Process for preparing artemisia apiacea powder by compound enzymolysis method

Abstract

The invention relates to the technical field of artemisia apiacea enzymolysis processes, and particularly discloses a process for preparing artemisia apiacea powder through a composite enzymolysis method.The process comprises the following steps of raw material pretreatment, enzymolysis optimization, inactivation, concentration and drying.A composite system of cellulase, pectinase, beta-glucanase and acid protease is adopted, the limitation of single enzymolysis is broken through, and the enzymolysis efficiency is improved; the release amount of reducing sugar is increased, and release of artemisinin, polysaccharide and flavonoid components is promoted; the material-to-liquid ratio is 1: 10 (artemisia apiacea powder: buffer solution), the enzymolysis efficiency and cost are balanced, the method is suitable for industrial production, the whole preparation process adopts a low-temperature technology, the biological activity is retained, the finished product is dry powder (the water content is less than or equal to 5%), the product is convenient to package, transport and store for a long time, and the raw material cost of enterprises in non-production areas is reduced.

Description

Technical Field

[0001] The present invention belongs to the technical field of artemisiae enzymatic hydrolysis process, and particularly relates to a process for preparing artemisiae powder by a composite enzymatic hydrolysis method. Background Art

[0002] Artemisia annua (sweet wormwood): As a traditional Chinese medicine with a long history, it was first recorded in "Prescriptions for Fifty-two Diseases" and "Pharmacopoeia". It is recorded that artemisia annua has the therapeutic effects of clearing away deficiency-heat, relieving summer heat, eliminating bone steaming, subsiding jaundice, and arresting malaria, and is mainly used to treat syndromes such as pathogenic heat damaging yin, feverishness in the evening and coolness in the early morning, yin deficiency fever, and bone-steaming fever. "Dietary Herbal" records the functions and indications of artemisia annua as "tonifying qi, promoting hair growth, tonifying the middle-jiao, improving eyesight, and expelling wind toxins. Treating bone steaming. Boiling the ash to obtain juice and decocting with lime to treat stubborn sores and scars." "Shennong's Herbal Classic" records that it is bitter, pungent, and cold in nature, and belongs to the liver and gallbladder meridians; it has the effects of clearing away deficiency-heat, relieving summer heat, eliminating bone steaming, and arresting malaria, and can effectively treat syndromes such as pathogenic heat damaging yin, yin deficiency fever, feverishness in the evening and coolness in the early morning, damp-heat jaundice, and summer-heat pathogen fever. Modern pharmacological studies have shown that the sesquiterpenoids and flavonoids contained in artemisia annua have good anti-malarial, anti-viral, anti-inflammatory and antipyretic, anti-inflammatory, and antibacterial and insecticidal effects. The volatile oil components can exert anti-tumor, antibacterial and insecticidal, and anti-inflammatory activities. The effects of these three types of components are basically consistent with the traditional efficacy of artemisia annua.

[0003] At present, the extraction of the active ingredients of artemisia annua mainly relies on the following methods: Organic solvent extraction method: There is a risk of solvent residue, and the active ingredients are easily damaged at high temperatures. Supercritical carbon dioxide method: The equipment cost is high, and the industrial promotion is limited. Intelligent continuous countercurrent extraction method: Complex equipment is required, and the energy consumption is relatively high. Ultrasonic and microwave assisted method: Although the efficiency can be improved, the protection of heat-sensitive components is insufficient.

[0004] When using the enzymatic hydrolysis method for preparation, only cellulase or pectinase is used as a single enzyme species, resulting in insufficient extraction rates of artemisinin, polysaccharides and other components. Direct use of artemisia annua by the folk has defects such as poor taste and easy deterioration, and the transportation cost of raw materials in non-producing areas is high. Summary of the Invention

[0005] The purpose of the present invention is to provide a process for preparing artemisia annua powder by a composite enzymatic hydrolysis method to solve the problems raised in the above background art.

