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Arsenic and other trace elements in the waters and organisms of an estuary in SW England
ARSENIC AND OTHER TRACE ELEMENTS IN THE WATERS AND ORGANISMS OF AN ESTUARY IN SW ENGLAND
D. W. KLUMPP & P. J. PETERSON
Department of Botany and Biochemistry, Westfield College, Kidderpore Arenue, London NW3 7ST, Great Britain
ABSTRACT
Arsenic, cadmium, copper and zinc occur at elevated levels in the waters of Carnon River and the upper reaches of Restronguet Creek, these concentrations decreasing progressively towards the estuary of Carrick Roads. While arsenite is the main form of As in the Carnon River, this is converted to arsenate in the lower estuary. Organisms analysed from Restronguet Creek reflect the As concentration in the water, this being most clearly evident in macrophytes. Although accumulated at all trophic levels, there was no evidence for biomagnification of As on an entire organism basis. The As was shown to exist in selected organisms as a compound(s) whichjorms on hydrolysis, dimethylarsinate and traces of arsenate and methylarsonate.
INTRODUCTION
Our knowledge of the biological cycling of arsenic in marine systems is limited. With the aid of recent improvements in the methods of As analysis it has been shown that As occurs in marine waters predominantly as arsenate with traces of arsenite, methylarsonic acid and dimethylarsinic acid (Braman & Foreback, 1974; Andreae, 1977, 1978). Arsenic is accumulated by a wide variety of marine organisms but in view of the known toxicity of high concentrations of arsenite and arsenate, it is unlikely to be accumulated in this form. Indeed, several workers have extracted organo-arsenic compounds from marine organisms and this information is summarised by Lunde (1977). Edmonds et al. (1977) have identified arsenobetaine in extracts of lobster. Reports that As is rapidly excreted by some marine organisms (Penrose, 1975; 11 Environ. Pollut. 19 (1979)--© Applied Science Publishers Ltd, England, 1979 Printed in Great Britain
12
D. W. KLUMPP, P. J. PETERSON
Penrose et al., 1977) support the view that this element is not bioaccumulated up the food chain. We have been examining the accumulation and metabolism of As in an estuarine food chain in Restronguet Creek and Carrick Roads. Pollution by As in the terrestrial area has already been reported from this laboratory (Porter & Peterson, 1975, 1977). The region drained by the Carnon River has a long history of mining activity dating back to the tin and copper mines of the Roman era. It extends to the present as alluvial tin operations and the re-opened tin mines at Mount Wellington and Wheal Jane. These activities, in addition to the natural processes of weathering, have resulted in considerable contamination by As and some trace metals in the waters and sediments of the Carnon River and its estuary (Hosking & Obial, 1966; Yim, 1972; Thornton et al., 1975). We report here the results of a survey where the concentrations of As, Cd, Cu and Zn were measured in waters and organisms from the Restronguet Creek system. The speciation of As in waters and its metabolism in several organisms are also mentioned.
MATERIALS AND METHODS
Water and selected organisms from the estuary (Fig. 1) were analysed for As and, on occasions, for Zn, Cu and Cd between November 1975 and September 1977. Observed effects of storage on the As species in water samples were essentially the same as those reported by Andreae (1977), although loss of arsenite was more rapid in the present study. To minimise changes, water samples were filtered (millipore HA, 0.45 pm) into polyethylene bottles in the field and analysed within 24h of collection. Intertidal organisms collected for analysis were transported to the laboratory, washed in filtered seawater and frozen until required. Samples of approximately 1 g dry weight (80 °C) were wet digested in concentrated nitric acid (BDH, AnalaR) followed by a 4:1 mixture of nitric and perchloric acid (BDH, AnalaR) until a clear solution was obtained. Final solutions were taken up in 1.5N hydrochloric acid. Reagent blanks were included in all digestion batches. Analytical procedures were tested by a comparative analysis of the standard materials Bowen's Kale and NBS Orchard Leaves. Results are presented for Zn, Cu, Cd and As in Bowen's Kale, and As in NBS Orchard Leaves (Table 1). Results obtained were in close agreement with those reported for both sample materials. A recovery of 92( + 2) ~ was obtained for As-spiked Kale.. Until May 1976 all water samples were treated by extraction of the APDCcomplex into chloroform as described by Mulford (1966). Arsenite was extracted from one aliquot while a second was reduced, then extracted, to determine total As (Strickland & Parsons, 1968). Elements were measured on a Varian Tectron AA5
ARSENIC AND METALS IN ESTUARINE ORGANISMS
5010' ¢
05' T
13
I"-"
sd
15/
Wheal
~, Jane Mt. Wel lington
1 I
16
0
2
I
I
kilometre
~
"~,~ ~/~
~~o
Falml English Channel STUDY AREA Fig. 1.
Map of the study site with sampling locations.
TABLE 1 C O M P A R A T I V E ANALYSIS W I T H S T A N D A R D REFERENC|" MATERIALS B O W E N ' S KALE u ~.ND NBS O R C H A R I ) [.IL~,VI-S b (IN PPM DRY W T )
Material
Kale
Orchard Leaves
Element
As Zn Cu Cd As
Present study ~
0.125 28.4 5"4 0.64 10-0
(0"012) (0,8) (0'5) (0,13) (0,1)
Bowen (1974). NBS (1976). "Values are the mean (n = 10) with the standard deviation in parentheses.
ReJerence ~'h
0.141 33.2 4-99 0.80 10.0 (20)
14
D.W. KLUMPP, P. J. PETERSON
atomic absorption spectrophotometer with hydride generation and electrically heated furnace adaptation for As determination ( T h o m p s o n & Thomerson, 1974; T h o m p s o n & Thoresby, 1977). F r o m M a y 1976, methylarsonate and dimethylarsinate, in addition to arsenate and arsenite, were measured in water using the more sensitive m e t h o d o f Andreae (1977). With A A S application o f this method, detection limits for arsine, m o n o - and dimethylarsine were 0.5, 0.7 and 1-2 ng, or 20, 28 and 48 ng/litre with a 25 ml sample. Selected organisms were homogenised in a blender with a 1:2 mixture o f water/methanol. The lipid fraction was separated by extracting with four volumes o f chloroform. The water/methanol fraction was analysed for As before and after boiling for 1 h with 1N sodium hydroxide. Part o f the initial extract was refluxed with concentrated nitric acid for 24h, taken to dryness, then dissolved in l'5N hydrochloric acid and analysed as previously described.
