Page 3 - TraceMetals_seagrass_algae

L. Campanella et al. / Environmental Pollution 111 (2001) 117±126 119

activities, while the ®fth site was the main harbour of acidi®ed, was stored at 4C to be subsequently miner-
the island, contaminated by naval trac emissions and alized in a microwave oven and analysed for total metal
domestic waste discharge. The sampling stations were concentrations.
carefully chosen on the basis of their exposure to dif-
ferent sea currents. While sampling the biota, special 2.2. Analysis
attention was paid to selecting individuals belonging to
a narrow size range, always collected at the same depth The seagrass individuals from each sampling station
and distance from the shoreline. were pooled and ®ve subsamples of about 0.7 g (dry
wt.) were digested in a microwave oven after 12 h pre-
Posidonia oceanica and Padina pavonica were hand- mineralization with ultrapure concentrated HNO3. The
picked in the subtidal zone at a depth of about 2 m. In same procedure was adopted for the algal material.
each station at least ®ve plants and 10 algae were sam- Only mature leaves and thalli of comparable length
pled. Individuals abundantly covered by epiphyta were were selected for analysis. Molluscs were analysed indi-
rejected during sampling. The samples were washed with vidually after digestion in a microwave oven. All
seawater and transferred to polyethylene bags. In the microwave-assisted mineralizations were conducted
laboratory epiphyta and sediments were removed from with a mixture of concentrated HNO3/H2O2 (5+2 ml,
the samples with nylon brushes under tap water. Posi- ultrapure grade). Separate dry weight determination
donia individuals were dissected in di€erent compart- was performed on the di€erent biota by oven drying at
ments: whole leaves, leaf tips (last 5 cm), leaf basal tissue 105C until constant weight.
(®rst 5 cm) and rhizomes. Then plant and algal material
was rinsed with Milli-Q water very quickly (to minimize Water samples were preconcentrated before analy-
any possible metal loss) and frozen until analysis. sis with the ammonium 1-pyrrolidinedithiocarbamate
(APDC) complexation method (Bruland et al., 1979;
About 20 individuals of Patella caerulea and Monodonta IRSA, 1984). This procedure involves metal chelation
turbinata were handpicked in the tidal zone from each with APDC, double extraction into chloroform and
station according to their availability. Organisms were back-extraction into nitric acid. Since APDC complex-
allowed to depurate for 2 days in ®ltered seawater col- ates Cr(VI) but not Cr(III) (Mizuike, 1983), in the
lected at the same site. Then the soft tissues were sepa- determination of the soluble metal content only the ®rst
rated from the shells, rinsed with Milli-Q water and species, toxicologically more relevant, was measured. As
frozen pending analysis. regards total (dissolved+particulate) metal concentra-
tions the chromium (Cr) levels measured can be
Water samples were collected from a small boat with assumed as good estimates of the whole Cr content
sampling bottles of 0.5 litre at a depth of 2 m. Salinity (Cr(III)+Cr(VI)), owing to the oxidizing treatment
and temperature were recorded. The sampling bottles during the mineralization step.
were rinsed twice with seawater and used to collect two
samples of 1 litre each in polyethylene containers. Sampling Cadmium (Cd), Cr, copper (Cu), lead (Pb) and zinc
bottles and polyethylene containers were previously (Zn) analysis was performed by electrothermal atomic
cleaned by soaking with 10% HNO3 for 2 days and absorption spectrometry (ETAAS). Whenever high
rinsed with Milli-Q water till neutral pH. In the labora- levels occurred, Cu and Zn in the biota were determined
tory one sample from each site was ®ltered a few hours by ¯ame atomic absorption spectrometry (FAAS).
after being collected through an acid pre-cleaned 0.45- Table 1 shows the detection limits and the coecients of
mm membrane ®lter, acidi®ed, and stored at 4C for variation for the di€erent matrices.
soluble metal analysis. The second sample, previously

Table 1
Detection limits (DL) and coecients of variation (CV) for the analysis performed by electrothermal atomic absorption spectrometry (ETAAS)a

Metal Macrophytes Molluscs Seawater

DL (mg gÀ1 dry wt.) CV DL (mg gÀ1 dry wt.) CV DL (ng lÀ1) CV

Cd 0.05 2.7 0.06 3.0 18 3.4
3.8 15 3.0
Cr 0.04 3.6 0.05 4.8 38 3.8
Cu 0.11b 4.5 0.14b 2.8 25 2.5
5.8 50 5.9
Pb 0.07 3.5 0.09
Zn 0.14b 5.3 0.18b

a DL, detection limit (calculated on the basis of 20 determinations of the blanks as three times the standard deviation of the blank). CV, coe-
cient of variation (%) (relative to 15 determinations performed on the same sample).

b DL for the analysis performed by ¯ame atomic absorption spectrometry (FAAS) (mg gÀ1 dry wt.): Cu=1.00 (macrophytes) and 1.27 (molluscs);
Zn=1.50 (macrophytes) and 1.91 (molluscs).