Method for evaluating quality of sorghum for wine and application of method

Abstract

The invention provides a method for evaluating the quality of sorghum for wine. The method comprises the following steps: (1) grain moistening simulation: performing grain moistening simulation treatment on sorghum to be evaluated; (2) sampling: sampling the sorghum in different simulated grain moistening stages, and respectively measuring the dulin content in the sorghum samples corresponding to different stages; and (3) evaluation: calculating the dulin content of the to-be-evaluated sorghum based on the dulin content in the sorghum samples corresponding to the different stages, which is measured in the step (2), and evaluating the quality of the to-be-evaluated sorghum based on the calculated dulin content of the to-be-evaluated sorghum. According to the evaluation method disclosed by the invention, the influence of dulin content change in the sorghum in the grain moistening process is considered, and the scientificity of the quality evaluation method of the sorghum for wine is improved.

Description

Technical Field

[0001] The present application relates to the technical field of quality evaluation of raw materials for brewing wine, and in particular to a method for evaluating the quality of sorghum for brewing wine. Background Art

[0002] Sorghum (Sorghum bicolor (L.) Moench) is the fifth largest cereal crop in the world. It can be used as food, feed or brewing raw material. It has the characteristics of good resistance to barrenness and stress. Sorghum is the main raw material for brewing Maotai-flavor liquor and the basis for brewing high-quality Maotai-flavor liquor. In recent years, with the development of the Maotai-flavor liquor industry, the demand for sorghum raw materials has been increasing, and the demand of brewing enterprises above a certain scale is hundreds of thousands of tons per year. There is also a certain amount of dullin in sorghum. According to relevant literature reports, dullin is derived from L-tyrosine through the action of three biosynthetic enzymes (two cytochrome P450, CYP79A1 and CYP71E1 enzymes and UDP-glycosyltransferase, UGT85B1). The genes encoding these three enzymes form a biosynthetic gene cluster, which is adjacent to each other in the sorghum genome; there is also a transporter (multidrug and toxic compound extrusion, MATE), whose function is to isolate potential self-toxic compounds in the vacuole of plant cells. During the liquor brewing process, the durin in sorghum grains can be hydrolyzed by β-glucosidase or pyrolysis to eventually generate a precursor of ethyl carbamate (EC). Due to its low boiling point, this substance can easily enter the base wine during the distillation process. This precursor finally generates EC in the wine, and a lower EC content is usually regarded as one of the higher product quality standards. Therefore, studying the durin content in sorghum to screen out sorghum with low durin content for brewing is of great significance for the control of EC in liquor.

[0003] Winemaking sorghum exhibits different durin production abilities due to the influence of factors such as variety and planting environment, and the prerequisite for screening winemaking raw material sorghum with low durin content is to have a corresponding evaluation method. Through literature search, it was found that the existing technology mainly uses the determination of the durin content in the initial state of sorghum as the basis for screening wine sorghum with low durin content, such as the document "Differential Analysis of the Content of Amygdalus Cyanogenides in Sorghum from Different Origins and Varieties-Zhou Hanling-China Brewing" disclosed: The durin content in 39 initial state sorghum samples in Northeast China and southern Sichuan was determined by high performance liquid chromatography-tandem mass spectrometry, and the differences in the content of amygdalus cyanogenides in sorghum from different origins and different varieties were compared and analyzed. However, there are certain limitations in the use of the durin content in the sorghum grains in the initial state of sorghum to evaluate and screen wine sorghum with low durin content, and it is impossible to more objectively reflect the durin content of wine sorghum. Summary of the invention

[0004] Based on this, the purpose of this application is to provide a more scientific method for evaluating the quality of sorghum for wine.

[0005] To achieve the above objectives, the present application provides a method for evaluating the quality of sorghum for wine, and the technical scheme adopted is as follows:

[0006] In some embodiments, the detection method of the present application comprises the following steps:

[0007] (1) Simulated grain moistening: simulated grain moistening of the sorghum to be evaluated;

[0008] (2) Sampling: Sampling sorghum at different simulated grain moistening stages, and measuring the durin content in the sorghum samples corresponding to different stages;

[0009] (3) Evaluation: Calculating the dulin content of the sorghum to be evaluated based on the dulin content in the sorghum samples corresponding to different stages measured in step (2), and evaluating the quality of the sorghum to be evaluated based on the calculated dulin content of the sorghum to be evaluated.

[0010] The present application has found through research that different storage times will cause different durin production abilities of sorghum for brewing. Compared with sorghum that has not been moistened, the durin content of sorghum that has been moistened varies greatly, and the durin content of sorghum in different moistening states is different. Regardless of whether it is a sauce-flavored, strong-flavored or light-flavored liquor, when sorghum is used as a raw material, the moistened sorghum is generally mixed with distiller's yeast during the brewing process. Therefore, moistening sorghum is an important and necessary step in liquor brewing. Therefore, the method of measuring the durin content of sorghum in a dry state and then evaluating the quality of sorghum based on the prior art has certain limitations, and cannot objectively reflect the durin content of sorghum for brewing, nor can it evaluate the quality of sorghum based on the measured durin content.

[0011] Based on this, this application proposes a new and more scientific method for evaluating the quality of sorghum for wine. By exploring the simulated grain moistening treatment conditions of sorghum and based on the durin content in sorghum in different states after simulated moistening treatment, the durin content in the sorghum to be evaluated is further calculated, thereby realizing the evaluation of sorghum quality and improving the scientific nature of the quality evaluation method of sorghum for wine.

[0012] In some embodiments, the simulated grain moistening method is the same as the grain moistening method in actual winemaking in terms of the following indicators, which include: one or more of water temperature, water addition amount, water addition times, water addition interval, culture time, culture temperature, and sorghum breakage;

[0013] The grain moistening methods in actual winemaking include the grain moistening methods used in the brewing process of liquors such as Maotai-flavor liquor, Qing-flavor liquor, Luzhou-flavor liquor, and Fengxiang-flavor liquor with sorghum as one of the raw materials;

[0014] In some embodiments, the simulated grain moistening process includes multiple waterings, and the sampling nodes include before each watering of the multiple waterings, within a period of time after the last watering, and when the grain moistening is completed.

