Thin-layer identification method and application of brown malt formula granules

Abstract

The invention provides a thin-layer method for identifying caramel malt formula granules and application, and the thin-layer method for identifying the caramel malt formula granules comprises the following steps: preparing a contrast sample application sample by using caramel malt decoction pieces, and preparing a sample application sample for test by using the caramel malt formula granules, a developing solvent adopted by the thin-layer method is n-hexane-ethyl acetate-methanol with the volume ratio of 5: (9-11): 1, after sample application, a developing plate is placed in an ammonium hydroxide pre-saturated developing cylinder to be developed, taken out, aired, sprayed with a potassium iodide iodine test solution, volatilized to be dry and placed under the sunlight to be inspected. According to the method, the caramel malt decoction pieces are used as control decoction pieces, the specificity is high, and the method is suitable for identifying related preparations such as caramel malt aqueous extracts and formula granules.

Description

Technical Field

[0001] This invention belongs to the field of traditional Chinese medicine detection and analysis technology, specifically providing a thin-layer chromatography identification method and application for charred malt formula granules. Background Technology

[0002] Malt is barley, a plant belonging to the Poaceae family. Hordeum vulgare L. malt is a processed product made by germinating and drying the mature fruit of L. Roasted malt is a type of processed malt. Malt is stir-fried in a wok over medium heat until it turns a dark brown color, then removed and cooled. Raw malt has the effects of strengthening the spleen and stomach, and soothing the liver and regulating qi. It is mainly used for spleen deficiency with poor appetite and milk stasis. Roasted malt is mainly used to promote digestion and resolve stagnation. It is used for indigestion and abdominal distension and pain. Roasted malt granules are preparations made under the guidance of traditional Chinese medicine theory from roasted malt slices through extraction, concentration, drying, and granulation.

[0003] Thin-layer chromatography (TLC) uses simple instruments and is easy to operate, allowing for the analysis of drug chemical components in a short time, and has been widely used in the analysis and testing of traditional Chinese medicine. The 2025 edition of the Chinese Pharmacopoeia specifies a TLC identification method using malt as a reference material under the malt section. However, this method involves complex sample preparation and requires two development steps, resulting in a long processing time. Furthermore, the developing solvent contains a large proportion of toluene, a highly toxic reagent that can cause skin irritation, drowsiness, or dizziness; it is also a precursor chemical for toxic substances. Moreover, this TLC method is not suitable for the identification of charred malt formulation granules, mainly because a certain amount of excipients needs to be added during the preparation of charred malt formulation granules, and these excipients can interfere with TLC separation. In addition, there are some easily confused products among charred malt formulation granules, such as charred barley sprout formulation granules and charred rice sprout formulation granules. These easily confused products are difficult to distinguish by appearance, necessitating a simple and quick method to control the quality of these formulation granules.

[0004] Therefore, establishing a method for identifying roasted malt and its preparations is of great significance for the quality control of roasted malt and its formulation granules. Summary of the Invention

[0005] This invention aims to at least partially solve one of the technical problems in the prior art. Therefore, one object of this invention is to provide a thin-layer chromatography method and its application for identifying roasted malt formulation particles.

[0006] In a first aspect, the present invention provides a thin-layer chromatography method for identifying granules of roasted malt formulations, comprising: The control sample was prepared using roasted malt slices, and the test sample was prepared using roasted malt granules. After spotting, the developing plate was placed in a developing tank pre-saturated with ammonia water for development. The developing solvent was n-hexane-ethyl acetate-methanol with a volume ratio of 5:(9-11):1, preferably 5:10:1. The plate was then removed, dried, sprayed with potassium iodide iodine solution, evaporated, and examined under sunlight.

[0007] The thin-layer chromatography method provided by this invention, using roasted malt slices as a reference, exhibits high specificity and is suitable for the identification of roasted malt water extracts, formulation granules, and other related preparations. The developing solvent used in this invention effectively separates the test sample and the reference slice solution, both exhibiting identical thin-layer spots. This effectively overcomes the interference of excipients in roasted malt formulation granules, enabling effective identification of roasted malt formulation granules. This invention requires only one development step, significantly shortening the experimental time and improving detection efficiency. It also avoids the use of toxic solvents such as benzene. The developing solvent used in this invention has low toxicity, good separation effect, and uses potassium iodide iodine solution as a specific colorimetric reagent, resulting in clear spots that can be examined under sunlight, making the operation simple.