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

[0007] A process for preparing artemisia annua powder by a composite enzymatic hydrolysis method, comprising the following steps:

[0008] Raw material pretreatment:

[0009] S01. Harvesting: Select artemisia annua for harvesting when it has grown for 130 - 150 days;

[0010] S02. Cleaning: Rinse fresh artemisia annua leaves with clean water to remove sediment and impurities;

[0011] S03, Drying: Bake at a low temperature of 50°C until the moisture content ≤ 8%;

[0012] S04, Crushing: Crush and sieve through a 40 - 60 mesh sieve;

[0013] Enzymatic hydrolysis optimization:

[0014] S05, Selection of composite enzyme species: The composite enzyme species include cellulase, pectinase, β - glucanase, and acid protease; The enzyme species are selected as cellulase (2000U / g) + pectinase (1500U / g) + β - glucanase (500U / g) or cellulase (2000U / g) + pectinase (1500U / g) + β - glucanase (500U / g) + acid protease (500U / g);

[0015] S06, Ratio of composite enzyme system:

[0016] The mass ratio of cellulase: pectinase: β - glucanase: acid protease is 3:2:0.5:2.5;

[0017] S07, Enzymatic hydrolysis buffer:

[0018] Use citric acid - disodium hydrogen phosphate buffer, add the above - mentioned composite enzyme and mix at 100ml / 0.3g, and adjust to pH 4.8 - 5.2 and temperature to 50 - 55°C;

[0019] S08, Enzymatic hydrolysis of Artemisia annua:

[0020] Add the Artemisia annua powder obtained in S04 to the enzymatic hydrolysis buffer in S07, material - liquid ratio: 1:10 (Artemisia annua powder: buffer), stir at a constant speed of 100 - 150 rpm, and carry out enzymatic hydrolysis for 2 - 3 hours;

[0021] Inactivation

[0022] S09, Terminate the reaction by water - bathing at 90°C for 10 minutes and cool to room temperature;

[0023] Concentration and drying

[0024] S10, Vacuum concentration: Concentrate under reduced pressure at 45°C until the solid content ≥ 30%;

[0025] S11, Vacuum - dry at 60 - 80°C to obtain the finished product.

[0026] Preferably, during S01 harvesting, avoid rainy days, and carry out harvesting on cloudy days, overcast days, sunny days. On sunny days, harvest before 11 am and after 5 pm. On cloudy days, harvest throughout the day. Cut down the above - ground part of Artemisia annua from the stem near the root, remove the main stem, and retain the side branches with Artemisia annua leaves.

[0027] Preferably, the drying equipment in S03 of the present invention includes a CT-C series hot air circulation oven, a multi-layer mesh belt dryer and a DZF series vacuum drying oven, and the crushing equipment in S04 includes a CT-C series hot air circulation crushing unit, a Baishun Da wall-breaking crusher and a 20B / 30B / 40B / 60B series crusher.

[0028] The enzymolysis tank in S08 includes an MJ type enzymolysis reaction tank and DZF-6020 (50L) and DZF-6050 (100L) enzymolysis tanks, and the vacuum concentration and drying equipment includes a DZF series vacuum drying oven, a disc vacuum dryer, and a Lima vacuum low-temperature dryer.

[0029] Compared with the prior art, the present invention has the following beneficial effects:

[0030] The invention adopts a composite system of cellulase, pectinase, beta-glucanase and acid protease, breaks through the limitation of single enzymolysis, improves the release of reducing sugar, and promotes the release of artemisinin, polysaccharides and flavonoid components; the material-liquid ratio of 1:10 (artemisia annua powder: buffer) balances the enzymolysis efficiency and cost, and is suitable for industrial production.

[0031] At the same time, the preparation process of the present invention adopts low-temperature technology throughout the whole process to retain biological activity, avoid thermal degradation of artemisinin, maintain enzyme activity and reduce the loss of heat-sensitive components; at the same time, the concentration process is concentrated under reduced pressure at 45°C to retain volatile components (artemisia annua volatile oil) and the integrity of active ingredients, avoiding secondary destruction of active ingredients by high-temperature sterilization.

