**1. Introduction**

The toxicity assessment of nanoparticles (NPs) is a relevant issue since many researchers are using, specially, luminescent nanoparticles for various applications, such as bioimaging or drug delivery for *in vivo* and *in vitro* applications [1–3]. In this chapter, we will analyze how the approach in this analysis has been carried out for upconversion luminescent nanoparticles (UCNPs), which are a special type of NPs since they can receive energy in the near-infrared region (NIR) and emit in the visible or NIR spectrum. These NPs are composed of a matrix cell that can be made of oxides, oxysulfides, oxyhalides, phosphates, molybdates, tungstates, gallates, vanadates, and fluorides. The UCNPs are doped with lanthanide elements such as: Yb3+, Er3+, Tm3+, and Ho3+, among others. It is common in the upconversion process to have lanthanide elements co-doped to bring about a photon transfer between energy levels. For example, for the doping of Yb/Er, the Yb3+ absorbs NIR radiation at 970–980 nm of

wavelength in its base state (2 F7/2–2 F5/2), then this energy is transferred to Er3+ and the electron is populated to level <sup>4</sup> I11/2, then, a second photon is absorbed and by Yb3+ and it is transferred to Er3+, so the electron is raised to level 4 F7/2. From this state, it decays rapidly to <sup>4</sup> S3/2, and the green emission happens (4 S3/2–4 I15/2), and this process is called the APTE (Addition de photons par transfert d′énergie, i.e., photon energies by adding transfers), as can be seen in **Figure 1**. There are more upconversion processes with different doping combinations and concentrations of the ions, where the percentage of doping directly affects the color of emission [4].

The UCNPs have emerged as a promising nanomaterial for identifying specific cells and for drug delivery. Unlike other dyes, UCNPs exhibit stable emission if the source of excitation is maintained, making them more reliable. There are other types of upconversion processes such as: two-step absorption, cooperative sensitization, cooperative luminescence, the second harmonic generation, and two-photon absorption [4].

One crucial aspect of using UCNPs in biomedical applications lies in ensuring their biocompatibility on cells and or organisms. To achieve this, UCNPs must be functionalized with different ligands that specifically target the desired cells and organs. Several chemical groups, including polyethylene glycol (PEG) [5], polyethyleneimine (PEI) [6], polyvinylpyrrolidone (PVP) [7], polyacrylic acid (PAA) [8], and silica [6], have been used for this purpose. However, it is important to highlight that the toxicology of UCNPs depends on their physicochemical and physiological properties. Physicochemical properties include size, shape, surface area, and chemical composition, while physiological properties refer to the disease conditions, genetics, and other factors [9]. The recommended size for optimal penetration of NPs is below 100 nm. However, this size may also pose a risk of toxicity due to their potential to penetrate cellular structures and organs via the circulatory system. Moreover, UCNPs may generate reactive oxygen species (ROS) that can induce DNA damage, which not only affects the cell growth by means of protein oxidation, but also impacts mitochondrial respiration [10].

**Figure 1.** *Upconversion process between Yb3+and Er3+ ions.*

Several toxicological studies have been conducted on both *in vivo* and *in vitro* human cell lines and organs to assess the potential harmful effects of UCNPs. These studies have evaluated the effects of gene expression, growth, and reproduction of the organisms. It is crucial to continue monitoring and evaluating the toxicity of UCNPs as their use becomes more prevalent in biomedical applications.
