Subcellular Localization of Sulfonated Tetraphenyl Porphines in Colon Carcinoma Cells by Spectrally Resolved Imaging

Subcellular localization of the dye, 5,10,15,20-tetra(4-sul-fonatophenyl)porphine (TPPS₄) and the more hydrophobic dye, 5,10,15,20-tetra(1-sulfonatophenyl)porphine (TPPS₁), in murine colon carcinoma cells was studied by spectrally resolved imaging (SRI) combined with image processing techniques. Spectrally resolved imaging enabled the acquisition of multipixel fluorescence spectra (>10⁴) from a single cell. Demarcation of specific localization sites and segregation of the irrelevant fluorescence were based on the pixel spectra and by operating the functions of spectral similarity mapping (SSM), principal component analysis (PCA) and spectral classification. The SRI revealed the fine details of the photochemical process that clarify some aspects of subcellular damage. The SRI depicted the differences between TPPS₄ and TPPS, with respect to their initial localization and their fate at the end of the photochemical effect. The dye TPPS₄ was localized initially in lysosomal vesicles, and upon irradiation fluorescence was seen in the nucleus as well as in vesicles. Some of the vesicles were closely related to the nucleus, as resolved by SSM, PCA and spectral classification. Additional light exposure stimulated relocalization of TPPS₄ into the nucleus as well as into the nucleolus, which was clearly depicted by SSM and PCA. Spectral classification showed a third, weak residual cytoplasmic array around the nucleus. The dye TPPS, concentrated in a Golgi-like complex and was resolved in the nuclear envelope and in small vesicles: it was not redistributed into other compartments upon photosensitization. Serum supplementation to the incubation media of colon carcinoma cells treated with TPPS₄ or TPPS, did not change the localization patterns. Pixel spectra of the two dyes in the cells showed spectral shifts and expanded shoulders due to microenvironmental effects. Thus, the chemical nature of the sulfonated phenyl porphines, and not their interaction with serum proteins, was the main determinant of their binding to the lysosomes, nucleus, nucleolus, nuclear envelope or Golgi.     

Vascular endothelial cells as targets for photochemical internalization (PCI)

Cancer treatment can be exerted by targeting both cancer cells and the vasculature supplying solid tumors. Photochemical internalization (PCI) is a modality for cytosolic drug delivery, but recent data on contrast‐enhanced MRI have indicated that the method also reduces blood perfusion in HT1080 fibrosarcoma xenografts. The present report aims to investigate if PCI may exert direct cytotoxic effects on endothelial cells. PCI of saporin was performed on endothelial human umbilical vein endothelial cell (HUVEC) and fibrosarcoma cells (HT1080) using two PCI‐relevant photosensitizers, TPPS_2a and AlPcS_2a. A 22‐ and 13‐fold higher photosensitizer uptake was detected in the endothelial cells compared with the HT1080 cells for AlPcS_2a and TPPS_2a, respectively. PCI of saporin was, however, found more effective in HT1080 cells. For HT1080 cells, PCI with saporin increased cell killing 1.9‐fold over photodynamic therapy alone, but under the same conditions, only increased HUVEC cell killing by 1.6‐ and 1.3‐fold with AlPcS_2a and TPPS_2a, respectively. Saporin uptake was higher in HUVECs than in the HT1080 cells, hence did not reflect the cell line differences in PCI efficacy. This is the first report on PCI‐mediated kill of endothelial cells and lays the foundation for further preclinical evaluation of the PCI technology as an antivascular strategy to ablate tumors.     

