Table 2.
In vivo genotoxicity studies with acrylamide (AA) and/or glycidamide (GA)
| Test | Species | Experimental design and doses | Results | Comments | Reference |
|---|---|---|---|---|---|
| Micronucleus test in RET |
F344 rats and B6C3F1 mice (M) (8 weeks old; 7 animal/dose group) |
Oral (drinking water) AA in rat: 0, 0.5, 1.5, 3, 6, 12 mg/kg bw per day AA in mice: 0, 0.5, 1.5, 3, 6, 12, 24 mg/kg bw per day For 30 days. Positive control: ENU |
Negative AA: in rats Positive AA: in mice (6–24 mg/kg per day) |
Hobbs et al. (2016), Chepelev et al. (2017) | |
| Mutation in RET and RBC (Pig‐a gene) |
Equivocal AA in rats: increased mutation frequency only at highest dose; only in RET (but not in RBC); only in 3 out of 7 animals. Equivocal AA in mice: increased mutation frequency only at an intermediate dose; only in RET (but not in RBC). |
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| Micronucleus test in bone marrow |
Swiss albino mice (M, F) (10–12 weeks of age, 4 animal/dose group) |
Oral (drinking water) AA: 0, 2, 4, 8, 16, 32 mg/kg bw per day Exposure: 30 days Analysis: 24 h after the last exposure Positive control: doxorubicin |
Positive AA: dose‐related increase from the lowest dose. | Algarni (2018) | |
| Chromosome aberrations in bone marrow |
Positive AA: dose‐dependent increase in polyploidy, chromatid gaps, Robertsonian centric fusions and stickiness. Decreased mitotic index (slow progression of cells from S‐ to M‐phase of the cell‐cycle). |
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| Micronucleus test in RET |
gpt delta mice (M) (8 weeks of age, 5–6 animals/dose group) |
Gavage AA: 7.5, 15, 30 mg/kg bw per day for 28 days Analysis: 3 days (for micronuclei) and 3 and 49 days (expression time for mutation) at the end of the treatment. Positive control: ENU |
Positive AA: dose‐ dependent increase in micronuclei (significant increases at 15 and 30 mg/kg bw per day). |
Changes in mutational classes: sperm (GC > TA, GC > AT, 1 bp deletions); AT > TA (lung). Important discussion on sensitivity of germ cells to AA genotoxicity (spermatogonial cells less sensitive). |
Hagio et al. (2021) |
| Mutations at the gpt gene in testes, sperm and lung |
Positive AA: increased mutation frequency Testis and lung: two‐ to threefold increases at 30 mg/kg bw per day. No difference between 3 and 49 days of analyses. Sperm: sixfold increase at 30 mg/kg per bw (only after 3 days) |
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| Micronucleus test in bone marrow |
Kunming mice (M) (6–7 weeks old; 10 animals/dose group) |
i.p. injection AA: 50 mg/kg bw per day, for 5 days. Oxidative stress markers: SOD, GSH‐Px, MDA. |
Positive AA |
Main focus on inhibitory role of several antioxidants on AA genotoxicity. See Table 4 for further information. |
Zhao et al. (2015a) |
| Comet assay in lymphocytes and liver | Positive AA: as measured by tail length, olive tail moment and tail % DNA. | ||||
|
Micronucleus test in bone marrow |
Kunming mice (M) (6–7 weeks old; 10 animals/dose group) |
i.p. injection AA: 50 mg/kg bw per day, for 7 days. Oxidative stress markers: ROS, SOD, GSH‐Px, GST, GSH, γ‐GCS, P450 2E1. |
Positive AA |
Main focus on inhibitory role of blueberry anthocyanin extract antioxidant properties on AA genotoxicity. See Table 4 for further information. |
Zhao et al. (2015b) |
| Comet assay in lymphocyte and liver cells | Positive AA: as measured by tail length, olive tail moment and tail % DNA | ||||
| Meiotic maturation in mouse oocytes | Germinal vesicle‐stage mouse oocytes from BALB/c mice (F) |
i.p. injection F BALB/c mice (n = 12) AA: 25 mg/kg per day, for 7 days. |