[0032] At the same time, the finished product of this product is a dry powder (moisture content ≤5%), which is convenient for packaging, transportation and long-term storage, reducing the raw material costs of enterprises in non-production areas; it can be directly used in the fields of medicine, food, feed, etc., or as a deep-processing raw material to broaden the application scenarios. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] Figure 1 It is a process flow chart of the present invention. DETAILED DESCRIPTION

[0034] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0035] Embodiment 1:

[0036] See also Figure 1 A composite enzymatic hydrolysis method for preparing artemisia annua powder comprises the following steps:

[0037] Preparation equipment: Model 40 crusher (40 - 60 mesh sieve, particle size uniformity ≥ 95%)

[0038] YZ - 50L constant temperature oscillating enzymatic hydrolysis tank (pH / temperature self - control, multi - parameter on - line monitoring)

[0039] ZW - 1000 vacuum extraction and concentration equipment (vacuum degree 0.09 MPa, jacket pressure 0.3 MPa, condensation area 10.0 ㎡)

[0040] Harvesting and processing

[0041] In July - August in the south (September - October in the north), select the above - ground part of Artemisia annua before 11 am or after 5 pm on a sunny day, and retain the leafy side branches.

[0042] Rinse the fresh Artemisia annua leaves with clear water to remove sediment and impurities.

[0043] Drying and crushing

[0044] Drying: Use the heating function of the YZ - 50L enzymatic hydrolysis tank, set the low - temperature drying at 50℃ until the moisture content ≤ 8% (about 6 - 8 hours).

[0045] Crushing: Pass through a 40 - mesh sieve with the XX - 40 crusher to obtain Artemisia annua powder (particle size ≤ 0.425 mm).

[0046] II. Enzymatic hydrolysis optimization

[0047] Configuration of composite enzyme system

[0048] Enzyme types and ratios:

[0049] Cellulase (2000 U / g): 0.1125 g / L

[0050] Pectinase (1500 U / g): 0.075 g / L

[0051] β - Glucanase (500 U / g): 0.01875 g / L

[0052] Acidic protease (500 U / g): 0.09375 g / L

[0053] Buffer solution: Citric acid - disodium hydrogen phosphate buffer solution (pH 4.8 - 5.2), adjusted according to the dose of 100 ml / 0.3 g, and heated to 50 - 55℃.

[0054] Enzymatic hydrolysis reaction

[0055] Ratio of material to liquid: Artemisia annua powder: buffer solution = 1:10 (e.g., 10 kg of Artemisia annua powder is mixed with 100 L of buffer solution).

[0056] Operating conditions:

[0057] Use the YZ-50L enzymatic hydrolysis tank, and set the temperature at 50°C, pH at 5.0, and stirring speed at 120 rpm.

[0058] Enzymatic hydrolysis time: 2.5 hours (online monitoring shows that the polysaccharide degradation rate is ≥85%).

[0059] III. Inactivation and Concentration

[0060] After the enzymatic hydrolysis is completed, raise the temperature of the enzymatic hydrolysis tank to 90°C, maintain for 10 minutes to terminate the reaction, and then cool to room temperature.

[0061] Vacuum Concentration

[0062] Transfer the enzymatic hydrolysate to the ZW-1000 vacuum concentration equipment, set the vacuum degree at 0.09 MPa and the jacket temperature at 45°C, and concentrate until the solid content is ≥30% (about 3 - 4 hours are required).

[0063] IV. Drying and Finished Product

[0064] Vacuum Drying

[0065] Continue to raise the temperature to 60 - 80°C in the ZW-1000 equipment, maintain the vacuum degree at 0.09 MPa, and dry until the moisture content is ≤5% (about 2 hours are required).

