Abstract: Brownies are one type of cake that has a characteristic blackish brown color. Brownies are made from a mixture of flour, eggs, margarine, chocolate powder and chocolate bars. The purpose of this study is to determined the level of acceptance of panelists and hardness in brownies substituted with seaweed flour Eucheuma cottonii. Variations in substitution of seaweed flour at the manufacture of steamed brownies had carried out with the percentage of seaweed flour at 0%, 5%, 10% and 15%. The research method used a completely randomized design of 4 treatments and 5 replication. Data were analyzed using the Kruskal-wallis test and the hardness analysis was analyzed using the One Way ANOVA test. The results showed that the substitution of Eucheuma cottonii seaweed flour has a significant effect on the hardness of brownies. The 10% concentration has a higher acceptability with a score of 3.54 with a hardness value of 48.48N.