Table 3.
Genotoxicity of silica in vitro (including data of non-food-grade and colloidal SAS)
| Test system | Test substance | Particle size and/or SSA | Source | Method/treatment/parameters studied | Results | References |
|---|---|---|---|---|---|---|
| In gene mutation—bacteria | ||||||
| S. typhimurium TA98, TA100, TA1535, TA1537, TA1538 | Pyrogenic SAS (CAB-O-SIL® EH-5) | 385 m2/g | Cabot | Standard plate, agar plate, only with S-9 | Negative | ECETOC (2006) |
| S. typhimurium TA98, TA100, TA1535, TA1537, TA1538 | Pyrogenic SAS (CAB-O-SIL® M-5) | 200 m2/g | Cabot | nr, ± S-9 (not defined) | Negative | |
| S. typhimurium TA98, TA100, TA1535, TA1537, TA1538 | Silica gel (Silcron G-910) | nr | nr | Standard plate, agar plate | Negative | |
| Escherichia coli WP2 | Silica gel (Silcron G-910) | nr | nr | Standard plate, agar plate | Negative | |
| S. typhimurium TA1530, G-46 | Silica gel (Syloid 244) | 2.5–3.7 µm | nr | Spot test | Negative | |
| S. typhimurium TA98, TA100, TA1535, TA1537, WP2uvrA | Colloidal silica* | 20, 100 nm | E&B Nanotech Co Ltd | OECD TG 471, GLP | Negative | Kwon et al. (2014) |
| Gene mutation—mammalian cells | ||||||
| CHO cells | Pyrogenic SAS (CAB-O-SIL® EH-5) | 385 m2/g | Cabot | OECD TG 476, GLP | Negative | ECETOC (2006) |
| L5178Y mouse lymphoma cells | Precipitated SAS (NM-200) | 190 m2/g | JRC | OECD TG 476, GLP | Negative | CEFIC (2012a) |
| L5178Y mouse lymphoma cells | Pyrogenic and precipitated SAS (NM-200, -201, -202, -203) | 10–22 nm | JRC | OECD TG 476 | Negative | NANOGENOTOX (2013) |
| V79 hamster lung fibroblasts, HPRT | 2 pyrogenic SAS, 1 precipitated SAS, 2 precipitated colloids* | 20 and 25/70 nm (pyrogenic), 20 nm (precip.), 15, 40/80 nm (colloid)/50–200 m2/g | Commercial | 12.5, 25, 50, 100 mg/L, 24 h | Negative | Guichard et al. (2015a) |
| Mouse embryonic fibroblast (MEF-LacZ cells) | Stöber silica without stabiliser* | 10, 30, 80, 400 nm (actual sizes 11, 34, 34 and 248 nm); SSA nr | Glantreo | 4, 40, 400 mg/L, 16 h | 10, 80, 400 nm: negative 30 nm: dose related ↑(2-threefold) at 40, 100, 400 mg/L; ↓viability (80 % at 85 mg/L): no particles in nucleus, implying secondary effect |
Park et al. (2011) |
| Mouse lung epithelial (FE1) cells derived from Muta™ Mouse | Colloidal SAS* and 2 µm SAS | 12, 5–15, 10–20 nm, 2 µm | Sigma-Aldrich, NIST | 12.5 mg/L for 8 consecutive times | Negative, concentrations higher than 12.5 mg/L were cytotoxic and not included in the analysis | Decan et al. (2016) |
| In vitro micronuclei induction | ||||||
| BEAS2B, 16HBE, A549, Caco-2 | Pyrogenic and precipitated SAS (NM-200, -201, -202, -203) | 10–22 nm | JRC | OECD TG 487, 24 h, then cytB added | A549: positive for NM-201 and NM-202; Caco-2: inconclusive (1 exp positive, 1 exp negative); BEAS2B, 16HBE: mostly negative |
NANOGENOTOX (2013) |
| BEAS2B | Precipitated silica (NM-200) and pyrogenic silica (NM-203) | 22 nm, 190 m2/g | JRC | OECD TG 487, 0.1–100 mg/L, 48 h, cytB added 6 h after the beginning of treatment | Negative | Zijno et al. (2016) |
| Caco-2 | Colloidal silica (Levasil® 50, Levasil® 200)* | 15, 55 nm | HC Starck | OECD TG 487, 24 h treatment, then cytB added | 15 nm: 1.5-fold↑ at 16 µg/mL and threefold↑ at 32 and 64 µg/mL in the presence of ↓RI (remained above 55 %); addition of FCS reduced effect by 50 % 55 nm: no effect; addition of |
