Alkaloid Extraction Protocol

One can find a number of methods of quinolizidine alkaloid extraction within existing literature. The most frequently applied methods are extraction by alcohols (50% methanol or 96% ethanol, or these alkaloids with a 1% addition of glacial acetic acid).

Wysocka and Przybyl presented an efficient extraction method of quinolizidine alkaloids245. The authors themselves developed and investigated this method, in which includes the following steps of extraction:

• maceration of the ground lupin seeds with 25% aq. KOH for about 36 hours in order to destroy the tissues and release the alkaloids from their salts;

• excess water present in the alkaline pulp is absorbed by diatomaceous earth;

• mass prepared in this way is poured into a linen sack and then placed in an extractor followed by elution with ethyl ether and then with methylene chloride;

• extracts are condensed to a volume of about 100 cm3 and the alkaloids are eluted with 2 N hydrochloric acid;

• in order to degrease, the acidic alkaloid solution is extracted with petroleum ether (bp 40-60 °C) until the fats have been removed from the organic layers;

• after degreasing, the aqueous solution is made alkaline with 50% aq. KOH and then eluted with ethyl ether and methylene chloride.

This method has since been modified for efficiency, resulting in the following steps:

1. Grinding of seeds

2. Degreasing of the ground seeds with petroleum ether

3. Drying of the seeds in the air

4. Maceration of the meal with 25% aq. KOH

5. Mixing of the macerated meal with diatomaceous earth

6. Elution of alkaloids with methylene chloride

7. Purification of the extract by filtration through a column with aluminium oxide (activity grade II)

8. Evaporation of the extract to a constant weight

9. Yellow solidifying oil.

This method is more sensitive than other established methods. The amount of alkaloids obtained through this method is considerably higher than that through alcohol extraction.

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