Bioavailability of organic pollutants in boreal waters with varying levels of dissolved organic material

W’al.Res. Vol. 25, No. 4, pp. 455-463, 1991 Britain. All rights reserved

Go43-1354/91 $3.00+ 0.00 Copyright Q 1991Pergamon Press plc

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BIOAVAILABILITY OF ORGANIC POLLUTANTS IN BOREAL WATERS WITH VARYING LEVELS OF DISSOLVED ORGANIC MATERIAL JUSSI KUKKONEN and AIMO OIKARI University of Joensuu, Department of Biology, P.O. Box 111, SF-80101 Joensuu, Finland (First received April 1990; accepted in revised form November

1990)

Abstract-Dissolved organic matter (DOM) in 20 surface waters in Eastern Finland were characterized to examine relationships between structural and compositional properties of DOM and partition coefficients (K,) describing sorption of four model contaminants to DOM and the bioavailability of contaminants by Daphniu magna. The hydrophobic acids (HbA), hydrophobic neutrals (HbN) and hydrophilic (HI) fractions of DOM were separated by XAD-8 resin. The K,s were measured by equilibrium dialysis. Model contaminants were benzo(a)pyrene (BaP), naphthalene (NAPH), 3,3’,4,4’-tetrachlorobiphenyl flCB) and dehydroabietic acid (DHAA). DOM concentrations varied from 2.0 to 38.3 mg org. C/l in the water series. The percentage of HbA and the aromaticity of DOM, as indicated by the absorptivity at 270 nm (A,,,) and hydrogen/carbon ratio (H/C ratio), increased with increasing DOM concentration. Significant correlations were observed between !$ of BAP, A,,, and HbA content of the DOM from different sources. For the other contaminants similar kmds of relationships between KPs and quality parameters of DOM could not be found. The bioavailability of model compounds was decreased by increasing DOM concentration in the water series. For all four model contaminants, measured bioconcentration factors (BCF) correlated well with the A,,, of a water and HbA content of the DOM. These results show that the total DOM concentration is an important factor controlling the bioavailability of xenobiotics in natural waters. Besides the quantity also the quality of DOM, like proportion of HbA, can contribute in bioavailability. Ke_y words-dissolved Daphnia magna

organic material, humus, organic pollutants,

INTRODUCTION The pool of dissolved natural waters consists

organic material (DOM) in of a variety of organic mol-

ecules. While some of these molecules have a defined chemical structure, most of the organic material in natural waters has no readily identifiable structure, and this heterogeneous group of organic macromolecules are referred to as humic substances. DOM is an important factor in water chemistry and aquatic toxicology because it has been demonstrated that DOM can bind both metals (Alberts and Giesy, 1983) and hydrophobic organic pollutants such as DDT, polycyclic aromatic hydrocarbons (PAHs) polychlorinated biphenyls (PCBs) and polychlorinated dibenzodioxins and -furans (Gjessing and Bergling, 1981; Carter and Suffet, 1982; Hassett and Milicic, 1985; McCarthy and Jimenez, 1985; Servos and Muir, 1989). Both commercially available (Aldrich) humic acid preparations and natural water samples bind organic pollutants, and the magnitude of the binding, expressed as the partition coefficient (I$), is related to the hydrophobicity of the contaminant (McCarthy and Jimenez, 1985; Chiou et al., 1986). However, the affinity of the organic matter for binding a given contaminant appears to vary among waters from different sources (Carter and Suffet, 1982; Whitehouse,

biouvailability,

natural waters,

1985; Morehead et ai., 1986; Kukkonen et al., 1989). The underlying causes of the observed variability in binding affinity of different waters for organic contaminants is not fully understood, and hampers attempts to describe and predict the importance of natural organic matter in the transport and fate of organic pollutants in aquatic systems. DOM also affects the bioavailability of organic pollutants. Both Aldrich humic acid and natural water samples reduce the bioavailability of organic pollutants, and the magnitude of the decrease is related to the extent of the binding between the contaminant and the organic matter (McCarthy et al., 1985; Black and McCarthy, 1988; Kukkonen and Oikari, 1987; Kukkonen et al., 1989). Thus, our success to predict the significance of organic macromolecules in the accumulation and toxicity of hydrophobic organic contaminants in aquatic environments is dependent on, and limited by, the poorly understood variability in the binding affinity among natural waters. One approach to elucidate the source of this variability is to chemically fractionate DOM and to determine if there are underlying similarities in binding affinities of functionally similar subgroups of the total DOM. Nonionic macroporous sorbents such as Amberlite XAD resins have been used to fractionate DOC into fractions based on the hydrophobicity and 455

