Solid loquat spice as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of tobacco flavors, and particularly relates to a solid loquat flavor as well as a preparation method and application thereof. The preparation method comprises the following steps: S1, preparing a loquat leach liquor; s2, adding a trichoderma harzianum suspension into the loquat leach liquor obtained in the S1 for fermentation, and adding a pichia pastoris suspension for fermentation to obtain a loquat concentrate; s3, mixing the loquat concentrate, amino acid and reducing sugar, and carrying out a Maillard reaction to obtain a liquid loquat spice; s4, preparing the liquid loquat spices into solid loquat spices; the trichoderma harzianum and the pichia pastoris which are added according to a specific sequence have a synergistic effect, the loquat extract can be fermented under the condition that an organic solvent is not added, the cost is low, the operation is simple, and the application prospect is wide; the prepared solid spice is convenient to transport and store.

Description

Technical Field

[0001] The invention belongs to the technical field of tobacco flavors, and particularly relates to a solid loquat flavor and a preparation method and application thereof. Background Art

[0002] In order to enhance the unique style of cigarettes or improve the sensory quality of low-tar cigarettes, cigarette flavoring has become a conventional technology in cigarette production. At present, many tobacco companies have explored and utilized a variety of natural flavors to enrich the aroma of cigarettes. These flavors mainly exist in various forms such as essential oils, extracts and tinctures. However, these flavors are usually liquid at room temperature, with a low boiling point and high volatility, which makes them susceptible to loss during transportation and storage. Therefore, exploring and developing high-quality solid flavors to reduce volatilization losses is of great significance to improving the overall quality of cigarettes.

[0003] Loquat, also known as Luju, Jinwan, etc., belongs to the genus Eriobotrya of the Rosaceae family. It is a common fruit tree in subtropical areas and has high economic and medicinal value. Using natural loquat as a flavoring, while retaining its unique aroma and incorporating new beneficial aroma components, can effectively improve the smoking quality of tobacco products and has good application prospects.

[0004] Microbial fermentation combined with thermal reaction to prepare spices is a method for producing spices that combines biotechnology and chemical technology. This method uses the biotransformation ability of microorganisms to produce spice precursors or intermediates through the fermentation process, and then further transforms and optimizes the chemical structure of the spices through thermal reactions to obtain the desired spice compounds. However, the existing fermentation method requires fermentation in an organic solvent, which has the problem of not being able to completely retain the characteristic aroma of loquat after fermentation and difficulty in removing the organic solvent. Summary of the invention

[0005] Therefore, the technical problem to be solved by the present invention is to overcome the defects of the existing fermentation technology that an organic solvent is required and the aroma of loquat cannot be completely retained after fermentation, thereby providing a solid loquat flavor and a preparation method and application thereof.

[0006] To this end, the present invention provides the following technical solutions:

[0007] The first aspect of the present invention protects a method for preparing solid loquat flavor, wherein the preparation method comprises the following steps:

[0008] S1, extracting loquat with water to obtain loquat extract;

[0009] S2, activating Trichoderma harzianum to obtain a Trichoderma harzianum suspension, activating Pichia pastoris to obtain a Pichia pastoris suspension, adding the Trichoderma harzianum suspension to the loquat extract of S1, performing a first fermentation, adding the Pichia pastoris suspension, performing a second fermentation, filtering, and concentrating to obtain a loquat concentrate;

[0010] S3, mixing loquat concentrate, amino acids and reducing sugars, and performing Maillard reaction to obtain liquid loquat flavor;

[0011] S4, preparing the liquid loquat flavor into solid loquat flavor.

[0012] In the present invention, the loquat is a conventional loquat in the art, typically but not limitingly, fresh loquat is used, and in a specific experiment, the loquat is cleaned and impurities removed (core removed and damaged); the water is conventional water, typically but not limitingly, the water is deionized water.

[0013] In the present invention, after extraction in S1, filtration is used for solid-liquid separation, and the filtration is conventional filtration in the art. Typically, but not limitedly, 50-mesh nylon filter cloth is used for filtration to obtain loquat extract.

[0014] According to the present invention, in S1, the mass ratio of water to loquat is 5-7:1.

[0015] According to the present invention, the extraction temperature is 60-75°C, the extraction time is 1-2h, and the stirring speed during extraction is 500-700rpm.

[0016] According to the present invention, based on the mass of the loquat extract, the mass of the Trichoderma harzianum suspension is 0.1-1wt%, and the mass of the Pichia pastoris suspension is 1-3wt%.

[0017] According to the present invention, based on the mass of the loquat extract, the mass of the Trichoderma harzianum suspension is 0.5-0.8wt%, and the mass of the Pichia pastoris suspension is 1.5-2.5wt%.