Tarantini et al. (2015b) |
| Human lymphocytes | Pyrogenic and precipitated SAS (NM-200, -201, -202, -203) | 10–22 nm, 160–230 m2/g | JRC | OECD TG 487, up to 1,250 mg/L, 24 h | Negative | Tavares et al. (2014) |
| Human lymphocytes | Colloidal silica (Levasil® 50, Levasil® 200)* | 15, 55 nm/200, 50 m2/g | HC Starck | OECD TG 487, 31.6–1000 mg/L | Negative | Downs et al. (2012) |
| Balb/3T3 mouse fibroblasts | Precipitated silica (NM-200) and pyrogenic silica (NM-203); and colloidal silica* |
5–90 nm/50–200 m2/g | JRC | OECD TG 487, 100 mg/L, 24 h, then cytB added | Negative | Uboldi et al. (2012) |
| V79 hamster lung fibroblasts | 2 pyrogenic, 1 precipitated, and 2 precipitated colloids* | 20 and 25/70 nm (pyrogenic), 20 nm (precip.), 15, 40/80 nm (colloid)/50–200 m2/g | Commercial | 12.5, 25, 50, 100 mg/L, 24 h | Negative | Guichard et al. (2015) |
| V79 hamster lung fibroblasts | Silica gel (Spherisorb® 5 µm)* | nr | nr | 24 h, 20–160 µg/cm2 | Weak but significant induction of micronuclei at cytotoxic doses | Liu et al. (1996) |
| Mouse embryonic fibroblast (MEF-LacZ cells) | Stöber silica without stabiliser* | 10, 30, 80, 400 nm (actual sizes 11, 34, 34 and 248 nm); SSA nr | Glantreo | 4, 40, 400 mg/L, 16 h | 10, 30, 400 nm: negative 80 nm: positive; no particles in nucleus, implying secondary effect |
Park et al. (2011) |
| A549 human epithelial lung carcinoma cells | Stöber silica* | 12–174 nm | Laboratory | OECD TG 487; 40 h | No significant induction of micronuclei; other weak chromosomal effects were observed, but again without reaching statistical significance; no cytotoxicity | Gonzalez et al. (2010, 2014) |
| In vitro chromosome aberration studies | ||||||
| Chromosome aberrations, CHO cells | Pyrogenic SAS (CAB-O-SIL® EH-5) | 385 m2/g | Cabot | Equivalent to OECD TG 473, GLP -S9: 16 h, 38–300 µg/ml; +S9: 2 h, 250–1000 µg/mL |
Negative | Cabot 1990 as cited in ECETOC (2006) |
| Chromosome aberrations, V79 cells |
Precipitated silica (NM-200) | 190 m2/g | JRC | OECD TG 473, GLP | Negative | CEFIC (2012b) |
| Chromosome aberrations, V79 cells |
Colloidal silica* | 20, 100 nm | E&B Nanotech Co Ltd | OECD TG 473, GLP | Negative | Kwon et al. (2014) |
| Chromosome aberrations, human embryonic lung cells (Wi-38) | Micronized silica gel (Syloid® 244) | 2.5–3.7 µm | nr | 24 h (presumably), only in the absence of S9, 1–1000 µg/mL | Negative | US-FDA 1974 as cited in ECETOC (2006) |
| In vitro UDS assays | ||||||
| Primary rat hepatocytes | Pyrogenic SAS (CAB-O-SIL® EH-5) | 385 m2/g | Cabot | 0.3–1000 µg/ml, with and without S9, exposure time 18–20 h | Negative | Cabot 1989 as cited in ECETOC (2006) |
| In vitro comet assays | ||||||
| HT-29 human colon carcinoma cell line | Pyrogenic SAS (AEROSIL® 200, AEROSIL® Ox50) | 12, 40 nm, 200, 50 m2/g | Evonik Industries | Cytotoxicity (±FCS, 1 and 10 %, 0.03–156.3 µg/cm2); comet assay with and without Fpg | Negative, no oxidative DNA damage | Gehrke et al. (2013) |
| Human Caco-2 intestinal cells (undifferentiated) | Pyrogenic silica* | 14 nm, SSA 200 m2/g | Sigma | 20, 80 µg/cm2; 4 and 24 h, cytotoxicity (LDH and WST-1); 20 µg/cm2 for Fpg–comet assay (4 h treatment); glutathione | Positive (20 µg/cm2) in the presence of cytotoxicity (cytotoxic at 20 µg/cm2 after 24 (LDH) or 4 h (WST-1); DNA damage only with Fpg; ↓glutathione | Gerloff (2010) and Gerloff et al. (2009) |
