Processing method of traditional Chinese medicinal material poecilobdella manillensis

Abstract

The invention discloses a processing method of traditional Chinese medicinal material poecilobdella manillensis, and belongs to the technical field of traditional Chinese medicines. The processing method disclosed by the invention comprises the following steps: (1) carrying out feed deprivation culture on living poecilobdella manillensis; (2) freezing the living poecilobdella manillensis after the feed deprivation culture is finished; (3) performing water bath ultrasonic treatment on the frozen poecilobdella manillensis; the frequency of ultrasonic waves during ultrasonic treatment is 20-40 kHz; and (4) freeze-drying the poecilobdella manillensis subjected to the water bath ultrasonic treatment, and crushing to obtain the poecilobdella manillensis freeze-dried powder The processing method provided by the invention effectively solves the problem that the activity of antithrombin is reduced due to high temperature, reduces the fishy smell caused by blood storage in the body of the poecilobdella manillensis, improves the efficacy and quality of the poecilobdella manillensis freeze-dried powder, and improves the acceptance of patients. The processing process is low in energy consumption, easy in production condition control, high in processing efficiency and suitable for popularization and application.

Description

Technical Field

[0001] The invention relates to the technical field of traditional Chinese medicine, and in particular to a method for preparing a traditional Chinese medicinal material Hirudo philadelphica. Background Art

[0002] Poecilobdella manillensis, commonly known as golden-edged leech, is the dried whole body of Poecilobdella manillensis, an animal of the Hirudinidae family, Hirudinales class. It is a traditional Chinese medicinal material, first recorded in Shennong's Herbal Classic. As a medicinal material, it has the effects of breaking blood and promoting menstruation, removing blood stasis and eliminating symptoms. It is often used to treat blood stasis, amenorrhea, lumps, hemiplegia caused by stroke, and injuries caused by falls. Modern medical experimental studies have shown that anticoagulant active ingredients such as hirudin, hirudin A, and hirudin B are the main active ingredients of Poecilobdella manillensis. The whole body has anticoagulant effects and has a high therapeutic effect on diseases such as cerebral thrombosis, cerebral hemorrhage, hyperviscosity, hyperlipidemia, hypertension, and cor pulmonale.

[0003] However, there are some problems with the existing processing methods of Philippine leeches, which affect the quality and efficacy of the medicinal materials. The traditional processing method is to catch Philippine leeches in summer and autumn, wash them, scald them to death with boiling water, and then dry them in the sun or at low temperature. This method has obvious defects: if the Philippine leeches are directly scalded to death with boiling water, due to the high temperature, the antithrombin activity in the body will be greatly reduced, that is, the hirudin content will be reduced, which will directly affect the efficacy of the medicinal materials; because the Philippine leeches have a strong fishy smell, the medicinal materials that are directly dried and crushed will make patients feel nauseous when taken, and even cause vomiting, which not only affects the patient's medication experience, but also reduces the efficacy of the medicinal materials.

[0004] In order to avoid the destruction of antithrombin by high temperature, technicians have developed modern processing methods, such as low-temperature freeze-drying, which can maximize the retention of active ingredients in Philippine cattle leeches, such as antithrombin, and avoid the problem of quality decline of medicinal materials caused by high temperature treatment. However, in actual operation, it is still necessary to strictly control temperature, time and other conditions to ensure the quality of medicinal materials. In addition, this method has high requirements for equipment and technology, which may increase production costs. In order to reduce the fishy smell of Philippine cattle leeches, modern processing methods adopt the method of draining the blood in the body of Philippine cattle leeches, but the blood of Philippine cattle leeches also contains hirudin, and draining and storing blood is undoubtedly a waste of antithrombin, which reduces the efficacy of Philippine cattle leeches. Therefore, it is urgent to provide a simple and convenient method for processing Chinese medicinal materials Philippine cattle leeches with low fishy smell and high antithrombin content. Summary of the invention

[0005] The purpose of the present invention is to provide a processing method of a Chinese medicinal material Hirudo philadelphica to solve the problems existing in the above-mentioned prior art. The processing method of the present invention effectively solves the problem of decreased antithrombin activity caused by high temperature, and at the same time reduces the fishy smell caused by blood in the Hirudo philadelphica body, improves the efficacy and quality of Hirudo philadelphica freeze-dried powder, and improves patient acceptance. The processing process has low energy consumption, easy to control production conditions, high processing efficiency, and is suitable for promotion and application.