[0018] According to the present invention, the specific steps of activating the Trichoderma harzianum are as follows: inoculating the Trichoderma harzianum onto a seed culture medium, with the mass of the Trichoderma harzianum being 1-3wt% based on the mass of the seed culture medium, performing a first activation at 25-30°C for 18-24h, centrifuging the product after the first activation and discarding the supernatant, collecting the first wet bacterial body, adding sterile water to the first wet bacterial body to prepare a Trichoderma harzianum suspension, wherein the concentration of the Trichoderma harzianum is 10 7 -10 9 CFU / mL.

[0019] In the present invention, the activation is carried out in a shaker, and the rotation speed of the shaker is conventional in the art, typically but not limitingly, the rotation speed of the shaker is 120-140 rpm / min.

[0020] According to the present invention, the preparation step of the seed culture medium comprises: mixing potatoes, glucose and water in a mass ratio of (100-150): (20-25): (1000-1200), sterilizing in steam at a temperature of 120-130°C and a pressure of 200-210 kPa for 30-40 minutes to obtain the seed culture medium.

[0021] According to the present invention, the specific steps of activating the Pichia pastoris yeast are: mixing the Pichia pastoris yeast and water in a mass ratio of 1:8-15, performing a second activation at 25-30°C for 18-24 hours, centrifuging the product after the second activation and discarding the supernatant, collecting the second wet cells, and adding sterile water to the second wet cells to prepare a Pichia pastoris yeast suspension, wherein the concentration of the Pichia pastoris yeast is 10 7 -10 9 CFU / mL.

[0022] In the present invention, the centrifugal treatment of the first activated product and the second activated product is a conventional treatment condition in the art, typically but not limited to, centrifuging at a speed of 5000-6000 r / min for 10-15 min.

[0023] According to the present invention, the temperature of the first fermentation is 25-30°C, the time is 5-6 days, and the stirring speed during the first fermentation is 150-180r / min.

[0024] According to the present invention, the temperature of the second fermentation is 25-30°C, the time is 46-49h, and the stirring speed during the second fermentation is 150-180r / min.

[0025] According to the present invention, in S2, filtration is performed using a ceramic membrane of 80-100 nm.

[0026] According to the present invention, the filtered product is concentrated to 1 / 6-1 / 4 of the mass of the filtered product.

[0027] According to the present invention, the mass ratio of loquat concentrate, amino acid and reducing sugar is (90-100):(1-3):(2-8).

[0028] According to the present invention, the amino acid includes at least one of glycine, glutamic acid, proline, lysine and valine.

[0029] According to the present invention, the reducing sugar includes at least one of glucose, fructose, maltose, arabinose and xylose.

[0030] According to the present invention, the temperature of the Maillard reaction is 90-150° C., the time is 80-140 min, and the pH is 6.5-7.5.

[0031] In the present invention, pH refers to the pH of the entire system.

[0032] It can be understood by those skilled in the art that the method of preparing liquid loquat flavor into solid loquat flavor is conventional in the field, for example, liquid loquat flavor can be mixed with acceptable excipients to prepare solid particles, and the excipients include but are not limited to at least one of disintegrants, binders, fillers and lubricants. According to a preferred embodiment of the present invention, the specific steps of preparing liquid loquat flavor into solid loquat flavor include: mixing liquid loquat flavor, microcrystalline cellulose, β-cyclodextrin and glycerol to prepare wet material, extruding and molding, rolling into spherical wet particles, and drying to obtain solid loquat flavor.

[0033] According to the present invention, the mass ratio of the liquid loquat flavor, microcrystalline cellulose, β-cyclodextrin and glycerol is (13-22): (10-20): (8-15): (0.8-2.5), and can be optionally (15-20): (12-15): (10-12): (1-2).

[0034] According to the present invention, the rotation speed during the extrusion molding is 100-120 r / min, and the rotation speed during the spheronization is 1000-1200 r / min.

[0035] According to the present invention, the drying temperature is 40-50° C. and the drying time is 1-5 hours.

[0036] The second aspect of the present invention protects a solid loquat flavor obtained by the aforementioned preparation method.

[0037] A third aspect of the present invention protects a cigarette filter rod, wherein the cigarette filter rod comprises the aforementioned solid loquat flavoring.

[0038] According to the present invention, the cigarette filter rod is applied to the field of cigarettes or heat-not-burn cigarettes.

[0039] According to the present invention, solid loquat flavor is dispersedly added to a carrier (such as acetate fiber tow) at an addition amount of 0.5-2.5 mg / mm to form a cigarette filter rod, wherein mm refers to the length of the cigarette filter rod.

[0040] In the present invention, the method of adding solid loquat flavor to the cigarette filter rod is a conventional method in the art. Typically, but not limitedly, the solid loquat flavor is added to the cigarette filter rod and placed in a constant temperature box with a relative humidity of (60±3)% and a temperature of (22±1)°C for 48 hours.

