FRET-Derived Ratiometric Fluorescent K+ Sensors Fabricated from Thermoresponsive Poly(N-isopropylacrylamide) Microgels Labeled with Crown Ether Moieties

Abstract

We report on the fabrication of ratiometric fluorescent K(+) sensors based on thermoresponsive poly(N-isopropylacrylamide) (PNIPAM) microgels covalently incorporated with K(+)-recognizing 4-acrylamidobenzo-18-crown-6 residues (B18C6Am), fluorescence resonance energy transfer (FRET) donor dyes, 4-(2-acryloyloxyethylamino)-7-nitro-2,1,3-benzoxadiazole (NBDAE), and rhodamine-B-based FRET acceptors (RhBEA) by utilizing K(+)-induced changes in microgel volume phase transition (VPT) temperatures. P(NIPAM-B18C6Am-NBDAE-RhBEA) microgels were synthesized via the free radical emulsion copolymerization technique. The spatial proximity between FRET pairs (NBDAE and RhBEA dyes) within microgels can be tuned via thermoinduced collapse and swelling of thermoresponsive microgels above and below VPT temperatures, leading to the facile modulation of FRET efficiencies. B18C6Am moieties within P(NIPAM-B18C6Am-NBDAE-RhBEA) microgels can preferentially capture K(+) via the formation of 1:1 molecular recognition complexes, resulting in the enhancement of microgel hydrophilicity and elevated VPT temperatures. Thus, the gradual addition of K(+) into microgel dispersions at intermediate temperatures, i.e., between VPT temperatures of P(NIPAM-B18C6Am-NBDAE-RhBEA) microgels in the absence and presence of K(+) ions, respectively, can directly lead to the reswelling of initially collapsed microgels. This process can be monitored by changes in fluorescence intensity ratios, i.e., FRET efficiencies. The presence of FRET pairs within P(NIPAM-B18C6Am-NBDAE-RhBEA) microgels allows for the facile in situ monitoring of thermoinduced and K(+)-induced VPTs of dually responsive microgels. The response time for fluorescent K(+)-sensing was further investigated via the stopped-flow technique, which reveals that the process completes within ∼4 s. This work represents the first report of thermoresponsive microgel-based ratiometric fluorescent sensors for both K(+) ions and temperatures.