Multi-source water pollution in a highly anthropized wetland system associated with the estuary of Huelva (SW Spain)

Marine Pollution Bulletin 60 (2010) 1259–1269

Contents lists available at ScienceDirect

Marine Pollution Bulletin journal homepage: www.elsevier.com/locate/marpolbul

Multi-source water pollution in a highly anthropized wetland system associated with the estuary of Huelva (SW Spain) C. Barba-Brioso a,*, J.C. Fernández-Caliani b, A. Miras a, J. Cornejo c, E. Galán a a

Dpto. Cristalografía, Mineralogía y Química Agrícola, Facultad de Química, Universidad de Sevilla, 41072 Sevilla, Spain Dpto. Geología, Facultad de Ciencias Experimentales, Universidad de Huelva, 21071 Huelva, Spain c Instituto de Recursos Naturales y Agrobiología de Sevilla, Apdo. 1052, 41080 Sevilla, Spain b

a r t i c l e Keywords: Water pollution Heavy metals Agrochemicals Estuary Tidal marsh Huelva

i n f o

a b s t r a c t Major ions, nutrients, trace elements and pesticides distribution were studied in a coastal wetland heavily impacted by human development in Spain. Past land use has altered the local hydrodynamics leading to the partitioning of the ecosystem into a tideland subject to marine influence, and an artificial freshwater reservoir created by stream impoundment. The tideland stretch is flooded twice a day with a heavy metal plume that emerges from the mine-polluted estuary of Huelva and propagates landward depicting the same dispersal trend of major seawater ions. Additionally, the tidal channel receives acid discharges from industrial point sources that contribute to metal enhancement. The impounded area and stream tributaries are affected by agrochemicals runoff (nitrate, phosphate, pendimethalin, simazine, diuron and therbuthylazine) from surrounding agricultural lands. The tidal regime plays a crucial role in the transport and dispersion of pollutants, except in the artificial reservoir where freshwater exhibits a seasonal mineralization pattern. Ó 2010 Elsevier Ltd. All rights reserved.

1. Introduction The impact from human-induced activities is noticeable in most regions of the world, but it is particularly evident in estuaries and coastal wetlands subject to multiple environmental pressures, such as: water abstraction, mining, industrial and domestic effluents, altered hydrology, habitat degradation and overharvest of resources (Butler, 2006). The Huelva estuary, on the southwestern coast of Spain (Gulf of Cádiz), is considered one of the most polluted estuaries in Europe, as a result of the acid mine drainage and disposal of industrial processing wastes (Nelson and Lamothe, 1993; Fernández-Caliani et al., 1997; Davis et al., 2000; Borrego et al., 2002; Achterberg et al., 2003; Sáinz et al., 2004; Ruiz et al., 2008). This estuary receives very acidic (pH values below 4) and heavily polluted discharges from the Odiel and Tinto rivers, after draining a large number of mining-impacted sites of the Iberian Pyrite Belt (e.g. Sánchez España et al., 2005; Fernández-Caliani et al., 2009). The mean annual discharges range widely from 3 to 15 m3 s 1 depending on seasonal conditions (Braungardt et al., 2003), with a

* Correspondence to: C. Barba-Brioso, Dpto. Geología, Facultad de Ciencias Experimentales, Campus Del Carmen, Universidad de Huelva, Avda/3 de marzo s/n, 21071 Huelva, Spain. Tel.: +34 959 219 831/959 219 820/661 308 314; fax: +34 959 219 810. E-mail addresses: [email protected], [email protected] (C. Barba-Brioso). 0025-326X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.marpolbul.2010.03.018

dissolved load of 7900 ton of Fe, 3500 ton of Zn and 1700 ton of Cu (Olías et al., 2006). Additionally, large amounts of industrial pollutants have been discharged since the mid-1960s from a variety of chemical factories (copper smelter, paper mill factory, phosphate fertiliser plants, chloro-alkali industries, etc.) and oil refineries. Moreover, the introduction of strawberry cultivation in the late half of the 1970s has increased the impacts on soil and water quality, due to intensive use of agrochemicals in the vast plantations that stretch over tens of square kilometres around the estuary. In this paper, a strongly anthropized wetland area associated with the Huelva estuary, so-called Domingo Rubio, has been selected in order to assess the effects of multiple source pollution on water quality. The primary objectives were: (a) to determine the influence of tidal and seasonal regimes on concentration and distribution of pollutants in the wetland; (b) to detect hydrochemical anomalies representing anthropogenic inputs; and (c) to identify the various pollution sources by apportionment methods. 2. Study area Domingo Rubio is a tidal system located in the left bank of the Tinto river before its confluence with the Odiel river to form the Huelva estuary along the mesotidal coast of southwestern Spain (Fig. 1). The Huelva estuary is a well mixed, partially dredged, estuary with a tendency to become stratified during spring tides (Braungardt et al., 2003). The Domingo Rubio wetland is connected

1260

C. Barba-Brioso et al. / Marine Pollution Bulletin 60 (2010) 1259–1269

Fig. 1. Location map of the Domingo Rubio wetland area in the Huelva estuary, showing the sampling sites.

to this estuary through a tidal channel across a salt-marsh surface developed in a small flooded valley, which is incised into Plio-Pleistocene detritic deposits. This system is clearly dominated by tidal processes owing to the protection afforded by nearby coastal sand barriers against wave action and the restricted fluvial supply. The tidal regime is semidiurnal, the mean amplitude reaching up to 2 m during a spring-neap cycle (Pendón et al., 1998). A comprehensive description of the depositional environment and Holocene evolution of the estuarine facies can be found in Pendón et al. (op. cit.) and Ruiz et al. (2005). Local hydrodynamics has been significantly altered in the last decades as a result of road constructions that form hydrological barriers to flow, giving place to a freshwater shallow lake in the upper course of the tidal channel. Hence, two zones with contrasting hydrological and environmental features can be distinguished in the wetland area: (a) a tideland zone, comprising broad saltmarsh bodies crossed by a meandering tidal channel and covered by Salic Fluvisols with abundant halophytic vegetation (Barba-Brioso et al., 2007); and (b) a lacustrine zone, consisting of a freshwa-

ter reservoir artificially created by stream impoundment, due to the construction of roads, with aquatic and riparian vegetation. A dendritic network of ephemeral stream tributaries flow into both zones from a watershed affected by intensive irrigated cropping operations and industrial activities. The tidal and lacustrine areas are connected through a drainage pipe that drains overflow from the lagoon into the tideland during rainy periods. Considering its high ecological value, the Regional Government of Andalusia has declared the tideland zone as a Natural Park and a special protection zone for birds. Despite this legal protection, the Domingo Rubio wetland is threatened by a number of anthropogenic factors potentially affecting water quality (Table 1), such as industrial development (Martín et al., 2002), hazardous wastes disposal, agriculture expansion and excessive extraction of sands and gravels resources. Recent studies have documented trace element pollution in soils and plants, particularly in the tideland zone (Barba-Brioso et al., 2009; Madejón et al., 2009), and biological effects of pollutants using several molecular biomarkers and proteomic approaches (Montes-Nieto et al., 2007).

