Phyllanthus emblica milk processing technology optimization method based on single factor and pharmacodynamic experiment

Abstract

The invention discloses an emblic leafflower fruit milk processing technology optimization method based on a single factor and pharmacodynamic experiment, and belongs to the technical field of traditional Chinese medicine processing, and the method comprises the following steps: screening out optimal technological parameters through the single factor experiment: mixing emblic leafflower fruit and raw milk according to a material-liquid ratio of 1: 15, soaking at 45 + / -5 DEG C for 12 hours, cleaning milk stains, naturally air-drying for 2 hours, and drying at 50 DEG C for 24-48 hours until the water content is less than 6%. A liquid chromatography-tandem high-resolution mass spectrometry technology is adopted for analysis and displaying that the content of gallic acid, ellagic acid and corilagin in a processed product is remarkably reduced. Pharmacodynamic experiments prove that the toxicity of the processed product to WRL68 hepatocytes and 293T renal cells is weakened, the survival rate of the hepatocytes and the renal cells is increased along with reduction of the concentration of the components, and the toxicity reduction effect is achieved. The technological parameters are clear, the quality is stable, and a scientific basis is provided for standardized production and clinical safe medication of emblic leafflower fruit milk processing.

Description

Technical Field

[0001] This invention relates to the field of traditional Chinese medicine processing technology, and in particular to an optimization method for processing Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments. Background Technology

[0002] Phyllanthus emblica, the dried, mature fruit of the Euphorbiaceae plant, is an important medicinal material in the Uyghur medicine system, and its medicinal value has been recorded in numerous historical documents. The Compendium of Materia Medica mentions that Phyllanthus emblica can be "preserved in honey or salt, and steamed with salt for best results." In traditional Uyghur medicine, milk soaking is a unique processing method, the core purpose of which is to reduce the toxicity of the medicinal material, enhance its efficacy, or regulate its properties through the action of milk, so as to better exert its effects such as "drying and cooling, purifying abnormal blood, nourishing the heart, brain, and liver, and enhancing stomach absorption." Modern pharmacological research has further confirmed that Phyllanthus emblica fruit is rich in tannins and phenolic acids, among which gallic acid, ellagic acid, and corilagin are key material bases for its protective effects on the gastrointestinal tract, anti-inflammatory, antibacterial, and antioxidant effects, especially showing clear activity in improving indigestion and other ailments.

[0003] However, although the milk processing technique for Phyllanthus emblica has been used in traditional medicine for a long time, there are many problems that urgently need to be solved in the current technology: First, the processing parameters are vague. Neither historical literature nor modern research has clearly defined key parameters such as soaking temperature, soaking time, material-to-liquid ratio, and type of milk, resulting in significant differences in the quality of processed products from different production batches, making it difficult to achieve standardized production; Second, there is a lack of systematic component analysis methods, making it impossible to accurately quantify the changes in the content of key components before and after processing, and making it difficult to establish scientific quality control standards; Third, pharmacodynamic verification is insufficient. The traditional "toxicity reduction" effect is only based on experience and lacks toxicity assessment data for targets such as liver and kidney cells, which cannot provide direct evidence for safe clinical use.

[0004] Therefore, it is urgent to establish an optimized processing method for Phyllanthus emblica in milk based on scientific experiments, clarify key parameters, and combine component analysis and pharmacodynamic verification to achieve process standardization and quality control, thereby providing a scientific basis for the clinical application of Phyllanthus emblica. Summary of the Invention

[0005] The purpose of this invention is to provide an optimization method for processing Phyllanthus emblica into milk based on single-factor and pharmacodynamic experiments, in order to solve the above-mentioned problems.

[0006] This invention provides an optimization method for processing Phyllanthus emblica into milk based on single-factor and pharmacodynamic experiments, comprising the following steps: S1. Single-factor experiment: Soaking temperature, soaking time, material-to-liquid ratio, and milk type were used as the factors to be investigated, and the contents of gallic acid, ellagic acid, corilagin, and moisture were used as comprehensive evaluation indicators to screen the optimal levels of each factor in the processing. S2. Component analysis: The chemical components of raw and processed Phyllanthus emblica were analyzed by liquid chromatography-tandem high-resolution mass spectrometry, characteristic peaks were identified and the content of target components was determined. S3. Pharmacodynamic experiments: Verify the toxic effects of processed products on liver and kidney cells through cell viability testing experiments; S4. Process Determination: Based on the results of single-factor experiments, component analysis and pharmacodynamic experiments, the optimal processing technology was determined as follows: Mix Phyllanthus emblica with milk according to the material-to-liquid ratio, soak at the soaking temperature and for the soaking time, wash off the milk stains after soaking, and dry until the moisture content is <6%.

