RESUMEN
BACKGROUND: Non-invasive indirect blood glucose monitoring can be realized by detecting low concentrations of glucose (0.05-5 mM) in tears, but sensitive optical indicators are required. The intensity of the phosphorescence of a candidate optical indicator, palladium hematoporphyrin monomethyl ether (Pd-HMME), is increased by oxygen consumption under sealed conditions in the presence of glucose and glucose oxidase. However, the glucose detection limit based on this mechanism is high (800 µM) because the phosphorescence is completely quenched under ambient oxygen conditions and hence a large amount of glucose is required to reduce the oxygen levels such that the phosphorescence signal is detectable. RESULTS: To improve the glucose detection limit of Pd-HMME phosphorescence-based methods, the triplet protector imidazole was introduced, and strong phosphorescence was observed under ambient oxygen conditions. Detectable phosphorescence enhancement occurred at low glucose concentrations (<200 µM). Linear correlation between the phosphorescence intensity and glucose concentration was observed in the range of 30-727 µM (R2 = 99.9 %), and the detection limit was â¼10 µM. The glucose sensor has a fast response time (â¼90 s) and excellent selectivity for glucose. SIGNIFICANCE AND NOVELTY: These results indicate the potential of the developed optical indicator for fast, selective, and reliable low-concentration glucose sensing.
Asunto(s)
Límite de Detección , Mediciones Luminiscentes , Mediciones Luminiscentes/métodos , Hematoporfirinas/química , Hematoporfirinas/análisis , Paladio/química , Glucosa/análisis , Glucosa Oxidasa/química , Glucosa Oxidasa/metabolismo , Glucemia/análisis , Imidazoles/química , Técnicas Biosensibles/métodos , Oxígeno/química , HumanosRESUMEN
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.
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Neoplasias de la Mama/diagnóstico por imagen , Mama/diagnóstico por imagen , Hematoporfirinas/análisis , Imagen Óptica/métodos , Fármacos Fotosensibilizantes/análisis , Espectrometría Raman/métodos , Femenino , Humanos , Espectrometría de Fluorescencia/métodosRESUMEN
The present article was designed to demonstrate the efficiency of application of fluorescent microscopy for histological studies of preparations of putrescent material suspected to contain effused blood. This method makes it possible to detect haematoporphirin, a product of haemoglobin degradation, the presence of which gives reliable evidence of the presence of effused blood in the study material.
Asunto(s)
Medicina Legal/métodos , Hematoporfirinas/análisis , Hemorragia/diagnóstico , Microscopía Fluorescente/métodos , Eritrocitos/química , HumanosRESUMEN
In this study, fluorescence excitation and emission matrices and multivariate curve resolution (PARAFAC) were used to detect and characterize active photosensitizers spectrally in butter. Butter samples were packed under high (air) and low oxygen (<0.05%) atmospheres and exposed to violet, green, or red light. Six photosensitizers were found: riboflavin, protoporphyrin, hematoporphyrin, a chlorophyll a-like molecule, and two unidentified tetrapyrrols. By estimation of relative concentrations, we could follow how each sensitizer was photodegraded as function of wavelength, oxygen level, and time. The degradation rate of protoporphyrin, hematoporphyrin, chlorophyll a, and one of the tetrapyrrols correlated well (0.83-0.91) with the formation of sensory measured oxidation. The results suggest that mainly type I photoreactions were responsible for the degradation of photosensitizers in both high and low oxygen atmosphere. Type II photoreactions (generation of singlet oxygen) were involved in the oxidation of butter stored in air. The study shows that PARAFAC modeling of fluorescence landscapes is an excellent tool for studying photooxidation in complex systems.
