances the depth and sweetness of the aroma, but also shows a significant improvement in the overall sensory quality.

[0041] In a fourth aspect, the present invention provides the application of the method of fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract as described above in improving the color of tobacco leaves; wherein improving the color of tobacco leaves includes deepening the color of tobacco leaves.

[0042] In this invention, Staphylococcus saprophyticus and Ganoderma lucidum extract are used for synergistic fermentation, which makes the tobacco leaves darker and more distinct, further improving the appearance quality of the cigar wrapper.

[0043] In this invention, the CX-14 cigar wrapper tobacco used in the experiment was sourced from Enshi, Hubei Province, and supplied by the Enshi cigar tobacco production base. (Ganoderma lucidum) Ganoderma lucidum The extract was purchased from Hai'aosi Herbal Essence Workshop.

[0044] Saprophytic Staphylococcus ( Staphylococcus saprophyticus(See the literature (Yao L, Zhao Z, Li L, et al. The impact of Staphylococcus saprophyticus on the fermentation of cigarette filler tobacco leaves and the dynamics of microbial community[J]. Frontiers in Bioengineering and Biotechnology, 2025, 13(13):1666879.DOI:10.3389 / fbioe.2025.1666879.), which can be obtained by the public from Hubei Provincial Tobacco Science Research Institute.)

[0045] In this invention, the volatile aroma compounds of cigar tobacco leaves were determined as follows: referring to the method in the literature (Ding Jingyi, Yu Jun, Yang Chunlei, et al. Effect of yeast fermentation of tobacco flower bud extract on cigar tobacco leaf fermentation. 2024), the volatile aroma components of fermented cigar tobacco leaves were determined by simultaneous distillation extraction combined with GC-MS.

[0046] In this invention, the color parameters are measured as follows: An HP-C210 colorimeter is used for colorimetric measurement. The measurement location is selected on the line connecting the midpoint between the leaf margin and the midrib on one side of the tobacco leaf. Measurements are taken at 1 / 3, 1 / 2, and 2 / 3 of the leaf length, respectively. A total of 5 points are measured on both sides of each tobacco leaf to ensure the comprehensiveness and accuracy of the data. During the measurement process, the tobacco leaf must be laid flat on the backing plate with the front side facing up, ensuring it is flat and wrinkle-free to guarantee measurement accuracy. The colorimetric indicators measured mainly include L... Value (brightness value, the higher the value, the brighter), a Value (redness value; the higher the value, the more reddish; a negative value indicates a more greenish hue), b The values ​​(yellowness value, the higher the value, the more yellow; negative values ​​indicate a bluish tint) and ΔE (total color difference value, the higher the value, the more significant the difference from the backing) are used to evaluate the color characteristics of tobacco leaves. ; Where: L0 a0 b0 These are the back panel color parameters, where L0 =90.6、a0 =-0.62、b0 =1.73.

[0047] In this invention, the sensory evaluation is as follows: tobacco leaf samples are hand-rolled into cigars, and their sensory quality is scored by experts from the Enshi Prefecture cigar tobacco production base in Hubei Province. A nine-point scoring system (see Table 1 below) is used, involving eight main indicators: aroma quality, aroma intensity, aftertaste, off-flavors, sweetness, irritation, combustibility, and ash color, to comprehensively evaluate the sensory quality of the tobacco leaves. The higher the score, the better the sensory quality of the tobacco leaves.

[0048] Table 1 Sensory Quality Evaluation Criteria for Cigar Tobacco Leaves

[0049] In this invention, data analysis was performed as follows: Chromatograms were analyzed using GC-MS Solution software (Agilent Technologies, USA). Mass spectrometry data were compared with mass spectra in the NIST reference library (NIST 14) within the GC-MS data system to identify volatile compounds. Differences in microbial communities before and after fermentation were processed using the bioinformatics analysis cloud platform of Beijing Novogene Technology Co., Ltd. (https: / / magic.novogene.com / ); changes in aroma substances, chemical components, and color parameters were processed using the CNSknowall platform (https: / / cnsknowall.com). Significance analysis was performed using Statistica 23.0 software (SPSS Inc., Chicago, IL, USA).

[0050] In this invention, the preparation of wet Staphylococcus saprophyticus cells includes the following steps: Take 1 mL of preserved saprophytic Staphylococcus ( Staphylococcus saprophyticus The bacterial culture was inoculated into liquid LB medium and cultured with shaking at 200 rpm for 12 h to obtain the seed culture. The seed culture was then transferred to a fermenter for large-scale culture. The bacterial culture was centrifuged, the supernatant was discarded, the bacterial precipitate was collected and stored at 4°C for later use.

