Isolation, Purification And Transformation Of Chlorogenic Acids From Artemisia Annua L

Artemisia annua L.is a plant used for the treatment of malaria and fever.As a natural resource of Chinese herbal medicine,it is used in the production of artemisin in the current industry.The work was carried out to study the separation?purification and transformation process of chlorogenic acids in Artemisia annua L.A method using Liquid Chromatography and Negtive Electrospary Ionization Tandem Mass Spectrometry method was established for the comprehensive and accurate identification of chlorogenic acids in the extract of A.annua.The mobile phase consisted of 0.05%(v/v)acetic acid in water(A)and acetonitrile(B).Thirty-six chlorogenic acids were successfully identified in 45 min using gradient elution.Fifteen were reported for the first time from A.annua.These chlorogenic acids comprised two monocaffeoylquinic acids,five dicaffeoylquinic acids,one feruloylquinic acid,three caffeoyl-feruloylquinic acids,two caffeoyl-dimethoxycinnamoylquinic acids,one dimethoxycinnamoylquinic acid and one coumaroylquinic acid.Especially,1,5-dicaffeoylquinic acid was the main composition of anti-aids drug-IBE-5,which is chemical synthesis drug with independent intellectual property rights in China,which provided a new sources for dicaffeoylquinic acid separation.The extraction of caffeoylquinic acids in Artemisia annua L.was investigated.Single factor test on the major influencing factors such as extraction solvent type,extraction concentration,liquid to material ratio and extraction time were studied.The extractions were as follows:ethanol-water(30:70,v/v)used as extraction solvent,extraction for 1.5 hour,liquid to material ratio 1:20.The total content of caffeoylquinic acids of Artemisia annua L.in different regions was between 0.4%and 5.8%.The maximum amount was 2%for 1,5-dicaffeoylquinic acid.The separation and purification of monocaffeoylquinic acids and dicaffeoylquinic acids in Artemisia anua L.residue were investigated.Static experiments showed that adsorption rate of polyamide was 89.4%,the adsorption capacity was 16.98 mg/g for dicaffeoylquinic acids.Polyamide is appropriate separation for dicaffeoylquinic acids in Artemisia annua L.The purity of dicaffeoylquinic acid had reached 45.7%and the yield was 66.8%.After the purification of polyamide,a column packed with SephadexLH-20 was used to perform the secondary purification.The purity of dicaffeoylquinic acids was 85.7%and the yield was 53.4%.The Aspergillus aculeatus and Aspergillus niger solid fermentation extracts were used to bioconvert chlorogenic acids.The Aspergillus aculeatus enzyme extracts could convert chlorogenic acids to 4-caffeoylquinic acid?4-ferulicquinic acid and 4-caffeoylquinic acid efficiently and specifically.The growth rate was 121.3%.50.4%and 47.3%,respectively.1,5-dicaffeoylquinic acid(87.9%)have untransformed and retained in solution.The conversion rate of 3,5-dicaffeoylquinic acid and 4,5-dicaffeoylquinic acid was 44.2%and 96.6%,respectively.The Aspergillus aculeatus enzyme extracts was able to convert 5-caffeoylquinic acid to caffeic acid.The conversion rate reached 100%in 0.5h.Aspergillus aculeatus enzyme could bioconvert chlorogenic acids to 4-caffeoylquinic acid,4-ferulicquinic acid and 4-caffeoylquinic acid.The growth rate was 131.4%,32.6%and 22.3%.The conversion rate of 1,5-dicaffeoylquinic acid 3,5-dicaffeoylquinic acid and 4,5-dicaffeoylquinic acid was 16.1%,42.8%and 25.3%respectively.The experimental result provided a potential new way for the low cost preparation of monocaffeoylquinic acids.