Asunto(s)
Mantequilla/análisis , Fármacos Fotosensibilizantes/análisis , Espectrometría de Fluorescencia , Clorofila/análisis , Clorofila A , Hematoporfirinas/análisis , Fotoquímica , Fármacos Fotosensibilizantes/química , Protoporfirinas/análisis , Riboflavina/análisisRESUMEN
Since the selectivity of photodynamic therapy (PDT) depends on the distribution of a photosensitizer in a tissue during the treatment, an investigation of drug distribution is a key step for performing PDT effectively. The distribution of photosensitizer absorbed in tissues is adjusted by the animal body system, so an apparatus that can measure the fluorescence intensity of photosensitizer in different tissues of the same body simultaneously is in demand. To achieve precise estimate of tissue selectivity of the photosensitizer, a spatially separated three-channel laser-induced fluorescence (LIF) detection system was set up and employed in the present study to measure the fluorescence intensity of Hematoporphyrin Monomethyl Ether (HMME) in different tissues of the same body simultaneously. The time-dependent variations in the concentrations of HMME within the skin,cartilage, normal synovium and inflammatory synovium of rabbit were monitored in vivo. The results obtained showed that the synovium has higher absorptivity of HMME thanthe skin and cartilage. The difference is distinct from the very beginning of injection. Although the quantity of HMME absorbed in the inflammatory synovium is not very high in the first 20 min, it is still 6 times higher than that in the skin and cartilage. In addition, the absorptivity of HMME is much stronger for the inflammatory synovium than that for the normal synovium. If thelaser beam irradiates outside the joint for the rheumatoid arthritis, tissues around the inflammatory synovium have less HMME, thereby causing weak PDT effect. This would help reduce the side effect of PDT. Thus we suggest that for PDT treated rheumatoid arthritis, taking the first 20 min after the injection for outside-the-joint excitation employing HMME maybe a good choice.
Asunto(s)
Artritis Reumatoide/metabolismo , Fluorescencia , Hematoporfirinas/análisis , Fármacos Fotosensibilizantes/análisis , Membrana Sinovial/metabolismo , Animales , Artritis Reumatoide/patología , Femenino , Hematoporfirinas/química , Humanos , Masculino , Modelos Animales , Fotoquímica/métodos , Fármacos Fotosensibilizantes/química , Conejos , Ratas , Ratas Endogámicas LewRESUMEN
The on-line incorporation of cloud point extraction (CPE) to flow injection analysis (FIA) is demonstrated for the first time. The technical difficulties of inducing the cloud point phenomenon, separating the surfactant-rich phase from the aqueous phase, and detecting trace amounts of analyte(s) in the presence of the highly scattering surfactant medium in an on-line FIA system were resolved by the following: (1) mixing the sample solution containing the analyte(s) and CPE surfactant with an appropriate salting-out agent, (2) using a collection column to entrap the analyte-containing surfactant aggregates, and (3) employing the peroxyoxalate chemiluminescence reaction for the sensitive and selective determination of the analyte(s) in the presence of surfactant micelles. The figures of merit for the determination of coproporphyrin in pretreated urine samples were as follows: precision, 1.1-2.2% (RSD); limit of detection, 2.0 microg/L; and the calibration curve was linear from 46 to 2319 (micro/L (r = 0.9996).
Asunto(s)
Coproporfirinas/orina , Análisis de Inyección de Flujo/métodos , Hematoporfirinas/análisis , Humanos , Estándares de Referencia , Solubilidad , Tensoactivos/químicaRESUMEN
Hematoporphyrin (HP) was used as a microenvironmental fluorescent probe to investigate the development of human atherosclerotic plaques. We compared the site of HP accumulation with changes in the HP fluorescence spectrum in atheromatous plaques and in a liposome control model, using a confocal laser scanning microscope equipped with a photonic multi-channel analyzer linked to a fluorescence spectrometer. Wavelength shifts of the two peaks of HP fluorescence (F1, 620 nm; and F2, 640-690 nm) were monitored, and the integrated F2/F1 ratio was calculated as a measure of HP fluorescence. The F1 peak is characteristic of a predominantly aqueous site. The ratio reflects selective changes in the distribution and overcrowding of HP molecules in the limited space of the artificial membrane model. Compared with a normal artery, the atherosclerotic lesions showed an increase in the area of HP fluorescence, increased fluorescence intensity, a red shift of the HP fluorescence spectrum, and an increased F2/F1 ratio. The F2/F1 ratio of the membranous structures was markedly greater in the cores of the fibrous plaques than in the other plaques. The F1 peak showed increased intensity in the atheromatous plaque core, whereas in hydrophilic fibrous regions, such as the cap of the plaque, the intensity of the F1 peak was lower than in the core region. HP aggregation was observed in damaged cells and in water surrounded by lipid in the atheromatous core. Using HP as a probe allowed us to determine not only the ionization or polarity of each region in the atherosclerotic plaques but also to detect the separation and fusion of the lipid bilayer or micelle lipids, as well as damage to the cellular membranes and cholesterol enrichment. These findings suggest that HP is useful for detecting clinically important changes in atherosclerotic lesions, to lipid-rich, unstable, and vulnerable plaques, which are closely associated with cardiovascular events.
