Polymorphonuclear leukocyte (PMNL) was obtained from peritoneal cavity in rat treated with casein. Effects of divalent cations and drugs on leukotriene $B_4(LTB_4)$ formation from arachidonic acid in the PMNL were determined by HPLC assay. 5-Lipoxygenase, a key enzyme for $LTB_4$ formation from arachidonic acid, exhibited $V_{max}$ at $1;{ imes};10^{-5}M$, and $K_m$ at $9.89;{ imes};10^{-5}M$ of arachidonic acid. Optimal $Ca^{++}$ concentration for the enzyme activity was $1;{ imes};10^{-5}M$ but $Zn^{++}$ did not show any significant effects on the $LTB_4$ formation. Indomethacin and caffeic acid exhibited inhibitory effects on the $LTB_4$ formation at $1;{ imes};10^{-5}M$ and $4;{ imes};10^{-5}M$, respectively. However, 6-aminohexanoic acid did not show any significant effects on the $LTB_4$ formation.