New application of rhizopus deelemar

Abstract

The invention relates to application of Rhizopus deelemar in a traditional Chinese medicine immature bitter orange decoction piece processing method, and belongs to the field of medicine processing. According to the processing method, fermentation strains are defined, interference of infectious microbes in air in the fermentation process is avoided, and the safety of the fructus aurantii immaturus decoction pieces is ensured. Meanwhile, key process parameters for fermenting and steaming the immature bitter oranges are optimized and determined, and the quality stability of the immature bitter orange decoction pieces is ensured. Compared with the current naturally fermented immature bitter orange decoction pieces, the immature bitter orange decoction pieces prepared by the method have the advantages that the content of synephrine and flavonoid active ingredients is obviously increased, harmful ingredients such as aflatoxin are not detected, the quality is better, and the method is suitable for large-scale production of the immature bitter orange decoction pieces.

Description

[0001] This application is a divisional application of the application with the application number 2023117002075, the application date of December 12, 2023, and the invention title "Application of Rhizopus delemar in the processing method of traditional Chinese medicine Fructus Aurantii Immaturus slices". Technical Field

[0002] The present invention relates to the application of Rhizopus delemar in the processing method of traditional Chinese medicine Fructus Aurantii Immaturus slices, belonging to the field of drug processing. Background Art

[0003] Fructus Aurantii Immaturus is the dried immature fruit of Citrus aurantium L. of the Rutaceae family and its cultivated varieties or Citrus sinensis Osbeck. The fruits that have fallen are collected in May or June, the impurities are removed, and they are cut transversely into two halves from the middle, then dried in the sun or by low-temperature drying. The smaller ones are directly dried in the sun or by low-temperature drying.

[0004] Currently, the main processing methods of Fructus Aurantii Immaturus are as follows: 1. Remove impurities, wash, moisten thoroughly, cut into thin slices, and dry. 2. Stir-fry Fructus Aurantii Immaturus with bran: Take Fructus Aurantii Immaturus slices and stir-fry them according to the bran-stir-frying method (a method of stir-frying the purified or cut drug with wheat bran) until the color becomes darker. Yu Xiaoli, Research on the processing technology and quality standard of Fructus Aurantii Immaturus, Master's thesis of Liaoning University of Traditional Chinese Medicine in 2010, determined the specific cutting process of Fructus Aurantii Immaturus slices, optimized the best processing technology of stir-fried Fructus Aurantii Immaturus, compared the effects on gastrointestinal function before and after processing Fructus Aurantii Immaturus, and formulated the quality standard of stir-fried Fructus Aurantii Immaturus. Huang Yanqiu, et al., Historical evolution and research progress of the processing technology of Fructus Aurantii Immaturus, Guangdong Chemical Industry, No. 8, 2023, Vol. 50, Total No. 490, summarized the processing methods in past dynasties, changes in chemical components before and after processing, pharmacological effects, and processing technology. Due to the complex chemical components of Fructus Aurantii Immaturus, there are certain differences in chemical components before and after processing. Most of the research is only limited to laboratory research and has not been scaled up, so it cannot fully reflect the actual situation of large-scale production. Wang Cheng, et al., Research progress on the processing of Fructus Aurantii Immaturus, Asia-Pacific Traditional Medicine, June 2023, Vol. 19, No. 6, disclosed more than 20 processing methods of Fructus Aurantii Immaturus in past dynasties, including purification, cutting, stir-frying (stir-frying yellow, stir-frying to charcoal), stir-frying with solid adjuvants (stir-frying with bran, stir-frying with earth, stir-frying with flour), roasting (roasting with wine, roasting with vinegar, roasting with ginger, roasting with honey), burning, steaming, baking, boiling, soaking in rice water, making into vinegar paste, etc.; modern pharmacopoeias and local processing specifications basically follow the traditional processing methods of Fructus Aurantii Immaturus, mainly including purification, cutting, and stir-frying with bran. The research on the processing technology of Fructus Aurantii Immaturus mainly focuses on stir-frying with bran, softening and cutting, and roasting with water. After processing Fructus Aurantii Immaturus, the content of volatile oil decreases, the types of components also change, and the contents of components such as hesperidin, naringin, and synephrine also decrease.

