TRICHLOROACETIC-ACID - INVESTIGATION INTO THE MECHANISM OF CHROMOSOMAL DAMAGE IN THE IN-VITRO HUMAN LYMPHOCYTE CYTOGENETIC ASSAY AND THE MOUSE BONE-MARROW MICRONUCLEUS TEST

Trichloroacetic acid (TCA) was tested for its ability to induce chromo somal damage in cultured human peripheral blood lymphocytes and in bon e marrow cells of male and female C57BL/6J(f)BL10/Alpk mice, Two in vi tro cytogenetic assays were conducted with TCA, In the first TCA, as f ree acid, was added to whole blood cultures at final concentrations of 500, 2000 and 3500 mu g/ml in the presence and absence of an auxiliar y metabolic activation system (rat liver S9-mix), Statistically signif icant increases in the percentage of aberrant cells compared with solv ent control values were observed in cultures treated with TCA at 2000 and 5000 mu g/ml. Investigation into the effects of TCA on the pH of t he culture medium revealed significant reductions in pH at both these TCA concentrations, Neutralized TCA was then tested at concentrations of 500, 2 000 and 5000 mu g/ml, also in the presence and absence of S9 -mix, No statistically or biologically significant increases in the pe rcentage of aberrant cells were observed in any of these cultures, In the mouse micronucleus test, neutralized TCA was administered in two e qual intraperitoneal doses 24 h apart to C57BL/6J(f)BL10/Alpk mice (33 7, 675 and 1080 mg/kg in males; 405, 810 and 1300mg/kg in females), Th ese dose levels represent 25%, 50% and 80% of the median lethal dose ( MLD) in this strain of mouse, Bone marrow samples were taken 6 and 24 h after the second dose and the chromosomal damage assessed by analysi s of the bone marrow for micronuclei, No statistically or biologically significant increases in the incidence of micronucleated polychromati c erythrocytes compared with the solvent control dosed animals were ob served in either sex at the 6 h sampling time or in the females at the 24 h sampling time. A small but statistically significant increase in micronucleated polychromatic erythrocytes was observed in male mice 2 4 h after a dose of 675 mg/kg (50% MLD), Since no increases were noted at the 25 or 80% MLD, and the levels recorded are within the range of the concurrent solvent control values, the small increase observed in the males at the 50% MLD is considered not to be biologically signifi cant, Flow cytometric studies on suspensions of isolated liver cell nu clei revealed that changes in FITC binding (indicating altered chromat in conformation) were induced by pH changes alone and were not caused by neutralized TCA, Consideration of all the data presented in this pa per indicates that increase in chromosomal damage observed in human ly mphocyte cultures treated with TCA is a result of pH changes inducing artefactual chromosomal damage rather than to any clastogenic effects of TCA, TCA is therefore considered not to be clastogenic when examine d in the in vitro human lymphocyte chromosomal aberration assay or in the mouse bone marrow micronucleus test.