[0007] Preferably, in the above-mentioned method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments, the soaking temperature in step S1 is 45±5℃.

[0008] Preferably, in the above-mentioned method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments, the soaking time in step S1 is 12 hours.

[0009] Preferably, in the above-mentioned method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments, the material-to-liquid ratio in step S1 is Phyllanthus emblica: milk = 1:15.

[0010] Preferably, in the above-mentioned method for optimizing the processing technology of Phyllanthus emblica milk based on single-factor and pharmacodynamic experiments, the type of milk mentioned in step S1 is raw milk.

[0011] Preferably, in the above-mentioned method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments, the liquid chromatography-tandem high-resolution mass spectrometry technique described in step S2 uses a Thermo QE plus liquid chromatography-tandem high-resolution mass spectrometer, combined with Compound Discover data processing software, mzcloud online database and locally built mzVault Chinese herbal natural product database for characteristic peak identification. The identification criteria are mass deviation < 5 ppm, conformity to isotope distribution and mzVault best match database matching score > 70 points.

[0012] Preferably, in the above-mentioned optimization method for processing Phyllanthus emblica into milk based on single-factor and pharmacodynamic experiments, the processed product obtained in step S4 contains 0.2%-1.0% gallic acid (C7H6O5) and 0.2% ellagic acid (C7H6O5). 14 The content of H6O8 is 0.05%-0.20%, and the content of Corilagin (C 27 H 22 O 18 The content is 0.2%-0.9%.

[0013] Preferably, in the above-mentioned method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments, the drying in step S4 includes: first air drying for 2 hours, and then drying at a low temperature of 50°C for 24-48 hours.

[0014] Preferably, in the above-mentioned method for optimizing the processing technology of Phyllanthus emblica milk based on single-factor and pharmacodynamic experiments, the cell viability detection experiment in step S3 adopts the CCK8 method, and the detection targets are WRL68 hepatocytes and 293T kidney cells, comparing the effects of raw Phyllanthus emblica, processed Phyllanthus emblica and control products on cell viability.

[0015] Preferably, in the above-mentioned optimization method for processing Phyllanthus emblica into milk based on single-factor and pharmacodynamic experiments, the characteristics of the processed product obtained in step S4 are as follows: the surface is brownish-yellow, almost tasteless after drying, lighter in weight, and brittle and easily crushed; when not dried, it has a slight milky taste, a clear sour and astringent taste, and is soft and full-bodied.

[0016] Therefore, this invention provides the above-mentioned optimization method for processing Phyllanthus emblica with milk based on single-factor and pharmacodynamic experiments. The optimal soaking temperature, soaking time, material-to-liquid ratio, and type of milk were screened through single-factor experiments, solving the problems of vague parameters and poor repeatability in traditional processing methods, and providing an operable standardized process for large-scale production. Liquid chromatography-tandem high-resolution mass spectrometry was used to analyze the components before and after processing, clarifying the content range of gallic acid, ellagic acid, and corilagin in the processed product. Combined with moisture control, precise control of the quality of the processed product was achieved. Pharmacodynamic experiments confirmed that the content of tannins in Phyllanthus emblica was significantly reduced after processing, and the survival rate of WRL68 hepatocytes and 293T kidney cells increased with decreasing component concentration, clearly verifying the detoxification effect of milk processing and improving drug safety. After processing, Phyllanthus emblica exhibits uniform properties, a brownish-yellow surface, and a brittle, easily crushed texture after drying. When undried, it has a slightly milky and distinct sour and astringent taste. The yield, moisture content, and component content are stable, meeting the quality consistency requirements for large-scale production.