A Study of the Uptake of Toluidine Blue O by Porphyromonas gingivalis and the Mechanism of Lethal Photosensitization

The purpose of the study was to determine the distribution of the photosensitizer toluidine blue O (TBO) within Porphyromonas gingivalis and the possible mechanism(s) involved in the lethal photosensitization of this organism. The distribution of TBO was determined by incubating P. gingivalis with tritiated TBO (³H-TBO) and fractionating the cells into outer membrane (OM), plasma membrane (PM), cytoplasmic proteins, other cytoplasmic constituents and DNA. The percentage of TBO in each of the fractions was found to be, 86.7, 5.4, 1.9, 5.7 and 0.3%, respectively. The involvement of cytotoxic species in the lethal photosensitization induced by light from a helium-neon (HeNe) laser and TBO was investigated by using deuterium oxide (D₂O), which prolongs the lifetime of singlet oxygen, and the free radical and singlet oxygen scavenger L-tryptophan. There were 9.0 log₁₀ and 2 log₁₀ reductions in the presence of D₂O and H₂O (saline solutions), respectively, at a light dose of 0.44 J (energy density = 0.22 J/cm²), suggesting the involvement of singlet oxygen. Decreased kills were attained in the presence of increasing concentrations of L-tryptophan. The effect of lethal photosensitization on whole cell proteins was determined by measuring tryptophan fluorescence, which decreased by 30% using 4.3 J (energy density = 4.3 J/ cm²) of light. Effects on the OM and PM proteins were determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. There was evidence of change in the molecular masses of several PM proteins and OM proteins compared to controls. There was evidence of damage to the DNA obtained from irradiated cells. Scanning electron microscopic studies showed that there was coaggre-gation of P. gingivalis cells when sensitized and then exposed to laser light. These results suggest that lethal photosensitization of P. gingivalis may involve changes in OM and/or PM proteins and DNA damage mediated by singlet oxygen.     

Photobiological effects of metal ions. Photosensitizing action of uranyl acetate on Escherichia coli K-12

Abstract— The Uranyl acetate sensitized killing of Escherichia coli K-12 by a light source approximating sunlight in both intensity and wavelength distribution is demonstrated to occur at a concentration of 5.0 × 10O^-4M uranyl acetate (pH 7.0). The photosensitized killing was evident after 150 min and almost complete within 320 min. Auxotrophic mutants were isolated from cultures incubated in both light and dark for 160 min at this concentration of uranyl acetate. Binding of UO₂²⁺ to E. coli is shown to occur with 82% of the UO₂²⁺ ions in a 5 × 10^-4M solution (pH 7.0) being bound to the cell wall. In the dark as well as at other pH values the extent of binding was much less. Most of the binding occurred in a time less than 30 min. The observation of rapid binding but delayed photosensitization is attributed to the necessity for penetration of uranyl ions into the cells' interior to effect photosensitization.     

MODE OF ACTION OF α-TERTHIENYL ON ESCHERICHIA COLI: EVIDENCE FOR A PHOTODYNAMIC EFFECT ON MEMBRANES

The photodynamic effects of α-terthienyl (αT) in near-UV light (UV-A) on Escherichia coli showed close agreement with the light absorption of αT at different wavelengths suggesting that αT is the primary absorbing molecule responsible for the photosensitized reaction. Studies with DNA repair deficient mutants of E. coli indicated that the bactericidal action of αT/UV-A was not mediated by DNA damage, in direct contrast to the well-known photosensitizer, 8-methoxypsoralen. By using a closed borosilicate glass reaction vessel and various gas mixtures, it was demonstrated that photosensitization of both E. coli and a more resistant bacterium, Pseudomonas aeruginosa , was absolutely dependent on the presence of oxygen. The rate of killing by αT/UV-A showed a rather small dependence on preincubation temperatures, with quite rapid killing at 5°C, suggesting that the movement of αT across the cytoplasmic membrane of E. coli is not the rate limiting step in killing and perhaps is not even necessary for killing. Sodium dodecyl sulphate-polyacrylamide gels of cell membrane proteins after 15 and 30min of treatment with αT/UV-A showed substantial random crosslinking of these proteins. The results taken overall suggest that αT is a photodynamic photosensitizer which exerts its primary effect at the level of the cytoplasmic membrane.     