C. Barba-Brioso et al. / Marine Pollution Bulletin 60 (2010) 1259–1269 Table 1 Main anthropogenic activities, sources of pollution and pollutants in the Domingo Rubio wetland. Wetland area

Anthropogenic activities

Type of pollution

Pollution source

Main pollutant

Tideland area

Point

Tideland area

Mining and metallurgy Industry

Overall wetland

Agriculture

Point

Tideland area

Mining and industry Industry and harbour activities

Diffuse

Sulfidic wastes Industrial effluents Agricultural stream and pipes Huelva estuary Atmospheric particulate matter

Heavy metals Heavy metals Pesticides and fertilizers Heavy metals Heavy metals

Overall wetland

Point

Diffuse

3. Sampling and methods 3.1. Water sampling Several sampling campaigns were carried out to study the water quality variation in the area. The tideland zone was sampled during both high tide (9 samples) and low tide (12 samples) in order to consider the effects of tidal regime. These estuarine samples were collected in the upper layer of the water column (0–20 cm) at each sampling site, using a Zodiac boat for access. Site depth decreased from about 4 m (sample AE1) to less than 1 m in the most upstream site (sample AE17). In addition, to investigate seasonal variations of freshwater, eight surface (0–20 cm) water samples were taken in the lacustrine zone (three in summer and five in winter), and 12 samples (three in summer and nine in winter) were collected in running water from tributary streams and agricultural open-pipes. The sampling design was unbalanced due to field constraints. Overall, 41 water samples were collected at 26 different geopositioned sites (Fig. 1). In-situ measurements of pH, redox potential (Eh), electrical conductivity, dissolved oxygen concentration and temperature were made with portable Crison instruments. Water samples were taken following general procedures described in many specific documents (Fresenius et al., 1988; Rump, 1999; Salminen, 2005), filtered through a 0.45 lm pore diameter membrane, stored in polyethylene bottles previously rinsed three times with the same water to be sampled, and refrigerated immediately in a cooler. The water samples were stored at 4 °C in the dark until analysis, after acidulation with HNO3 (at pH < 2) for metal determination. The samples selected for pesticide recovery were collected one month after application and stored in topaz glass bottles. 3.2. Analytical methods 3.2.1. Anions and nutrients The concentrations of sulfate, nitrate, nitrite, chloride and bromide were determined by ion chromatography on a Dionex DX120 apparatus fitted with an IonPac AS9-HC of 4  250 mm separation column and 4-mm ASRS-ULTRA II suppressing membrane. Standards for external calibration were prepared from certified referenced solutions Merck 1000 ppm for chromatography. Detection limits were 0.1 mg L 1 for bromide and nitrite, 0.3 mg L 1 for nitrate, and 0.5 mg L 1 for sulfate and chloride. Bicarbonate determination was done by the method 2320 B of the Standard Methods for the Examination of Water and Wastewater (Eaton et al., 1998), consisting in a volumetric titration with previously standardized 0.01 N HCl, using bromocresol green as indicator. Each sample was measured in triplicate and the mean was taken as the representative value, achieving a precision of ±0.5%. Phosphate

1261

concentration was measured by colorimetry in a Shimadzu UV– Visible spectrophotometer, model UV mini-1240, after formation of antimony–phospho-molybdate complex reduced to an intensely blue-coloured complex by ascorbic acid (EPA method 365.4). The instrumental detection limit (0.03 mg L 1) was calculated as three times the standard deviation of seven blank values (USEPA, 1984; Miller and Miller, 2000). 3.2.2. Major and trace elements The concentrations of Ca and Mg were determined by atomic absorption spectroscopy (AAS), after addition of La2O3 to the samples to prevent any chemical interference, while the Na and K determinations were done by atomic emission spectroscopy (AES), by adding LiCl to avoid partial ionization (Fresenius et al., 1988). The analyses were conducted in a Perkin Elmer 2380 spectrophotometer. The system was calibrated by the measure of single standards of the elements containing 1, 3, 5, 10, 20 and 50 ppm, prepared from single certified standard solutions (Merck VI for ICP-MS) of each one of the analyzed elements. Dissolved trace element analyses (As, Cd, Co, Cr, Cu, Ni, Pb and Zn) were conducted by inductively coupled plasma-mass spectrometry (ICP-MS) in a Hewlett Packard 4500 instrument. The multi-element calibration standards were prepared with 0, 5, 10, 25, 50, 100 and 200 ppb from a multi-element certified standard solution (Merck VI for ICP-MS) containing 1000 ppm of all studied elements, except As which used individual solutions. Certified reference material SRM-1640 NIST was also measured to correct for any drift during the analysis. Multi-element reference standards were run after nine samples. The mean precision and the accuracy fall within the range of 3–10% for all the analysis. The detection limits of the technique are optimized to 0.01 ppb for all the analyzed trace elements, but they were also statistically calculated for each element, obtaining values lower than 2 ppb in all cases. 3.2.3. Pesticides Solid-phase extraction (SPE) procedure and liquid chromatographic techniques (HPLC) were applied to quantify all the pesticides studied (atrazine, simazine, terbuthylazine, deethylsimazine, deethylterbuthylazine, diuron, napropamide and pendimethalin). Automated SPE was performed with the Visiprep SPE Vacuum Manifold (SUPELCO, Bellefonte, PA) apparatus. For triazines and diuron the cartridges used (Supelclean ENVI-18 SPE Tubes, SUPELCO) contain 500 mg of C18 bonded silica material and with a capacity of 6 ml. Four ml of methanol and 4 ml of water were used for conditioning the cartridge. Four replicates of 250 ml of water sample were passed through the corresponding cartridges at a flow-rate of 2.5 ml min 1. Water residues from cartridges were removed by 10 min under a flow of nitrogen. Elution was carried out with 4 ml of ethyl acetate at a flow-rate of 1 ml min 1. For napropamide and pendimenthalin the cartridges used (Bond Elut Plexa Tubes, VARIAN) contain 60 mg of C18 bonded silica material and with a capacity of 3 ml. One ml of methanol and 1 ml of water were used for conditioning the cartridge. Four replicates of 250 ml of water sample were passed through the corresponding cartridges at a flow-rate of 4 ml min 1. Water residues from cartridges were removed by 10 min under a flow of nitrogen. Elution was carried out with 2 ml of acetone at a flow-rate of 1 ml min 1. For all samples extracted evaporation of the solvent was performed also under a stream of nitrogen until dryness. The final extract was recovered with 0.25 ml of methanol and analyzed by high performance liquid chromatography (HPLC). The eluent was delivered by a gradient system from a Waters 600 pump associated to a 717 Automatical injector and a 996 Photo diode array detector (Waters, Milford, MA). A Waters Nova-Pak C18 cartridge column (3.9  150 mm) was used. The gradient elution was carried out with a binary gradient composed by

