SPDS SET 2: ASH-01

c) Procedure: 20 µ l of each individual standard solution, mixed standard solutions, sample

solutions were injected into two different HPLC’s [(i) LC2010A equipped with UV detector

and (ii) LC10ADVP equipped with PDA detector)] on different date and time and the chromatograms were recorded. Relative retention time of different withanolides were

calculated with respect to Withanolide A . .

4.1.5) Observation: Retention time of Withanoside IV is about 15.5 minutes, Withanoside V is about 19.7 minutes, Withaferin A is about 20.3 minutes, 12-Deoxy withastramonolide is

about 21.4 minutes, Withanolide A is about 22.2 minutes and Withanolide B is about 25.5

minutes (Refer chromatogram-1). In the sample chromatogram all the individual

withanolides are well resolved. (Refer chromatogram-2).

Chromatogram – 1: Standard mix chromatogram

Chromatogram – 2: Sample chromatogram

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3DJH RI