456

Ju~l KUKKONEN and AIMO OIKARI

charge of the molecules (Leenheer and Huffman, 1979). K u k k o n e n et al. (1990) have reported differences between three X A D - 8 fractions of natural D O M in their binding affinities for benzo(a)pyrene and 2,2',5,5'-tetrachlorobiphenyl. It was also shown that the measured binding (K v value) of the xenobiotics in original water samples can be counted as a sum of the relative Kp values of the fractions. On the other hand, Gauthier et al. (1987) have reported for pyrene and McCarthy et al. (1989) for benzo(a)pyrene that the binding coefficients for those compounds were correlated with the aromaticity of the D O M or with the relative abundance of different X A D - 8 fractions, respectively. It is important to study different types of pollutants because evidently their hydrophobicity is one of the main factors affecting the extent of interaction between pollutants and D O M . This generalization works in large scale or within one type of pollutant, like P A H s (McCarthy et al., 1985). But, the hydrophobicity of pollutants is not the only factor governing their interaction with D O M (Lee and Farmer, 1989; Kukkonen et al. (1990). The objective of this study is to extend these observations concerning the bioavailability of selected model compounds in a series of natural surface waters having a large variation in D O M concentrations. MATERIALS AND METHODS Chemicals and waters

Radiolabeled model compounds used in this study were [14C]benzo(a)pyrene (BaP; 52.0 mCi/mmol (1.92 GBq/mmol); Amersham, U.K.), [t4C]naphthalene [NAPH; 59.0 mCi/mmol (2.19 GBq/mmol); CEA, France], [14C]3,3',4,4'-tetrachlorobipbenyl [TCB; 37.1 mCi/mmol (1.37GBq/mmol); Sigma, U.S.A.], and [3H]dehydroabietic acid DHAA; 2.3 mCi/mmol (84.9 MBq/mmol); labeled according to Kutney et al. (1981)]. Table I lists some chemical characteristics of the model compounds. The stocks were diluted in ethanol for bioaccumulation experiments. Aqueous solutions of radiolabeled contaminants were prepared by addition of the stock carrier solution. The model compounds were chosen, because they represent environmentally important classes of contaminants (PAH, PCB and resin acid) and offer contrasts in their chemical structure, hydrophobicity and affinity for binding to dissolved organic material. Artificial organic-free control water was made up of Milli-Q grade water (DOC ~<0.3 mg C/l) and the following reagent-grade salts: CaC12.2I-I20, 58.8 rag/l; MgSO4' 7H20, 24.7 rag/l; KCI, 1.1 rag/l; and NaHCO3, 13.0 mg/l (Ca + Mg hardness = 0.5 raM, pH adjusted to 6.5). Natural waters used in the experiments were collected in July 1988 in North Karelia, Eastern Finland (Fig. 1). All waters were filtered (Whatman GF/C) within 24 h after sampling. Previous to experiments the samples were stored in brown glass bottles for 2-4 weeks in the dark at 4°C. Characteristics of the waters are shown in Table 2. Two of the samples were brook

Fig. 1. Map showing the origin of the sampled waters. The numbers are listed in the Table 2. waters having a very high DOM load. The catchment areas of these brooks are small and contain mainly forest or peat land (Ahtiainen et al., 1988). DOM of these brook waters can be considered as a material freshly extracted from the soil of the area. The rest of the samples were lake waters having a varying DOC concentration, pH and conductivity representing the variation of waters in the small lakes in Eastern Finland. Characterization o f D O M

The XAD-8 fractionation of DOM was performed to a slightly modified version of the method of Leenheer and Huffman (1979). Water samples were filtered through precombusted glass fiber filters (Whatman GF/C). Samples (150 ml) were acidified (pH ~<2, cone. H2SO4) and applied to the column of purified XAD-8 resin (5 ml bed volume) at a flow rate of 1.2 ml/min. The hydrophilic fraction (HI) of the DOM is defined as the fraction in the acidified sample not retained by the column. The hydrophobic acids fraction (HbA) is defined as the fraction eluted from the column with 0.1 N NaOH. The hydrophobic neutrals fraction (HbN) is defined as the fraction retained by the XAD-8 and not eluted with base. The dissolved organic carbon (DOC) concentration was measured after Salonen (1979). The waters were characterized by u.v.-vis spectroscopy. The absorbance of the samples was measured at 250, 270, 365, 465 and 665 nm at the original water pH using a Hitachi U-2000 spectrophotometer and 10ram quartz cuvettes.

