Effects of pH in the in vitro chromosomal aberration test

The relation between the pH of the medium and clastogenic activity was studied in Chinese hamster ovary (CHO) K1 cells in vitro. The pH was adjusted with NaOH, KOH, HCl or H2SO4. No clastogenic activity was observed over the initial pH range of 7.3-10.9 without S9 mix, but a few chromosomal aberrations were induced at pH 10.4 with S9 mix. The frequency of aberrations increased with the increase in amount of S9. At acidic pH, many chromatid breaks were induced at initiatial pH 5.5 or below without S9 mix, and aberrations such as chromatid breaks and chromatid exchanges were induced at initial pH 6.2 or below with S9 mix. Using MES and Bis-Tris as buffers instead of sodium bicarbonate, we observed that aberrations of the chromatid break type were inducible at pH 6.2 or below. These results show that the combination of strong alkalinity and S9 is clastogenic to CHO-K1 cells, and also that weakly acidic media are genetically active. The results indicate that incubations at non-physiological pH might give false-positive responses.