Benchmark performance of four sampling schemes (SS, I-IV) on 5/5/5 small/medium/large-sized reactions. The Schemes are summarized in Fig. . SSI is the lowest cost and performs minimal conformational sampling, SRNum4 is the most costly and performs the most conformational sampling. In the large-sized systems, direct activation energy estimation using the model reactions is no longer reliable. For example, two intramolecular Diels-Alder reactions (R3-1 and R3-2) with distinct activation energies share the same inter-molecular Diels-Alder model reaction, which can not capture the steric hindrance effect of forming a fused-ring structure in R3-2. Another source of error comes from the underestimation of the reactant energy. For instance, the model reactants in R3-1 and R3-5 simplify the original long-chain reactants into much smaller bimolecular reactants. Because the reactant energy of bimolecular reactants is defined as the summation of each separate reactant, the model reactants ignore the intramolecular interactions of the original reactants, resulting in a lower reactant energy and higher activation energy estimation. To achieve higher accuracy for complex reactions, the exact TSs need to be located. It is observed that all four sampling schemes successfully locate the most stable TSs for R3-4 and R3-5. SSIII and SSIV identify TSs with even lower energy for R3-1, due to the inclusion of constrained conformational sampling on the TS initial guesses. However, only SSIV identifies the most stable TS of R3-2 and R3-3. In fact, exploring the most stable TSs for these two reactions is an extremely challenging task and is analyzed in detail in SI Section 4.