[0041] The technical solution of the present invention has the following advantages:

[0042] 1. The present invention provides a preparation method of solid loquat flavor, wherein the preparation method comprises the following steps: S1, extracting loquat with water to obtain loquat extract; adding Trichoderma harzianum suspension to the loquat extract of S1, performing a first fermentation, adding Pichia pastoris suspension, performing a second fermentation, filtering and concentrating to obtain a loquat concentrate; S3, mixing the loquat concentrate, amino acids and reducing sugars, performing Maillard reaction, and obtaining liquid loquat flavor; S4, preparing the liquid loquat flavor into solid loquat flavor; in the present invention, Trichoderma harzianum and Pichia pastoris added in a specific order of addition act synergistically, can smoothly prepare loquat flavor without adding an organic solvent, and has low cost, simple operation and great application prospects; in the present invention, the Maillard reaction can add a caramel sweet aroma on the basis of the characteristic aroma of loquat, enriching the style of the flavor.

[0043] 2. Existing fermentation technology requires a large amount of microorganisms, and the transportation and storage conditions of microorganisms are relatively strict. As the amount used increases, the cost is greatly increased, which is not conducive to large-scale production. The specific amount of Trichoderma harzianum and Pichia pastoris in the present invention can achieve the fermentation of loquat extract with a smaller amount, further reducing the production cost.

[0044] 3. The specific dosage of the liquid loquat flavor, microcrystalline cellulose, β-cyclodextrin and glycerol of the present invention enables the wet material to be smoothly extruded and formed, and the synergistic effect of the liquid loquat flavor, microcrystalline cellulose, β-cyclodextrin and glycerol also enhances stability, sustained release and moisture retention; after spheronization, a solid flavor liquid loquat flavor that is convenient for transportation and storage is obtained. DETAILED DESCRIPTION

[0045] The following examples are provided for a better understanding of the present invention, but are not intended to limit the best mode of implementation, nor to limit the content and protection scope of the present invention. Any product identical or similar to the present invention obtained by anyone under the inspiration of the present invention or by combining the features of the present invention with other prior arts shall fall within the protection scope of the present invention.

[0046] If no specific experimental steps or conditions are specified in the examples, the conventional experimental steps or conditions described in the literature in the field can be used. If no manufacturer is specified for the reagents or instruments used, they are all conventional reagent products that can be obtained commercially.

[0047] Trichoderma harzianum (CICC 13010, China Industrial Microbiological Culture Collection Center);

[0048] Pichia pastoris (CICC 1958, China Industrial Microbiological Culture Collection Center);

[0049] Saccharomyces cerevisiae (CICC 1388, China Industrial Microbiological Culture Collection Center);

[0050] The extruder is an extruder E-25; the rounding machine is a rounding machine Mini S;

[0051] The loquats in the embodiments and comparative examples are loquats that have been cleaned and removed of impurities (cores, broken and damaged);

[0052] Sterile water is obtained after water is sterilized by high-temperature steam, wherein the steam pressure is 200 kPa, the temperature is 120°C, and the sterilization time is 30 minutes;

[0053] The preparation steps of the seed culture medium include: mixing potatoes, glucose and water in a mass ratio of 100:20:1000, sterilizing in steam at a temperature of 121° C. and a pressure of 205.8 kPa for 30 minutes, and preparing the seed culture medium.

[0054] Example 1

[0055] This embodiment provides a method for preparing solid loquat flavor, comprising the following steps:

[0056] S1, crushing loquats, adding deionized water, the mass ratio of deionized water to loquats is 5:1, stirring and extracting at 60°C and 500 rpm for 1 hour, filtering through a 50-mesh nylon filter cloth to obtain a loquat extract;

[0057] S2, inoculating the Trichoderma harzianum preserved under laboratory conditions onto a seed culture medium, with the mass of the seed culture medium as a benchmark, the mass of the Trichoderma harzianum being 2wt%, and culturing on a shaker (rotation speed of 140rpm / min) at 28°C for a first activation for 24h, centrifuging the first activated product at a rotation speed of 5000r / min for 10min, discarding the supernatant, collecting a first wet cell, adding sterile water to the first wet cell to prepare a Trichoderma harzianum suspension, wherein the concentration of the Trichoderma harzianum is 10 8 CFU / mL, the Pichia pastoris preserved under laboratory conditions was mixed with water at a mass ratio of 1:10, and the second activation was carried out at 28°C for 24 hours. The second activated product was centrifuged at 5000r / min for 10 minutes, and the supernatant was discarded. The second wet cells were collected, and sterile water was added to the second wet cells to prepare a Pichia pastoris suspension, wherein the concentration of Pichia pastoris was 10 8CFU / mL; adding the Trichoderma harzianum suspension to the loquat extract of S1, the mass of the Trichoderma harzianum suspension is 0.7wt% based on the mass of the loquat extract, and shaking and fermenting for 5 days at 28°C and 170r / min; adding the Pichia pastoris suspension to the loquat extract fermented by Trichoderma harzianum, the mass of the Pichia pastoris suspension is 2wt% based on the mass of the loquat extract, and shaking and fermenting for 48h at 30°C and 170r / min. After the fermentation is completed, centrifugation is performed to obtain the supernatant, and the supernatant is filtered using a 100nm ceramic membrane and concentrated to 1 / 5 of the mass of the filtered product to obtain a loquat concentrate;