[0008] It should be noted that the thin-layer method of the present invention can be used not only for roasted malt formulation granules, but also for the identification of roasted malt water extracts, which are the materials that were not added with excipients in the early stage of the preparation of roasted malt formulation granules.

[0009] In some embodiments of the present invention, the method for preparing test sample spotting using charred malt formulation granules includes: After grinding the malt extract granules, add ammonia water and soak in a sealed container. Then add dichloromethane for ultrasonic extraction. After filtration, evaporate the filtrate and dissolve the residue.

[0010] Compared to other methods, the sample prepared using the method described above in this invention reduces impurities such as acids, resulting in a cleaner background and clearer spots in the thin-layer chromatography (TLC) image. If the sample contains alkaloids, the weakly alkaline environment provided by ammonia can release the potential alkaloids. Furthermore, dichloromethane has good solubility for neutral and weakly polar organic molecules, allowing this preparation method to maximize sample extraction.

[0011] In some embodiments of the present invention, the mass ratio of the roasted malt formula granules to the volume ratio of the ammonia water is (2-6) g: 10 ml, and preferably the concentration of the ammonia water is 25-28%.

[0012] In some embodiments of the present invention, the soaking time is 15-30 minutes.

[0013] In some embodiments of the present invention, the volume ratio of the ammonia water to the dichloromethane is 1:(2-4).

[0014] In some embodiments of the present invention, the solvent for dissolving the filtrate residue is a mixed solution of isopropanol and dichloromethane, preferably with a volume ratio of (1-2):(1-2), more preferably 1:1. The present invention uses a mixed solution of isopropanol and dichloromethane as the solvent for dissolving the residue. Isopropanol is highly polar and can help dissolve components that are difficult to dissolve in dichloromethane. Using either isopropanol alone would reduce the development effect of the sample. If isopropanol is replaced with methanol, although methanol is more polar, its volatility is poor, which would make it difficult for the spots to dry during spotting, affecting the color development.

[0015] In some embodiments of the present invention, the ultrasonic extraction time is 10-30 min.

[0016] In some embodiments of the present invention, the method for preparing the control spotting sample using roasted malt slices includes: The roasted malt slices were heated under reflux with water, centrifuged to obtain the supernatant, and then concentrated to dryness under reduced pressure. The first residue was soaked in ammonia water, then treated with dichloromethane by ultrasound, filtered, and the filtrate was evaporated to dissolve the second residue.

[0017] Compared to other methods, the control spotting sample prepared by the method described above in this invention can maintain the same number of spots as the sample from charred malt granules or other preparations. Since charred malt granules are water extracts, the preparation of the sample after water extraction can better serve as a control for characterization.

[0018] In some embodiments of the present invention, based on 10g of the roasted malt slices, the amount of water added is 80-120ml, the volume of ammonia added is 15-25ml, the volume of dichloromethane added is 20-40ml, and preferably the concentration of ammonia is 25-28%.

[0019] In some embodiments of the present invention, the heating reflux time is 0.8-1.5 h.

[0020] In some embodiments of the present invention, the soaking time is 10-30 minutes.

[0021] In some embodiments of the present invention, the ultrasonic treatment time is 10-30 min.

[0022] In some embodiments of the present invention, the solvent for dissolving the second residue is a mixed solution of isopropanol and dichloromethane, preferably the volume ratio of isopropanol to dichloromethane in the mixed solution is (1-2):(1-2), more preferably 1:1.

[0023] In some embodiments of the present invention, the amount of the sample spotting specimen is 20-40 µl.

[0024] In some embodiments of the present invention, the amount of the control sample is 20-40 µl.

[0025] In some embodiments of the present invention, the ambient temperature for unfolding the thin-layer plate is 8-23°C.

[0026] In some embodiments of the present invention, the ambient humidity during the unfolding of the thin-layer plate is 22-79%.

