Polymer brushes of polymethacrylic acid (PMAA) and PMAA-associated polymer blends of PMAA/PNIPAM (poly-N-isopropylacrylamide) were prepared on porous silicon for further investigation of the EDC/NHS (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide/N-hydroxysuccinimide) activation mechanisms by infrared spectroscopy. When the fragmentation degree of PMAA blocks in PMAA-associated polymer blends is increased, the production of anhydride wanes from dominant to recessive, whereas a complementary product of the NHS-ester waxes from recessive to dominant. The Thorpe-Ingold effect was applied to explain the formation of anhydride: the gem-dialkyl groups of PMAA next to the carboxylic acids compress the acid side chains close to each other; thus, once the intermediate of O-acylisourea forms, it will be attacked by the intramolecular neighboring acid much faster than any other nucleophiles such as NHS and water, and therefore the six-membered ring of the anhydride will be formed. All acid side chains in PMAA standing next to each other will form an anhydride, primarily due to the Thorpe-Ingold effect, unless they are sterically hindered, whereas only isolated acid side chains form the NHS-ester. The EDC/NHS activation results for four small molecules of dicarboxylic acids in aqueous media, namely, glutaric acid and 2,2-dimethyl glutaric acid, which generate disuccinimidyl ester with high yield, and succinic acid and 2,2-dimethyl succinic acid, which remain intact, can also be explained by the Thorpe-Ingold effect. A clear understanding of the EDC/NHS activation mechanisms of PMAA will take us a step closer for resolving the mechanistic ambiguity of the carbodiimide/additive coupling reactions for amide bond formation.