Asunto(s)
Aorta Abdominal/patología , Arteriosclerosis/patología , Colesterol/análisis , Hematoporfirinas/farmacocinética , Adulto , Anciano , Aorta Abdominal/citología , Arteriosclerosis/clasificación , Colorantes , Femenino , Hematoporfirinas/análisis , Histocitoquímica/métodos , Humanos , Liposomas , Masculino , Microscopía Confocal , Persona de Mediana Edad , Valores de Referencia , Espectrometría de Fluorescencia/métodosRESUMEN
Photo-detection using in vivo fluorescence was studied for different stages of chemically induced premalignant lesions and squamous cell carcinoma (SCC) of the Wistar rat palatal mucosa. It was found that the epithelial dysplasia (numerically expressed in the epithelial atypia index (EAI) of the rat palate, induced by repeated application of the carcinogen 4-nitroquinoline 1-oxide (4NQO), showed an increase approximately proportional to the duration of the application period. Photo-detection of the lesions using Photofrin-induced fluorescence was studied with dual-wavelength excitation and the subtraction of images, in an attempt to reduce the autofluorescence. The Photofrin dose was 2.5 mg kg-1. This was based on a dose-response study for normal tissue damage by photodynamic therapy (PDT) in this animal model, because the underlying rationale was to study photo-detection as a method of locating additional (early) malignancies in patients treated by PDT. Fluorescence intensities 24 and 48 h after injection of Photofrin were shown to increase with the duration of 4NQO application and with increasing EAI. For an EAI greater than 15, there was a statistically significant difference (p < 0.01) between the fluorescence signals obtained with and without the injection of Photofrin. Fluorescence signals of these lesions without the use of Photofrin (autofluorescence) also showed an increase with increasing stages of epithelial dysplasia of the rat palate. However, the fluorescence signals obtained with Photofrin were always higher than those of the autofluorescence. From this study, we conclude that photo-detection with Photofrin has potential in distinguishing chemically induced premalignant lesions and squamous cell carcinomas from the normal rat palatal mucosa. Photofrin (2.5 mg per kg of body weight) certainly adds to the sensitivity of photo-detection, but autofluorescence alone also has promising features for detecting premalignant and malignant lesions of the oral mucosa.
