HPLC/LC/MS Analysis
The extracted product was analyzed using an LC/MS technique performed on an HPLC system
with an Evaporative Light Scattering Detector (Waters 2420) with analog – digital convertor
(ADC), as from UV analysis the sample found to be UV inactive which is coupled to a mass
spectrometer (Agilent Technologies, 1200 Series) with a mass spectrum detector (MSD). The
mobile phase consisted of 0.10% formic acid in HPLC grade deionised water (A) (milli-q-water
(subjected to IR radiation under 3.5micron filter) and Methanol (B) taken in the stationary phase
of Atlantis dc 18 column (50 x 4.6mm - 5μm). The gradient program was as follows: 10% B to
95% B in 4 min, 95% B to 95% B in 1 min, 95% B to 10% B in 0.5 min followed by 10% B in
1.5 min at a flow rate of 1.2 mL min-1. The column oven temperature was kept at 40°C and the
injection volume was 2.0 μL. Product mass spectra were recorded in the range of m/z 150-1000.
The instrumental parameters were optimized before the run.