[0017] The technical solution of the present invention will be further described in detail below with reference to the accompanying drawings and embodiments. Attached Figure Description

[0018] Figure 1 This is a graph showing the trend of the percentage content of corilagin, gallic acid and ellagic acid as a function of soaking time in Example 1; Figure 2 This is a graph showing the trend of processing yield as a function of soaking time in Example 1; Figure 3 This is a graph showing the trend of gallic acid percentage content as a function of soaking temperature in Example 1; Figure 4 This is a graph showing the trend of processing yield as a function of soaking temperature in Example 1; Figure 5This is a graph showing the variation of gallic acid, ellagic acid, and corilagin content with material ratio in Example 1. Figure 6 This is a graph showing the processing yield corresponding to different material ratios in Example 1; Figure 7 This is a graph showing the trend of gallic acid, ellagic acid, and corilagin content with different milk brands in Example 1. Figure 8 The image shows the TIC (total ion current) chromatogram of the Phyllanthus emblica sample in positive ion mode in Example 2. Figure 9 The image shows the TIC diagram of the Phyllanthus emblica sample under negative ion mode in Example 2. Figure 10 The image shows the TIC diagram of the processed Phyllanthus emblica sample in positive ion mode in Example 2. Figure 11 The image shows the TIC diagram of the processed Phyllanthus emblica sample under negative ion mode in Example 2. Figure 12 This is the TIC image of the blank sample in positive ion mode in Example 2; Figure 13 The TIC image is shown for the blank sample in negative ion mode in Example 2. Figure 14 The effect of raw and processed Phyllanthus emblica on hepatocyte survival rate in Example 3; Figure 15 The effect of the control in Example 3 on hepatocyte survival rate; Figure 16 The effect of raw and processed Phyllanthus emblica on renal cell viability in Example 3; Figure 17 This is the effect of the control substance in Example 3 on the survival rate of kidney cells. Detailed Implementation

[0019] To better understand the above technical solutions, a detailed description of the solutions will be provided below in conjunction with the accompanying drawings and specific embodiments. Obviously, the described embodiments are merely some, not all, of the embodiments of the present invention. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the scope of protection of the present invention.

[0020] The terminology used in the embodiments of this invention is for the purpose of describing particular embodiments only and is not intended to limit the invention. The singular forms “a,” “the,” and “the” as used in the embodiments of this invention and the appended claims are also intended to include the plural forms, and “multiple” generally includes at least two unless the context clearly indicates otherwise.

[0021] It should also be noted that the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that an article or device that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such an article or device. Without further limitation, an element defined by the phrase "comprising one..." does not exclude the presence of other identical elements in the article or device that includes said element.

[0022] This invention provides an optimization method for processing Phyllanthus emblica into milk based on single-factor and pharmacodynamic experiments, comprising the following steps: S1. Single-factor experiment: Soaking temperature, soaking time, material-to-liquid ratio, and milk type were used as the factors to be investigated, and the contents of gallic acid, ellagic acid, corilagin, and moisture were used as comprehensive evaluation indicators to screen the optimal levels of each factor in the processing. S2. Component analysis: The chemical components of raw and processed Phyllanthus emblica were analyzed by liquid chromatography-tandem high-resolution mass spectrometry, characteristic peaks were identified and the content of target components was determined. S3. Pharmacodynamic experiments: Verify the toxic effects of processed products on liver and kidney cells through cell viability testing experiments; S4. Process Determination: Based on the results of single-factor experiments, component analysis and pharmacodynamic experiments, the optimal processing technology was determined as follows: Mix Phyllanthus emblica with milk according to the material-to-liquid ratio, soak at the soaking temperature and for the soaking time, wash off the milk stains after soaking, and dry until the moisture content is <6%.

[0023] Preferably, in the above-mentioned method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments, the soaking temperature in step S1 is 45±5℃.

[0024] Preferably, in the above-mentioned method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments, the soaking time in step S1 is 12 hours.

[0025] Preferably, in the above-mentioned method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments, the material-to-liquid ratio in step S1 is Phyllanthus emblica: milk = 1:15.

[0026] Preferably, in the above-mentioned method for optimizing the processing technology of Phyllanthus emblica milk based on single-factor and pharmacodynamic experiments, the type of milk mentioned in step S1 is raw milk.

[0027] Preferably, in the above-mentioned method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments, the liquid chromatography-tandem high-resolution mass spectrometry technique described in step S2 uses a Thermo QE plus liquid chromatography-tandem high-resolution mass spectrometer, combined with Compound Discover data processing software, mzcloud online database and locally built mzVault Chinese herbal natural product database for characteristic peak identification. The identification criteria are mass deviation < 5 ppm, conformity to isotope distribution and mzVault best match database matching score > 70 points.