[0015] In the brewing of sauce-flavor, strong-flavor, light-flavor, and phoenix-flavor liquors, the grain needs to be moistened with water many times to allow it to fully absorb water, and the grain may be left for a period of time after moistening to better carry out the subsequent fermentation process. Therefore, the above sampling method can take into account the effects of different water addition stages and different standing times after water addition on the formation of durin in sorghum.

[0016] In some embodiments, the simulated grain moistening treatment of the sorghum to be evaluated includes sequentially performing simulated watering treatment on the sorghum to be evaluated.

[0017] In some embodiments, the simulated watering treatment includes: a first simulated watering treatment, a second simulated watering treatment, and a third simulated watering treatment; the crushing degree of the sorghum to be evaluated in the simulated watering treatment is 25%-35%;

[0018] In some embodiments, based on the grain moistening conditions of the Maotai-flavor liquor sand making round, simulated grain moistening is performed, and the proportion of broken sorghum in the total amount can be 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, or a range consisting of any two of the above values;

[0019] In some embodiments, simulated grain moistening is performed based on the grain moistening conditions of the Maotai-flavor liquor sand making round, and the broken sorghum accounts for 30% of the total amount;

[0020] In some embodiments, simulating water addition comprises the following steps: adding water to the sorghum sample to be evaluated, and placing it in an incubator for culturing;

[0021] In some embodiments, the simulated grain moistening treatment of the sorghum to be evaluated includes sequentially performing simulated watering treatment after crushing the sorghum to be evaluated;

[0022] In some embodiments, the simulated grain moistening treatment of the sorghum to be evaluated includes successively performing simulated watering treatment after crushing the sorghum to be evaluated; the crushing degree of the sorghum to be evaluated is 20% to 100%;

[0023] In some embodiments, the simulated grain moistening treatment of the sorghum to be evaluated includes successively performing simulated watering treatment after crushing the sorghum to be evaluated; the crushing degree of the sorghum to be evaluated is 20% (sanding round) or 30% (sand making round); at this time, the crushing degree of this embodiment is suitable for brewing Maotai-flavor liquor;

[0024] In some embodiments, the simulated grain moistening treatment of the sorghum to be evaluated includes crushing the sorghum to be evaluated and then performing simulated watering treatment in sequence; the crushing degree of the sorghum to be evaluated is 100%; at this time, the crushing degree of this embodiment is suitable for the brewing of strong-flavor liquor (light-flavor liquor, glutinous rice wine, Daqu wine, etc., which require the sorghum to be completely crushed). In some embodiments, the watering ratio used in the first simulated watering is 30-40% of the weight of the sorghum to be evaluated; the temperature of the water used in the first simulated watering is 90-100°C; the culture temperature of the first simulated watering is 38-45°C;

[0025] In some embodiments, the watering ratio used in the second simulated watering is 8-15% of the weight of the sorghum to be evaluated, the temperature of the water used in the second simulated watering is 90-100° C.; the cultivation temperature of the second simulated watering is 38-45° C.;

[0026] In some embodiments, the watering ratio used in the third simulated watering is 8-15% of the weight of the sorghum to be evaluated, and the temperature of the water used in the second simulated watering is 90-100°C; the culture temperature of the third simulated watering is 38-45°C; in some embodiments, the first watering ratio is 34% of the weight of the sorghum to be evaluated, the temperature of the water used in the first simulated watering is 95°C, and the culture temperature of the first simulated watering is 42°C; the second watering ratio is 12% of the weight of the sorghum to be evaluated, the temperature of the water used in the second simulated watering is 95°C, and the culture temperature of the second simulated watering is 42°C; the third watering ratio is 10% of the weight of the sorghum to be evaluated, the temperature of the water used in the third simulated watering is 95°C, and the culture temperature of the second simulated watering is 42°C;

[0027] In some embodiments, the sampling of sorghum at different simulated grain moistening stages includes: sampling the sorghum to be evaluated in the simulated grain moistening process before the first simulated watering, 1.0-1.5 hours after the first simulated watering, 6.0-6.5 hours after the first simulated watering, 20-28 hours after the first simulated watering, and 50-54 hours after the first simulated watering;

[0028] In some embodiments, the sampling of sorghum at different simulated grain moistening stages includes: sampling the sorghum to be evaluated in the simulated grain moistening process before the first simulated watering, 1.3 hours after the first simulated watering, 6.3 hours after the first simulated watering, 24 hours after the first simulated watering, and 54 hours after the first simulated watering;

[0029] In some embodiments, the durin content of the sorghum to be evaluated is calculated based on the durin content in the sorghum samples corresponding to different stages determined in step (2), including: using a dynamically changing weighted summation method, among the different sorghum samples obtained, a smaller weighting coefficient is assigned to the durin content of the sorghum samples close to the start stage of simulated grain moistening, and a larger weighting coefficient is assigned to the durin content of the sorghum samples close to the end stage of simulated grain moistening.

[0030] In some embodiments, the step of calculating the durin content of the sorghum to be evaluated based on the durin content in the sorghum samples corresponding to different stages determined in step (2) comprises: calculating using the following calculation formula: E = X1C1 + ... + X n Cn, where E is the dullin content of the sorghum to be evaluated, 1 to n represent the different simulated grain moistening stages of the samples taken from the beginning to the end of the simulated grain moistening, C represents the dullin content measured in the sorghum samples at the corresponding simulated grain moistening stages, and X represents the weighting coefficient, where X1+…+X n＝ 100%; in some embodiments, X n Greater than X1 to X n-1 In some embodiments, X n ≥(X1+…+X n-1 ); in some embodiments, n can be an integer greater than or equal to 1; in some embodiments, E=X1C1, that is, at this time n=1, X1=100%, when n=1, the simulated grain moistening stage represented by n is close to the end stage of simulated grain moistening, or the simulated grain moistening stage represented by n is in a simulated grain moistening state before steaming. The reason is that the closer to the steaming stage, the easier it is for the dulin in this state to enter the white wine. Therefore, the weight of the dulin content of the sorghum to be evaluated near the steaming stage should be larger.