[0005] There are reports in the literature on the processing of Fructus Aurantii Immaturus by fermentation. For example, Xia Quan, etc., explored the characteristics of Lingnan processing from the perspective of Lingnan medicine. Chinese Traditional and Herbal Drugs, Vol. 40, No. 6, June 2017, reported that the processing method of Fructus Aurantii Immaturus is steaming after fermentation, aiming to alleviate the pungent, dry, and drastic medicinal properties. The fermentation and steaming process of Fructus Aurantii Immaturus slices is a characteristic processing technology in the Lingnan region. Its standard process flow is included in the item of Fructus Aurantii Immaturus in the Guangdong Provincial Chinese Medicine Processing Specification (1984 Edition). The processing method is stipulated as follows: Fructus Aurantii Immaturus, remove impurities, wash, soak until 70% permeated, take out, wait for fermentation for 3 - 4 days, wash, steam for 4 - 6 hours, close for 1 night, until the inside is dark purple, take out, slice, and dry. According to the standard record, this process is still relatively rough, and the process details such as fermentation and steaming are not clear. The processing time, temperature, and degree are mostly determined by the experience of processing workers. There is a lack of standardized and normalized processes during processing, and the quality lacks stability and controllability. At the same time, due to the existence of many harmful bacteria in the air, during the actual fermentation process of Fructus Aurantii Immaturus, it is easy to be accompanied by the growth of miscellaneous bacteria, and a large amount of harmful toxins (such as aflatoxin) will be produced after the growth of miscellaneous bacteria, affecting the safety of the clinical application of fermented and steamed Fructus Aurantii Immaturus slices. All these are not conducive to the promotion and large-scale processing of Lingnan characteristic Fructus Aurantii Immaturus slices. Summary of the Invention

[0006] Aiming at the problems existing in the fermentation and steaming process of Fructus Aurantii Immaturus slices, the present invention preferably selects a Rhizopus delemar strain suitable for the fermentation of Fructus Aurantii Immaturus slices, and determines the preparation process of fermenting and steaming Fructus Aurantii Immaturus slices using the Rhizopus delemar strain through process parameter screening experiments. It solves the deficiencies of the existing processing specifications, is conducive to ensuring the quality stability and safety of Lingnan characteristic Fructus Aurantii Immaturus slices, and is conducive to the large-scale production of Fructus Aurantii Immaturus slices.

[0007] The present invention provides the application of Rhizopus delemar in the processing method of Chinese medicine Fructus Aurantii Immaturus slices. The registration number of the Rhizopus delemar in the China Center for Industrial Culture Collection of Microorganisms is CICC40308.

[0008] The present invention also provides a processing method of Fructus Aurantii Immaturus, which is fermented and processed using Rhizopus delemar CICC40308, including the following steps:

[0009] a. Take the sterilized Fructus Aurantii Immaturus medicinal materials and moisten them thoroughly with water;

[0010] b. Take Rhizopus delemar, after activation treatment, prepare it into a strain activation solution, and the amount of bacteria in the activation solution is (2.0 ± 2.0×10%)×10 8 CFU / mL, dilute with water, and evenly spray it into the medicinal materials;

[0011] c. Fermentation: temperature 20 - 30°C, relative humidity 70 - 90%, fermentation time 3 - 5d;

[0012] d. Steaming: Steam the medicinal materials fermented in step c in a water bath for 4 - 6 hours;

[0013] e. Moistening:

[0014] f. Slicing and drying for 3.5 - 8 h.

[0015] Further preferably, the sterilization method in step a is ultraviolet sterilization with a sterilization time of 30 min; the penetration time is 5 h; the water addition amount is 60% w / w of the weight of Fructus Aurantii Immaturus; the dosages of the strain activation solution, water and Fructus Aurantii Immaturus in step b are: 10 ml of the strain activation solution and 40 ml of water per kg of Fructus Aurantii Immaturus; the fermentation conditions in step c are a temperature of 25°C and a relative humidity of 90% with a fermentation time of 4 d; the penetration time in step e is 5 h; the drying temperature in step f is: 60°C and the drying time is 4 h.

[0016] The present invention also provides Fructus Aurantii Immaturus slices prepared by the processing method of Fructus Aurantii Immaturus, wherein the content of synephrine (mg / g) is not less than 3.0 mg / g; the contents of narirutin, naringin, hesperidin, neohesperidin (mg / g) are not less than 160 mg / g.

[0017] Further preferably, the content of synephrine (mg / g) is not less than 4.0 mg / g.

