Maleimide-Functionalized Photochromic Spirodihydroindolizines

Two photochromic spirodihydroindolizine/betaine systems for tethering to peptides and proteins via a maleimide function have been prepared. The absorption spectra of the betaines are in the red region of the visible spectrum and in the near-IR spectral domain, which are suitable energies of light for future in vivo applications. The half-times of cyclization have been determined for both DHI/betaine systems. The findings are consistent with a thermal barrier of varying size between the <i>transoid</i> and <i>cisoid</i> conformers of the betaines

<i>In-vitro</i> tests of the cell based CHX drug delivery system on <i>E</i>. <i>coli</i> C600N.

<p>Here <i>E</i>. <i>coli</i> C600N was exposed to the following test groups: A is the CHX—<i>M</i>. <i>luteus</i>—neutrophil delivery system, B is the first control group with neutrophils loaded with unmodified <i>M</i>. <i>luteus</i>, C is the second control group with just unmodified neutrophils and D is the third control group with plain RPMI culture medium.</p

Comparison of the fluorescence of an unloaded neutrophil population with a neutrophil population loaded with rhodamine-modified <i>M</i>. <i>luteus</i>.

<p>Panel on the right shows neutrophil survival after 3 and 6 hours for ratios of 10, 20 and 100 bacteria/neutrophil determined by PI/annexin 5 apoptosis assay and flow cytometry.</p

Image A shows a mouse liver from the control group with lesions (marked by the arrows) caused by <i>Fusobacterium necrophorum</i>.

<p>Image B shows a liver from the CHX treatment group with no visible lesions.</p

Results of the <i>in-vivo</i> experiment.

<p>MP + CHX is the treatment group containing neutrophils loaded with CHX modified <i>M</i>. <i>luteus</i>. The PBS group is the control group containing pure RPMI medium while PMN and PMN + MP represent the neutrophil control groups without and with unmodified <i>M</i>. <i>luteus</i>.</p

<i>In-vitro</i> tests of the cell based CHX drug delivery system on <i>Fusobacterium necrophorum</i>.

<p>The bacterium was exposed to the following test groups: A is the CHX—<i>M</i>. <i>luteus</i>—neutrophil delivery system, B is the first control group with neutrophils loaded with unmodified <i>M</i>. <i>luteus</i>, C is the second control group with just unmodified neutrophils and D is the third control group with plain RPMI culture medium.</p

Zeta potential data (surface charge) for <i>M</i>. <i>luteus</i> loaded with CHX-hydrochloride over a range of concentrations from 0 μg/ml to 1000 μg/ml CHX.

<p>The data has been normalized (0%-100% equals an increase of the zeta potential from -32.86 ± 1.88 mV to -9.63 ± 2.22 mV) in order to fit a logarithmic function (%ζ = 33.48•log(CHX-concentration)). The panel on the right shows correlation of the measured zeta potential (x-axis) with the measured absorption of CHX in the lysate (y-axis). A linear trend with an R² value of 0.9045 is also shown.</p

Channel Blocking of MspA Revisited

Porin A from Mycobacterium smegmatis (MspA) is a highly stable, octameric channel protein, which acts as the main transporter of electrolytes across the cell membrane. MspA features a narrow, negatively charged constriction zone, allowing stable binding of various analytes thereby blocking the channel. Investigation of channel blocking of mycobacterial porins is of significance in developing alternate treatment methods for tuberculosis. The concept that ruthenium­(II)­quaterpyridinium complexes have the capability to act as efficient channel blockers for MspA and related porins, emerged after very high binding constants were measured by high-performance liquid chromatography and steady-state luminescence studies. Consequently, the interactions between the ruthenium­(II) complex RuC2 molecules and MspA, leading to RuC2@MspA assemblies, have been studied utilizing time-resolved absorption/emission, atomic force microscopy, dynamic light scattering, ζ potential measurements, and isothermal titration calorimetry. The results obtained provide evidence for the formation of clusters/large aggregates of RuC2 and MspA. The results are of interest with respect to utilizing prospective channel blockers in porins. The combination of results from conceptually different techniques shed some light onto the chemical nature of MspA–channel blocker interactions thus contributing to the development of a paradigm for channel blocking