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Effect of calcium phosphate and alumina C γ gels on the mutagenicity and cytotoxicity of dimethylnitrosamine as studied in the CHO/HGPRT system
Authors:Eng-Lay Tan  Abraham W Hsie
Institution:The University of Tennessee-Oak Ridge Graduate School of Boimedical Sciences and Biology Division, Oak Ridge National Laboratory, Oak Ridge, TN 37830, U.S.A.
Abstract:When CaCl2 was added in increasing concentrations to a rat liver metabolic activation system (S9) buffered with sodium phosphate, the mutagenic activity and cytotoxicity of dimethylnitrosamine (DMN) in the Chinese hamster ovary cell/hypoxanthine-guanine phosphoribosyl transferase (CHO/HGPRT) system were greatly increased. This effect was not observed with an S9 mix buffered with N-2-hydroxyethylpiperazine-N′-2-ethanesulfonic acid (HEPES). The calcium phosphate gel precipitate of the S9 mix possessed approximately built13 of the total activity of the mix, while the supernatant had only slight activity. However, when the calcium phosphate gel precipitate of a solution of S9 salts (without S9 protein) was added to the supernatant, the remaining 23 of the activity was recovered. Commercially obtained calcium phosphate, tricalcium phosphate, and alumina C γ gels could substitute for CaCl2 in the S9 mix, but diethylaminoethyl cellulose (DEAE cellulose) could not. Alumina C γ gel can exert its effect in the absence of both CaCl2 and phosphate in the S9 mix. Increasing the time of contact between the S9 protein and the S9 salts increased the efficacy with which the S9 mix activated DMN; this is indicative of an adsorptive process by calcium phosphate gel.
Keywords:CHO/HGPRT  Chinese hamster ovary cell/hypoxanthine-guanine phosphoribosyl transferase system  DEAE cellulose  diethylaminoethyl cellulose  DMN  F12  Ham's F12 medium  F12FBS10  Ham's F12 medium supplemented with 10% fetal bovine serum  F12FCM5  Ham's F12 mediu supplemented with 5% heat-inactivated  dialysed fetal bovine serum  HEPES
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