| A549, HT29, and HaCat | Colloidal silica (Ludox SM-30)* | 14 nm, agglomerated to 500 nm in medium | Sigma | 24 h, 0.01–10 µg/mL | Significant increases in DNA damage at ≥0.1 mg/L in all tested cell types; cytotoxicity ≥1 mg/L | Mu et al. (2012) |
| 3T3-L1 fibroblasts | Colloidal silica (LUDOX® CL and CL-X and non-stabilised SAS particles)* | 20, 30, 80, 400 nm | Commercial and laboratory | 4 and 40 μg/mL in DMEM; 3, 6, and 24 h incubation | Negative | Barnes et al. (2008) |
| SH-SY5Y neuronal cell line | Colloidal silica (LUDOX® AS-20, CL and AM, polygon)* | 12 nm (nominal) | Commercial | 48 h, up to 1000 ppm | Inconclusive | Kim et al. (2010) |
| Primary rat alveolar macrophages | Precipitated silica (NM-200) | 230 m2/g | JRC | 4 and 24 h incubation; 0, 0.01, 0.05, 0.25, 2.5, and 10 (10 only for 4 h incubations) µg/cm2; positive control D12 (25 µg/cm2; 4 h) | Negative, no oxidative DNA lesions; cytotoxic at highest dose level tested | CEFIC (2012c), |
| A549 | Colloidal silica (Levasil®)* | 9, 15, 30, 55 nm | AkzoNobel | Alkaline unwinding, 100–300 µg/mL | 30, 55 nm: >50 µg/mL DNA damage; 9, 15 nm: at higher concentrations, at 100 µg/mL oxidative damage | Maser et al. (2015) |
| Rat, lung | Colloidal silica (Levasil®)* | 15, 55 nm | AkzoNobel | Precision cut slices, 10–300 µg/mL | 15, 55 nm: >100 µg/mL DNA damage; no overt cytotoxicity | Maser et al. (2015) |
| V79 hamster lung fibroblasts | Colloidal silica (Levasil®)* | 15, 55 nm | AkzoNobel | Alkaline comet assay, alkaline unwinding assay, 100–300 µg/mL, ± Fpg |
15 nm: ↑strand breaks at 100 µg/mL (>twofold); no oxidative damage; no cytotoxicity 55 nm: ↑strand breaks at 300 µg/mL (>twofold); no oxidative damage; no cytotoxicity |
Maser et al. (2015) |
| V79 hamster lung fibroblasts | Silica gel (Spherisorb®)* | 5 µm | Commercial | 3 h treatment | Positive at ≥68.9 µg/cm2 | Zhong et al. (1997) |
| V79 hamster lung fibroblasts | 2 pyrogenic, 1 precipitated, and 2 precipitated colloids* | 20 and 25/70 nm (pyrogenic), 20 nm (precip.), 15, 40/80 nm (colloid)/50–200 m2/g | Commercial | 12.5, 25, 50, 100 mg/L, 24 h, comet assay with and without Fpg | Positive only with Pyr20 and Col15 in the presence of cytotoxicity and with Fpg, but no change in ROS; indicating indirect mechanisms | Guichard et al. (2015) |
| Phosphorylated gamma-H2Ax foci | ||||||
| Caco-2 cell line | Colloidal silica (Levasil® 50, Levasil® 200)* | 15, 55 nm | HC Starck | Phosphorylated gamma-H2Ax foci, 24 h, 4–64 µg/mL/1.25–20 µg/cm2 | 15 nm: threefold↑ at 32 µg/mL and fivefold↑ at 64 µg/mL; likely a result of apoptosis as the caspase was also↑ 55 nm: no effect |
Tarantini et al. (2015b) |
| Human HT-29 intestinal epithelial cell line | Mesoporous silica, core dye doped with two different labels* | 25, 100 nm | Laboratory | Phosphorylated gamma-H2Ax foci, 24 h, 10, 50, 150 µg/mL | 25 nm: 10, 50 (↑), 150 ↑ 100 nm: 10, 50 ↑, 150 no effect |
Sergent et al. (2012) |
cytB Cytochalasin B, HPRT hypoxanthine guanine phosphoribosyltransferase, NIST National Institute of Standards and Technology (USA), ROS reactive oxygen species
* Substance which does not fulfil the current EU criteria for E 551 (no star does, however, not implicate that the substance would be in compliance with EU E 551 specifications)