RESULTS AND DISCUSSION E l e m e n t in w a t e r s
Dissolved As levels (Table 2) in the waters o f the lower estuary (Fig. 1) are close to published figures (Onishi, 1969; Andreae, 1978). In comparison, As was at considerably elevated concentrations in the upper reaches o f Restronguet Creek and in the C a r n o n River (Table 2). The high concentration o f As seen for station 1 in C a r n o n River water decreases markedly a r o u n d station 2 where river and marine waters mix. The d r o p in concentrations shown at station 2 m a y be due to dilution o f As-rich river water with that o f marine origin, a l t h o u g h precipitation o f the element TABLE 2 DISSOLVED ARSENIC SPECIES IN WATERS a W I T H LOCATION IN T H E C A R N O N ESTUARY SYSTEM
Station b'C
1 2 LT 2 HT 3LT 3 HT 4LT 4 HT 5 LT 5 HT 6LT 6 HT 7 LT 7 HT
January 1976
Concentration (#g/litre) May 1976
March 1977
25"7 (16) 16.0 (19) 3-5 (63) 11.8 (35) 4.0 (95) 9.0 (40) 5.0 (96) 5.9 (86) 3.5 (94) 4.1 (61) 2.8 (100) 2.3 (100) 2.3 (100)
42-1 (5) 22.6 (7) 10.6 (60) 14.9 (22) 4.6 (72) 8.1 (53) 5.2 (86) 7.1 (48) 3.4 (88) 6-8 (63) 3.7 (92) 4-8 (100) 1.9 (100)
34"3 (9) 18.3 (0) 7.9 (64) 15-2 (33) * 11.6 (71) 6.7 (100) 9.9 (84) 1.8 (100) 9.9 (66) 2.6 (100) 8.3 (100) 4.8 (100)
aValues are the mean (n --- 5) total arsenic (arsenate + arsenite) with % arsenate in parentheses. bRefer to Fig. 1 for location. cCollected at high tide (HT) and low tide (LT). * No data.
ARSENIC AND METALS IN ESTUARINE ORGANISMS
15
out of solution may be important. Mean values of over 1000 ppm As are reported (Thornton et al., 1975) in sediments of this estuary, providing a possible source of As to the water. Arsenate is the major form of As detected in the waters of the estuary, while arsenite predominates in the Carnon River. Arsenite occurs in waters of the estuary in decreasing concentrations towards the sea. Samples of water taken at low tide at station 2 contain mainly arsenite but at high tide arsenate predominates. Levels of both inorganic species are important at stations 3 and 4 during low tide while stations further down the estuary are consistently low in arsenite. Arsenite was not detected at high or low tide in the main estuary of Carrick Roads. These findings are summarised in Table 2. Results suggest that As enters the system as arsenite in Carnon River waters and is then either converted to arsenate or diluted when mixed with saline waters. Both Johnson (1971) and Andreae (1977, 1978) have reported arsenate as the main form of As in marine waters. The presence of arsenite in marine waters is thought to be a product of biological action (Johnson, 1972). Indeed, phytoplankton grown at natural As levels have been shown to release arsenite and methylated arsenics into water (details to be published later). Methylated As species were not detected in open waters of the estuary. However, dimethylarsonate (0.002 ppm) was found in one sample of rock pool water. This initiated a more thorough examination of waters from rock pools at a range of tidal heights and with varying degrees of algal growth. Methyl arsenics were not detected and the pools were found to contain As as arsenate and arsenite at levels seen in the estuary for that tidal position (see Table 2). Concentrations ofCd, Cu and Zn in waters of this system agree with that reported by Thornton et al. (1975). Cadmium, Cu and Zn were more heavily concentrated in the Carnon River compared with the estuary (Table 3). Elements in organisms High levels of Zn and Cu were found in the oyster Ostrea edulis and to a lesser extent in macrophytes (Table 4). Enteromorpha intestinalis showed the least TABLE 3 DISSOLVED Z I N C , C O P P E R A N D C A D M I U M IN WATERS a OF T H E C A R N O N ESTUARY SYSTEM
Location River C a r n o n Stn 1 Restronguet Creek (low tide) Stn 4 Carrick Roads (low tide) Stn 6
Zinc
Concentration (~g/litre) Copper
Cadmium
5820 3692-22000) 285 (12.9-305) 230 (16-500)
236 (32.5 395.7) 16 (2.1 21-0) 13 (2.8 20)
5.3 (4.3 7.4) 2.8 (nd 3-71 2"6 (nd 3.1)
~Values are the mean and range in parentheses from samples collected over 1976 77. n d = Not detected.
PETERSON
D. W . KLUMPP, P. J.
16
TABLE 4 ELEMENTS IN THE ENTIRE SOFT PARTS OF S E L E C T E D ORGANISMS FROM RESTRONGUET CREEK
Speciesb Arsenic Fucus serratus F. vesiculosus F. spiralis Ascophyllum nodosum Peh,etia canaliculata Enteromorpha intestinalis Tealiafelina
Stn 6 Ostrea edulis Mytilus edulis Patella vulgata
Stn 6 Carcinus maenas Littorina obtusata Nucella lapillus
141.4 (103.3-189.3) 105.3 (94.7-126.0) 103.6 (96.3-113.4) 91.7 (69-7--118.3) 65.0 (59.1-71.0) 83.7 (83.3-85-5) 55.2 (53.3 5 9 . 0 ) 17.0 (16.9-17.2) 16.9 (16.8-17.1) 36.7 (35.3 41.0) 22.3 (19.9-34,5)
Concentration (ppm dry wt) a Zinc Copper
771 65 (727-1321) (62.4~68.2) 719 89 (709.8 7 3 8 ) (86.2-91-3) 735 78"6 (718-746) (71-3-86.2) 938 96.2 (513 1441) (89.4~228.3) 194.5 67.6 (182-206.5) (62.3-69.8) 114-4 60.9 (103.7-118) (60.1-61.8) 288.8 7.7 (286-294.5) (7.6-7.9) 7167 2161 (7087-7207) (2132-2220) 166.6 14.8 (160.2 170-5) (14.6-14.9) 243.2 44'7 (240.2-246-2) (42.3 4 5 . 9 ) 198.2 221.2 (188-8211.6) (148.9-276.1)
51.5
--
(48.5-59.8) 48.0 (38-1 64.5)
. --
Cadmium
1.1 (1.0-1.4) 1.2 (1-1-1-3) 1.1 (0.9-1.3) 1,3 (0.2--3.8) 0.4 (0.3-0.8) 0.2 (0.1-0.3) 0.4 (0-3-0.4) 3.0 (2.9 3.2) 1-0 (0-8-1.2) 15.6 (13.2-17-7) nd
--
.