[0058] S3, mixing loquat concentrate, glycine and glucose in a mass ratio of 90:1:2 to obtain a Maillard reaction substrate, adjusting the pH value to 7, and performing a Maillard reaction in an oil bath at a reaction temperature of 90° C. for a reaction time of 80 min to obtain a liquid loquat flavor;

[0059] S4, mixing liquid loquat flavor, microcrystalline cellulose, β-cyclodextrin and glycerol in a mass ratio of 17:14:11:1.5, stirring well to form a wet material, adding the wet material to an extruder, extruding at a speed of 100 r / min, and then placing the extrudate in a spheronizer and spheronizing at a speed of 1000 r / min into spherical wet particles; drying the prepared spherical wet particles in an oven at 40°C for 2 h to obtain solid loquat flavor.

[0060] Example 2

[0061] This embodiment provides a method for preparing solid loquat flavor, comprising the following steps:

[0062] S1, crushing loquats, adding deionized water, the mass ratio of deionized water to loquats is 5:1, stirring and extracting at 60°C and 500 rpm for 1 hour, filtering through a 50-mesh nylon filter cloth to obtain a loquat extract;

[0063] S2, inoculating the Trichoderma harzianum preserved under laboratory conditions onto a seed culture medium, with the mass of the seed culture medium as a benchmark, the mass of the Trichoderma harzianum being 1.5wt%, and culturing on a shaker (rotation speed of 140rpm / min) at 25°C for a first activation for 24h, centrifuging the first activated product at a rotation speed of 5500r / min for 10min and discarding the supernatant, collecting the first wet cells, adding sterile water to the first wet cells to prepare a Trichoderma harzianum suspension, wherein the concentration of the Trichoderma harzianum is 10 7CFU / mL, the Pichia pastoris preserved under laboratory conditions was mixed with water at a mass ratio of 1:10, and the second activation was carried out at 28°C for 24 hours. The second activated product was centrifuged at 5000r / min for 10 minutes, and the supernatant was discarded. The second wet cells were collected, and sterile water was added to the second wet cells to prepare a Pichia pastoris suspension, wherein the concentration of Pichia pastoris was 10 8 CFU / mL; adding the Trichoderma harzianum suspension to the loquat extract of S1, the mass of the Trichoderma harzianum suspension is 0.3wt% based on the mass of the loquat extract, and shaking and fermenting for 5 days at 28°C and 170r / min; adding the Pichia pastoris suspension to the loquat extract fermented by Trichoderma harzianum, the mass of the Pichia pastoris suspension is 2wt% based on the mass of the loquat extract, and shaking and fermenting for 46 hours at 30°C and 170r / min. After the fermentation is completed, centrifugation is performed to obtain the supernatant, and the supernatant is filtered using a 100nm ceramic membrane and concentrated to 1 / 5 of the mass of the filtered product to obtain a loquat concentrate;

[0064] S3, mixing loquat concentrate, glutamic acid and glucose in a mass ratio of 95:2:5 to obtain a Maillard reaction substrate, adjusting the pH value to 6.5, and performing a Maillard reaction in an oil bath at a reaction temperature of 120° C. for a reaction time of 100 min to obtain a liquid loquat flavor;

[0065] S4, mixing liquid loquat flavor, microcrystalline cellulose, β-cyclodextrin and glycerol in a mass ratio of 15:12:10:1, stirring well to form a wet material, adding the wet material to an extruder, extruding at a speed of 100 r / min, and then placing the extrudate in a spheronizer and spheronizing at a speed of 1000 r / min into spherical wet particles; drying the prepared spherical wet particles in an oven at 40°C for 2 h to obtain solid loquat flavor.

[0066] Example 3

[0067] This embodiment provides a method for preparing solid loquat flavor, comprising the following steps:

[0068] S1, crushing loquats, adding deionized water, the mass ratio of deionized water to loquats is 5:1, stirring and extracting at 60°C and 500 rpm for 1 hour, filtering through a 50-mesh nylon filter cloth to obtain a loquat extract;

