A biological processing method for Huajuhong

Abstract

The present invention relates to a biological preparation method of tangerine peel, which can significantly improve the content of active ingredients such as total flavonoids, naringin, wild sumac glycoside, melittin, prunin and new eriocin of tangerine peel by using various microorganisms including Lactobacillus plantarum, Leuconostoc mesenteroides, Bacillus subtilis and Rhizopus sinensis to prepare tangerine peel, and enhance its antioxidant and anti-inflammatory effects. In addition, the present invention can realize standardized production by controlling fermentation conditions, ensure the quality consistency of each batch of products, improve the overall quality of tangerine peel and the stability of drug efficacy, and is suitable for large-scale industrial production.

Description

Technical Field

[0001] The invention belongs to the technical field of traditional Chinese medicine processing, and particularly relates to a biological processing method of Huajuhong and Huajuhong obtained by the method. Background Art

[0002] Chinese medicine processing is a traditional Chinese medicine pharmaceutical technology unique to my country, which contains profound Chinese medicine theory and rich experience and skills. This technology uses specific processing methods to process Chinese medicine to reduce its toxicity or enhance its efficacy. The processing methods and processes of the processing process not only affect the appearance of Chinese medicinal materials, but are also directly related to the content and activity of its key ingredients, which ultimately affects the efficacy and quality of Chinese medicine. However, the traditional processing process of Chinese medicine often relies too much on manual experience and judgment, and there are differences in processing technology in various regions. There is a lack of objective and quantitative indicators, resulting in uneven processing quality. These problems not only limit the standardized production of Chinese medicine processing, but also affect the international competitiveness of Chinese medicine products.

[0003] With the development and application of bioengineering technology, biological processing has become an emerging research field in the field of Chinese medicine processing in recent years. Biological processing mainly refers to a method of using microorganisms or enzymes to process Chinese medicinal materials to improve their quality. This process can include biochemical changes such as fermentation, composting, and enzymolysis. Biological processing has many advantages. It can decompose macromolecules in Chinese medicinal materials into small molecules, making them easier to absorb; through the action of microorganisms, it can change the performance of Chinese medicine, improve the efficacy or reduce toxicity; and change the properties of medicinal materials during the fermentation process. For example, the originally cold medicinal materials become mild, which is in line with the theory of "nature, flavor and meridian" in traditional Chinese medicine. Therefore, the use of biological processing methods to process Chinese medicine can not only retain the characteristics of Chinese medicine, but also improve its quality, safety and efficacy, which represents an important direction for the development of Chinese medicine processing technology.

[0004] For example, Chinese patent document CN106879798A discloses a method for processing tangerine peel slices containing probiotic bacteria, which includes the following steps: a. Pretreatment and killing: put the tangerine peel fruit into a container containing inert gas within 1 hour, take it out and wash it with acid solution, and put it into saturated steam to kill and soften it; b. Slice: cut into slices; c. Enzymatic hydrolysis: dry it under a blast environment until the moisture content is 38-46%, spray the probiotic enzyme solution evenly, and press it into shape; d. Dry the tangerine peel slices: dry it under a temperature of 55-65℃, spray the alkaline solution, and continue to dry it until the moisture content is less than 13%, and finally obtain dry tangerine peel slices; e. Finished product packaging. The prepared hairy tangerine peel slices can effectively remove the bitter taste in the hairy tangerine peel slices, have a good taste after brewing, dissolve the effective ingredients quickly, and contain probiotics that are beneficial to the human body.