Asunto(s)
Carcinoma de Células Escamosas/diagnóstico , Fotoquimioterapia , 4-Nitroquinolina-1-Óxido/farmacología , Animales , Carcinógenos/farmacología , Carcinoma de Células Escamosas/inducido químicamente , Hematoporfirinas/análisis , Hematoporfirinas/química , Aumento de la Imagen , Masculino , Mucosa Bucal/patología , Neoplasias Palatinas/inducido químicamente , Neoplasias Palatinas/diagnóstico , Neoplasias Palatinas/patología , Lesiones Precancerosas/inducido químicamente , Lesiones Precancerosas/diagnóstico , Ratas , Ratas Wistar , Espectrometría de FluorescenciaRESUMEN
Photodynamic therapy (PDT) of cancer typically involves systemic administration of tumor-localizing photosensitizers followed 48-72 h later by exposure to light of appropriate wavelengths. Knowledge about the distribution of photosensitizers in tissues is still fragmentary. In particular, little is known as to the detailed localization patterns of photosensitizers in neoplastic and normal tissues as well as the relationship between such patterns and the actual targets for the photosensitizing effect. This review focuses on ultrastructural features seen in treated cells and tumors. An attempt is made to correlate these findings with the subcellular/intratumoral localization pattern of the photosensitizers in tumor cell lines in vitro and in tumor models in vivo. Several subcellular sites are main targets of PDT with different sulfonated aluminum phthalocyanines (AIPcSn) in the human tumor cell line LOX. Nuclei are not among the primary targets. Overall, the ultrastructural changes correlate well with the data about the subcellular localization patterns for each analogue of AIPcSn in the same cell line. Similar findings are also obtained for the family of sulfonated mesotetraphenylporphines (TPPSn) in the NHIK 3025 cell line. The mechanisms involved in the killing of tumors by PDT seem to be a complex interplay between direct and indirect (via vascular damage) effects on neoplastic cells according to the intratumoral localization pattern of the applied dye. Several factors can affect the localization pattern of a drug, such as its chemical character, the mode of drug delivery, the time interval between drug administration and light exposure, and tumor type. Furthermore, whether local immune reactions (such as macrophages) and apoptosis (programmed cell death) are involved in the destruction of neoplastic cells by PDT in vivo is still an enigma. A general model for PDT-induced tumor destruction is suggested.
Asunto(s)
Hematoporfirinas/análisis , Neoplasias Experimentales/tratamiento farmacológico , Neoplasias Experimentales/ultraestructura , Fotoquimioterapia , Fármacos Fotosensibilizantes/análisis , Animales , Humanos , Fracciones Subcelulares/ultraestructuraRESUMEN
Photodynamic therapy (PDT) was performed in vitro and in vivo using monoclonal antibody conjugated to hematoporphyrin (HP). The antibody (45-2D9) recognized a cell surface glycoprotein on cells derived from NIH 3T3 cells which were transformed with the ras oncogene (45-342). Radionuclide imaging with either In111 or I125 chelated to 45-2D9 or the isotype identical (IgG1) antibody MOPPC-21 revealed selectivity of 45-2D9 for 45-342 flank tumours in nude mice, and minimal targetting for a 45-342 clone which did not express the cell surface glycoprotein. The 45-2D9-HP conjugate resulted in selective killing of the 45-342 line compared with the parent line in vitro. At HP concentrations of 76 micrograms ml-1, the 45-2D9-HP conjugate resulted in significantly more long-term cures of PDT treated flank tumours compared with free HP at the same concentration. 45-2D9 alone had no effect on tumour growth. The antibody-HP conjugate resulted in significantly less local toxicity compared with standard Photofrin II PDT, and also achieved a greater number of long-term cures. This 'photoimmunotherapy' demonstrates the ability to treat established tumours with greater efficacy and decreased morbidity, probably due to specific sensitizer targetting which allows normal surrounding tissue to be spared upon illumination.
Asunto(s)
Anticuerpos Monoclonales/uso terapéutico , Éter de Dihematoporfirina/uso terapéutico , Neoplasias Experimentales/tratamiento farmacológico , Fotoquimioterapia , Células 3T3 , Animales , Éter de Dihematoporfirina/toxicidad , Hematoporfirinas/análisis , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Músculos/química , Neoplasias Experimentales/química , Piel/química , Resultado del TratamientoRESUMEN
By means of laser scanning fluorescence microscopy the intratumoral localization patterns of several photosensitizers in LOX tumors in nude mice were studied. Lipophilic dyes such as TPPS1 (tetraphenylporphine monosulfonate), TPPS2a (tetraphenylporphine disulfonates with the sulfonate groups on adjacent rings), AlPCS1 (aluminium phthalocyanine monosulfonate) and AlPCS2 (aluminium phthalocyanine disulfonates) localized mainly in tumor cells. The fluorescence intensity of these dyes increased from 4 h to 48 h postinjection and the fluorescence was still observable 120 h postinjection. The more hydrophilic dyes such as TPPS3 (tetraphenylporphine trisulfonates), TPPS4 (tetraphenylporphine tetrasulfonates), and AlPCS4 (aluminium phthalocyanine tetrasulfonates) localized mainly extracellularly in the tumorous stroma. The fluorescence intensity of these dyes decreased from 4 h to 48 h postinjection. 120 h postinjection no significant fluorescence of these dyes could be seen in the tumors. P-II (Photofrin II), 3-THPP [tetra(3-hydroxyphenyl)porphine], TPPS2o (tetraphenylporphine disulfonates with the sulfonate groups on opposite rings) and AlPCS3 (aluminum phthalocyanine trisulfonates) had a combined localization pattern, i.e. a strongly cytoplasmic membrane-localizing pattern and a weakly intracellular distribution pattern, although some fluorescence could be seen in the tumorous stroma. The data are discussed in relation to what is known about the in vivo photosensitizing efficiency of some of the dyes.