[0027] In some embodiments of the present invention, the thin-layer plate used in the thin-layer method includes a silicone G plate.

[0028] In some embodiments of the present invention, the spot shift values ​​on the thin-layer plate of the roasted malt formulation granules are 25-28% from the spotting point to the development direction of the developing agent.

[0029] In a second aspect, the present invention proposes the application of the above-mentioned thin-layer chromatography method for identifying malt-based formulation particles in the identification of malt-based formulation particles, malt-based rice formulation particles, and malt-based barley formulation particles.

[0030] Compared with the prior art, the present invention can achieve at least one of the following beneficial effects: (1) This invention uses roasted malt slices as a reference slice, which has strong specificity and is suitable for the identification of roasted malt water extract, formula granules and other related preparations. The developing solvent used in this invention can effectively separate the test sample and the reference slice solution. The two have the same thin-layer spots, which effectively overcomes the interference of excipients in roasted malt formula granules and can effectively identify roasted malt formula granules.

[0031] (2) This invention only requires one development, which greatly shortens the experimental time, improves the detection efficiency, and avoids the use of benzene-based toxic solvents. The developing agent used in this invention has low toxicity and good separation effect. Furthermore, potassium iodide iodine solution is used as the exclusive colorimetric agent, and the spots are clearer and can be inspected under sunlight. The operation is simple. Attached Figure Description

[0032] Figure 1 This is a thin-layer chromatogram of Example 1; Figure 2 The thin-layer chromatogram of Example 2; Figure 3 The thin-layer chromatogram of Example 3; Figure 4 The thin-layer chromatogram is shown in Example 4. Figure 5 The thin-layer chromatogram of Example 5; Figure 6 The thin-layer chromatogram of Example 6; Figure 7 The thin-layer chromatogram of Example 7; Figure 8 The thin-layer chromatogram of Example 8; Figure 9 The thin-layer chromatogram of Example 9; Figure 10 The thin-layer chromatogram of Example 10; Figure 11 The thin-layer chromatogram of Example 11; Figure 12 This is the thin-layer chromatogram of Comparative Example 1; Figure 13 This is the thin-layer chromatogram of Comparative Example 2; Figure 14 This is the thin-layer chromatogram of Comparative Example 3; Figure 15 This is the thin-layer chromatogram of Comparative Example 4. Detailed Implementation

[0033] Based on the embodiments of this invention, all other embodiments obtained by those skilled in the art without inventive effort are within the scope of protection of this invention. The invention will now be described with reference to specific embodiments. It should be noted that these embodiments are merely descriptive and do not limit the invention in any way.

[0034] In the embodiments and comparative examples of the present invention: Roasted malt slices (roasted malt reference slices) are prepared by stir-frying clean malt according to the stir-frying method (General Chapter 0213 of the 2025 edition of the Chinese Pharmacopoeia) until it turns dark brown, cooling it, and sifting out the ash.

[0035] The freeze-dried powder of standard roasted malt decoction is made by soaking 100g of roasted malt slices for 30 minutes, then decocting twice. For the first decoction, add 9 times the amount of water as the amount of malt slices and decoct for 30 minutes. For the second decoction, add 6 times the amount of water as the amount of malt slices and decoct for 20 minutes. Filter the decoction while it is still hot through a 150-mesh filter cloth. Combine the two decoctions, concentrate under reduced pressure until the material-to-liquid ratio is about 1:1, and then freeze-dry to obtain the final product.

[0036] The roasted malt formula granules are made by decocting 4700g of roasted malt slices with water, filtering, concentrating the filtrate into a clear extract (dry extract yield is 11% to 19%), adding an appropriate amount of excipients, drying (or drying and pulverizing), adding another appropriate amount of excipients, mixing well, granulating, and making 1000g of the product.

[0037] The granules of roasted barley sprouts are made by decocting roasted barley sprout slices with water, filtering, concentrating the filtrate into a clear paste, adding an appropriate amount of excipients, drying (or drying and pulverizing), adding an appropriate amount of excipients again, mixing well, and granulating to obtain the final product.