1262 Table 2 Basic descriptive statistics of hydrochemical parameters, major cations, anions and dissolved trace element concentrations in waters of the Domingo Rubio wetland, arranged according to tidal and seasonal conditions prevailing during sampling. pH

high tide 9 9 7.8 386 7.8 382 7.7 369 8.0 425 0.1 19

Tidal channel low tide Valid N 12 12 Mean 7.2 375 Median 7.6 358 Minimum 5.3 334 Maximum 7.8 439 Std. Dev. 0.9 38

F

Cl

NO2

Br

NO3

SO42

HCO3

Ca2+

Mg2+

Na+

K+

DO (%)

EC (mS/ cm)

9 103 113 82 123 16

9 32.7 39.2 3.0 47.5 15.7

9 0.0 0.0 0.0 0.3 0.1

9 10414 12351 614 16390 5469

9 0.0 0.0 0.0 0.0 0.0

9 36.2 43.0 3.3 56.4 18.6

9 1.6 0.0 0.0 6.4 2.5

9 1502 1759 158 2426 757

9 196 209 115 248 40.9

9 1.7 1.5 0.7 4.1 1.0

9 240 266 56.0 332 90.4

9 666 760 49.0 1020 320

9 6333 7200 326 9725 3092

9 243 301 29.5 368 115

0

12 8.1 3.8 0.9 28.5 9.0

12 0.0 0.0 0.0 0.2 0.1

12 2781 1134 103 10054 3221

12 0.0 0.0 0.0 0.0 0.0

12 10.4 4.8 1.4 34.7 11.2

12 8.8 6.3 0.0 25.0 7.4

12 520 240 90.2 1843 567

12 110 116 16.0 172 42

12 0.3 0.3 0.1 0.5 0.1

12 95.8 43.8 18.8 496 132

12 196 67.5 13.8 750 248

12 1568 608 94.6 5845 1936

PO34

Cr

Mn

Fe

Co

Ni

Cu

Zn

As

9 46.8 50.7 2.4 77.1 20.9

9 232 197 161 506 109

9 745 728 326 1170 252

9 2.7 1.9 1.0 7.7 2.0

12 29.6 12.1 3.7 107 34.6

12 16.7 17.0 3.5 30.6 7.3

12 291 189 47.4 882 241

12 590 126 59.5 3433 1018

Cd

Pb

9 7.3 6.6 5.4 10.7 2.0

9 576 590 36.0 1239 353

9 73.1 67.0 27.0 113 25.8

9 74.1 78.9 10.3 124 36.1

9 2.0 1.9 0.3 4.1 1.4

9 0.2 0.2 0.2 0.2 0.0

12 4.2 0.2 0.2 46.0 13.2

12 10.6 8.1 4.6 32.4 7.6

12 352 88.7 20.9 1715 557

12 256 18.9 0.7 1570 488

12 19.3 19.0 4.3 39.9 9.8

12 11.6 11.8 1.0 23.3 6.1

12 3.1 0.2 0.2 11.9 4.5

(lg/L)

(mg/L)

Lagoon summer Valid N 3 Mean 8.4 Median 8.4 Minimum 8.3 Maximum 8.5 Std. Dev. 0.1

3 318 314 311 330 10

3 103 104 72 134 31

3 1.2 1.2 1.2 1.3 0.1

3 0.2 0.2 0.1 0.2 0.0

3 163 162 150 178 13.7

3 0.0 0.0 0.0 0.0 0.0

3 1.7 1.7 1.6 1.7 0.0

3 11.8 7.1 0.7 27.8 14.2

3 102 101 96.8 109 6.1

3 224 230 209 233 12.9

3 0.5 0.4 0.4 0.8 0.2

3 50.7 50.5 50.0 51.5 0.8

3 22.5 22.0 22.0 23.5 0.9

3 90.0 86.9 83.6 99.6 8.4

3 18.1 18.0 17.7 18.7 0.5

3 0.6 0.5 0.2 1.0 0.4

3 209 204 179 246 33.9

3 1338 1303 1211 1499 147

3 0.2 0.2 0.2 0.3 0.0

3 5.1 4.9 4.6 6.0 0.8

3 14.4 11.7 10.1 21.3 6.1

3 15.3 10.0 10.0 26.0 9.2

3 6.9 6.7 6.4 7.7 0.7

3 0.3 0.3 0.1 0.7 0.3

3 1.1 0.4 0.3 2.5 1.3

Lagoon winter Valid N 5 Mean 7.5 Median 7.5 Minimum 7.4 Maximum 7.7 Std. Dev. 0.1

5 350 375 225 398 72

0

5 0.7 0.7 0.6 0.8 0.0

5 0.1 0.0 0.0 0.2 0.1

5 116 111 106 131 10.0

5 0.1 0.0 0.0 0.2 0.1

5 1.1 1.1 1.0 1.3 0.1

5 6.4 6.6 5.4 6.9 0.6

5 90.4 92.8 70.2 111 15.8

5 136 137 110 160 18.0

5 0.1 0.1 0.0 0.2 0.1

5 35.3 33.8 26.3 41.3 6.3

5 18.3 18.8 13.8 21.3 3.3

5 65.5 64.9 51.9 77.1 9.1

5 9.1 9.6 6.7 10.4 1.5

5 2.2 1.9 1.6 3.0 0.6

5 102 118 53.4 133 31.5

5 195 128 75.5 366 128

5 0.2 0.2 0.2 0.2 0.0

5 3.4 2.7 2.1 5.5 1.5

5 17.4 19.7 7.2 27.7 7.8

5 1.3 0.7 0.7 3.2 1.1

5 2.2 1.9 1.7 2.9 0.6

5 1.0 1.0 1.0 1.1 0.1

5 0.2 0.2 0.2 0.2 0.0

Tributaries summer Valid N 3 3 Mean 7.5 327 Median 7.7 325 Minimum 7.2 310 Maximum 7.8 347 Std. Dev. 0.3 19

3 70 68 60 82 11

3 1.1 1.1 0.9 1.3 0.2

3 0.2 0.2 0.1 0.3 0.1

3 120 123 101 137 18.5

3 0.9 0.5 0.3 1.8 0.8

3 1.3 1.1 1.1 1.8 0.4

3 46.0 34.7 22.1 81.2 31.1

3 96.7 74.9 66.1 149 45.5

3 199 208 126 263 68.6

3 5.0 3.1 0.7 11.3 5.6

3 43.0 33.0 19.5 76.5 29.8

3 18.5 19.5 15.5 20.5 2.6

3 79.4 83.4 65.4 89.4 12.5

3 21.0 18.8 16.5 27.7 5.9

3 0.7 0.7 0.2 1.2 0.5

3 89.9 121 1.3 148 77.9

3 613 664 176 999 414

3 4.1 2.9 1.1 8.4 3.8

3 12.4 7.5 4.0 25.8 11.7

3 21.0 17.7 14.3 31.1 8.9

3 11.3 12.0 10.0 12.0 1.2

3 10.7 11.5 6.3 14.3 4.0

3 0.6 0.7 0.3 0.8 0.3

3 0.2 0.2 0.2 0.2 0.0

Tributaries winter Valid N 9 Mean 6.9 Median 7.7 Minimum 3.8 Maximum 8.0 Std. Dev. 1.7