Asunto(s)
Colorantes Fluorescentes , Hematoporfirinas/análisis , Indoles/análisis , Melanoma/metabolismo , Compuestos Organometálicos/análisis , Porfirinas/análisis , Fármacos Sensibilizantes a Radiaciones/análisis , Animales , Línea Celular , Éter de Dihematoporfirina , Femenino , Humanos , Rayos Láser , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Microscopía de Contraste de Fase/métodos , Fotoquimioterapia/métodosRESUMEN
The patterns of in vitro intracellular and in vivo intratumoral localization of Photofrin II (P-II) and aluminum phthalocyanine tetrasulfonate (AlPCS4) in human melanoma LOX were studied by means of computer-enhanced video fluorescence microscopy (CEVFM). The hydrophobic drug P-II localized diffusely in the perinuclear fraction of the cytoplasm of the LOX cells cultivated in vitro. Light exposure did not result in any observable change in the localization pattern. The hydrophilic dye AlPCS4 was distributed as granular and grain patterns in the cytoplasm before light exposure, in exactly the same pattern as that of acridine orange incubated in the same cells, which is known to emit red fluorescence from lysosomes, thus indicating that AlPCS4 was also primarily localized in the lysosomes of the LOX cells. After light exposure the distribution of the intracellular AlPCS4 fluorescence was altered and the intensity increased. In vivo, P-II had a combined cellular localization pattern (i.e. a strongly cytoplasmic membrane-localizing pattern and a weakly intracellular distribution pattern) and an extracellular distribution pattern in the tumor tissue, while the AlPCS4 fluorescence was seen mainly in the stroma of the tumor. The total fluorescence intensity of P-II and AlPCS4 in the LOX tumor tissue at different times after injection was quantitatively determined by means of CEVFM.
Asunto(s)
Hematoporfirinas/análisis , Indoles/análisis , Melanoma/metabolismo , Microscopía Fluorescente/métodos , Compuestos Organometálicos/análisis , Animales , Línea Celular , Computadores , Citoplasma/metabolismo , Éter de Dihematoporfirina , Femenino , Humanos , Lisosomas/metabolismo , Ratones , Ratones Endogámicos BALB C , Fotoquimioterapia/métodos , Células Tumorales Cultivadas , Grabación en VideoAsunto(s)
Terapia por Láser , Neoplasias/diagnóstico , Neoplasias/terapia , Fotoquimioterapia , Porfirinas , Animales , Hematoporfirinas/análisis , Hematoporfirinas/uso terapéutico , Humanos , Neoplasias/radioterapia , Neoplasias Experimentales/diagnóstico , Neoplasias Experimentales/radioterapia , Neoplasias Experimentales/terapia , Porfirinas/análisis , Porfirinas/uso terapéutico , RatasRESUMEN
By means of laser scanning fluorescence microscopy the intratumoral localization patterns of several photosensitizers in LOX tumors in nude mice were studied. Lipophilic dyes such as P-II (Photofrin II), 3-THPP tetra(3-hydroxyphenyl)porphin, TPPS1 (tetraphenylporphine monosulfonate), TPPS2a (tetraphenylporphine disulfonates with the sulfonate groups on adjacent rings), A1PCS1 (aluminium phthalocyanine monosulfonate) and A1PCS2 (aluminium phthalocyanine disulfonates) localized mainly in tumor cells. The fluorescence intensity of these dyes increased from 4 h to 48 h post-injection and the fluorescence was still observable 120 h post-injection. The more hydrophilic dyes such as TPPS2o (tetraphenylporphine disulfonates with the sulfonates groups on opposite rings), TPPS3 (tetraphenylporphine trisulfoantes), TPPS4 (tetraphenylporphine tetrasulfonates), A1PCS3 (aluminium phthalocyanine trisulfonates) and A1PCS4 (aluminium phthalocyanine tetrasulfonates) localized mainly extracellularly in the tumorous stroma. The fluorescence intensity of these dyes decreased from 4 h to 48 h post-injection. 120 h post-injection no significant fluorescence of these dyes could be seen in the tumors. The data are discussed in relation to what is known about the in vivo photosensitizing efficiency of some of the dyes.
