Viability of enzymatically isolated and purified oat (Avena sativa L. ‘Markton’) mesophyll protoplasts was established by their ability to photosynthetically fix CO2 and to accumulate neutral red. When protoplasts were treated with 10-3 M alachlor [2-chloro-2′,6′-diethyl-N-(methoxymethyl)acetanilide], barban (4-chloro-2-butynyl m-chlorocarbanilate), dalapon (2,2-dichloropropionic acid), dicamba (3,6-dichloro-o-anisic acid), glyphosate [N-(phosphonomethyl)glycine], oryzalin (3,5-dinitro-N
4-dipropylsulfanilamide), paraquat (1,1′-dimethyl-4,4′-bipyridinium ion), picloram (4-amino-3,5,6-trichloropicolinic acid), or trifluralin [α,α,α-trifluroro-2,6-dinitro-N,N-dipropyl-p-toluidine), only alachlor and barban caused significant leakage of 14C-labeled material from the protoplasts as compared to controls. An abrupt increase in leakage occurred when protoplasts were treated with Tween 20 (polyoxyethylene 20 sorbitan monolaurate) at concentrations above 0.01% (v/v). The amount of leakage from protoplasts treated with dalapon or paraquat in combination with Tween 20 appears to be equivalent to the sum of the individual treatments. Treatment of the plasma membrane ATPase with the above named herbicides during the assay period shows that alachlor, barban, and dicamba reduces enzyme activity whereas glyphosate increases the activity. Pretreatment of the membrane ATPase with the herbicides in assay buffer [36mM Tris [tris-(hydroxymethyl)-aminomethane]-MES [2-(N-morpholino)ethanesulfonic acid], 3mM MgSO4, 50mM KC1] did not inhibit the enzyme activity during the assay period. Pretreatment with dicamba, glyphosate, and picloram in gradient buffer [1mM Tris-MES, 1mM MgSO4, 1mM dithiothreitol] did inhibit enzyme acitivty during the assay period. However, these herbicides cause the pH of the gradient buffer to be acidic.