[0028] Preferably, in the above-mentioned optimization method for processing Phyllanthus emblica into milk based on single-factor and pharmacodynamic experiments, the processed product obtained in step S4 contains 0.2%-1.0% gallic acid (C7H6O5) and 0.2% ellagic acid (C7H6O5). 14 The content of H6O8 is 0.05%-0.20%, and the content of Corilagin (C 27 H 22 O 18 The content is 0.2%-0.9%.

[0029] Preferably, in the above-mentioned method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments, the drying in step S4 includes: first air drying for 2 hours, and then drying at a low temperature of 50°C for 24-48 hours.

[0030] Preferably, in the above-mentioned method for optimizing the processing technology of Phyllanthus emblica milk based on single-factor and pharmacodynamic experiments, the cell viability detection experiment in step S3 adopts the CCK8 method, and the detection targets are WRL68 hepatocytes and 293T kidney cells, comparing the effects of raw Phyllanthus emblica, processed Phyllanthus emblica and control products on cell viability.

[0031] Preferably, in the above-mentioned optimization method for processing Phyllanthus emblica into milk based on single-factor and pharmacodynamic experiments, the characteristics of the processed product obtained in step S4 are as follows: the surface is brownish-yellow, almost tasteless after drying, lighter in weight, and brittle and easily crushed; when not dried, it has a slight milky taste, a clear sour and astringent taste, and is soft and full-bodied.

[0032] To more clearly and in detail introduce the optimization method for processing Phyllanthus emblica into milk based on single-factor and pharmacodynamic experiments provided in the embodiments of the present invention, the following description will be based on specific embodiments.

[0033] Example 1 Single-factor experiments were conducted to screen the optimal processing parameters, and the effects of soaking temperature, soaking time, material-to-liquid ratio, and type of milk on the quality of the processed products were investigated to determine the optimal level of each factor.

[0034] Soaking time study: Weigh the Phyllanthus emblica herb into a beaker, add raw milk (material-to-liquid ratio 1:20), shake well, seal with plastic wrap, poke small holes to allow air to pass through, and place the conical flask in a 45℃ water bath. The water level in the water bath should be higher than the liquid level in the conical flask. Soaking times were 0h, 4h, 8h, 12h, 16h, 20h, and 24h. After the time was up, the flask was removed, rinsed promptly, and excess milk stains were washed away. The flask was then spread out on newspaper and allowed to air dry naturally. Air drying took about 2 days in summer and 3-5 days in winter. The flask was weighed, the processing yield was calculated, and the contents of gallic acid, corilagin, and ellagic acid in the processed product were determined.

[0035] like Figure 1-2 As shown, the contents of gallic acid, ellagic acid, and corilagin in processed Phyllanthus emblica generally decreased with prolonged soaking time. The highest gallic acid content was observed after 4 hours of soaking, and the lowest after 12 hours; the highest ellagic acid content was observed after 0 hours of soaking, and the lowest after 24 hours, with similar contents at 12 and 16 hours; the highest corilagin content was observed after 0 hours of soaking, and the lowest after 24 hours. The overall yield trend was a decrease with prolonged soaking time. Therefore, considering the above three components and yield as indicators, the optimal soaking time for processing Phyllanthus emblica is 12 hours.

[0036] Soaking temperature investigation: Weigh the Phyllanthus emblica material into a beaker, add raw milk (material-to-liquid ratio 1:10), shake well, seal with plastic wrap, poke small holes for ventilation, place the conical flask in a water bath, ensuring the water level in the water bath is higher than the liquid level in the conical flask, soak for 12 hours at temperatures of 25℃, 35℃, 45℃, and 55℃. After the time is up, remove the flask, rinse it promptly to remove excess milk stains, spread it out on newspaper, and air dry naturally (about 2 days in summer, 3-5 days in winter). Weigh the flask, calculate the processing yield, and determine the gallic acid content in the processed product.

[0037] like Figure 3-4 As shown, the gallic acid content in processed Phyllanthus emblica decreased continuously before the soaking temperature reached 45℃. At 45℃, the gallic acid content reached its lowest point, less than half that of the raw product. The gallic acid content significantly increased after the temperature reached 55℃. The processing yield decreased with increasing soaking temperature, remaining above 50% before 45℃. Using the changes in gallic acid content and yield as indicators, subsequent single-factor experiments comparing material ratios and milk were conducted, with a soaking temperature of 45℃ and a soaking time of 12 hours.

