마우스 [NIH (GP)] 태아세포 일차 배양계에 투여된 Benzo(a)pyrene과 세포내 고분자 화합물의 결합량 측정을 시도하였다. $0.5;{mu}M;-8.0;{mu}M$의 $^3H$-B(a)P 첨가 배지로 24시간 배양한 후, $^3H$-B(a)P의 세포내외 분포도를 조사한 결과, 가해준 $^3H$-B(a)P의 90% 이상은 배지내에 잔류되어 있었고, 2-10%는 세포 표면에 가볍게 부착되었으며 (alcohol soluble fraction) 0.5%미만은 세포내 고분자 화합물에 결함되었다(alcohol insoluble fraction). B(a)P 24시간 처리군의 세포내 DNA, RNA 단백철 농도는 대조군에 비하여 변화가 없었다. 이때 DNA에 결합되어 냐타나는 $^3H$-B(a)P의 비활성도는 첨가된 B(a)P 농도에 비례하여 증가하다가 $2;{mu}M$이상 투여시 $24;{mumu}moles/mg$으로 포화됨을 확인하였다. 단백질과의 결합량은 B(a)p $2;{mu}m$ 까지는 농도에 비례증가하였으며, $8;{mu}m$까지 B(a)P와 단백질 결합이 계속 증가하는 것을 확인 하였다(r=0.99).
We describe here the binding of benzo(a)pyrene to the cellular macromolecules of mouse embryonic fibroblasts in primary culture. On the 3rd day of the culture, DME medium containing 10% FCS was removed and the cells were incubated with fresh medium containing $1;{mu}M$, $2;{mu}M$ and $4;{mu}M$ of benzo(a)pyrene for 24 hours. Cell growh was inhibited more than 20% after the first 48 hours and more than 50% after 96 hours. B(a)p cytotoxicity was not so severe with the 24 hour treatment. $^3H$-B(a)P was applied to the cell monolayers and more than 90% remained in the culture medium after a 24 hr incubation and about 2 to 10% was loosely bound to the cell surface. The $^3H$-B(a)P tightly bound to the cellular macromolecules was less than 0.5%. The contents of cellular DNA, RNA and protein were not changed with the treatment of 2 to $8;{mu}M$ of B(a)P for 24 hours. Specific binding of $^3H$-B(a)P to DNA was linearly increased depending on the concentration of B(a)p lower than $2;{mu}M$, and it was saturated at $24;{mumu}moles/mg$ with higher than $2;{mu}m$ of B(a)p. Specific activity of $^3H$-B(a)P-protein-protein adduct was also lineraly increased with $2;{mu}m$ of B(a)p, but it was not saturated up to $8;{mu}M$ of B(a)p.