[0005] Huajuhong is the immature or nearly mature dried outer peel of Citrus grandis'Tomentosa' or Citrus grandis (L.) Osbeck, a plant of the Rutaceae family. It is one of the important medicinal materials in traditional Chinese medicine and has the effects of relieving cough and reducing phlegm, strengthening the stomach and digestion. Huajuhong is named because it is mainly produced in Huazhou, Maoming, Guangdong. Huajuhong was originally a wild pomelo tree. It has a unique geographical growth environment in Huazhou and has special effects after a long evolution. It is one of the "Four Major Southern Chinese Medicines" and "Ten Major Guangdong Medicines" and has won the reputation of "Southern Ginseng". Traditional processing methods have a significant impact on the efficacy of Huajuhong. Studies have shown that as time goes by, the total flavonoid content of Huajuhong increases, the aroma is fragrant, the bitter and numb taste decreases, and it is easier to be absorbed by the human body, and the effect is doubled. However, due to the lack of scientific quality control standards and systems, the consistency and effect of the processing process are difficult to guarantee, which affects the efficacy and market value of Huajuhong.

[0006] Therefore, modernizing the processing method of Huajuhong through biological processing is of great significance for improving the content of biologically active ingredients in Huajuhong and standardizing quality control. Summary of the invention

[0007] In view of the above problems existing in the prior art, the present invention provides a biological processing method for tangerine peel, which can significantly increase the content of active ingredients such as total flavonoids, naringin, wild sumac glycoside, bee venom glycoside, prunin and new north american erythrin in tangerine peel, and enhance its antioxidant and anti-inflammatory effects.

[0008] The present invention adopts the following technical solution:

[0009] The first aspect of the present invention provides a biological processing method of Citrus aurantium, comprising the following steps:

[0010] S1: Take fresh Huazhou pomelo or pomelo, wash, remove the inner pulp, cut into strips / slices, and then sterilize at high temperature, and then dry at 35-55°C to a moisture content of 60-70%, to obtain preliminarily processed Huajuhong slices;

[0011] S2: placing the Huajuhong tablets obtained in step S1 in an anaerobic environment, spraying 1-4wt% of the composite fermentation liquid A for biological processing, and performing microbial sterilization after the processing, wherein the composite fermentation liquid A is composed of Lactobacillus plantarum and Leuconostoc mesenteroides;

[0012] S3: placing the Huajuhong tablets processed in step S2 in an aerobic environment, spraying 3-8 wt % of the composite fermentation liquid B for biological processing, and sterilizing the microorganisms after the processing, wherein the composite fermentation liquid B is composed of Bacillus subtilis and Rhizopus chinensis;

[0013] S4: freeze-dry and dehydrate the prepared Huajuhong tablets to obtain a finished product.

[0014] Preferably, in step S1, the high temperature sterilization treatment temperature is 110-130° C., and the time is 10-20 min.

[0015] Preferably, in step S1, the mixture is dried at 40-45° C. until the moisture content is 60-70%.

[0016] Preferably, in step S2, the mass ratio of Lactobacillus plantarum to Leuconostoc mesenteroides in the composite fermentation broth A is (1-4):(3-7).

[0017] Preferably, in step S2, the number of viable bacteria in the plant lactobacillus powder in the composite fermentation liquid A is greater than 10 9 CFU / g, such as 1~10×10 9 CFU / g, the number of live bacteria in Leuconostoc mesenteroides powder is >10 11 CFU / g, such as 1~10×10 11 CFU / g.

[0018] Preferably, in step S2, the biological processing is carried out at a temperature of 28-32° C. and a humidity of 85-95% for 3-7 days, and the composite fermentation liquid A is sprayed every day during the fermentation.

[0019] Preferably, in step S3, the mass ratio of Bacillus subtilis to Rhizopus chinensis in the composite fermentation liquid B is (3-5): (4-9).

[0020] Preferably, in step S3, the number of viable bacteria in the Bacillus subtilis powder in the composite fermentation liquid B is greater than 10 8 CFU / g, such as 1~10×10 8 CFU / g, the number of viable bacteria in Rhizopus sinensis powder is more than 10 7 CFU / g, such as 1~10×10 7 CFU / g.

[0021] Preferably, in step S3, the biological preparation is carried out at a temperature of 25-30° C. and a humidity of 80-90% for 5-10 days, and the composite fermentation liquid B is sprayed every day during the fermentation.

[0022] Another aspect of the present invention also provides the Huajuhong prepared by the biological processing method.