[0018] The beneficial effects of the present invention are:

[0019] The processing method of the present invention defines the fermentation strain, avoids the interference of miscellaneous bacteria in the air during the fermentation process, and ensures the safety of Fructus Aurantii Immaturus slices. At the same time, the key process parameters of fermenting and steaming Fructus Aurantii Immaturus are optimized and determined, ensuring the stability of the quality of Fructus Aurantii Immaturus slices. Compared with the currently naturally fermented Fructus Aurantii Immaturus slices prepared by the method of the present invention, the contents of effective components such as synephrine and flavonoids are significantly increased, and harmful components such as aflatoxin are not detected, with better quality and being suitable for large-scale production of Fructus Aurantii Immaturus slices. Specific embodiments

[0020] Example 1 Preparation process flow of fermented Fructus Aurantii Immaturus slices

[0021] Take the Fructus Aurantii Immaturus herbs, classify them according to size, wash them with sterile purified water, sterilize them under an ultraviolet lamp for 30 min, mix them evenly with 0.6 times the amount of sterile purified water, cover them and moisten them for 5 h until there is no white core inside, and place them in a disinfected and clean incubator. Take Rhizopus delemar (strain number CICC40308) and activate it using Sabouraud liquid medium (10 g of peptone, 40 g of glucose, 0.1 g of chloramphenicol, made up to 1000 ml with distilled water). The activation conditions are a temperature of 25 °C and shaking culture for 48 h. The amount of bacteria after activation is (2.0 ± 2.0×10%)×10 8 CFU / mL. Take 10 ml of the above-mentioned activated bacterial solution, add 40 ml of distilled water, shake well, spray it into 1 kg of clean Fructus Aurantii Immaturus herbs, mix evenly, pad and cover with a wet gauze, and cover well. Place it in an incubator and culture it at a temperature of 25 °C and a relative humidity of 90% for 4 d. Take it out, wash the fermented Fructus Aurantii Immaturus clean, steam it in a steamer over water for 4 - 6 h, steam it overnight (12 h), take it out and slice it, and dry it at 60 °C.

[0022] The Rhizopus delemar CICC40308 used in the present invention, with the Latin name Rhizopus delemar, is a strain of the China Center of Industrial Culture Collection (CICC, located at Building 6, No. 24 Courtyard, Jiuxianqiao Middle Road, Chaoyang District, Beijing). The Rhizopus delemar strain CICC40308 described in the present invention could be purchased through domestic and foreign commercial channels before the filing date of the present invention application, or the strain could be purchased from the China Center of Industrial Culture Collection. CICC has a dedicated website at the address: http: / / www.china - cicc.org, and the public can directly place an order for the bacterial strain online, and its website address is

[0023] http: / / m.china-cicc.org / search / ?classtype=0&keyword=40308 。

[0024] Example 2 Screening Experiment on Strains for Preparing Fermented Fructus Aurantii Immaturus Slices

[0025] Take the Fructus Aurantii Immaturus herbs, classify them according to size, wash them with sterile purified water, sterilize them under an ultraviolet lamp for 30 minutes, mix them evenly with 1 times the amount of sterile purified water, cover and moisten them thoroughly until there is no white core inside, and place them separately in 3 disinfected and clean incubators. One portion is fermented naturally; one portion takes the activated bacterial solution of Rhizopus delemar strain [the amount of bacteria is (2.0 ± 2.0×10%)×10 8CFU / mL], add 4 times the volume of sterile purified water, mix well, and evenly spray into the medicinal materials at a ratio of 100 g:5 mL; take 1 portion of Mucor powder (spore content 10 billion / g, manufacturer: Jining Xiangyuan Biotechnology Co., Ltd.) accounting for 0.4% of the weight of Fructus Aurantii Immaturus, add 20 times the volume of sterile purified water, and evenly spray into the medicinal materials. The above three samples of Fructus Aurantii Immaturus are covered with wet gauze and the lids are closed. Place them in an incubator and culture at a temperature of 25 °C and a relative humidity of 90% for 4 days. Wash the fermented Fructus Aurantii Immaturus, steam it in a steamer over water for 4 - 6 hours, leave it sealed overnight (12 hours), take it out and slice, and dry at 60 °C.

[0026] Evaluation indicators:

[0027] 1. Appearance evaluation: Describe the properties of the prepared slices according to whether miscellaneous bacteria grow during the fermentation process, and score them in grades. The scoring criteria are shown in Table 1.

[0028] Table 1 Scoring table for the fermentation process of Fructus Aurantii Immaturus

[0029]

[0030] 2. Determination of synephrine content: Determine according to the high performance liquid chromatography method (General Principles 0512) in Part IV of the Chinese Pharmacopoeia 2020 Edition.