[0031] In some embodiments, the calculating of the durin content of the sorghum to be evaluated based on the durin content in the sorghum samples corresponding to different stages determined in step (2) comprises: calculating using the following calculation formula: E=X1C1+X2C2+X3C3+X4C4+X5C5; wherein E is the durin content of the sorghum to be evaluated, the value range of X1 is 8%-15%, the value range of X2 is 2%-8%, the value range of X3 is 2%-8%, the value range of X4 is 25%-35%, the value range of X5 is 40%-55%, X1+X2+X3+X4+X5C5 is 10%. 5＝ 100%, C1 is the content of dolin measured by sampling before the first simulated watering, C2 is the content of dolin measured by sampling 1.0-1.5h after the first simulated watering, C3 is the content of dolin measured by sampling 6.0-6.5h after the first simulated watering, C4 is the content of dolin measured by sampling 20-28h after the first simulated watering, and C5 is the content of dolin measured by sampling 50-54h after the first simulated watering;

[0032] In some embodiments, the calculation of the dulin content of the sorghum to be evaluated based on the dulin content in the sorghum samples corresponding to different stages determined in step (2) includes: calculating using the following calculation formula: E=10%C1+5%C2+5%C3+32%C4+48%C5; wherein E is the dulin content of the sorghum to be evaluated, C1 is the dulin content measured by sampling before the first simulated watering, C2 is the dulin content measured by sampling 1.0-1.5h after the first simulated watering, C3 is the dulin content measured by sampling 6.0-6.5h after the first simulated watering, C4 is the dulin content measured by sampling 20-28h after the first simulated watering, and C5 is the dulin content measured by sampling 50-54h after the first simulated watering;

[0033] In some embodiments, the simulated grain moistening treatment of the sorghum to be evaluated further includes placing the sorghum to be evaluated in a container and sequentially performing simulated watering treatment on the sorghum to be evaluated; the ratio of the weight of the sorghum to be evaluated to the capacity of the container is: 1:4 g / mL to 1:2 g / mL;

[0034] In some embodiments, the weight of the sorghum sample to be evaluated is 350g-500g;

[0035] In some embodiments, the weight of the sorghum sample to be evaluated can be 350 g, 360 g, 370 g, 380 g, 390 g, 400 g, 410 g, 420 g, 430 g, 440 g, 450 g, 460 g, 470 g, 500 g, or a range consisting of any two of the above values;

[0036] When setting the sampling weight of the sorghum to be evaluated, when the sample amount is too small, the water content of the sample is easily lost, and multiple samplings will reduce the total amount of the sample to a certain extent, thereby increasing the difference between samples; when the sample amount is too large, it will take up too much constant temperature incubator space and waste raw materials; combined with the actual conditions such as experimental operability and convenience, therefore, in some embodiments, the sampling weight of the sorghum to be evaluated is 400g, and the container capacity is 1000mL;

[0037] In some embodiments, the determining the durin content in the sorghum samples corresponding to different stages respectively comprises: determining the durin content in the sorghum samples based on chromatography-mass spectrometry;

[0038] In some embodiments, the pretreatment comprises: taking 2 g of sorghum in a 50 mL centrifuge tube, adding 20 mL of 95% ethanol, homogenizing with an automatic homogenizer for 50 seconds, vortexing at 2000 rpm for 5 minutes, ultrasonicating in an ice bath for 20 minutes, then vortexing at 2000 rpm for 5 minutes, centrifuging at 6000 rpm for 5 minutes, and filtering the supernatant to obtain a solution to be analyzed;

[0039] In some embodiments, the chromatography-mass spectrometry is ultra-high performance liquid chromatography-high resolution mass spectrometry, and the parameters of the ultra-high performance liquid chromatography used include: the chromatographic column is: Hypersil Gold aQ liquid chromatography column;

[0040] In some embodiments, the chromatographic parameters further include: injection volume 3.5 to 7.5 μL;

[0041] In some embodiments, the parameters of the chromatography further include: mobile phase: phase A is ultrapure water containing 5 mmol / L ammonium acetate and 1‰ formic acid, and phase B is methanol;

[0042] In some embodiments, the parameters of the chromatography also include: flow rate: 300-400 μL / min;

[0043] In some embodiments, the parameters of the ultra-high performance liquid chromatography are as follows: the ultra-high performance liquid chromatography conditions are as follows: a Hypersil Gold aQ liquid chromatography column (2.1 mm × 150 mm, 1.9 μm) is selected, the injection volume is 5 μL; the mobile phase: phase A is ultrapure water containing 5 mmol / L ammonium acetate and 1‰ formic acid, and phase B is methanol; flow rate: 350 μL / min;

[0044] In some embodiments, the mass spectrometer is a quadrupole electrostatic field orbital trap high-resolution mass spectrometer, and the quadrupole electrostatic field orbital trap high-resolution mass spectrometer conditions include: positive ion mode, sheath gas flow rate of 30-50arb, auxiliary gas flow rate of 10-20arb, purge gas of 0arb, ion transfer tube temperature of 260-300°C, spray voltage of 4-5kV, back purge temperature of 360-400°C, scan mode of PRM, NCE of 10, wherein the parent ion of Du Lin is m / z: 329.1343, and the daughter ion is m / z: 132.0441;

[0045] In some embodiments, the quadrupole electrostatic field orbital trap high-resolution mass spectrometry conditions include: the specific parameters of the positive ion mode are as follows: the sheath gas flow rate is 40arb, the auxiliary gas flow rate is 15arb, the purge gas is 0arb, the ion transfer tube temperature is 280°C, the spray voltage is 4.5kV, the back purge temperature is 380°C, the scan mode is PRM, and the NCE is 10, wherein the parent ion of Du Lin is m / z: 329.1343, and the daughter ion is m / z: 132.0441;

[0046] In some embodiments, the sample is pre-treated before the determination, and the pre-treatment includes: adding ethanol to the sampled sorghum, homogenizing, vortexing, centrifuging, taking the supernatant, filtering the membrane, and obtaining the liquid to be analyzed;

[0047] In some embodiments, the pretreatment includes: weighing 1-5 g of sorghum sample, adding 10-50 mL of 95% ethanol, homogenizing with an automated homogenizer for 45-120 s, then vortexing for 3-10 minutes, then ultrasonicating in an ice bath for 15-35 minutes, then vortexing for 3-10 minutes, then centrifuging for 1-10 minutes, and taking the supernatant and filtering the membrane to obtain the liquid to be analyzed.

