Preparation Technology And Quality Standard Of Total Flavonoids From Juniperus Sabina L.

Juniperus sabina L.belongs to Cupressaceae Juniperus plants,which is its branches,leaves and fruits and is used to treat rheumatic autoimmune disease.With tha study object of total flavonoids from Juniperus sabina L.,this paper studies that the extraction and purification process of total flavonoids from Juniperus sabina L.were optimized by single factor tests and orthogonal experiment design;identification flavonoids were separated from total flavonoids by high-speed counter-current chromatography(HSCCC),and their structures were identified by high performance liquid chromatography(HPLC)and nuclear magnetic resonance(NMR);on the base of the studies of preparation technology and chemical components,the quality standard of total flavonoids from Juniperus sabina L.was researched and drafted out.The conclusions were as follows:(1)The determination method of three characteric ingredients in Juniperus sabina L.leaves was established by HPLC,and chromatographic conditions were showed as follows: Phenomenex Gemini-NX C18 as chromatographic column(250mm×4.6mm,5?m),the acetonitrile-0.2% phosphoric acid as the mobile phase(0 min~2 min~20 min~25 min,19%A~20%A~ 24%A~19%A),the temperature was 30?,the flow rate was 1.0 m L/min,and detection wavelength was 360 nm.Rutin,isoquercitrin and quercitrin were effectively separated with content determinations of stability.The three kinds of flavonoid glycosides contents from Juniperus sabina L.leaves were different.This method can provide base in order to effective substance development of Juniperus sabina L.(2)The optimum technological conditions for extracting total flavonoids from J.sabina(JSTF)were obtained as follows: ethanol concentration 50%,solid-to-liquid ratio 1:25,extraction time 1.5h and extraction times 3.Under these conditions,the extraction yield of JSTF was 7.47%.The optimal parameters for purified JSTF by D101 macroporous resin were as follows: the sample concentration of 1.2256 mg/m L,the sample flow rate of 1.0 m L/min,5 BV water consumption on removing impurity,eluting at 50% ethanol with 4 BV,desorption rate at 1.0 mL/min.Under these conditions,JSTF had a recovery rate of 88.36 % and a purity of 69.96 %.The experimental results of pilot amplification showed that extraction and purification process was stable and feasible.(3)Three compounds were separated from JSTF by HSCCC including isoquercitrin(91.89%),quercitrin(98.45%),quercetin-3-O-(6?-O-acetyl)-?-Dglucopyranoside(95.35%).Therefore,this method was fast and convient,and can effectively separate flavonoids in Juniperus sabina L.with the purity of 90% above.(4)The quality evaluation method of JSTF has been eatablished by the study of physical and chemical identification,thin-layer chromatography examination and content determination according to Chinese Pharmacopoeia(2015).The quality control of JSTF was expressed as follows: moisture content less than 7.5%,burn residue less than 0.9%,heavy metal less than 15 parts per million and arsenic salt was under 0.5 parts per million;total flavonoid content in JSTF was above 50%;the contents of rutin,isoquercitrin and quercitrin were not less than 21.4 mg/g,6.8 mg/g,17.7 mg/g.The appearance and color of JSTF were not obviously affected by the influence factors experiments(high temperature,high humidity and strong light),at same time the contents of JSTF and identify component has no change and stable character.