Positive AA: significant reduction in % of metaphase II oocytes compared to controls, meiotic spindle mass reduced, chromosome misalignments. | Reduced maturation in in vivo experiments but negative results in in vitro experiments. | Aras et al. (2017) |
| Micronucleus test in bone marrow RET |
Wistar rat (M) (6 animals/dose group) |
Gavage AA: 0, 10, 15, 20 mg/kg bw, for 28 days. Markers of oxidative stress: MDA, GSH, SOD. Positive control: Cyclophosphamide |
Positive AA: dose‐dependent increase from 10 mg/kg per bw; altered PCE/NCE. |
Alterations: in several liver haematological parameters; in brain, spinal cord, liver, kidney, myocardium by histopathology; in neurobehavioural parameters. No significant changes in oxidative stress markers in liver and kidney. |
Jangir et al. (2016) |
| Micronucleus test in bone marrow |
Wistar rats (M) (5‐week‐old) |
Oral AA: 100 mg/kg bw per day Unclear exposure time |
Positive AA: significant increase in micronuclei. | Main focus on horseradish allyl isothiocyanate antioxidant properties in inhibiting AA‐induced DNA damage (micronuclei and comet) | Shimamura et al. (2017) |
| Comet assay in liver, kidney, brain | Positive AA: significant induction of DNA measured by tail intensity in all organs. No difference in bw and relative weights of liver and kidney. | ||||
| Micronucleus test in bone marrow |
Sprague‐Dawley rats (F) (12–14 week of age< 5 animals/dose group) |
Oral AA: 50 mg/kg bw per day, for 30 days. |
Positive AA: increased micronuclei at decreased PCE/NCE ratio. Measurements of MPO activity, urinary 8‐OHdG, levels of GSH, MDA, PCO, TBARS. |
Main focus on argan oil antioxidant properties on AA‐induced clastogenicity. See Table 4 for further information. |
Şekeroğlu et al. (2017) |
| Micronucleus test in peripheral blood |
F344 rats (M) (8 weeks old; 6 animals/dose group) |
Gavage AA: 0, 0.33, 0.66, 1.32, 2.7, 5, 10, 20 mg/kg per day, for up to 29 days Analysis micronuclei: day 4 and 29. Positive control: scoring controls. |
Negative AA. | Dobrovolsky et al. (2016) | |
| Comet assay in liver, kidney, bone marrow |
Analysis Comet: day 29 (analysis 3 h after final dose) For short‐term comet assay: 250 mg/kg per day GA for 3 days starting at ~12 weeks of age. Positive control: MMS |
Positive AA: in liver (weak), but not kidney and bone marrow. | |||
|
Mutation in RET and RBC (Pig ‐a gene) |
Analysis Pig‐a: day 15, 29, 56. Positive control: mutant‐mimic samples. |
Positive AA: only at 20 mg/kg per day at day 56, only in RBC. | |||
| Micronucleus test in RET | F344/DuCrl rats (M) |
Drinking water AA: 0, 0.5, 1.5, 3, 6, 12 mg/kg bw per day for 30 days. |
Negative AA |
No changes in final bw or bw gain in exposed versus control animals. Data on transcriptional profiling. See Table 4 for further information. |
Chepelev et al. (2017), Hobbs et al. (2016) |
| Mutation in RET and RBC (Pig‐a gene) | Equivocal AA | ||||
| Comet assay in liver cells |
Wistar rats (M) (6 animals/ dose group) |
i.p. injection AA: 50 mg/kg bw Analysis: 48 h after injection Oxidative stress markers, GST, 8‐OHdG; Histopathological findings |
Positive AA: as measured by tail length, tail moment and tail % DNA |
Main focus on inhibitory role of quercetin antioxidant properties on AA genotoxicity. See Table 4 for further information. |
Ansar et al. (2016) |
| Comet assay in mouse spermatozoa |
Swiss CD1 mice (M) (3 animals/dose group) |
Oral (drinking water) AA: 1 ug/mL, for 6 months |
Positive AA: increased DNA breaks in spermatozoa without a concomitant reduction in overall fertility. Increased DNA breaks also in the spermatozoa of offspring (F1). Small increase in DNA 8‐OHdG. |