4 68 68 65 70 2

9 1.0 1.1 0.4 2.3 0.6

9 0.1 0.0 0.0 0.2 0.1

9 86.4 86.8 10.4 166 50.9

9 0.2 0.0 0.0 1.3 0.4

9 1.1 1.2 0.0 3.0 1.0

9 39.6 24.8 4.9 96.8 35.8

9 209 108 21.5 758 242

9 126 95.9 1.0 458 143

9 3.9 0.3 0.1 18.5 6.6

9 42.3 38.8 18.8 67.5 18.1

9 25.2 20.0 13.8 65.0 16.0

9 66.9 74.4 11.3 95.4 28.8

9 13.5 13.7 4.9 24.3 6.7

9 4.1 1.9 1.5 19.9 6.0

9 462 182 27.4 1539 609

9 465 226 120 1722 509

9 43.6 3.9 0.2 267 89.9

9 124 7.9 2.0 771 262

9 23559 19.3 16.6 168000 56067

9 5223 27.2 5.6 33600 11426

9 33.4 13.5 2.2 219 69.9

9 807 1.1 1.0 4371 1642

9 32.8 0.2 0.1 198 66.5

9 389 353 299 585 106

C. Barba-Brioso et al. / Marine Pollution Bulletin 60 (2010) 1259–1269

Tidal channel Valid N Mean Median Minimum Maximum Std. Dev.

Eh (mV)

1263

C. Barba-Brioso et al. / Marine Pollution Bulletin 60 (2010) 1259–1269

a

9.0

b 50

8.0

40

pH

EC

7.0 6.0

30

20

4.0

10 -

5.0

3.0

0 0

2

4

6

8

10

12

14

0

c

18000

3000

2

4

6

8

10

12

14

d

16000

2500 14000

2000

12000

Cl

-

SO4 =

10000 8000

1500 1000

6000 4000

500 2000

0

0 0

2

4

6

8

10

12

0

14

e

60

40

200

6

8

2

4

6

10

12

14

HCO3

Br

-

250

4

f

300

50

2

30

150

20

100

10

50

0

0

0 4.5

2

4

6

8

10

12

14

0 30

g

4

8

10

12

14

h

25 3.5

20 -

2.5

NO3

PO4 3-

3

2

15 10

1.5 1

5

0.5

0

0 0

2

4

6

8

10

12

0

14

2

Distance (Km)

4

6

8

10

12

14

Distance (Km)

high tide

low tide

Fig. 2. Spatial distribution of master variables across the Domingo Rubio wetland. Distances are expressed in km from the tidal channel mouth. EC in mS cm concentrations in mg L 1, and trace element concentrations in lg L 1.

acetonitrile–water. The initial mobile phase was 85–15 water–acetonitrile. During the 40 min run of each sample, the ratio of sol-

1

, ionic

vents was changed with a linear gradient to 100% acetonitrile. The flow-rate was 1 ml min 1, injection volume was 25 ll and

1264

C. Barba-Brioso et al. / Marine Pollution Bulletin 60 (2010) 1259–1269

2000

10000

i

1800

1600

1600

1000

1400

1400 1200

1200

1000

Cu

100

1000

Zn

Fe

k

2000

j

1800

800

800

10

600

600 400

400

1

200

200 0

0

0.1 0

2

4

6

8

10

12

14

0

l

140

30

120

2

4

6

8

10

12

0

14

2

4

6

8

10

12

14

m

25

100

20

High tide

As

Cd

80 15

60

Low tide 10

40

5

20

0

0

0 100

2

4

6

8

10

12

0

14 1000

n

4

6

8

10

12

14 90

o

900

10

2

70

700

60 50 Cr

Mn

Co

600 500 400 1

200

20

100

10 0

0 0

2

4

6

8

10

12

14

40 30

300

0.1

p

80

800

0

2

4

Distance (Km)

6 8 10 Distance (Km)

12

14

0

2

4

6

8

10

12

14

Distance (Km)

Fig. 2 (continued)

the analysis was carried out at a wavelength of 225 nm for triazines and 250 nm for diuron. For napropamide and pendimenthalin the eluent was delivered by a isocratic system from a Waters 600 pump associated to a 717 Automatical injector and a 996 Photo diode array detector (Waters, Milford, MA). A Waters Nova-Pak C18 cartridge column (3.9  150 mm) was used. The isocratic elution was carried out with a mobile phase of 70% acetonitrile and 30% water. The flow-rate was 1 ml min 1, injection volume was 25 ll and the analysis was carried out at a wavelength of 240 nm for both pesticides. The retention times were 2.3 and 6.8 min for napropamide and pendimethalin, respectively. Quantification was made by external calibration using over 99% purity standards purchased from Dr. Ehrenstorfer GmbH (Augsburg, Germany). Identification was based on chromatographic retention times. Confirmation of peak identity was achieved checking diode array spectra, as appropriate and according to instrument availability (Aguer et al., 2000; Cabrera et al., 2007).

tive statistics, normality tests (Kolmogorov–Smirnov and Shapiro– Wilk) and hierarchical cluster analysis. A statistic technique based on Mahalanobis distance was applied for detection of extremes, which were depicted on a box and whisker diagram. A Spearman correlation matrix was used to determine the relationships among the analyzed variables. Because of the requirements of the parametric analyses were not accomplished, according to the normality tests, a cluster analysis using average linkage was done to identify different geochemical associations, as shown by a dendrogram. 4. Results A summary and comparison of water quality parameters and chemical composition of the samples in terms of basic descriptive statistics is given in Table 2, and the spatial distribution of master variables (hydrochemical parameters, major cations, anions, nutrients and trace elements) in the tidal area is depicted in Fig. 2. 4.1. Hydrochemical parameters

3.3. Statistical analysis The obtained experimental results were treated with the software StatSoft Statistica 7 in order to recognize the temporal and spatial trends in the water quality parameters of the Domingo Rubio wetland. Five variables (Pb, Co, NO2 , F and dissolved O2) were excluded due to insufficient data. Statistics analysis included descrip-

The estuarine waters of the tidal channel are slightly alkaline (Fig. 2a), with pH mean values ranging from 7.2 (low tide) to 7.8 (high tide). However, acidic values were also recorded locally at the mouth of the tidal channel under low tide conditions (pH 5.3), and in some tributaries receiving chemical effluents and leachates from abandoned sulfide wastes (pH 3.8–4.1). In the

1265

C. Barba-Brioso et al. / Marine Pollution Bulletin 60 (2010) 1259–1269

1400

tidal channel high tide tidal channel low tide tributaries summer tributaries winter lagoon summer lagoon winter

1200

1000

800

AA22 AA27 AE18 AE1

Eh (mV)

600

400

200

0 -200

-400

All the analyzed water samples are oxidizing, but there are redox potential differences between the estuarine tidal channel (Eh mean value around +380 mV) and the poorly drained lacustrine zone, where Eh values as low as +225 mV were measured. The higher values of Eh (up to +585 mV) were registered in the most acidic waters, resulting in a deviation from the general pattern shown by the dominant waters in the wetland area (Fig. 3). The electrical conductivity (EC) of water declined gradually in the upstream direction, from 47.5 mS cm 1 (estuarine zone) to values around 1 mS cm 1 (lacustrine zone), thus reflecting the tidal interaction effect. The maximum value was recorded during high tide at the confluence of the Domingo Rubio stream and the Tinto river (Fig. 2b). The longitudinal gradient of salinity varied from about 30‰ to less than 0.5‰, within 6 km of distance along the tidal channel. The EC values reached up to 28.5 mS cm 1 in some temporary floodplain ponds (sample AE 18) affected by acid leachates from nearby sulfide wastes. The dissolved oxygen (DO) concentration for tidal channel waters during morning high tide ranged between 6.8 and 10.2 mg L 1, with a mean value of 8.6 mg L 1. However, the streams draining across the agricultural fields showed concentrations of DO (from 4.9 to 6.8 mg L 1) lower than tidal channel into which they discharge.