[0069] S2, inoculating the Trichoderma harzianum preserved under laboratory conditions onto a seed culture medium, with the mass of the seed culture medium as a benchmark, the mass of the Trichoderma harzianum being 2wt%, and culturing on a shaker (rotation speed of 140rpm / min) at 28°C for a first activation for 24h, centrifuging the first activated product at a rotation speed of 5000r / min for 10min, discarding the supernatant, collecting a first wet cell, adding sterile water to the first wet cell to prepare a Trichoderma harzianum suspension, wherein the concentration of the Trichoderma harzianum is 10 8 CFU / mL, the Pichia pastoris preserved under laboratory conditions was mixed with water at a mass ratio of 1:10, and the second activation was carried out at 28°C for 24 hours. The second activated product was centrifuged at 5000r / min for 10 minutes, and the supernatant was discarded. The second wet cells were collected, and sterile water was added to the second wet cells to prepare a Pichia pastoris suspension, wherein the concentration of Pichia pastoris was 10 8 CFU / mL; adding the Trichoderma harzianum suspension to the loquat extract of S1, the mass of the Trichoderma harzianum suspension is 0.5wt% based on the mass of the loquat extract, and shaking and fermenting for 5 days at 28°C and 170r / min; adding the Pichia pastoris suspension to the loquat extract fermented by Trichoderma harzianum, the mass of the Pichia pastoris suspension is 2wt% based on the mass of the loquat extract, and shaking and fermenting for 48h at 30°C and 170r / min. After the fermentation is completed, centrifugation is performed to obtain the supernatant, and the supernatant is filtered using a 100nm ceramic membrane and concentrated to 1 / 5 of the mass of the filtered product to obtain a loquat concentrate;

[0070] S3, mixing loquat concentrate, proline and fructose in a mass ratio of 100:2:8 to obtain a Maillard reaction substrate, adjusting the pH value to 7, and performing a Maillard reaction in an oil bath at a reaction temperature of 130° C. for a reaction time of 120 min to obtain a liquid loquat flavor;

[0071] S4, mixing liquid loquat flavor, microcrystalline cellulose, β-cyclodextrin and glycerol in a mass ratio of 17:14:11:1.5, stirring well to form a wet material, adding the wet material to an extruder, extruding at a speed of 100 r / min, and then placing the extrudate in a spheronizer and spheronizing at a speed of 1000 r / min into spherical wet particles; drying the prepared spherical wet particles in an oven at 40°C for 2 h to obtain solid loquat flavor.

[0072] Example 4

[0073] This embodiment provides a method for preparing solid loquat flavor, comprising the following steps:

[0074] S1, crushing loquats, adding deionized water, the mass ratio of deionized water to loquats is 5:1, stirring and extracting at 60°C and 500 rpm for 1 hour, filtering through a 50-mesh nylon filter cloth to obtain a loquat extract;

[0075] S2, inoculating the Trichoderma harzianum preserved under laboratory conditions onto a seed culture medium, with the mass of the seed culture medium as a benchmark, the mass of the Trichoderma harzianum being 2wt%, and culturing on a shaker (rotation speed of 140rpm / min) at 28°C for a first activation for 24h, centrifuging the first activated product at a rotation speed of 5000r / min for 10min, discarding the supernatant, collecting a first wet cell, adding sterile water to the first wet cell to prepare a Trichoderma harzianum suspension, wherein the concentration of the Trichoderma harzianum is 10 8 CFU / mL, the Pichia pastoris preserved under laboratory conditions was mixed with water at a mass ratio of 1:15, and the second activation was carried out at 28°C for 24 hours. The second activated product was centrifuged at 5500r / min for 10 minutes, and the supernatant was discarded. The second wet cells were collected, and sterile water was added to the second wet cells to prepare a Pichia pastoris suspension, wherein the concentration of Pichia pastoris was 10 7 CFU / mL; adding the Trichoderma harzianum suspension to the loquat extract of S1, the mass of the Trichoderma harzianum suspension is 0.7wt% based on the mass of the loquat extract, and shaking and fermenting for 5 days at 28°C and 170r / min; adding the Pichia pastoris suspension to the loquat extract fermented by Trichoderma harzianum, the mass of the Pichia pastoris suspension is 3wt% based on the mass of the loquat extract, and shaking and fermenting for 48h at 30°C and 170r / min. After the fermentation is completed, centrifugation is performed to obtain a supernatant, and the supernatant is filtered using a 100nm ceramic membrane and concentrated to 1 / 5 of the mass of the filtered product to obtain a loquat concentrate;

[0076] S3, mixing loquat concentrate, lysine and maltose in a mass ratio of 98:3:8 to obtain a Maillard reaction substrate, adjusting the pH value to 7.5, and performing a Maillard reaction in an oil bath at a reaction temperature of 150° C. for a reaction time of 100 min to obtain a liquid loquat flavor;

[0077] S4, mixing liquid loquat flavor, microcrystalline cellulose, β-cyclodextrin and glycerol in a mass ratio of 16:13:12:2, stirring well to form a wet material, adding the wet material to an extruder, extruding at a speed of 100 r / min, and then placing the extrudate in a spheronizer and spheronizing at a speed of 1000 r / min into spherical wet particles; drying the prepared spherical wet particles in an oven at 40°C for 2 h to obtain solid loquat flavor.

