# Comparison of Genotoxicity and Pulmonary Toxicity Study of Modified SiO2 Nanomaterials
## 4 Discussion  
Nano-silicas are one of the most commonly used nano-additives in many products. Silicas are divided into three major forms: crystalline, cryptocrystalline, and amorphous. Crystalline silicas can be further divided into three crystalline forms, such as quartz, cristobalite, and tridymite. Quartz is one of the most abundant minerals in the Earth's crust and is, therefore, also the most common dust that miners are exposed to [27]. Long-term exposure to silica dust may cause silica to be deposited in the alveolar space and induce silicosis or lung cancer. Silicosis may be further classified as simple (nodular) silicosis, acute silicosis, and complex pneumoconiosis [28]. In addition to silicosis, silicas may also induce many autoimmune diseases, such as systemic sclerosis, rheumatoid arthritis, systemic lupus erythematosus, etc. [29]. When compared to amorphous silicas, crystalline silicas show higher cytotoxicity due to irregular particles with sharp edges, stable surface radicals, and sustained release of HO radicals via a Fenton-like mechanism [30]. To compare the genotoxicity of the unmodified and modified silica samples, we conducted an Ames test, chromosomal aberration test, and micronucleus test. The Ames test is a quick and simple genotoxicity test conducted with mutated _Salmonella typhimurium_ strains. The TA98 strain has an allele of hisD3052, which is sensitive to frameshift mutations. On the other hand, the TA100 strain has an allele of hisG46, which is sensitive to base-pair substitution mutations of the C-G base pair [31]. In our study, SB induced bacterial reverse mutation in the TA100 strain but not in the TA 98 strain. The positive result in TA100 and the negative result in TA98 in the Ames test indicated that SB might induce base-pair substitution mutations, not frameshift mutations, in _Salmonella typhimurium_. Some reports have indicated that no mutagenic potential is observed in the Ames test with or without metabolic activation [32; 33; 34]. One study showed the genotoxic potential of exfoliated silicate nanoclay [35]. The chromosomal aberration test is an in vitro genotoxicity test conducted in a CHO-K1 cell line. In this test, the samples were tested to see if they could induce chromosome recombination, breaks, polyploidy, or other morphological abnormalities in rapidly dividing mammal cells. These morphological abnormalities are also called chromosome aberrations. To induce chromosome aberrations, at least one double-strand-break (DSB) of DNA is required [36]. Chromosome aberrations can be further divided into chromosome-type aberrations (CSAs) and chromatid-type aberrations (CTAs) according to the mechanisms that induce the chromosome aberration. CSAs are mainly induced by ionizing radiation. On the other hand, CTAs are mainly produced by chemical injury [37; 38]. In a previous study, the CA test of the nano-silica in CHO-K1 cells showed that nano-silica did not cause the ratio of chromosomal aberrations to increase with or without S9 metabolism [39]. However, in our study, SB induced significant increases in the frequency of chromosome aberrations. On the other hand, the major type of chromosome aberration that SB induced was gaps, which belong to CSAs. The results of the CA test indicated that SB may induce ionizing radiation damage to the genes of CHO-K1 cells. The micronucleus test is an in vivo genotoxicity test conducted by observing the frequency of micronuclei in the peripheral blood of rodents. The majority of mature RBCs  
\begin{table}
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**Group** & **M (10\({}^{5}\))** & **N (10\({}^{5}\))** & **L (10\({}^{5}\))** & **E (10\({}^{5}\))** \\ \hline NC 1 [FOOTNOTE:1]Footnote 1: NC: negative control, R.D.W, 0.5 mL/kg bw, i.t. ; SA: sample A, 0.645 mg/kg bw, 320\(\times\) diluted, 0.5 mL/kg bw, i.t. ; mSA: modified sample, 0.06 mg/kg bw, 320\(\times\) diluted, 0.5 mL/kg bw, i.t. ; SB: sample B, 2.24 mg/kg bw, 80\(\times\) diluted, 0.5 mL/kg bw, i.t. ; mSB: modified sample B, 0.24 mg/kg bw, 80\(\times\) diluted, 0.5 mL/kg bw, it.