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@#An analytical method was developed for the determination of five carbohydrate impurities in amino acid drug substances by high performance liquid chromatography-evaporative light scattering detection (HPLC-ELSD). Sugar impurities in the amino acid sample were separated and enriched by cation exchange resin. A Lichropher NH2 column (4.6 mm × 250 mm, 5 μm) was used for chromatographic separation, and a gradient elution was performed using acetonitrile-water as mobile phase. The drift tube temperature was 40 oC, the gain value was 8, and nitrogen (350 kPa) was auxiliary gas. Method validation results showed that the limits of detection for fructose, glucose, sucrose, maltose and lactose were in the range of 20.8-75.0 mg/kg and that the limits of quantitation were in the range of 96.2-238.8 mg/kg. Good linear relationship (r ≥ 0.999) were in the linear range for the five sugars, and the recoveries ranged from 84.9%-107.8%. With easy operation, high sensitivity, good precision and reliable accuracy, the method can be used for analysis of residual sugar impurities in amino acid drug bulk drug.
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OBJECTIVE:To optimize the extraction technology for flavonoids in leaves and seeds of Hippophae rhamnoides. METHODS:Using the total extraction rate of 6 flavonoid aglycones(catechins,rutin,myricetin,quercetin,kaempferol,isorham-netin)as index,ethanol volume fraction,extraction time,extraction times,material-lipid ratio as investigation indexes,L9(34)or-thogonal test was designed to optimize the extraction technology of flavonoids in leaves and seeds of H. rhamnoides,and verifica-tion test was carried out. RESULTS:The optimum extraction technology for flavonoids in leaves of H. rhamnoides was ethanol vol-ume fraction of 70%,extracting for 3 times with material-lipid ratio of 1:16,and 2.0 h each time;and that of seeds was ethanol volume fraction of 50%,extracting for 3 times with material-lipid ratio of 1:24,and 1.5 h each time. In verification test,the total extraction rate of 6 flavonoid aglycones was 56.4 mg/g in the leaves (RSD=1.4%,n=3) and 15.4 mg/g in the seeds (RSD=3.4%,n=3). CONCLUSIONS:Optimized extraction technology is simple,stable,feasible,and can be used for extracting the fla-vonoids in leaves and seeds of H. rhamnoides.
RESUMO
To establish a rapid determination method of six flavonoids:catechin,rutin,myricetin,quercetin,kaempferol and isorhamnetin,from seabuckthorn leaves by RP-HPLC-DAD.The seabuckthorn leaves were first degreased by petroleum ether,extracted by ethanol,and determined by RP-HPLC-DAD.The six flavonoids were separated and eluted by a Shimadzu C18(150 mm ×2.1 mm,5 μm) column with methanol-water (0.1％ phos phoric acid) (60∶ 40) at a flow rate of 1.0 mL/min.The detection wavelength were as follow:catechin 208 nm,rutin 257 nm,myricetin 373 nm,quercetin 371 nm,kaempferol 367 nm,and isorhamnetin 371 nm,respectively.The injection volume was 20 μL.The contents of the six flavonoids were in the range of 0.47 to 30.00 μ,g/mL with good linearity.The validation of the method,including precision,stability and recovery rate,was acceptable.The established method can be used for fast determination of the content of six flavonoids in seabuckthorn leaves.