Table I. Molecularweights, octanol-water partition coefficientsand solubilitiesof model compounds Compound MW log Ko~ Solubility Reference Bcnzo(a)pyrene 252 5.98 3.7 gg/l Miller et al. (1985) Naphthalene 128 3.3 30 mg/l Miller et al. (1985) 3,3',4,4'-Tetrachlorobiphenyl 292 6.5 10/~g/l Shiu and Mackay (1986) Dehydroabietic acid 300 4.8* 6.6 mg/I Nyr~nand Back 0958) *Professor B. Holmbom (University of Abo Akademi), personal communication.

Fate of organic pollutants in humic lakes

457

Table 2. Chemical characterization of natural waters used in the experiments(na = not analyzed). The number is the same as in Fig. 1 DOC (mgC/l)

Sample I. Brook Vfilioja 2. Brook Liuhapuro 3. Lake Ahvenlampi 4. Lake Louhilampi 5. Lake Mekrijiirvi 6. Lake lso-Sormunen 7. Lake Melalampi 8. Lake Piim/ijiirvi 9. Lake Koitere 10. Lake H6yti~iinen 11. Lake Riihilampi 12. Lake Viinij/irvi(point 5) 13. Lake Iso-Hietaj/irvi 14. Lake Tammalammit 15. Lake Valkialammit 16. Lake Viinij/irvi(point 1) 17. Lake Likolampi 18. Lake Miilunlampi 19. Lake Kakkisenlampi 20. Lake Kuorinka

38.3 32.2 20.4 18.1 15.1 13.5 9.5 9.6 7.5 7.5 6.8 8.4 6.0 5.0 4.9 4.2 4.9 2.8 2.0 3.0

pH 4.3 4.6 6.5 5.1 5.2 5.4 5.6 7.1 6.9 6.3 ' 5.6 6.9 6.7 5.6 5.8 7.4 5.6 6.9 4.9 6.6

Conductivity Color COD ( m S / m ) (ragPt/l) (mgO:/l) 3.2 2.7 5.4 2.3 2.8 1.8 1.8 5.7 2.1 4.7 2.1 6.7 1.7 2.1 1.8 6.7 1.9 7.2 1.3 3.7

350 400 120 200 180 100 70 60 70 50 65 na 30 25 10 10 20 20 10 5

Also, infrared spectra (Nicolet 20SXC FT-IR Spectrometer) and elemental composition (Carlo Erba elemental analyzer mod. 1106) of freeze dried samples were measured.

Determination of partition coeJ~cients (Kp) Equilibrium dialysis (Carter and Suffet, 1982; McCarthy and Jimenez, 1985) was used to determine the Kv between model compounds and the DOM. 5 ml of a sample at the ambient pH was added to a dialysis bag (Speetra/Por 6, mol. wt cutoff 1000 Da) and placed in a 200-ml glass jar containing radiolabeled compound dissolved in distilled water (180ml). Sodium azide (0.002%) was added to inhibit microbial activity. The jar was sealed with a Teflon-lined cap and shaken in the dark at 20°C for 4 days. At least three replicate determinations were made. After dialysis solutions inside and outside the dialysis bag were analyzed for ~4Cactivity using a scintillation cocktail (Luma Gel, Lumac, NL) and a liquid scintillation counter (1217 Racbeta, Wallac LKB, Finland). The outside concentration (Co, ng/ml) is the freely dissolved organic pollutant, while the difference between the inside and outside concentration (Cp, ng/ml) is the pollutant bound to organic matter in the bag. Kp was calculated as:

Kp = Cp/(C o × DOC)

(1)

where DOC is the concentration of dissolved organic carbon (kg carbon/l) and Kp has the unit of l/kg.

