You are overwriting the fill aesthetic inside the geom_col() function, instead of that you have to set the colors at the scale level (also notice the syntax differences in my example).
library(ggplot2)
library(scales)
library(forcats)
freqHMatM <- data.frame(
stringsAsFactors = FALSE, NA,
HMatM = c(3L,4L,5L,2L,3L,4L,5L),
Gender = c("Homem", "Homem","Homem", "Mulher","Mulher", "Mulher","Mulher"),
n = c(5L,14L, 40L,2L,4L,11L,40L),
Frequency = c(8.47, 23.73, 67.80, 3.51,7.02, 19.30, 70.17)
)
colors <- setNames(c("mistyrose3", "mistyrose2", "mistyrose1", "mistyrose4",
"mistyrose3", "mistyrose2", "mistyrose1"),
freqHMatM$Frequency)
ggplot(freqHMatM, aes(x = Gender,
y = Frequency,
fill = fct_rev(fct_inseq(as_factor(Frequency))),
label = round(Frequency, 4))) +
geom_col(position ="fill",
color = "white") +
geom_text(position = position_fill(vjust = 0.5)) +
labs(title = "HMatM",
x = NULL,
y = "Frequency",
fill = "Frequency") +
scale_y_continuous(labels = percent) +
scale_fill_manual(values = colors) +
theme_classic(base_size = 18)

Created on 2020-11-29 by the reprex package (v0.3.0.9001)
Another thing to have in mind is that you are mapping a continuos variable to a discrete scale (by setting a discrete selection of colors), this doesn't seem like a good idea, consider this approach instead.
library(ggplot2)
library(scales)
freqHMatM <- data.frame(
stringsAsFactors = FALSE, NA,
HMatM = c(3L,4L,5L,2L,3L,4L,5L),
Gender = c("Homem", "Homem","Homem", "Mulher","Mulher", "Mulher","Mulher"),
n = c(5L,14L, 40L,2L,4L,11L,40L),
Frequency = c(8.47, 23.73, 67.80, 3.51,7.02, 19.30, 70.17)
)
ggplot(freqHMatM, aes(x = Gender,
y = Frequency,
fill = Frequency,
label = round(Frequency, 4))) +
geom_col(position ="fill",
color = "white") +
geom_text(position = position_fill(vjust = 0.5)) +
labs(title = "HMatM",
x = NULL,
y = "Frequency",
fill = "Frequency") +
scale_y_continuous(labels = percent) +
scale_fill_gradient(low = "mistyrose4", high = "mistyrose1") +
theme_classic(base_size = 18)

Created on 2020-11-29 by the reprex package (v0.3.0.9001)