Provitamin A biofortification of staple crops may decrease the prevalence of vitamin A (VA) deficiency if widely adopted in target countries. To assess the impact of processing methods on the VA value of plant foods, the unique bioefficacies of cis-βC isomers (formed during cooking) compared with all-trans (at) β-carotene (βC) must be determined. The bioefficacies of 9-cis (9c)- and 13-cis (13c)-βC isomers were compared with those of the at-βC isomer and VA positive (VA+) and negative (VA - ) controls in VA-depleted Mongolian gerbils (Meriones unguiculatus) in two experimental studies (study 1, n 56; study 2, n 57). A 3- or 4-week depletion period was followed by a 3- or 4-week treatment period in which the groups received oral doses of the 9c-, 13c- or at-βC isomers in cottonseed oil (study 1, 15 nmol/d; study 2, 30 nmol/d). In study 1, the βC isomers did not maintain baseline liver VA stores in all groups (0.69 (SD 0.20) μmol/liver) except in the VA+group (0.56 (SD 0.10) μmol/liver) (P= 0.0026). The βC groups were similar to the VA+group, but the 9c- and 13c-βC groups did not differ from the VA - group (0.39 (SD 0.09) μmol/liver). In study 2, the βC isomers maintained baseline liver VA stores in all the βC groups (0.35 (SD 0.13) μmol/liver), and in the VA+group, the VA supplement (0.54 (SD 0.19) μmol/liver) exceeded the baseline VA status (0.38 (SD 0.15) μmol/liver) (P< 0.0001); however, the 9c-βC group did not differ from the VA - group (0.20 (SD 0.07) μmol/liver). In vivo isomerisation of βC was confirmed in both experimental studies. Lower VA bioconversion factor values were obtained for the cis-βC isomers in study 2 when compared with study 1, but higher values were obtained for the at-βC isomer. Dose and VA status clearly affect bioconversion factors. In conclusion, the cis-βC isomers yielded similar liver VA stores to the at-βC isomer in Mongolian gerbils, and liver VA stores of the 9c- and 13c-βC groups did not differ when the doses were provided at physiological levels over time in two studies.