The invention provides a preparation method of iron mine collector for reverse flotation and an application thereof and belongs to the technical field of beneficiation. The preparation method comprises the following steps of: (1) mixin0g drainage oil, methanol and concentrated sulfuric acid in a mass ratio of (1.0-1.2): (1.1-1.8): (0.05-0.09), and stirring the mixture for 5-12h at 60-90 DEG C to obtain a product 1; (2) mixing hydroxylamine hydrochloride, water and sodium hydroxide in a mass ratio of (1.0-1.3): (6.4-9.3): (1.9-2.4), and stirring the mixture for 1-3h at 20-30 DEG C to obtain a product 2; and (3) adding the product 1 into the product 2 in a weight ratio of (1.0-1.2): (0.28-0.48), and stirring the mixture for 4-6h at 40-60 DEG C to obtain the collector. The collector providedby the invention is wide in source of raw materials and simple in preparation process, has the characteristics of being good in dissolving property, good in selectivity, small in dosage and environmental-friendly, is suitable for low-temperature reverse flotation desilicication of low grade iron mine, effectively reduces the warming energy consumption, and is remarkable in economical benefit.