.
. ---
--
~Values are the mean (n = 10) and range in parentheses. bCollected at Station 5 unless otherwise shown. nd = Not detected. --- = Not analysed. a c c u m u l a t i o n o f the elements analysed. A n o t h e r bivalve, M y t i l u s edulis, c o n t a i n e d low levels o f Zn a n d Cu. C a d m i u m was c o n c e n t r a t e d to similar levels, a p p r o x i m a t e l y 1 p p m , in all a n i m a l species with slightly higher levels in m a c r o p h y t e s . The limpet P a t e l l a v u l g a t a c o n c e n t r a t e s C d to 15 p p m c o m p a r e d with the 3 p p m seen in the E n t e r o m o r p h a on which it feeds. M a c r o p h y t e s were f o u n d to be i m p o r t a n t a c c u m u l a t o r s o f As (Table 5) as r e p o r t e d by Jones (1922), V i n o g r a d o v (1953) a n d L u n d e (1970). M e a n levels a p p r o a c h i n g 100 p p m A s were f o u n d in the fucoides while levels in excess o f this occur in those m a c r o p h y t e s growing in A s - r i c h waters o f the e s t u a r y (Table 5). The A s c o n c e n t r a t i o n s showed c o n s i d e r a b l e v a r i a b i l i t y within a n d a m o n g s t species, with algae having the greatest r a n g e o f c o n c e n t r a t i o n . Both P a t e l l a a n d the snail L i t t o r i n a o b t u s a t a graze u p o n the m a c r o p h y t e s yet show no b i o m a g n i f i c a t i o n o f As on an entire a n i m a l basis. G a s t r o p o d s f r o m v a r i o u s t r o p h i c levels a c c u m u l a t e d As to the same extent. A l t h o u g h A s is a c c u m u l a t e d by higher t r o p h i c levels this element does n o t a p p e a r to b i o m a g n i f y up the f o o d chain. A t this stage it is not k n o w n w h e t h e r w a t e r o r f o o d is the m a i n source o f such a c c u m u l a t e d As.
17
ARSENIC AND METALS IN ESTUARINE ORGANISMS
TABLE 5 ARSENIC IN O R G A N I S M S FROM D I F F E R E N T L O C A T I O N S IN R E S T R O N G U E T CREEK
Location b
3J 3D 4J 4D 5J 5D 6J 6D
Fucus serratus
Fucus vesiculosus
* * 143.2 147.0 140.1 76.9 62.4 55" 1
184.4 79.5 117.6 88.3 104"2 38"5 * 46.3
T o t a l As (ppm d r y wt) ~ Fucus Ascophyllum Enterospiralis morpha
173.6 71.2 * 68.1 100.6 41.8 * 45.2
79-1 87.4 92-1 52.2 93.5 88-4 32"7 45.1
113"6 102.1 84.9 *. 93-7 82.6 18"0 15.3
Carciuus
Linorina
54.1 61.8 * * 50.1 * 27-0 22-4
97.7 84.6 93.8 63.5 86-2 55.5 * 46./t
" V a l u e s are the m e a n (n = 5). hSee Fig. 1 for location. J is near low and D near high tide level. * N o t o c c u r r i n g in zone at time of collection.
The trend in As concentration in selected species with position in the estuary on a horizontal and vertical basis is presented in Table 5. Macrophytes, Littorina and the crab Carcinus maenas were widely distributed throughout the estuary and, therefore, provided the best comparison. The As concentrations in these organisms decrease with distance down the estuary, apparently reflecting As levels in the water. In contrast, a Newfoundland study, including some of the above species, found considerably lower As levels in water and organisms, with no enrichment of As in organisms close to the source (Penrose et al., 1975). Separate parts of the thallus of the macrophytes Fucus t~esiculosus, F. serratus and Ascophyllum nodosum were analysed for their As content (Fig. 2). Except for slightly elevated levels in the basal stem of Ascophyllum, and the reproductive tissue of F. t,esiculosus, there appears to be a uniform concentration of As throughout the thallus. When water/methanol extracts of F. spiralis, Ascophyllum, Littorina and Nucella lapillus were analysed it was found that only trace amounts of the total As could be detected (Table 6). This was mainly in the form of arsenate with traces of arsenite and dimethylarsinate. Alkali digestion of these extracts released about 60 i~,' of the As in Fucus, Ascophyllum, and Littorina and 50 °/o in Nucella, and this was mainly in the form ofdimethylarsinate. Treatment with concentrated nitric acid/concentrated perchloric acid left most of the As as dimethylarsinate. This shows that the organoAs compounds in nature can be degraded to simple methylated derivatives. This has also been shown by Crecelius (1977). Furthermore, there are analytical complications which arise from these results. Direct atomic absorption determination of As in tissue by hydride generation will underestimate levels of this element, depending on the completeness of organic degradation of the sample. The problem is that sodium borohydride generates the As species at different rates. The solution would be to relate As concentration to peak area or ensure complete oxidisation of the sample.
18
D.W.
K L U M P P , P. J. P E T E R S O N
66 7 03"4) 76"9 19"1)
60-6 (6"9)
83"0
(7"5)
Fucus serratus 82"9
(7"2)
~ ~ p l
57.8
76.8 I t7.8)
70.9 (12.31.
(5.4)
L_._1
82"5 (9.2)
E
8O-'9 (5"71
Ascophyllum 74"4(5.0)Fucus vesiculosus 82.3 Fig. 2.
