**1. Introduction**

Dental caries is one of the most common diseases affecting humans. Traditionally, treatment involved the removal of all the affected tooth structure followed by replacement with amalgam. Silver mercury amalgam is a highly effective restorative material due to both its good mechanical and antibacterial properties. Following the Minamata agreemen<sup>t</sup> in 2013 on global mercury reduction signed by 140 counties, amalgam is being phased out. In Europe, amalgam use in primary teeth was banned in 2018 [1].

Modern dentistry advocates for the removal of highly infected surface dentine but not underlying demineralized and less infected dentine that may be close to the pulp [2]. Effective cavity sealing with Glass Ionomer Cement (GIC) restorations can reduce any underlying residual bacterial contamination. It can also encourage remineralization but have insufficient strength for larger cavities.

Conversely, dental composites have greater strength, but their placement is complex and their technique sensitive. Multiple adhesion-promoting steps are required, in addition to material placement and light curing in several increments to reduce polymerization shrinkage consequences [3]. Composite

failure is usually due to secondary caries at the restoration margins. This is attributed to a combination of residual bacteria, shrinkage and enzyme-activated degradation of demineralized dentine, which may compromise the bonding interface, enabling further bacterial penetration and growth [4]. A composite material that could provide surface release of antibacterial and remineralizing agents upon damage of the tooth/restoration interface might therefore be beneficial.

Antibacterial components such as chlorhexidine, fluoride, quaternary ammonium methacrylate, silver and triclosan have previously been added to dental composites [5–9]. Polylysine (PLS) is an alternative antibacterial agen<sup>t</sup> that has been included in composites more recently. Polylysine-greater eukaryotic cell-compatibility may address the limited biocompatibility of some other antibacterial agents [10,11]. Studies sugges<sup>t</sup> electrostatic adsorption of polylysine on bacterial surfaces, and its abnormal distribution within their cytoplasm, leads to their cell damage [12,13]. Polylysine, however, is generally recognized as safe (GRAS). It is a natural polypeptide that is biodegradable, water-soluble, nontoxic and edible. Additionally, polylysine is extensively used in eukaryotic cell culture to promote cell attachment and as a food preservative [12,14]. In the human body, polylysine degrades harmlessly to give the essential amino acid, lysine [12,14]. Furthermore, polylysine addition into calcium phosphate-containing composites can enhance apatite precipitation on their surfaces from simulated body fluid. Polylysine may therefore also enhance composite remineralizing potential [10].

Whilst many properties of polylysine-containing dental composites have now been investigated [10,11], publications on their actions against bacteria are limited. In this study, therefore, the ability of composites with increasing levels of polylysine to a ffect bacterial growth was investigated. Effects on planktonic bacteria are assessed to model the feasibility of surface polylysine release reducing bacterial microleakage in gaps at the tooth restoration interface. Polylysine release kinetics are provided to help explain the varying levels of antibacterial action. Potential polylysine addition and release effects on set material stability are monitored through mass and volume changes in water.

#### **2. Materials and Methods**