[0023] Compared with the prior art, the present invention has the following beneficial effects:

[0024] The present invention uses a variety of microorganisms including Lactobacillus plantarum, Leuconostoc mesenteroides, Bacillus subtilis and Rhizopus sinensis to process the tangerine peel biologically, which can significantly increase the content of active ingredients such as total flavonoids, naringin, wild sumac glycoside, bee venom glycoside, prunin and new North American eriocin in the tangerine peel, and enhance its antioxidant and anti-inflammatory effects. In some embodiments of the present invention, the total flavonoids content in the tangerine peel is ≥13%, the naringin content is ≥10%, the wild sumac glycoside content is ≥0.15%, the bee venom glycoside content is ≥0.6%, the prunin content is ≥0.08%, and the new North American eriocin content is ≥0.09%.

[0025] In addition, the present invention realizes standardized production by controlling fermentation conditions (temperature, humidity, fermentation time, etc.), ensures the quality consistency of each batch of products, improves the overall quality of Huajuhong and the stability of its efficacy, and is suitable for large-scale industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 To investigate the effects of different processing methods of Citrus aurantium extract on NO. DETAILED DESCRIPTION

[0027] The present invention is further described in detail below in conjunction with the embodiments. It should be noted that the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments, and the present application should not be construed as being limited to the specific embodiments described. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in the art without creative work are within the scope of protection of the present invention.

[0028] 1. Preparation of Huajuhong

[0029] Example 1

[0030] This embodiment provides a biological processing method of Huajuhong, comprising the following steps:

[0031] S1: Take fresh Huazhou pomelo, wash it, remove the inner pulp, cut it into strips, and sterilize it at high temperature at 121°C for 15 minutes. Then dry it at 45°C until the water content is 65%, and obtain the preliminarily processed Huajuhong slices;

[0032] S2: Place the above-mentioned Huajuhong tablets in an anaerobic biological fermentation tank, spray 2 wt% of the composite fermentation liquid A for biological processing, the processing temperature is 30°C, the humidity is 90%, the processing time is 5 days, and the composite fermentation liquid A is sprayed every day during the fermentation. After the processing, microbial sterilization is performed, wherein the composite fermentation liquid A is composed of Lactobacillus plantarum and Leuconostoc mesenteroides, and the mass ratio is 2:3; the number of viable bacteria in the Lactobacillus plantarum powder is 2×10 9 CFU / g, the number of viable bacteria in the powder of Leuconostoc mesenteroides is 3×10 11CFU / g.

[0033] S3: Place the processed Huajuhong tablets in an aerobic biological fermentation tank and spray 5 wt% of the composite fermentation liquid B for biological processing. The processing temperature is 28°C, the humidity is 85%, and the processing time is 7 days. During the fermentation, the composite fermentation liquid B is sprayed every day. After the processing, microbial sterilization is performed. The composite fermentation liquid B is composed of Bacillus subtilis and Rhizopus chinensis, and the mass ratio is 3:5; the number of viable bacteria in the Bacillus subtilis powder is 2×10 8 CFU / g, the number of viable bacteria in Rhizopus sinensis powder is 1.5×10 7 CFU / g.

[0034] S4: placing the processed Huajuhong tablets in a freeze dryer for vacuum drying and dehydration to obtain a finished product.

[0035] Example 2

[0036] This embodiment provides a biological processing method of Huajuhong, comprising the following steps:

[0037] S1: Take fresh Huazhou pomelo, wash it, remove the inner pulp, cut it into strips, and sterilize it at high temperature at 121°C for 15 minutes. Then dry it at 40°C until the water content is 70%, and obtain the preliminarily processed Huajuhong slices;

[0038] S2: Place the above-mentioned Huajuhong tablets in an anaerobic biological fermentation tank, spray 2 wt% of the composite fermentation liquid A for biological processing, the processing temperature is 32°C, the humidity is 95%, the processing time is 4 days, and the composite fermentation liquid A is sprayed every day during the fermentation. After the processing, microbial sterilization is performed, wherein the composite fermentation liquid A is composed of Lactobacillus plantarum and Leuconostoc mesenteroides, and the mass ratio is 1:2; the number of viable bacteria in the Lactobacillus plantarum powder is 2×10 9 CFU / g, the number of viable bacteria in the powder of Leuconostoc mesenteroides is 3×10 11 CFU / g.