[0006] To achieve the above object, the present invention provides the following solutions:

[0007] The present invention provides a method for preparing a Chinese medicinal material Hirudo philadelphica, comprising the following steps:

[0008] (1) Stop feeding the live Philippine leeches;

[0009] (2) After the feeding period is over, the live Philippine leeches are frozen;

[0010] (3) subjecting the frozen leeches to water bath ultrasonic treatment, wherein the frequency of the ultrasonic wave during the ultrasonic treatment is 20-40 kHz;

[0011] (4) freeze-drying the leech after water bath ultrasonic treatment and then crushing it to obtain leech freeze-dried powder.

[0012] Optionally, the feeding-free culture period is 10-20 days.

[0013] Optionally, the temperature of the fasting culture is 20-25°C.

[0014] Optionally, the freezing temperature is -20°C and the freezing time is 30-50 minutes.

[0015] Furthermore, the temperature of the water bath ultrasonic treatment is 0-4°C.

[0016] Furthermore, the water bath ultrasonic treatment time is 10-15 minutes.

[0017] Furthermore, the freeze-drying procedure is as follows: the pre-freezing end point temperature is -40°C, the time is 1-2h; the sublimation drying temperature is -28°C to 5°C, the vacuum degree is below 20Pa, and the time is 10-12h; the analytical drying temperature is 10-40°C, the vacuum degree is below 25Pa, and the time is 8-12h.

[0018] Optionally, the particle size of the leech freeze-dried powder is less than 200 μm.

[0019] The present invention discloses the following technical effects:

[0020] The processing method of the Philippine leech of the present invention adopts the technology of fasting breeding, freezing ultrasound and freeze drying, effectively solves the problem of high temperature causing the antithrombin activity to decrease, reduces the fishy smell caused by blood in the Philippine leech, avoids the use of reagents such as flavoring agents, chemical reagents or auxiliary materials, ensures the naturalness of the medicinal material, improves the safety of the drug, improves the efficacy and quality of the Philippine leech freeze-dried powder, and improves the patient acceptance. The processing method of the present invention also shortens the freeze drying time, reduces energy consumption, is easy to control the production conditions, has stable product quality, and improves the processing efficiency of the Philippine leech, is more suitable for factory production, and provides new experience technology for the processing of animal medicinal materials in the field of traditional Chinese medicine. DETAILED DESCRIPTION

[0021] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but should be understood as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0022] It should be understood that the terms described in the present invention are only for describing a particular embodiment and are not intended to limit the present invention. In addition, for the numerical range in the present invention, it should be understood that each intermediate value between the upper and lower limits of the scope is also specifically disclosed. The intermediate value in any stated value or stated range, and each smaller range between any other stated value or intermediate value in the described range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

[0023] Unless otherwise indicated, all technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials associated with the documents. In the event of a conflict with any incorporated document, the content of this specification shall prevail.

[0024] It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments of the present invention description without departing from the scope or spirit of the present invention. Other embodiments derived from the present invention description will be apparent to those skilled in the art. The present invention description and examples are exemplary only.

[0025] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0026] The preparation steps of the leech freeze-dried powder of the present invention are as follows:

[0027] 1. Keep the live leeches at room temperature without feeding for 10-20 days to slow down the metabolic rate of the leeches and reduce the production of fishy substances. After the feeding period, wash the leeches with running water to remove impurities on the body surface.