[0051] Example 1 This embodiment provides a method for fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract, including the following steps: S1. Place 100kg of eggplant-wrapped tobacco leaves with a moisture content of 29-31% into a fermentation box (2m×1m×1m); S2. Continue to add 1.3% of the wetted Staphylococcus saprophyticus cells and 1% of the Ganoderma lucidum extract by weight of the tobacco leaves to the fermentation chamber, and ferment for 25 days at a temperature of 37℃. First, turn the chamber over on the 10th day of fermentation, then turn it over on the 17th day of fermentation, and finally turn it over on the 25th day. Turning the chamber over follows the principle of "from top to bottom and from outside to inside". Set up 3 biological replicates to obtain the fermented tobacco leaves.

[0052] Among them, tobacco leaf samples were randomly collected after three rounds of turning over the box, and were marked as JZ1-JZ3 respectively, for subsequent index detection and analysis.

[0053] Comparative Example 1 In this comparative example, the method of fermenting cigar tobacco leaves is basically the same as in Example 1, except that the Staphylococcus saprophyticus and Ganoderma lucidum extract are replaced with an equal amount of water.

[0054] Among them, tobacco leaf samples were randomly collected before fermentation and after three turnings in the box, and were recorded as CK0-CK3 respectively.

[0055] Comparative Example 2 In this comparative example, the method of fermenting cigar tobacco leaves is basically the same as in Example 1, except that only wet Staphylococcus saprophyticus cells are added.

[0056] Among them, tobacco leaf samples were randomly collected after three rounds of turning over the box, and were recorded as J1-J3 respectively.

[0057] Comparative Example 3 In this comparative example, the method of fermenting cigar tobacco leaves is basically the same as in Example 1, except that only Ganoderma lucidum extract is added.

[0058] Among them, tobacco leaf samples were randomly collected after three rounds of turning over the box, and were recorded as Z1-Z3 respectively.

[0059] Performance Test 1 The aroma of cigar wrapper tobacco is mainly composed of various volatile substances, including alcohols, aldehydes, esters, and heterocyclic compounds. These compounds mostly originate from the degradation and transformation processes of chlorophyll, carotenoids, phenylalanine, sipranones, and sugar-amino compounds.

[0060] Based on this, the aroma compounds in the tobacco leaf samples collected in Example 1 and Comparative Examples 1-3 were tested, and the results are as follows: Figure 1 As shown.

[0061] from Figure 1As can be seen, in terms of aroma precursor conversion, chlorophyll degradation products accounted for the main proportion in all four groups, with the highest accumulation in group JZ, especially reaching 1557.57 μg / g in stage JZ-2, which is nearly 69% higher than CK-2. Carotenoid degradation products were significantly increased in groups Z and JZ, reaching 175.31 μg / g in stage JZ-2, which is nearly 77% higher than the control group. Dihydroactinolone and farnesylacetone were the main contributors, giving cigar tobacco leaves distinct floral and fruity aroma characteristics.

[0062] Phenylalanine conversion products accumulated significantly in stage JZ-3 (90.50 μg / g), an increase of 154% compared to CK-3. Representative substances, phenylacetaldehyde and phenylethanol, enhanced sweet and floral aromas, improving the smoothness and harmony of the tobacco leaves. Meanwhile, Maillard reaction products increased significantly in stage JZ-1, indicating that the addition of exogenous microorganisms may have accelerated the glycoamine reaction, generating furfural, furfuryl alcohol, and other substances that impart nutty and roasted aromas to the tobacco leaves.

[0063] Furthermore, the heat map of the changes in aroma substances in the above-mentioned tobacco leaf samples is as follows: Figure 2 As shown.

[0064] from Figure 2 As can be seen, the levels of neophytadiene, phytone, and phenylacetaldehyde were significantly increased in the JZ group. Neophytadiene had the highest content in the JZ-3 stage (1252.35 μg / g), becoming an important source of woody aroma and fullness. Phenylene alcohol accumulation in the JZ-3 stage was also significantly higher than in other groups, adding floral and sweet aromas to the cigar tobacco. Notably, indole also accumulated significantly in the JZ group, providing a unique jasmine aroma to the tobacco. In contrast, the accumulation of these substances was limited in the control group, indicating that natural fermentation has a limited effect on enhancing the aroma of tobacco.

[0065] In summary, the combined fermentation of Ganoderma lucidum extract and Staphylococcus saprophyticus not only promoted the degradation of chlorophyll and carotenoids, reduced grassy and unpleasant odors, and released floral and fruity aromas, but also enhanced phenylalanine conversion and Maillard reactions, significantly increasing the accumulation of sweet, nutty, and roasted aroma compounds. Particularly in the later stages of fermentation, group JZ showed an advantage in the content of multiple characteristic aroma compounds, resulting in a richer, more harmonious, and longer-lasting aroma in the cigar wrapper tobacco, providing strong support for its high-end quality.

