1. Introduction

A plethora of methods have been developed to precisely control the size, shape, and surface chemistry of AuNPs. These range from green synthesis methods, where the AuNPs are produced either by microorganisms or plant extracts [56,57,58], to physical methods such as laser ablation [59,60,61], thermal decomposition [62,63], and mechanical milling [64], and finally, chemical synthesis methods. Chemical synthesis methods are perhaps the most widely employed methods, owing to the vast array of physicochemical properties which can be achieved and the specificity with which they can be obtained. Many approaches exist to chemically synthesize AuNPs; however, they all proceed via essentially the same steps (Figure 1):

• Reduction of Au³⁺—afforded by a gold salt, usually HAuCl₄—to atomic Au⁰; this process is rapid and continues until the concentration of gold atoms in solution reaches supersaturation.
• Nucleation of gold atoms into gold clusters; the number of nucleation sites determines the number concentration of AuNPs, i.e., for a fixed mass concentration more nucleation events results in smaller particles and vice versa.
• Growth via coalescence of gold clusters and diffusion of remaining soluble gold atoms onto the surface of gold agglomerates.

The following sections will outline some of the most common methods used to synthesize ultrasmall AuNPs; they have been divided into the four main methods used in the literature—Turkevich/Frens, reduction by sodium borohydride, Brust–Schiffrin, and seeded growth. Where reagent names have been abbreviated, the meaning can be found in Table 1.

Turkevich/Frens (Figure 2A) synthesis is the classical method of producing AuNPs. It was one of the first systematic approaches to the size-controlled synthesis of AuNPs and is still popular today, owing largely to its simplicity and reliability. The method was pioneered Turkevich et al. in 1951 [65], producing 15–24 nm AuNPs, and later refined by Frens in 1973 [66], extending the size range to 16–147 nm. In this synthesis, citrate is used as both reducing agent and capping agent; however, citrate is not a strong enough reducing agent to rapidly generate atomic gold at room temperature; therefore, the synthesis is carried out at elevated temperature, typically boiling. The AuNPs’ size is controlled predominantly by the ratio of citrate:Au, where more citrate results in more rapid nucleation and, subsequently, smaller particles. Particle size and distribution may also be controlled by pH [67], temperature [68], and order of reagent addition [69]. While AuNPs with an average diameter of 4 nm have been reportedly synthesized by the Turkevich method with minor modifications [70], it is far more common for particles to be larger than 10 nm in diameter.

Sodium borohydride (Figure 2B) is often implemented as a strong reducing agent in the synthesis of AuNPs enabling the reaction to be performed at room temperature and allowing for rapid nucleation and formation of smaller AuNPs, frequently sub 5 nm. Like the Turkevich method, citrate may be included; however, when NaBH₄ is used as a reducing agent, citrate severs solely as a capping agent [71]. Alternatively, citrate can be replaced by other hydrophilic capping agents such as alginate [72] or chitosan [73]. The synthesis may also be performed in non-polar solvents such as chloroform, implementing hydrophobic capping agents such as CTAB [74] and ODA [75,76]. Finally, capping agents may be omitted entirely to produce “bare” AuNPs [77].

Brust–Schiffrin (Figure 2C) synthesis is a two-phase approach to produce alkanethiol-capped AuNPs which are soluble in hydrophobic solvents. TOAB is employed to transfer AuCl₄^- from the aqueous phase to an organic phase, typically toluene; NaBH₄ is used to reduce the gold salt in the presence of a capping agent, traditionally an alkanethiol. The capping agent first used, and still commonly used today, is the alkanethiol dodecanethiol [78]; however, this may be replaced with other alkanethiols such as pentanethiol [79] or hexanethiol [80], surfactants such as CTAB or CTAC [81], or even ionizable molecules for the synthesis of water soluble AuNPs, for example via the use of MPA [82].

Seeded growth (Figure 2D) is synthetic process of first producing Au⁰ clusters, often via NaBH₄, although Turkevich/Frens particles may also be used as seeds, which are then introduced as presynthesized nuclei into a growth solution. Essentially, the particle number concentration of the resulting solution can be finely controlled by varying the number of nuclei introduced and the final particle size is regulated by the gold concentration in the growth solution. This approach is not particularly well suited to the formation of ultrasmall AuNPs and is more commonly employed for the preparation of particles over a large size range [83,84]. As well as being applicable over large size ranges, seeded growth is also capable of producing a wide variety of shapes by using different shape directing agents, such as CTAC for spheres [85] and cubes [86], CTAC/NaI for triangles [87], CTAB/CTAC/HQL for bipyramids/javelins [88], PVP for stars [89], and BDAC/CTAB for rods [90].

The methods outlined in the previous sections are summarized in Table 2.