Size of TiO2 nanoparticles influences their phototoxicity: an in vitro investigation

To uncover the size influence of TiO₂ nanoparticles on their potential toxicity, the cytotoxicity of different-sized TiO₂ nanoparticles with and without photoactivation was tested. It was demonstrated that without photoactivation, TiO₂ nanoparticles were inert up to 100 μg/ml. On the contrary, with photoactivation, the toxicity of TiO₂ nanoparticles significantly increased, which correlated well with the specific surface area of the particles. Our results also suggest that the generation of hydroxyl radicals and reactive oxygen species (ROS)-mediated damage to the surface-adsorbed biomolecules could be the two major reasons for the cytotoxicity of TiO₂ nanoparticles after photoactivation. Higher ROS generation from smaller particles was detected under both biotic and abiotic conditions. Smaller particles could adsorb more proteins, which was confirmed by thermogravimetric analysis. To further investigate the influence of the generation of hydroxyl radicals and adsorption of protein, poly (ethylene-alt-maleic anhydride) (PEMA) and chitosan were used to coat TiO₂ nanoparticles. The results confirmed that surface coating of TiO₂ nanoparticles could reduce such toxicity after photoactivation, by hindering adsorption of biomolecules and generation of hydroxyl radical (·OH) during photoactivation.     

Identification of the thiazide diuretic drugs

A method for detecting and identifying ten thiazide diuretic drugs in tablets, gastric washings and urine by ultraviolet spectrophotometry and paper chromatography is described. The thiazides can be detected by absorption maxima in the region 264 to 294mμ at concentrations of 5 μg/ml or lower. Identification is by ascending high temperature reverse phase paper chromatography using tributyrin treated paper and developing for 20 min at 90° with a phosphate buffer (pH 7614). Further differentiation is obtained with the solvent system amyl alcohol—0 880 ammonia (9:1) on Whatman No. 1 paper. The thiazides are located as absorbing or fluorescent spots in ultraviolet light (2534 Å) and these are confirmed by the stable red colour given by an alkaline sodium 1,2-naphthaquinone-4-sulphonate spray reagent. Many of these diuretics are co-extracted with barbiturates and may interfere with barbiturate determinations.     

噻嗪类利尿剂治疗高血压的定位

<正>噻嗪类利尿剂包括噻嗪类及噻嗪类似物,于1956年问世后[1],与利血平、肼屈嗪、地巴唑等成为当时高血压治疗的主要药物。数10年来,随着众多新药诞生,许多传统降压药物或因疗效、脏器保护缺陷,或因不良反应突出而日渐衰退,而噻嗪类利尿剂尽管因某些不良反应存在争议,但至今仍在高血压治疗中占有重要的基础地位。     

吡非尼酮灌胃小鼠的急性光毒性

目的:研究一种新的抗纤维化试验药物吡非尼酮(PFD)灌胃(ig)小鼠的急性光毒性反应。方法:设小鼠igPFD100、200、400mg·kg-1·d-13个剂量组;2个溶媒对照组:0.5%羧甲基纤维素钠(CMC-Na)溶液对照组,生理盐水对照组;盐酸氯丙嗪注射液30mg·kg-1·d-1阳性对照组、空白对照组共7个组,每组10只小鼠,连续给药7d后,于第7天给药15min后,用黑光灯(220V,40W,波长320~450nm,峰值360nm,滤去致正常组织红斑的290~340nm波长)连续照射24h,观察小鼠igPFD后两耳有无红斑、水肿、皮肤瘙痒和色素沉着甚至局部坏死、溃烂、表皮脱落等光毒反应,共严密观察7d。结果:阳性对照组小鼠双耳出现中度充血、水肿等反应,3d后症状消失。PFD各剂量组和2个溶媒对照组、空白对照组小鼠在长波紫外线照射后7d内均未出现急性光毒反应。结论:连续7digPFD小鼠未出现急性光毒反应。     