[0031] Chromatographic conditions: Chromatographic column Agilent Eclipse plus C 18 (4.6×250 mm, 5 μm); the mobile phase is a mixture of methanol - potassium dihydrogen phosphate solution (dissolve 0.6 g of potassium dihydrogen phosphate, 1.0 g of sodium dodecyl sulfate, and 1 mL of glacial acetic acid in water and dilute to 1000 mL) (50:50); the detection wavelength is 275 nm; the flow rate is 1 mL / min; the column temperature is 30 °C; the number of theoretical plates calculated based on the synephrine peak should be not less than 2000.

[0032] Preparation of reference substance solution: Take an appropriate amount of synephrine reference substance, weigh it accurately. Dissolve it in water to prepare a solution containing 30 μg per 1 mL, and you will get it.

[0033] Preparation method of test sample: Take about 0.5 g of the middle powder of this product, weigh it accurately, place it in a stoppered conical flask, accurately add 25 mL of methanol, ultrasonically treat for 30 minutes, cool and filter, take 5 mL of the filtrate, dilute it with water and make the volume up to 20 mL, shake well, and you will get it.

[0034] Determination method: Accurately pipette 10 μL of the reference substance solution and the test sample solution respectively, inject them into the liquid chromatograph, and determine to get it.

[0035] 3. Determination of flavonoid components (naringin, naringin, hesperidin, neohesperidin) content

[0036] Chromatographic conditions: Chromatographic column: Agilent Eclipse plus C18 (4.6×250 mm, 5 μm); Mobile phase: acetonitrile (A) - 0.1% formic acid aqueous solution (B); Gradient elution conditions: 0 - 10 min, 10% - 19% A; 10 - 25 min, 19% A; 25 - 27 min, 19% - 30% A; 27 - 50 min, 30% - 53% A; 50 - 51 min, 53% - 75% A; 51 - 56 min, 75% A; 56 - 57 min, 75% - 10% A; 57 - 62 min, 10% A. Detection wavelength: 330 nm; Flow rate: 1 mL / min; Column temperature: 35 °C; The number of theoretical plates calculated by synephrine peak should be not less than 2000.

[0037] Preparation of reference substance solution: Appropriate amounts of rutacridone epoxide, naringin, hesperidin, and neohesperidin reference substances were accurately weighed and dissolved in methanol to prepare a mixed reference substance solution with mass concentrations of 100 μg / mL (rutacridone epoxide, naringin) and 200 μg / mL (hesperidin, neohesperidin), respectively.

[0038] Preparation method of test sample: 0.5 g of Fructus Aurantii Immaturus powder was accurately weighed and placed in a stoppered conical flask. 25 ml of methanol was accurately added, and the weight was weighed. It was ultrasonically treated for 30 min, cooled, and then weighed again. The lost weight was made up with methanol, shaken well, filtered, and 5 ml of the subsequent filtrate was accurately measured and transferred to a 25 ml volumetric flask. Methanol was added to the scale and shaken well to obtain the solution.

[0039] Determination method: 10 μl of the reference substance solution and the test sample solution were accurately pipetted and injected into the liquid chromatograph for determination to obtain the results.

[0040] 4. Aflatoxin inspection was carried out according to the method of Determination of Aflatoxin, the second method (liquid chromatography - tandem mass spectrometry) in the method of Determination of Mycotoxins in the fourth part of Chinese Pharmacopoeia 2020 edition.

[0041] The determination results are shown in Table 2.

[0042] Table 2 Screening experiment of strains for preparing fermented Fructus Aurantii Immaturus slices

[0043]

[0044] In addition, according to Chinese Pharmacopoeia 2020 edition, the content of synephrine in Fructus Aurantii Immaturus herbs and slices should be not less than 3 mg / g (0.3%), and there is no specified limit for flavonoid components. It can be seen from Table 2 that the indexes of the slices obtained by fermentation with Rhizopus delemar are better than those of other fermentation methods.

[0045] Example 3 Screening experiment on the preparation process of fermented Fructus Aurantii Immaturus slices

[0046] 1. Research on moistening process

[0047] 1.1 Water absorption test

[0048] Take 50 g of net Fructus Aurantii Immaturus, weigh it, add 3 times the amount of water, let it stand for 4 hours, filter off the water, weigh the Fructus Aurantii Immaturus after water absorption, and calculate the water absorption. The test results are shown in Table 3.

[0049] Table 3 Water absorption test of Fructus Aurantii Immaturus

[0050]

[0051] According to the experimental results, the water absorption of Fructus Aurantii Immaturus is about 60%, so 60% of the weight of Fructus Aurantii Immaturus can be added with water for moistening.

