In Situ Primary Productivity Protocol
A Description of Actual Procedures Used on the 1992 EqPac Survey and
Time Series Cruises
Richard T. Barber
The conceptual basis for the anti-contamination procedures are, of course,
the classic paper by Fitzwater, Knauer and Martin (1982), but other
important comments on inhibition of phytoplankton are given in Chavez and
Barber (1987), Price et al. (1986), Williams and Robertson (1989)
and Marra and Heinemann (1987).
- I. C Solution
- A. Anhydrous crystalline sodium carbonate is added to stock carbonate
solution of 0.3 g carbonate in 1.0 liter of Nanopure water. The lot
number of the NEN (New England Nuclear) Na CO was
2653-074; the specific activity was 55.0 mCi/mmol. The designation number
- B. The solution is made up and stored in Teflon containers that are
cleaned as described below.
- C. The C solution is refrigerated, but allowed to come to room
temperature before addition to the seawater.
- D. The intended activity of the C solution is 100 µCi/ml;
however, in our procedure the activity added is measured for each profile,
so variations in the initial activity are not a problem.
- II. Cleaning
- A. New bottles and labware are cleaned as follows: soak in a 2 % Micro
solution for three days. Rinse three times with DI water, then soak
overnight in DI water. Rinse again, then soak for two days in 0.5N HCl
(Fisher trace metal grade). Rinse three times with Nanopure water.
- B. After each use, the incubation bottles are soaked in a 10 % acid
wash for 12 to 24 hours and then rinsed well (3 times) with Nanopure water.
- C. Teflon stock bottles are cleaned by performing Micro and DI soaks as
above. Then sequential three-day soakings are performed, filling first
with 6 N HCl for three days, then with 2 N HNO for three days, and
then with the cleanest available 0.5 N HNO for three days. Each
filling should be followed with Nanopure rinses. Oven dry on a plastic
tray at 65 C.
- D. Polyethylene or vinyl disposable gloves without talc are worn during
Rosette handling and all other procedures.
- III. Sampling
- A. Eight light depths are sampled down to the 0.1 % I light depth
with custom made Go-Flo bottles on a ``trace metal clean'' General
Oceanics rosette. The rosette was made by General Oceanics according to
Moss Landing Marine Laboratory's specification; it is called the EqPac
``clean rosette.'' The rosette was lowered on a Kevlar conducting
hydroline with non-metal sheaves and a dedicated winch.
- B. Samples were taken before dawn, usually at 3:00 to 4:00 am.
- C. The 280 ml polycarbonate bottles are rinsed three times with sample
water and filled. The water ``fell'' from the Go-Flo spout into the
bottle without the use of a ``filling tube''. (McCarthy et al and
Landry et al. always use a ``filling tube'' to reduce shear and
turbulence that hurts microflagellates and ciliates. To avoid potential
contamination, we do not use a ``filling tube.'')
- D. Inoculation of 100 µl of C solution is done in the
radioactivity van with an Eppendorf disposable tip dispenser.
- E. In addition to two bottles from each light depth a third bottle is
taken from surface (``100 % I '') and the 8 % I depth for the
determination of time zero particulate C counts. The 100 % and 8 %
samples are inoculated and immediately filtered and treated identically to
the incubated filters.
- IV. Incubation
- A. The two replicate bottles from each depth are placed in a nylon mesh
bag and closed with polyethylene cable ties.
- B. Each bag is attached to the polypropylene array line by ``tuna''
snaps that clip into rings that are spliced in the line at one-meter intervals.
- C. The array line has a 50-lb lead weight at the bottom and two floats
at the top.
- D. A 20 m tag line connects the array line to the array spar buoy which
has an aluminum radar reflector, 3M light reflector sheets, a reflective
International Orange flag, a Novatech VHF transmitter and a Novatech xenon
flasher. Batteries on the VHF and flasher are changed at each deployment.
- E. The array is picked up after about 24 hours or about 4 am. The
nylon bags are taken off and placed in a box as the array is recovered.
The recovery takes about 20 to 30 minutes after the spar buoy is caught
with grapnel hooks.
- V. Filtration
- A. Following retrieval of the nylon bags from the array line the bags
and bottles are taken to the radioactivity van. One ml is taken from the
100 % and 8 % I depth bottles and added to scintillation vials
containing 1 ml of beta phenethylamine then 10 ml of Ecolume is added.
The purpose of this procedure is to determine an added C activity.
- B. The samples are then filtered through Whatman GFF filters.
- C. The filters are placed in scintillation vials and 0.5 ml of 0.5
added. The acidified filters are left for 24 hours in the hood.
- D. 10 ml of Ecolume is then added and the vials capped and left for 24 hours.
- VI. Counting and Calculations
- A. The time zero, total activity and incubated samples are counted on
the liquid scintillation counter with a wide window.
- B. Carbon uptake for each light level is calculated as follows:
Carbon = ([DPM - DPM] 1.05 24000)/DPM time)
DPM = CPM/efficiency of filters
DPM = CPM/efficiency of filters
DPM = (CPM/efficiency tot) bottle volume
1.05 = factor for preferential uptake of C over C
24000 = weight in mg/m of the inorganic carbon in seawater
- C. Carbon uptake for the water column down to the 1.0 % and 0.1 % light
level is calculated using a trapezoidal integration. We note that each
P.I. in JGOFS uses a different integration scheme. This is an area where
some discussion might be useful.
- Chavez, F.P. and R.T. Barber (1987).
- An estimate of new production
in the equatorial Pacific. Deep-Sea Research, 34: 1229--1243.
- Fitzwater, S.E., G.A. Knauer and J.H. Martin (1982).
contamination and its effects on primary production measurements.
Limnology and Oceanography, 27: 544--551.
- Marra, J. and K. R. Heinemann (1987).
- Primary production in the
north Pacific central gyre. Deep-Sea Research, 43: 1821--1829.
- Price, N. M., P. J. Harrison, M. R. Landry, F. Azam and K. J. F.
- Toxic effect of latex and Tygon tubing on phytoplankton,
zooplankton and bacteria. Marine Ecology: Progress Series, 34: 41.
- Williams, P. J. LeB. and N. I. Robertson (1989).
- A serious
inhibition problem from a Niskin sampler during plankton productivity
studies. Limnology and Oceanography, 34: 1300--1304.