-600

4.2. Major cations, anions and nutrients -800 0

1

2

3

4

5

6

7

8

9

1 0 11 12 13 14

pH Fig. 3. Water samples plotted on an Eh–pH water stability diagram.

lacustrine wetland, the mean pH values vary in a wider range depending on the season (7.5 in summer and 8.4 in winter).

As expected in a well mixed estuarine system, Na+ is by far the most prevalent major cation dissolved in the estuarine water. The maximum concentrations of Na+ were found in the mouth of the Domingo Rubio tidal channel (sample AE1), although they varied widely between high tide (up to 9725 mg L 1) and low tide (up to 4278 mg L 1). The content of Na+ decreases with increasing distance away from the estuary, and drops dramatically in the lacustrine

Fig. 4. Piper diagram depicting the relative ionic composition of the water samples. The proportions are expressed as% meq L

1

.

1266

C. Barba-Brioso et al. / Marine Pollution Bulletin 60 (2010) 1259–1269

An upstream gradient of decreasing concentrations also exists regarding major anions. The high levels of chloride (up to 16390 mg L 1), sulfate (up to 2426 mg L 1) and bromide (up to 56.4 mg L 1) were measured at high tide in estuarine waters (Fig. 2c,d,e). These values are close to average concentrations in seawater (Hem, 1992), and contrast markedly with those found in freshwaters of the artificial lake and stream tributaries. The bicarbonate anion showed a more homogeneous distribution pattern within the wetland area (Fig. 2f), with values generally lower than 200 mg L 1. In most water samples the fluoride content was below the detection limit of the method (<0.1 mg L 1), except some tributaries that contained up to 0.3 mg L 1. The hydrochemical data plotted on a Piper diagram (Fig. 4) of relative proportions of dissolved major cations and anions reveals two dominant water types: calcium–magnesium sulfate–chloride mixed and sodium chloride– sulfate. In terms of nutrient anions, the quantity of phosphate dissolved in the estuarine water was significantly greater at high tide than at

Table 3 Concentrations of pesticides detected in water at different sampling sites. Pesticides (lg L

Simazine Pendimethaline Diuron Terbuthylazine

1

)

Tideland zone

Lacustrine zone

Tributary streams

AE6

AE16

AL12

AL24

AA10

AA11

AA20

0.62 0.125 nd nd

nd nd nd nd

0.78 nd nd nd

0.42 0.37 nd nd

0.52 0.072 nd nd

0.5 0.078 nd nd

0.65 nd 21.8 52

zone, where the mean value (below 100 mg L 1) changes seasonally rather than tidally. A similar spatial and seasonal distribution pattern is apparent for the other major cations. The largest concentrations of Mg2+ (1020 mg L 1), Ca2+ (496 mg L 1) and K+ (368 mg L 1) were recorded in the stream mouth area during the high tide period. These values are within the range of concentrations obtained by Grande et al. (1999) as typical of the Huelva estuary.

2AA22 2AA272AA27

50000,00

2AA22

Concentration ( g L-1 )

5000,00

2AA27 2AA22

2AA27 2AA22 2AE1

2AE1 2AE18

2AA22 2AA27

500,00

2AA22

50,00

5,00

Median Interquartile range Non-Outlier Range Extremes

0,50

0,05 Cr

Mn

Fe

Ni

Cu

Zn

As

Cd

Fig. 5. Box and whisker plot for trace element concentrations showing the median value within a box defined by the interquartile range, the extreme values that exceed the expected distribution range, and the whiskers representing the non-outlier range.

Rescaled Distance Cluster Combine 0

5

10

15

Cl Br EC Na Mg Ca 2SO4 K Cr Fe Ni Cu As Zn Cd Eh Mn NO3 PO43pH HCO32Fig. 6. Dendrogram obtained from the cluster analysis.

20

25

1.000 0.413 0.572 1.000 0.471 0.842 0.665 1.000 0.474 0.043 0.517 0.388 1.000

0.290 0.208 0.297 0.535 0.165 0.005 0.427 0.268 0.128 0.180 0.510 0.195 0.330 0.114 0.074 0.264 0.134 0.153 0.339 0.088 0.047 0.470 0.004 0.503

1267

low tide, and decreased upstream from 4.1 mg L 1 to 0.7 mg L 1 following a longitudinal gradient along the tidal channel (Fig. 2g). Instead, nitrate distribution showed an opposite trend, with higher concentration during the low tide period increasing in the upstream direction (Fig. 2h), from less than 0.3 mg L 1 (mouth area) to 15.1 mg L 1. Nevertheless, the highest levels of phosphate (up to 18.5 mg L 1), nitrate (up to 96.8 mg L 1) and nitrite (up to 1.82 mg L 1) were detected in small streams that drain agricultural landscapes (samples AA10 and AA11). 4.3. Trace elements Iron concentrations in waters of the Domingo Rubio wetland are highly variable, from less than 100 to over 3000 lg L 1. Elevated contents of Fe were measured in estuarine water at the confluence of the tidal channel and the Tinto river (up to 1802 lg L 1). In the tideland area, the Fe showed decreasing upstream concentrations during high tide, but followed the opposite trend during the low tide period (Fig. 2i). Enhancement values (up to 1499 lg L 1) also occurred at the lacustrine zone in summer. However, the highest Fe concentration (3433 lg L 1) was measured in stream water receiving acid leachates derived from sulfide waste dumps (sample AE18). On the other hand, the distribution of various potentially toxic elements (Cu, As, Zn, Co and Cd) follows closely the pattern of major cations and anions, depicting a decreasing upstream trend along the tidal channel during both flood and ebb tides (Fig. 2j– n). The abundances of Mn and Cr (Fig. 2o,p) did not reveal any systematic distribution pattern, and most water samples showed Pb concentrations below the analytical detection limit (0.2 lg L 1) due to its low solubility under neutral aqueous conditions. However, the largest amounts of trace elements were confined to some well-defined stream tributaries (samples AA22 and AA27) affected by acid spills from an abandoned chemical plant (up to 168 mg L 1 of Cu; 33.6 mg L 1 of Zn; 4.37 mg L 1 of Cd; 1.54 mg L 1 of Mn; 770.5 lg L 1 of Ni; 267.2 lg L 1 of Co; 219.1 lg L 1 of As; and 198.3 lg L 1 of Pb). Very high values of various metals (especially Cu, Pb and Zn) were also found in stream water receiving acid discharges from sulfidic wastes (sample AE18) and industrial effluents from the petrochemical industry (sample AA19).

