Supercritical fluid extraction of artemisinin, CO2 as a solvent is low in price, non-toxic, non-flammable, can be recycled, and production does not cause environmental pollution. CO2 extraction artemisinin process is simple, the cycle is short, the operating temperature is close to room temperature, artemisinin almost no chemical changes such as thermal cracking. By changing the CO2 density and operating parameters (extraction pressure, temperature), the selectivity of CO2 for artemisinin dissolution can be improved, the content of impurities (wax) in the extract is low, the purification and purification of artemisinin is simple, and the yield is high. Good quality and good industrial application prospects.
1 test device and operation
The volume of the extraction tank of the supercritical fluid extraction device of this experiment was 4 L.
The raw material used for the experiment was Artemisia annua, which was produced in Baise, Guangxi and had a moisture content of 8% to 12% (air-dry) and an artemisinin content of 0.296%. CO2 is a product of Guangxi Hechi Chemical Industry Group, with a purity of ≥99.5%, a moisture content of ≤0.02%, and no odor.
After being crushed and sieved, Artemisia sphaerocephala is dropped into an extraction tank. CO2 from the cylinder is filtered by a filter and compressed by a compressor to a set pressure. The temperature is controlled and stabilized by the circulating water through the extraction tank jacket. When the extraction tank reaches the required pressure and temperature, cycle extraction is started and the extraction time is calculated. The artemisinin-containing CO2 is depressurized and enters the separation tank. Artemisinin (and impurities) is separated and separated in the separation tank, and CO2 is sent to the compressor for recycling. The extract is simply isolated and purified to obtain pure artemisinin.
2 Extract Processing and Analysis
2.1 Extract treatment: The extract is taken out of the separation tank, added several times the amount of ethanol solution is dissolved at a certain temperature, and an appropriate amount of powdered activated carbon is added. The activated carbon is removed by decoloring and hot filtration, and the activated carbon is washed with a small amount of hot ethanol solution for 3 times. The liquid and the filtrate were combined and allowed to stand, and the artemisinin crystals were precipitated and filtered, and the crystals were recrystallized with a 50% ethanol solution. Recrystallized in a constant temperature oven at 60 °C ~ 70 °C drying to get pure artemisinin.
2.2 Artemisinin Analysis
2.2.1 Thin Layer Chromatography (TLC): TLC analysis of the product with artemisinin standards, color development with iodine ultraviolet light or peroxide reagent (ammonium thiocyanate 0.625 g, ferrous sulfate 0.875 g, dissolve in 12 mL of distilled water and add 0.1 mL of concentrated sulfuric acid to prepare a solution.  Spray to develop color. Both have the same Rf value. With a mixture of 60% petroleum ether (40[deg.] C. to 60[deg.] C.) and 40% diethyl ether as developing solvent, the Rf value was 0.36. Each sample TLC analysis showed a single dot.
2.2.2 Spectral analysis: The product was confirmed by IR, NMR (1H, 13C) and MS analysis for the artemisinin structure.
2.2.3 Analysis of quality indicators: Product quality indicators: melting point 152°C to 154°C, optical rotation [α]25D=+75.3° (c, 1.644, chloroform), content 99.42%.
3 Results and Discussion
3.1 Gracilaria particle size test: Tests have shown that the appropriate raw material particle size can shorten the extraction time, which is beneficial to increase the yield of artemisinin. It shows that the supercritical CO2 extraction of artemisinin mass transfer process is controlled by intracellular diffusion of the plant. Therefore, the test material must be broken beforehand. In this study, 0.25 to 2 mm particles of Artemisia sphaerocephala were used for the experiment.
3.2 Relationship between artemisinin yield and extraction time: the same raw material particle size (0.2-2 mm) at the same extraction temperature (333 K), extraction pressure (20 MPa) and CO2 circulation (2.0-2.5 kg/h) Next, the relationship between extraction time and artemisinin yield was tested. The test results showed that the yield of artemisinin increased with the extension of extraction time when the extraction began. When the extraction time reached 2 h, the yield reached a maximum, and the yield of the extended extraction time increased little. Therefore, the subsequent tests were carried out by extraction for 2 h.
3.3 Relationship between artemisinin yield and extraction pressure: Extraction pressures of three extraction temperatures (308 K, 318 K, 333 K) and Artemisia annua were performed at the same extraction conditions with the same raw material particle size (0.2-2 mm). Yield test. From the test results, the yield of artemisinin increased with the increase of extraction pressure under the same extraction conditions. When the extraction pressure reached 20 MPa, the yield of artemisinin reached the maximum at the test temperature.
3.4 The relationship between the yield of artemisinin and the extraction temperature: Under the same extraction conditions, the yield of artemisinin increased significantly with increasing temperature within a certain temperature (308K ~ 333K). If the extraction pressure is 20 MPa, the yield of artemisinin is 72.3% at 308 K, 85.5% at 318 K, and 94.6% at 333 K. Raising the extraction temperature favors the supercritical CO2 extraction of artemisinin. However, as the temperature rises, the content of impurities in the extract also increases, which makes it more difficult to purify artemisinin. At the same time, the temperature is too high, and artemisinin can undergo chemical changes such as thermal cracking. Therefore, the extraction temperature should not be too high. When the extraction temperature is 333K, the yield has reached 94.6%, basically reaching the extraction requirements.
4.1 Artemisinin is extracted from Artemisia annua with supercritical CO2, and its mass transfer is controlled by the internal diffusion of plant cells. Therefore, it is necessary to smash or pre-soak Arty total turfgrass.
4.2 The extraction operation conditions have a great influence on the yield of artemisinin. Under a certain extraction temperature, the extraction pressure is increased and the yield is increased. Similarly, under a certain extraction pressure, increasing the extraction temperature, the artemisinin yield also increased. That is, changing the extraction pressure and temperature can yield higher yields, but there is a limit. In this study, the yield of artemisinin was greater than 95% under the optimum extraction conditions.
4.3 Supercritical CO2 has good solubility to artemisinin and its extraction cycle is short. By changing the supercritical CO2 extraction process conditions, the artemisinin solubilization selectivity can be improved, and the purification and refining of artemisinin can be simple and easy. The artemisinin product has high purity and good quality.