CYTOTOXICITY STUDIES OF ANCHORAGE-INDEPENDENT LS-L-929 MOUSE FIBROBLASTS USING MEMBRANE INTEGRITY, ATP CONTENT AND ATP ADP RATIO AS DETERMINANTS - INCLUDING THE XENOBIOTIC EFFECTS OF COMPONENTS IN AN EXPERIMENTAL SHAMPOO FORMULATION

Cell death (CD50), as measured by loss of membrane integrity using flu diacetate and ethidium bromide, and reduction in ATP content (ATP50), were used as endpoints in a comparative study of the cytotoxicity of 40 chemicals (supplied by FRAME) to anchorage-independent LS-L-929 mou se fibroblasts. The rank order correlation of the two test was strong (r = 0.98), but the ATP50 values only showed a weak correlation with r at acute lethality (LD50) data (r = 0.43). More-mechanistic endpoints were sought in order to characterise cellular energy status, and the e ffects of acetaminophen, acetylsalicylic acid, diazepam, digoxin and e thanol on ATP/ADP ratio, phosphorylation potential and adenylate energ y charge were examined in detail. The first two ratios were very sensi tive to toxic insult to the cells and gave results which more closely mimicked differences in human acute oral lethality than did the ATP50 and CD50. In addition, there was evidence that digoxin and diazepam af fected oxidative phosphorylation, which suggested an alteration to mit ochondrial function. Energy charge appeared less sensitive to the xeno biotics, probably owing to the dampening effect of the denominator in the ratio. The effects of individual components of an experimental det ergent-based formulation on cell death were examined. It was found tha t one detergent gave different results, depending on the presence or a bsence of a second detergent. A commercial cream containing varying am ounts of glyceryl monostearate contaminated with varying levels of sod ium dodecyl sulphate gave results which reinforced the point that cyto toxic assays are valuable in quality assurance.