[0028] 2. Freeze the live leeches after the food withdrawal treatment at -20°C for 30-50 minutes, take them out, and treat them with ultrasound in a water bath with the temperature of the water bath controlled at 0-4°C. The frequency of ultrasound is set to 20-40kHz for 10-15 minutes. Ultrasonic waves within this frequency range can produce a cavitation effect, which can remove dirt on the surface of the frozen leeches on the one hand, and on the other hand, the cavitation effect acts on the cells of the leeches, which can destroy the cell structure and release volatile substances with fishy smell (trimethylamine, etc.) during the gradual thawing process of the leeches to reduce the fishy smell of the finished product. At the same time, in the frozen state, active ingredients such as hirudin are not easy to lose.

[0029] 3. After the ultrasonic treatment, drain the leeches and place them in a vacuum freeze dryer for freeze drying; during the freeze drying process, set the pre-freezing end point temperature to -40°C and the time to 1-2h; the temperature of sublimation drying is -28°C to 5°C, the vacuum degree is below 20Pa, and the time is 10-12h; the temperature of analytical drying is 10-40°C, the vacuum degree is below 25Pa, and the time is 8-12h. Since the live leeches after the food withdrawal treatment are frozen, the leeches are still in a semi-frozen state after water bath ultrasound. Therefore, in the subsequent vacuum freeze drying process, the pre-freezing time can be shortened, and the energy consumption and time of the freeze drying process can be reduced.

[0030] 4. The freeze-dried leech is crushed to a particle size of less than 200 μm to obtain freeze-dried leech powder. The freeze-dried leech powder with this particle size has high solubility and higher bioavailability.

[0031] Example 1

[0032] 1. Keep the live leeches at room temperature without feeding for 15 days. After the feeding period, wash the leeches with running water to remove impurities on the body surface.

[0033] 2. Freeze the live leeches at -20°C for 40 minutes, take them out, and treat them with ultrasound in an ice water bath. The frequency of the ultrasound is set to 30kHz and the time is 10 minutes.

[0034] 3. After the ultrasonic treatment, drain the leeches and place them in a vacuum freeze dryer for freeze drying; during the freeze drying process, set the pre-freezing end temperature to -40°C and the time to 1.5h; the sublimation drying temperature is -28°C-5°C, the vacuum degree is below 20Pa, and the time is 12h; the analytical drying temperature is 10-40°C, the vacuum degree is below 25Pa, and the time is 8h.

[0035] 4. The freeze-dried leech is crushed to a particle size of less than 200 μm to obtain freeze-dried leech powder.

[0036] Example 2

[0037] 1. Keep the live leeches at room temperature without feeding for 10 days. After the feeding period, wash the leeches with running water to remove impurities on the body surface.

[0038] 2. Freeze the live leeches at -20°C for 30 minutes, take them out, and treat them with ultrasound in a water bath at 4°C. The frequency of ultrasound is set to 20kHz and the time is 15 minutes.

[0039] 3. After the ultrasonic treatment, drain the leeches and place them in a vacuum freeze dryer for freeze drying; during the freeze drying process, set the pre-freezing end temperature to -40°C and the time to 1.5h; the sublimation drying temperature is -28°C-5°C, the vacuum degree is below 20Pa, and the time is 12h; the analytical drying temperature is 10-40°C, the vacuum degree is below 25Pa, and the time is 8h.

[0040] 4. The freeze-dried leech is crushed to a particle size of less than 200 μm to obtain freeze-dried leech powder.

[0041] Example 3

[0042] 1. Keep the live leeches at room temperature without feeding for 20 days. After the feeding period, wash the leeches with running water to remove impurities on the body surface.

[0043] 2. Freeze the live leeches at -20°C for 50 minutes, take them out, and treat them with ultrasound in an ice water bath. The frequency of the ultrasound is set to 40kHz and the time is 10 minutes.