[0078] Example 5

[0079] This embodiment provides a method for preparing solid loquat flavor, comprising the following steps:

[0080] S1, crushing loquats, adding deionized water, the mass ratio of deionized water to loquats is 5:1, stirring and extracting at 60°C and 500 rpm for 1 hour, filtering through a 50-mesh nylon filter cloth to obtain a loquat extract;

[0081] S2, inoculating the Trichoderma harzianum preserved under laboratory conditions onto a seed culture medium, with the mass of the seed culture medium as a benchmark, the mass of the Trichoderma harzianum being 3wt%, and culturing on a shaker (rotation speed of 140rpm / min) at 28°C for a first activation for 24h, centrifuging the first activated product at a rotation speed of 5000r / min for 10min and discarding the supernatant, collecting a first wet cell, adding sterile water to the first wet cell to prepare a Trichoderma harzianum suspension, wherein the concentration of the Trichoderma harzianum is 10 9 CFU / mL, the Pichia pastoris preserved under laboratory conditions was mixed with water at a mass ratio of 1:8, and the second activation was carried out at 28°C for 24 hours. The second activated product was centrifuged at 5000r / min for 10 minutes, and the supernatant was discarded. The second wet cells were collected, and sterile water was added to the second wet cells to prepare a Pichia pastoris suspension, wherein the concentration of Pichia pastoris was 10 9 CFU / mL; adding the Trichoderma harzianum suspension to the loquat extract of S1, the mass of the Trichoderma harzianum suspension is 1wt% based on the mass of the loquat extract, and shaking and fermenting for 5 days at 28°C and 170r / min; adding the Pichia pastoris suspension to the loquat extract fermented by Trichoderma harzianum, the mass of the Pichia pastoris suspension is 3wt% based on the mass of the loquat extract, and shaking and fermenting for 48h at 30°C and 170r / min. After the fermentation is completed, centrifugation is performed to obtain the supernatant, and the supernatant is filtered using a 100nm ceramic membrane and concentrated to 1 / 5 of the mass of the filtered product to obtain a loquat concentrate;

[0082] S3, mixing loquat concentrate, valine and xylose in a mass ratio of 100:2:4 to obtain a Maillard reaction substrate, adjusting the pH value to 7, and performing a Maillard reaction in an oil bath at a reaction temperature of 100° C. for a reaction time of 140 min to obtain a liquid loquat flavor;

[0083] S4, mixing liquid loquat flavor, microcrystalline cellulose, β-cyclodextrin and glycerol in a mass ratio of 20:15:12:2, stirring well to form a wet material, adding the wet material to an extruder, extruding at a speed of 100 r / min, and then placing the extrudate in a spheronizer and spheronizing at a speed of 1000 r / min into spherical wet particles; drying the prepared spherical wet particles in an oven at 40°C for 2 h to obtain solid loquat flavor.

[0084] Example 6

[0085] The present embodiment provides a preparation method of solid loquat flavor. The specific steps are the same as those of Embodiment 1, except that in S4, "liquid loquat flavor, microcrystalline cellulose, β-cyclodextrin and glycerol are mixed in a mass ratio of 17:14:11:1.5" is changed to "liquid loquat flavor, microcrystalline cellulose, β-cyclodextrin and glycerol are mixed in a mass ratio of 22:20:12:2".

[0086] Comparative Example 1

[0087] The present comparative example provides a method for preparing solid loquat flavor. The specific steps are the same as those of Example 1, except that, in S2, the Pichia pastoris preserved under laboratory conditions is mixed with water in a mass ratio of 1:10, and activated at 28°C for 24 hours. The activated product is centrifuged at a speed of 5000 r / min for 10 minutes, and the supernatant is discarded. The wet cells are collected, and sterile water is added to the wet cells to prepare a Pichia pastoris suspension, wherein the concentration of the Pichia pastoris is 10 8 CFU / mL, the Pichia pastoris suspension was added to the loquat extract, the mass of the Pichia pastoris suspension was 2.7wt% based on the mass of the loquat extract, and the fermentation was carried out at 30°C and 170r / min for 7 days. After the fermentation was completed, the supernatant was obtained by centrifugation, and the supernatant was filtered through a 100nm ceramic membrane and concentrated to 1 / 5 of the mass of the filtered product to obtain a loquat concentrate.

