| Determination of Berberine by HPLC
a. Acetronitrile (HPLC) b. Nanopure water c.SDS d.Phosphate acid(A.R)
HPLC (Shimadzu) Analytical balance (1/10000) Ultrasonic cleaner
0.45μm PTFE filter 50ml volumetric flask Screw cap vials
3) HPLC condition
Column: Hypersil ODS C18 5μm , 4.6×200mm
Detector: UV 282nm
Mobile Phase: A=water ( including 0.02ml/L phosphate acid and 0.2% SDS)
Flow Rate: 0.8ml/min
Accurately weigh 30mg extraction into 50ml volumetric flask. Add 0.1%mol/L Phosphate acid to volume and sonicate to mix, filter through a 0.45μm PTFE filter into a screw cap vial and proceed with the analysis.
Standard preparation /calibration
Prepare a standard solution (approximately 0.6mg/ml in 0.1%mol/L Phosphate acid) using a commercial reference standard of berberine，Sonicate for about 10 minutes and transfer to screw-cap vials and can be stored in the refrigeration.
Determine the peak areas of the berberine on the sample chromatogram and compare to the corresponding peaks in the chromatogram for the reference standard solution. Calculate the weight percent.