Highly efficient and photostable photosensitizer based on BODIPY chromophore

Photosensitizers are reagents that produce reactive oxygen species upon light illumination and are commonly used to study oxidative stress or for photodynamic therapy. There are many available photosensitizers, but most have limitations, such as low photostability, structural instability, or a limited usable range of solvent conditions. Here, we describe a novel photosensitizer scaffold (2I-BDP) based on the unique characteristics of the BODIPY chromophore (i.e., high extinction coefficient, high photostability, and insensitivity to solvent environment). 2I-BDP shows stronger near-infrared singlet oxygen luminescence emission and higher photostability than the well-known photosensitizer, Rose Bengal. Unlike other photosensitizers, this scaffold is widely applicable under various conditions, including lipophilic and aqueous environments. HeLa cells loaded with 2I-BDP could be photosensitized by light illumination, demonstrating that 2I-BDP is potentially useful as a reagent for cell photosensitization, oxidative stress studies, or PDT.     

Calcium as a modulator of photosensitized killing of H9c2 cardiac cells.

Illumination of H9c2 rat heart cells in the presence of Rose Bengal resulted in dose-dependent cell killing (assessed by trypan blue staining) and modification of ionic currents flowing through the heart cell membrane. Inhibitors of voltage-gated ionic currents were shown to have little effect on cell killing. Ionic current measurements were used to assess the increase in leak conductance of these cells, which has been suggested to be a causal factor in killing of other cell types (1). Inhibitors of voltage-gated ionic currents, including the sodium channel blocker tetrodotoxin (100 microM) and the calcium channel blocker lanthanum (10 microM) were shown to have little effect on cell killing. The potassium channel inhibitor tetraethylammonium (20 mM) inhibited cell killing, but the effect is viewed as being caused by an inhibition of leak current. The time course of block of voltage-activated ionic currents during illumination, in the presence of Rose Bengal, was rapid compared with that for induction of leak current and for cell killing. These observations are consistent with a role for leak current in photosensitized killing of cardiac cells. They are interpreted with respect to calcium influx through the leak current pathway as a trigger for the cellular response.     

PHOTOSENSITIZED INACTIVATION OF CHINESE HAMSTER CELLS BY PHTHALOCYANINES

Chloroaluminum phthalocyanine was found to sensitize cultured Chinese hamster cells upon exposure to white fluorescent light. Elimination of wavelengths below 370 nm did not reduce the effect significantly, indicating that the effective wavelengths were those absorbed by the Q band (600–700 nm) of phthalocyanine. The magnitude of the photosensitizing effect increased with the dye concentration and the time of its contact with the cells prior to light exposure. Although photosensitization was drastically reduced in the absence of oxygen, the lack of effect of glycerol and D₂0 during exposure suggests that neither hydroxyl radicals nor ¹O₂ are responsible for the cytotoxic response. The efficiency of the photosensitized induced cell killing did not vary with the position of the cells in the cell cycle, in contrast to exposure to X-rays. The improved spectral properties, the reported low toxicity and the selective retention by neoplasms, make phthalocyanines promising candidates for use in photodynamic therapy of cancer.     

The Effect of Photodynamic Action on Leakage of Ions Through Liposomal Membranes that Contain Oxidatively Modified Lipids

Singlet oxygen, created in photosensitization, peroxidizes unsaturated fatty acids of the membrane's lipids. This generates alcoholic or aldehyde groups at double bonds' breakage points. In a previous study, we examined the leakage of a K⁺‐induced cross‐membrane electric potential of liposomes that undergo photosensitization. The question remains to what extent peroxidized lipids can compromise the stability of the membrane. In this study, we studied the effect of the oxidatively modified lipids PGPC and ALDOPC in the membrane on its stability, by monitoring the membrane electric potential with the potentiometric dye DiSC₂(5). As the content of the modified lipids increases the membrane becomes less stable, and even at just 2% of the modified lipids the membrane's integrity is affected, in respect to the leakage of ions through it. When the liposomes that contain the modified lipids undergo photosensitization by hematoporphyrin, the lipid bilayer becomes even more unstable and passage of ions is accelerated. We conclude that the existence of lipids with a shortened fatty acid that is terminated by a carboxylic acid or an aldehyde and more so when photosensitized damage occurs to unsaturated fatty acids in lecithin, add up to a critical alteration of the membrane, which becomes leaky to ions.     