[0088] Comparative Example 2

[0089] The present comparative example provides a preparation method of solid loquat flavor, and the specific steps are the same as those of Example 1, except that, in S2, the Trichoderma harzianum preserved under laboratory conditions is inoculated onto a seed culture medium, and the mass of the Trichoderma harzianum is 2wt% based on the mass of the seed culture medium, and the culture is activated on a shaker (with a rotation speed of 140 rpm / min) at 28°C for 24 hours, and the activated product is centrifuged at a speed of 5000 r / min for 10 minutes, and the supernatant is discarded, the wet cells are collected, and sterile water is added to the wet cells to prepare a Trichoderma harzianum suspension, wherein the concentration of the Trichoderma harzianum is 10 8CFU / mL, the Trichoderma harzianum suspension was added to the loquat extract of S1, the mass of the Trichoderma harzianum suspension was 2.7wt% based on the mass of the loquat extract, and the mixture was shaken and fermented at 28°C and 170r / min for 7 days. After the fermentation, the supernatant was obtained by centrifugation, which was filtered through a 100nm ceramic membrane and concentrated to 1 / 5 of the mass of the filtered product to obtain a loquat concentrate.

[0090] Comparative Example 3

[0091] The present comparative example provides a preparation method of solid loquat flavor. The specific steps are the same as those of Example 1, except that in S2, "Pichia pastoris" is changed to "Saccharomyces cerevisiae", Saccharomyces cerevisiae preserved under laboratory conditions is mixed with water in a mass ratio of 1:10, and activated at 28°C for 24 hours. The activated liquid is centrifuged at 5000 r / min for 10 minutes, and the supernatant is discarded. The wet cells are collected, and sterile water is added to the wet cells to prepare a Saccharomyces cerevisiae suspension, wherein the concentration of Saccharomyces cerevisiae is 10 8 CFU / mL; based on the mass of loquat extract, the mass of brewer's yeast suspension is 2.7wt%.

[0092] Comparative Example 4

[0093] The present comparative example provides a method for preparing solid loquat flavor, and the specific steps are the same as those of Example 1, except that, in S2, the Trichoderma harzianum preserved under laboratory conditions is inoculated onto a seed culture medium, and the mass of the Trichoderma harzianum is 2wt% based on the mass of the seed culture medium, and the first activation is carried out on a shaker (with a rotation speed of 140 rpm / min) at 28°C for 24 hours, the first activated product is centrifuged at a speed of 5000 r / min for 10 minutes, the supernatant is discarded, the first wet cells are collected, and sterile water is added to the first wet cells to prepare a Trichoderma harzianum suspension, wherein the concentration of the Trichoderma harzianum is 10 8 CFU / mL, the Pichia pastoris preserved under laboratory conditions was mixed with water at a mass ratio of 1:10, and the second activation was carried out at 28°C for 24 hours. The second activated product was centrifuged at 5000r / min for 10 minutes, and the supernatant was discarded. The second wet cells were collected, and sterile water was added to the second wet cells to prepare a Pichia pastoris suspension, wherein the concentration of Pichia pastoris was 10 8CFU / mL. The Pichia pastoris suspension was added to the loquat extract of S1, and the mass of the Pichia pastoris suspension was 2wt% based on the mass of the loquat extract. The mixture was shaken and fermented at 30°C and 170r / min for 48h. The Trichoderma harzianum suspension was added to the loquat extract fermented with the Pichia pastoris suspension, and the mass of the Trichoderma harzianum suspension was 0.7wt% based on the mass of the loquat extract. The mixture was shaken and fermented at 28°C and 170r / min for 5d. After the fermentation was completed, the supernatant was obtained by centrifugation, filtered using a 100nm ceramic membrane, and concentrated to 1 / 5 of the mass of the filtered product to obtain a loquat concentrate.

[0094] Test Case

[0095] The solid loquat flavor prepared in the embodiment and the comparative example was dispersedly added to the acetate fiber tow in an addition amount of 0.5 mg / mm, 1 mg / mm, 1.5 mg / mm, 2.0 mg / mm and 2.5 mg / mm to form a cigarette filter rod, which was placed in a constant temperature box with a relative humidity of (60±3)% and a temperature of (22±1)°C for 48 hours; the sensory evaluation was conducted by the smoking testers in accordance with GBYC / T 5604.4-2005 "Cigarettes Part 4: Sensory Technical Requirements" for traditional cigarettes. The evaluation team consisted of 10 persons with smoking evaluation qualifications, and the smoking evaluation results were the average values ​​of the tests of samples No. 1 to 5; the control sample was a blank control without adding solid loquat flavor, and the specific test results are shown in Table 1.

[0096] Table 1

[0097]

[0098]

[0099] The Trichoderma harzianum and Pichia pastoris added in a specific order of addition in the present invention work synergistically, and can smoothly prepare loquat flavor without adding an organic solvent, with low cost, simple operation and great application prospect.

[0100] Comparison between Example 1 and Comparative Example 3 shows that, when Saccharomyces cerevisiae is used, although the quality of Saccharomyces cerevisiae is improved, it can be seen from the final score in Table 1 that it is still not as high as the score of Example 1.

[0101] Obviously, the above embodiments are merely examples for the purpose of clear explanation, and are not intended to limit the implementation methods. For those skilled in the art, other different forms of changes or modifications can be made based on the above description. It is not necessary and impossible to list all the implementation methods here. The obvious changes or modifications derived from these are still within the scope of protection of the invention.