[0052] 1.2 Moistening time test

[0053] Take 50 g of net Fructus Aurantii Immaturus, weigh it, wash it, add 60% of the weight of Fructus Aurantii Immaturus (30 g) of water, observe the moistening degree every 60 minutes until there is no white core inside and it feels not hard when pinched by hand. Determine the appropriate moistening time. The test results are shown in Table 4.

[0054] Table 4 Moistening time test of Fructus Aurantii Immaturus

[0055]

[0056] According to the experimental results, it is determined that the moistening time of Fructus Aurantii Immaturus is about 5 hours until all the water is absorbed.

[0057] 2. Fermentation process research

[0058] 2.1 Fermentation process conditions

[0059] In the preparation process of fermented Fructus Aurantii Immaturus, fermentation temperature, fermentation humidity, and fermentation time are the main influencing factors. Therefore, in the research on the fermentation process of Fructus Aurantii Immaturus, taking appearance, synephrine content, and flavonoid component content (naringin, naringin, hesperidin, neohesperidin) as evaluation indexes, and selecting fermentation temperature, fermentation humidity, and fermentation time as investigation factors, orthogonal research is carried out. The process parameter design is shown in Table 5, and the process condition design table is shown in Table 6.

[0060] Table 5 Factor - level table of Fructus Aurantii Immaturus fermentation process

[0061]

[0062] Table 6 Process condition table of Fructus Aurantii Immaturus fermentation

[0063]

[0064] 2.2 Fermentation process test method

[0065] All processing tools need to be boiled in water (boiling water) for disinfection (15 - 20 minutes). Take 9 portions of Fructus Aurantii Immaturus herbs, classified by size, with each portion being 50 g. Wash them with sterile purified water, sterilize under an ultraviolet lamp for 30 min, add 30 g of sterile purified water and mix evenly. Cover and moisten for 5 h until there is no white core inside. Place them in a disinfected and clean culture vessel, line with a wet gauze pad, and cover well. Then place the culture vessel in an incubator and ferment according to the conditions in Table 5 (check the humidity of the gauze during the fermentation process and spray distilled water appropriately to keep it in a moist state). After fermentation is completed, wash the Fructus Aurantii Immaturus clean, steam it in a steamer over water for 5 hours, and leave it covered overnight (12 hours). Take it out, slice it, and dry at 60 °C.

[0066] 2.3 Evaluation Indexes and Determination Methods for Fermentation Process

[0067] To objectively and comprehensively evaluate the fermentation process of Fructus Aurantii Immaturus, three evaluation indexes are selected, namely appearance, synephrine content, and flavonoid content (naringin, naringin, hesperidin, neohesperidin), and the comprehensive score is calculated according to the weights.

[0068] (1) Appearance: Describe the traits of the cut herbs according to whether there is any contamination by miscellaneous bacteria during the fermentation process, and score them by grade. The determination method is shown in Example 2. It accounts for 40% in the comprehensive score of the fermentation process evaluation of Fructus Aurantii Immaturus.

[0069] (2) Synephrine content: The determination method is shown in Example 2. It accounts for 30% in the comprehensive score of the fermentation process evaluation of Fructus Aurantii Immaturus.

[0070] (3) Flavonoid content (naringin, naringin, hesperidin, neohesperidin): The determination method is shown in Example 2. It accounts for 30% in the comprehensive score of the fermentation process evaluation of Fructus Aurantii Immaturus.

[0071] (4) Comprehensive score: Calculate according to the following formula.

[0072]

[0073] 2.4 Results of Fermentation Process

[0074] The determination results are shown in Table 7.

[0075] Table 7 Determination Results of Fermentation Process of Fructus Aurantii Immaturus

[0076]

[0077] Considering the test results comprehensively, the optimal process parameters for the fermentation of Fructus Aurantii Immaturus are determined as A2B3C2. That is: ferment at a temperature of 25 °C and a relative humidity of 90% for 4 days (check the humidity of the gauze during the fermentation process and spray distilled water appropriately to keep it in a moist state).

[0078] 3. Study on Steaming Time

[0079] Prepare a batch of fermented Fructus Aurantii Immaturus (100 g) according to the determined optimal fermentation process, divide it into 5 portions, each portion being 20 g, and steam them for 0 hour, 3 hours, 4 hours, 5 hours, and 6 hours respectively, then seal them for one night (12 hours) until the interior turns dark purple. Take them out, slice them, and place them in an oven at 60 °C to dry for 4 hours for standby.

