Alternatives to titanium dioxide in tablet coating.

Titanium dioxide (TiO2) is one of the most commonly used pharmaceutical excipients. It is widely used as a white pigment in tablet and pellet coatings. However, it has recently been under massive criticism as a number of studies suggest a cancerogenic potential. It can therefore no longer be taken for granted that TiO2 will continue to be universally available for drug products. Finding suitable alternatives is hence of special relevance. In this study, a number of different pigments were coated on tablets and their covering potential analyzed. None of the alternative pigments showed comparable effectiveness and efficiency to TiO2, though the CaCO3/CaHPO4-based coating showed the second-best results. Regarding the ability to protect photosensitive active ingredients, ZnO showed a comparable potential as TiO2, while all other pigments failed. Using the alternative pigments as markers for in-line Raman spectroscopy as a process analytical technology was challenging and led to increased prediction errors. Again, the CaCO3/CaHPO4-based coating was the only of the tested alternatives with satisfying results, while all other pigments led to unacceptably high prediction errors.

Matrix-dependent absorption of 8-methoxypsoralen in extracorporeal photopheresis.

BACKGROUND: Extracorporeal photopheresis (ECP) is an effective immunomodulatory therapy for various diseases. Autologous leukocytes are collected, photoactivated with a photosensitizer (8-methoxypsoralen, 8-MOP) and UVA light, and subsequently reinfused back to the patient. Leukapheresis and UVA irradiation systems can be integrated into one device (inline) or handled by two separate devices (offline). ECP works via intercalation of 8-MOP into DNA helices and UVA-based interactions to inhibit DNA replication. 8-MOP is known to adhere to plastic materials, which might reduce its availability for intercalation. In the present study we examined the bioavailability of 8-MOP when different plastic materials and solvents are used as matrices. METHODS: Varying amounts of shredded ethylene vinyl acetate (EVA) and polyvinylchloride (PVC) from the MacoGenic irradiation bag (EVA1), UVA PIT irradiation bag (EVA2), UVA PIT recirculation bag (PVC A) and UVA PIT tubing (PVC B) by MacoPharma and PIT Medical Systems, respectively, were incubated with 200 ng mL-1 8-MOP dissolved in diisopropyl ether (DIPE) plus toluene 90/10 vol%, deionized water or plasma. After 2 h 8-MOP concentrations were determined by GC-MS. RESULTS: After incubation, 8-MOP concentrations varied depending on the amount and type of plastic (PVC > EVA) and solvent (water > plasma > DIPE/toluene). Absorption to 200 mg EVA1 or EVA2 resulted in 8-MOP concentrations of 57% or 32% in water, 91% or 80% in plasma, and 93% or 92% in DIPE/toluene, while 200 mg PVC A and PVC B yielded recovery rates of 26% and 10% in water, 76% and 75% in plasma, and 55% and 30% in DIPE/toluene, respectively. Remaining 8-MOP differed significantly between container materials (EVA vs. PVC; p < 0.022) but not manufacturers (MacoPharma vs. PIT Medical Systems). CONCLUSION: Absorption loss of 8-MOP depends on the type of plastic and solvent and is more pronounced with water than with plasma. As the DNA binding effect of 8-MOP is dose-dependent, ECP starting doses should be adjusted to ensure that a sufficient concentration of free bioavailable 8-MOP is present during UV irradiation.

Enhanced Ferroptosis by Oxygen-Boosted Phototherapy Based on a 2-in-1 Nanoplatform of Ferrous Hemoglobin for Tumor Synergistic Therapy.

