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HPLC Phytoplankton Pigment Measurements: Continuation of Climate Quality Data Records (CQDR) in Support of NASA’s Earth Science Enterprise (ESE) Chuck Trees and Jason Perl Center for Hydro-Optics and Remote Sensing (CHORS) San Diego State University (ctrees@chors.sdsu.edu)
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HPLC Phytoplankton Pigment Measurements: Continuation of Climate Quality Data Records (CQDR) in Support of NASA’s Earth Science Enterprise (ESE) Chuck Trees and Jason Perl Center for Hydro-Optics and Remote Sensing (CHORS) San Diego State University (ctrees@chors.sdsu.edu) (jperl@chors.sdsu.edu)
HPLC Phytoplankton Pigment Protocols (Bidigare et al., 2003; NASA/TM-2003-211621/Rev 4 - Vol. V) EOS Terra & Aqua Data Products Program - 3,000 samples/yr Augmentation for an additional 1,800/yr= (4,800/yr) 1. Van Heukelem and Thomas (2001) Filtration 1. GF/F glass fiber filters (25 mm) 2. Positive pressure filtration 3. Filtration Volumes a. Oligotrophic waters - 3-4 liters b. Mesotrophic waters - 1-2 liters c. Eutrophic waters - 0.5-1 liter
HPLC Phytoplankton Pigment Protocols Sample Handling, Storage and Shipping 1. Fold filters in half with filtered sides facing in. 2. Place in aluminum foil envelops, cryogenic vials or HistoPrep tissue capsules. 3. Label samples containers with a sequential number 4. Place and ship (dry shipper) in liquid nitrogen (LN2). Recordkeeping (electronically transmitted to CHORS). 1. For each sample record volume filtered, station number, date, time, depth, lat and long.
HPLC System Calibrations 145 pigment compound calibration curves with 6-10 injections per curve (870-1,450 std injections or 24-40 days of continuous running standards).
Fluorometer Calibration
Number of Samples by Date and PI.
Vitamin E Internal Standard (284 nm) trans--apo-8'-carotenal (Fluka Chemical Company) Problems - Vitamin E Internal Standard Vitamin E Monovinyl Chl a Divinyl Chl a Divinyl Chl b Zeaxanthin b,b Carotene
Mixing the sample with the buffer. Heukelem and Thomas (2001) method picks up in a ‘train’ (150-75-75-75-150). It is mixed when injected onto the column. CHORS HPLC System mixes in a 2nd vial. Minimized the mixing time, but the uncertainty increased 3-4%. 2. UV/VIS halogen lamp. Faulty connection at the base of the lamp which would cause erratic readings. Solution was to have several lamps in stock to test when the lamp failed. Other Problems with HPLC Method and System
Culture Mixture (450 nm) Mixture of Pigments from Cultures Fucoxanthin Monovinyl Chl a Chl c3 Injection Spike Diadinoxanthin 19’-Hexanoyloxyfucoxanthin Monovinyl Chl b Peridinin Chl c2 Prasinoxanthin b.b carotene Zeaxanthin Monovinyl Chls a’, a” Alloxanthin Diatoxanthin b.e carotene
AGU/ASLO/TOS Ocean Sciences Meeting (Honolulu, 20-24 Feb 06) “To Separate or Not to Separate? That is the HPLC Question: A comparison between C18 and C8 methods” J. Perl, Q. Williams, and C. Trees “Particulate absorption: a pigment reconstruction analysis” Q. Williams, J. Perl, and C. Trees