[0038] The formula granules for roasted rice sprouts are made by decocting roasted rice sprout slices with water, filtering, concentrating the filtrate into a clear paste, adding an appropriate amount of excipients, drying (or drying and pulverizing), adding an appropriate amount of excipients again, mixing well, and granulating to obtain the final product.

[0039] Potassium iodide iodine test solution: Dissolve 0.5 g of iodine and 1.5 g of potassium iodide in 25 ml of water.

[0040] The hexanol, ethyl acetate, methanol, dichloromethane, and ammonia (ammonia concentration 25-28%) used in the examples and comparative examples are all commercially available products.

[0041] Example 1 (1) Preparation of the sample solution for spotting the test sample: Take 4g of malt formula granules, grind them finely, add 10ml of ammonia water, seal tightly, soak for 20 minutes, add 30ml of dichloromethane, sonicate for 20 minutes, filter, evaporate the filtrate to dryness, add 1ml of isopropanol:dichloromethane (volume ratio of 1:1) mixed solution to dissolve the residue, and use it as the sample solution for spotting the test sample.

[0042] (2) Preparation of control spotting sample solution: Take 10g of roasted malt slices (passed through No. 1 sieve), add 100ml of water, heat under reflux for 1 hour, centrifuge (3500 rpm) for 10 minutes, take the supernatant, concentrate under reduced pressure to dryness, add 20ml of ammonia water to the residue, seal tightly, soak for 20 minutes, add 30ml of dichloromethane, sonicate for 20 minutes, filter, evaporate the filtrate to dryness, add 0.5ml of isopropanol:dichloromethane (volume ratio of 1:1) mixed solution to dissolve the residue, and use it as the control spotting sample solution.

[0043] (3) Thin-layer chromatography conditions: Perform thin-layer chromatography (General Chapter 0502 of Chinese Pharmacopoeia 2025). Take 30 μl of the test sample solution and the control sample solution and spot them on the same silica gel G thin-layer plate. Use n-hexane-ethyl acetate-methanol (volume ratio of 5:10:1) as the developing solvent. Develop in a developing tank pre-saturated with ammonia (double tank developing tank, add 5 ml of ammonia to the tank and pre-saturate for 20 minutes). Remove, air dry, spray with potassium iodide iodine test solution, evaporate to dryness, and examine under sunlight.

[0044] See results Figure 1 , Figure 1 In the diagram, 1 represents roasted malt slices, 2 represents roasted malt granules (K395CP01), 3 represents roasted malt granules (K395CP02), and 4 represents roasted malt granules (K395CP03). The thin-layer chromatographic spots of all three batches of roasted malt formulation granules were clear and reproducible, with the main spot showing R0. f The value is moderate, R f The values ​​of 0.20, 0.39, and 0.46 showed spots of the same color at the corresponding positions as the control granules. Therefore, this thin-layer chromatography method can be used as a thin-layer identification method for charred malt formulation granules.

[0045] Example 2 The difference between Example 2 and Example 1 is that a blank solvent (a mixture of isopropanol and dichloromethane (volume ratio 1:1)) and a negative control (an appropriate amount of maltodextrin was prepared using the same method as the sample solution preparation method for the test sample, and used as a negative control solution) were added.

[0046] See results Figure 2 , Figure 2 In the sample, 1 is the blank solvent, 2 is the negative control, 3 is the roasted malt slices, and 4 is the roasted malt granules test sample. Figure 2 It can be seen that no spots were displayed in the blank solvent area, the negative control did not interfere with the separation spot information, and the spots of the test sample solution of roasted malt slices and roasted malt formula granules were clear and had good specificity.

[0047] Example 3 The difference between Example 3 and Example 1 is that 2g, 4g and 6g of roasted malt formula granules (batch number K395CP01) were taken and the test sample solution was prepared according to the preparation method of Example 1.

[0048] See results Figure 3 , Figure 3 1 is roasted malt slices, 2 is roasted malt formula granules (2g), 3 is roasted malt formula granules (4g), and 6g is roasted malt formula granules. Figure 3 It can be seen that when the sample weight of the roasted malt granules is 4g and 6g, the thin-layer chromatographic spots are clearer and the separation is better. f Moderate.