0.718 0.833 0.876 0.668 0.755 0.485 0.034 0.487 0.793 0.317 0.756 0.481 0.363 0.290 0.319 0.500 0.233 0.212 0.289 0.422 0.321 0.000 0.342 0.253 0.766 0.881 0.935 0.689 0.782 0.532 0.092 0.539 0.824 0.330 0.821 0.490 0.681 0.845 0.935 0.686 0.779 0.523 0.106 0.392 0.782 0.350 0.817 0.468

0.247

0.255 0.390 0.455 0.279 0.563 0.450 0.051 0.130 0.512 0.223 0.313 0.306

0.303

0.336 0.176 0.135 0.198 0.062 0.068 0.553 0.041 0.080 0.101 0.151 0.277

PO34 Ca Mg Na K Cr Mn Fe Ni Cu Zn As Cd

0.761 0.873 0.975 0.741 0.782 0.522 0.194 0.563 0.803 0.278 0.879 0.450

0.167

0.187 0.336 0.244 0.226

0.071 0.570

SO24

HCO23

0.352

0.032 0.182 0.080

1.000 0.356 1.000 0.355 0.973 1.000 0.954 0.277 0.936 1.000 0.951 0.962 0.326 0.907 1.000 0.464 0.490 0.504 0.042 0.536 pH Eh EC Cl Br NO

1.000 0.131 0.157 0.109 0.192 0.091 0.141

0.164

1.000

4.4. Pesticides

SO24 NO3 Br Cl EC Eh pH Variable

Table 4 Spearman’s correlation matrix. Figures in bold are significant <0.01.

0.253

HCO3

1.000

PO34

Ca

1.000 0.850 0.756 0.759 0.534 0.383 0.303 0.501 0.628 0.171 0.639 0.136

Mg

1.000 0.900 0.668 0.753 0.398 0.133 0.411 0.769 0.345 0.735 0.436

Na

1.000 0.707 0.831 0.521 0.160 0.514 0.792 0.314 0.864 0.490

K

1.000 0.419 0.499 0.393 0.405 0.517 0.023 0.701 0.042

Cr

1.000 0.299 0.091 0.413 0.875 0.450 0.812 0.666

Mn

1.000 0.253 0.308 0.360 0.028 0.342 0.071

Fe

1.000 0.140 0.179 0.187 0.287 0.276

Zn Cu Ni

As

1.000 0.505

Cd

1.000

C. Barba-Brioso et al. / Marine Pollution Bulletin 60 (2010) 1259–1269

Nine herbicides are actually authorised for application in strawberry orchards in this region. Napropamide and pendimethalin are two of them found in the study area. The others herbicides found in waters are not generally used for strawberry protection but they are widely used in other crops or even as industrial herbicides. The levels of pesticide found in selected sampling sites are given in Table 3. For the rest of the analyzed pesticides (atrazine, napropamide, desethylsimazine and desethylterbuthylazine), the concentrations fall below the detection limit of the method in all the analyzed water samples. Simazine concentrations had always levels higher than the 0.1 lg L 1, according to the vulnerability limit set by the Directive 98/83/CE (CEC, 1998). The highest amount of this herbicide (0.78 lg L 1) was detected in lacustrine waters. However, diuron and pendimethaline concentrations were lower than 0.1 lg L 1, except those values corresponding to samples AE6 and AL24 for pendimethaline and AA20 for diuron. The latter location shows an unusual high value for diuron (21.8 lg L 1) and terbuthylazine (52 lg L 1) concentrations, probably related to a punctual unvoluntary discharge of these two herbicides into the tributary stream coming from an industrial area. Pendimethaline was found in almost all sampling points affected by the strawberry production. Herbicide concentrations measured in these waters are similar to those found in others surface waters in Spain (Hildebrandt et al., 2008).

1268

C. Barba-Brioso et al. / Marine Pollution Bulletin 60 (2010) 1259–1269

5. Discussion Although there is no clear evidence of eutrophication or harmful algal blooms in water bodies, the amounts of certain aquatic nutrients (particularly nitrates and phosphates) are well above average values reported for natural surface waters (Meybeck, 1982; Salminen, 2005). High concentrations of chlorophyll-a (85–700 mg m 3) were detected in the recent past, which are indicative of hypertrophic conditions during the assessment period (Junta de Andalucía, 2004). Nitrogen fertilizer is widely used in the watershed area to increase the production of strawberry crops. Nitrate median value (34.68 mg L 1 in summer and 24.81 mg L 1 in winter) for tributary streams that drain lands under intensive farming is certainly anomalous when compared with that of stream water (2.8 mg L 1) in Western Europe (Salminen, 2005). In some tributaries, nitrate concentrations exceed the safety limit of 50 mg L 1 established by the World Health Organization for drinking water. Surface runoff from agricultural fields contributes significantly to enhancement of nitrate loads in both lacustrine and estuarine waters of the Domingo Rubio wetland, which represents the main transport pathways by which agrochemicals reach the Huelva estuary. Therefore, the high proportion of nitrates detected in this estuarine environment (Grande et al., 1999; Grande et al., 2003; Elbaz-Poulichet et al., 1999; Morillo et al., 2005) seems to be linked to agricultural supplies rather than urban and industrial effluents. Anomalous isolated values of nitrites found in some tributaries (up to 1.82 mg L 1) could also be related to fertilizer application or sewage effluents, as documented by Mendiguchía et al. (2007). Phosphorous is another important plant nutrient used as a fertiliser in the catchment. Phosphate levels in natural freshwater are typically low (median value 0.02 mg L 1, according to Reimann and De Caritat, 1998), because of its tendency to form poorly soluble complexes with major cations. In comparison to natural values, some tributaries that discharge into the Domingo Rubio system had concentrations as high as 18.5 mg L 1, as a probable result of excessive fertilizer application. High but decreasing upstream concentrations (from 4.1 mg L 1 to 0.7 mg L 1) in estuarine waters during flood tide suggest that nearby phosphogypsum wastes stockpiled on the Tinto river bank (Elbaz-Poulichet et al., 2000; Braungardt et al., 2003) should be an additional anthropogenic source of phosphate pollution. In addition, relatively large proportions of potentially toxic heavy metals and metalloids are being periodically supplied by the contaminated Odiel–Tinto river and estuary system. The influx of metallic pollutants entering the Domingo Rubio tideland is effective during flood tidal periods, when the dissolved load of sulfide-associated elements (Cu, Zn, As and Cd) in estuarine water exceed by at least one order of magnitude the typical values reported for uncontaminated natural water (e.g. Hem, 1992; Drever, 1997). The occurrence of Ni and Cr (non-sulfide source metals) in the dissolved phase is indicative of some additional industrial input. Despite the low solubility of most metals under neutral conditions and low Eh values (Perelman, 1986), the observed enhancement levels could be explained by the abundance of inorganic complexing agents in the estuarine water, mainly chloride and sulfate anions, which may keep the metals in solution. The decline in salinity may not only diminish the complexing capacity of water but also increase the extent of adsorption by particulate matter, thus reducing the dissolved heavy metal load. The increase of pH values in the upstream direction could also enhance the sorption capacity of suspended material. Controls on dissolved Fe and Mn concentrations are more complex because their solubilities are strongly influenced by redox processes (Bourg, 1995). The highest