Accumulation experiments Water samples were filtered (Nucleopore, 0.22 #m) and pH adjusted to 6.5 with 0.1 N NaOH and HCI. Aqueous concentrations of BaP, TCB, NAPH and DHAA were l, 2, 5 and 70/~g/l, respectively, all well below the published water solubility limits for each compound (Table 1). D. magna were obtained from a culture maintained at the University of Joensuu and were fed with a culture of Monoraphidium contortum. Animals used in the experiments were approx. 7-8 days old and did not have eggs in the brood chamber. Before exposures, daphnids were held for I h in the clean control water to clear their gut contents. Groups of five D. magna were transferred to 100-ml glass beakers containing 50 ml water sample with one of the radiolabeled contaminants. Four replicate determinations were made for each sample. Beakers were kept in the dark at 20°C. After 24 h animals were removed from the water with a widemouthed pipet, collected on filter paper, briefly rinsed in 50 ml of distilled water, blotted dry and all animals from each beaker were weighed together on a microbalance (Sartorius model 4503). The five animals were added to 10 ml of Lipo Luma (Lumac), and analyzed for radioactivity. The radio-

64 22 24 na na 18 13 12 12 na 9.6 9.7 5.0 4.8 4.5 3.3 4.4 2.7 1.4 1.8

Fe (#g/I)

Mn (#g/l)

Tot. P (#g/l)

Tot. N (#g/I)

1750 1270 250 780 na 180 380 470 360 70 300 430 85 400 170 75 200 500 185 30

43 45 60 na na 48 54 90 50 na 45 na 14 90 60 12 110 130 24 na

45 50 22 na na 13 10 27 12 na 12 19 9 30 14 5 37 8 15 8

890 6 l0 1090 na na 460 290 700 250 na 250 450 185 400 400 280 590 200 170 260

activity remaining in the exposure water was determined. Each experiment included a parallel control experiment using organic-free control water. The results are reported as a 24-h bioconcentration factor (BCF) calculated as the ratio of the concentration of the pollutant in the animals (ng/g wet wt) and in the water after the experiment (ng/ml), calculated from the known specific activities of the compounds. The fitting of regression lines and all statistical analyses were performed using SAS software (SAS Institute Inc., 1985) on VAX 11/785 computer.

RESULTS AND DISCUSSION

Characterization o f D O M Dissolved organic matter ( D O M ) in 20 surface waters in Eastern Finland was characterized to examine relationships between structural and compositional properties o f D O M and partition coefficients describing sorption o f h y d r o p h o b i c model c o m p o u n d s and the bioavailability o f these c o m p o u n d s . D O M p r o p erties o f interest are shown in Table 3 and includes percentages o f different X A D - 8 fractions, absorptivity at 270 n m (A270), ratio o f absorbance at 250 n m and absorbance at 365 nm (E2/E3), ratio o f absorbance at 465 n m and absorbance at 665 nm (E4/Er) and h y d r o g e n / c a r b o n (H/C) atomic ratios. Percentages o f h y d r o p h o b i c acids (HbA) varied in the water series from 77% in dark colored b r o o k waters having a high D O C concentration to 19% in very clear water having a real low D O C concentration. Percentages o f hydrophobic neutrals (HbN) increased from 2% in dark colored b r o o k waters to 39% in the clearest lake water, but the trend is not that clear when compared to the decrease of the percentage o f H b A in the same series. The percentage of hydrophillics varied from 18 to 59%. The percentage of total hydrophobics varied from a b o u t 80% in the colored waters to a b o u t 50% in clear waters. The correlations between the X A D - 8 fractions and the other water quality parameters measured are given in the Table 4. The absorptivity o f D O M , E2/E3 ratio and E4/E 6 ratio varied in the sample series from 11 to 43, from

