Use of bisabolane-type sesquiterpenoids in the preparation of anti-complement drugs
A technology of bisabolane and sesquiterpenoids, applied in the field of bisabolane-type sesquiterpenoids and preparation of anti-complement drugs
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Embodiment 1
[0024] Example 1 Preparation of bisabolane-type sesquiterpenoids
[0025] Take 3.5 kg of dried roots and rhizomes of L. tekesi, crush them, and use petroleum ether: ether: methanol = 1:1:1 to repeatedly cold soak and extract 3 times at room temperature, soaking each time for 7 days. The solvent was recovered under reduced pressure to obtain 130 g of the total extract. 130g of total extract and 100-200 mesh silica gel 1:1 mixed sample, wet column (5 kg SiO 2 ), petroleum ether-acetone gradient elution (50:1, 30:1, 15:1, 9:1, 7:1, 5:1, 3:1, 2:1, 1:1) fractions were subjected to TLC The detection was pooled into 11 fractions (Fr.1-11). Fraction Fr.7 (2.5g), was loaded on a forward silica gel column by dry method, eluted with petroleum ether-acetone (10:1), obtained fractions, sampled, detected by TLC and concentrated sulfuric acid-vanilla The aldehyde reagent was used for color observation, and the samples were combined to obtain 3 parts, which were eluted by silica gel column...
Embodiment 2
[0026] Example 2 Anti-complement classical pathway test in vitro
[0027] Take 0.1ml of complement (guinea pig serum), add barbiturate buffer solution (BBS) to prepare a 1:5 solution, and double-dilute with BBS to 1:10, 1:20, 1:40, 1:80, 1: 160, 1:320 and 1:640 solutions; take 1:1000 hemolysin, each concentration of complement and 2% sheep red blood cell (SRBC) each 0.1ml dissolved in 0.3ml BBS, mix well, 37 ℃ water bath for 30min, then put into low temperature Centrifuge in a high-speed centrifuge at 5000rpm and 4°C for 10min, take 0.2ml of the supernatant from each tube and put it in a 96-well plate, measure its absorbance at 405nm, and set up a full hemolysis group (0.1ml of 12% SRBC dissolved in 0.5ml of three Distilled water), using the absorbance of three-distilled water lysed blood vessels as the standard of full hemolysis, calculate the hemolysis rate, take the dilution of complement as the X-axis, and the percentage of hemolysis caused by each dilution of complement a...
Embodiment 3
[0028] Example 3 Anti-complement alternative pathway test in vitro
[0029] Take 0.2ml of complement (human serum), add AP diluent (barbital buffer, pH 7.4, containing 5 mMg 2+ , 8mMEGTA) was prepared into a 1:5 solution, and double-diluted into 1:10, 1:20, 1:40, 1:80, 1:160, 1:320 and 1:640 solutions, and each concentration of complement 0.15ml, 0.15ml of AP diluent and 0.20ml of 0.5% rabbit red blood cells (RE), mix well, put in a low-temperature high-speed centrifuge after 30min in 37℃ water bath, centrifuge at 5000rpm, 4℃ for 10min, take each tube Put 0.2 ml of the supernatant in a 96-well plate, and measure the absorbance at 405 nm. At the same time, a complete hemolysis group (0.20 ml 0.5% RE dissolved in 0.3 ml triple-distilled water) was set up in the experiment. The absorbance of the three-distilled water lysed blood vessel was used as the standard of total hemolysis to calculate the hemolysis rate , with the complement dilution as the X-axis, the percentage of hemol...
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