(8.1)
A r s e n i c ( p p m ) in the m a c r o p h y t e t h a l l u s - - m e a n (n: I0) w i t h s t a n d a r d e r r o r .
TABLE
6
ARSENIC FORMS IN TREATED WATER/METHANOL EXTRACTS
Species F. spiralis Ascophyllum Littorina Nucella
Concentration a (pg/g dry wt) Water~methanol Alkali hydrolysed AsV A s l l l MA DMA AsV MA DMA 0.25 0.14 0.43 0-73
AsV arsenate. MA methylarsonate. -- = Not detected.
0.03 0.02 0"11 0.25
-----
0.0018 0.0012 ---
2-1 2.5 3.3 3-1
0-4 -0.2 --
66.2 56.4 20.2 5.7
A s l l l arsenite. DMA dimethylarsinate.
Nitric/perchloric acid AsV MA DMA 1.2 2-5 2.8 1.9
1.7 2.1 0-9 0.5
105.4 82-2 34.4 15.3
ARSENIC AND METALSIN ESTUARINEORGANISMS
19
Prolonged nitric/perchloric acid digestion or dry ashing, as proposed by Leblank & Jackson (1973), will ensure this.
CONCLUSIONS
Data from this study on the relative proportions of arsenic species in waters support the view that arsenate is the stable form of this element in seawater. The results presented demonstrate that arsenic is accumulated by marine organisms relative to levels in their environment with concentration factors of up to 10,000 reported. The effects of such levels of concentration upon aquatic organisms cannot be adequately tested until the chemical forms of assimilated arsenic are known. It has been established that arsenic in marine organisms exists mainly in organic forms that can be hydrolysed to methylated arsenicals. There is no evidence for bioaccumulation of arsenic in food chains. However, it remains to be shown whether food or water is the important source of this element to marine animals. This and other studies of aquatic ecosystems indicate that there is an arsenic cycle in which this element undergoes several chemical transformations. In order to characterise this cycle, further study is required on the bioconcentration kinetics of arsenic and those factors affecting it.
ACKNOWLEDGEMENTS
The authors are grateful for financial support from Westfield College during the course of this work.
REFERENCES ANDREAE, M. O. (1977). Determination of arsenic species in natural waters. Analyt. Chem., 49, 820-3. ANDREAE,M. O. (1978). Distribution and speciation of arsenic in natural waters and some marine algae. Deep-Sea Res., 25, 391-402. BOWEN, H. J. M. (1974). Problems in the elementary analysis of standard biological materials. J. Radioanalyt. Chem., 19, 215-26. BRAMAN,R.. S. & FOREBACK,C. C. (I 974). Methylated forms of arsenic in the environment. Science, N. Y., 82, 1247-9. CRECELIUS, E. A. (1977). Changes in the chemical speciation of arsenic following ingestion by Man. Environ. Hlth Perspectives, 19, 147-52. EDMONDS)J. S., FRANCESCONI,K. A., CANNON, J. R., RASTON,C. L., SKELTON,B. W. • WHITE, A. H. (1977). Isolation, crystal structure and synthesis of arsenobetaine, the arsenical constituent of the western rock lobster, Panulirus longipes. Tetrahedron Letters, 18, 1543-6. HOSKING, K. F. G. & OmAL, R. (1966). A preliminary study of the distribution of certain metals of economic interest in the sediments and waters of Carrick, and of its "feeder' rivers. Camborne Sch. Mines Mag., 66, 17-37. JOHNSON, D. L. (1971). Simultaneous determination of arsenate and phosphate in natural waters. Environ. Sci. & Technol., 5, 411 14.
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D. W. KLUMPP, P. J. PETERSON
JOHNSON, D. L. (1972). Bacterial reduction of arsenate in sea water. Nature, Lond., 240, 44-5. JONES, A. J. (1922). The arsenic content of some of the marine algae. Pharm. J., 109, 86-7. LEBLANK,P. J. & JACKSON,A. L. (1973). Arsenic in marine fish and invertebrates. Mar. Pollut. Bull., 4(6), 88-90. LVNDE, G. (1970). Analysis of trace elements in seaweed. J. Sci. Food Agric., 21,416 18. LUNDE, G. (1977). Occurrence and transformation of arsenic in the marine environment. Environ. Hlth Perspectiees, 19, 47 52. MULVORP, C. E. (1966). Solvent extraction techniques for atomic absorption spectroscopy. At. Absorption Newsl., 5, 88-90. NATIONAL BUREAU Or STANDARDS(1976). Certificate of analysis standard reference material 1571. Washington, DC. US Department of Commerce. ONIsm, H. (1969). Arsenic. In Handbook of geochemistry, ed. by K. H. Wedepohl, 2(3), 33-B-1-33-O-1. Berlin, Springer-Verlag. PENROSE, W. R. (1975). Biosynthesis of organic arsenic compounds in brown trout (Salmo trutta). J. Fish. Res. Bd Can., 32, 2385-90. PENROSE,W. R., BLACK,R. & HAYWARD,M. J. (1975). Limited arsenic dispersion in seawater, sediments and biota near a continuous source. J. Fish. Res. Bd Can., 32, 1275-81. PENROSE,W. R., CONACHER,H. B. S., BLACK,R., MERANGER,J. C., MILES,W., CUNNINGHAM,H. M. & SQVmES, W. R. (1977). Implications of inorganic/organic interconversion on fluxes of arsenic in marine food webs. Ent,iron. HIth Perapectit~es, 19, 53-9. PORTER, E. K. & PETERSON, P. J. (1975). Arsenic accumulation by plants on mine-waste (United Kingdom). J. Sci. Tot. Environ., 4, 365-71. PORTER, E. K, & PETERSON,P. J. (1977). Arsenic tolerance in grasses growing on mine waste. Environ. Pollut., 14, 255 65. STRICKLAND,J. D. H. & PARSONS,T. R. (1968). A practical handbook of seawater analysis. Bull. Fish Res. Bd Can., 167. THOMPSON, K. C. & THOMERSON, D. R. (1974). Atomic-absorption studies on the determination of antimony, arsenic, bismuth, germanium, lead, selenium, tellurium and tin by utilizing the generation of covalent hydrides. Analyst, Lond., 99, 595-601. THOMPSON,A. J. & THORESBY,A. (1977). Determination of arsenic in soil and plant materials by atomicabsorption spectrophotometry with electrothermal atomization. Analyst, Lond., 102, 9-16. THORNTON, 1., WATLING,H. & DARRACOTT,A. (1975). Geochemical studies in several rivers and estuaries used for oyster rearing. J. Sci. Tot. Ent,iron., 4, 325-45. VINOGRADOV,A. P. (1953). Arsenic in various algae. In The elementary chemical composition of marine organisms. Mar. Res. Mere., 2, New Haven, Sears Found. Y1M, W. S. (1972). A further investigation on the distribution of certain elements in the sediments of the Fal Estuary, Cornwall. Camborne Sch. Mines Mag., 66, 17-37.