[0049] Example 4 The difference between Example 4 and Example 1 is as follows: Take 4g of roasted malt formula granule powder (batch number K395CP01) in three portions, add 10ml of ammonia water, seal tightly, soak for 20 minutes, add 30ml of dichloromethane, and ultrasonically treat for 10 minutes, 20 minutes and 30 minutes respectively. Then prepare the test sample solution according to the preparation method of Example 1.

[0050] See results Figure 4 , Figure 4 In the test samples, 1 is roasted malt slices, 2 is roasted malt granules (10 minutes), 3 is roasted malt granules (20 minutes), and 4 is roasted malt granules (30 minutes). Figure 4 It can be seen that the thin-layer chromatographic spots obtained from the charred malt granule test sample at different ultrasonic times were all relatively clear, with good separation. f Moderate.

[0051] Example 5 (Investigation of different sample sizes) The difference between Example 5 and Example 1 is as follows: 20µl, 30µl, and 40µl of the charred malt formula granule test solution (batch number K395CP01) and 20µl, 30µl, and 40µl of the charred malt slice solution were respectively taken and developed under the thin-layer chromatography conditions of Example 1 for color development.

[0052] See results Figure 5 , Figure 5 In the sample, 1 is 20µl of roasted malt granules, 2 is 30µl of roasted malt granules, 3 is 40µl of roasted malt granules, 4 is 20µl of roasted malt slices, 5 is 30µl of roasted malt slices, and 6 is 40µl of roasted malt slices. Figure 5 It can be seen that when the sample volume of the roasted malt granule test solution and the sample volume of roasted malt slices are both 30-40 µl, the thin-layer chromatographic spots are clear and the resolution is good. f Moderate.

[0053] Example 6 (Reproducibility Study) Reproducibility experiments were conducted by different experimenters using the method described in Example 2.

[0054] See results Figure 6 , Figure 6 In the sample, 1 is the blank solvent, 2 is the negative control, 3 is the roasted malt slices, and 4 is the roasted malt granules test sample. Figure 6 It can be seen that, after different experimental operations, no spots were displayed in the blank solvent area, the negative control did not interfere with the separation spot information, and the spots of the test sample solution of roasted malt slices and roasted malt formula granules were clear, meeting the reproducibility requirements.

[0055] Example 7 (Durability Test) The control decoction piece solution and the test solution (batch number K395CP01) were spotted onto the same silica gel G thin-layer plate, and developed under the thin-layer chromatographic conditions of Example 1 at room temperature (23℃) and low temperature (8℃), respectively.

[0056] See results Figure 7 1 is roasted malt slices, and 2 is roasted malt granules. Figure 7 It can be seen that the separation effect of chromatographic spots of malt formulation particles is good under both room temperature and low temperature conditions, indicating that temperature has no significant effect on the thin-layer identification of malt formulation particles, and that the thin-layer identification method has good temperature durability.

[0057] Example 8 (Investigation at different humidity levels) The control decoction piece solution and the test solution (batch number K395CP01) were spotted onto the same silica gel G thin-layer plate and developed under different humidity conditions according to the thin-layer chromatography conditions of Example 1.

[0058] See results Figure 8 , Figure 8 In the sample, 1 is roasted malt slices and 2 is roasted malt granules. Figure 8 It can be seen that the chromatographic spot separation effect of the malt formulation particles is good under both high and low humidity conditions, indicating that different humidity levels do not have a significant impact on the separation effect of the malt formulation particles, and the method has good durability under different humidity conditions.

[0059] Example 9 (Investigation of different thin-layer plates) The control decoction piece solution and the test solution (batch number K395CP01) were spotted onto ordinary silica gel G thin-layer plates from different manufacturers, and the thin-layer chromatography conditions of Example 1 were followed.

[0060] See results Figure 9 , Figure 9 In the sample, 1 is roasted malt slices and 2 is roasted malt granules. Figure 9 It can be seen that the malt formulation granules showed good separation effect on thin-layer plates from three manufacturers, and the difference was not significant, indicating that the method has good durability on silica gel G thin-layer plates from different manufacturers.

