A lysosome specific theranostic NO donor inhibits cancer cells by stimuli responsive molecular self-decomposition with an on-demand fluorescence pattern.

The anticancer mechanism of NO is difficult to study owing to its short lifetime and high reactivity. Thus, a theranostic anticancer NO donor assembled with NO on-demand release abilities, accurate lysosome location capabilities and signal feedback behavior was developed. Profiting from the theranostic properties, the specific mechanism was comprehensively studied. Spectral and cell imaging studies revealed that the as prepared NO donors could release NO in solution or within cancer cells. Fluorescence co-dyeing experiments demonstrated that Mo-Nap-NO entered lysosomes specifically and disrupted them after being triggered by light. Upon irradiation with 460 nm visible light, both the donors demonstrated considerable in vitro anticancer effects. A further mechanistic study showed that after entering the lysosome and being triggered by 460 nm irradiation, NO ruptured the lysosome, resulting in the release of cathepsin D into the cytosol, which activated the caspase3 mediated apoptosis pathway.

A molecular hybrid producing simultaneously singlet oxygen and nitric oxide by single photon excitation with green light.

Combination of photosensitizers (PS) for photodynamic therapy with NO photodonors (NOPD) is opening intriguing horizons towards new and still underexplored multimodal anticancer and antibacterial treatments not based on "conventional" drugs and entirely controlled by light stimuli. In this contribution, we report an intriguing molecular hybrid based on a BODIPY light-harvesting antenna that acts simultaneously as PS and NOPD upon single photon excitation with the highly biocompatible green light. The presented hybrid offers a combination of superior advantages with respect to the other rare cases reported to date, meeting most of the key criteria for both PSs and NOPDs in the same molecular entity such as: (i) capability to generate 1O2 and NO with single photon excitation of biocompatible visible light, (ii) excellent 1O2 quantum yield and NO quantum efficiency, (iii) photogeneration of NO independent from the presence of oxygen, (iv) large light harvesting properties in the green region. Furthermore, this compound together with its stable photoproduct, is well tolerated by both normal and cancer cells in the dark and exhibits bimodal photomortality of cancer cells under green light excitation due to the combined action of the cytotoxic 1O2 and NO.

Super-Resolution Monitoring of Mitochondrial Dynamics upon Time-Gated Photo-Triggered Release of Nitric Oxide.

Nitric oxide (NO) potentially plays a regulatory role in mitochondrial fusion and fission, which are vital to cell survival and implicated in health, disease, and aging. Molecular tools facilitating the study of the relationship between NO and mitochondrial dynamics are in need. We have recently developed a novel NO donor (NOD550). Upon photoactivation, NOD550 decomposes to release two NO molecules and a fluorophore. The NO release could be spatially mapped with subdiffraction resolution and with a temporal resolution of 10 s. Due to the preferential localization of NOD550 at mitochondria, morphology and dynamics of mitochondria could be monitored upon NO release from NOD550.

Factors affecting ultraviolet irradiation/hydrogen peroxide (UV/H2O2) degradation of mixed N-nitrosamines in water.

Disinfection by-products (DBPs) are a great challenge to our drinking water security. Particularly, nitrosamines (NAms), as emerging DBPs, are potently carcinogenic, mutagenic, and teratogenic, and have increasingly attained public attention. This study was to evaluate the performance of the NAms degradation by the ultraviolet (UV) irradiation (253.7 nm) in the presence of hydrogen peroxide (H(2)O(2)). In the UV/H(2)O(2) system, hydroxyl radicals (OH), a type of nonselective and powerful oxidant, was produced to attack the molecules of NAms. Factors affecting the treatment efficiency, including the H(2)O(2) dosage, initial NAms concentration, UV irradiation intensity, initial solution pH, and inorganic anions present in water, were evaluated. All the NAms degradation exhibited a pseudo-first-order kinetics pattern. Within 60 min, 0.1 mg/L of any NAms could be almost decomposed except NDPhA that required 120 min for complete removal, at 25 µmol/L H(2)O(2) and at initial pH 7. Results demonstrate that the UV/H(2)O(2) treatment is a viable option to control NAms in water.

UVA activation of N-dialkylnitrosamines releasing nitric oxide, producing strand breaks as well as oxidative damages in DNA, and inducing mutations in the Ames test.