D. W. KLUMPP & P. J. PETERSON
Department of Botany and Biochemistry, Westfield College, Kidderpore Arenue, London NW3 7ST, Great Britain
ABSTRACT
Arsenic, cadmium, copper and zinc occur at elevated levels in the waters of Carnon River and the upper reaches of Restronguet Creek, these concentrations decreasing progressively towards the estuary of Carrick Roads. While arsenite is the main form of As in the Carnon River, this is converted to arsenate in the lower estuary. Organisms analysed from Restronguet Creek reflect the As concentration in the water, this being most clearly evident in macrophytes. Although accumulated at all trophic levels, there was no evidence for biomagnification of As on an entire organism basis. The As was shown to exist in selected organisms as a compound(s) whichjorms on hydrolysis, dimethylarsinate and traces of arsenate and methylarsonate.
INTRODUCTION
Our knowledge of the biological cycling of arsenic in marine systems is limited. With the aid of recent improvements in the methods of As analysis it has been shown that As occurs in marine waters predominantly as arsenate with traces of arsenite, methylarsonic acid and dimethylarsinic acid (Braman & Foreback, 1974; Andreae, 1977, 1978). Arsenic is accumulated by a wide variety of marine organisms but in view of the known toxicity of high concentrations of arsenite and arsenate, it is unlikely to be accumulated in this form. Indeed, several workers have extracted organo-arsenic compounds from marine organisms and this information is summarised by Lunde (1977). Edmonds et al. (1977) have identified arsenobetaine in extracts of lobster. Reports that As is rapidly excreted by some marine organisms (Penrose, 1975; 11 Environ. Pollut. 19 (1979)--© Applied Science Publishers Ltd, England, 1979 Printed in Great Britain
12
D. W. KLUMPP, P. J. PETERSON
Penrose et al., 1977) support the view that this element is not bioaccumulated up the food chain. We have been examining the accumulation and metabolism of As in an estuarine food chain in Restronguet Creek and Carrick Roads. Pollution by As in the terrestrial area has already been reported from this laboratory (Porter & Peterson, 1975, 1977). The region drained by the Carnon River has a long history of mining activity dating back to the tin and copper mines of the Roman era. It extends to the present as alluvial tin operations and the re-opened tin mines at Mount Wellington and Wheal Jane. These activities, in addition to the natural processes of weathering, have resulted in considerable contamination by As and some trace metals in the waters and sediments of the Carnon River and its estuary (Hosking & Obial, 1966; Yim, 1972; Thornton et al., 1975). We report here the results of a survey where the concentrations of As, Cd, Cu and Zn were measured in waters and organisms from the Restronguet Creek system. The speciation of As in waters and its metabolism in several organisms are also mentioned.
MATERIALS AND METHODS
Water and selected organisms from the estuary (Fig. 1) were analysed for As and, on occasions, for Zn, Cu and Cd between November 1975 and September 1977. Observed effects of storage on the As species in water samples were essentially the same as those reported by Andreae (1977), although loss of arsenite was more rapid in the present study. To minimise changes, water samples were filtered (millipore HA, 0.45 pm) into polyethylene bottles in the field and analysed within 24h of collection. Intertidal organisms collected for analysis were transported to the laboratory, washed in filtered seawater and frozen until required. Samples of approximately 1 g dry weight (80 °C) were wet digested in concentrated nitric acid (BDH, AnalaR) followed by a 4:1 mixture of nitric and perchloric acid (BDH, AnalaR) until a clear solution was obtained. Final solutions were taken up in 1.5N hydrochloric acid. Reagent blanks were included in all digestion batches. Analytical procedures were tested by a comparative analysis of the standard materials Bowen's Kale and NBS Orchard Leaves. Results are presented for Zn, Cu, Cd and As in Bowen's Kale, and As in NBS Orchard Leaves (Table 1). Results obtained were in close agreement with those reported for both sample materials. A recovery of 92( + 2) ~ was obtained for As-spiked Kale.. Until May 1976 all water samples were treated by extraction of the APDCcomplex into chloroform as described by Mulford (1966). Arsenite was extracted from one aliquot while a second was reduced, then extracted, to determine total As (Strickland & Parsons, 1968). Elements were measured on a Varian Tectron AA5
ARSENIC AND METALS IN ESTUARINE ORGANISMS
5010' ¢
05' T
13
I"-"
sd
15/
Wheal
~, Jane Mt. Wel lington
1 I
16
0
2
I
I
kilometre
~
"~,~ ~/~
~~o
Falml English Channel STUDY AREA Fig. 1.
Map of the study site with sampling locations.
TABLE 1 C O M P A R A T I V E ANALYSIS W I T H S T A N D A R D REFERENC|" MATERIALS B O W E N ' S KALE u ~.ND NBS O R C H A R I ) [.IL~,VI-S b (IN PPM DRY W T )
Material
Kale
Orchard Leaves
Element
As Zn Cu Cd As
Present study ~
0.125 28.4 5"4 0.64 10-0
(0"012) (0,8) (0'5) (0,13) (0,1)
Bowen (1974). NBS (1976). "Values are the mean (n = 10) with the standard deviation in parentheses.