458

JuSSl KUKKONENand AIMO OIKARI

Table 3. Characterization of DOM in the waters studied Sample %HI* %HbAt %HbN %Tot. Hb~ A270¶ E2/E3]I EJEr** H/Ctt 1. Brook V/ilioja 23 75 2 77 37.9 4.47 9.55 0.80 2. Brook Liuhapuro 20 77 3 80 43.4 4.49 12.4 0.85 3. Lake Ahvenlampi 30 63 7 70 29.5 4.71 3.64 1.11 4. Lake Louhilampi 18 61 20 81 32.7 4.78 8.20 0.94 5. Lake Mekrij/irvi 27 63 10 73 35.9 4.49 2.65 0.95 6. Lake Iso-Sormunen 22 51 27 78 30.6 4.88 3.33 0.97 7. Lake Melalampi 28 59 13 72 35.3 4.90 2.78 1.08 8. Lake Piim~ij~irvi 25 55 20 75 31.7 5.17 2.56 1.37 9. Lake Koitere 34 51 I5 66 38.7 5.62 3.67 1.02 I0. Lake HiSyti~inen 29 40 31 71 36.9 5.63 4.00 1.18 11. Lake Riihilampi 32 44 24 68 35.1 4.63 2.50 0.98 12. Lake Viinij/irvi(point 5) 32 39 28 67 26.4 5.88 5.00 1.33 13. Lake Iso-Hietaj~irvi 39 41 20 61 18.3 6.36 1.00 1.14 14. Lake Tammalammit 36 46 17 63 22.1 4.34 1.86 1.20 15. Lake Valkialampi 35 45 20 65 18.5 4.46 1.71 1.19 16. Lake Viinij~irvi(point 1) 59 28 13 41 12.6 5.46 2.00 1.70 17. Lake Likolampi 48 22 30 52 10.8 4.25 1.50 1.46 18. Lake Miilunlampi 49 37 14 51 17.9 3.76 4.00 1.61 19. Lake Kakkisenlampi 49 38 13 51 22.4 3.29 1.43 1.71 20. Lake Kuorinka 42 19 39 58 13.4 3.79 1.33 na *Percentage of hydrophilic fraction; tpercentage of hydrophobic acids fraction, ~:pereentageof hydrophobic neutrals fraction; §percentage of hydrophobics (HbA + HbN); ¶absorptivity at 270 nm (units of l/mgC x crn x 103); IIE2/E3ratio is the ratio of the absorbances at 250 and 365 nm; **/~/E6 ratio is the ratio of the absorbances at 465 and 665 nm; ttH/C is the atomic ratio of hydrogen and carbon.

3.3 to 5.6 a n d from !.0 to 12.4, respectively. The h y d r o g e n c a r b o n ratio varied from 0.8 to 1.7 suggesting large variation from mainly a r o m a t i c c o m p o u n d s or conjugated alkenes to r a t h e r simple molecules. A c c o r d i n g to C h e n et al. (1977), a high E 4 / E 6 ratio is p r o p o r t i o n a l to the degree of humification a n d to the molecular weight of humic substances, as measured by the colligative properties of material extracted from soil. Schnitzer (1977) f o u n d t h a t the E4/E6 values for humic acids a n d fulvic acids extracted from soils, formed u n d e r widely differing conditions, are within the range of 3.8-5.8 a n d 7.6-11.5, respectively. According to De H a a n (1983), fulvic acids from strongly humic and oligotrophic waters are characterized by a E 2/E 3 ratio of a b o u t four. The absorptivity o f the sample at 2 7 0 n m (A270; in units o f 1/mgC x cm × 103) reflects a b s o r b a n c e of p i - p i ' transitions in substituted benzenes a n d m o s t polyenes a n d have been related to the a r o m a t i c content o f isolated soil humic acids ( G a u t h i e r et al., 1987; T r a i n a et al., 1990). The water samples having the highest D O M concentrations a n d representing " f r e s h " D O M extracted from the c a t c h m e n t area (Brooks L i u h a p u r o and V/ilioja) had an extremely low H b N portion and a high H b A p o r t i o n (Table 3). The correlations between the percentage of X A D - 8 fractions in the water samples a n d the s p e c t r o p h o t o m e t r i c p a r a m e t e r s show the d o m i n a n t role of H b A in the waters (Table 4). The increase o f D O M c o n c e n t r a t i o n is mainly due to

increasing a m o u n t o f H b A , which can be recognized as humic substances. This data suggests also t h a t the H b A fraction is enriched with a r o m a t i c constituents. The infrared spectra of the seven selected samples (Fig. 2) show characteristically b r o a d a b s o r p t i o n b a n d s typical for humic substances (Stevenson, 1982; M a c C a r t h y and Rice, 1985). Some differences between spectras can be notified. The peak at 1720 cm -~ is clear only in the spectra n u m b e r 2 (Brook Liuhapuro); this b a n d in humic substances is normally attributed to the C-------K) stretching vibration due mainly to unionized carboxyl groups (Stevenson, M a c C a r t h y a n d Rice, 1985). The b a n d at 1 6 3 0 - 1 6 0 0 c m - l is clear in each spectra but the intensity of the peak increases from the spectra 18-2 (from low D O M c o n c e n t r a t i o n to high D O M c o n c e n t r a t i o n in the original water). The b a n d on this region in humic substances is usually attributed to C------C vibrations of a r o m a t i c structures, a l t h o u g h several o t h e r reasons can also be given for a b s o r p t i o n o f this region, However, the increase of the intensity o f this peak is well in accordance with the absorptivity (A270) a n d elemental (H/C) datas, which also describes the aromaticity of the D O M (Table 3). Binding o f contaminants to D O M