[0066] Performance Test 2 Sensory evaluations were performed on the tobacco leaf samples collected in Example 1 and Comparative Examples 1-3, and the results are shown in Table 2 below.

[0067] Table 2 Sensory evaluation scores of cigar wrapper tobacco samples

[0068] As shown in Table 2, the pre-fermentation sample group (CK-0), serving as the baseline, exhibited weak aroma quality, insufficient aroma quantity, insignificant sweetness, and a moderate grayness. Specifically, the pre-fermentation sample group (CK-0) scored 30 points, with aroma quantity at 4 points, aroma quantity at 3 points, sweetness at 2 points, and grayness at 6 points. This indicates that unfermented cigar wrapper tobacco is generally rather bland in terms of sensory experience.

[0069] In contrast, the fermentation treatment groups (J-1, J-2, J-3) with added Staphylococcus saprophyticus showed significantly improved sensory quality. For example, in group J-3, aroma quality, aroma intensity, aftertaste, and sweetness were all significantly improved. Aroma quality and aroma intensity scores were 7 and 6 respectively, significantly higher than the 4 and 3 of the unfermented group, indicating that the addition of Staphylococcus saprophyticus promoted the enhancement of aroma and sweetness. The overall score was 52 points, an increase of 22 points compared to the unfermented group (30 points), indicating that Staphylococcus saprophyticus has a positive impact on the aroma and taste of cigar wrapper tobacco.

[0070] The fermentation treatment groups (Z-1, Z-2, and Z-3) with only Ganoderma lucidum extract added also showed significant improvements. The overall score of group Z-3 was 54 points, an increase of 24 points compared to the unfermented sample group (30 points). Its aroma quality, aroma intensity, and sweetness scores were 7, 6, and 7 respectively, all significantly higher than the unfermented group. The addition of Ganoderma lucidum extract further enhanced the depth and sweetness of the aroma, particularly in the gray tone, demonstrating the beneficial effect of Ganoderma lucidum on the sensory quality of tobacco leaves.

[0071] Among the mixed fermentation treatment groups (JZ-1, JZ-2, and JZ-3) of Staphylococcus saprophyticus and Ganoderma lucidum extract, the sensory quality performance was the best. Specifically, the overall score of group JZ-3 was 56 points, an increase of 26 points compared to the unfermented sample group (30 points), with aroma intensity and aroma quantity of 8 and 7 respectively, significantly better than other groups. In addition, group JZ-3 also performed well in terms of sweetness and grayness, scoring 7 and 6 respectively, further demonstrating the synergistic effect of mixed fermentation. The mixed fermentation treatment not only significantly enhanced the depth and sweetness of the aroma, but also showed a significant improvement in overall sensory quality.

[0072] The above results indicate that the synergistic effect of Staphylococcus saprophyticus and Ganoderma lucidum extract is of great significance in enhancing the aroma, taste, and visual perception of cigar wrapper tobacco.

[0073] Performance Test 3 The color parameters of the tobacco leaf samples collected in Example 1 and Comparative Examples 1-3 were measured, and the results are as follows: Figures 3A-3D As shown.

[0074] from Figures 3A-3D As can be seen from this, L in all treatment groups a and b The value continuously decreases during fermentation, indicating that the color of the tobacco leaves gradually deepens; the value continuously increases, reflecting that the color difference intensifies as fermentation progresses.

[0075] During the third turning of the naturally fermented group (CK), L a and b The values ​​decreased by 6.1%, 9.4%, and 16.4% respectively compared to the initial values, while the ΔE value increased by 4.8%, indicating that natural fermentation can promote color deepening, but the effect is limited. Compared with the CK group, the addition of Staphylococcus saprophyticus (J group) or Ganoderma lucidum extract (Z group) significantly enhanced the browning of tobacco leaves. In the J group, L... a and b The values ​​decreased by 10.1%, 11.2%, and 20.0% respectively, while the ΔE value increased by 3.4%. The decrease was even greater in group Z, with values ​​of 12.9%, 15.1%, and 23.5% respectively, and the ΔE value increased by 4.8%, indicating that Ganoderma lucidum extract was more effective in promoting pigment degradation and transformation.

[0076] The color change was most significant in the mixed fermentation group (JZ). During the third turning of the fermentation tank, L... a and b Compared with before fermentation, the values ​​decreased by 18.5%, 15.6%, and 27.0%, respectively, while the ΔE value increased by 8.9%. Among them, the enzymatic hydrolysis of Ganoderma lucidum extract can accelerate the oxidation of polyphenols into quinones and further polymerize to generate brown substances, while the strong carbon and nitrogen metabolism of Staphylococcus saprophyticus provides additional substrates, accelerates the Maillard reaction, and deepens the browning degree. Therefore, the synergistic effect of the two not only significantly promotes the transformation of aroma, but also makes the tobacco leaf color darker and more distinct, providing direct evidence for the improvement of the appearance quality of cigar wrappers.

