Isolation of three antibacterial naphthoquinones from Plumbago indica roots and development of a validated quantitative HPLC analytical method.

Three naphthoquinones, plumbagin (1), 3,3'-biplumbagin (2) and elliptinone (3), isolated from Plumbago indica roots by antibacterial bioassay-guided isolation, were used as standard markers for quantitative determination. A reversed-phase HPLC method was established for the simultaneous determination of the naphthoquinones in P. indica root extracts. The method utilised a Phenomenex® ODS column (4.6 × 150 mm, 5 µm) at 25°C with a mixture of methanol and 5% aqueous acetic acid (80 : 20 v/v) as the mobile phase at a flow rate of 0.85 mL/min, and UV detection at 260 nm. The parameters of linearity, precision, accuracy specificity and sensitivity of the method were evaluated. The recovery of the method was 98.6-100.6% with good linearity (r (2 )≥ 0.9997) for all three naphthoquinones. A high degree of sensitivity, specificity as well as repeatability and reproducibility (R.S.D. values less than 5%) were also achieved.