Recognizing the biocompatibility of various carboxylic acid-based molecules and protein stabilization by pH controls, this work demonstrates acid-catalyzed acylation reaction in nonaqueous acid media (Figure ). Our previous studies demonstrated protein bioconjugation in protein-compatible nonaqueous media such as ionic liquid and fluoroalcohol solvents. Notably, the urea-forming lysine modification process in ionic liquid did not cause any noticeable effects on native protein abilities such as streptavidin and antibody binding capabilities even by introducing the strong base/nucleophile iminophosphorane (pKa of its conjugate acid is >20 ). In aqueous media, extremely acidic and basic conditions are often avoided for studying proteins in part due to the potential hydrolysis of amide backbones, though those extreme pH ranges could be helpful for the prevention of protein precipitation in terms of their isoelectric points (Figure ). Because a hydrolysis process would not be a favorable chemical event in nonaqueous media with a substantially low concentration of water compared to aqueous solutions, we hypothesized that extreme pH conditions could be potentially protein-compatible, which has been demonstrated by the iminophosphorane-based bioconjugation in ionic liquid as an extremely basic process. Carboxylic acid-based molecules are often biocompatible and used as common buffer components for biochemical studies (e.g., acetate, glycine, and citrate buffers), and therefore, we examined alcohol-selective chemical reactions in nonaqueous carboxylic acid-based media as an extremely acidic bioconjugation process. In particular, our focus was an acid-catalyzed acylation approach (i.e., Fischer esterification) that would be not only potentially biomolecule-compatible due to the abovementioned reasons, but would the acidic media also suppress the reactivity of many intrinsically basic/nucleophilic amino acid side chains through their protonation. Catalytic acylation processes in an acidic medium enabled chemoselective labeling toward the alkyl hydroxyl groups over other nucleophilic side chains at an amino-acid level (Figures ). To test our hypothesis, N-protected canonical amino acids were subjected to the acylation processes with acetic acid as a solvent and a catalytic amount of scandium triflate. The acylation products were observed only for serine and threonine in liquid-chromatography mass spectrometry (LC-MS) analysis with a higher selectivity toward serine over threonine (Figure ). In contrast, other nucleophilic amino acids including tyrosine, lysine, arginine, histidine, tryptophan, glutamine, glutamic acid, and methionine did not show meaningful product formation (Figures ), indicative of the high chemoselectivity through suppression of the reactivities of those typical nucleophiles. Peptide-substrate scope investigation and tandem-mass spectrometry (MS/MS) also suggested chemoselective modification of alkyl hydroxyl groups and suggested more efficient modification of serine compared to threonine (Table , Figures ).