ReJerence ~'h
0.141 33.2 4-99 0.80 10.0 (20)
14
D.W. KLUMPP, P. J. PETERSON
atomic absorption spectrophotometer with hydride generation and electrically heated furnace adaptation for As determination ( T h o m p s o n & Thomerson, 1974; T h o m p s o n & Thoresby, 1977). F r o m M a y 1976, methylarsonate and dimethylarsinate, in addition to arsenate and arsenite, were measured in water using the more sensitive m e t h o d o f Andreae (1977). With A A S application o f this method, detection limits for arsine, m o n o - and dimethylarsine were 0.5, 0.7 and 1-2 ng, or 20, 28 and 48 ng/litre with a 25 ml sample. Selected organisms were homogenised in a blender with a 1:2 mixture o f water/methanol. The lipid fraction was separated by extracting with four volumes o f chloroform. The water/methanol fraction was analysed for As before and after boiling for 1 h with 1N sodium hydroxide. Part o f the initial extract was refluxed with concentrated nitric acid for 24h, taken to dryness, then dissolved in l'5N hydrochloric acid and analysed as previously described.
RESULTS AND DISCUSSION E l e m e n t in w a t e r s
Dissolved As levels (Table 2) in the waters o f the lower estuary (Fig. 1) are close to published figures (Onishi, 1969; Andreae, 1978). In comparison, As was at considerably elevated concentrations in the upper reaches o f Restronguet Creek and in the C a r n o n River (Table 2). The high concentration o f As seen for station 1 in C a r n o n River water decreases markedly a r o u n d station 2 where river and marine waters mix. The d r o p in concentrations shown at station 2 m a y be due to dilution o f As-rich river water with that o f marine origin, a l t h o u g h precipitation o f the element TABLE 2 DISSOLVED ARSENIC SPECIES IN WATERS a W I T H LOCATION IN T H E C A R N O N ESTUARY SYSTEM
Station b'C
1 2 LT 2 HT 3LT 3 HT 4LT 4 HT 5 LT 5 HT 6LT 6 HT 7 LT 7 HT
January 1976
Concentration (#g/litre) May 1976
March 1977
25"7 (16) 16.0 (19) 3-5 (63) 11.8 (35) 4.0 (95) 9.0 (40) 5.0 (96) 5.9 (86) 3.5 (94) 4.1 (61) 2.8 (100) 2.3 (100) 2.3 (100)
42-1 (5) 22.6 (7) 10.6 (60) 14.9 (22) 4.6 (72) 8.1 (53) 5.2 (86) 7.1 (48) 3.4 (88) 6-8 (63) 3.7 (92) 4-8 (100) 1.9 (100)
34"3 (9) 18.3 (0) 7.9 (64) 15-2 (33) * 11.6 (71) 6.7 (100) 9.9 (84) 1.8 (100) 9.9 (66) 2.6 (100) 8.3 (100) 4.8 (100)
aValues are the mean (n --- 5) total arsenic (arsenate + arsenite) with % arsenate in parentheses. bRefer to Fig. 1 for location. cCollected at high tide (HT) and low tide (LT). * No data.
ARSENIC AND METALS IN ESTUARINE ORGANISMS
15
out of solution may be important. Mean values of over 1000 ppm As are reported (Thornton et al., 1975) in sediments of this estuary, providing a possible source of As to the water. Arsenate is the major form of As detected in the waters of the estuary, while arsenite predominates in the Carnon River. Arsenite occurs in waters of the estuary in decreasing concentrations towards the sea. Samples of water taken at low tide at station 2 contain mainly arsenite but at high tide arsenate predominates. Levels of both inorganic species are important at stations 3 and 4 during low tide while stations further down the estuary are consistently low in arsenite. Arsenite was not detected at high or low tide in the main estuary of Carrick Roads. These findings are summarised in Table 2. Results suggest that As enters the system as arsenite in Carnon River waters and is then either converted to arsenate or diluted when mixed with saline waters. Both Johnson (1971) and Andreae (1977, 1978) have reported arsenate as the main form of As in marine waters. The presence of arsenite in marine waters is thought to be a product of biological action (Johnson, 1972). Indeed, phytoplankton grown at natural As levels have been shown to release arsenite and methylated arsenics into water (details to be published later). Methylated As species were not detected in open waters of the estuary. However, dimethylarsonate (0.002 ppm) was found in one sample of rock pool water. This initiated a more thorough examination of waters from rock pools at a range of tidal heights and with varying degrees of algal growth. Methyl arsenics were not detected and the pools were found to contain As as arsenate and arsenite at levels seen in the estuary for that tidal position (see Table 2). Concentrations ofCd, Cu and Zn in waters of this system agree with that reported by Thornton et al. (1975). Cadmium, Cu and Zn were more heavily concentrated in the Carnon River compared with the estuary (Table 3). Elements in organisms High levels of Zn and Cu were found in the oyster Ostrea edulis and to a lesser extent in macrophytes (Table 4). Enteromorpha intestinalis showed the least TABLE 3 DISSOLVED Z I N C , C O P P E R A N D C A D M I U M IN WATERS a OF T H E C A R N O N ESTUARY SYSTEM
Location River C a r n o n Stn 1 Restronguet Creek (low tide) Stn 4 Carrick Roads (low tide) Stn 6
Zinc
Concentration (~g/litre) Copper
Cadmium
5820 3692-22000) 285 (12.9-305) 230 (16-500)
236 (32.5 395.7) 16 (2.1 21-0) 13 (2.8 20)
5.3 (4.3 7.4) 2.8 (nd 3-71 2"6 (nd 3.1)
~Values are the mean and range in parentheses from samples collected over 1976 77. n d = Not detected.