We investigated the photo-mutagenicity and photo-genotoxicity of N-dialkylnitrosamines and its mechanisms of UVA activation. With simultaneous irradiation of UVA, photo-mutagenicity of seven N-dialkylnitrosamines was observed in Ames bacteria (Salmonella typhimurium TA1535) in the absence of metabolic activation. Mutagenicity of pre-irradiated N-dialkylnitrosamines was also observed with S. typhimurium hisG46, TA100, TA102 and YG7108 in the absence of metabolic activation. UVA-mediated mutation with N-nitrosodimethylamine (NDMA) and N-nitrosodiethylamine (NDEA) decreased by adding either the NO or OH radical scavenger. When superhelical DNA was irradiated with N-dialkylnitrosamines, nicked circular DNA appeared. Ten N-dialkylnitrosamines examined produced strand breaks in the treated DNA in the presence of UVA. The level of single-strand breaks in phiX174 DNA mediated by N-nitrosomorpholine (NMOR) and UVA decreased by adding either a radical scavenger or superoxide dismutase. When calf thymus DNA was treated with N-dialkylnitrosamines (NDMA, NDEA, NMOR, N-nitrosopyrrolidine (NPYR) and N-nitrosopiperidine (NPIP)) and UVA, the ratio of 8-oxodG/dG in the DNA increased. Action spectra were obtained to determine if nitrosamine acts as a sensitizer of UVA. Both mutation frequency and NO formation were highest at the absorption maximum of nitrosamines, approximately 340 nm. The plots of NO formation and mutation frequency align with the absorption curve of NPYR, NMOR and NDMA. A significant linear correlation between the optical density of N-dialkynitrosamines at 340 nm and NO formation in each irradiated solution was revealed by ANOVA. We would like to propose the hypothesis that the N-nitroso moiety of N-dialkylnitrosamines absorbs UVA photons, UVA-photolysis of N-dialkylnitrosamines brings release of nitric oxide, and subsequent production of alkyl radical cations and active oxygen species follow as secondary events, which cause DNA strand breaks, oxidative and alkylative DNA damages and mutation.

N-nitrosodimethylamine (NDMA) removal by reverse osmosis and UV treatment and analysis via LC-MS/MS.

N-nitrosodimethylamine (NDMA) is a probable human carcinogen found in ng/l concentrations in chlorinated and chloraminated water. A method was developed for the determination of ng/l levels of NDMA using liquid chromatography-tandem mass spectrometry (LC-MS/MS) preceded by sample concentration via solid-phase extraction with activated charcoal. Recoveries were greater than 90% and allowed a method reporting limit as low as 2ng/l. Using this method, the removal of NDMA was determined for the Interim Water Purification Facility (IWPF), an advanced wastewater treatment facility operated by the Orange County Water District (OCWD) in Southern California. The facility treats effluent from an activated sludge treatment plant with microfiltration (MF), reverse osmosis (RO), and an ultraviolet-hydrogen peroxide advanced oxidation process (UV-AOP). Six nitrosamines were surveyed: NDMA, N-nitrosomethylethylamine (NMEA), N-nitrosodiethylamine (NDEA), N-nitrosodi-n-propylamine (NDPA), N-nitrosopiperidine (NPip), and N-nitrosopyrrolidine (NPyr). Only NDMA was detected and at all treatment steps in the IWPF, with influent concentrations ranging from 20 to 59 ng/l. Removals for RO and UV ranged from 24% to 56% and 43% to 66%, respectively. Overall, 69+/-7% of the original NDMA concentration was removed from the product water across the advanced treatment process and, in combination with blending, the final concentration did not exceed the California drinking water notification level of 10 ng/l. NDMA removal data are consistent with findings reviewed for other advanced treatment facilities and laboratory studies.

Effect of modified atmosphere packaging and irradiation in combination on content of nitrosamines in cooked pork sausage.

The effect of modified atmosphere packaging and irradiation in combination on nitrosodimethylamine (NDMA) and nitrosopyrrolidine (NPYR) levels in pork sausage was studied. Emulsion-type cooked pork sausage was manufactured and packaged in aerobic, CO2 (100%), N2 (100%), and CO2/N2 (25%/75%) environments, respectively, and irradiated at 0, 5, 10, and 20 kGy with gamma irradiation. The nitrosamine contents were significantly reduced by irradiation, and the reduction of nitrosamines was more extensive with modified atmosphere packaging than with aerobic packaging. The correlation coefficient between irradiation dose and nitrosamine content indicated that irradiation can reduce the levels of nitrosamines. The combination of irradiation and modified atmosphere packaging is effective in enhancing the chemical safety of sausage by reducing nitrosamines, if present, as well as enhancing the microbial safety of cooked pork sausage.

