Shaun Whalley's arrest follows that of striker Ricky Miller.
A club statement said Whalley has been suspended pending a police inquiry.
A Bedfordshire Police spokesman said a 27-year-old from Hitchin in Hertfordshire had been arrested on suspicion of assault.
Luton have launched an internal investigation and the statement added that the club "takes any allegation against a player or member of staff very seriously".
Miller, who has also been suspended by the League Two club, has been released on police bail.
A second Luton Town player has been arrested on suspicion of assault following an incident in the early hours of Monday morning.