[0048] On the other hand, the present application provides an application of the aforementioned method in the quality evaluation of sorghum for wine.

[0049] In some embodiments, according to the method described above, the present application screens sorghums with different vitality, and the sorghums for wine with different vitality include sorghums with different storage conditions such as storage time, temperature, and humidity. By screening sorghums with low dullin content or actively reducing the dullin production capacity in sorghum by frying, etc., less dullin enters the liquor brewing system, thereby improving the quality of liquor. Therefore, the method for evaluating the quality of sorghum for wine in the present application can make a more objective and scientific evaluation of sorghums of different varieties, different origins, and different processing conditions.

[0050] Beneficial effects: The present application measures the durin content of sorghum at different simulated grain-moistening stages and calculates the durin content of the sorghum to be evaluated based on the measurement results. The research of the present application found that the grain-moistening process may affect the activities of various enzymes in sorghum, thereby affecting the durin content in sorghum. Grain-moistening is an important step in the brewing of various types of liquor using sorghum as raw material. Therefore, the evaluation method of the present application takes into account the impact of changes in the durin content in sorghum during the grain-moistening process, thereby improving the scientific nature of the quality evaluation method of sorghum for liquor. BRIEF DESCRIPTION OF THE DRAWINGS

[0051] Figure 1 The dullin content E of the sorghum to be evaluated is measured at different simulated grain moistening time stages for sorghum stored for one year, sorghum stored for three months, and sorghum stored for one year and fried;

[0052] Figure 2 The sorghum is roasted and stored for one year;

[0053] Figure 3 This is the state diagram of sorghum stored for one year at 54 hours after the first watering;

[0054] Figure 4 This is the state diagram of sorghum stored for 3 months at 54h after the first watering;

[0055] Figure 5 This is the state diagram of the roasted sorghum stored for one year at 54 hours after the first watering; DETAILED DESCRIPTION

[0056] The technical solution of the present invention is further described below by specific embodiments, which do not limit the protection scope of the present invention. Some non-essential modifications and adjustments made by others based on the concept of the present invention still fall within the protection scope of the present invention.

[0057] The equipment and reagent information used in the embodiments of the present invention are as follows:

[0058] Equipment: 1290 ultra-high performance liquid chromatography: Agilent Technologies, USA; Q-Exactive quadrupole electrostatic field orbital trap high-resolution mass spectrometer: Thermo Corporation, USA; constant temperature incubator: Shanghai Foma Experimental Equipment Co., Ltd.; centrifuge: Beckman Coulter Co., Ltd.; fully automatic homogenizer (AH-40): Ruike Group (Xiamen) Co., Ltd.; Milli-Q ultrapure water instrument: Millipore, USA; 0.22μm microporous filter membrane: Agilent, USA; XP205 analytical balance: Shanghai Mettler-Toledo Instrument Co., Ltd.

[0059] Reagents: acetonitrile, methanol (chromatographic grade): Merck, Germany; 95% ethanol (analytical grade): Sinopharm Chemical Reagent Co., Ltd., Dulin: Beijing Bailingwei Technology Co., Ltd., phosphoric acid (85%-90%, Aladdin); laboratory water was ultrapure water (18.2 MΩ·cm).

[0060] Example 1

[0061] This embodiment provides a method for evaluating the quality of sorghum for wine, comprising the following steps:

[0062] 1. Grinding grain

[0063] 1500g of sorghum samples were taken respectively (the sorghum samples included sorghum with a storage time of 1 year, sorghum with a storage time of 3 months, and sorghum with a storage time of 1 year after being fried), and the sorghum was crushed according to the sand-running grain process requirements for making Maotai-flavor liquor, and the crushing degree was controlled to be 30%; wherein, the specific operation of crushing was as follows: 450g of sorghum was crushed with a crusher, 1050g of whole sorghum was added and mixed evenly, and a crushed sorghum sample with a crushing degree of 30% was obtained; the frying method of the sorghum with a storage time of 1 year after being fried was as follows: 500g of sorghum was fried with an induction cooker until it was crisp, fried, not mushy, and not bubbling, and fried over low heat, specifically 600W for 3min, and mixed evenly after three fryings. Figure 2 As shown;

[0064] 2. Simulate grain moistening

[0065] 400 g of the sorghum samples with a crushing degree of 30% prepared in step 1 were taken respectively and placed in a 1000 mL beaker. The sorghum samples were subjected to simulated grain moistening treatment according to the watering ratio and watering time interval of Maotai-flavor liquor. Three parallel experimental groups were set up in the experiment. The specific simulated grain moistening treatment steps are as follows:

[0066] The first simulated watering: the watering ratio is 34%, and the watering amount is 400g*34%=136g, that is, 136g of 95℃ water is added to each sorghum sample, and the mixture is stirred in time. After stirring, a layer of wet gauze is covered on the sample to prevent water loss. The sample is placed in a 42℃ constant temperature incubator and the timing is started;

[0067] Second simulated watering: 1.3 hours after the first watering, the second watering was carried out, the watering ratio was 12%, and the watering amount was 400g*12%=48g, that is, 1.3 hours after the first watering, 48g of 95°C water was added to each sorghum sample, and the samples were stirred in time. After the stirring, the samples were placed in a constant temperature incubator at 42°C;