[0080] Determination indicators: Describe the decoction pieces by observation; for the synephrine content and flavonoid content (naringin, naringin, hesperidin, neohesperidin), the determination method is the same as in Example 2. The determination results are shown in Table 8.

[0081] Table 8 Determination Results of Fermented Fructus Aurantii Immaturus Decoction Pieces with Different Steaming Times

[0082]

[0083] The results show that the quality indicators of the fermented Fructus Aurantii Immaturus decoction pieces obtained by steaming for 4 - 6 hours are relatively excellent.

[0084] 4. Research on Drying Methods

[0085] Prepare a batch of fermented Fructus Aurantii Immaturus (100 g) according to the determined optimal fermentation process and steaming process, take it out, cut it into thin slices, divide it into 4 portions, and place them in an oven to dry (at 40 °C, 50 °C, 60 °C, 70 °C) respectively.

[0086] Determination indicators:

[0087] (1) Drying time: Examine the drying time required when the moisture content of the medicinal materials is less than 10%.

[0088] (2) Synephrine content: The same as in Example 2.

[0089] (3) Contents of naringin, naringin, hesperidin, and neohesperidin: The same as in Example 2.

[0090] The determination results are shown in Table 9.

[0091] Table 9 Determination Results of Fermented Fructus Aurantii Immaturus Decoction Pieces Obtained at Different Drying Temperatures

[0092]

[0093] The results show that the content of active ingredients in the fermented Fructus Aurantii Immaturus decoction pieces obtained at 40, 50, and 60 °C is relatively high, and the drying time required under the condition of 60 °C is shorter. Considering comprehensively, the drying temperature is determined to be 60 °C.

[0094] Example 4 Verification Experiment on the Preparation Process of Fermented Fructus Aurantii Immaturus Decoction Pieces

[0095] According to the optimal process conditions determined in Example 3, three batches of fermented Fructus Aurantii Immaturus decoction pieces were trial-produced, with 200 g per batch. The appearance of the decoction pieces, synephrine content, flavonoid content (naringin, naringin, hesperidin, neohesperidin), aflatoxin limit and other indicators were measured to comprehensively evaluate the repeatability of the process. The measurement results are shown in Table 10.

[0096] Table 10 Results of the validation experiment for the preparation process of fermented Fructus Aurantii Immaturus decoction pieces

[0097]

[0098] In summary, compared with the currently naturally fermented Fructus Aurantii Immaturus decoction pieces, the content of effective components such as synephrine and flavonoids in the Fructus Aurantii Immaturus decoction pieces prepared by the method of the present invention is significantly increased, and harmful components such as aflatoxin are not detected, which is suitable for the large-scale production of Fructus Aurantii Immaturus decoction pieces.

Claims

1. Use of Rhizopus delemar in the processing method of traditional Chinese medicine Fructus Aurantii Immaturus slices, and the registration number of Rhizopus delemar in the China Center for Industrial Culture Collection of Microorganisms is CICC40308.

2. The use according to claim 1, characterized in that: It is fermented and processed by Rhizopus delemarCICC40308, and includes the following steps: a. Take the sterilized Zhishi medicinal material and moisten it with water; b. Take Rhizopus delemar, after activation treatment, prepare it into a bacterial strain activation solution, and the amount of bacteria in the activation solution is (2.0 ± 2.0×10%)×10 8 CFU / mL, dilute it with water, and evenly spray it into the medicinal materials; c. Fermentation: temperature 20-30°C, relative humidity 70-90%, fermentation time 3-5 days; d. Steaming: Steam the fermented medicinal materials in step c for 4-6 hours; e.Muffled: f. Slice and dry for 3.5-8 hours.

3. The use according to claim 2, characterized in that: The sterilization method described in step a is ultraviolet sterilization, and the sterilization time is 30 minutes; the soaking time is 5 hours; and the amount of water added is 60% w / w of the weight of the Citrus aurantium.

4. The use according to claim 2, characterized in that: The dosage of the bacterial strain activation liquid, water and Citrus aurantium in step b is: 10 ml of bacterial strain activation liquid and 40 ml of water per kg of Citrus aurantium.

5. The use according to claim 2, characterized in that: The fermentation conditions in step c are a temperature of 25° C., a relative humidity of 90%, and a fermentation time of 4 days.

6. The use according to claim 2, characterized in that: The soaking time described in step e is 5 hours.

7. The use according to claim 2, characterized in that: The drying temperature in step f is 60°C and the drying time is 4 hours.

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