Contaminant pathways in Port Curtis: Final report - OzCoasts

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Contaminant pathways in Port Curtis: Final report8: Effect of pulse eventsDissolved copper, µg/L3530(i)25201510500 12 24 36 48 60 72Time, hDissolved copper, µg/L605040302010(ii)00 12 24 36 48 60 72Time, hDissolved copper, µg/L2001601208040(iii)00 12 24 36 48 60 72Time, hFigure 8.1. Copper exposure scenarios tested: (i) equivalent copper ‘dose’, but varying durationand magnitude; (ii) equivalent copper ‘dose’ and concentration, but varying pulse frequency;and (iii) high copper concentration pulses of decreasing pulse duration8.2.4 Intracellular and extracellular copper determinationsTo investigate the uptake of copper during pulsed and continuous exposure, thecopper bound to the surface of the algae (extracellular) and the internalisedcopper (intracellular) were determined at various exposure times. Extracellularcopper was isolated by suspending the algal pellet in 20 ml of 0.01 M EDTA inNaCl solution (3.5% m/v) for 35 min. Following centrifugation, a portion of thisextract was then acidified with 1% HNO 3 to pH 3 and analysed by SW-ASV. Theresidual algal pellet was then washed with approximately 10 mL of clean seawater, acidified (2 mL of concentrated HNO 3 ), and was left for at least 30 min priorto microwave digestion (90 W for 5 min, then cooling to room temperature). Thedigests were then diluted to 20 mL with deionised water and analysed by GF-AAS.This fraction was deemed the “intracellular Cu.” The environmentally realistic, lowalgal cell densities used in these experiments meant that multiple flasks often hadto be combined so that the extracted intracellular and extracellular copper wereabove the detection limit of the instruments used in their analysis.99
Contaminant pathways in Port Curtis: Final report8: Effect of pulse events8.2.5 Modelling bioassay response with fluctuating copper concentrationsIn the absence of a toxicant, and under constant light, temperature and nutrientconditions (assuming no lag phase), algal growth is exponential (Nyholm 1985)and the growth rate (µ t ) is constant. A plot of log cell density (log N) versus timeis linear with slope equal to µ t (equation 1):N t = N 0·e µt (t-t 0 ) (Eqn 1)Algal growth rate, µ t , decreases with increasing toxicant concentration and wasmodelled using a four-parameter logistic model (Nyholm et al. 1992; Simpson etal. 2003) according to:µt = α – β/(1 + δ·e(-φ·Ct)) (Eqn 2)where µ t is the growth rate at time t, C t is the toxicant concentration at time t, α, β,δ and φ are constants.In the model, C t , and subsequently µ t were calculated at hourly intervals over the72-h test period and algal cell densities were calculated according to equation 3.Nt = Ntd·eµt (t - td) (Eqn 3)where N t is the number of cells at any given time t, N td is the number of cells attime td (td< t), µ t is the growth rate at time t (varying due to changingconcentration, C t ).Measurements of algal biomass at time periods t = 0, 24, 48, and 72 h duringexposure of the algae to copper concentrations of 0, 2, 4, 8, 16, 30 and 50 µg/Lcopper was used to develop a relationship between the algal growth-rateparameter, µ t , and copper. The fit between the measured algal biomass and themodel predictions was optimised using the Solver tool application of MicrosoftExcel ® which minimises the sum-of-the-squares of difference between themeasured and the model algal biomass data by changing the variables in themodel (equations 2 and 3). Only the effects-data for the continuous exposureexperiments were used for optimising the model fit.Two exposure effect models were developed. In Model 1, the external copperconcentration in solution was treated as the ‘exposure’ and the cause of the toxiceffects to algal growth rate. In Model 2, the intracellular copper concentration,within the algae, was treated as the ‘exposure’ and the cause of the toxic effects toalgal growth rate. In Model 2, the intracellular exposure was calculated bymultiplying the external copper concentration in solution by an uptake rate100
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Contaminant pathways in Port Curtis: Final report - OzCoasts

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