Photodynamic therapy (PDT) combined with oxygenating strategies is widely employed in cancer treatment; however, oxygen-boosted PDT has failed to achieve an ideal effect due to the complexity, heterogeneity, and irreversible hypoxic environment generated by tumor tissues. With the emergence of Fe-dependent ferroptosis boasting reactive oxygen species (ROS) cytotoxicity as well, such a chemodynamic approach to cancer therapy has drawn extensive attention. In this study, hemoglobin (Hb) is connected with the photosensitizer chlorin e6 (Ce6) to construct a 2-in-1 nanoplatform (SRF@Hb-Ce6) with Sorafenib (SRF, ferroptosis promotor) loaded, combining oxygen-boosted PDT and potent ferroptosis. Benefiting from the intrinsic presence of Fe capable of binding oxygen, hemoglobin concurrently furnishes oxygen for oxygen-dependent PDT and Fe for Fe-dependent ferroptosis. Furthermore, amphiphilic MMP2-responsive peptide is incorporated into the skeleton of the nanoplatform to ensure drug-release specificity for safety improvement. Correlative measurements demonstrate the potentiation of PDT and ferroptosis with SRF@Hb-Ce6. More importantly, PDT strengthens ferroptosis by recruiting immune cells to secrete IFN-γ, which can sensitize the tumor to ferroptosis in our findings. The therapeutic effect of synergistic treatment with SRF@Hb-Ce6 in vitro and in vivo was proven significant, revealing the promising prospects of combined PDT and ferroptosis therapy with the 2-in-1 nanoplatform.

Expulsion of a potent cancer-cell photosensitizer from its micelle-bound state using ß-cyclodextrin: A tenable model for efficient drug release.

The present study delves into the interaction of a potent cancer-cell photosensitizer Norharmane (NHM) with non-ionic triblock copolymer P123, followed by the assessment of the stability of the formed complex in the presence of ß-cyclodextrin (ß-CD). Spectroscopic results unveil the modulation of the prototropic equilibrium of NHM within the constrained microheterogeneous medium of the copolymer micelle to be favoured towards the neutral species of NHM over the cationic counterpart; which has been aptly rationalized invoking the key role of hydrophobic interaction in the association process and is further reinforced from steady-state and time-resolved spectroscopic measurements. The micropolarity of the probe-binding site has been evaluated by the archetypal ET(30) analysis revealing that the cationic probe remains in the corona region of the micelle instead of penetrating deeper into the micellar core. Moreover, the effect of ß-CD on the stability of the NHM-bound P123 aggregates has also been investigated, revealing that ß-CD can be used as a potential host for the release of the micelle-encapsulated drug through an inclusion complex formation with the P123 monomers. The result is expected to be of potential interest from medical perspective owing to the context of efficient drug release at their potential sites.

Characterization of Protoporphyrin IX Species in Vitro Using Fluorescence Spectroscopy and Polar Plot Analysis.

Protoporphyrin IX (PPIX) is a photodynamic therapy (PDT) agent for the treatment of various types of cancer. The effectiveness of PDT is believed to be associated with aggregation of PPIX in cells. However, the aggregation equilibrium of PPIX in the cellular environment and in solution is still poorly understood. This is attributed by the lack of a method that allows for controllable generation of PPIX aggregates and robust analysis technique for measuring their photophysical properties. In this study, the dynamics of PPIX aggregation were investigated under high pressure and different solvent conditions using time-resolved fluorescence spectroscopy. The data were analyzed on a polar plot, a model-free analysis method that has become increasingly popular for fluorescence lifetime studies. We discovered that increasing hydrostatic pressure enhanced the formation of J-type aggregates based on measured absorbance, spectra, and lifetime features. Formation of large aggregates, which have a subnanosecond lifetime in the excited state, was observed under the increasing concentration of divalent cations as well as under a solvent of around neutral pH. PPIX monomerizes from the aggregate as pH becomes more basic, not dimerization as proposed by previous studies. Here, we demonstrate that the combination of time-resolved measurement and polar plot analysis is very robust for monitoring the presence of different types of PPIX aggregates formed in various chemical environments.

Comparative study of ink photoinitiators in food packages using gas chromatography with vacuum ultraviolet detection and gas chromatography with mass spectrometry.

