Abstract
Trifluoromethyl phenyl diazirine (TPD) molecules are relatively stable carbene precursors, that readily form carbon covalent bonds with proteins. The stability of the diazoalkane intermediates is unknown, as are the factors which control carbene/diazoalkane ratios. This leads to incomplete carbene insertion onto desired compounds. Herein, stability and decay kinetics of diazoalkanes are evaluated from TPDs with various electron drawing groups, including 3-Phenyl-3-(trifluoromethyl)-3H-diazirine (TPD-H), p-benzyl alcohol (TPD-CH2OH), p-4-benzoic acid (TPD-COOH), and p-benzyl bromide (TPD-CH2Br). The spectroscopic analysis before and after UVA activation is performed both in dilute chloroform and neat by 19F NMR and ATR-FTIR, respectively. The increase of diazoalkane concentration after UVA exposure was in the order of: TPD-H > TPD-CH2Br > TPD-CH2OH > TPD-COOH. Indirect carbene/diazoalkane ratios ranged from 6: 1 to 3:1. Diazoalkane was relatively stable over the evaluation period of 30 min (post-UVA activation) in all compounds except TPD-CH2Br, which exhibits an 11 min half-life.
Highlights
Photoaffinity labelling and tissue adhesives exploit diazirine's ability of non-specific covalent insertion mediated photoactivation [1,2,3]
1) is compared to Trifluoromethyl phenyl diazirine (TPD)-H, as control. 19F NMR results of TPDs (1–4; referred to as dilute further in text) in Fig. 1A–D indicate a singlet diazirine peak at −65 ppm and no presence of diazoalkane intermediate that is normally positioned at −56 to −59 ppm [30] (1H NMR spectra and peak assignment with exact values of chemical shifts are presented in Supplementary information; Fig. S1)
Spectrum of dilute 1 shows the ~5% of azine adduct and TPD 2 shows a doublet peak at −76.4 ppm assigned to ether adduct as a results of carbene insertion into –OH group of 2 [30]
Summary
Photoaffinity labelling and tissue adhesives exploit diazirine's ability of non-specific covalent insertion mediated photoactivation [1,2,3]. Exploitation of carbene insertion reactions has evolved new types of voltage- and UVAactivated adhesive formulations [2,7,8,9,10] Both voltage- and UVAgenerated carbenes covalently insert into tissue proteins providing new strategies for adhering to wet tissue substrates. For these reasons, the quantitative determination of diazoalkane decay kinetics is necessary for accurate photoaffinity labelling as well as effective tissue proximation by interfacial carbene insertion
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