UA Biophysics:Protocols:Carotenoid Extration

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Carotenoid Extraction, old protocol

  • Pigments are extracted from pellet of planktonic
  • 50mg of the cell pellets are dissolved in 1ml of methanol vortexed for 30 seconds
  • Centrifuge at 10,000rpm for 10 min
  • The supernatant containing the pigments is shaken with 1.7M NaCl and ethyl acetate (3:1 v/v) and allow to stand for 10 min to allow a sufficient phase separation
  • The ethyl acetate layer is then carefully drawn from the top, dried with anhydrous Na2SO4, and decant into a clean glass tube
  • The solvent containing the pigment is remove with N2, and the residue dissolved in equal volumes of benzene and acetone
  • This solution was then cooled to -20°C to precipitate the phospholipids, dried with N2 again, and dissolved in benzene for absorption measurement
  • The content of carotenoids is quantified by absorption from 400 to 550nm

Carotenoid Extraction, New protocol

  • Pigments are extracted from pellet of planktonic
  • 50mg of the cell pellets are dissolved in 1ml of methanol vortexed for 5 minutes with glass beads
  • Centrifuge at 8500 rpm for 10 min
  • The supernatant containing the pigments is shaken with 1.7M NaCl and ethyl acetate (3:1 v/v) (first adding ethyl acetate) and the sample is centrifuge at 8500 rpm for 5 min
  • The upper (ethyl acetate) phase is is then carefully drawn from the top and the lower phase is reextracte by addition of 1.0 mL of ethyl acetate (perform in duplicate)
  • Both organic phases is combine, dried with anhydrous Na2SO4, and decant into a clean glass tube
  • The organic solvent containing the pigment is remove with N2 gaseous, and the residue dissolved in equal volumes of benzene and acetone
  • This solution was then cooled to -20°C to precipitate the phospholipids, dried with N2 again, and dissolved in benzene for absorption measurement
  • The content of carotenoids is quantified by absorption from 400 to 550nm


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