Photodynamic inactivation of isolated crayfish mechanoreceptor neuron: different death modes under different photosensitizer concentrations

To study the mechanism of photodynamic nerve cell killing, isolated crayfish mechanoreceptor neurons were photosensitized by the sulfonated aluminum ophthalocyanine Photosens. Neuron activity was continuously recorded until irreversible abolition. Intense (10(-5) M Photosens) or weak (10(-7) M Photosens) photosensitization induced different bioelectric neuron responses: firing activation followed by irreversible depolarization block or gradual inhibition until firing abolition, respectively. These bioelectric responses were accompanied by different biochemical and morphological changes. In the case of intense photosensitization, neuron nuclei swelled and then shrank. Succinate dehydrogenase (SDH) was inhibited, and the plasma membrane was compromised just after firing cessation. Weak photosensitization did not induce these changes but caused swelling of the endoplasmic reticulum and destruction of the matrix, cristae and membranes in some of the mitochondria. Other mitochondria, however, retained the normal structure. Plasma membrane damage, SDH inhibition, nucleus shrinkage and impairment of the nuclear border occurred after 2-4 h. It is concluded that intense photosensitization induced necrotic processes during irradiation, whereas weaker impact caused delayed necrosis 2-4 h later. The observed electrophysiological neuron responses to photodynamic therapy may be considered as early hallmarks of different modes of forthcoming cell death.     

Mechanistic aspects of photoinactivation of Candida albicans by exogenous porphyrins

The mechanism of photoinactivation of Candida albicans by 3.5 μM uncharged, cationic or anionic porphyrins under blue light (407-420 nm) was found to be dependent on the uptake of porphyrins into yeast cells, and was also dependent on the presence or absence of proteins in the photosensitization medium. In a very protein-rich medium, a decrease in viability was observed only with the uncharged porphyrin. Photoinactivation by uncharged or cationic porphyrins in a protein-poorer medium resulted in total eradication, whereas no significant decrease was observed with the anionic porphyrin. Phototreatment in PBS resulted in eradication with all three porphyrins. X-ray microanalysis after phototreatment by the uncharged or cationic porphyrins in the protein-poor medium exhibited ion loss, indicating cell-membrane damage. Transmission electron microscopy indicated cellular and chromosomal damage. No ion loss or cell damage was observed in this medium with the anionic porphyrin. The efficiency of photoeradication of C. albicans is dependent on porphyrin uptake, which might lead (upon illumination) to processes that facilitate the formation of reactive oxygen species that damage the cells. Uptake of charged porphyrins is dependent on protein quantity and quality in the photosensitization microenvironment. This fact must be taken into account when using charged photosensitizers.     

GH3 Cells, Ionic Currents and Cell Killing: Photomodification Sensitized by Rose Bengal

Photosensitization using Rose Bengal (RB) modifies membrane ionic currents and kills cultured mouse pituitary, GH₃, cells. Here we investigate the dose-response relationship for ionic current modification and for cell killing to assess a possible causal link. When exposed to 0.5 μM RB and 6.5 mW/cm² of visible light, calcium current was blocked in 1.9 ± 0.2 min (meanSEM; 0.74 ± 0.08 J/ cm²; n = 18), a transient component of potassium current, tentatively identified as a delayed-rectifier potassium current, disappeared in 52 ± 8 s (0.34 ± 0.05 J/cm²; n = 10) and a steady-state component of potassium current, largely a calcium-activated potassium current, disappeared in 3.5 ± 0.4 min (1.37 ± 0.16 J/cm²; n = 11). Conversely, the background leak current increased in magnitude. At 5 min of illumination, the longest time studied here, it continued to increase nearly linearly, making it the only current component studied that is still changing after 5 min of light. Under the conditions used, cell killing increased to 100% in the exposure range of4–10 min of illumination (1.6 J/cm² to 3.9 J/cm²) when assessed using fluorescent markers, ethidium homodimer and calcein and required slightly longer exposure times when assessed using trypan blue. Thus, it is difficult to ascribe a causal role in cell killing by photosensitization to alterations of standard ion channels and known ionic currents. However, the increase in leak current has the correct dose-response characteristics to be involved.     