The Kp values for benzo(a)pyrene (BaP; Fig. 3) were similar as reported earlier for natural D O M samples ( L a n d r u m et al., 1984; K u k k o n e n et al,, 1989, 1990; M c C a r t h y et al., 1989). Also, the Kps for

Table 4. Correlation coefficients (r) between percentages of the XAD-8 fractions and the parameters describing the quantity or the quality of dissolved organic carbon in the water samples (n = 20) H/C atomic %HI %HbA %HbN ratio A270 E4/E6 E2/E3 DOC -0.673(0.001) 0.844(0.001) -0.622(0.003) -0.654(0.002) 0.645(0.002) 0.854(<0.001) -0.002(0.992) E2/E3 -0.196(0.409) -0.057(0.810) 0.124(0.602) -0.334(0.150) 0.201(0.395) -0.004(0.986) E4/E6 -0.597(0.006) 0.711 (<0.001) -0.490(0.028) -0.535(0.015) 0.625(0.003) A270 -0.815(<0.001) 0.823(<0.001) -0.430(0.059) -0.772(<0.001) H/C atomic ratio 0.765(<0.001) -0.811 (<0.001) 0.466(0.038)

Fate of organic pollutants in humic lakes 25

459

BaP

NAPH

8000-

6000xS'~oo2ooo~ lOOO-

X

5oJ

* , , *

*

o 5 I) 1 6 ~ ~ o

TCB

5-

t

*

DHAA

4~3X

, * ""

1o

* 0~

o 5 l o = ~ ~

DOC ~

DOC rn~/L

Fig. 3. The binding coefficients (Kp) of model compounds in different waters. Each point represents the mean of four replicates.

4000

32100

24~00 ~ 1600

800

WAVENUMBER (cm")

Fig. 2. Infra-red spectra of the selected samples. The number of the spectra is the sample number in Tables 2 and 3. 3,3',4,4'-tetraclorobiphenyl (TCB) and naphthalene (NAPH) agreed well with the published partition coefficients for the similar compounds (Landrum et al., 1984; Hassett and Milicic, 1985; Kukkonen et al., 1990). The Kps for dehydroabietic acid (DHAA) are the first published. For BaP it was possible to measure the Kp value in every water sample but for the other model compounds especially NAPH and DHAA, which have much lower Kp values i.e. they do not have much interaction with DOM, the dialysis method was not sensitive enough to obtain very accurate measurements in the water samples with low DOM concentration. However, the K~s of BaP and TCB binding to the DOM from different sources confirm that hydro-

phobicity of the pollutants (TCB > BaP; Table 1) is not the only factor affecting the association of the pollutants and DOM (Kps: BaP > TCB) in the water phase. The same conclusion was made by Kukkonen et al. (1990), when they studied the binding capacities of different XAD-8 fractions of DOM, and by Lee and Farmer (1989) in their study on interactions of nonionic pesticides with DOM. There were linear relationships between the K~ values of BaP and the absorptivity at 270 nm (A270, r = 0.87, P < 0.001) as well as hydrophobic acid (HbA) content of the natural waters (r =0.77, P < 0.001) (Fig. 4). Also, there was a negative correlation between the Kp values of BaP and the hydrogen/ carbon ratio of DOM (r = 0.76, P < 0.001) (Fig. 4). These results on BaP agree well with the observations reported earlier by Gauthier et al. (1987) and by McCarthy et al. (1989). Gauthier et al. (1987) showed a strong correlation between both aromatic content and absorptivity at 270nm of humic materials extracted from soils and sediments (14 different samples) and the partition coefficient for binding of pyrene. In a study of 11 natural water samples McCarthy et al. (1989) reported good correlations between Kp for BaP and the size of humic molecules, HbA content of the DOM as well as A270. On the other hand, Kukkonen et al. (1990) showed that the HbA fraction binds most of BaP in the natural water having high DOM concentration. Taken all together, we conclude that the observed differences in affinities of BaP, and also of some other organic pollutants (pyrene, PCBs), between different water sources are largely explained by different proportions of hydrophobic acids and the degree of aromaticity of DOM. Bioavailability o f contaminants to D. m a g n a

Accumulation of model compounds by D. m a g n a was reduced by increasing the concentration of DOC

JUSSI KUKKONEN a n d A~MO OIKARI

460

Kp x 10 ~ 2620"

