Heteroarylcarbene-Arylnitrene Radical Cation Isomerizations.

5-Phenyltetrazole 1e is an important source of phenylnitrene or the phenylnitrene radical cation ( m/ z 91) under thermal, photochemical, and electron impact conditions. Similarly, 3- or 4-(5-tetrazolyl)pyridines 12b,c yield pyridylnitrene radical cations 9a•+ ( m/ z 92) upon electron impact. In contrast, 2-(5-tetrazolyl)pyridine 12a•+ generates 2-pyridyldiazomethane 24•+ and 2-pyridylcarbene 26•+ radical cations ( m/ z 119 and 91) upon electron impact. The 2-pyridylcarbene radical cation undergoes a carbene-nitrene rearrangement to yield the phenylnitrene radical cation. Calculations at the B3LYP/6-311G(d,p) level have revealed facile H-transfer from the tetrazole to the pyridine ring in 2-(5-tetrazolyl)pyridine, 12a•+ → 21•+, taking place in the radical cations. Subsequent losses of N2 generate the pyridinium diazomethyl radical 22•+ or pyridinium-2-carbyne 23•+. These two ions can isomerize to 2-pyridyldiazomethane 24•+ and 2-pyridylcarbene 26•+, the latter rearranging to the phenylnitrene radical cations 9a•+. 13C-labeling of the tetrazole rings confirmed that 2-(5-tetrazolyl)pyridine 12a generates 2-pyridylcarbene/phenylnitrene radical cations retaining the 13C label, but 4-(5-tetrazolyl)pyridine 12c generates 4-pyridylnitrene 18c•+, which has lost the 13C label. 2-Pyridylcarbene/phenylnitrene radical cations ( m/ z 91) also constitute the base peak in the mass spectrum of 1,2,3-triazolo[1,5- a]pyridine 34. Similarly, 4-pyridylnitrene radical cation 18c•+ or its isomers ( m/ z 92) is obtained from 1,2,3-triazolo[1,5- a]pyrazine 36. Several other α-heteroaryltetrazoles behave in the same way as 2-(5-tetrazolyl)pyridine, yielding heteroarylcarbene/arylnitrene radical cations in the mass spectrometer, and this was confirmed by 13C-labeling in the case of 1-(5-tetrazolyl)isoquinoline 42-13C. In general, 5-aryltetrazoles generate arylnitrene radical cations under electron impact, but α-heteroaryltetrazoles generate α-heteroarylcarbene radical cations.

Phenylnitrene Radical Cation and Its Isomers from Tetrazoles, Nitrile Imines, Indazole, and Benzimidazole.

Phenylnitrene radical cations m/ z 91, C6H5N, 8a•+ are observed in the mass spectra of 1-, 2-, and 5-phenyltetrazoles, even though no C-N bond is present in 5-phenyltetrazole. Calculations at the B3LYP/6-311G(d,p) level of theory indicate that initial formation of the C-phenylimidoylnitrene 13•+ and/or benzonitrile imine radical cation 19•+ from 1 H- and 2 H-5-phenyltetrazoles 11 and 12 is followed by isomerizations of 13•+ to the phenylcyanamide ion 15•+ over a low barrier. A cyclization of imidoylnitrene ion 13•+ onto the benzene ring offers alternate, very facile routes to the phenylnitrene ion 8a•+ and the phenylcarbodiimide ion 14•+ via the azabicyclooctadienimine 16•+. Eliminations of HNC or HCN from 14•+ and 15•+ again yield the phenylnitrene radical cation 8a•+. A direct 1,3-H shift isomerizing phenylcarbodiimide ion 14•+ to the phenylcyanamide ion 15•+ requires a very high activation energy of 114 kcal/mol, and this reaction needs not be involved. The benzonitrile imine -3-phenyl-1 H-diazirine-phenylimidoylnitrene-phenylcarbodiimide/phenylcyanamide rearrangement has parallels in thermal and photochemical processes, but the facile cyclization of imidoylnitrene 13•+ to azabicyclooctadienimine 16•+ is facilitated by the positive charge making the nitrene more electrophilic. Furthermore, the benzonitrile imine radical cation 19•+ can cyclize to indazole 24•+, and a series of intramolecular rearrangements via hydrogen shifts, ring-openings and ring closures allow the interconversion of numerous ions of composition C7H6N2•+, including 19•+, 24•+, the benzimidazole ion 38•+ and o-aminobenzonitrile ion 40•+, all of which can eliminate either HCN or HNC to yield the C6H5N•+ ions of phenylnitrene, 8a•+, and/or iminocyclohexadienylidene, 34•+. Moreover, benzonitrile imine 19•+ can behave like a benzylic carbenium ion, undergoing a novel ring expansion to cycloheptatetraenyldiazene 45•+. The N-phenylnitrile imine ion 2d•+ derived from 2-phenyltetrazole 1d cleaves efficiently to the phenylnitrene ion 8a•+ but may also cyclize to the indazole ion 24•+. The N-phenylimidoylnitrene 59•+ derived from 1-phenyltetrazole 5d undergoes facile isomerization to the phenylcyanamide ion 15•+ and hence phenylnitrene radical cation 8a•+.