mean value (1338 lg L 1) for Fe was obtained at the poorly drained lacustrine zone, where the relatively low Eh values may give rise to high concentrations. Because of estuarine water body moves in and out with the tide, the trace element concentrations vary notably along the tidal channel, following an upstream decreasing trend for most metals. Therefore, tidal regime seems to be the main factor controlling the dispersion of pollutants in the tideland zone, whereas seasonal changes may affect relative abundance of heavy metals and agrochemicals in the lacustrine zone and its inflowing streams. In these freshwater environments, mineralization peaks in summer conditions of low water flow due to the high concentration of dissolved salts. However, considering as thresholds the extreme values that exceed the non-outlier range of all water samples (Fig. 5), the most anomalous metal concentrations correspond to stream water receiving acid drainage from sulfide wastes (sample AE18) and industrial effluents and spills (samples AA19, AA22 and AA27). In these highly anomalous sampling sites, the metals are easily mobilized and readily available for ecological uptake because of the high acidity of the stream water. On the other hand, the extreme value recorded at the mouth of the Domingo Rubio tidal channel (sample AE1) reflects primarily the heavy metal pollution of the Huelva estuary. Structure of the variable dependence was found by Spearman correlation matrix (Table 4). A dendrogram obtained from the results of the cluster analysis provides an easy way to visualize the relationships existing among the hydrochemical variables (Fig. 6). Three cluster were identified. Cluster 1 is dominated by major cations (Na+, K+, Mg2+, Ca2+), seawater anions (Cl , Br , SO24 ), electrical conductivity, Fe, and Cr which usually form water-soluble oxyanion species under circumneutral pH conditions. This group is interpreted to reflect the influx of high-salinity polluted water coming from the estuary of Huelva. Cluster 2 is defined by Eh values, Mn and certain trace elements, such as As, Cd, Cu, Ni and Zn. This cluster represents the anthropogenic assemblage of pollutants released from industrial effluents and spills, and sulfide oxidation reactions in abandoned waste dumps. Finally, the association of nutrient anions (NO3 and PO34 ), HCO3 and pH values is explained by the Cluster 3. This group reflects, at least in part, the fertilizers drained from the agricultural soils.

6. Conclusions Based on the monitoring results, the following conclusions can be drawn:  The Domingo Rubio wetland is subject to diffuse water pollution caused by the combined effect of multiple anthropogenic sources. The tideland stretch is semidiurnally flooded with saline water containing typical concentrations of seawater ions and high levels of dissolved Cu, As, Cd, Co, Cr, Ni and Zn, as a result of the estuarine flushing. The mine-polluted estuary of Huelva constitutes, therefore, a significant source of dissolved metals to the studied tidal system.  Additionally, the tidal channel receives acid discharges arising from specific point sources, such as sulfide wastes dumps, industrial effluents pipes and abandoned chemical plants, thus contributing to metal enhancement.  Agriculture is other main source of diffuse pollution in the wetland area. During periods of high freshwater influx, water quality is affected by inputs of nitrate and phosphate through agricultural runoff. The Domingo Rubio channel tidal is a major contributor of nutrients into the Huelva estuary, but it also receives phosphate fluxes from phosphogypsum wastes dumped in the estuary.

C. Barba-Brioso et al. / Marine Pollution Bulletin 60 (2010) 1259–1269

 Water contamination with pesticides (mostly simazine and pendimethalin) is generally higher than the CEC vulnerability level in the wetland area, although high levels of diuron and therbuthylazine were found locally. This contamination by agrochemicals could be avoided if a more controlled herbicides and fertilizers application be considered by the farmers.  The transport and dispersion of pollutants released by the different sources is mainly controlled by the tidal regime, except in the artificially created reservoir where freshwater exhibits a seasonal mineralization gradient.

Acknowledgment This work was financially supported by the Regional Government of Andalusia through the Project RNM 05-523 ‘‘Aproximación multidisciplinar para el estudio de la contaminación y sus efectos en organismos. Aplicación al estero Domingo Rubio (Parque Natural Marismas del Odiel)”. References Achterberg, E.P., Herzl, V.M.C., Braungardt, C.B., Millward, G.E., 2003. Metal behaviour in an estuary polluted by acid mine drainage: the role of particulate matter. Environ. Pollut. 121, 283–292. Aguer, J.P., Cox, L., Richard, C., Hermosin, M.C., Cornejo, J., 2000. Sorption and photolysis studies in soil and sediment of the herbicide napropamide. J. Envion. Sci. Health-Part B 35, 725–738. Barba-Brioso, C., Fernández-Caliani, J.C., Miras, A., Galán, E., 2007. Mineralogía de los suelos y sedimentos actuales del estero Domingo Rubio (estuario de Huelva). Macla 7, 60. Barba-Brioso, C., Fernández-Caliani, J.C., Miras, A., Galán, E., 2009. Evaluation of labile metal pools in soils from a highly industrialized wetland area of southwestern Spain by single and sequential extraction methods. Geochim. et Cosmochim. Acta 73(13S), 85A. Borrego, J., Morales, J.A., De La Torre, M.L., Grande, J.A., 2002. Geochemical characteristics of heavy metal pollution in surface sediments of the Tinto and Odiel river estuary (southwestern Spain). Environ. Geol. 41, 785–796. Bourg, A.C.M., 1995. Speciation of heavy metals in soils and groundwater and implications for their natural and provoked mobility. In: Salomon, W., Förstner, U., Mader, P. (Eds.), Heavy metals. Problems and Solutions. Springer-Verlag Publisher, pp. 19–31. Braungardt, C.B., Achterberg, E.P., Elbaz-Poulichet, F., Morley, N.H., 2003. Metal geochemistry in a mine-polluted estuarine system in Spain. Appl. Geochem. 18, 1757–1771. Butler, E.C.V., 2006. The tail of two rivers in Tasmania: the Derwent and Huon estuaries. In: Wangersky, P. (Ed.), Estuaries, vol. 5 H. Hdb. Environ. Chem. Springer-Verlag Publisher, pp. 1–49. Cabrera, A., Cox, L., Velarde, P., Koskinen, W.C., Cornejo, J., 2007. Fate of diuron and terbuthylazine in soils amended with two-phase olive oil mill waste. J. Agric. Food Chem. 55, 4828–4834. CEC (Commission of the European Communities), 1998. Council Directive 98/83/ EEC of 3 November 1998 related to the water quality for human consuming. Official J. L 330, pp. 32–54. Davis Jr., R.A., Welty, A.T., Borrego, J., Morales, J.A., Pendón, J.G., Ryan, J.G., 2000. Rio Tinto estuary (Spain): 5000 years of pollution. Environ. Geol. 39, 1107–1126. Drever, J.I., 1997. The geochemistry of natural waters. Surface and Groundwater Environments, third ed. Prentice Hall. Eaton, A.D., Clesceri, L.S., Greenberg, A.E. (Eds.), 1998. Standard Methods for the Examination of Water and Wastewater, 20th ed. American Public Health Association, American Water Works Association and Water Environment Federation, Washington, DC. Elbaz-Poulichet, F., Morley, N.H., Cruzado, A., Velasquez, Z., Achterberg, E.P., Braungardt, C.B., 1999. Trace metal and nutrient distribution in an extremely low pH (2.5) river–estuarine system, the Ría of Huelva (South-West Spain). Sci. Total Environ. 227, 73–83. Elbaz-Poulichet, F., Dupuy, C., Cruzado, A., Velasquez, Z., Achterberg, E.P., Braungardt, C.B., 2000. Influence of sorption processes by iron oxides and algae fixation on arsenic and phosphate cycle in an acidic estuary (Tinto river, Spain). Water Res. 34, 3222–3230.