BaP 120-

,

NAPH

100- ,

r • 0.87

~

15

~ ~

10

0-

5

o 5 ~ ~

0

~

20000- ~ , 10

20

Kp x 104 2520-

30

40

50

o 5

~

"

TCB

~ DHAA

le1~-

ABS=ro

8000-

r " 0.77

15-

0-

o5

105-

o

20

40

60

80

%HbA

100

Kp x 10 ~ 25r - -0.76

2015-

10~ 5 o.5

1.o

1.5

2.0

H / C atomic ratio

Fig. 4. The binding coefficients (Kv) for binding of benzo(a)pyrene to different sources of DOM is linearly related to A270, percentage of hydrophobic acids and hydrogen/carbon atomic ratio.

in the water series (Fig. 5). The BCF values for model compounds in the organic-free control water were, in most cases, the same or significantly (P < 0.05) higher than BCFs for water samples with DOM. It is important to realize that the relationship between water DOC concentration and BCFs for BaP in animals is similar to the logarithmic type for this series of natural waters from a boreal area that can be produced by diluting a single humus water or humus preparation with control water (Kukkonen et al., 1989, 1990). As an exception, however, BCF of NAPH in the waters having a low DOM concentration ( < 4 mgC/1) revealed significantly (P < 0.05) higher BCF values than the control water (DOC ~<0.3, mgC/l). Similar type effect of DOM for NAPH was earlier detected by us by diluting a natural water sample (Kukkonen et al., 1990). The bioavailability of methylcholanthrene to D. m a g n a is also reported to increase by Aldrich humic acid (Leversee et al., 1983), but another study by McCarthy et al. (1985) showed opposite results for this compound. Kukkonen et al. (1989) showed

O-

~ ~2b~sb~

Fig. 5. Bioavailabilityof model compounds as a function of DOC in waters. Each point represents the mean of four replicate determinations. The arrow indicates the bioconcentration factor (BCF) in the organic-free control water. slightly increased accumulation of BaP at really low concentrations (DOC ~ 1 mgC/1) of natural humic material by D. m a g n a compared to organic-free control water. The bioavailability of DHAA was reduced, even in the waters containing low DOM concentration, but the effect of increasing DOM concentration is not as distinct as it is for the other model compounds. On the other hand, DHAA has very low Kp values (Fig. 3), which means that one can not expect very much effects of DOM on the bioavailability of DHAA either. DHAA is also a weak organic acid having a pK, value of 5.7 (Nyrrn and Back, 1958), which means that at the experimental pH 6.5 more than 50% of DHAA is in the ionized, more hydrophilic, form which does not accumulate so well (Kukkonen, 1991). The observed bioaccumulation of BaP in lake waters can be compared to the predicted values based on the assumption that BaP bound to the DOM is unavailable for uptake. Accordingly, bioaccumulation in water containing DOM will be proportional to the

,,," , .,-""

/

o-

/

//

/ /' b

~

~ ' ~ ' ~ ' ~

Fig. 6. Measured BCF values plotted against predicted BCF values for BaP of water samples from different sources. Prediction is based on the measured K~ values. The 95% confidence interval (. . . . ) of the regression line ( ) overlaps the 1: 1 line (- - -) predicted by the hypothesis that only the freely dissolved BaP is available for D. rnagna.

461

Fate of organic pollutants in humic lakes fraction of the contaminant that is freely dissolved

(hr.): predicted BCF in presence of DOM = control BCF x ff~= (2) w h e r e f ~ is calculated from the measured Kv (Fig. 3) and DOC concentration of each lake water: ./free = 1/(1 + Kp x DOC)

(3)

where DOC concentration is expressed as kg/1 and dimension of Kp is 1/kg. For BaP, the measured BCF values agreed well with the predicted BCF values from equation (2) (Fig. 6) and the 95% confidence limits of the regression overlaps the 1 : 1 line, which is predicted by the hypothesis expressed in equation (2). This result is consistent with those for BaP in D. magna (McCarthy et al., 1985; Kukkonen et al., 1990), Pontoporeia hoyi (Landrum et al., 1985) and rainbow trout (Black and McCarthy, 1988). Our study extends these observations to a geographic series of natural waters having both quantitative and qualitative differences in content of DOM. It also provides additional confirmation that the effects of natural DOM on bioaccumulation of BaP can be predicted from physicochemical measurements of K~s. On the other hand, the presented conclusion on BaP can not be extended to the other compounds of this study because of the difficulties to measure the exact Kp values. One limitation for accurate K~ measurement was the low natural DOM concentrations in some samples, however, we did not want to concentrate natural samples to avoid possible changes of the physico-chemical characteristics. Black and McCarthy (1988) got similar results for 2,2'5,5'-tetrachlorobiphenyl as we did for BaP, but they measured the K, value using a reverse-phase separation method. Moreover they used a Aldrich humic acid, known to have higher affinity to xenobiotics than natural