Claims

1. A method for preparing solid loquat flavor, characterized in that: The preparation method comprises the following steps: S1, extracting loquat with water to obtain loquat extract; S2, activating Trichoderma harzianum to obtain a Trichoderma harzianum suspension, activating Pichia pastoris to obtain a Pichia pastoris suspension, adding the Trichoderma harzianum suspension to the loquat extract of S1, performing a first fermentation, adding the Pichia pastoris suspension, performing a second fermentation, filtering, and concentrating to obtain a loquat concentrate; S3, mixing loquat concentrate, amino acids and reducing sugars, and performing Maillard reaction to obtain liquid loquat flavor; S4, preparing the liquid loquat flavor into solid loquat flavor.

2. The preparation method according to claim 1, characterized in that: In S1, the mass ratio of water to loquat is 5-7:1; And / or, the extraction temperature is 60-75°C and the extraction time is 1-2h.

3. The preparation method according to claim 1 or 2, characterized in that: Based on the mass of the loquat extract, the mass of the Trichoderma harzianum suspension is 0.1-1wt%, and the mass of the Pichia pastoris suspension is 1-3wt%; Optionally, based on the mass of the loquat extract, the mass of the Trichoderma harzianum suspension is 0.5-0.8wt%, and the mass of the Pichia pastoris suspension is 1.5-2.5wt%; And / or, the activation of Trichoderma harzianum specifically comprises the following steps: inoculating Trichoderma harzianum onto a seed culture medium, with the mass of the Trichoderma harzianum being 1-3wt% based on the mass of the seed culture medium, performing a first activation at 25-30°C for 18-24h, centrifuging the product after the first activation and discarding the supernatant, collecting a first wet cell, and adding sterile water to the first wet cell to prepare a Trichoderma harzianum suspension, wherein the concentration of Trichoderma harzianum is 10 7 -10 9 CFU / mL; Optionally, the step of preparing the seed culture medium comprises: mixing potatoes, glucose and water in a mass ratio of (100-150): (20-25): (1000-1200), sterilizing in steam at a temperature of 120-130° C. and a pressure of 200-210 kPa for 30-40 minutes to prepare the seed culture medium; And / or, the specific steps of activating the Pichia pastoris yeast are: mixing the Pichia pastoris yeast and water in a mass ratio of 1:8-15, performing a second activation at 25-30°C for 18-24 hours, centrifuging the product after the second activation and discarding the supernatant, collecting the second wet cells, adding sterile water to the second wet cells to prepare a Pichia pastoris yeast suspension, wherein the concentration of the Pichia pastoris yeast is 10 7 -10 9 CFU / mL.

4. The preparation method according to claim 1 or 2, characterized in that: The temperature of the first fermentation is 25-30°C and the time is 5-6 days; And / or, the temperature of the second fermentation is 25-30°C and the time is 46-49h; And / or, in S2, the filtering is performed using a ceramic membrane of 80-100 nm; And / or, in S2, the filtered product is concentrated to 1 / 6-1 / 4 of the mass of the filtered product.

5. The preparation method according to any one of claims 1 to 4, characterized in that: The mass ratio of loquat concentrate, amino acid and reducing sugar is (90-100):(1-3):(2-8); and / or, the amino acid comprises at least one of glycine, glutamic acid, proline, lysine and valine; And / or, the reducing sugar includes at least one of glucose, fructose, maltose, arabinose and xylose.

6. The preparation method according to any one of claims 1 to 5, characterized in that: The temperature of the Maillard reaction is 90-150° C., the time is 80-140 min, and the pH is 6.5-7.

5.

7. The preparation method according to any one of claims 1 to 6, characterized in that: The specific steps of preparing the liquid loquat flavor into solid loquat flavor include: mixing the liquid loquat flavor, microcrystalline cellulose, β-cyclodextrin and glycerol to prepare a wet material, extruding and molding, rolling into spherical wet particles, and drying to obtain the solid loquat flavor; Optionally, the mass ratio of the liquid loquat flavor, microcrystalline cellulose, β-cyclodextrin and glycerol is (13-22):(10-20):(8-15):(0.8-2.5), or optionally (15-20):(12-15):(10-12):(1-2).

8. The preparation method according to claim 7, characterized in that: The rotation speed during the extrusion molding is 100-120r / min, and the rotation speed during the spheronization is 1000-1200r / min; And / or, the drying temperature is 40-50° C. and the drying time is 1-5 h.

9. A solid loquat flavor prepared by the preparation method according to any one of claims 1 to 8.

10. A cigarette filter rod, characterized in that: The cigarette filter rod comprises the solid loquat flavoring as claimed in claim 9; Optionally, the cigarette filter rod is used in the field of cigarettes or heat-not-burn cigarettes.

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