Winterization is a simple method to remove saturated fatty acid contents in biodiesel fuels for improving their cold flow properties. In this work, biodiesel fuels with different initial long-chain (C16 and above) saturated fatty acid constituents (Si) were prepared from blends of palm, canola, and corn oils. The prepared biodiesels were treated at various winterization temperatures (Tw) to investigate the effect of Tw and Si on the final saturated fatty acid constituents (Sw) of the winterized biodiesel fuel. Optical microscopy showed that ball-like crystals formed with fluid regions at moderate cooling rates (-6 °C/h) could allow solid-liquid separation by filtration. A saturated fatty acid reduction ratio, Rs, defined as (Si - Sw)/Si × 100, was used with the experimental results on large samples (ca. 600 mL) to develop a correlation for winterization temperature as Tw (°C) = 0.659 Si (wt%) - 0.104 Rs (wt%) - 10.197. The correlation can provide estimation of the required winterization temperature for reducing a specified ratio of fatty acids in a biodiesel fuel that mainly contains long-chain fatty acids from the initial saturated fatty acid constituents. When used with literature relationships for cold filter plugging point (CFPP) and Sw, estimation of the CFPP of winterized biodiesel fuels is possible without requiring actual winterization treatment.