Asunto(s)
Colorantes Fluorescentes , Hematoporfirinas/análisis , Indoles/análisis , Melanoma/metabolismo , Compuestos Organometálicos/análisis , Porfirinas/análisis , Fármacos Sensibilizantes a Radiaciones/análisis , Animales , Línea Celular , Éter de Dihematoporfirina , Femenino , Humanos , Rayos Láser , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Microscopía de Contraste de Fase/métodos , Fotoquimioterapia/métodosRESUMEN
A comparative kinetic observation of the in vivo biolocalization of Photofrin II (P-II) and aluminum phthalocyanine tetrasulfonate (AIPCS4) in a transplanted human malignant tumor LOX and in normal tissues of nude mice has been made by means of highly light-sensitive video intensification microscopy at various intervals after i.p. administration. In the human tumor LOX, transplanted to athymic nude mice, fluorescence of P-II was observed on the membrane and in the cytoplasm of tumor cells, and in the stroma 4-48 hr post-injection. From 72 hr post-injection almost all fluorescing P-II had disappeared from the membrane of the tumor cells while strong fluorescence was still found in the stroma. AIPCS4 fluorescence was seen mainly in tumorous stroma with none detected in the tumor cells. Almost no fluorescence was found in the tumorous stroma 24 hr after injection. In most normal tissues observed, P-II was eliminated at a much slower rate than AIPCS4, but the in vivo biolocalization of the 2 drugs was similar. They were observed primarily where collagenous proteins are normally found, i.e. basal lamina, collagenous connective tissue, and in keratinized epithelium, renal epithelium, mononuclear phagocyte system and on the membrane of muscular cells. In addition, AIPCS4 had a strong affinity for the bronchiogenic epithelium. In the skin, P-II was distributed in keratinized epithelium, hair, hair follicles and their accessory, collagenous connective tissue of dermis, whereas AIPCS4 was present only in hair and collagenous connective tissue of dermis. No fluorescence of P-II or of AIPCS4 was found in the skin epidermis, nor in the transitional epithelium of the bladder mucosa.
Asunto(s)
Hematoporfirinas/farmacocinética , Indoles/farmacocinética , Melanoma/patología , Compuestos Organometálicos/farmacocinética , Fármacos Sensibilizantes a Radiaciones/farmacocinética , Animales , Línea Celular , Éter de Dihematoporfirina , Femenino , Hematoporfirinas/análisis , Humanos , Indoles/análisis , Ratones , Ratones Desnudos , Microscopía Fluorescente/métodos , Trasplante de Neoplasias , Compuestos Organometálicos/análisis , Piel/patología , Distribución Tisular , Trasplante HeterólogoRESUMEN
The inhibitory effects of synthetic phenolic compounds on benzo(a)pyrene-induced neoplasia of the mouse forestomach have been measured by Wattenberg et al. The efficiency of this inhibition has been estimated for each phenol, using R, the ratio of the number of tumors per mouse in the protected group over the number of tumours per mouse in the control group. We have observed a linear correlation between the chemoprotection efficiency R and the logarithm of the rate of quenching of singlet oxygen, k, by this family of phenols, log k being itself correlated with the one-electron oxidation potential of the phenols. These correlations suggest a charge transfer mechanism for the inhibition of neoplasia induced by benzo(a)pyrene. The correlations described provide a theoretical basis for scaling the inhibitors of mutagenicity induced by polycyclic aromatic compounds in terms of their oxidation potentials.
