May 23, 2023
Extraction of 1-HP derivatives and HPLC preparation
- Muhammad Zaka Asif1,
- Man Shah1,
- Yosef Smadi1
- 1Edison Lab, UGA
Protocol Citation: Muhammad Zaka Asif, Man Shah, Yosef Smadi 2023. Extraction of 1-HP derivatives and HPLC preparation. protocols.io https://dx.doi.org/10.17504/protocols.io.5jyl8j43dg2w/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: May 23, 2023
Last Modified: May 23, 2023
Protocol Integer ID: 82352
Abstract
This protocol describes day 6 of our workflow to grow worms in liquid culture to induce the production of natural products (in this case, 1-HP derivatives). In this protocol, we extract 1-HP and its derivatives and then prepare the sample for HPLC injection.
Remove the sample from the lyophilizer. It should appear to be a powder.
Resuspend in minimal methanol (enough to submerge the pellet)
Vortex the tube with methanol to aid in dissolution.
The HPLC injection sample caps out at 90 uL, so minimizing the solution volume here will minimize the total number of injections needed.
Transfer the solution to a 2 mL microcentrifuge tube. Centrifuge at 20,800 RCF for 30 minutes.
Transfer only the supernatant into HPLC tubes (leave out the solids if any at the bottom of the microcentrifuge tubes). Keep track of the volume so that the injection volume for HPLC will be accurate.
Place HPLC tubes in the correct loading sites of the machine and run the protocol.
While the HPLC is ongoing, collect fractions if desired (either manually or automatically). Once collected, fractions can be stored at -80˚C for NMR prep later.