Photosensitization Reactions of Fluoroquinolones and Their Biological Consequences

This review focuses on damage photosensitized by the fluoroquinolone (FQ) antibacterial drugs. Different models are employed to study biosubstrate photodamage mediated by FQs (organisms, cells, isolated biomolecules and super molecules). Being that the effect of environment (polarity of the medium, ions, pH, binding with bio‐molecules, etc.) is crucial in FQ photochemistry, photobiological reactions can be consequently dramatically influenced. Thus, the photosensitization processes induced by FQs are here discussed taking into account that such extensive and cross‐targeted pathological implications request an excursus covering photosensitization in systems of increasing molecular complexity. In vivo and in vitro evidences for photoallergy, phototoxicity, photomutagenesis and photocarcinogenesis mediated by FQs are discussed.     

Porphyrin Nanorods Modified Glassy Carbon Electrode for the Electrocatalysis of Dioxygen,Methanol and Hydrazine

Porphyrin nanorods (PNR) were prepared by ionic self‐assembly of two oppositely charged porphyrin molecules consisting of free base meso‐tetraphenylsulfonate porphyrin (H₄TPPS₄^2?) and meso‐tetra(N‐methyl‐4‐pyridyl) porphyrin (MTMePyP⁴⁺M=Sn, Mn, In, Co). These consist of H₄TPPS₄^2?? SnTMePyP⁴⁺, H₄TPPS₄^2?? CoTMePyP⁴⁺, H₄TPPS₄^2?? InTMePyP⁴⁺ and H₄TPPS₄^2?? MnTMePyP⁴⁺ porphyrin nanorods. The absorption spectra and transmission electron microscopic (TEM) images of these structures were obtained. These porphyrin nanostructures were used to modify a glassy carbon electrode for the electrocatalytic reduction of oxygen, and the oxidation of hydrazine and methanol at low pH. The cyclic voltammogram of PNR‐modified GCE in pH 2 buffer solution has five irreversible processes, two distinct reduction processes and three oxidation processes. The porphyrin nanorods modified GCE produce good responses especially towards oxygen reduction at ?0.50 V vs. Ag|AgCl (3 M KCl). The process of electrocatalytic oxidation of methanol using PNR‐modified GCE begins at 0.71 V vs. Ag|AgCl (3 M KCl). The electrochemical oxidation of hydrazine began at around 0.36 V on H₄TPPS₄^2?? SnTMePyP⁴⁺ modified GCE. The GCE modified with H₄TPPS₄^2?? CoTMePyP⁴⁺ H₄TPPS₄^2?? InTMePyP⁴⁺ and H₄TPPS₄^2?? MnTMePyP⁴⁺ porphyrin nanorods began oxidizing hydrazine at 0.54 V, 0.59 V and 0.56 V, respectively.     

Photosensitization of Chinese hamster cells by water-soluble phthalocyanines

Abstract— Phthalocyanines are efficient photosensitizers of cultured mammalian cells and are considered for use in photodynamic therapy. The photobiological properties of chloroaluminum phthalocyanine sulfonate (AIPCS) were compared to those of the unsubstituted, water-insoluble derivative (AIPC). The development of photosensitization after addition of the dye into growth medium is ca. 8 times more rapid for AlPC than for AIPCS. Conversely, the loss of photosensitization when cells are incubated in a dye-free growth medium following a period of dye uptake, is also faster for AIPC. The dye uptake followed a kinetic behavior similar to the development of photosensitivity, but the loss of dye was too slow for both AlPC and AIPCS to explain loss of photosensitivity. When cells are incubated prior to illumination with AIPCS in phosphate buffered saline instead of growth medium, shorter time and smaller amount of dye are required to achieve the same level of photosensitization. The dependence of photosensitivity on dye concentration is linear for both AIPC and AIPCS. As already found for AIPC, photosensitization by the water-soluble derivative is also not enhanced in D₂O, suggesting that singlet oxygen is not involved in the cytotoxic response. Sodium salicylate, which was found to enhance the effect of AlPC was also effective with AIPCS. This effect is quite specific since the meta and para isomers had no effect. The metal atom complexed with the phthalocyanine ring is significant for the photobiological activity. Among the compounds tested, those containing Al or Zn are most active.     