RAFT polymer cross-coupling with boronic acids.

The ability to modify the thiocarbonylthio end-groups of RAFT polymers is important for applications where an inert or highly functionalised material is required. Here we report a copper promoted cross-coupling reaction between RAFT polymer end-groups and aryl boronic acids. This method gives high conversion to the modified polymers, and is compatible with a wide variety of functional molecules.

Biocompatibility and modification of the protein-based adhesive secreted by the Australian frog Notaden bennetti.

When provoked, Notaden bennetti frogs secrete a proteinaceous exudate, which rapidly forms a tacky and elastic glue. This material has potential in biomedical applications. Cultured cells attached and proliferated well on glue-coated tissue culture polystyrene, but migrated somewhat slower than on uncoated surfaces. In organ culture, dissolved glue successfully adhered collagen-coated perfluoropolyether lenses to debrided bovine corneas and supported epithelial regrowth. Small pellets of glue implanted subcutaneously into mice were resorbed by surrounding tissues, and all of the animals made a full recovery. An initial but transient skin necrosis at the implant site was probably caused by some of the potentially toxic metabolites present in the frog secretion; these include sterols and carotenoids, as well as fatty alcohols, aldehydes, ketones, acids, and aromatic compounds. Removal of the carotenoid pigments did not significantly alter the glue's material properties. In contrast, peroxidase treatment of dissolved glue introduced unnatural crosslinks between molecules of the major protein (Nb-1R) and resulted in the formation of a soft hydrogel, which was very different to the original material.

Purification and characterization of recombinant ligand-binding domains from the ecdysone receptors of four pest insects.

Cloned EcR and USP cDNAs encoding the ecdysone receptors of four insect pests (Lucilia cuprina, Myzus persicae, Bemisia tabaci, Helicoverpa armigera) were manipulated to allow the co-expression of their ligand binding domains (LBDs) in insect cells using a baculovirus vector. Recombinant DE/F segment pairs (and additionally, for H. armigera, an E/F segment pair) from the EcR and USP proteins associated spontaneously with high affinity to form heterodimers that avidly bound an ecdysteroid ligand. This shows that neither ligand nor D-regions are essential for the formation of tightly associated and functional LBD heterodimers. Expression levels ranged up to 16.6mg of functional apo-LBD (i.e., unliganded LBD) heterodimer per liter of recombinant insect cell culture. Each recombinant heterodimer was affinity-purified via an oligo-histidine tag at the N-terminus of the EcR subunit, and could be purified further by ion exchange and/or gel filtration chromatography. The apo-LBD heterodimers appeared to be more easily inactivated than their ligand-containing counterparts: after purification, populations of the former were <40% active, whereas for the latter >70% could be obtained as the ligand-LBD heterodimer complex. Interestingly, we found that the amount of ligand bound by recombinant LBD heterodimer preparations could be enhanced by the non-denaturing detergent CHAPS (3-[(3-cholamidopropyl)dimethyl-ammonio]-1-propanesulfonate). Purity, integrity, size and charge data are reported for the recombinant proteins under native and denaturing conditions. Certain intra- and intermolecular disulfide bonds were observed to form in the absence of reducing agents, and thiol-specific alkylation was shown to suppress this phenomenon but to introduce microheterogeneity.