[0039] S3: Place the processed Huajuhong tablets in an aerobic biological fermentation tank and spray 5 wt% of the composite fermentation liquid B for biological processing. The processing temperature is 30°C, the humidity is 80%, and the processing time is 5 days. During the fermentation, the composite fermentation liquid B is sprayed every day. After the processing, microbial sterilization is performed. The composite fermentation liquid B is composed of Bacillus subtilis and Rhizopus chinensis, and the mass ratio is 4:5; the number of viable bacteria in the Bacillus subtilis powder is 2×10 8 CFU / g, the number of viable bacteria in Rhizopus sinensis powder is 1.5×10 7 CFU / g.

[0040] S4: placing the processed Huajuhong tablets in a freeze dryer for vacuum drying and dehydration to obtain a finished product.

[0041] Example 3

[0042] This embodiment provides a biological processing method of Huajuhong, comprising the following steps:

[0043] S1: Take fresh Huazhou pomelo, wash it, remove the inner pulp, cut it into strips, and sterilize it at high temperature at 121°C for 15 minutes. Then dry it at 45°C until the water content is 65%, and obtain the preliminarily processed Huajuhong slices;

[0044] S2: Place the above-mentioned Huajuhong tablets in an anaerobic biological fermentation tank, spray 2 wt% of the composite fermentation liquid A for biological processing, the processing temperature is 27 ° C, the humidity is 85%, the processing time is 6 days, and the composite fermentation liquid A is sprayed every day during the fermentation. After the processing, microbial sterilization is performed, wherein the composite fermentation liquid A is composed of Lactobacillus plantarum and Leuconostoc mesenteroides, and the mass ratio is 1:3; the number of viable bacteria in the Lactobacillus plantarum powder is 2×10 9 CFU / g, the number of viable bacteria in the powder of Leuconostoc mesenteroides is 3×10 11 CFU / g.

[0045] S3: Place the processed Huajuhong tablets in an aerobic biological fermentation tank and spray 5 wt% of the composite fermentation liquid B for biological processing. The processing temperature is 29°C, the humidity is 80%, and the processing time is 9 days. During the fermentation, the composite fermentation liquid B is sprayed every day. After the processing, microbial sterilization is performed. The composite fermentation liquid B is composed of Bacillus subtilis and Rhizopus chinensis, and the mass ratio is 4:5; the number of viable bacteria in the Bacillus subtilis powder is 2×10 8 CFU / g, the number of viable bacteria in Rhizopus sinensis powder is 1.5×10 7 CFU / g.

[0046] S4: placing the processed Huajuhong tablets in a freeze dryer for vacuum drying and dehydration to obtain a finished product.

[0047] Comparative Example 1

[0048] S1: Take fresh Huazhou pomelo, wash it, remove the inner pulp, cut it into strips, and sterilize it at high temperature at 121°C for 15 minutes. Then dry it at 45°C until the water content is 65%, and obtain the preliminarily processed Huajuhong slices;

[0049] S2: Place the processed Huajuhong tablets in an aerobic biological fermentation tank and spray 5 wt% of the composite fermentation liquid B for biological processing. The processing temperature is 28°C, the humidity is 85%, and the processing time is 7 days. During the fermentation, the composite fermentation liquid B is sprayed every day. After the processing, microbial sterilization is performed. The composite fermentation liquid B is composed of Bacillus subtilis and Rhizopus chinensis, and the mass ratio is 3:5; the number of viable bacteria in the Bacillus subtilis powder is 2×10 8 CFU / g, the number of viable bacteria in Rhizopus sinensis powder is 1.5×10 7 CFU / g.

