2.1. Rendering the Sapounidou Scheme as Structural Alerts and Subsequent Implementation in the Form of KNIME Workflow

A detailed description of the rationale underpinning the construction and content of the Sapounidou scheme is presented in Sapounidou et al.³³ It should be noted that a selection of minor amendments has since been made to its composition—the majority concerning only terminology. These are referenced explicitly in Table S2.

An overview of the key features within the scheme, as it exists in its present form, is provided in Table 1. In brief, it is structured to incorporate three initial tiers, each offering a progressively enhanced level of mechanistic resolution. Three broad top-level domains are present at Tier 1—“narcosis” (nonspecific effects typically manifesting as “baseline toxicity”), “reactive” (emerging as a product of intrinsic, nontargeted chemical reactivity), and “specific” (targeted interaction at a defined biomolecule, receptor, or pathway). Beneath this, within Tier 2, sit ten mechanistic groups. These are further divided across 25 mechanistic subgroups, together forming Tier 3. Each subgroup is anchored in turn within (potentially several) MIEs, which are themselves defined at the finest level by structural alerts.

Implementation to form a practical in silico profiling tool was achieved through construction of a workflow within KNIME analytic software (v.4.3.1; www.knime.com).³⁴ This was constituted such that it returns all accompanying information associated with given assignments—including the domain of taxonomical applicability—as presented in Table S3. Structural alerts were compiled from expert knowledge of chemistry surrounding those molecular initiating events established, within the literature, as holding relevance to aquatic toxicology. Their form was tailored such that both excessive exclusivity and generality in terms of potentially matched compounds were minimized. Ultimately, each was coded in the form of SMILES Arbitrary Target Specification (SMARTS) (www.daylight.com). Where possible, rules and alerts were adapted from existing schemes, with adjustments made to ensure coverage of a more appropriate spectrum of chemicals (as supported by existing knowledge). Alerts relating to narcosis were, for example, drawn primarily from Verhaar et al.²⁵—supplemented by the addition of rules covering carboxylic acid esters and various forms of ionic and nonionic surfactants.