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Using Chinese hamster ovary K1 cells, chromosomal aberration tests were carried out with formic acid, acetic acid and lactic acid, and the relationship between the pH of the medium and the clastogenic activity was examined. The medium used was Ham's F12 supplemented with 17 mM NaHCO3 and 10% fetal calf serum. All of these acids induced chromosomal aberrations at the initial pH of ca. 6.0 or below (about 10-14 mM of each acid) both with and without S9 mix. Exposure of cells to about pH 5.7 or below (about 12-16 mM of each acid) was found to be toxic. When the culture medium was first acidified with each of these acids and then neutralized to pH 6.4 or pH 7.2 with NaOH, no clastogenic activity was observed. Using F12 medium supplemented with 34 mM NaHCO3 as a buffer, no clastogenic activity was observed at doses up to 25 mM of these acids (initial pH 5.8-6.0). However, it was found that about 10% of the cells had aberrations at pH 5.7 or below (27.5-32.5 mM of each acid). Furthermore, when 30 mM HEPES was used as a buffer, chromosomal aberrations were not induced at doses up to 20 mM formic acid and acetic acid (initial pH 7.0-7.1), and at doses up to 30 mM lactic acid (initial pH 6.6). In the initial pH range of 6.4-6.7 (25-32.5 mM of each acid), chromosomal aberrations were observed. The above results show that these acids themselves are non-clastogenic, and the pseudo-positive reactions attributable to non-physiological pH could be eliminated by either neutralization of the treatment medium or enhancement of the buffering ability.

Quantitative determination of the blood level of a heparin-like substancewas made using a group of three calves with experimentally-induced bracken poisoning.The average blood level of the heparin-like substance in eight normal calves was 3.2+0.95 jtg / mZ (mdhn -4- s.d.). The blond level 61 the hepaririLIikc substance exceeded 6.0 7tg /mZ in all the affected animals in the hemorrhagic stage. It did not increase in the initialstage, when the clotting time prolonged mildly. This was due to the effect of a decreasein the platelet count observed in this stage. It was very likely that the prolonged clottingtime in the hemorrhagic stage might have been caused by the synergistic etrect of anincrease in the blood level of the heparin-like substance and the stimulated sensitivity ofthe affected blood to the heparin-like substance as a result of the decrease in the plateletcount.