[0068] The third simulated watering: the third watering was carried out 5 hours after the second watering, the watering ratio was 10%, and the watering amount was 400g*10%=40g, that is, 5 hours after the second watering, 40g of 95°C water was added to each sorghum sample, and the mixture was stirred in time. After the stirring, the samples were placed in a constant temperature incubator at 42°C;

[0069] 3. Sampling

[0070] Sampling of each sorghum sample was carried out before the first simulated watering, 1.3 h after the first simulated watering (i.e. before the second simulated watering), 6.3 h after the first simulated watering (i.e. before the third simulated watering), 24 h after the first simulated watering, and 54 h after the first simulated watering in the simulated grain moistening process. 10 g of sorghum was sampled each time. After sampling, the sorghum samples were immediately stored in a 4°C refrigerator. The state of the samples at the C5 stage was as follows: Figure 2 As shown;

[0071] 4. Detection of dolin content

[0072] The dulin content of the sorghum samples was tested separately, and the specific steps were as follows:

[0073] (1) Sample pretreatment

[0074] Take 2g of the sorghum sample in a 50mL centrifuge tube, add 20mL of 95% ethanol, homogenize with an automatic homogenizer for 50s, vortex at 2000rpm for 5min, ultrasonicate in an ice bath for 20min, vortex at 2000rpm for 5min, centrifuge at 6000rpm for 5min, take the supernatant and filter through a membrane to obtain the test solution;

[0075] (2) Determination

[0076] The content of dulin in the test solution obtained in step (1) is measured based on chromatography-mass spectrometry technology to obtain the dulin content corresponding to the sorghum samples under different simulated grain moistening conditions;

[0077] Among them, the dullin content of the sorghum sample taken before the first simulated watering was recorded as C1, the dullin content of the sorghum sample taken 1.3 hours after the first simulated watering was recorded as C1, the dullin content of the sorghum sample taken 6.3 hours after the first simulated watering was recorded as C3, the dullin content of the sorghum sample taken 24 hours after the first simulated watering was recorded as C4, and the dullin content of the sorghum sample taken 54 hours after the first simulated watering was recorded as C5;

[0078] The ultra-high performance liquid chromatography conditions used in the chromatography-mass spectrometry technology are as follows: a Hypersil Gold aQ liquid chromatography column (2.1 mm × 150 mm, 1.9 μm) was selected, the injection volume was 5 μL; the mobile phase: phase A was ultrapure water containing 5 mmol / L ammonium acetate and 1‰ formic acid, and phase B was methanol; flow rate: 350 μL / min; gradient elution: the elution program is shown in Table 1; the entire analysis process took a total of 10 min.

[0079] Table 1 Gradient elution program

[0080]

[0081] The conditions of the quadrupole electrostatic field orbital trap high-resolution mass spectrometry used in the chromatography-mass spectrometry technology are: positive ion mode, sheath gas flow rate of 40arb; auxiliary gas flow rate of 15arb; purge gas of 0arb; ion transfer tube temperature of 280℃; spray voltage of 4.5kV; scan mode of PRM, NCE of 10, where the parent ion of Dulin is m / z: 329.1343, and the daughter ion is m / z: 132.0441;

[0082] 5. Calculation of Durin Content

[0083] Based on the dulin content in each sorghum sample measured in the above manner, the dulin content in the dry basis sample was obtained after correction with the sample moisture content. The dulin content obtained from three parallel samples was averaged to obtain C1, C2, C3, C4, and C5 corresponding to different sorghum samples (sorghum stored for 1 year, sorghum stored for 3 months, and sorghum stored for 1 year after being fried). The dulin content (dry basis) corresponding to each sorghum sample is shown in Table 2:

[0084] Table 2 Results of the dulin content of each sorghum sample at each sampling stage

[0085]

[0086] Substitute the data of the dullin content of the above sorghum samples obtained at different sampling stages into the following sorghum dullin content calculation formula: E = 10% C1 + 5% C2 + 5% C3 + 32% C4 + 48% C5, and the dullin content of each sorghum sample is shown in Table 3:

[0087] Table 3 Results of the dullin content of each sorghum sample

[0088] Serial number sample Dulin content (μg / kg) 1 1 year sorghum 13327.8 2 March Sorghum 40853.9 3 1 year old sorghum after frying 7945.7

[0089] In order to evaluate the authenticity of the simulated grain moistening step, this application takes sorghum with a storage time of 1 year as an example, and measures the change of its durin content during the actual production process. The comparison between its durin content and the simulated grain moistening state is shown in Table 4:

[0090] Table 4: Comparison of dullin content in sorghum stored for one year under simulated moistening conditions and during actual production

[0091]

[0092] In the Durin calculation formula provided in this embodiment, the initial value in the sorghum sample has a weight of 10%. The main reason is that the present application found that the Durin content is a dynamically changing process during the winemaking process. At the beginning, the vitality of the sorghum is average, and there is still a long time before the start of steaming. Therefore, the initial Durin value before moistening the grain has a limited effect on the amount of Durin that eventually enters the winemaking system; in the C2 and C3 stages, due to the addition of water above 95°C and the high temperature, the background value of Durin in the sorghum decomposes at high temperature, resulting in a decrease in the Durin content, and the duration is only 1.3 hours and 6.3 hours, and there is still a long time before the start of steaming. The amount of Durin that eventually enters the winemaking system is limited, but it cannot be without weight, because the absorption process of sorghum is also It is the preparation stage for the life activities of sorghum. Therefore, before the second and third watering, the weight of the durin content of the durin content is set to 5% in the present invention; after the third watering, the sorghum fully absorbs water, the life activities of the sorghum grains are started, and durin is gradually generated under the action of enzymes. Sorghum of different varieties, different processing technologies, and different storage time conditions show different vitality at this stage, which leads to different abilities to generate durin. The present application has found that the closer the sorghum durin content is to the upper steaming stage, the greater the impact on the final result, and the greater the possibility that durin will eventually enter the brewing system. Therefore, the weight of the sorghum durin content in the 24th hour and the 54th hour is set to 32% and 48% respectively. In summary, the present invention further sets the calculation formula of the durin content of sorghum as:

[0093] E=10%C1+5%C2+5%C3+32%C4+48%C5.