[0061] Example 10 (1) Preparation of the sample solution for spotting the test sample: Take 4g of lyophilized malt standard powder sample, grind it finely, add 10ml of ammonia water, seal it tightly, soak for 20 minutes, add 30ml of dichloromethane, sonicate for 20 minutes, filter, evaporate the filtrate to dryness, add 1ml of isopropanol:dichloromethane (volume ratio of 1:1) mixed solution to dissolve the residue, and use it as the sample solution for spotting the test sample.

[0062] (2) The preparation process of the control spotting sample solution is the same as that in Example 1.

[0063] (3) The thin-layer chromatography test method is the same as that in Example 1.

[0064] See results Figure 10 , Figure 10In the table, 1 represents roasted malt slices, 2 represents roasted malt standard freeze-dried powder (K395BJ01), 3 represents roasted malt standard freeze-dried powder (K395BJ02), 4 represents roasted malt standard freeze-dried powder (K395BJ03), 5 represents roasted malt standard freeze-dried powder (K395BJ05), 6 represents roasted malt standard freeze-dried powder (K395BJ06), 7 represents roasted malt standard freeze-dried powder (K395BJ07), 8 represents roasted malt standard freeze-dried powder (K395BJ08), and 9 represents roasted malt standard freeze-dried powder. Powder (K395BJ09), 10 is lyophilized malt standard powder (K395BJ10), 11 is lyophilized malt standard powder (K395BJ11), 12 is lyophilized malt standard powder (K395BJ12), 13 is lyophilized malt standard powder (K395BJ16), 14 is lyophilized malt standard powder (K395BJ17), 15 is lyophilized malt standard powder (K395BJ18), and 16 is lyophilized malt standard powder (K395BJ19). Figure 10 It can be seen that the lyophilized powder test solutions of 15 batches of roasted malt standard decoction showed spots of the same color at the corresponding positions as the control decoction pieces in the chromatogram. The spots were clear, and R... f The value is moderate, and the separation effect is good.

[0065] Example 11 Similar to roasted malt, easily confused with roasted barley sprouts and roasted rice sprouts are other traditional Chinese medicines. This study used the thin-layer chromatography identification method established in Example 1 to identify roasted malt, roasted rice sprout, and roasted barley sprout formulations. Results are shown below. Figure 11 , Figure 11 In the table, 1 represents roasted malt slices; 2-4 represent roasted malt granules; 5-7 represent roasted rice sprout granules; and 8-10 represent roasted rice sprout granules. Figure 11 It can be seen that the thin-layer chromatography of charred malt granules, charred rice sprout granules, and charred rice sprout granules shows significant differences, and the test solution of charred malt granules exhibits distinct characteristics in R... f Clear color spots were observed at the 0.45~0.50 position, while no spots were observed at the corresponding positions in the test solutions of roasted rice sprouts and roasted barley sprouts. Therefore, this method can be used to distinguish roasted malt granules from roasted rice sprouts and roasted barley sprouts.

[0066] Comparative Example 1 Thin-layer chromatography identification method for malt in the 2025 edition of the Chinese Pharmacopoeia: (1) Preparation of the sample solution for spotting the test sample: Take 5g of the powder of this product, add 30ml of anhydrous ethanol, sonicate for 40 minutes, filter, add 1.5ml of 50% potassium hydroxide solution to the filtrate, heat under reflux for 15 minutes, cool in an ice bath for 5 minutes, extract with petroleum ether (30-60℃) three times, 10ml each time, combine the petroleum ether solutions, evaporate to dryness, add 1ml of ethyl acetate to the residue to dissolve, and use as the test sample solution.

[0067] (2) Preparation of control point sample solution: Take 5g of malt control medicinal material and prepare control medicinal material solution in the same way.

[0068] (3) Thin-layer chromatography conditions: Perform thin-layer chromatography (General Chapter 0502 of Chinese Pharmacopoeia 2025). Take 2 μl of each of the above two solutions and spot them separately on the same silica gel G thin-layer plate to form strips. Use toluene-chloroform-ethyl acetate (10:10:2) as the developing solvent, develop, remove, and air dry. Then use toluene-chloroform-ethyl acetate (10:10:1) as the developing solvent, develop, remove, and air dry. Spray with 15% nitric acid ethanol solution and heat at 100℃ until the spots are clearly visible. Examine under ultraviolet light (365nm).