[0066] Finished Product Packaging

[0067] Discharge, crush, and pass through an 80-mesh sieve to obtain the finished product of artemisinin extract, and store it sealed and away from light.

[0068] Among them, the equipment regulations are as follows:

[0069]

[0070] Example 2:

[0071] A process for preparing artemisinin powder by a composite enzymatic hydrolysis method includes the following steps:

[0072] S01. Harvesting: Artemisia annua can be harvested after growing for 130 - 150 days (in the south from July to August, in the north from September to October). Avoid harvesting on rainy days, and it can be harvested on cloudy days, overcast days, or sunny days. Before 11 am and after 5 pm on sunny days, and throughout the day on cloudy days, cut down the above-ground part of Artemisia annua from the stem near the root, remove the main stem, and retain the side branches with Artemisia annua leaves;

[0073] S02. Cleaning. Rinse the fresh Artemisia annua leaves with clean water to remove sediment and impurities;

[0074] S03. Drying. Dry at a low temperature of 50°C until the moisture content is ≤8% to avoid destroying the active ingredients at high temperatures;

[0075] S04. Crushing. Crush and pass through a 40-mesh sieve to increase the contact area with the enzyme;

[0076] S05. Enzyme selection. Cellulase (2000 U / g) + pectinase (1500 U / g) + β-glucanase (500 U / g), which synergistically decompose cell wall polysaccharides; Auxiliary enzyme (optional): Acid protease (500 U / g) is used to decompose conjugated proteins and improve the artemisinin release rate;

[0077] S06. Ratio of the complex enzyme system. Cellulase: Pectinase: β-glucanase: Acid protease

[0078] S07. Enzymatic hydrolysis buffer. Use citric acid-disodium hydrogen phosphate buffer, add the above complex enzyme at a ratio of 100 ml / 0.3 g (citric acid-disodium hydrogen phosphate buffer / complex enzyme) for mixing, and adjust to pH 4.8 and a temperature of 50 °C;

[0079] S08. Enzymatic hydrolysis of Artemisia annua. Add the Artemisia annua powder obtained in (1) to the enzymatic hydrolysis buffer in (S7), with a material-liquid ratio of 1:10 (Artemisia annua powder: buffer), stir at a constant speed of 100 - 150 rpm, and perform enzymatic hydrolysis for 2 - 3 h;

[0080] S09. Inactivation. Terminate the reaction by a 10-minute water bath at 90 °C and cool to room temperature.

[0081] S10. Concentration. Vacuum concentration: Concentrate under reduced pressure at 45 °C until the solid content ≥ 30%.

[0082] S11. Drying. Vacuum dry at 60 °C to obtain the finished product.

[0083] Example 3:

[0084] A process for preparing Artemisia annua powder by a complex enzymatic hydrolysis method includes the following steps:

[0085] S01. Harvesting: Artemisia annua can be harvested after growing for 130 - 150 days (July - August in the south, September - October in the north). Avoid harvesting on rainy days, and it can be harvested on cloudy days, overcast days, or sunny days. Before 11 am and after 5 pm on sunny days, and throughout the day on cloudy days. Cut down the above-ground part of Artemisia annua from the stem near the root, remove the main stem, and retain the lateral branches with Artemisia annua leaves;

[0086] S02. Cleaning. Rinse the fresh Artemisia annua leaves with clean water to remove sediment and impurities;

[0087] S03. Drying. Dry at a low temperature of 50 °C until the moisture content ≤ 8%, to avoid destroying the active ingredients at high temperatures;

[0088] S04. Crushing. Crush and pass through a 60-mesh sieve to increase the enzyme contact area;

[0089] S05. Enzyme selection. Cellulase (2000 U / g) + pectinase (1500 U / g) + β-glucanase (500 U / g), which synergistically decompose cell wall polysaccharides; Auxiliary enzyme (optional): Acid protease (500 U / g) is used to decompose conjugated proteins and improve the artemisinin release rate;