[0094] According to the results presented in Table 3, after storing sorghum for different periods of time or frying sorghum, the content of sorghum was measured and it was found that the ability of sorghum to generate sorghum in the process of simulating grain moistening was different and the difference was large. Since the storage time and frying had an impact on the vitality of sorghum, this method took into account the impact of the vitality of sorghum used for brewing on the generation of sorghum, and the evaluation method was more scientific. In addition, the sorghum sample with a storage period of 3 months had a higher ability to generate sorghum than the sorghum of 1 year, indicating that proper storage of sorghum can reduce the generation of sorghum in the process of moistening to a certain extent, thereby reducing the generation of EC precursors in liquor, which has positive significance for liquor brewing. The content of sorghum after frying was significantly lower than that of sorghum without frying, indicating that the same kind of sorghum will affect the ability of sorghum to generate sorghum under different processing conditions. It is suggested that when the present invention is applied to the screening of sorghum planting areas with low durlin production ability, under the condition that the yield and quality of sorghum are not affected, the lower the durlin content, the more beneficial it is to control EC, and the higher the sorghum quality of the corresponding planting area is; when the present invention is applied to the breeding of sorghum varieties, under the condition that the output of liquor and the flavor of base liquor are not affected, the lower the durlin content, the more beneficial it is to control EC, and the higher the sorghum quality of the corresponding sorghum varieties is; when the present invention is applied to the screening of sorghum processing technology, under the condition that the output of liquor and the flavor of base liquor are not affected, the lower the durlin content, the more beneficial it is to control EC, and the higher the sorghum quality under the corresponding processing technology conditions is; when the present invention is applied to the optimization of sorghum storage time, under the condition that the output of liquor and the flavor of base liquor are not affected, the lower the durlin content, the more beneficial it is to control EC, and the higher the sorghum quality of the corresponding storage time is.

[0095] In addition, according to the results presented in Table 4, the method provided by the present invention takes into account the possibility of changes in the durin content of sorghum during the actual production process, implements dynamic measurement of the durin content of sorghum during the actual production process, and improves the scientific nature of the method for evaluating the quality of sorghum for wine.

[0096] In addition, in order to accurately and objectively realize the quality evaluation of sorghum samples corresponding to different storage times, the present invention further explores the simulated grain moistening process and parameters provided by the present invention: such as the confirmation of the sampling amount of sorghum samples, the reasonable amount of sorghum samples in the simulated grain moistening experiment is one of the key factors for the success of the experiment. When the sample amount is too small, firstly, the water content of the sample is easily lost, and secondly, because a certain amount of sample is taken from the sample for measurement each time, 5 samplings will bring a certain degree of error to the total amount of sample; but when the sample amount is too large, it will take up too much space in the constant temperature incubator and there will be a certain degree of food waste; combined with the actual situation of experimental operability and convenience, this experiment In the embodiment, the sample size of the simulated experiment of the sorghum sample is 400g, and the experimental container is a 1000mL beaker; in addition, the watering ratio and watering time are determined. Water is one of the key factors affecting the vitality of sorghum. The watering temperature, watering ratio and watering time interval determine the water absorption of the sorghum sample, which in turn affects the vitality of sorghum. Different vitality affects the sorghum's ability to generate turin. In order to simulate the actual grain moistening conditions as much as possible, the present invention explores the watering temperature, watering ratio and watering time interval. Finally, it is found that the watering temperature is selected to be greater than 95°C, and the watering is divided into three times. The first watering is set to 34%, and the first watering is set to 100%. After stirring evenly, the second watering was carried out after an interval of 120 minutes, and the mixture was stirred immediately after the second watering. After the second watering, the third watering was carried out after an interval of 300 minutes, and the mixture was stirred immediately after the third watering. After mixing evenly, the mixture was placed in a constant temperature box at 42°C for cultivation. The data on the content of durin in the obtained samples was closest to the content of durin measured under the actual conditions of sand making and grain moistening. For the determination of the sampling time node, the sampling time was set at a time that did not affect the sample cultivation and reflected the situation of the generation of durin at each key node. Therefore, the sampling time was set at the 0th hour, that is, before the first watering. At this time, the sorghum sample reflected the initial value. The content in the sorghum sample at this time was not high, and the sorghum grains were in The dormant state, at which the content of durin reflects the initial value of the sorghum sample; the second sampling point is set before the second watering, and the content of durin in the sorghum sample is measured 120 minutes after the first watering. The actual data show that due to the thermal decomposition of durin, the content of durin in the sorghum sample will decrease before the second watering; the third sampling point is set before the third watering, which reflects the content of durin in the sorghum sample 300 minutes after the second watering; combined with the actual situation, the second and third data show the content of durin in the process of sorghum absorbing water. At this time, it is still in the water absorption stage, and the germination ability of sorghum has not been fully triggered;The fourth and fifth sampling time points were set at 24h and 54h after the first watering. The sorghum samples entered the germination life stage and showed obvious germination. At this time, the metabolic activity in the sorghum samples was vigorous, and different varieties of sorghum samples metabolized different levels of dolin content. Therefore, two nodes were set to determine the dolin content level of sorghum in the germination stage. By the 54th hour, it had reached the last time limit before the sauce-flavor liquor was steamed, so the fifth sampling time point was set at the 54th hour. ;

[0097] Effect example

[0098] 1. Standard working curve, linear relationship, and quantitative limit

[0099] Preparation of standard solution: Accurately weigh a certain amount of Duling standard, dissolve it with 0.1mol / L phosphoric acid water and dilute it to a 100mL volumetric flask to obtain 5mg / L Duling standard stock solution, and store it in a refrigerator at 4℃. Dilute the standard stock solution step by step with 95% ethanol to obtain standard working solutions with a mass concentration gradient of 10μg / L, 25μg / L, 50μg / L, 100μg / L, 250μg / L, 500μg / L, and 1000μg / L;

[0100] Based on the detection method in Example 1, the standard working solutions of different concentrations were detected, and a standard curve was drawn with the peak area (Y) as the ordinate and the mass concentration (X) as the abscissa to obtain the linear regression equation and correlation coefficient (R 2 ), the method detection limit (LOD) was determined according to the signal to noise ratio (S / N) = 3, and the method quantitation limit (LOQ) was determined according to S / N = 10. The results are shown in Table 5. As shown in Table 5, Du Lin has a good linear relationship within the mass concentration range, with a correlation coefficient of 0.9999; the method detection limit is 0.6μg / kg, and the quantitation limit is 4.5μg / kg.