[0044] 3. After the ultrasonic treatment, drain the leeches and place them in a vacuum freeze dryer for freeze drying; during the freeze drying process, set the pre-freezing end temperature to -40°C and the time to 1.5h; the sublimation drying temperature is -28°C-5°C, the vacuum degree is below 20Pa, and the time is 12h; the analytical drying temperature is 10-40°C, the vacuum degree is below 25Pa, and the time is 8h.

[0045] 4. The freeze-dried leech is crushed to a particle size of less than 200 μm to obtain freeze-dried leech powder.

[0046] Comparative Example 1

[0047] The difference from Example 1 is that the process of stopping feeding and cultivating is omitted. Other operations are the same.

[0048] Comparative Example 2

[0049] The difference from Example 1 is that the ultrasonic treatment process is omitted. The other operations are the same.

[0050] Comparative Example 3

[0051] The difference from Example 1 is that the freezing in step 2 is omitted and the water bath ultrasonic treatment is directly performed. The other operations are the same.

[0052] Comparative Example 4

[0053] The difference from Example 1 is that the frequency of the ultrasonic wave is adjusted to 60 kHz. The other operations are the same.

[0054] Comparative Example 5

[0055] The difference from Example 1 is that the leech treated with ultrasound is bled and then vacuum freeze-dried. Other operations are the same. The specific method of bleeding is: the leech is completely thawed, hung up to drain blood, and squeezed when necessary to make the blood in the leech flow out.

[0056] Test Example 1

[0057] 1. Detection of antithrombin activity

[0058] The antithrombin activity in the lyophilized powder of Hirudo philadelphica of Examples 1-3 and Comparative Examples 1-5 was determined according to the method in the 2020 edition of the Chinese Pharmacopoeia. The test results are shown in Table 1.

[0059] Table 1 Antithrombin activity test results of leech freeze-dried powder

[0060] Philippine leech freeze-dried powder Antithrombin activity (U / g) Example 1 1203.5 Example 2 1232.4 Example 3 1182.0 Comparative Example 1 984.2 Comparative Example 2 1012.1 Comparative Example 3 642.8 Comparative Example 4 811.5 Comparative Example 5 517.4

[0061] According to Table 1, the antithrombin activity in the leech freeze-dried powder prepared in Examples 1-3 is relatively high, meeting the requirements for antithrombin activity in the leech freeze-dried powder decoction piece standard (Yun YPBZ-0199-2013). Compared with Examples 1-3, Comparative Example 1 omits the process of stopping feeding and breeding, which has little effect on the activity of antithrombin; Comparative Example 2 omits ultrasonic treatment, which has little effect on the activity of antithrombin; Comparative Example 3 omits freezing and directly performs water bath ultrasound, and it can be seen that the activity of antithrombin has decreased significantly, and during the ultrasonic treatment, the surface bleeding of the leech is obvious. It is speculated that it may be due to the strong effect of ultrasound on the living cells of the leech, which directly leads to serious lysis of the leech cells and loss of hirudin, resulting in a decrease in antithrombin; Comparative Example 4 increases the frequency of ultrasound, and the activity of antithrombin has also decreased, indicating that the ultrasound frequency will also affect the effect on the leech cells; Comparative Example 5 adopts the method of draining and storing blood, but this method obviously leads to a significant decrease in antithrombin activity.

[0062] 2. Sensory evaluation of fishy smell

[0063] The sensory evaluation of the fishy smell of the freeze-dried leech powder of Examples 1-3 and Comparative Examples 1-5 was performed, and the specific process was as follows:

[0064] 5 g of the lyophilized powder of Hirudo philadelphicae in Examples 1-3 and Comparative Examples 1-5 were taken respectively, put into 50 mL of warm water (40° C.), and stirred evenly to prepare a sample solution. 12 assessors (half male and half female, with age, sensory sensitivity, etc. kept consistent as much as possible) were selected to assess the sample solution, and the scoring standard was: 0-2 points for basically no fishy smell; 2-4 points for a slight fishy smell; 4-6 points for a fishy smell; 6-8 points for a relatively strong fishy smell; 8-10 points for a strong fishy smell.