[0038] Material-liquid ratio analysis: Weigh the Phyllanthus emblica medicinal material into a beaker, add raw milk, and mix in material-liquid ratios of 1:5, 1:10, 1:15, and 1:20. Shake well, seal with plastic wrap, and poke small holes to allow air to pass through. Place the conical flask in a 45°C water bath, ensuring the water level in the water bath is higher than the liquid level in the conical flask. Soak for 12 hours. After the time is up, remove the flasks, rinse them promptly, remove excess milk stains, spread them out on newspaper, and air dry naturally. This takes about 2 days in summer and 3-5 days in winter. Weigh the flasks, calculate the processing yield, and determine the content of gallic acid, ellagic acid, and corilagin in the processed products.

[0039] like Figure 5-6 As shown, the contents of gallic acid, ellagic acid, and corilagin in processed Phyllanthus emblica decreased with increasing material ratio, reaching their lowest levels at a material ratio of 1:15, and increasing further at 1:20. In the material ratio analysis, the yield of processed Phyllanthus emblica was consistently >50%, showing no significant correlation with the material ratio. In conclusion, using low levels of gallic acid, ellagic acid, and corilagin as indicators, the optimal material ratio for processed Phyllanthus emblica is 1:15.

[0040] Milk type investigation: Raw amla was taken and raw milk and different brands of sterilized / pasteurized milk were added at a material-to-liquid ratio of 1:15. The mixture was soaked in a 45℃ water bath for 12 hours to investigate the effect of milk type on the content of gallic acid, corilagin, and ellagic acid in amla.

[0041] like Figure 7 As shown, the changes in gallic acid, ellagic acid, and corilagin content in processed amla products were not significantly different from those of milk types, but were related to soaking time, soaking temperature, and material-to-liquid ratio. To facilitate later production and save costs, raw milk was ultimately selected as the processing auxiliary material.

[0042] In summary, based on single-factor analysis and taking the low content of gallic acid, ellagic acid, and corilagin as indicators, the optimal processing parameters for Phyllanthus emblica are as follows: use raw milk with 15 times the weight of the medicinal material, soak the Phyllanthus emblica in a beaker, place the beaker in a 45℃ water bath for 12 hours, remove it, rinse off the milk stains, and dry it until the moisture content is <6%.

[0043] Example 2 Liquid chromatography-tandem high-resolution mass spectrometry was used to analyze the components of raw and processed Phyllanthus emblica and to verify the changes in components before and after processing.

[0044] Preparation of raw Phyllanthus emblica test sample: Accurately weigh 1.0 g of raw Phyllanthus emblica and place it in a 50 mL centrifuge tube. Add 20 mL of 70% methanol solution and extract by sonication for 30 minutes. After extraction, centrifuge at 5000 r / min for 10 minutes at 4℃ and collect the supernatant. Add 20 mL of 70% methanol to the residue and extract once more. Combine the two supernatants and evaporate by rotary evaporation until nearly dry. Make up to 10 mL with 70% methanol and filter through a 0.22 μm organic filter membrane to obtain the raw product test sample solution. Store at 4℃ for later use.

[0045] Preparation of processed Phyllanthus emblica sample: The processed product was prepared according to the optimal process determined in Example 1. Phyllanthus emblica was mixed with raw milk at a ratio of 1:15, soaked in a water bath at 45°C for 12 hours, rinsed with purified water until no milk residue remained, air-dried naturally for 2 hours, and then dried at 50°C for 24-48 hours until the moisture content was <6%. After pulverization, it was passed through an 80-mesh sieve. Subsequent extraction, centrifugation, volume adjustment, and filtration were performed in the same manner as the preparation of the raw product sample, thus obtaining the processed product sample solution.

[0046] Blank sample preparation: Take the same volume of 70% methanol solution as the test sample extract, and process it according to the above steps of "extraction, centrifugation, merging, volume adjustment, and membrane filtration". Do not add raw or processed Phyllanthus emblica to obtain a blank sample solution, which is used to eliminate interference from solvent and extraction process.

