|Determination of catechine by HPLC
a.Methanol (HPLC) b.Acetic acid (A.R) c.DMF(A.R) d. Nanopure water
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 278nm
Mobile Phase: A=water B= DMF:MeOH:HAC(40:2:1)
Time（min） 0 2 10 12 22 26 27 30
%A 84 84 79 79 68 68 84 stop
%B 16 16 21 21 32 32 16
Flow Rate: 1.1ml/min
Accurately weigh 30mg extraction into 50ml volumetric flask. Add nanopoure water 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 an mixed standard solution (approximately 0.6mg/ml in water) using a commercial reference standard of catechine，Sonicate for about 10 minutes and transfer to screw-cap vials and can be stored in the refrigeration.