DOM (McCarthy et al., 1989; Eadie et al. 1990). A "biological Kp" for TCB in the present study can be backcalculated based on the reduction in BCF. The biological K~ values are higher than those shown in Fig. 3 for TCB with the dialysis technique, but these biological Kv values did not correlate with any measured quality parameters of DOM. Besides the total DOM concentration also the quality of DOM affects very clearly the bioavailability of organic xenobiotics. The measured BCF values correlated well with the percentage of hydrophobic acids (Fig. 7) and absorptivity at 270 nm (Fig. 8). It is important to notice, that the bioavailability of NAPH and DHAA correlated well with the chemical parameters of DOM. However, the Kp values of NAPH and DHAA did not reveal any relationship with the measured quality parameters of DOM. In fact, the results obtained by measuring bioaccumulation were clearer and more informative than the results obtained by Kp measurements. Environmental implications

This research has extended previous observations on the role of natural DOM in binding hydrophobic organic compounds and, more importantly, altering their availability for uptake by biota over a wide range of boreal waters. The binding capacity of BaP to DOM is clearly related to the aromaticity of DOM and the percentage of hydrophobic acids in waters. This result is confirmatory to the study of Kukkonen et al. (1990), who stated, based on one humic water sample, that hydrophobic acids are the main DOM fraction binding BaP. It is shown here that the total concentration of DOM is one of the main factors, maybe the most important one, controlling the bioavailability of xenobiotics. Besides the quantity, the quality of DOM i.e. the aromaticity and the portion of hydrophobic acids can lay an important role. Thus 5000-

BaP 1201

~**

=i 00

100, ~ ,

i

,

NAPH

i

.

J

20000-

i

. ~

o-

TCB 100]

B,P

•

0-0

r= -0~65

*

x-~,

120]

•

NAPH

4000 ~

~ooo.

r.;. '

*

DHAA

20000-

mooo~ moo~

:I

r:-0.78

,

~*

t r =-0.66

TCB 100-

**

*

DHAA

80-

**~'

*

***

8o 2 40-

4ooo~

0- r,-0.84 ~ ~ o

r= -o,r~

~ ~o

X I-~A Fig. 7. The BCF values in the four model compounds are correlated well with the percentage of hydrophobic acids in the series of natural water samples.

o

0-

r=-0.79 10 20 30 40 60

r=-0.38 0

10 20 30 40 60

7. HbA

Fig. 8. The BCF values in the four model compounds in water samples were directly related to the u.v.-absorptivity of the organic material in the water sample at 270 nm.

462

JUSSl KUKKONENand AIMOO1KARI

waters having about the same D O M concentration may exhibit different affinity to xenobiotics and the bioavailability of xenobiotics may be different also. Our data show that even low natural D O M concentrations can affect the bioavailability of xenobiotics and this fact should be noticed when predicting the fate of the organic xenobiotics in aquatic environments. Besides D O M , also particulate and colloidal organic material and sediments affect the transport and fate of xenobiotics. Adequate prediction of transport and availability to biota requires exact information on all these pools of organic matter. An ecotoxicologically interesting result is that certain contaminants, like naphthalene, revealed increased bioavailability in waters having a low D O M concentration compared to humus-free control water. Although there is not yet an explanation for this "low D O M effect", it may have distinct environmental implications in natural waters with low D O M . So, it is important to confirm this result with further experiments and with other compounds having about similar chemical characteristics as N A P H . It is also important to expand the study to the toxic effects of these moderately water soluble compounds, because D O M can enhance the toxicity of organic xenobiotics in natural waters (Kukkonen and Oikari, 1987; Virtanen et al., 1989; Oikari et al., 1991). Acknowledgements--We wish to thank Miss T. Pet/inen for

technical assistance, Dr K. H/inninen for helping to run i.r. spectra and Dr M. C. Black for reviewing the manuscript. This work was financed by the Academy of Finland/Research Council for Environmental Sciences (project 06/133).

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