Release of nitric oxide together with carbon-centered radicals from N-nitrosamines by ultraviolet light irradiation.

Solutions of N-nitrosamines, N-nitrosodimethylamine, N-nitrosodiethylamine, N-nitrosomorpholine and N-nitrosopyrrolidine in phosphate buffer (pH 7.4) were irradiated by ultraviolet (UV) light at room temperature. The N-nitrosamines were extensively degraded due to irradiation for 120 min in a time-dependent fashion as monitored by UV-absorption or high performance liquid chromatographic analysis. Carbon-centered radicals were generated from four N-nitrosamines during the short time irradiation of 10-60 s as monitored by electron spin resonance (ESR) technique using 5,5-dimethyl-1-pyrroline N-oxide and N-tert-butyl-alpha-phenylnitrone as spin traps. Nitric oxide (NO) was generated during the short time irradiation as monitored by ESR technique using cysteine-Fe(II) complex, N-methyl-D-glucamine dithiocarbamate and 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide. Significant amounts of nitrite (4-16%) from four N-nitrosamines and also a significant amount of nitrate (4%) was produced from N-nitrosodimethylamine during the irradiation time of 120 min. Released NO from the N-nitrosamines must be converted into nitrite through intermediary reactive nitrogen oxide species including nitrogen dioxide and dinitrogen trioxide in contact with dissolved oxygen.

Synergistic action of N-nitrosodialkylamines and near-UV in the induction of chromosome aberrations in Chinese hamster lung fibroblasts in vitro.

N-Nitrosodialkylamines are promutagens and proclastogens, requiring metabolic activation for their actions. Previously, we showed that direct-acting bacterial mutagens can be formed from N-nitrosodialkylamines on exposure to near-UV. We have now found that N-nitrosodialkylamines with near-UV irradiation are clastogenic to Chinese hamster lung cells. When the cells in culture were irradiated with near-UV for 3 h in the presence of N-nitrosodimethylamine (NDMA), N-nitrosodiethylamine (NDEA), N-nitrosopyrrolidine (NPYR), N-nitrosopiperidine (NPIP) or N-nitrosomorpholine (NMOR), and then further incubated for a total period of 24 h with the N-nitrosodialkylamines, chromosome aberrations were induced. Neither the N-nitrosodialkylamine nor near-UV alone were clastogenic. Severe clastogenicity (> 50% of cells examined showing aberrations) was observed for 0.5 mM NDEA, NPYR and NPIP. The order of the clastogenic activity was NDEA, NPYR > NPIP, NDMA > NMOR. This order differed from that of bacterial mutagenicity previously reported for these N-nitrosodialkylamines plus near-UV, in which NMOR gave the strongest activity. The chromosome aberrations induced by the NPYR and NDEA plus near-UV in CHL-cells were inhibited by superoxide dismutase, glutathione and L-cysteine. Dimethylsulfoxide or D-mannitol, scavengers of hydroxy radical, and L-histidine, a scavenger of single oxygen, were ineffective. These results suggest that superoxide formed by a synergistic action of an N-nitrosodialkylamine and near-UV is the cause of the chromosome aberrations observed, an assumption consistent with the known ability of superoxide to cleave DNA.

[The effect of ionizing radiation on nitrosamine formation from anthropogenic precursors]. / Vliianie ioniziruiushchego izlucheniia na obrazovanie nitrozoaminov iz antropogennykh predshestvennikov.

The kinetics of formation of nitrosoamines from precursors under the effect of ionizing radiation has first been quantitatively estimated, and the optimal conditions of their formation determined. The possibility of formation of nitrosoamines under the effect of low-level radiation has been investigated. Using the mathematical functions obtained from the experiments, the number of nitrosoamines, that may form under the effect of radiation in the aqueous medium, has been determined.

Formation of directly mutagenic alpha-hydroxy-N-nitrosopiperidine phosphate ester by near-ultraviolet irradiation of N-nitrosopiperidine in phosphate buffer.

Previously we found that direct-acting mutagens can be formed from N-nitrosodialkylamines on exposure to near-ultraviolet light in the presence of phosphates. We have now isolated the mutagenic photoproduct formed from N-nitrosopiperidine and inorganic phosphate and identified its structure as the phosphate ester of alpha-hydroxy-N-nitrosopiperidine. This reaction represents a new, non-enzymatic activation of promutagenic N-nitrosodialkylamines.