1269

Fernández-Caliani, J.C., Ruiz, F., Galán, E., 1997. Clay mineral and heavy metal distributions in the lower estuary of Huelva and adjacent Atlantic shelf. Sci. Total Environ. 198, 181–200. Fernández-Caliani, J.C., Barba-Brioso, C., González, I., Galán, E., 2009. Heavy metal pollution in soils around the abandoned mine sites of the Iberian Pyrite Belt (South-West Spain). Water Air Soil Pollut. 200, 211–226. Fresenius, K.E. Quentin, W. Schneider (Eds.), 1988. Water Analysis: A Practical Guide to Physico-Chemical, Chemical and Microbiological Water Examination and Quality Assurance. Springer-Verlag, cop, Berlin, 804 p. Grande, J.A., Borrego, J., Morales, J.A., 1999. A study of heavy metal pollution in the Tinto–Odiel estuary in southwestern Spain using factor analysis. Environ. Geol. 39, 1095–1101. Grande, J.A., Borrego, J., De La Torre, M.L., Sáinz, A., 2003. Application of cluster analysis to the geochemistry zonation of the estuary waters in the Tinto and Odiel rivers (Huelva, Spain). Environ. Geochem. Health 25, 233–246. Hem, J.D., 1992. Study and Interpretation of the Chemical Characteristics of Natural Waters, third ed. USGS Water-Supply Paper. Hildebrandt, A., Guillamon, M., Lacorte, S., Tauler, R., Barceló, D., 2008. Impact of pesticides used in agriculture and vineyards to surface and groundwater quality (North Spain). Water Res. 42, 3315–3326. Junta de Andalucía, 2004. Plan Andaluz de Humedales. Consejería de Medio Ambiente. Madejón, P., Burgos, P., Murillo, J.M., Cabrera, F., Madejón, E., 2009. Bioavailability and accumulation of trace elements in soils and plants of a highly contaminated estuary (Domingo Rubio tidal channel, SW Spain). Environ. Geochem. Health 31, 629–642. Martín, J.E., García-Tenorio, R., San Miguel, E.G., Respaldiza, M.A., Bolívar, J.P., Alves, L.C., da Silva, M.F., 2002. Historical impact in an estuary of some mining and industrial activities evaluated through the analysis by TTPIXE of a dated sediment core. Nucl. Instrum. Methods Phys. Res. B 189, 153–157. Mendiguchía, C., Moreno, C., García-Vargas, M., 2007. Evaluation of natural and anthropogenic influences on the Guadalquivir river (Spain) by dissolved heavy metals and nutrients. Chemosphere 69, 1509–1517. Meybeck, M., 1982. Carbon, nitrogen and phosphorous transport by world rivers. Am. J. Sci. 282, 401–450. Miller, J.C., Miller, J.N., 2000. Statistics and Chemometrics for Analytical Chemistry, fourth ed. Prentice Hall, London. Montes-Nieto, R., Fuentes-Almagro, C.A., Bonilla-Valverde, D., Prieto-Alamo, M.J., Jurado, J., Carrascal, M., Gómez-Ariza, J.L., López-Barea, J., Pueyo, C., 2007. Proteomics in free-living Mus spretus to monitor terrestrial ecosystems. Proteomics 23, 4376–4387. Morillo, J., Usero, J., Gracia, I., 2005. Biomonitoring of trace metals in a minepolluted estuarine system (Spain). Chemosphere 58, 1421–1430. Nelson, C.H., Lamothe, P.J., 1993. Heavy metal anomalies in the Tinto and Odiel river and estuary system. Spain Estuaries 16, 496–511. Olías, M., Cánovas, C.R., Nieto, J.M., Sarmiento, A.M., 2006. Evaluation of the dissolved contaminant load transported by the Tinto and Odiel rivers (South West Spain). Appl. Geochem. 21, 1733–1749. Pendón, J.G., Morales, J.A., Borrego, J., Jiménez, I., López, M., 1998. Evolution of estuarine facies in a tidal channel environment, SW Spain: evidence for a change from tide- to wave-domination. Mar. Geol. 147, 43–62. Perelman, A.I., 1986. Geochemical barriers: theory and practical applications. Appl. Geochem. 1, 669–680. Reimann, C., De Caritat, P., 1998. Chemical Elements in the Environment. SpringerVerlag, Berlin. Ruiz, F., González-Regalado, M.L., Pendón, J.G., Abad, M., Olías, M., Muñoz, J.M., 2005. Correlation between foraminifera and sedimentary environments in recent estuaries of Southwestern Spain: applications to Holocene reconstructions. Quatern. Int. 140–141, pp. 21–36. Ruiz, F., Borrego, J., González-Regalado, M.L., López-González, N., Carro, B., Abad, M., 2008. Impact of millennial mining activities on sediments and microfauna of the Tinto River estuary (SW Spain). Mar. Pollut. Bull. 56, 1258–1264. Rump, H.H., 1999. Laboratory Manual for the Examination of Water, Waste Water and Soil, third ed. Wiley-VCH, cop, Weinheim, 225 p. Sáinz, A., Grande, J.A., De La Torre, M.L., 2004. Characterisation of heavy metal discharge into the Ria of Huelva. Environ. Int. 30, 557–566. Salminen, R., 2005. Geochemical Atlas of Europe. Part 1: Background Information, Methodology and Maps. Geological Survey of Finland . Sánchez España, J., López Pamo, E., Santofimia, E., Aduvire, O., Reyes, J., Barettino, D., 2005. Acid mine drainage in the Iberian Pyrite Belt (Odiel river watershed, Huelva, SW Spain): geochemistry, mineralogy and environmental implications. Appl. Geochem. 20, 1320–1356. USEPA, 1984. Appendix B to Part 136 – definition and procedure for the determination of the method detection limit – revision 1.11. Fed. Regist. 49 (209), 43430.