This work focused on the development and validation of the analytical procedure using gas chromatography equipped with vacuum-ultraviolet detection for the specific and sensitive determination of nine photoinitiators in food packages. Subsequently, a comparison of the combination of vacuum ultraviolet spectroscopy with gas chromatography and a developed gas chromatography with mass spectrometry method was performed. The vacuum-ultraviolet spectra of all tested photoinitiators were collected and found to be highly distinct, even for isomers. Under the optimal conditions, the limits of detection for nine photoinitiators ranged from 1 to 5 mg/L using vacuum ultraviolet detection and from 0.15 to 0.5 mg/L using mass spectrometric detection. Both techniques were successfully applied for screening of photoinitiators in seven kinds of food packages and the obtained data showed good agreement (the relative difference was between 3 and 18%). The variability in concentrations found in triplicate samples was assessed to be below 18%. Predominantly benzophenone was found in all analysed samples in the range of 0.31-4.23 mg/kg. It appears to be preferably selected by food packaging manufacturers. This study proposes a new simple and sensitive technique used for analysis of photoinitiators that could be a good alternative to gas chromatography with mass spectrometry.

Fotossensibilização primária em bovinos leiteiros causada por Froelichia humboldtiana / Primary photosensitization in dairy cattle caused by Froelichia humboldtiana

O presente trabalho foi conduzido com o objetivo de relatar um surto de fotossensibilização causado por Froelichia humboldtiana em bovinos leiteiros no Estado do Rio Grande do Norte, Brasil. Foram examinados animais de uma propriedade rural que apresentavam sintomatologia compatível com fotodermatite. Procedeu-se a coleta de amostras de sangue periférico de cinco bovinos para análise das atividades das enzimas hepáticas gamaglutamiltransferase e aspartatoaminotransferase, além da concentração de bilirrubina total, direta e indireta. Das áreas de pele com lesões de dois animais foram realizadas biópsias. Constatou-se que 15 animais de um rebanho composto por 40 animais apresentaram fotossemsibilização. Os animais tinham histórico de apresentar lesões de fotodermatite aproximadamente 10 dias após pastarem em áreas invadidas por F. humboldtiana. Ao exame clínico dos bovinos leiteiros notou-se que inicialmente apresentavam prurido e hiperemia nas áreas de pele despigmentadas do dorso e úbere, também havia alterações do comportamento. Posteriormente, as áreas hiperêmicas se apresentavam com edema que evoluíam para dermatite ulcerativa, necrotizante e exudativa, com perda de extensas áreas da epiderme. As úlceras eram mais graves nos quatro bovinos que apresentavam automutilação por lambedura. Esses quatro animais foram retirados do pasto e abrigados em local sombreado. Uma semana após, o prurido regrediu e as fissuras da pele passaram a cicatrizar. Porém, as lesões reapareceram logo após os bovinos serem reintroduzidas no pasto infestado por F. humbolditiana. Percebeu-se ainda queda na produção leiteira (redução de 50-60%) das vacas após a instalação de fotodermatite. Porém, os bezerros que ainda eram lactantes e ingeriam o leite nas vacas acometidas por fotossensibilização, não apresentaram sinais de fotodermatite. A histopatologia de biópsias de pele revelou inflamação na derme superficial constituída por mastócitos, linfócitos, e alguns plasmócitos. Na epiderme haviam extensas úlceras, recobertas por crostas, associada a infiltrado neutrofílico. As atividades séricas de AST, GGT e as concentrações de bilirrubina estavam dentro dos valores de referência normais para a espécie bovina. O diagnóstico de fotossensibilização primária associada à ingestão de F. humboldtiana foi baseado na epidemiologia, sinais clínicos, bioquímica sérica, biópsia de pele e reocorrência das lesões após os animais serem reintroduzidos no pasto invadido pela planta. Conclui-se que a F. humboldtiana é uma importante causa de fotossensibilização primária em bovinos leiteiros no semiárido brasileiro e que sua toxina provavelmente não é excretada pelo leite bovino.(AU)

