14The infections causes by Chikungunya virus (CHIKV), family Togaviridae, genus Alfavirus, have become 15 a health problem around the world, due to its widespread occurrence, the high morbidity rate and the 16 absence of vaccines or antiviral treatments. We analyzed a competitive inhibitor of ornithine 17 decarboxylase, enzyme key in the biosynthesis of Polyamines (PAs), N-ω-chloroacetyl-L-ornithine 18 (NCAO), as a possible inhibitor of CHIKV replication because intracellular polyamines participate in the 19 transcription and in vitro translation of CHIKV. NCAO does not have any cytotoxic effect on C6/36 cells 20 even at a concentration of 1000μM at 72h after post-exposure but in Vero cells the cytotoxic effect was 21 presented above 380μM at 48h post-exposure, which was considered when determining the inhibitory 22 effect on viral replication. We demonstrate that NCAO inhibits the replication of CHIKV in Vero and C6/36 23 cells in a dose-dependent manner causing a decrease in the PFU/mL of at least 4-logarithm (p <0.01) in 24 both cell lines. Viral yields were restored by the addition of exogenous polyamines, mainly putrescine. 25 HPLC analyses it shown that NCAO decrease content of intracellular PAs even though in infected cells 26 mainly decreased spermidine and spermine, so NCAO inhibits CHIKV replication, by depleting the 27 intracellular polyamines in Vero and C6/36 cells, suggesting that this compound is a possible antiviral in 28 CHIKV infections. 29 30 Importance 31 The infections caused by Chikungunya virus (CHIKV), genus Alfavirus, have become a worldwide health 32 problem, due to its high morbidity rate and the absence of vaccines or antiviral treatments. It is known 33 that CHIKV use intracellular poliamines during transcription and traduction process, so the depletion of 34 intracellular putrescine, spermidine and spermine reduce the viral replication. N-ω-chloroacetyl-L-35 ornithine (NCAO) is known as a competitive inhibitor of ornithine decarboxylase, key enzyme in 36 Polyamines biosynthesis, but no studies have proven its activity on the inhibition of polyamine-dependent 37 viral replication. Here we demonstrate NCAO inhibits in vitro CHIKV replication, so we propose NCAO as 38 an antiviral candidate.39