[0069] Figure 12 This is a thin-layer chromatography image of a charred malt sample obtained using pharmacopoeia methods. Figure 12 In the diagram, 1 represents maltodextrin; 2 represents blank solvent; 3 represents charred malt granules (K395CP01); 4 represents charred malt granules (K395CP02); and 5 represents charred malt granules (K395CP03). It can be seen that sample 1 and charred malt granules 3, 4, and 5 all exhibit the same spots at the same locations. Sample 1, being maltodextrin, is an excipient added during the preparation of charred malt granules. If an excipient can show the same spots as the granules, this thin-layer chromatography method cannot identify whether the sample is a charred malt formulation granule. Furthermore, the spots exhibited by charred malt granules under these conditions are not clear enough, with relatively dense spots in the middle section. Therefore, this method is not suitable for the thin-layer chromatography identification of charred malt formulation granules.

[0070] Comparative Example 2 The preparation process of the test sample solution and the control sample solution is the same as in Example 1.

[0071] Take the control decoction piece solution and the test solution (batch number K395CP01) and spot them on the same silica gel G thin layer plate. Under the thin layer chromatographic conditions of Example 1, with n-hexane-ethyl acetate-methanol (volume ratio of 4.5:4.5:2) as the developing solvent, develop in a developing tank pre-saturated with ammonia water (double tank developing tank, add 5 ml of ammonia water to the tank and pre-saturate for 20 minutes). Remove, air dry, spray with potassium iodide iodine test solution, evaporate to dryness, and examine under sunlight.

[0072] Figure 13 In the table, 1 represents the reference roasted malt slices, and 2 represents the roasted malt formulation granules. For example... Figure 13 As shown, the sample spots are relatively concentrated, with multiple spots close together, resulting in poor separation.

[0073] Comparative Example 3 The preparation process of the test sample solution and the control sample solution is the same as in Example 1.

[0074] Take the control decoction piece solution and the test solution (batch number K395CP01) and spot them on the same silica gel G thin layer plate. Under the thin layer chromatographic conditions of Example 1, with n-hexane-ethyl acetate-methanol (volume ratio of 5:13:1) as the developing solvent, develop in a developing tank pre-saturated with ammonia water (double tank developing tank, add 5 ml of ammonia water to the tank and pre-saturate for 20 minutes). Remove, air dry, spray with potassium iodide iodine test solution, evaporate to dryness, and examine under sunlight.

[0075] Figure 14 In the table, 1 represents the reference roasted malt slices, and 2 represents the roasted malt formulation granules. For example... Figure 14 As shown, the two spots in the middle section of the chromatogram have poor separation, and the background color in the lower middle section is darker, with no clear spots.

[0076] Comparative Example 4 The preparation process of the test sample solution and the control sample solution is the same as in Example 1.

[0077] Take the control decoction piece solution and the test sample solution (batch number K395CP01) and spot them on the same silica gel G thin layer plate. Perform thin layer chromatography under the conditions of Example 1, spraying with potassium bismuth iodide reagent, evaporating to dryness, and examining under sunlight.

[0078] Figure 15 In the table, 1 represents the reference roasted malt slices, and 2 represents the roasted malt formulation granules. For example... Figure 15 As shown, the thin layer has not yet shown any spots, so the color developer is not suitable.

[0079] Any numerical value mentioned in this invention, if there is only a two-unit interval between any minimum and any maximum value, includes all values ​​that increase by one unit each time from the minimum to the maximum value. For example, if the amount of a component, or the value of a process variable such as temperature, pressure, or time, is stated as 50-90, in this specification it means specifically listing values ​​such as 51-89, 52-88… and 69-71 and 70-71, etc. For non-integer values, it may be appropriately considered that a unit is 0.1, 0.01, 0.001, or 0.0001. These are merely some specifically specified examples. In this application, in a similar manner, all possible combinations of numerical values ​​between the listed minimum and maximum values ​​are considered to have been disclosed.