The present study was conducted with the objective to report an outbreak of photosensitization caused by Froelichia humboldtiana in dairy cattle in the State of Rio Grande do Norte, Brazil. Animals from a rural property with symptoms compatible with photodermatitis were examined. Peripheral blood samples from five cattle were collected for the analysis of the activities of hepatic enzymes gammaglutamyltransferase and aspartate aminotransferase, in addition were also analysed the concentration of total, direct and indirect bilirubin. From the areas of skin with lesions of two animals, biopsies were performed. It was verified that 15 animals from a herd composed by 40 animals presented photosensitization. The animals had a history of photodermatitis lesions approximately 10 days after grazing in areas invaded by F. humboldtiana. Clinical examination of dairy cattle showed that they initially had pruritus and hyperemia in the depigmented areas of the dorsum and udder, and there were also behavioral changes. Subsequently, the hyperemic areas presented edema that evolved to ulcerative, necrotizing and exudative dermatitis, with loss of extensive areas of the epidermis. The ulcers were more severe in four bovines that had self-mutilation by licking. These four animals were removed from the pasture and sheltered in a shady location. A week later, the pruritus regressed and the fissures of the skin began to heal. However, the lesions reappeared after the cattle were reintroduced in the grass infested by F. humbolditiana. There was also a decrease in milk production (reduction of 50-60%) of cows after the installation of photodermatitis. However, calves that were still lactating and ingested the milk in photosensitized cows, showed no signs of photodermatitis. Histopathology of skin biopsies revealed inflammation in the superficial dermis consisting of mast cells, lymphocytes, and some plasma cells. In the epidermis there were extensive ulcers, covered by crusts, associated with neutrophilic infiltrate. Serum activities of AST, GGT and bilirubin concentrations were within normal reference values for the bovine species. The diagnosis of primary photosensitization associated with F. humboldtiana ingestion was based on epidemiology, clinical signs, serum biochemistry, skin biopsy and lesion reoccurrence after the animals were reintroduced in the pasture invaded by the plant. It is concluded that F. humboldtiana is an important cause of primary photosensitization in dairy cattle in the Brazilian semi-arid region and that its toxin is probably not excreted by bovine milk.(AU)

Scanning Fiber Endoscope Improves Detection of 5-Aminolevulinic Acid-Induced Protoporphyrin IX Fluorescence at the Boundary of Infiltrative Glioma.

OBJECTIVE: Fluorescence-guided surgery with protoporphyrin IX (PpIX) as a photodiagnostic marker is gaining acceptance for resection of malignant gliomas. Current wide-field imaging technologies do not have sufficient sensitivity to detect low PpIX concentrations. We evaluated a scanning fiber endoscope (SFE) for detection of PpIX fluorescence in gliomas and compared it to an operating microscope (OPMI) equipped with a fluorescence module and to a benchtop confocal laser scanning microscope (CLSM). METHODS: 5-Aminolevulinic acid-induced PpIX fluorescence was assessed in GL261-Luc2 cells in vitro and in vivo after implantation in mouse brains, at an invading glioma growth stage, simulating residual tumor. Intraoperative fluorescence of high and low PpIX concentrations in normal brain and tumor regions with SFE, OPMI, CLSM, and histopathology were compared. RESULTS: SFE imaging of PpIX correlated to CLSM at the cellular level. PpIX accumulated in normal brain cells but significantly less than in glioma cells. SFE was more sensitive to accumulated PpIX in fluorescent brain areas than OPMI (P < 0.01) and dramatically increased imaging time (>6×) before tumor-to-background contrast was diminished because of photobleaching. CONCLUSIONS: SFE provides new endoscopic capabilities to view PpIX-fluorescing tumor regions at cellular resolution. SFE may allow accurate imaging of 5-aminolevulinic acid labeling of gliomas and other tumor types when current detection techniques have failed to provide reliable visualization. SFE was significantly more sensitive than OPMI to low PpIX concentrations, which is relevant to identifying the leading edge or metastasizing cells of malignant glioma or to treating low-grade gliomas. This new application has the potential to benefit surgical outcomes.

Smart activatable and traceable dual-prodrug for image-guided combination photodynamic and chemo-therapy.