Asunto(s)
Antineoplásicos , Neoplasias Experimentales/tratamiento farmacológico , Fenoles/análisis , Animales , Antioxidantes/farmacología , Benzo(a)pireno , Tampones (Química) , Electroquímica , Hematoporfirinas/análisis , Ratones , Neoplasias Experimentales/inducido químicamente , Oxidación-Reducción , Oxígeno/análisis , Fenoles/farmacología , Radiólisis de ImpulsoRESUMEN
The oligomeric composition of HpD, Photofrin II and other hematoporphyrin derivatives useful for the diagnosis and therapy of tumors has been studied. Gel chromatographic procedures were used that excluded porphyrin aggregation. Photofrin and hematoporphyrin derivatives were shown to contain different quantities of monomer, dimer and other oligomeric porphyrins.
Asunto(s)
Hematoporfirinas/análisis , Cromatografía en Gel , Cromatografía en Capa Delgada , Estabilidad de Medicamentos , Polímeros , SolucionesRESUMEN
Flow cytometry (FCM) has been used to investigate the intracellular fluorescence of hematoporphyrin derivative (HPD) in monolayer and spheroid cultures of WiDr cells. For exponentially-growing monolayer cultures mean cellular fluorescence was directly proportion to the external HPD levels in the range 5-100 micrograms ml-1 (r = 0.99). Heterogeneity of cellular fluorescence was quantified by determining the ratio of the fluorescence value below which were observed values for 98% of the cell population compared to the fluorescence value for 2%. In exponentially-growing cultures, decreasing levels of HPD in the medium led to an increase in the 98:2% ratio, i.e. an increase in heterogeneity of intracellular drug levels. The growth of cells as multicellular spheroids confers a spheroid-size-dependent resistance to photodynamic treatment. With increasing spheroid size (100, 250, 500, 750 and 1000 microns diam.) there was a decrease in mean intracellular HPD levels and a large linear increase in the 98:2% ratio (r = 0.94).
Asunto(s)
Adenocarcinoma/análisis , Neoplasias del Colon/análisis , Citometría de Flujo , Hematoporfirinas/análisis , Adenocarcinoma/metabolismo , Adenocarcinoma/patología , Neoplasias del Colon/metabolismo , Neoplasias del Colon/patología , Resistencia a Medicamentos , Derivado de la Hematoporfirina , Hematoporfirinas/metabolismo , Humanos , Fotoquimioterapia , Células Tumorales CultivadasRESUMEN
Dilute aqueous solutions of hematoporphyrin (Hp) and its derivative (Hpd or PF II) have been found to undergo a transformation (aging) on keeping at room temperature leading to (i) shift of the Soret band from 395 nm to 405 nm, (ii) disappearance of visible bands I (610 nm) and IV (503 nm) and (iii) shift of the first emission band from 615 nm to 580 nm. The transformation was concentration dependent. The effects of concentration and temperature on the absorption spectra were much more pronounced in Hp than in Photofrin II (PF II). Variation of pH resulted in changes in the relative intensities of the absorption bands, possibly due to formation of different ionic species at different pH. The rate of transformation was accelerated in the presence of Zn ions (0.01 microM) and considerably increased at higher (50 microM) concentration. The effect of Cu ions was different from the effect of aging. It formed the metal-chelate even when present in very small amounts. The results (absorption and fluorescence analysis) suggest that in dilute solutions (conc. less than or equal to 2 microM) of Hp and PF II, Zn ions present in glass and water as impurity, deform the porphyrin nucleus leading to changes in the conjugated ring symmetry and hence changes in the absorption and fluorescence spectra, while in higher concentrations (greater than 2 microM) it forms the metal chelate as evidenced by their absorption and fluorescence spectra.