Abstract
Carbonate removal using acids is a common practice in ecological studies. The effects, however, of acid pre-treatment on the elemental and isotopic composition of marine invertebrates as well as how these effects vary according to species’ carbonate content is little known. We examined the effects of acid pre-treatment on the elemental (%C, %N, C:N ratio (%C:%N)) and isotopic composition (δ13C, δ15N) of 28 lightly- and heavily-calcified species from Cnidaria, Mollusca, Arthropoda, Bryozoa, Echinodermata and Chordata. The present study showed that acid pre-treatment modified the elemental and isotopic composition of lightly- and heavily-calcified marine invertebrates. The shifts were clearly seen as a decrease in the %C and δ13C of heavily-calcified species while we did not detect a clear pattern for %N and δ15N (in both lightly- and heavily calcified species). Apart from carbonates, acid pre-treatment caused also the loss of organic compounds, thus confounding the interpretation of carbonate proxy (CP) -a widely used proxy for carbonate content. We recommend the use of CP solely with heavily-calcified species. For the first time it was shown that the use of δ15N values from acidified samples can introduce substantial bias in our perception about the number of trophic levels, the distribution of species and distribution of biomass across the trophic levels in a community. We have uncovered and elucidated previously unknown aspects and highlighted the challenge posed when predicting shifts in elemental and isotopic composition of species following acid pre-treatment. The present findings should be considered in future studies using acid pre-treatment as they can contribute to the optimum use of samples while avoiding bias in the interpretation of findings.
Highlights
The elemental and isotopic composition of organisms’ organic compounds can provide ecologists with useful information about food-web structure and cycling of organic matter
The possible distortion of stable isotope ratios due to preservation in formalin could introduce some uncertainty in the examination of a community’s trophic structure; it should be mentioned that a) several studies have shown that the effects of formalin preservation on δ15Ν values were minor compared to a commonly-used trophic fractionation factor (i.e. +3.4‰, DeNiro and Epstein 1981; Post 2002) enabling the allocation of species to trophic levels (Fanelli et al 2010) and b) a confounding effect on our findings should not be expected since both acidified and non-acidified subsamples have been preserved in formalin
Effects of carbonate removal on elemental composition In 14 taxa there was a statistically-significant decrease in their %C while nonstatistically-significant shifts were found in 4 taxa (Table 1)
Summary
The elemental and isotopic composition of organisms’ organic compounds can provide ecologists with useful information about food-web structure and cycling of organic matter. In contrast to the organic carbon fractions, the inorganic forms of carbon (i.e. carbonates, CaCO3) do not reflect dietary sources They exhibit enriched (i.e. higher) δ13C values and their presence can confound findings about species’ diet and community trophic structure Carbonate removal can be carried out mechanically (e.g. shells of large-sized bivalves) This approach is not always possible for small-sized invertebrates and some Phyla (e.g. echinoderms) (Mateo et al 2008; Schlacher and Connolly 2014). In these cases, carbonate removal is carried out using acids which leads to the release of carbonates. Apart from the type of acid used, there is variability in terms of the acid concentration used (e.g. 1–36% in the case of HCl, Brodie et al 2011 and references therein) and the means of application (e.g. direct application of the acid in the capsule containing the sample vs. acid fumigation, addition of a specific volume of acid vs. the addition of acid until the cessation of effervescence) (Bosley and Wainright 1999; Jacob et al 2005; Søreide et al 2006; Jaschinski et al 2008; Serrano et al 2008; Vafeiadou et al 2013)
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