Activatable photosensitizers (PSs) and chemo-prodrugs are highly desirable for anti-cancer therapy to reduce systemic toxicity. However, it is difficult to integrate both together into a molecular probe for combination therapy due to the complexity of introducing PS, singlet oxygen quencher, chemo-drug, chemo-drug inhibitor and active linker at the same time. To realize activatable PS and chemo-prodrug combination therapy, we develop a smart therapeutic platform in which the chemo-prodrug serves as the singlet oxygen quencher for the PS. Specifically, the photosensitizing activity and fluorescence of the PS (TPEPY-SH) are blocked by the chemo-prodrug (Mitomycin C, MMC) in the probe. Meanwhile, the cytotoxicity of MMC is also inhibited by the electron-withdrawing acyl at the nitrogen position next to the linker. Upon glutathione activation, TPEPY-S-MMC can simultaneously release active PS and MMC for combination therapy. The restored fluorescence of TPEPY-SH is also used to report the activation for both PS and MMC as well as to guide the photodynamic therapy.

Singlet Oxygen Detection Using Red Wine Extracts as Photosensitizers.

Moderate consumption of red wine provides beneficial effects to health. This is attributed to polyphenol compounds present in wine such as resveratrol, quercetin, gallic acid, rutin, and vanillic acid. The amount of these antioxidants is variable; nevertheless, the main beneficial effects of red wine are attributed to resveratrol. However, it has been found that resveratrol and quercetin are able to photosensitize singlet oxygen generation and conversely, gallic acid acts as quencher. Therefore, and since resveratrol and quercetin are some of the most important antioxidants reported in red wines, the aim of this research was to evaluate the photosensitizing ability of 12 red wine extracts through photo-oxidation of ergosterol. The presence of 1 O2 was detected by ergosterol conversion into peroxide of ergosterol through 1 H NMR analysis. Our results showed that 10 wine extracts were able to act as photosensitizers in the generation of singlet oxygen. The presence of 1 O2 can damage other compounds of red wine and cause possible organoleptic alterations. Finally, although the reaction conditions employed in this research do not resemble the inherent conditions in wine making processing or storing, or even during its consumption, this knowledge could be useful to prevent possible pro-oxidant effects and avoid detrimental effects in red wines.

A New Green Titania with Enhanced NIR Absorption for Mitochondria-Targeted Cancer Therapy.

A new kind of green titania (G-TiO2-x ) with obvious green color was facilely synthesized from black titania (B-TiO2-x ) through subsequently strong ultrasonication. Comparatively, this stable G-TiO2-x shows much enhanced near infrared (NIR) absorption, especially around 920 nm, which can be ascribed to the obvious change of TiO2-x lattice order owing to the effect of ultrasonication. This feature enables G-TiO2-x to be stimulated with 980 nm laser in the combined photodynamic therapy (PDT) and photothermal therapy (PTT), which is greatly beneficial for improving tissue penetration depth. Furthermore, since mitochondria are preferred subcellular organelles for PDT/PTT, G-TiO2-x was further designed to conjugate with triphenylphosphonium (TPP) ligand for mitochondria-targeted PDT/PTT to obtain precise cancer treatment. Attributing to the high mitochondria-targeting efficiency and simultaneously synergistic PDT/PTT, high phototherapeutic efficacy and safety with a much lower laser power density (980 nm, 0.72 W cm-2) and low materials dosage were achieved both in vitro and in vivo. In addition, negligible toxicity was found, indicating high biocompatibility. This novel G-TiO2-x could provide new strategies for future precise minimal/non-invasive tumor treatment.

Photosensitizer-mediated mitochondria-targeting nanosized drug carriers: Subcellular targeting, therapeutic, and imaging potentials.