PETERSON
D. W . KLUMPP, P. J.
16
TABLE 4 ELEMENTS IN THE ENTIRE SOFT PARTS OF S E L E C T E D ORGANISMS FROM RESTRONGUET CREEK
Speciesb Arsenic Fucus serratus F. vesiculosus F. spiralis Ascophyllum nodosum Peh,etia canaliculata Enteromorpha intestinalis Tealiafelina
Stn 6 Ostrea edulis Mytilus edulis Patella vulgata
Stn 6 Carcinus maenas Littorina obtusata Nucella lapillus
141.4 (103.3-189.3) 105.3 (94.7-126.0) 103.6 (96.3-113.4) 91.7 (69-7--118.3) 65.0 (59.1-71.0) 83.7 (83.3-85-5) 55.2 (53.3 5 9 . 0 ) 17.0 (16.9-17.2) 16.9 (16.8-17.1) 36.7 (35.3 41.0) 22.3 (19.9-34,5)
Concentration (ppm dry wt) a Zinc Copper
771 65 (727-1321) (62.4~68.2) 719 89 (709.8 7 3 8 ) (86.2-91-3) 735 78"6 (718-746) (71-3-86.2) 938 96.2 (513 1441) (89.4~228.3) 194.5 67.6 (182-206.5) (62.3-69.8) 114-4 60.9 (103.7-118) (60.1-61.8) 288.8 7.7 (286-294.5) (7.6-7.9) 7167 2161 (7087-7207) (2132-2220) 166.6 14.8 (160.2 170-5) (14.6-14.9) 243.2 44'7 (240.2-246-2) (42.3 4 5 . 9 ) 198.2 221.2 (188-8211.6) (148.9-276.1)
51.5
--
(48.5-59.8) 48.0 (38-1 64.5)
. --
Cadmium
1.1 (1.0-1.4) 1.2 (1-1-1-3) 1.1 (0.9-1.3) 1,3 (0.2--3.8) 0.4 (0.3-0.8) 0.2 (0.1-0.3) 0.4 (0-3-0.4) 3.0 (2.9 3.2) 1-0 (0-8-1.2) 15.6 (13.2-17-7) nd
--
.
.
. ---
--
~Values are the mean (n = 10) and range in parentheses. bCollected at Station 5 unless otherwise shown. nd = Not detected. --- = Not analysed. a c c u m u l a t i o n o f the elements analysed. A n o t h e r bivalve, M y t i l u s edulis, c o n t a i n e d low levels o f Zn a n d Cu. C a d m i u m was c o n c e n t r a t e d to similar levels, a p p r o x i m a t e l y 1 p p m , in all a n i m a l species with slightly higher levels in m a c r o p h y t e s . The limpet P a t e l l a v u l g a t a c o n c e n t r a t e s C d to 15 p p m c o m p a r e d with the 3 p p m seen in the E n t e r o m o r p h a on which it feeds. M a c r o p h y t e s were f o u n d to be i m p o r t a n t a c c u m u l a t o r s o f As (Table 5) as r e p o r t e d by Jones (1922), V i n o g r a d o v (1953) a n d L u n d e (1970). M e a n levels a p p r o a c h i n g 100 p p m A s were f o u n d in the fucoides while levels in excess o f this occur in those m a c r o p h y t e s growing in A s - r i c h waters o f the e s t u a r y (Table 5). The A s c o n c e n t r a t i o n s showed c o n s i d e r a b l e v a r i a b i l i t y within a n d a m o n g s t species, with algae having the greatest r a n g e o f c o n c e n t r a t i o n . Both P a t e l l a a n d the snail L i t t o r i n a o b t u s a t a graze u p o n the m a c r o p h y t e s yet show no b i o m a g n i f i c a t i o n o f As on an entire a n i m a l basis. G a s t r o p o d s f r o m v a r i o u s t r o p h i c levels a c c u m u l a t e d As to the same extent. A l t h o u g h A s is a c c u m u l a t e d by higher t r o p h i c levels this element does n o t a p p e a r to b i o m a g n i f y up the f o o d chain. A t this stage it is not k n o w n w h e t h e r w a t e r o r f o o d is the m a i n source o f such a c c u m u l a t e d As.
17
ARSENIC AND METALS IN ESTUARINE ORGANISMS
TABLE 5 ARSENIC IN O R G A N I S M S FROM D I F F E R E N T L O C A T I O N S IN R E S T R O N G U E T CREEK
Location b
3J 3D 4J 4D 5J 5D 6J 6D
Fucus serratus
Fucus vesiculosus
* * 143.2 147.0 140.1 76.9 62.4 55" 1
184.4 79.5 117.6 88.3 104"2 38"5 * 46.3
T o t a l As (ppm d r y wt) ~ Fucus Ascophyllum Enterospiralis morpha
173.6 71.2 * 68.1 100.6 41.8 * 45.2
79-1 87.4 92-1 52.2 93.5 88-4 32"7 45.1
113"6 102.1 84.9 *. 93-7 82.6 18"0 15.3
Carciuus
Linorina
54.1 61.8 * * 50.1 * 27-0 22-4
97.7 84.6 93.8 63.5 86-2 55.5 * 46./t
" V a l u e s are the m e a n (n = 5). hSee Fig. 1 for location. J is near low and D near high tide level. * N o t o c c u r r i n g in zone at time of collection.
The trend in As concentration in selected species with position in the estuary on a horizontal and vertical basis is presented in Table 5. Macrophytes, Littorina and the crab Carcinus maenas were widely distributed throughout the estuary and, therefore, provided the best comparison. The As concentrations in these organisms decrease with distance down the estuary, apparently reflecting As levels in the water. In contrast, a Newfoundland study, including some of the above species, found considerably lower As levels in water and organisms, with no enrichment of As in organisms close to the source (Penrose et al., 1975). Separate parts of the thallus of the macrophytes Fucus t~esiculosus, F. serratus and Ascophyllum nodosum were analysed for their As content (Fig. 2). Except for slightly elevated levels in the basal stem of Ascophyllum, and the reproductive tissue of F. t,esiculosus, there appears to be a uniform concentration of As throughout the thallus. When water/methanol extracts of F. spiralis, Ascophyllum, Littorina and Nucella lapillus were analysed it was found that only trace amounts of the total As could be detected (Table 6). This was mainly in the form of arsenate with traces of arsenite and dimethylarsinate. Alkali digestion of these extracts released about 60 i~,' of the As in Fucus, Ascophyllum, and Littorina and 50 °/o in Nucella, and this was mainly in the form ofdimethylarsinate. Treatment with concentrated nitric acid/concentrated perchloric acid left most of the As as dimethylarsinate. This shows that the organoAs compounds in nature can be degraded to simple methylated derivatives. This has also been shown by Crecelius (1977). Furthermore, there are analytical complications which arise from these results. Direct atomic absorption determination of As in tissue by hydride generation will underestimate levels of this element, depending on the completeness of organic degradation of the sample. The problem is that sodium borohydride generates the As species at different rates. The solution would be to relate As concentration to peak area or ensure complete oxidisation of the sample.
