Chloropigments and Amino Acid Concentrations from MULVFS Sampler
version Nov 11, 2001
Cindy Lee
Pigment Analysis: Chloropigments and fucoxanthin samples were measured
using reverse-phase high pressure liquid chromatography (HPLC) (Mantoura and
Llewellyn, 1983; Bidigare et al., 1985) as described in Lee et al. (2000).
Chloropigments measured include chlorophyll a, pheophorbide a,
pyropheophorbide a, and pheophytin a. Monovinyl and divinyl chlorophylls
(Bidigare and Ondrusek, 1996) were not separated.
Filter plugs were sonciated in 100% acetone to extract pigments. Extracted
plugs were immediately freeze-dried and stored in a -20C freezer for amino
acid analyses. Acetone extracts of the plugs were filtered through 0.2-µm
Zetapor membrane filters and injected with 20% MilliQ water onto a 5-µm
Adsorbosphere C-18 column. Chloropigments were identified using a
fluorescence detector (excitation = 440nm, emission = 660nm), and
fucoxanthin was identified using an absorbance detector (= 446nm).
Chloropigment and fucoxanthin concentrations were determined by comparison
of sample peaks and pigment standards. Pigment standards were either
purchased (Turner Design chlorophyll a and Porphyrin Products pheophorbide
a) or prepared (pyropheophorbide a, pheophytin a, and fucoxanthin).
Pyropheophorbide a and pheophytin a were prepared as in King (1993), and
fucoxanthin was prepared from extracted Pheaodactylum tricornutum cultures.
Prepared standards were compared, using HPLC, to archived chloropigment
standards and a fucoxanthin standard from Horn Point Laboratories in
Maryland, USA.
Bidigare, R.R., Kennicutt, M.C., Brookes, J.M., 1985. Rapid determination
of chlorophylls and their degradation products by high performance liquid
chromatography. Limnology and Oceanography 30, 432-435.
Bidigare, R.R., Ondrusek, M.E., 1996. Spatial and temporal variability of
phytoplankton pigment distributions in the central equatorial Pacific
Ocean. Deep-Sea Research II 43, 809-834.
King, L., 1993. Chlorophyll diagenesis in the water column and sediments
of the Black Sea. Ph.D. Thesis, W.H.O.I./M.I.T., unpublished.
Lee, C., Wakeham, S.G., Hedges, J.I., 2000. Composition and flux of
particulate amino acids and chloropigments in equatorial Pacific seawater
and sediments. Deep-Sea Research 47, 1535-1568.
Mantoura, R.F.C., Llewellyn, C.A., 1983. The rapid determination of algal
chlorophyll and carotenoid pigments and their breakdown products in natural
waters by reverse-phase high performance liquid chromatography. Analytica
Chimica Acta 151, 297-314.