Mitochondria-targeting drug carriers have considerable potential because of the presence of many molecular drug targets in the mitochondria and their pivotal roles in cellular viability, metabolism, maintenance, and death. To compare the mitochondria-targeting abilities of triphenylphosphonium (TPP) and pheophorbide a (PhA) in nanoparticles (NPs), this study prepared mitochondria-targeting NPs using mixtures of methoxy poly(ethylene glycol)-(SS-PhA)2 [mPEG-(SS-PhA)2 or PPA] and TPP-b-poly(ε-caprolactone)-b-TPP [TPP-b-PCL-b-TPP or TPCL], which were designated PPAn-TPCL4-n (0≤n≤4) NPs. With increasing TPCL content, the formed PPAn-TPCL4-n NPs decreased in size from 33nm to 18nm and increased in terms of positive zeta-potentials from -12mV to 33mV. Although the increased TPCL content caused some dark toxicity of the PPAn-TPCL4-n NPs due to the intrinsic positive character of TPCL, the NPs showed strong light-induced killing effects in tumor cells. In addition, the mitochondrial distribution of the PPAn-TPCL4-n NPs was analyzed and imaged by flow cytometry and confocal microscopy, respectively. Thus, the PhA-containing NPs specifically targeted the mitochondria, and light stimulation caused PhA-mediated therapeutic effects and imaging functions. Expanding the capabilities of these nanocarriers by incorporating other drugs should enable multiple potential applications (e.g., targeting, therapy, and imaging) for combination and synergistic treatments.

A method for the quantification of 8-methoxypsoralen by mass spectrometry for offline extracorporeal photopheresis.

BACKGROUND: Extracorporeal photopheresis (ECP) is an efficient method to treat various autoimmune diseases, cutaneous T-cell lymphoma, and graft-versus-host disease. It is based on the ex vivo inactivation of lymphocytes by 8-methoxypsoralen (8-MOP)/UV light treatment. Despite the adhesive, lipophilic nature of 8-MOP, no quality control is established for the ECP procedure. METHODS: We developed a sensitive high-performance liquid chromatography/tandem mass spectrometry (HPLC-MS/MS) assay to monitor residual 8-MOP concentration after UVA irradiation in the whole blood supernatant after acetonitrile precipitation. RESULTS: The preanalytical stability of 8-MOP exceeded 7 days, allowing batch mode analysis. Linearity was determined with R2 above 0.99. The 8-MOP concentrations decreased exponentially after UV exposure, with decay constants of 0.0259 in plasma and 0.0528 in saline. The recovery of 8-MOP in photopheresates was about 68%, indicating binding to DNA as well as to plastic structures. UVA induced no 8-MOP fragmentation, but caused self-adducts under extreme conditions (10-fold UV dosage). CONCLUSIONS: Detection of 8-MOP proved to be feasible and demonstrated that the doses were in the pharmaceutically active range.

Raman microspectroscopy of Hematoporphyrins. Imaging of the noncancerous and the cancerous human breast tissues with photosensitizers.

Raman microspectroscopy combined with fluorescence were used to study the distribution of Hematoporphyrin (Hp) in noncancerous and cancerous breast tissues. The results demonstrate the ability of Raman spectroscopy to distinguish between noncancerous and cancerous human breast tissue and to identify differences in the distribution and photodegradation of Hematoporphyrin, which is a photosensitizer in photodynamic therapy (PDT), photodynamic diagnosis (PDD) and photoimmunotherapy (PIT) of cancer. Presented results show that Hematoporphyrin level in the noncancerous breast tissue is lower compared to the cancerous one. We have proved also that the Raman intensity of lipids and proteins doesn't change dramatically after laser light irradiation, which indicates that the PDT treatment destroys preferably cancer cells, in which the photosensitizer is accumulated. The specific subcellular localization of photosensitizer for breast tissues samples soaked with Hematoporphyrin was not observed.

Photodynamic diagnosis of pleural malignant lesions with a combination of 5-aminolevulinic acid and intrinsic fluorescence observation systems.