[0065] The assessors tasted 1 mL of sample solution each time according to the specified scoring criteria. After tasting a sample, they rinsed their mouths with water until there was no medicine taste in their mouths before tasting the next sample. The scores of each assessor for each sample were recorded, and the average was calculated as the final score. The evaluation results are shown in Table 2.

[0066] Table 2 Evaluation results

[0067] Philippine leech freeze-dried powder Sensory Rating Example 1 1.75 Example 2 1.83 Example 3 1.83 Comparative Example 1 4.00 Comparative Example 2 5.58 Comparative Example 3 2.58 Comparative Example 4 2.08 Comparative Example 5 1.58

[0068] According to Table 2, the fishy smell of the freeze-dried leech powder prepared in Examples 1-3 is not obvious and can be basically ignored. Compared with Examples 1-3, the freeze-dried leech powder of Comparative Examples 1 and 2 has obvious fishy smell, the freeze-dried leech powder of Comparative Examples 3 and 4 has less fishy smell, and the freeze-dried leech powder of Comparative Example 5 has less fishy smell. Combining the data in Tables 1 and 2, the freeze-dried leech powder prepared in Examples 1-3 has the best quality, higher antithrombin activity, and less fishy smell.

[0069] 3. Quality inspection of Philippine leech freeze-dried powder

[0070] The moisture, total ash, acid-insoluble ash and pH of the leech freeze-dried powder of Example 1-3 were determined according to the method in the 2020 edition of the Chinese Pharmacopoeia. The test results are shown in Table 3.

[0071] Table 3 Quality inspection results of leech freeze-dried powder

[0072] index Example 1 Example 2 Example 3 Moisture 3.17％ 2.54％ 3.12％ Total ash 5.7％ 5.3％ 5.4％ Acid insoluble ash 0.5％ 0.4％ 0.5％ pH 6.2 6.5 6.6

[0073] It can be seen from Table 3 that the leech freeze-dried powder prepared in Examples 1-3 meets the quality requirements of the 2020 edition of the "Chinese Pharmacopoeia", and also meets the Yunnan Province Leech Freeze-dried Powder Decoction Standard (Yun YPBZ-0199-2013), and meets the listing standards.

[0074] The embodiments described above are only descriptions of the preferred modes of the present invention, and are not intended to limit the scope of the present invention. Without departing from the design spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by ordinary technicians in this field should all fall within the protection scope determined by the claims of the present invention.

Claims

1. A method for preparing the Chinese medicinal material Hirudo philadelphica, characterized in that: The steps include: (1) Stop feeding the live Philippine leeches; (2) After the feeding is stopped, the live Philippine leeches are frozen; (3) subjecting the frozen leeches to water bath ultrasonic treatment, wherein the frequency of the ultrasonic wave during the ultrasonic treatment is 20-40 kHz; (4) freeze-drying the leech after water bath ultrasonic treatment and then crushing it to obtain leech freeze-dried powder.

2. The processing method according to claim 1, characterized in that: The time of the feeding-free culture is 10-20 days.

3. The processing method according to claim 1, characterized in that: The temperature of the feeding-fasting culture is 20-25°C.

4. The processing method according to claim 1, characterized in that: The freezing temperature is -20°C and the freezing time is 30-50 minutes.

5. The processing method according to claim 1, characterized in that: The temperature of the water bath ultrasonic treatment is 0-4°C.

6. The processing method according to claim 1, characterized in that: The water bath ultrasonic treatment lasts for 10-15 minutes.

7. The processing method according to claim 1, characterized in that: The freeze-drying procedure is as follows: the pre-freezing end point temperature is -40°C, the time is 1-2h; the sublimation drying temperature is -28°C to 5°C, the vacuum degree is below 20Pa, and the time is 10-12h; the analytical drying temperature is 10-40°C, the vacuum degree is below 25Pa, and the time is 8-12h.

8. The processing method according to claim 1, characterized in that: The particle size of the leech freeze-dried powder is less than 200 μm.