Use of enzymes in different parts of textile finishing processes has become popular, and several enzymes have been introduced into the textile industry. This study aimed to carry out the whole fabric pretreatment process in a single bath containing different enzymes. Ultrasonic cavitation was also tested to show its effect on enzyme-based finishing of cotton fabric. After optimization of the enzyme-based ultrasound-aided finishing, the fabrics were also colored in the same bath using natural dyes of pomegranate peel, nutshell, orange tree leaf, and alkanet root. Finally, it was observed that ultrasound-aided biopreparation of cotton fabric could provide sufficient pretreatment results. Coloration of these fabrics could be achieved with the use of natural dyes in the same bath as biopreparation.