[0047] Mass spectrometry data acquisition was performed using Q Exactive Orbitrap high-resolution mass spectrometry, with FullMS-ddMS detection mode. 2 Positive and negative ion modes were scanned separately, with a scan range of m / z 100-1200, MS 1 Resolution set to 70000, MS 2 Resolution was set to 17500, ion source voltage to 3.2 kV, capillary ion transfer tube temperature to 320°C, auxiliary gas heater temperature to 350°C, sheath gas flow rate to 40 L / min, auxiliary gas flow rate to 15 L / min, AGCT target to 1e6, TopN to 5, and MS triggering. 2 The collision energy for the scan was measured using a stepped fragmentation voltage NCE, set to 30, 40, and 50.

[0048] Characteristic peaks were extracted from raw RAW mass spectrometry data using Compound Discover 3.2 software. Mass deviations for characteristic peak elemental matching, molecular formula prediction, and isotope distribution matching were all set to within 5 ppm. Characteristic peak identification was performed using the mzcloud online database and the locally built mzVault database of traditional Chinese medicine and natural products. Positive results were selected based on a mass deviation <5 ppm, conformity to isotope distribution, and a matching score >70 in the mzVault best match database. Results included the compound's English and Chinese names, molecular formula, structural formula, peak area, TIC chromatogram, and MS data. 1 and MS 2 Mass spectrum.

[0049] like Figure 8-13 As shown in Tables 1-2, 40 components, including ellagic acid, gallic acid, and corilagin, were identified in the raw product; the peak areas of the above three components were significantly reduced in the processed product, and new components such as betaine and trigonelline hydrochloride were added; the blank sample confirmed the absence of interfering peaks, ensuring the accuracy of the analysis.

[0050] Table 1. Identification of medicinal components of Phyllanthus emblica

[0051] Table 2. Identification of components in processed Phyllanthus emblica products

[0052] Example 3 Pharmacodynamic experiments were conducted to verify the toxicity reduction effect and to investigate the toxic effects of processed Phyllanthus emblica on hepatocytes and kidney cells. The CCK8 assay was used to detect the effects of raw Phyllanthus emblica, processed Phyllanthus emblica, and the control compounds geranylin, ellagic acid, L-hydroxyproline, and taraxacin on the survival rate of WRL68 hepatocytes and 293T kidney cells. The preparation methods for raw and processed Phyllanthus emblica were consistent with those in Example 2. Raw and processed Phyllanthus emblica, along with the control compounds geranylin, ellagic acid, L-hydroxyproline, and taraxacin, were applied to WRL68 hepatocytes and 293T kidney cells. Based on a highly sensitive cell proliferation and toxicity detection method using water-soluble tetrazolium salts, the reducing ability of mitochondrial dehydrogenases to the WST-8 reagent was detected, generating an orange-yellow formazan product. The concentration of this product was directly proportional to the number of viable cells, yielding the following results.

[0053] like Figure 14-17 As shown, the cell survival rate of the processed product group was significantly higher than that of the raw product group, and the cell survival rate increased in a dose-dependent manner as the concentration of components such as gallic acid decreased, confirming that the liver and kidney toxicity was reduced after processing.

[0054] Example 4 Ten batches of processed products were subjected to quality verification to confirm the stability and repeatability of the optimal process. Following the optimal process, ten batches of processed products were prepared by soaking in raw milk at a ratio of 1:15 at 45℃ for 12 hours, air-drying for 2 hours, and then drying at 50℃ for 24-48 hours until the moisture content was <6%. The yield, properties, and components such as gallic acid, ellagic acid, and corilagin were compared, and the results are shown in Table 3.

[0055] Table 3 Quality verification results of 10 batches of processed products

[0056] It is evident that the quality of the processed Phyllanthus emblica products was stable. The moisture content decreased significantly after processing. Ten batches of processed Phyllanthus emblica products contained gallic acid (0.2%-1.0%), ellagic acid (0.05%-0.20%), and corilagin (0.2%-0.9%). The processed Phyllanthus emblica products had a brownish-yellow surface, a slightly milky aroma, a more pronounced sour and astringent taste, and were soft and plump. The dried processed products were almost tasteless, significantly lighter, and brittle, easily crushed. Compared to the raw product, the processed products showed a significant reduction in gallic acid, ellagic acid, and corilagin content, resulting in reduced hepatotoxicity and nephrotoxicity.

