For bile acid extraction and HPLC analysis from fecal samples, acetonitrile is a suitable solvent for the mobile phase due to its effective separation capabilities. Alternatively, a combination of methanol with phosphate buffer can also be used for optimal bile acid separation.

To extract bile acids from human fecal samples for HPLC analysis, weigh about 100–200 mg of frozen feces into a centrifuge tube and add 1–2 mL of methanol:water (1:1, v/v) with 0.1% formic acid. Vortex the mixture for 2–5 minutes to homogenize, then sonicate it for 10–15 minutes to improve extraction efficiency. Centrifuge the sample at 12,000 g for 10–15 minutes at 4°C, and carefully collect the supernatant. Filter the supernatant through a 0.22 µm syringe filter and transfer it to an HPLC vial. Bile acids can be analyzed using a C18 reverse-phase HPLC column with a gradient mobile phase of water and acetonitrile, both acidified with 0.1% formic acid, and detected by UV (typically at 210 nm) or mass spectrometry for greater sensitivity.

Is it good to use a ready to use Bile acid standard mixture from Sigma for hplc from human faecal samples ?? And if used do we have to filter it and process before using or can it be used directly?