MULTIPLE MODALITY TREATMENT OF CARCINOMA CELLS WITH Pt(Rh-123)2 PLUS X-RAY PLUS LIGHT

A complex of platinum tetrachloride with two molecules of rhodamine-123 (Rh-123), Pt(Rh-123)2, has been reported to act as hypoxic cell radiosensitizer of carcinoma cells in vitro and in vivo. In the present paper we report that Pt(Rh-123)2 photosensitizes human mammary carcinoma (MCF-7) cells and cis-platinum resistant human mammary carcinoma (MCF-7/CP) cells to 400-800 nm light in vitro. The efficiency of photosensitization by Pt(Rh-123)2 was 10 times greater than for Rh-123. Combination therapy using Pt(Rh-123)2 plus x-ray plus light was also much more effective compared to the combination therapy using Rh-123 plus x-ray plus light. After 15 microM of Rh-123 plus x-ray (0-8 Gy) plus light (5 J/cm2) treatment, cell survival curve was parallel to the x-ray cell survival curve with an initial decrease in the surviving fraction corresponding to the drug plus light mediated killing. Cell killing caused by Rh-123 (15 microM) plus x-ray (0-8 Gy) plus light (5 J/cm2) was additive as determined by the product of the surviving fraction after Rh-123 plus light and x-ray. In contrast, for 15 microM of Pt(Rh-123)2 plus x-ray (8 Gy) plus light (5 J/cm2) treatment, whereas additive killing predicts a survival fraction of approximately 0.024, in reality, the combination therapy caused the survival fraction to decrease to 0.0012, implying that the cell killing was enhanced by a factor of 20. Using Pt(Rh-123)2 plus x-ray plus light, supra-additive cell killing was also observed under hypoxic conditions, although compared to normally oxygenated conditions the degree of cytotoxicity was significantly reduced.(ABSTRACT TRUNCATED AT 250 WORDS)     

PHOTOSENSITIZING EFFICIENCIES, TUMOR- and CELLULAR UPTAKE OF DIFFERENT PHOTOSENSITIZING DRUGS RELEVANT FOR PHOTODYNAMIC THERAPY OF CANCER

Abstract Several parameters of the following dyes, all relevant as sensitizers for photochemotherapy of cancer, have been studied: Photofrin II (PII), hematoporphyrin (HP)-di-hexyl-ether, HP-di-ethyl-ether, tetra (3-hydroxyphenyl) porphyrin, (3THPP), tetraphenyl porphine tetrasulphonate (TPPS₄) aluminium phthalocyanine tetrasulfonate (A1PCTS), aluminium phthalocyanine (A1PC), chlorin e, (Chi e₆) and merocyanine 540 (MC 540). The following parameters and features of these dyes were studied: (1) Tumor uptake in C3H mouse mammary carcinomas. (2) Skin/tumor concentration ratio in the same animal system. (3) Triton X-114/H₂0 partition coefficients at different pH-values. (4) Uptake of the dyes by human cells of the line NHIK 3025. (5) Relative fluorescence quantum yields of the dyes bound to cells. (6) Absorption-, fluorescence-excitation- and fluorescence-emission spectra of the cell-bound dyes. (7) Relative quantum yields for photoinactivation of cells after 18 h incubation with the dyes. (8) Relative quantum yields of photodegradation of the singlet oxygen trap 1,3-diphenylisobenzofuran (DPBF) in cells after 18 h incubation with the dyes. The following main conclusions were drawn: (1) 3THPP was the best and most selective tumor localizer of the dyes tested, followed by AIPCTS, TPPS₄, PII and Chi e.,. (2) The Triton X-114/H₂0 partition coefficient of most of the dyes decreased with increasing pH. (3) The cellular uptake of the dyes (18 h incubation in medium with 3% serum) increased with increasing Triton X-114/H₂0 partition coefficient. (4) HP-di-hexyl-ether had the highest quantum yields both for photoinactivation of cells and degradation of cell-bound DPBF, followed by the other lipophilic porphyrins and Chi e₆. The water-soluble dyes TPPS₄ and AIPCTS had quantum yields of the order of ten times lower than those of the lipophilic porphyrins. (5) There was a clear correlation between the quantum yields for cell-inactivation and those for photodegradation of DPBF, suggesting that the same reactive photoinduced species is involved in both processes. This suggestion was strengthened by the observation that DPBF reduced the quantum yield of cell inactivation. Thus, all the tested dyes seem to act via a type II process. (6) All of the dyes, even the water-soluble TPPS₄ and AIPCTS, are aggregated in aqueous solutions, and the cells bind both monomers and aggregates. (7) A significant fraction of the cell-bound dyes was located close to tryptophan-containing proteins. (8) Cell-bound Chi e,₆ had the highest fluorescence quantum yield of the dyes.     