[0057] Therefore, this invention provides the above-mentioned optimization method for processing Phyllanthus emblica with milk based on single-factor and pharmacodynamic experiments. The optimal soaking temperature, soaking time, material-to-liquid ratio, and type of milk were screened through single-factor experiments, solving the problems of vague parameters and poor repeatability in traditional processing methods, and providing an operable standardized process for large-scale production. Liquid chromatography-tandem high-resolution mass spectrometry was used to analyze the components before and after processing, clarifying the content range of gallic acid, ellagic acid, and corilagin in the processed product. Combined with moisture control, precise control of the quality of the processed product was achieved. Pharmacodynamic experiments confirmed that the content of tannins in Phyllanthus emblica was significantly reduced after processing, and the survival rate of WRL68 hepatocytes and 293T kidney cells increased with decreasing component concentration, clearly verifying the detoxification effect of milk processing and improving drug safety. After processing, Phyllanthus emblica exhibits uniform properties, a brownish-yellow surface, and a brittle, easily crushed texture after drying. When undried, it has a slightly milky and distinct sour and astringent taste. The yield, moisture content, and component content are stable, meeting the quality consistency requirements for large-scale production.

[0058] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and not to limit them. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can still be made to the technical solutions of the present invention, and these modifications or equivalent substitutions cannot cause the modified technical solutions to deviate from the spirit and scope of the technical solutions of the present invention.

Claims

1. An optimization method for processing Phyllanthus emblica into milk based on single-factor and pharmacodynamic experiments, characterized in that, Includes the following steps: S1. Single-factor experiment: Soaking temperature, soaking time, material-to-liquid ratio, and milk type were used as the factors to be investigated, and the contents of gallic acid, ellagic acid, corilagin, and moisture were used as comprehensive evaluation indicators to screen the optimal levels of each factor in the processing. S2. Component analysis: The chemical components of raw and processed Phyllanthus emblica were analyzed by liquid chromatography-tandem high-resolution mass spectrometry, characteristic peaks were identified and the content of target components was determined. S3. Pharmacodynamic experiments: Verify the toxic effects of processed products on liver and kidney cells through cell viability testing experiments; S4. Process Determination: Based on the results of single-factor experiments, component analysis and pharmacodynamic experiments, the optimal processing technology was determined as follows: Mix Phyllanthus emblica with milk according to the material-to-liquid ratio, soak at the soaking temperature and for the soaking time, wash off the milk stains after soaking, and dry until the moisture content is <6%.

2. The method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments according to claim 1, characterized in that, The soaking temperature in step S1 is 45±5℃.

3. The method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments according to claim 1, characterized in that, The soaking time in step S1 is 12 hours.

4. The method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments according to claim 1, characterized in that, The material-to-liquid ratio mentioned in step S1 is Phyllanthus emblica: milk = 1:

15.

5. The method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments according to claim 1, characterized in that, The type of milk mentioned in step S1 is raw milk.

6. The method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments according to claim 1, characterized in that, The liquid chromatography-tandem high-resolution mass spectrometry technique described in step S2 uses a Thermo QE plus liquid chromatography-tandem high-resolution mass spectrometer, combined with Compound Discover data processing software, the mzcloud online database, and the locally built mzVault database of traditional Chinese medicine natural products for characteristic peak identification. The identification criteria are mass deviation < 5 ppm, conformity to isotope distribution, and a matching score of > 70 points in the mzVault best match database.

7. The method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments according to claim 1, characterized in that, The processed product obtained in step S4 contains gallic acid of 0.2%-1.0%, ellagic acid of 0.05%-0.20%, and corilagin of 0.2%-0.9%.

8. The method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments according to claim 1, characterized in that, The drying process described in step S4 includes: first air drying for 2 hours, and then drying at a low temperature of 50°C for 24-48 hours.

9. The method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments according to claim 1, characterized in that, The cell viability assay described in step S3 uses the CCK8 method, and the assay subjects are WRL68 hepatocytes and 293T kidney cells, comparing the effects of raw Phyllanthus emblica, processed Phyllanthus emblica, and control on cell viability.

10. The method for optimizing the processing technology of Phyllanthus emblica in milk based on single-factor and pharmacodynamic experiments according to claim 1, characterized in that, The processed product obtained in step S4 has the following characteristics: the surface is brownish-yellow, it is almost tasteless after drying, its weight is reduced, and its texture is crisp and easy to crush; when not dried, it has a slightly milky taste, a clear sour and astringent taste, and is soft and full-bodied.

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