BACKGROUND: We have developed a new diagnostic method using the photosensitizer 5-aminolevulinic acid (5ALA) for diagnosing intrathoracic malignant lesions. When ingested exogenously, 5ALA is metabolized to a heme precursor, protoporphyrin IX, which stays in malignant cells and emits red to pink luminescence of about 630 nm. METHODS: We enrolled 40 patients who underwent respiratory surgery and consented to participate in this study. Twenty-eight patients had primary lung cancer, 8 metastatic lung tumors, 2 malignant pleural tumors, and 2 benign tumors. Localization of malignant lesions was attempted by observing such lesions with an autofluorescence imaging system and by comparing the color tone of the autofluorescence between malignant lesions and normal tissues after oral administration of 5ALA. Malignant lesions on the pleural surface emitted pink autofluorescence in contrast to the green autofluorescence of the surrounding normal tissues. RESULTS: When 28 patients with primary lung cancer were examined according to the degree of pleural infiltration (pl), red fluorescence was confirmed in 10 of 10 patients (100%) with p11-p13 and 5 of 18 patients (27.7%) with p10. The latter 5 patients had been diagnosed with PL1 preoperatively or intraoperatively. CONCLUSION: This system achieved accurate localization of malignant lesions, suggesting that it may also be applicable to photodynamic therapy.

Nano-enabled SERS reporting photosensitizers.

To impart effective cellular damage via photodynamic therapy (PDT), it is vital to deliver the appropriate light dose and photosensitizer concentration, and to monitor the PDT dose delivered at the site of interest. In vivo monitoring of photosensitizers has in large part relied on their fluorescence emission. Palladium-containing photosensitizers have shown promising clinical results by demonstrating near full conversion of light to PDT activity at the cost of having undetectable fluorescence. We demonstrate that, through the coupling of plasmonic nanoparticles with palladium-photosensitizers, surface-enhanced Raman scattering (SERS) provides both reporting and monitoring capability to otherwise quiescent molecules. Nano-enabled SERS reporting of photosensitizers allows for the decoupling of the therapeutic and imaging mechanisms so that both phenomena can be optimized independently. Most importantly, the design enables the use of the same laser wavelength to stimulate both the PDT and imaging features, opening the potential for real-time dosimetry of photosensitizer concentration and PDT dose delivery by SERS monitoring.

Laser-induced autofluorescence spectroscopy: can it be of importance in detection of bladder lesions?

BACKGROUND: Non-muscle invasive bladder cancer can be missed during white light endoscopy in up to 50% of cases. We aimed to test whether or not we could find a difference between benign and cancerous tissue wavelengths using laser induced autofluorescence spectroscopy can increase cancer detection. MATERIALS AND METHODS: We analysed 67 tissue samples using spectral analysis. The WavSTAT (Spectra Science) optical biopsy device was used to record fluorescence spectra from biopsied tissue enabling calculation of an AUC for each spectrum, a measure of the mean spectral wavelength (λ¯ (nm)) and a dimensionless fluorescence ratio. Mann-Whitney test was used to compare the two groups. RESULTS: We found that 49.3% (33/67) of the tissue was benign, 44.8% (30/67) was CIS/cancerous tissue, and the remaining 4/67 samples were atypia (2) and dysplasia (2). The median AUC for the benign tissue was 19.53 (interquartile range [IQR]: 5.35-30.39) and that for CIS/cancerous tissue was 7.05 (IQR: 2.89-14.24) (P=0.002). The median wavelengths for the benign tissue and malignant tissue were 502.4nm (IQR: 500.3-504.3nm) and 505.2nm (IQR: 502.1-513.2nm), respectively (P=0.003). The median fluorescence ratio was 0.080 (IQR: 0.070-0.088) for benign tissue and 0.096 (IQR: 0.079-0.221) for CIS/cancerous tissue (P=0.002). CONCLUSIONS: We found statistical differences between the median AUC calculations and median wavelengths for the benign and cancerous tissue. We also found a statistical difference between the fluorescence ratios between the two tissue types. There seems to be a role for optical spectroscopy in verifying bladder lesions.

Photoreactive stapled peptides to identify and characterize BCL-2 family interaction sites by mass spectrometry.

