Extraction And Enrichment Of Main Active Components Of Dandelion And Preliminary Activity Evaluation

In this paper,a rapid and effective HPLC method for the determination of four target active components in dandelion leaves,namely chlorogenic acid,isochlorogenic acid,cichoric acid and luteolin,was established.Dandelion was extracted by ultrasonic assisted method with low co solvent,and then enriched by macroporous adsorption resin,and its antibacterial,antioxidant and anti-inflammatory activities were detected,Finally,the dandelion extract was made into a spray film agent and its antibacterial activity was studied.The main findings are as follows:1.A HPLC method was developed for the simultaneous detection of chlorogenic acid,isochlorogenic acid,chicoric acid,and luteolin in dandelion samples:Column:CuroSil-PFP 5u reversed-phase column(250 mm×4.6 mm id,5 μm,KYA);mobile phase:0.1%(v/v)formic acid aqueous solution(A)and acetonitrile(B);column temperature is 30℃;injection volume:20 μL;flow rate:1 mL/min;detection wavelength is 254 nm;gradient elution conditions:0-7 min,95-90%(A);7-25 min,90-75%(A);25-30 min,75-70%(A);30-37 min,70-60%(A);37-40 min,60-40%(A).The linear relationship,repeatability,precision,stability and recovery of chlorogenic acid,isochlorogenic acid,chicory acid and luteolin in the detection range all meet the experimental requirements.2.Single factor and BDD experiments were carried out to optimize the process parameters of ultrasonic assisted extraction of four active components from dandelion:the type and ratio of the deep eutectic solvents,the water content of the deep eutectic solvents,the extraction temperature,and the extraction time And liquid-solid ratio,the effects of chlorogenic acid,isochlorogenic acid,chicoric acid and luteolin in dandelion were determined experimentally,and the optimal extraction process parameters were determined:Deep eutectic solvents:Choline chloride:Ethylene glycol=1:1(molar ratio,water content 20%)Extraction temperature:29℃Extraction time:38 minLiquid-solid ratio:18 mL/gExtraction rates are obtained respectively:chlorogenic acid was 2.792 mg/g,isochlorogenic acid was 0.647 mg/g,chicoric acid was 9.824 mg/g,and luteolin was 0.281.mg/g.And the D101 resin,the best macroporous adsorption resin,was screened for the enrichment and separation of dandelion extract.The recoveries of chlorogenic acid,isochlorogenic acid,chicoric acid and luteolin were 78.23%,75.69%,84.15%and 73.42%,respectively,the enrichment contents were 5.74%,3.54%,9.87%and 2.72%,respectively.3.Perform in vitro antibacterial,anti-oxidant and anti-inflammatory activities on enriched and isolated dandelion extracts.Firstly,the antibacterial effect of dandelion extract on S.aureus,C.albicans and E.coli was determined.The data of the antibacterial experiment of dandelion extract on S.aureus and C.albicans showed that the diameter of the antibacterial circle of dandelion extract on S.aureus and C.albicans were 6.7 mm and 7.1 mm respectively Mm,MIC value of dandelion extract to S.aureus and C.albicans were 34.3075 mg/ml and 68.615 mg/ml respectively,MBC value of dandelion extract to S.aureus and C.albicans were 137.23 mg/ml and 137.23 mg/ml respectively,and no significant antibacterial effect on E.coli was detected;The DPPH scavenging ability of dandelion extract was determined.The IC50 value of dandelion extract was 0.661 mg/ml,which proved that dandelion extract had certain antioxidant activity.Finally,the anti-inflammatory effect of dandelion extract on RAW 264.7 macrophages was determined by measuring the anti-inflammatory effect of NO in LPS-induced RAW 264.7 macrophages.The dandelion extract can effectively inhibit the secretion of NO at a concentration of 12.5-50 ng/mL and has an anti-inflammatory effect.Later,the dandelion extract was made into a spray film-forming agent product,and its antibacterial effect was evaluated.C.albicans can be completely killed at 40 minutes,and S.aureus can be completely killed at 60 minutes.It has no obvious inhibitory effect on E.coli within 60 minutes.In this paper,the deep eutectic solvents-assisted extraction of active ingredients in dandelion can effectively improve yield,reduce costs,and ensure green pollution-free.After that,the extract was enriched and separated to obtain high activity of dandelion extract,and the natural green was preliminarily developed to make dandelion extract film spraying agentIt provides an important application basis for the reasonable development of dandelion resources.