It is important to determine the cytochrome P450 (CYP) contribution of certain drugs by taking into consideration the attrition due to issues such as genetic polymorphism and inter-individual variation. In many cases in the early discovery stage, the metabolites of a new chemical have not been identified. Therefore, the present paper devised an approach in which the in vitro intrinsic clearance (CLint) value for new chemicals was determined by measuring substrate depletion. The following prediction methods were compared to calculate CLint using data from recombinant CYP enzymes: (1) the relative CYP content in human liver microsomes; (2) the relative activity factor (RAF) based on the Vmax value; and (3) the RAF value based on the CLint value. The most accurate prediction method was RAF based on CLint. This method would be useful in the early drug-discovery process in cases in which the main metabolite is not identified.