An unsaturated fatty acid auxotroph of Escherichia coli was grown with a series of cis-octadecenoate isomers in which the location of the double bond varied from positions 3 to 17. Each of these fatty acid isomers was incorporated into the cellular lipids, but cyclopropane derivatives were formed to at least a 3-fold greater extent from the cis-9 and cis-11 isomers than from any other positional isomers. The extent of cyclopropane acid formation was observed to be highly dependent on the rate of shaking of the culture. A culture shaking at 340 rev./min converted 8.7% of its oleate to the cyclopropane derivative at stationary phase, whereas a parallel culture shaken at 110 rev./min converted 66% of the oleate to a cyclopropane acid. The inability to observe selectivity or form derivatives from isomers other than the cis-9 and cis-11 isomers seems to be due to enzyme specificity rather than a secondary affect of the abnormal unconverted fatty acids on the cell, because the cis-9 isomer is converted to its cyclopropane derivative even in cells grown with abnormal unreactive positional isomers. The preferred substrates for cyclopropanecarboxylic acid formation contained a cis ethylenic bond at either the 9 position or the (n-7) position. In combination with results of previous studies the specificity reported here supports a concetpt that two different enzymes may participate in cyclopropane ring synthesis. One enzyme activity may recognize its substrate by the distance from the pi-bond to the carboxyl group and the other by the distance to the methyl group.