Protein interactions dictate a myriad of cellular activities that maintain health or cause disease. Dissecting these binding partnerships, and especially their sites of interaction, fuels the discovery of signaling pathways, disease mechanisms, and next-generation therapeutics. We previously applied all-hydrocarbon peptide stapling to chemically restore α-helical shape to bioactive motifs that become unfolded when taken out of context from native signaling proteins. For example, we developed stabilized alpha-helices of BCL-2 domains (SAHBs) to dissect and target protein interactions of the BCL-2 family, a critical network that regulates the apoptotic pathway. SAHBs are α-helical surrogates that bind both stable and transient physiologic interactors and have effectively uncovered novel sites of BCL-2 family protein interaction. To leverage stapled peptides for proteomic discovery, we describe our conversion of SAHBs into photoreactive agents that irreversibly capture their protein targets and facilitate rapid identification of the peptide helix binding sites. We envision that the development of photoreactive stapled peptides will accelerate the discovery of novel and unanticipated protein interactions and how they impact health and disease.

Hypericum perforatum: Influences of the habitat on chemical composition, photo-induced cytotoxicity, and antiradical activity.

CONTEXT: Hypericin, isolated from Hypericum perforatum L. and about another 300 Hypericum species (Guttiferae), is one of the most powerful photosensitizers found in nature. OBJECTIVE: The aim of this study was to assess the variability of chemical composition and biological activities of four H. perforatum samples, collected at different altitudes in the South Apennine of Italy. MATERIALS AND METHODS: MTT assay was used to evaluate the antiproliferative activity of different samples concentrations (0.6-100 µg/mL) after irradiation at 365 nm. The inhibition of nitric oxide production was evaluated after 24 h of incubation using the macrophage cell line RAW 264.7 and sample solutions ranging from 12.5 to 1000 µg/mL. Antioxidant activities were evaluated using 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay and ß-carotene bleaching test (ranges were 12.5-1000 and 1-400 µg/mL, respectively). Chemical composition was evaluated through HPTLC, and different contents of hypericin and rutin have been observed. RESULTS: The most phototoxic sample was collected from Zumpano (no. 1 at 370 m), with IC50 values of 24.61 ± 0.02 µg/mL. Sample no. 1 showed also the best radical scavenging activity (IC50 value of 9.18 ± 0.03 µg/mL) and the best antioxidant activity (IC50 value of 10.04 ± 0.03 µg/mL after 30 min of incubation). Best activity of extract no. 1 was well in accordance with chemical data, including the phenolic total content and particular metabolome profile. DISCUSSION AND CONCLUSION: This paper confirms the usefulness in maintaining the exploration of H. perforatum activities, in order to confirm its potentiality as a multipurpose plant.

A high-throughput photodynamic therapy screening platform with on-chip control of multiple microenvironmental factors.

We present a novel high-throughput microfluidic platform that enables the evaluation of the anticancer efficacy of photodynamic therapy (PDT) drugs over multiple microenvironmental factors. PDT is uniquely complex, originating from its dependence on three separate but essential elements: drug (also called photosensitizer), oxygen, and light. Thus, obtaining a reliable evaluation of PDT efficacy is highly challenging, requiring considerable effort and time to evaluate all three interdependent parameters. In this paper, we report a high-throughput efficacy screening platform that we implemented by developing microfluidic components that individually control basic PDT elements (photosensitizer concentrations, oxygen levels, and light fluence) and then integrating them into a single triple-layer device. The integrated microfluidic chip consists of an array of small compartments, each corresponding to a specific combination of these three variables. This allows for more than 1000 different conditions being tested in parallel. Cancer cells are cultured within the device, exposed to different PDT conditions, and then monitored for their viability using live/dead fluorescence staining. The entire screening assay takes only 1 hour, and the collected PDT outcomes (cell viability) for combinatorial screening are analysed and reported as traditional dose-response curves or 3D bubble charts using custom software. As a proof of concept, methylene blue is adopted as a photosensitizer and its drug efficacy on C6 glioma cells has been successfully evaluated for a total of 324 PDT conditions using the fabricated chip. This platform can facilitate not only the development of new photosensitizers but also the optimization of current PDT protocols.