We present a detailed investigation of Photofrin photobleaching and photoproduct accumulation. Fisher rats were sensitized with 10 mg kg(-1) Photofrin and irradiated 24 h later with 514 nm light at 5 or 100 mW cm(-2). Fluorescence spectra were collected from the skin throughout treatment, and sensitizer bleaching and fluorescent photoproduct formation were quantified using spectral analysis. Photofrin bleaching was slightly more rapid at the higher irradiance under these conditions. However, accumulation of photoproduct was significantly enhanced at lower irradiance. To interpret these unexpected findings, we developed a new mathematical model in which reactions between singlet oxygen (1O2) and the photosensitizer and reactions between the sensitizer triplet and biological targets are both allowed to contribute to bleaching. Predictions of this model were tested in experiments performed on EMT6 spheroids sensitized with concentrations of 2.5, 10 and 30 microg mL(-1) Photofrin and subjected to PDT. Photofrin bleaching and photoproduct formation in these spheroids were measured using confocal fluorescence spectroscopy. In qualitative agreement with the mixed-mechanism model predictions, at the highest drug concentration Photofrin bleaching was more efficient via 1O2 reactions, while at the lowest concentration triplet reactions were more efficient. At all concentrations, photoproduct accumulation was greater under conditions of abundant oxygen.

In this review we focus on the widely used Suzuki–Miyaura reaction and show that Pd/C is an efficient catalyst for carbon–carbon bond formation. The advantages and limitations of Pd/C are presented through selected examples. Mechanistic as well as practical aspects are also discussed. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2006)

Treatment of mucocutaneous and cutaneous Candida albicans infections with photosensitizing agents and light, termed photodynamic therapy (PDT), offers an alternative to conventional treatments. Initial studies using the clinically approved photosensitizer Photofrin demonstrated the susceptibility of C. albicans to its photodynamic effects. In the present study, we have further refined parameters for Photofrin-mediated photodynamic action against C. albicans and examined whether mechanisms commonly used by microorganisms to subvert either antimicrobial oxidative defenses or antimicrobial therapy, including biofilm formation, were operative. In buffer and defined medium, germ tubes preloaded with Photofrin retained their photosensitivity for up to 2 hours, indicating the absence of degradation or export of Photofrin by the organism. The addition of serum resulted in a gradual loss of photosensitivity over 2 hours. In contrast to an adaptive response by germ tubes to oxidative stress by hydrogen peroxide, there was no adaptive response to singlet oxygen-mediated stress by photodynamic action. C. albicans biofilms were sensitive to Photofrin-mediated phototoxicity in a dose-dependent manner. Finally, the metabolic activity of C. albicans biofilms following photodynamic insult was significantly lower than that of biofilms treated with amphotericin B for the same time period. These results demonstrate that several of the mechanisms microorganisms use to subvert either antimicrobial oxidative defenses or antimicrobial therapy are apparently not operative during Photofrin-mediated photodynamic treatment of C. albicans. These observations provide support and rationale for the continued investigation of PDT as an adjunctive, or possibly alternative, mode of therapy against cutaneous and mucocutaneous candidiasis.

No abstract

Substitution of an alanine for leucine (shown in light blue) in the hydrophobic interior of designed three-stranded coiled coils allows for the control of metal ion coordination number and geometry. The influence of this perturbation by a noncoordinating residue can be monitored by the dramatic impact on the 113Cd NMR spectrum. The structural effect occurs even when the residue substitution is as much as 7 A from the metal binding site.

We report the influence of fluence rate on the photobleaching and cell survival in Colo 26 multicell spheroids photosensitized by meta-tetra-(hydroxyphenyl)chlorin (mTHPC). Photosensitizer degradation and therapeutic efficacy increased dramatically and progressively when the fluence rate was reduced over the range from 90 to 5 mW cm-2. These experimental results were compared to a mathematical model of photobleaching based on self-sensitized singlet oxygen reactions with the photosensitizer ground state. This model incorporates photophysical parameters obtained from microelectrode measurements of oxygen depletion at the surface of mTHPC-sensitized spheroids and was refined by including the inhomogeneous distribution of mTHPC in spheroids and oxygen depletion in the bulk medium. Since the model is consistent with the experimental data we conclude that the fluence rate dependence of the cell survival and of mTHPC photobleaching is due to photochemical oxygen consumption and a predominantly singlet oxygen-mediated mechanism of mTHPC photobleaching. The threshold dose of reacting singlet oxygen was calculated to be 7.9 +/- 2.2 mM in this system.

Phosphorescence quenching of certain metalloporphyrins is used to measure tissue and microvascular pO(2). Oxygen quenching of metalloporphyrin triplet states creates singlet oxygen, which is highly reactive in biological systems, and these oxygen-consuming reactions are capable of perturbing tissue oxygenation. Kinetics of photochemical oxygen consumption were measured for a Pd-porphyrin in two model systems in vitro over a range of irradiances (1.34-134 mW cm(-2)). For a given irradiance, and, after correction for differing porphyrin concentrations, rates of oxygen consumption were similar when the Pd-porphyrin was bound to bovine serum albumin and when it was taken up by tumor cells in spheroids. At irradiances comparable to those used in imaging superficial anatomy, rates of oxygen consumption were sufficiently low (2.5 microM s(-1)) that tissue oxygenation would be reduced by a maximum of 6%. An irradiance of 20 mW cm(-2), however, initiated a rate of oxygen consumption capable of reducing tissue pO(2) by at least 20-40%. These measured rates of consumption impose limitations on the use of phosphorescence quenching in thick tissues. The irreversible photobleaching of the Pd-porphyrin was also measured indirectly. The bleaching branching ratio, 23 M(-1), is significantly lower than that of porphyrin photodynamic agents.

Recently, we published an improved mathematical model of photodynamic therapy ͑PDT͒ dose deposition on length scales corresponding to intercapillary distances.1 This model describes the spatial and temporal dynamics of oxygen ͑ 3 O 2 ͒ consumption and transport and microscopic singlet oxygen ͑ 1 O 2 ͒ dose deposition during PDT treatment. It also enables simulation of volume-averaged quantities like hemoglobin oxygen saturation ͑SO 2 ͒ and photosensitizer fluorescence photobleaching, which are accessible experimentally. In a subsequent modeling study of the kinetics of the recovery of SO 2 following the interruption of PDT irradiation, we were troubled by what appeared to be physically unreasonable simulation results. The origin of these erroneous results was a sign error in one of the terms buried deep in the original code, which had the effect of creating a tumor microenvironment that was more hypoxic that we had specified as an initial condition. We emphasize that in our previously published article on this model, there were no errors in what was described, but the plots presented in that article were quantitatively affected by this sign error. Specifically, this error was in the numerator of Eq. ͑2͒ of Wang et al., 1 which was correct in the text; the error was only in the code. In this erratum, we show the revised figures generated by using the same photophysical and physiological parameters described previously.1 We also use this occasion to describe a few relatively minor improvements to the model.The oxygen transport equation in the capillary is revised slightly from the original and is now written ͑1͒whereHere, we include a new term, ͑1+S͒, on the left-hand side of the transport equation for the capillary ͓Eq. ͑1͔͒, which more correctly provides for the dynamic unloading of 3 O 2 from hemoglobin.2 Further, the revised code uses MichaelisMenten kinetics to describe the rate of metabolic 3 O 2 consumption not only in the calculation of the time-dependent state, as was done previously, but also in the calculation of the steady-state with axial diffusion. The equation for the tissue region remains as previously describedThe definitions and the values of the parameters were previously described.