Elsevier

Aquatic Toxicology

Volume 96, Issue 2, 31 January 2010, Pages 135-144
Aquatic Toxicology

Influence of bicarbonate and humic acid on effects of chronic waterborne lead exposure to the fathead minnow (Pimephales promelas)

https://doi.org/10.1016/j.aquatox.2009.10.012Get rights and content

Abstract

Historically, the USEPA has only considered water hardness when establishing acute and chronic water quality criteria (WQC) for lead (Pb) in freshwater. Yet, recent evidence suggests that hardness may not be protective during chronic Pb exposure and that other factors (e.g., dissolved organic carbon (DOC) and alkalinity) influence toxicity. In fact, we have recently shown that Ca2+ (as CaSO4) does not protect against Pb accumulation in fathead minnows (Pimephales promelas) during chronic exposures whereas DOC as humic acid (HA) clearly does. To more clearly define the water chemistry parameters mediating chronic Pb toxicity we carried out 300 d exposures to study the influence of DOC and alkalinity on Pb accumulation and toxicity to fathead minnows at 2 different Pb concentrations (170 and 580 nM (35 and 120 μg/L)). Alkalinity was adjusted by addition of 500 μM NaHCO3 and DOC by addition of 4 mg/L HA. Fish were collected at 4, 30, 150 and 300 d of exposure to measure growth and Pb accumulation. Breeding assays (21 d) were performed at the end of these exposures to assess reproductive and larval behavioral endpoints. To determine whether effects were acute or chronic, switched breeding exposures were performed in which control breeders were transferred to either high or low Pb conditions and Pb-exposed breeders transferred to tap water without Pb. Mortality and growth effects were observed primarily in the high Pb treatments and within the first 10 d of exposure. Strong protection against Pb accumulation was afforded by increased DOC at both Pb concentrations. Increased alkalinity also appeared to moderately reduce Pb accumulation although not to the level of statistical significance. Tissue distribution of Pb was analyzed at 300 d and was found to accumulate mostly in bone, gill, intestine and kidney. Unexpectedly, high Pb reduced total reproductive output and increased average egg mass in the HCO3 and DOC treatments but not in the control water (+Pb) treatments. No statistically significant differences in egg hatchability or egg Pb accumulation were observed. Results from switched exposures suggest that embryo Pb accumulation arose from acute exposure to embryos rather than parental transfer. Finally, prey capture assays revealed potential Pb-induced motor/behavioral impairment in 10-d-old F1 larvae exposed to high Pb in all water treatments.

Introduction

Lead (Pb), a non-essential metal, is of primary interest to the USEPA (Fairbrother et al., 2007) ranking behind only copper (Cu) as one of the most highly reported causes of metal impairment to water quality (Reiley, 2007). As with other metals, the toxicity of Pb can vary greatly depending on effects that differences in local water quality may have on its speciation. Previous studies have shown that acute toxicity of Pb decreases with increasing hardness or alkalinity/pH (Davies et al., 1976, Schubauer-Berigan et al., 1993, Stouthart et al., 1994). For hardness, the protective effect is most likely due to antagonistic binding of Ca2+ to a shared channel for Pb at the gill (Busselberg et al., 1991, Rogers and Wood, 2004). Alkalinity, on the other hand, affords protection by the formation of Pb carbonate complexes that sequester free ionic Pb, presumably rendering it unavailable for uptake:Pb2+ + CO32−  PbCO3;Pb2+ + 2CO32−  Pb(CO3)22−.

More recently, it has become clear that other influences, such as complexation by dissolved organic carbon (DOC) or other organic/inorganic species, may also be important in determining Pb toxicity (Macdonald et al., 2002). In the case of DOC, protection is attributed to the high number of binding sites (carboxyl, phenolic, amino and sulfhydryl groups) that chelate metals and other cations from the water (Filella and Town, 2000).