Photolysis of nitrosamines and nitrosamides at neutral pH: a spin-trap study.

A model system has been used to study the types of radicals formed on denitrosation of N-nitroso compounds. Free radicals were formed at room temperature (22 degrees-23 degrees C) and neutral pH by photolytic cleavage of N-nitroso bonds and were partially characterized following their addition to the spin traps 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) and N-tert-butyl-alpha-phenyl-nitrone (PBN). Carbon-centered radical adducts were obtained during nitrosamine photolysis and nitrogen-centered radical adducts during nitrosamide photolysis. Since both the nitrosamines and nitrosamides initially form nitrogen-centered radicals on photolysis, a secondary reaction or rearrangement must occur after initial N-nitroso bond cleavage in the nitrosamines. Mechanisms are proposed to account for these results.

Activation of N-nitrosodialkylamines by near-ultraviolet irradiation: formation of directly-acting mutagens and DNA-damaging products.

On near-ultraviolet (UVA) irradiation in a phosphate buffer, N-nitrosomorpholine (NMOR) and N-nitrosopyrrolidine (NPYR) were converted into directly-acting mutagens. The activated NPYR was fractionated, and the active product was isolated. The compound was shown to be identical to alpha-phosphonooxy NPYR on the basis of several properties: retention times in high-performance liquid chromatograms, mutagenic specificity and potency, ultraviolet spectrum, and inactivation by phosphatase treatment. Photoactivation was inhibited by superoxide dismutase, and therefore superoxide is implicated as playing a key role in mutagen formation. N-Nitrosoproline (NPRO) and eighteen other N-nitrosodialkylamines were irradiated with UVA in the presence of phi X174 RFI DNA. The DNA underwent single-strand breaks to give RFII DNA, indicating that N-nitrosodialkylamines in general have this property. DNA chain cleavage was inhibited both by superoxide dismutase and hydroxyl-radical scavengers. These results provide new information on the genotoxic mechanism of action of N-nitrosodialkylamines.

Mutagenicity arising from near-ultraviolet irradiation of N-nitrosomorpholine.

When a mixture of N-nitrosomorpholine and S. typhimurium TA100 in saline was irradiated with near-ultraviolet light, mutagenesis of the bacteria took place. The same observation was made with S. typhimurium TA1535, E. coli WP2 uvrA, pKM101 and uvrA/pKM101. Several other nitrosamines showed ed the same, but weaker, effect. Evidence is presented to indicate that the mutagenicity arises from the cellular phosphate-mediated photochemical formation of direct-acting mutagen from the nitrosamine.

Activation of promutagenic N-nitrosomorpholine and other N-nitrosoalkylamines by near-ultraviolet irradiation in the presence of phosphates.

When a neutral solution of N-nitrosomorpholine, N-nitrosopyrrolidine, or N-nitrosopiperidine was irradiated with near-ultraviolet light, the solution became mutagenic to Salmonella typhimurium TA100 in the absence of metabolic activation. The formation of the active compounds required the presence of phosphate or its esters such as adenosine 5'-triphosphate during the irradiation.

[Ultraviolet-induced effect of N-nitrosamines on the in vitro parameters of DNA fusion]. / Indutsirovannoe ul'trafioletom vliianie N-nitrozaminov na parametry plavleniia DNK in vitro.

The results of studies have shown the UV-induced decrease of melting temperatures of the DNA of E. coli and chick erythrocytes under the influence of simple N-nitrosamines (NDMA, NDEA, NDPA). Either UV or nitrosoamines separately failed to effect the DNA, or their action was insignificant. It is suggested that this effect may be partly due to the action of UV on DNA.

Detection and determination of N-nitrosamines by thin-layer chromatography using fluorescamine.

A novel procedure is described for the detection and determination of N-nitrosamines (NAs) on thin-layer chromatographic plates. Ultraviolet irradiation of NAs on activated plates (silica gel or aluminum oxide) yields primary or secondary amines, which after spraying with fluorescamine reagent give fluorescent or non-fluorescent products, respectively. The majority of the 24 NAs examined afforded NAs, although the volatile dimethyl-, diethyl- and pyrrolidine-N-nitrosamines gave limits of 500, 40, and 40 ng, respectively. Spectrophotometric determinations of the relative fluorescence of 0.1-40 nmoles of NAs gave rise to nearly linear calibration curves when plotted on a log-log scale.