[0080] It should be noted that the embodiments described above are only for explaining the present invention and do not constitute any limitation on the present invention. The present invention has been described with reference to typical embodiments, but it should be understood that the words used therein are descriptive and explanatory terms, not limiting terms. Modifications can be made to the present invention within the scope of the claims, and revisions can be made to the present invention without departing from the scope and spirit of the present invention. Although the present invention described herein relates to specific methods, materials, and embodiments, it does not mean that the present invention is limited to the specific examples disclosed herein; on the contrary, the present invention can be extended to all other methods and applications with the same function.

Claims

1. A thin-layer chromatography method for identifying granules of roasted malt formulation, characterized in that, include: The control sample was prepared using roasted malt slices, and the test sample was prepared using roasted malt granules. After spotting, the developing plate was placed in a developing tank pre-saturated with ammonia water and developed. The developing solvent was n-hexane-ethyl acetate-methanol with a volume ratio of 5:(9-11):

1. The plate was then removed, dried, sprayed with potassium iodide iodine solution, evaporated, and examined under sunlight.

2. The thin-layer method according to claim 1, characterized in that, The methods for preparing test sample spotting using charred malt formulation granules include: After grinding the malt extract granules, add ammonia water and soak in a sealed container. Then add dichloromethane for ultrasonic extraction. After filtration, evaporate the filtrate and dissolve the residue.

3. The thin-layer method according to claim 2, characterized in that, The mass ratio of the roasted malt granules to the volume of the ammonia solution is (2-6) g: 10 ml, and the concentration of the ammonia solution is preferably 25-28%. And / or, the soaking time is 15-30 minutes; And / or, the volume ratio of the ammonia to the dichloromethane is 1:(2-4).

4. The thin-layer method according to claim 2, characterized in that, The solvent for dissolving the filtrate residue is a mixed solution of isopropanol and dichloromethane, preferably with a volume ratio of (1-2):(1-2), more preferably 1:1; And / or, the ultrasonic extraction time is 10-30 min.

5. The thin-layer method according to claim 1, characterized in that, The method for preparing the control spotting sample using roasted malt slices includes: Take roasted malt slices, add water, heat and reflux, centrifuge to collect the supernatant, then concentrate under reduced pressure to dryness, soak the first residue in ammonia water, add dichloromethane and sonicate, filter, evaporate the filtrate, and dissolve the second residue.

6. The thin-layer method according to claim 5, characterized in that, Based on a mass of 10g of the roasted malt slices, the amount of water added is 80-120ml, the volume of ammonia added is 15-25ml, and the volume of dichloromethane added is 20-40ml. Preferably, the concentration of the ammonia is 25-28%. And / or, the heating reflux time is 0.8-1.5 h; And / or, the soaking time is 10-30 minutes; And / or, the duration of the ultrasonic treatment is 10-30 min; And / or, the solvent for dissolving the second residue is a mixed solution of isopropanol and dichloromethane, preferably the volume ratio of isopropanol to dichloromethane in the mixed solution is (1-2):(1-2), more preferably 1:

1.

7. The thin-layer method according to any one of claims 1-6, characterized in that, The sample volume for the test sample is 20-40 µl; And / or, the amount of the control sample spotting is 20-40 µl.

8. The thin-layer method according to any one of claims 1-6, characterized in that, The ambient temperature for thin-layer plate development is 8-23℃; And / or, the ambient humidity for thin-layer plate development is 22-79%; And / or, the thin-layer plate used in the thin-layer method includes a silicone G plate.

9. The thin-layer method according to any one of claims 1-6, characterized in that, On the thin-layer plate of the roasted malt formulation granules, the spot shift values ​​from the spotting point to the development direction of the developing agent are 0.15~0.20, 0.35~0.40, and 0.45~0.50, respectively.

10. The application of the thin-layer chromatography method for identifying roasted malt formulation particles according to any one of claims 1-9 in identifying roasted malt formulation particles, roasted rice sprout formulation particles, and roasted barley sprout formulation particles.

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