Hetero-selective crosslinking of primary and secondary alkyl amines with CS2 under ambient aerobic oxidation for bioconjugation

The advancement of modern biotherapeutics demands robust tools to precisely engineer complex molecular constructs. Among these, efficient methods for crosslinking biorelevant molecules (BMs), in- cluding native biomolecules and synthetic drugs and probes, are foundational for constructing multifunc- tional bioconjugates. Despite decades of development, practical crosslinking strategies remain limited, largely relying on reactions with highly reactive but scarce thiol groups and often requiring wasteful, stepwise operations. Expanding the scope to include less reactive but more abundant amino groups could significantly enhance the bioconjugation toolbox, while the ability to differentiate amine subtypes would further refine chemoselectivity and improve precision in crosslinking. Umpolung of primary amines via isothiocyanation presents a promising approach for linking BMs through two nucleophilic handles. How- ever, conventional isothiocyanation reactions typically require highly electrophilic reagents or strongly oxidative conditions, rendering them unsuitable for complex molecular systems. Herein, we present a robust and non-chemist-friendly protocol for the isothiocyanation of primary alkyl amines using CS2 un- der ambient aerobic oxidation conditions, eliminating the need for additional promoters. These mild con- ditions not only ensure broad compatibility with endogenous functional groups in BMs but also unlock a powerful new capability: selective differentiation between primary and secondary alkyl amines. Notably, the unique suppression of homo-linking for both primary and secondary amines under these conditions enables near-perfect hetero-selectivity in cross-linking two different BMs via their abundant amino han- dles in a one-pot operation, offering a streamlined and thiol-independent approach to bioconjugation.