1. Household blender.
2. Lab mill.
3. Lyophilizer.
2.2 Extraction
of Carotenoids
1. Homogenizer (Ultraturrax model T-25 or similar).
2. Solvent for extraction: Acetone containing 0.1% butylated
hydroxytoluene (BHT).
3. Diethyl ether containing 6 ppm BHT (see Note 3).
4. NaCl solution (10% w/v).
5. Anhydrous Na 2 SO 4 solution (2% w/v).
6. Refrigerated centrifuge with rotor for 50 mL and 15 mL tubes.
7. Refrigerated centrifuge with rotor for microcentrifuge tubes
(1.5–2 mL) with a minimum centrifuge force of 12,000 Â g.
8. 15 mL and 50 mL centrifuge tubes (Falcon type or similar).
9. 1.5 mL microcentrifuge tubes (Eppendorf type or similar).
10. Separation funnel (1 L).
11. Vortex.
12. Anhydrous Na 2 SO 4 (solid).
13. Rotary evaporator.
14. Nitrogen (gas).
15. Acetone (HPLC grade).
16. Sonication bath.
17. Sample vials for HPLC (see Note 4).
2.3 Saponification
of Carotenoid Extract
1. 15 mL centrifuge tube (Falcon type or similar).
2. Diethyl ether (containing 6 ppm BHT).
3. KOH–methanol solution (10% w/v).
4. Vortex.
5. pH test strips.
6. Anhydrous Na 2 SO 4 (solid).
7. Nitrogen (gas).
8. Acetone (HPLC-grade).
2.4 HighPerformance Liquid
Chromatography
(HPLC)
1. HPLC equipment with a diode array detector (see Note 5). In
our case the HPLC system consisted of a Waters 2695 Alliance
chromatograph fitted with a Waters 2998 photodiode array
detector (DAD), and controlled with Empower2 software
(Waters Corporation, Milford Massachusetts, USA).
2. Solvent A: Acetone (HPLC-grade).
HPLC Analysis of Carotenoids
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