Study On Quality Evaluation Of Lonicerae Japonicae Flos Samples From Different Area And The High- Effective Extraction Of Its Organic Acid

Lonicerae japonicae Flos, a well-known traditional Chinese medicinal plant, is the dried buds of Lonicera japonica Tilwnb. It is a commonly used traditional Chinese herb listed in Chinese Pharmacopoeia. According to the pharmacological studies, Lonicerae japonicae Flos possessed anti-bacterial, anti-inflammatory, anti-viral, anti-endotoxin, blood fat reducing, antipyretic, etc. In China, it is involved in healthy tea and TCM prescriptions. The extraction of Lonicerae japonicae Flos was widely used in many Chinese medicine preparations and functional food additives such as Jin Yin Hua and Shuang Huang Lian oral liquid, Wang Lao Ji and He Qi Zheng herb teas, etc. The main active compounds in Lonicerae japonicae Flos were neochlorogenic acid (NA), chlorogenic acid (CA),3,5-dicaffeoylquinic acid(3.5-DCA),3,4-dicaffeoylquinic acid(3.4-DCA) and 4,3-dicaffeoylquinic acid(4.3-DCA). The content of these compounds is an important index for the quality analysis of Lonicerae japonicae Flos. In the present study, we investigated the quality evaluation of Lonicerae japonicae Flos and Lonicera hypoglauca from the three main distribution provinces (Hebei, Henan and Shandong). And then our study focused on extraction, and enrichment and separation of these five important organic acids from Lonicerae japonicae Flos, developed an easy analysis method of HPLC, and established an efficient extraction process.The main findings are as follows:1. A rapid analysis method of HPLC was established for determination of chlorogenic acid, caffeic acid,3,4-dicaffeoylquinic acid,3,5-dicaffeoylquinic acid and 4,5-dicaffeoylquinic acid.Column:Diamonsil C18 (250 mm×4.6 mm i.d.,5μm); mobile phase:0.1% (v/v) acetic acid water (A) and 0.1% formic acid in acetonitrile-methanol,3:1v/v (B); Gradient elution conditions:0-5 min,2-8% B; 5-15 min,8-12% B; 15-30 min,12-15% B; 30-35 min,15-21% B; 35-46 min,21-24% B; 46-52 min,24% B; 52-66 min,24-36% B; 66-70 min,36-100% B; 70-75 min,100% B. Flow rate:1mL/min; column temperature:30 ℃; detection wavelength:320 run; injection volume:5 μL.2. The quality evaluation study of Lonicerae japonicae Flos from different distribution provincesThe Shangdong (Pingyi) sample had higher total phenolic and flavonoid contents and remarkable good antioxidant activity. In the ABTS test, the sample from Hebei was 0.810± 0.053 mmol/g which was the highest. Compared with other sites samples, Shangdong (Pingyi) sample had the highest reducing activity with the IC50 value 0.127 ± 0.011. Moreover, the three principal organic acids of different Lonicerae japonicae Flos samples were quantified. Chlorogenic acid (CA) and 3,5-dicaffeoylquinic acid (3.5DCA) were the most abundant components in Lonicerae japonicae Flos, and Shangdong (Pingyi) sample had the highest content of them. Based on the above results, Hunan (Shaoyang) sample had the best quality.3. The optimal parameters of microwave assisted assisted deep eutectic solvent extraction process:Deep eutectic solvents:1,3-Butanediol/ChCl (6:1, mol/mol)The ratio of deep eutectic solvent to water:10%Microwave-assisted extraction method:extraction temperature 60 ℃ liquid/soid ratio 9:1 mL/g extraction time 20 min Extraction cycle:3 cyclesUnder the above optimized conditions, the extraction yield of chlorogenic acid, caffeic acid,3,4-dicaffeoylquinic acid,3,5-dicaffeoylquinic acid and 4,5-dicaffeoylquinic acid 26.07+ 1.25,0.148±0.007,0.930±0.018,23.67±1.03 and 8.85+0.38 mg/g, respectively. NKA-9 macroporous resin made the recovery yields of chlorogenic acid, caffeic acid,3,4-dicaffeoylquinic acid,3,5-dicaffeoylquinic acid and 4,5-dicaffeoylquinic acid reached 79.25%, 80.03%,85.96%,86.01% and 85.52%, respectively. Recovery rates were all about to 80%. It is proved that phenolic acid compounds could be efficiently recovered from DES extracts.