[0050] S3: placing the processed Huajuhong tablets in a freeze dryer for vacuum drying and dehydration to obtain a finished product.

[0051] Comparative Example 2

[0052] S1: Take fresh Huazhou pomelo, wash it, remove the inner pulp, cut it into strips, and sterilize it at high temperature at 121°C for 15 minutes. Then dry it at 45°C until the water content is 65%, and obtain the preliminarily processed Huajuhong slices;

[0053] S2: Place the above-mentioned Huajuhong tablets in an anaerobic biological fermentation tank, spray 2 wt% of the composite fermentation liquid A for biological processing, the processing temperature is 30°C, the humidity is 90%, the processing time is 5 days, and the composite fermentation liquid A is sprayed every day during the fermentation. After the processing, microbial sterilization is performed, wherein the composite fermentation liquid A is composed of Lactobacillus plantarum and Leuconostoc mesenteroides, and the mass ratio is 2:3; the number of viable bacteria in the Lactobacillus plantarum powder is 2×10 9 CFU / g, the number of viable bacteria in the powder of Leuconostoc mesenteroides is 3×10 11 CFU / g.

[0054] S3: placing the processed Huajuhong tablets in a freeze dryer for vacuum drying and dehydration to obtain a finished product.

[0055] Comparative Example 3

[0056] S1: Take fresh Huazhou pomelo, wash it, remove the inner pulp, cut it into strips, and sterilize it at high temperature at 121°C for 15 minutes. Then dry it at 45°C until the water content is 65%, and obtain the preliminarily processed Huajuhong slices;

[0057] S2: Place the processed Huajuhong tablets in a freeze dryer for vacuum drying and dehydration to obtain the finished product.

[0058] 2. Efficacy Experiment

[0059] 1. Effects of different processing methods on the contents of total flavonoids and naringin in Citrus aurantium

[0060] Experimental method: After crushing the tangerine peel sample, take 1g of sample powder, add methanol at a solid-liquid ratio of 1:100, ultrasonically extract for 45min, and filter. After cooling to room temperature, add methanol to 100mL to obtain the tangerine peel extract filtrate. Use a pipette to draw 1mL of filtrate, add 9mL of 50% methanol for dilution, and determine the content of total flavonoids and naringin. Among them, the determination of the content of total flavonoids refers to: (Li Xingsui, Huang Shuhui, Guo Lilin. Determination of the content of total flavonoids in tangerine peel by ultraviolet-visible spectrophotometry [J]. Guangzhou Chemical Industry, 2022, 50(19): 127-129.), and the determination of naringin refers to the method of [Content Determination] under the tangerine peel item included in the 2020 edition of the "Chinese Pharmacopoeia".

[0061] Experimental results: As shown in Table 1, the contents of total flavonoids, naringin, naringin, bee venom glycoside, prunin and new erythrin in the example group are higher than those in the control example, indicating that the contents of active ingredients in the tangerine peel are increased after the co-biological processing of the composite fermentation broth A and the composite fermentation broth B, among which the effects of Examples 1 and 3 are better.

[0062] Table 1 Changes in the contents of total flavonoids and naringin in Citrus aurantium in different processing methods

[0063]

[0064] 2. Effects of different processing methods on the antioxidant capacity of Huajuhong

[0065] Experimental method: The antioxidant capacity of Citrus aurantium extract was evaluated by DPPH free radical scavenging test. 0.1 mMDPPH (dissolved in methanol) was added to a 96-well plate at a volume of 150 μL per well. Subsequently, 1 μL of the sample solution of Citrus aurantium extract to be tested was added. The 96-well plate was protected from light and incubated at room temperature for 30 minutes. The absorbance value was measured at 517 nm using a microplate reader to calculate the DPPH scavenging capacity.

