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Yang et al. Soft Sci 2024;4:9 https://dx.doi.org/10.20517/ss.2023.43 Page 11 of 26

evaluated by pre-soaking LMs within the growth media for 24 h, followed by co-culturing cells with these
media for different days and viability characterization. Their results showed that bulk EGaIn had little
impact on the cell viability and proliferation. However, when LMs were sonicated to micro-/nano-particles,
3+
the concentration of In was increased more than 1,000 times within 20 min of sonication [Figure 4A]. This
was probably ascribed to the increased surface area-to-volume ratio, which resulted in increased interfacial
interaction between LMs and solutions. This process also allowed more In to appear at the surface of LM
particles and interact with the solution. In vitro assays indicated significant damage to cells caused by
releases from LM nanoparticles (with a mean size of less than 500 nm) within only one day of co-culture
[Figure 4B]. Given the fact that ultrasonication caused a significant change in In rather than Ga and led
3+
3+
3+
to a decrease in cell metabolic activity, it could be speculated that In gave rise to more critical cytotoxicity
3+
than Ga .
In recent years, there has been much research that adopted LMs in biomedical applications and studied
their biosafety in different hierarchies. At the cellular level, their results are generally in accordance with the
above discussion. Large-scale LMs tend to show no significant damage to various cell lines [72,84,85] .
Conversely, LM nanoparticles showed more or less damage to cells; e.g., nanospheres prepared by Hou et al.
and nanorices prepared by Yan et al. both exhibited inhibition to cellular metabolic capability [86,87] . However,
it is noteworthy that Hou et al. only took a small amount of suspension after sonication to operate
lyophilization and later re-dispersed LMNPs in cell culture media, which resulted in the dilution effect to
relieve their in vitro toxicities . As conjectured by Kim et al., the sonication process generated extremely
[87]
[83]
localized heat and pressure and induced chemical reactions, which might encourage ion release . Then, we
can regard the eventual ion number of LM nanoparticles suspension as two parts: (i) the ion number right
at the timepoint that sonication terminated (n ) and (ii) the number of ions released during the static
son
standing of the post-sonication suspension (n ). According to the result of Kim et al., we can speculate that
sta
[83]
n ＞＞ n , indicating that dilution of primary dissolvents could probably remit the ionic toxicity . To
son
sta
further tackle the ion-release problem, biocompatible encapsulation was applied to restrain the release of
ions; e.g., Wang et al. prepared calcium alginate hydrogel packaged magnetic LM nanoparticles and verified
their negligible toxicity to different cell lines with the concentration from 2.5 to 200 mg/mL [Figure 4C].
[57]
For in vivo injection applications, the impact of LMNPs on the immune system of the human body is a
critical issue. Zhang et al. studied ex vivo engulfing behavior of macrophages to different inorganic
particles . They observed that whether as individual particles or aggregated particle clusters, Ga NPs and
[88]
other studied particles (Silica, Sn, In, Bi, and Au) could all be engulfed by macrophages. In terms of the
cellular viability of macrophages incubated with these particles, Ga showed little cytotoxicity in 12 h with a
-1
safe threshold of 244.2 μg·mL . Generally speaking, Ga showed better cellular compatibility than In and Bi
but is more toxic than Au and Sn. The authors also reported that particles with higher density cause a more
pronounced decrease in the migration ability of macrophages. Among these particles, the density of Ga
particles is only larger than silica and smaller than other metallic materials, and they were found to have a
milder impact on macrophage movement. This work indicates that injection of particles might cause
immunoreaction at cellular levels. Extensive work should be further carried out to verify their impact on the
immune system in living bodies.

In vivo biocompatibility
Although the cellular toxicity of bulk LMs (and encapsulated LM particles) is evidently proved to be
negligible, the in vivo biosafety is still of great concern. Cogitation includes inflammatory reaction,
mechanical damages, the metastasis of materials and concentration effect, organ toxicity and hematic
toxicity, etc.