A study of the genotoxicity of polyhydroxybutyrate
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Date
2009
Authors
Ali Mohammed, Abdulaziz Qaid
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Abstract
The purpose of this study is to determine the genotoxicity of polyhydroxybutyrate
(PHB), produced in short solid fibre form and manufactured by School of Biological
Sciences, Universiti Sains Malaysia, Penang, Malaysia, using three different tests: The
Salmonella mutagenicity test (Ames test), in vitro mammalian chromosomal aberration test
and gene expression analysis.
PHB was tested at various concentrations (0.3125, 0.625, 1.25, 2.5 and 5 mg/ml) in
all the three tests. For the Ames test, PHB was incubated with special genotype variants of
the bacterium Salmonella typhimurium, which carry mutations in several genes. Five tester
strains (TA1535, TA1537, TA1538, TA98 and TA100) were used, both with and without
metabolic activation system S9 mix and the test was assessed based on the number of
revertant colonies. Simultaneously, negative control tests using sterile distilled water and
positive control tests using sodium azide, 9-aminoacridine hydrochloride monohydrate
(98%) and 4-Nitro-O-phenylenediamine (98%) without metabolic activation system (S9)
and 2-aminoanthracene with metabolic activation system (S9) were carried out. To assess
the chromosomal aberrations, human osteoblast cell lines (HOS, CRL-1543) from
American type cell culture were exposed to PHB, sterile distilled water (Negative control)
and Mitomycin C (Positive control), both with and without metabolic activation system S9
mix. For the gene expression analysis, the fibroblast cell lines MRC-5 were treated with
PHB and incubated for 1, 12, 24 and 48 hours separately for each concentration. Total
RNA was isolated and analysed for the expression of p53, c-myc, bcl-xl and bcl-xs genes.
The results of Ames test showed that the average number of revertant colonies per
plate treated with PHB (28 to 344 colonies/plate without S9 and 150 to 499 colonies/plate
with S9) was less than double as compared to that of negative control (147 to 346
colonies/plate without S9 and 340 to 443 colonies/plate with S9). The absences of
increase in the number of revertant colonies by at least double with the test material
indicate that PHB was non-mutagenic. In the case of chromosome aberration test, there
was no indication of any mutagenicity due to PHB on the HOS cell line as revealed by the
mitotic index values [3.55(0.06) to 4.95(0.77) per cent without S9 and 3.10(0.14) to
5.20(0.98) per cent with S9]. Also, no chromosome aberrations were observed in the HOS
cell lines treated with PHB. The results of gene expression analysis carried out on
fibroblast cell line (MRC-5) treated with PHB at different timings did not show over or under
expression of the genes, p53 (Mean IDV range from 36100 to 36295), c-myc (Mean IDV
range from 33110 to 33270), bcl-xl (Mean IDV range from 31230 to 31443) and bcl-xs
(Mean IDV range from 33103 to 33290) as compared to that of negative controls (Mean
IDV range from 31230 to 36240).
Hence, the present study indicates that PHB produced by School of Biological
Sciences, Universiti Sains Malaysia, Penang, Malaysia is non-genotoxic under the present
test conditions.
Description
PhD
Keywords
Biological Science , Genotoxicity , Biomaterial