[0066] Experimental results: As shown in Table 2, the DPPH radical scavenging rate of the Citrus aurantium extract in the example groups was higher than that in the control group, indicating that after the Citrus aurantium was processed by the composite fermentation broth A and the composite fermentation broth B, the radical scavenging ability of the Citrus aurantium was significantly improved, among which the effects of Examples 1 and 3 were better.

[0067] Table 2 DPPH radical scavenging rate of Citrus aurantium extracts prepared by different methods

[0068]

[0069] 3. Effects of different processing methods on the anti-inflammatory effect of Citrus aurantium extract

[0070] Experimental method: RAW264.7 cells were seeded at a density of 500,000 / mL in a 24-well plate, and blank group, model group, Example 1 group, Example 2 group, Example 3 group, Comparative Example 1 group, Comparative Example 2 group and Comparative Example 3 group were set. After culturing overnight, the supernatant was discarded, and fresh complete medium was added to the blank group; after the model group replaced the fresh medium for 2 hours, LPS (1 μg / mL) was added; Example 1~3 groups and Comparative Example 1~3 groups were pretreated with the corresponding 50μg / mL of Citrus aurantium extract for 2 hours, and then LPS (1 μg / mL) was added. After incubation for 24 hours in each group, the supernatant was taken, and the NO content was determined using the NO determination kit according to the instructions.

[0071] Experimental results: Figure 1 As shown, the inhibitory effect of the extract of Citrus aurantium in the example group on the NO production of RAW264.7 cells induced by LPS is better than that of the control group, indicating that the anti-inflammatory effect of Citrus aurantium is significantly improved after being biologically processed with the composite fermentation broth A and the composite fermentation broth B.

[0072] The above is a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A biological processing method of Citrus aurantium, characterized in that: The following steps are involved: S1: Take fresh pomelo, wash it, remove the inner pulp, cut it into strips / slices, and then sterilize it at high temperature, and then dry it at 35-55°C to a moisture content of 60-70%, to obtain preliminarily processed Huajuhong slices; S2: placing the Huajuhong tablets obtained in step S1 in an anaerobic environment, spraying 1-4wt% of the composite fermentation liquid A for biological processing, and performing microbial sterilization after the processing, wherein the composite fermentation liquid A is composed of Lactobacillus plantarum and Leuconostoc mesenteroides in a mass ratio of 1-2: 2-3; wherein the number of viable bacteria in the Lactobacillus plantarum powder is 2×10 9 CFU / g, the number of live bacteria in the powder of Leuconostoc mesenteroides is 3×10 11 CFU / g; S3: placing the Huajuhong tablets processed in step S2 in an aerobic environment, spraying 3-8 wt% of the composite fermentation liquid B for biological processing, and performing microbial sterilization after the processing, wherein the composite fermentation liquid B is composed of Bacillus subtilis and Rhizopus chinensis in a mass ratio of 3-4:5; wherein the number of viable bacteria in the Bacillus subtilis powder is 2×10 8 CFU / g, the number of viable bacteria in Rhizopus sinensis powder is 1.5×10 7 CFU / g; S4: freeze-dry and dehydrate the prepared Huajuhong tablets to obtain a finished product.

2. The biological processing method of Citrus aurantium according to claim 1, characterized in that: In step S1, the high temperature sterilization treatment temperature is 110-130° C. and the time is 10-20 minutes.

3. The biological processing method of Citrus aurantium according to claim 1, characterized in that: In the step S1, the mixture is dried at 40-45° C. until the moisture content is 60-70%.

4. The biological processing method of Citrus aurantium according to claim 1, characterized in that: In step S2, the biological preparation is carried out at a temperature of 28-32° C. and a humidity of 85-95% for 3-7 days, and the composite fermentation liquid A is sprayed every day during the fermentation.

5. The biological processing method of Citrus aurantium according to claim 1, characterized in that: In step S3, the biological preparation is carried out at a temperature of 25-30° C. and a humidity of 80-90% for 5-10 days, and the composite fermentation liquid B is sprayed every day during the fermentation.

6. The Citrus aurantium wilfordii obtained by the biological processing method according to any one of claims 1 to 5.

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