1) Extraction: Get 1 kg of saffron powder and also essence twice, each use 1L mass focus of 70% ethanol aqueous solution, 45 ° (: mixing and also extracting 2h, filter, the filtrate of twice filtering is merged, under reduced pressure condition Spin to recoup solvent as well as concentrate to 2L.
2) Removal: the focused option obtained in step 1) was adjusted to pH 2 with 1mol/L hydrochloric acid, drawn out 3 times with 2L water-saturated n-butanol, and the natural layers were integrated as well as focused to 400mL;
3) Crystallization for the first time: the concentrated solution obtained in step 2) was included in 960 mL of acetone as well as 240 mL of methyl tert-butyl ether mixed solvent, warmed to reflux under mixing, then cooled down to 20 ° C to separate out a white precipitate, filteringed system, 45 ° C dried to offer 49g of white crystals;
4) Crystallization for the 2nd time: 49g crystals acquired by step 3) were liquified in 100 ° C of hot water of 200mL, mixed for 20min, then the remedy was cooled down to 0 ° C, a white strong was precipitated, suction filteringed system, as well as the solid was dissolved in Drying out under the condition of 45 DEG C, obtain β-escin 32g;
5) Preparation of aescin ended up item: Dissolve the 32g3-aescin acquired in step 4) in a mixed solvent of 300mL ethanol as well as 15mL n-hexane, add 4.8gL-proline to the above system, warmth and reflux for 30min, as well as dissolve the system Cool to 50 ° C, include 0.2 g of escin B with a pureness of 98% as seed crystals, continue to stir and cool down to 20 ° C, take shape up until white crystals no longer speed up, filter with suction, and dry the strong at 45 ° C After drying out, 12.5 g of white crystals of aescin B were obtained.
In this embodiment, the function of action I) is to remove and divide the total saponins from the seeds of sage, which is a standard process for drawing out the total saponins from the seeds, and also the removal approach can be reflux, soaking, leak, etc generally utilized in this field. method. Among them, the focus of ethanol will have an important influence on the pureness and also return of complete aescin. If the concentration of ethanol is also reduced, the removal of the product will certainly want, and also if the focus of ethanol is too expensive, the material of impurities will boost.
The removal of action 2) as well as the condensation functions of steps 3) and 4) are all to detoxify the essence to obtain high-purity β-escin. Amongst them, the purpose of readjusting pH is to boost the security and extraction efficiency of aescin. If the pH is expensive, the water solubility of aescin will enhance, and the removal performance of n-butanol will decrease; if the pH is too low, the security of aescin will be reduced. The ratio of acetone as well as methyl tert-butyl ether symphonious (3) will certainly influence the precipitation rate and also yield of aescin crystals.
The natural solvent symphonious 5) can also be picked from methanol, ethanol, isopropanol, acetone, tetrahydrofuran, methyl tert-butyl ether, n-hexane, n-heptane, and so on. Several arbitrary combinations, the objective is to make β-Aescin can dissolve well, as well as at the same time, it can much better separate out Aescin B13 in the generated recrystallization. The quantity of L-proline straight influences the splitting up as well as rainfall of Aescin B. Phylloside B can not just selectively as well as smoothly speed up crystals, yet also has high return and also pureness, and great crystal morphology.
Embrace high performance fluid phase approach to find aescin B in the product, particular condition is: chromatographic column is C18 message, as well as fragment diameter is 5um; Mobile phase is acetonitrile and also 0.5% phosphoric acid liquid service, and also the quantity proportion of acetonitrile as well as phosphoric acid water is 35:65, mobile stage PH= 2.,5; discovery wavelength is 220nm; circulation rate is 1.0 ml/min, shot volume is 10 μl;
The content of aescin B in the item was 99.7%, and the yield (the overall quantity of aescin B in the item accounted for the percent of the overall amount of aescin B in the feeding sage) was 61%.