Nope. It's basically referring to the hex code you use when everything is opaque. Adding transparency doesn't change the hex code. 

If you created a sprite using only black and put it on a background of white, using an alpha channel would enable you to create a full grayscale sprite with only two colors and an alpha, which is technically in line with the stated rules, but goes against the spirit of the rules. This was clarified in another post. Using sprites to get multiple shades in this way would be considered using multiple colors.