[0090] S06. Ratio of the complex enzyme system. Cellulase: Pectinase: β-glucanase: Acid protease (3:2:0.5:2.5);

[0091] S07. Enzymatic hydrolysis buffer. Use citric acid-disodium hydrogen phosphate buffer, add the above complex enzyme at a ratio of 100 ml / 0.3 g (citric acid-disodium hydrogen phosphate buffer / complex enzyme) for blending, and adjust to pH 5.0 and a temperature of 55 °C;

[0092] S08. Enzymatic hydrolysis of Artemisia annua. Add the Artemisia annua powder obtained in (1) to the enzymatic hydrolysis buffer in (S7), material-liquid ratio: 1:10 (Artemisia annua powder: buffer), stir at a constant speed of 100 - 150 rpm, and perform enzymatic hydrolysis for 2 h - 3 h;

[0093] S09. Inactivation. Terminate the reaction by a 10-minute water bath at 90 °C and cool to room temperature.

[0094] S10. Concentration. Vacuum concentration: Concentrate under reduced pressure at 45 °C until the solid content ≥ 30%.

[0095] S11. Drying. Vacuum dry at 70 °C to obtain the finished product.

[0096] It should be understood that in the development process of any actual implementation, such as in any engineering or design project, a large number of specific implementation decisions can be made. Such development efforts may be complex and time-consuming, but for those of ordinary skill in the art who benefit from this disclosure, without excessive experimentation, such development efforts will be a routine task of design, manufacture, and production.

[0097] 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 the preferred embodiments, those of ordinary skill in the art should understand that the technical solutions of the present invention can be modified or equivalently replaced without departing from the spirit and scope of the technical solutions of the present invention, and they should all be covered within the scope of the claims of the present invention.

Claims

1. A process for preparing artemisia annua powder by a composite enzymatic hydrolysis method, characterized in that: The following steps are involved: Raw material pretreatment: S01. Harvesting: Harvesting Artemisia annua at 130-150 days of growth; S02. Cleaning: Wash the fresh Artemisia annua leaves with clean water to remove sand and impurities; S03, Drying: Dry at 50℃ until the moisture content is ≤8%; S04, crushing: crush through 40-60 mesh sieve; Enzymatic hydrolysis optimization: S05. Selection of compound enzymes: the compound enzymes include cellulase, pectinase, β-glucanase and acid protease; the enzymes are selected from cellulase (2000U / g) + pectinase (1500U / g) + β-glucanase (500U / g) or cellulase (2000U / g) + pectinase (1500U / g) + β-glucanase (500U / g) + acid protease (500U / g); S06, compound enzyme system ratio: The mass ratio of cellulase: pectinase: β-glucanase: acid protease is 3:2:0.5:2.5; S07, Enzyme lysis buffer: Use citric acid-sodium hydrogen phosphate buffer, add the above complex enzyme at 100ml / 0.3g, and adjust to pH 4.8-5.2 and temperature to 50-55℃; S08, Enzymatic hydrolysis of Artemisia annua: Add the Artemisia annua powder obtained in S04 to the enzymolysis buffer in S07, with a solid-liquid ratio of 1:10 (Artemisia annua powder: buffer), stir at a constant speed of 100-150 rpm, and perform enzymolysis for 2h-3h; Inactivation S09, 90℃ water bath for 10 minutes to terminate the reaction, and cool to room temperature; Concentration and drying S10, vacuum concentration: concentrate at 45℃ under reduced pressure until the solid content is ≥30%; S11. Vacuum drying at 60-80°C to obtain the finished product.

2. The process for preparing artemisia annua powder by a composite enzymatic hydrolysis method according to claim 1, characterized in that: The S01 is harvested on rainy days, and is harvested on cloudy days, overcast days, and sunny days. It is harvested before 11 a.m. and after 5 p.m. on sunny days, and all day on cloudy days. The above-ground part of the Artemisia annua is cut down from the stem near the root, the main stem is removed, and the side branches with Artemisia annua leaves are retained.

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