The influence of water chemistry on chronic Pb toxicity is less clear than for acute toxicity due to the relative paucity of chronic studies. Consequently, acute toxicity data is heavily relied upon for establishing chronic water quality criteria (WQC) leading to potentially uncertain and/or inappropriate levels of environmental protection. From the limited studies available it would seem that hardness and increased pH/alkalinity are protective against chronic Pb toxicity in fish (Davies et al., 1976, Hodson et al., 1978). However, since CaCO3 contributes significantly to both hardness and alkalinity, and changes in these parameters commonly co-vary in natural waters and laboratory experiments, there remains uncertainty as to the protective contribution of each.

One of the main reasons that chronic studies evaluating reproductive toxicity in fish are lacking is the time and effort required to perform such experiments. Hence, we previously conducted short-term exposures to identify the likely key water chemistry parameters influencing chronic Pb toxicity prior to undertaking exposures through reproductive maturity. These efforts demonstrated protective effects by Ca2+ (as CaSO4) and DOC (as Aldrich humic acid (HA)) against acute Pb toxicity (Grosell et al., 2006), as well as against chronic Pb accumulation by HA but not Ca2+ in fathead minnows (Mager et al., 2008). Reproduction was not evaluated in the latter study and, aside from Pb-induced transcriptional responses, no other toxic effects were observed under the conditions examined. Still, these findings shed some doubt as to the protective influence of Ca2+ on chronic Pb toxicity while further supporting DOC as an important protective component that warrants greater consideration. However, because CaSO4 was used to explicitly study the effects of increased hardness without increasing alkalinity in these experiments, the influence of alkalinity alone on chronic Pb accumulation and toxicity remains unclear.

Having narrowed the field for potential key water chemistry parameters, we proceeded with the present study aimed primarily at investigating water chemistry influences on Pb-induced reproductive effects. Specifically, we again evaluated the influence of HA to determine whether the protection against whole body Pb accumulation observed previously translated into protection against full-term reproductive effects. We also investigated the effect of increased alkalinity (as NaHCO3) in lieu of Ca2+ given its previous failure to protect against chronic Pb accumulation. The reproductive endpoints of fecundity, hatchability, egg mass, egg Pb accumulation and attachment of eggs to breeding substrate were monitored. Additionally, growth, Pb accumulation and potential Pb-induced neurological impairment in larval offspring were assessed.

Section snippets

Experimental design

The main goal of this study was to examine the influence of DOC (as HA) and alkalinity (as NaHCO3) on the reproductive toxicity of chronic Pb exposure to fathead minnows (Fig. 1). To this end, Pb exposures were administered in 3 different laboratory waters (described below) to 8-d-old fathead minnow larvae for 230–300 d and subsequently through 3 sequential rounds of 21 d breeding assays. During these breeding assays eggs were counted daily to assess fecundity and collected for hatchability or

Water chemistry

In a previous 150 d study (Mager et al., 2008) we used a 2:1 deionized water:dechlorinated tap water mixture to achieve a moderately soft base water for investigating hardness and DOC effects on Pb toxicity. Due to the much larger scale and duration of this study, we chose instead to use full strength tap water to eliminate the difficulties associated with higher flow demands of deionized water. Accordingly, this led to a base water with approximately 3-fold higher concentrations of all water

Conclusions

We have demonstrated that increased HCO3 and DOC (as HA) protect against chronic Pb accumulation by fathead minnows. Yet paradoxically these same parameters appear to augment reproductive toxicity at high Pb concentrations. Indeed, the influences of HCO3 and HA on the effects of Pb exposure throughout this study were unexpected and somewhat puzzling. However, these influences were consistent across several of the endpoints examined lending credence to the connectivity of the combined impacts

Acknowledgements

This work was funded by the International Lead Zinc Research Organization (ILZRO) and the United States Environmental Protection Agency (USEPA) under the Science to Achieve Results (STAR) Graduate Fellowship Program (awarded to EMM). USEPA has not officially endorsed this publication and the views expressed herein may not reflect the views of the USEPA. KVB was supported by a University of Miami Maytag Fellowship. The authors would like to thank Dr. Adam Ryan (HydroQual, Inc.) for Pb speciation

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