The L(-)-carnitine production by biotransformation using the recombinant strain Escherichia coli pT7-5KE32 has been studied and optimized with crotonobetaine and D(+)-camitine as substrates. A resting rather than a growing cells system for L(-)-carnitine production was chosen, crotonobetaine being the best substrate. High biocatalytic activity was obtained after growing the cells under anaerobic conditions at 37 degreesC and with crotonobetaine or L(-)-carnitine as inducer. The growth incubation temperature (37 degreesC) was high enough as to activate the heat-inducible hp, promoter inserted in the plasmid pGPI-2. The best biotransformation conditions were with resting cells, under aerobiosis, with 3 g l(-1) and 100 mM biomass and substrate concentrations respectively. Under these conditions the biotransformation time (1 h) was shorter and the L(-)-carnitine yield (70%)) higher than previously reported. Consequently productivity value (11.3 g l(-1)h(-1)) was highly improved when comparing with other published works. The resting cells could be reused until eight times maintaining product yield levels well over 50% that meant to increase ten times the L(-)-carnitine obtained per gram of biomass.