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The present work investigates liquid antisolvent crystallization of cholesterol from a water (antisolvent)–ethanol (solvent)–cholesterol (solute) solution. It focuses on (i) a proposed mechanism of nucleation based on visual observations, (ii) mathematical modeling to validate the mechanism, and (iii) experimental parametric variations of particle size. The experiments demonstrate a 2-step sequence of primary and secondary nucleation. By employing the solid–liquid interfacial energy estimated from experimental induction time and kinetic parameters selected from the literature, the model predicts a particle diameter of 3.5 μm at 299.5 K after 30 s, in agreement with the experimental value of 3.8 μm, and two-level nucleation events of cholesterol to affirm the proposed mechanism. Experimental studies confirm that particle size decreases with initial supersaturation and antisolvent addition rate, and increases with temperature and batch time. At 290.5–299.5 K, the particle length increases from 6.1 to 7.2 μm after 4 h for the antisolvent to solution mode of addition, while it decreases to 5.7 μm at 299.5 K for the solution to antisolvent mode.

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