An immunomodulator from Tinospora cordifolia with antioxidant activity in cell-free systems

Several plant products are known to exhibit immense medicinal value against human diseases. Our earlier studies showed that dry stem crude extract (DSCE) ofTinospora cordifolia contained a polyclonal B cell mitogen, G1-4A. DSCE as well as G1-4A also enhanced immune response in mice. In order to explore the possibility of using G1-4A/PPI (partially purified immunomodulator) to modulate radiation induced immunosuppression, the antioxidant effect of PPI from this plant was examined against reactive oxygen and nitrogen species (ROS/RNS), generated by photosensitization/peroxynitrite. Levels of lipid peroxidation products, superoxide dismutase (SOD) and catalase in liver/spleen homogenate from mouse were monitored. Photosensitization induced significant increase in thiobarbituric acid reactive substances (TBARS) in liver. The activities of SOD and catalase were reduced considerably. PPI, present during photosensitisation, prevented lipid peroxidation and restored the activities of both the enzymes. Likewise, oxidative damage induced by peroxynitrite was inhibited by PPI. The degradation of proteins due to photosensitization as assessed by SDS-PAGE was effectively reduced by simultaneous treatment with PPI during photosensitization. Selective inhibitors of ROS like mannitol, SOD, sodium azide and antioxidants, GSH and vitamin C brought about significant inhibition of formation of TBARS suggesting possible involvement of O₂ ^•,^•OH and¹O₂. Photosensitization in deuterated buffer enhanced formation of TBARS thus indicating generation of¹O₂. Thus, the action of PPI may be against oxidative damage through Type I and II photosensitization mechanisms. Therefore, the immunomodulator fromTinospora cordifolia may also be beneficial as an antioxidant.     

Self‐Assembly of Anionic Porphyrins and Alkaline or Alkaline Earth Metal Ions Mediated by Cucurbit[7,8]uril

Nanoscaled coordination polymers based on biologically prevalent ions have potential applications in drug delivery and biomedical imaging. Herein, coordination polymer nanoparticles of anionic porphyrins, including meso‐tetra(4‐carboxyphenyl)‐porphyrin (H₂TCPP^4?) and meso‐tetra(4‐sulfonatophenyl)‐porphyrin (H₂TPPS^4?), and alkaline or alkaline earth metal cations, such as K⁺ and Ca²⁺, were constructed in aqueous solution in the presence of cucurbit[7]uril (CB7) or cucurbit[8]uril (CB8). UV/Vis absorption and fluorescence spectroscopy, dynamic light scattering (DLS), scanning electron spectroscopy (SEM), and atomic force microscopy (AFM) were applied to explore the assembly and particle formation of porphyrin anions and metal cations mediated by CBn. The particle size depends on the kinds of CBn and metal cations and their concentrations. The uptake of H₂TPPS^4? particles by tumor cells (A549 cells) was found to be more efficient than H₂TPPS^4? at 37 °C, showing the application potential of such assembled particles in biology and medicine.