See Table 4 for further information. | Katen et al. (2016a) |
| Comet assay in spermatozoa |
Swiss CD‐1 mice (M) (5–6 week of age; 3 and 6 mice for 3 and 6 month time points, respectively) |
Oral (drinking water) AA: 0.18 mg/kg bw per day Exposure: 3 and 6 months +/− resveratrol: once a week. |
Positive AA |
See Table 4 for further information (small increases in 8‐OHdG and γ‐H2AX foci) |
Katen et al. (2016b) |
| 8‐OHdG by immunostaining | Positive AA: both at 3 and 6 months AA exposure. | ||||
| γ‐H2AX by immuno‐fluorescence | Positive AA: increase in post‐meiotic germ cells (late spermatids and spermatozoa) both at 3 and 6 months AA exposure. | ||||
| Comet assay in spermatocytes and spermatozoa of mice |
Swiss CD‐1 mice (M) (6 animals/dose group) |
i.p. injection AA: 25 mg/kg bw per day, for 5 consecutive days Analysis: 3–5 days (spermatozoa) or 24–26 days (spermatocytes) following the last injection. |
Positive AA: at both stages of spermatogenesis. |
Epididymal CYP2E1 plays a critical role in AA‐induced DNA damage in spermatozoa and paternally mediated embryonic resorptions. See Table 4 for further information. |
Katen et al. (2017) |
| Comet assay in corneal epithelial cells |
Japanese white rabbits (M) (11–12 weeks of age, 2‐4 animals/dose group) |
Eye instillation AA: 0.6% and 3% AA, 50 µL per eye. Analysis: 2 h after instillation Also tested: ethidium bromide, paraquat, MMS, 4‐NQO |
Negative AA: comet assay as measured by % tail DNA. | Limited information from this study because of unusual animal model for genotoxicity testing and AA delivery. | Tahara et al. (2019) |
| γ‐H2AX foci in the urinary bladder |
B6C3F1 mice (M) (5 weeks of age; 10 animals/dose group) |
Oral (drinking water) AA: 0.005% (corresponding to 9.85 mg/kg bw daily intake) Exposure time: 4 weeks |
Negative AA | Limited information from this study because of the use of AA as a negative control (a genotoxic non bladder carcinogen). | Sone et al. (2019) |
|
UDS in corneal epithelial cells |
Japanese white rabbits (M) (11–12 weeks of age, 2‐4 animals/dose group) |
Eye instillation AA: 3% AA, 50 µL per eye. Analysis: 2 h after instillation Also tested: paraquat, acridine orange, ethidium bromide, 4‐NQO |
Negative AA | Limited information from this study because of unusual animal model for genotoxicity testing and AA delivery. | Tahara et al. (2021) |
| Mutation in RET and RBC (Pig‐a gene) | F344 rats (M) |
Oral (drinking water) 0, 25, 50, 100, 137.5, 175 mg/kg per day, for 28 days Analysis: 2, 7, 14, 28 days Positive control: ENU |
Negative AA | Body weight gain reduced in two highest dose groups; evidence of reticulocytosis at high doses. | Horibata et al. (2016) |
| Mutation in brain (cII gene) | Big Blue mice (M, F) |
Oral (drinking water) AA and GA: 0, 1.4, 7.0 mM, for 4 weeks |
Positive AA and GA: increased mutation frequency only in males at 7.0 mM. twofold increase in GC > TA; AT > TA and AT > CG. |
Similar spectra for AA and GA. Data also for lung, liver and testis from Wang et al. (2010); Manjanatha et al. (2015). | Li et al. (2016) |
AA: acrylamide; bw: body weight; ENU: N‐ethyl‐N‐nitrosourea; F: female; GA: glycidamide; γ‐GCS: γ‐glutamylcysteine synthetase; GST: gluthathione S‐transferase; GSH: glutathione; GSH‐Px: glutathione peroxidase; 8‐OHdG: 8‐hydroxy‐2'‐deoxyguanosine; M: male; MDA: malonaldehyde; MMS: methyl methanesulfonate; MPO: myeloperoxidase; 4‐NQO: 4‐nitroquinoline 1‐oxide; PCE/NCE: polychromatic erythrocytes/normochromatic erythrocytes; PCO: protein carbonyl; ROS: reactive oxygen species; RBC: red blood cells; RET: reticulocytes; SOD: Superoxide dismutase; TBARS: thiobarbituric acid reactive substances; UDS: unscheduled DNA synthesis.