Sample Preparation for 2D-HPLC-MS of Aconitum pendulum

Begin sample preparation by weighing the aconitum pendulum bush, or APB. Using a one by 10, 000 sensitivity balance, weigh 0.25 grams of dry APB in a three milliliter micro centrifuge tube. Add 2.5 milliliters of methanol to it and then sonicate for 30 minutes at a power of 240 watts and a frequency of 40 kilohertz.

Centrifuge the sonicated sample at 1.2G for five minutes, and collect the supernatant. Filter it through a 0.22 micron membrane filter. Next, prepare the two-dimensional mobile phase by using acetyl nitrile as the organic phase or phase B, and ultra pure water containing 0.1%formic acid as the aqueous phase or phase A.After performing two-phase filtration through the 0.22 micron filter, subject the mobile phases to ultrasonic sonication at 40 kilohertz for 15 minutes.

Purge the replaced mobile phase to remove the bubbles. Select a C18 column as the 1D column and a hydrophilic column as the 2D column, and mount them to the instrument. To adjust the appropriate shunt ratio, connect the 2D-LC instrument outline through a T to the mass inlet and the other end of the T to a shunted line.

Adjust the flow rate to a suitable value to ensure the appropriate flow rate for the mass spectrum.