Research and clinical studies show that the 2-hydroxylated estrogens (2-OH E2 and 2-OH E1) are a safer pathway of hydroxylation than the 4-hydroxyestrogens (4-OH E2 and 4-OH E1), which bind to and damage DNA, leading to mutations that are associated with increased breast cancer risk. If either 4-OH-E2 or 4-OH-E1 are higher than reference range, and not well methylated, this may indicate higher risk for DNA damage, mutations, and risk for developing breast cancer.

Hydroxylation of estrogens in the 2-position of estradiol (2-OH-E2) and estrone (2-OH-E1) is considered a safer pathway for metabolism than the 4-position hydroxylations of these estrogens. The 2-position hydroxylations (considered safer) are increased with cruciferous vegetables (and extracts of them) as well as iodine. The most commonly used are indole-3-carbinol (I3C) and its metabolite diindolylmethane (DIM). Iodine also increases the 2-hydroxylation of estrogens, with a slight increase in 4-hydroxylation. The more dangerous 4-hydroxylated estrogen metabolism is enhanced by exposure to environmental toxins, mostly petrochemical-based products but also heavy metals, that induce 4-hydroxylation pathway enzymes, and cause formation of Reactive Oxygen Species (ROS) that co-oxidize the catechol estrogens to more dangerous estrogen quinones.