[0101] Table 5 Standard curve parameter information table

[0102]

[0103] 2. Precision and recovery experiment

[0104] The standard solution of dolin was added to the sorghum sample, and the spiked mass concentrations were 1.0 mg / kg, 3.0 mg / kg, and 5.0 mg / kg, respectively. The pretreatment was carried out according to the above method, and then the instrument analysis was repeated 6 times. The recovery rate and relative standard deviation (RSD) of the method were calculated, and the results are shown in Table 6. As can be seen from Table 6, the recovery rates of the three added levels of dolin are between 93.5% and 108.3%, and the relative standard deviation of the precision experimental results is between 3.4% and 6.8%. It shows that this method has good accuracy, precision and repeatability, and can meet the quantitative determination of dolin in sorghum samples.

[0105] Table 6 Du Lin spike recovery

[0106]

[0107] Comparative Example 2:

[0108] Based on Example 1, this comparative example further explores the influence of using different methods to calculate the durin content of different sorghum samples (the sorghum samples include sorghum stored for 1 year, sorghum stored for 3 months, and sorghum stored for 1 year after being stir-fried) on the evaluation of sorghum quality. The difference between the comparative example and Example 1 is that in step 5, the durin calculation method used is different, as follows:

[0109] Calculation method 1: E1 = 100% C1;

[0110] Calculation method 2: E2 = 20% C1 + 20% C2 + 20% C3 + 20% C4 + 20% C5;

[0111] Calculation method 3: E3 = 10% C1 + 5% C2 + 5% C3 + 38% C4 + 42% C5;

[0112] Calculation method 4: E4 = 20% C1 + 5% C2 + 5% C3 + 20% C4 + 50% C5;

[0113] Calculation method 5: E5 = 20% C1 + 10% C2 + 10% C3 + 20% C4 + 40% C5;

[0114] Calculation method 6: E6 = 10% C1 + 10% C2 + 10% C3 + 30% C4 + 40% C5;

[0115] The dolin content in each sorghum sample was calculated by the above method, and the results are shown in the following table:

[0116] Table 7 The influence of different calculation methods of sorghum dorlin content E on the final calculation results

[0117]

[0118] If the evaluation is conducted in the first way, it is found that the E value of sorghum in March is the smallest, which is inconsistent with the actual sorghum content of sorghum in production. Therefore, it is not scientific to evaluate sorghum only by considering the sorghum content in the initial value. If the evaluation is conducted in the second way, the weights are evenly distributed, and the rationality, scientificity and logic of the stage of sorghum content in the life state of each node are not taken into account. If the evaluation is conducted in the third way, the influence of possible life activities on sorghum production in C4 and C5 stages is taken into account. This calculation method can not only distinguish the sorghum content of each sorghum sample well, but also well reflects the scientificity and rationality.

[0119] Comparative Example 2: Exploring the effects of different culture temperatures on the dullin content of sorghum

[0120] For the determination of the temperature conditions of the constant temperature incubator, the simulated grain moistening temperature is one of the decisive factors for the success of the simulation experiment. This comparative example is based on Example 1, and further explores the influence of different culture temperatures on the evaluation results of sorghum dullin content in the simulated grain moistening process. The difference between this comparative example and Example 1 is that the culture temperature is different, and the dullin content is compared based on the 54th hour dullin content (dry basis), specifically: at 4°C, 20°C, and 42°C, after three waterings according to the experimental conditions, the dullin content data is measured at the 54th hour. The results are shown in Table 8 below:

[0121] Table 8 Du Lin content at 54h after simulated grain moistening under different culture temperature conditions

[0122]

[0123] According to the results shown in Table 8, the data on the dullin content in the sample obtained at 54 hours after grain moistening at 42°C is closest to the dullin content measured under actual sand-making and grain moistening conditions. Therefore, choosing 42°C as the temperature condition for the grain moistening simulation experiment can truly reflect the dullin formation in sorghum.

[0124] It is to be understood that the present invention is described by some embodiments, and it is known to those skilled in the art that various changes or equivalent substitutions may be made to these features and embodiments without departing from the spirit and scope of the present invention. In addition, under the teachings of the present invention, these features and embodiments may be modified to adapt to specific circumstances and materials without departing from the spirit and scope of the present invention. Therefore, the present invention is not limited by the specific embodiments disclosed herein, and all embodiments falling within the scope of the claims of this application are within the scope of protection of the present invention.

Claims

1. A method for evaluating the quality of sorghum for wine, characterized in that: The steps include: (1) Simulated grain moistening: simulated grain moistening of the sorghum to be evaluated; (2) Sampling: Sampling sorghum at different simulated grain moistening stages, and measuring the durin content in the sorghum samples corresponding to different stages; (3) Evaluation: Calculating the dulin content of the sorghum to be evaluated based on the dulin content in the sorghum samples corresponding to different stages measured in step (2), and evaluating the quality of the sorghum to be evaluated based on the calculated dulin content of the sorghum to be evaluated.

2. The method according to claim 1, characterized in that The simulated grain moistening treatment of the sorghum to be evaluated includes sequentially performing simulated watering treatments on the sorghum to be evaluated.