18
D.W.
K L U M P P , P. J. P E T E R S O N
66 7 03"4) 76"9 19"1)
60-6 (6"9)
83"0
(7"5)
Fucus serratus 82"9
(7"2)
~ ~ p l
57.8
76.8 I t7.8)
70.9 (12.31.
(5.4)
L_._1
82"5 (9.2)
E
8O-'9 (5"71
Ascophyllum 74"4(5.0)Fucus vesiculosus 82.3 Fig. 2.
(8.1)
A r s e n i c ( p p m ) in the m a c r o p h y t e t h a l l u s - - m e a n (n: I0) w i t h s t a n d a r d e r r o r .
TABLE
6
ARSENIC FORMS IN TREATED WATER/METHANOL EXTRACTS
Species F. spiralis Ascophyllum Littorina Nucella
Concentration a (pg/g dry wt) Water~methanol Alkali hydrolysed AsV A s l l l MA DMA AsV MA DMA 0.25 0.14 0.43 0-73
AsV arsenate. MA methylarsonate. -- = Not detected.
0.03 0.02 0"11 0.25
-----
0.0018 0.0012 ---
2-1 2.5 3.3 3-1
0-4 -0.2 --
66.2 56.4 20.2 5.7
A s l l l arsenite. DMA dimethylarsinate.
Nitric/perchloric acid AsV MA DMA 1.2 2-5 2.8 1.9
1.7 2.1 0-9 0.5
105.4 82-2 34.4 15.3
ARSENIC AND METALSIN ESTUARINEORGANISMS
19
Prolonged nitric/perchloric acid digestion or dry ashing, as proposed by Leblank & Jackson (1973), will ensure this.
CONCLUSIONS
Data from this study on the relative proportions of arsenic species in waters support the view that arsenate is the stable form of this element in seawater. The results presented demonstrate that arsenic is accumulated by marine organisms relative to levels in their environment with concentration factors of up to 10,000 reported. The effects of such levels of concentration upon aquatic organisms cannot be adequately tested until the chemical forms of assimilated arsenic are known. It has been established that arsenic in marine organisms exists mainly in organic forms that can be hydrolysed to methylated arsenicals. There is no evidence for bioaccumulation of arsenic in food chains. However, it remains to be shown whether food or water is the important source of this element to marine animals. This and other studies of aquatic ecosystems indicate that there is an arsenic cycle in which this element undergoes several chemical transformations. In order to characterise this cycle, further study is required on the bioconcentration kinetics of arsenic and those factors affecting it.
ACKNOWLEDGEMENTS
The authors are grateful for financial support from Westfield College during the course of this work.
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20
D. W. KLUMPP, P. J. PETERSON
JOHNSON, D. L. (1972). Bacterial reduction of arsenate in sea water. Nature, Lond., 240, 44-5. JONES, A. J. (1922). The arsenic content of some of the marine algae. Pharm. J., 109, 86-7. LEBLANK,P. J. & JACKSON,A. L. (1973). Arsenic in marine fish and invertebrates. Mar. Pollut. Bull., 4(6), 88-90. LVNDE, G. (1970). Analysis of trace elements in seaweed. J. Sci. Food Agric., 21,416 18. LUNDE, G. (1977). Occurrence and transformation of arsenic in the marine environment. Environ. Hlth Perspectiees, 19, 47 52. MULVORP, C. E. (1966). Solvent extraction techniques for atomic absorption spectroscopy. At. Absorption Newsl., 5, 88-90. NATIONAL BUREAU Or STANDARDS(1976). Certificate of analysis standard reference material 1571. Washington, DC. US Department of Commerce. ONIsm, H. (1969). Arsenic. In Handbook of geochemistry, ed. by K. H. Wedepohl, 2(3), 33-B-1-33-O-1. Berlin, Springer-Verlag. PENROSE, W. R. (1975). Biosynthesis of organic arsenic compounds in brown trout (Salmo trutta). J. Fish. Res. Bd Can., 32, 2385-90. PENROSE,W. R., BLACK,R. & HAYWARD,M. J. (1975). Limited arsenic dispersion in seawater, sediments and biota near a continuous source. J. Fish. Res. Bd Can., 32, 1275-81. PENROSE,W. R., CONACHER,H. B. S., BLACK,R., MERANGER,J. C., MILES,W., CUNNINGHAM,H. M. & SQVmES, W. R. (1977). Implications of inorganic/organic interconversion on fluxes of arsenic in marine food webs. Ent,iron. HIth Perapectit~es, 19, 53-9. PORTER, E. K. & PETERSON, P. J. (1975). Arsenic accumulation by plants on mine-waste (United Kingdom). J. Sci. Tot. Environ., 4, 365-71. PORTER, E. K, & PETERSON,P. J. (1977). Arsenic tolerance in grasses growing on mine waste. Environ. Pollut., 14, 255 65. STRICKLAND,J. D. H. & PARSONS,T. R. (1968). A practical handbook of seawater analysis. Bull. Fish Res. Bd Can., 167. THOMPSON, K. C. & THOMERSON, D. R. (1974). Atomic-absorption studies on the determination of antimony, arsenic, bismuth, germanium, lead, selenium, tellurium and tin by utilizing the generation of covalent hydrides. Analyst, Lond., 99, 595-601. THOMPSON,A. J. & THORESBY,A. (1977). Determination of arsenic in soil and plant materials by atomicabsorption spectrophotometry with electrothermal atomization. Analyst, Lond., 102, 9-16. THORNTON, 1., WATLING,H. & DARRACOTT,A. (1975). Geochemical studies in several rivers and estuaries used for oyster rearing. J. Sci. Tot. Ent,iron., 4, 325-45. VINOGRADOV,A. P. (1953). Arsenic in various algae. In The elementary chemical composition of marine organisms. Mar. Res. Mere., 2, New Haven, Sears Found. Y1M, W. S. (1972). A further investigation on the distribution of certain elements in the sediments of the Fal Estuary, Cornwall. Camborne Sch. Mines Mag., 66, 17-37.