[0077] In summary, this invention constructed a mixed fermentation system of *Staphylococcus saprophyticus* and *Ganoderma lucidum* extract, and systematically verified its role in improving the quality of cigar wrappers. This system significantly promotes the production of aroma compounds, with the content of phenylalanine derivatives significantly increasing in the later stages of fermentation compared to the control group, accompanied by the earlier accumulation of Maillard products, resulting in a more balanced presentation of sweet, floral, and roasted aromas in the tobacco leaves. Simultaneously, changes in color parameters indicate that mixed fermentation significantly accelerated the degradation of chlorophyll and carotenoids, with L... a b The decrease in ΔE value and the increase in ΔE value were both greater than those of the single treatment group, reflecting a deeper and more consistent color. Sensory evaluation further confirmed this trend, with the mixed fermentation treatment achieving the highest scores in key indicators such as aroma quality and aroma quantity, and an overall score increase of nearly 26 points compared to before fermentation. Therefore, the synergistic effect of Staphylococcus saprophyticus and Ganoderma lucidum extract can not only effectively enhance the aroma complexity of the wrapper but also improve its color, thus providing a new technological approach for the application of domestically produced wrappers in the production of high-end cigars.

[0078] It should be noted that all the above embodiments belong to the same inventive concept, and the descriptions of each embodiment have different focuses. Where the description in a particular embodiment is not detailed, please refer to the description in other embodiments.

[0079] The embodiments described above are merely illustrative of implementation methods of the present invention, and while the descriptions are specific and detailed, they should not be construed as limiting the scope of the invention patent. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these all fall within the protection scope of the present invention. Therefore, the protection scope of this invention patent should be determined by the appended claims.

Claims

1. A method for fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract, characterized in that, Includes the following steps: Provide shredded tobacco leaves; After adding Staphylococcus saprophyticus wet cells and Ganoderma lucidum extract to the eggplant-coated tobacco leaves, fermentation was carried out, and the leaves were turned over during fermentation to obtain fermented tobacco leaves. In the step of adding Staphylococcus saprophyticus wet cells and Ganoderma lucidum extract to the eggplant leaves and then performing fermentation treatment, the amount of Staphylococcus saprophyticus wet cells added is 1-1.5% of the mass of the eggplant leaves, and the amount of Ganoderma lucidum extract added is 0.5-1.5% of the mass of the eggplant leaves.

2. The method for fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract according to claim 1, characterized in that, In the step of providing the veneer tobacco leaves, the moisture content of the veneer tobacco leaves is 29-31%.

3. The method for fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract according to claim 1, characterized in that, The fermentation process, which involves adding wet Staphylococcus saprophyticus cells and Ganoderma lucidum extract to the tobacco leaves and then performing a turning process during fermentation, specifically includes fermenting for 23-27 days at a temperature of 36-38℃.

4. The method for fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract according to claim 3, characterized in that, The fermentation process specifically includes: first, a first fermentation is carried out on day 9-11 of fermentation; then, a second fermentation is carried out on day 16-18 of fermentation; and finally, a third fermentation is carried out on day 24-26 of fermentation.

5. The method of fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract as described in any one of claims 1-4, in any of the following: A1) Increase the content of aroma precursors in tobacco leaves; A2) Increase the content of aroma substances in tobacco leaves.

6. The application according to claim 5, characterized in that, In A1), the tobacco aroma precursors include at least one of chlorophyll degradation products, carotenoid degradation products, phenylalanine conversion products, and Maillard reaction products.

7. The application according to claim 6, characterized in that, The carotenoid degradation products include at least one of dihydroactinolone and farnesylacetone; and / or, The phenylalanine conversion product includes at least one of phenylacetaldehyde and phenylethanol; and / or, The Maillard reaction products include at least one of furfural and furfuryl alcohol.

8. The application according to claim 5, characterized in that, In A2), the aroma substances of tobacco leaves include at least one of neophytadiene, phytone, phenylacetaldehyde, phenylethanol, and indole.

9. The method of fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract as described in any one of claims 1-4 in improving the sensory quality of tobacco leaves; in, The sensory quality of the tobacco leaves includes at least one of the following: aroma quality, aroma intensity, aftertaste, off-flavors, sweetness, irritation, and combustibility.

10. The method of fermenting cigar tobacco leaves using a mixture of Staphylococcus saprophyticus and Ganoderma lucidum extract as described in any one of claims 1-4, in improving the color of tobacco leaves; in, The improvement of tobacco leaf color includes deepening the color of the tobacco leaves.

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