As saponins occur as complex mixtures their isolation in a state of purity is somewhat tedious. In general, various chromatographic techniques are employed for their isolation and acetylation renders them crystallizable. Different extraction procedures are followed depending on the nature of saponin, amount present, and nature of the plant tissue. For the isolation of steroidal sapogenins, fresh plant material is usually used, though it is not uncommon to extract them from dried-size reduced plant material.

Ideally, fresh plant tissues are homogenized in aqueous solution and set aside for the autolysis of the glycosides through the action of endogenous glyosidase enzymes present in plant tissues. This also achieves cyclization of open-chain saponins in addition to cleaving off the sugar attached, yielding the sapogenin. This is a standard approach employed in the commercial production of steroidal sapogenins. Several newer methods of separation such as high speed counter current chromatography coupled with evaporative light scattering detector are being employed for the isolation of spaonins.

Following is a general method that may be adapted for the isolation of steroidal sapogenins:

1. 1 kg of the fresh plant material sufficiently size reduced is extracted with 95% ethanol at 100°C for 12 hours.
2. Filter hot and evaporate the filtrate under vacuum to a pasty residue.
3. Add 300 ml 1M Ethanolic hydrochloric acid and heat under reflux for 2 hours.
4. Filter, cool, and add 400 ml ether. Treat the solution with equal volume of water and discard the aqueous layer. Some methods adapt chromatographic separation of the saponin on a column of silica gel at this stage.
5. Wash the solution with 5% sodium hydroxide solution and again with water. Reject the aqueous layers and evaporate the solution to dryness.
6. Hydrolyze the dried residue with 10% alcoholic potassium hydroxide solution for 30 minutes and cool the solution.
7. Extract with 4 × 25 ml ether and evaporate the ether layers.
8. The product may be purified by recrystallization from acetone.