**1. Introduction**

Marine biofouling consists of the settlement and gradual accumulation of micro- and macro-organisms, such as bacteria, fungi, spores of algae, and invertebrate larvae in water submerged surfaces [1–3]. However, despite this natural process starting with the attachment and proliferation of

bacteria, it is the growth of macrofouling organisms that most concerns marine industries. Their dense layers can cause a reduction in or even blockage of water flow in pipes, mechanical damage, corrosion, and the failure of equipment [4]. In other words, marine biofouling can increase maintenance costs and fuel consumption [5,6].

Chemical control is the principal strategy to combat this issue, combining traditional antifouling (AF) coatings, as is the case of polydimethylsiloxane (PDMS) and polyurethane (PU)-based coatings, with biocides, which are released over time [7,8]. Although most of them are presented as non-persistent biocides, several occurrence studies have concluded that, in fact, booster biocides persist, owing to their high release in biocide-release-based AF coatings [9–12]. In addition, due to their low water solubility and hydrophobic behavior, booster biocides tend to bioaccumulate, causing environmental damage [13–15]. Fortunately, there is increasing concern about the influence of copper and booster biocides in the marine environment and the e ffort to find ecological alternatives has led many researchers to develop greener AF approaches in order to reduce biocide release and consequently their persistence in the ecosystem [16–20]. A good AF agen<sup>t</sup> must prevent fouling without persisting at concentrations greater than those that can cause detrimental e ffects to the environment.

A large variety of microorganisms and sessile marine organisms, such as sponges, corals, and algae, usually free of fouling on their surfaces, has been described to produce secondary metabolites to fight this natural process [21], as is the case of zosteric acid (ZA), a sulfated phenolic acid produced by the seagrass *Zostera marina* [22,23]. Inspired by the ZA structure, AF properties of a small library of synthetic sulfated small molecules were studied and gallic acid persulfate (GAP) was found to be one of the most promising compounds [24]. Furthermore, gallic acid, its starting material, is commercially available and easily accessible since it can be obtained from several sources, such as winery waste [25]. In contrast to ZA, GAP showed anti-settlement activity against the adhesive larvae of *Mytilus galloprovincialis*, one of the most aggressive invasive species in the world, according to the International Union for Conservation of Nature (IUCN) [26], without causing ecotoxicity against this species and other non-target species (Figure 1) [24].

**Figure 1.** Antifouling (AF) activity and ecotoxicity previously discovered for gallic acid persulfate (GAP) [24].

To assess GAP suitability as an AF agent, it is necessary to evaluate compound stability and degradation pathways, optimize its large-scale synthesis, and analyze its behavior after incorporation in selected coating formulations. In this study, both the solubility of GAP in ultra-pure water (UPW) and in sterilized natural seawater (snSW), as well as the stability of sulfate groups after exposure to varying conditions of light and temperature for a period of several months, were examined. GAP synthesis was optimized through a microwave (MW) reaction to obtain high amounts of the compound and allow further immobilization in traditional marine coatings. Coating formulations with new synthesized GAP were developed using both direct incorporation (DI) and chemical immobilization (CI) strategies into two representative marine coatings: PDMS and PU-based coatings. The purpose of the chemical immobilization of GAP was to achieve maximal compatibility between this polar molecule and polymeric coatings, as well as provide long-lasting antifouling e ffects with a non-release strategy. Leaching assays were performed to confirm the minimal release of GAP into the aquatic environment. As the targets of any preventive technology are the colonizing stages [27], the settlement inhibition of *Mytilus galloprovincialis* larvae on the several GAP-based coating formulations was evaluated.

During these studies, two analytical methods were developed in order to: (1) directly analyze this sulfated polar compound in snSW, through an ion pair reversed-phase high performance liquid chromatography (IP-RP-HPLC) method (stability assays) [28] and (2) concentrate and extract this analyte from the leached artificial seawater (ASW), through a weak anionic exchange (WAX) method preceding IP-RP-HPLC quantification (leaching assays).