3. The method according to claim 2, characterized in that The simulated watering comprises the following steps: adding water to the sorghum sample to be evaluated and placing it in an incubator for incubation; Preferably, the simulated grain moistening treatment of the sorghum to be evaluated includes sequentially performing simulated watering treatment after crushing the sorghum to be evaluated; Preferably, the simulated grain moistening treatment of the sorghum to be evaluated includes sequentially performing simulated watering treatment after crushing the sorghum to be evaluated; The crushing degree of the sorghum to be evaluated is 20% to 100%; preferably, the simulated watering treatment includes: a first simulated watering, a second simulated watering, and a third simulated watering; the crushing degree of the sorghum to be evaluated in the simulated watering treatment is 25%-35%; Preferably, the watering ratio used in the first simulated watering is 30-40% of the weight of the sorghum to be evaluated; the temperature of the water used in the first simulated watering is 90-100°C; the culture temperature of the first simulated watering is 38-45°C; Preferably, the watering ratio used in the second simulated watering is 8-15% of the weight of the sorghum to be evaluated, the temperature of the water used in the second simulated watering is 90-100° C., and the cultivation temperature of the second simulated watering is 38-45° C.; Preferably, the watering ratio used in the third simulated watering is 8-15% of the weight of the sorghum to be evaluated, the temperature of the water used in the second simulated watering is 90-100°C; and the cultivation temperature of the third simulated watering is 38-45°C.

4. The method according to claim 1, characterized in that: The sampling of sorghum at different simulated grain moistening stages includes: sampling the sorghum to be evaluated in the simulated grain moistening process before the first simulated watering, 1.0-1.5h after the first simulated watering, 6.0-6.5h after the first simulated watering, 20-28h after the first simulated watering, and 50-54h after the first simulated watering.

5. The method according to claim 1, characterized in that The method of calculating the durin content of the sorghum to be evaluated based on the durin content in the sorghum samples corresponding to different stages determined in step (2) comprises: using a dynamically changing weighted summation method, assigning a smaller weighting coefficient to the durin content of the sorghum samples close to the start stage of simulated grain moistening, and assigning a larger weighting coefficient to the durin content of the sorghum samples close to the end stage of simulated grain moistening among the different sorghum samples obtained.

6. The method according to claim 1, characterized in that The step of calculating the durin content of the sorghum to be evaluated based on the durin content in the sorghum samples corresponding to different stages measured in step (2) comprises: The following calculation formula is used for calculation: E = X1C1 + ... + X n Cn, where E is the dullin content of the sorghum to be evaluated, 1 to n represent the different simulated grain moistening stages of the samples taken from the beginning to the end of the simulated grain moistening, C represents the dullin content measured in the sorghum samples at the corresponding simulated grain moistening stages, and X represents the weighting coefficient, where X1+…+X n＝ 100%; Preferably, the step of calculating the durin content of the sorghum to be evaluated based on the durin content in the sorghum samples corresponding to different stages determined in step (2) comprises: calculating using the following calculation formula: E=X1C1+X2C2+X3C3+X4C4+X5C5; wherein E is the durin content of the sorghum to be evaluated, the value range of X1 is 8%-15%, the value range of X2 is 2%-8%, the value range of X3 is 2%-8%, the value range of X4 is 25%-35%, the value range of X5 is 40%-55%, and the value range of X1+X2+X3+X4+X5C5 is 10%. 5＝ 100%; C1 is the content of dolin measured by sampling before the first simulated watering, C2 is the content of dolin measured by sampling 1.0-1.5h after the first simulated watering, C3 is the content of dolin measured by sampling 6.0-6.5h after the first simulated watering, C4 is the content of dolin measured by sampling 20-28h after the first simulated watering, and C5 is the content of dolin measured by sampling 50-54h after the first simulated watering.

7. The method according to claim 1, characterized in that The simulated grain moistening treatment of the sorghum to be evaluated also includes placing the sorghum to be evaluated in a container and performing simulated watering treatment on the sorghum to be evaluated in sequence; the ratio of the weight of the sorghum to be evaluated to the capacity of the container is: 1:4g / mL to 1:2g / mL.

8. The method according to claim 1, characterized in that The determining of the durin content in the sorghum samples corresponding to different stages respectively comprises: determining the durin content in the sorghum samples based on chromatography-mass spectrometry technology; Preferably, the chromatography-mass spectrometry is ultra-high performance liquid chromatography-high resolution mass spectrometry, and the parameters of the ultra-high performance liquid chromatography used include: the chromatographic column is: Hypersil Gold aQ liquid chromatography column; Preferably, the chromatographic parameters also include: injection volume 3.5 to 7.5 μL; Preferably, the chromatographic parameters also include: mobile phase: phase A is ultrapure water containing 5mmol / L ammonium acetate and 1‰ formic acid, and phase B is methanol; Preferably, the chromatographic parameters also include: flow rate: 300-400 μL / min; Preferably, the mass spectrometer is a quadrupole electrostatic field orbital trap high-resolution mass spectrometer, and the conditions of the quadrupole electrostatic field orbital trap high-resolution mass spectrometer include: positive ion mode, sheath gas flow rate of 30-50arb, auxiliary gas flow rate of 10-20arb, purge gas of 0arb, ion transfer tube temperature of 260-300°C, spray voltage of 4-5kV, back purge temperature of 360-400°C, scan mode of PRM, NCE of 10, wherein the parent ion of Du Lin is m / z: 329.1343, and the daughter ion is m / z: 132.0441.

9. The method according to claim 1, characterized in that: The determination of the durin content in the sorghum samples corresponding to different stages comprises: pre-treating the samples before the determination, wherein the pre-treatment comprises: adding ethanol to the sampled sorghum, homogenizing, vortexing, centrifuging, taking the supernatant, filtering the membrane, and obtaining the liquid to be analyzed; Preferably, the pretreatment includes: weighing 1-5 g of sorghum sample, adding 10-50 mL of 95% ethanol, homogenizing with an automated homogenizer for 45-120 s, then vortexing for 3-10 minutes, then ultrasonicating in an ice bath for 15-35 minutes, then vortexing for 3-10 minutes, then centrifuging for 1-10 minutes, and taking the supernatant and filtering it through a membrane to obtain the liquid to be analyzed.

10. Use of the method according to any one of claims 1 to 9 in the quality evaluation of sorghum for wine.

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