Chemoenzymatic Synthesis of 3‘-O-(Carboxyalkyl)fluorescein Labels

Abstract

A general and versatile method is described for the synthesis of fluorescent labels. Coupling of the 3'-phenol of fluorescein methyl ester with hydroxyalkyl benzyl esters, followed by benzyl ester hydrolysis, provided a series of fluorescein carboxyalkyl ethers. Use of the Mitsunobu reaction allowed for the introduction of linkers of different lengths onto the 3'-phenol of fluorescein. Chemoenzymatic benzyl ester hydrolysis was achieved with LPL-80 lipase, providing pH-independent labels useful for the preparation of fluorescent conjugates.

Labeling Tetracysteine‐Tagged Proteins with a SplAsH of Color: A Modular Approach to Bis‐Arsenical Fluorophores

“…Synthesis of 2-((2-tert-butyl carbamato)ethyl)-3',6'-dihydroxyspiro[1H-isoindole-1,9'-(9H)-xanthene-3(2H)-one], fluorescein spirolactam 2: By following the general procedure of Adamczyk and Grote, [15] fluorescein methyl ester [16] (0.52 g, 1.5 mmol) was dis- Fluorescein spirolactam 4',5'-Bis(1,3,2-dithiaarsolan-2-yl), SplAsH-Boc, 4: Compound 3 (0.5 g, 0.50 mmol) was suspended in THF (10 mL), and N,N-diisopropylethylamine (DIPEA; 0.7 mL, 4 mmol), arsenic trichloride (0.8 mL, 10 mmol), and PdA C H T U N G T R E N N U N G (OAc) 2 (5 mg) were added; this resulted in complete dissolution of 3 in THF. The reaction mixture was heated at 50 8C for 2 h, cooled to RT, and then left at RT overnight.…”

Efficient Fluorescein Spirolactam and Bis-Spirolactam Synthesis

“…We chose fluorescein methyl ester since it represented a known open form of fluorescein. 10,11 We were pleasantly surprised to find that fluorescein methyl ester reacted rapidly with a variety of primary amines in DMF at ambient temperature rather than at the higher temperatures usually required for simple esters. However, the intensely red-orange fluorescein methyl ester solution was converted into a nearly colorless solution.…”

Quantitative fluorescence assays using a self-powered paper-based microfluidic device and a camera-equipped cellular phone

“…We prepared reagent 1 by coupling 3 17 with 4 18 through an S N 2 reaction (Scheme 1). The basic conditions of this reaction also removed the acetate protecting groups from the galactose substrate.…”