Photodegradation and in vitro phototoxicity of aceclofenac

Aceclofenac (Airtal) (1) is a photoallergic and phototoxic anti-inflammatory and analgesic agent. This drug is photolabile under aerobic and anaerobic conditions. Irradiation of an ethanol-solution of aceclofenac under oxygen or argon at 290-320 nm affords via decarboxlation compound 2 as the main isolated and spectroscopically identified photoproduct, besides hydroxylamine derivates 3 and 4. A radical intermediate was evidenced spectrophotometrically with GSH and DTNB, as well as by the dimerization of cysteine. Red blood cell lysis photosensitized by 1-4 was investigated. Furthermore, in a lipid-photoperoxidation test with linoleic acid the in vitro phototoxicity of aceclofenac was also verified. The photoinduced generation of peroxide by compound 1 was determined during the irradiation in presence of NADPH by chemiluminescence. In relation to the photoallergic activity of this drug, the interaction of aceclofenac with human serum albumin (HSA) has been studied through fluorescence spectroscopy. No photoinduced binding was observed after irradiation of compounds 1 in the presence of human serum albumin.     

Comparison of an in vitro cellular phototoxicity model against controlled clinical trials of fluoroquinolone skin phototoxicity.

Many therapeutic drugs induce phototoxic skin responses following exposure to solar or artificial ultraviolet radiation sources. Several in vitro model systems have been developed to predict drug phototoxicity but none have been conducted in parallel with controlled clinical phototoxicity studies on systemically administered pharmaceuticals. The in vitro phototoxicity of eight fluoroquinolone (FQ) antibiotics (ciprofloxacin, grepafloxacin, lomefloxacin, norfloxacin, ofloxacin, trovafloxacin, BAYy3118, moxifloxacin) was determined by exposing Chinese hamster fibroblasts to UVA radiation. Cell damage was quantified with standard MTT or neutral red assays and an in vitro phototoxic index calculated (PI(vit)=% cell viability with UVA alone /% cell viability with UVA+FQ) for each endpoint. Clinical photosensitizing ability of the eight systemically administered FQ was investigated using double-blind, placebo and positive controlled, clinical skin phototesting of normal subjects. Minimal erythema doses at 365+/-30nm were determined before and after 6-7 days of FQ ingestion and PI(clin) (minimal erythema dose without FQ/minimal erythema dose with FQ) calculated. Linear regression analysis of PI(vit) vs PI(clin) gave correlations of up to 0.893. Principal components analysis of PI(vit), daily dose, plasma levels and photophysical (absorption) properties of the eight FQ showed that phototoxic (arbitrarily defined as PI(clin)> or =2) and non-phototoxic (PI(clin)<2) FQ could be completely discriminated using these parameters, and that the in vitro models were able to rank the relative phototoxic potential of the eight FQ.     

Quantitative assessment of phototoxicity in the skin of hairless mice

For determining threshold levels of inflammation resulting from a phototoxic regimen, measurement of ear thickness with a hand-held micrometer was compared with assessment by eye of the reaction on the dorsa of hairless mice. The ear-measurement technique was less sensitive with respect to the minimum concentration of 8-methoxypsoralen at which phototoxicity could be detected. However this technique was more quantifiable and more objective than detection of threshold responses by eye. Changes in ear thickness were detectable over an extended period of time with a maximum increase at 16 days; return to normal thickness occurred by 35 days. Compressibility of affected ears changed during the course of measurement, but repetitive measurement of the ears did not alter the development or resolution of the inflammatory response.     

Studies on the photostability and in vitro phototoxicity of Labetalol

The purpose of this study was to obtain information on the photochemical and phototoxic properties of Labetalol, a beta-blocker drug. Preliminary information on the drug photoreactivity was achieved using a flow system with a photochemical reactor on-line with a diode array detection system. Photophysical and photochemical investigations on the drug were performed in aqueous solutions at different pH values using spectrophotometric and fluorimetric methods; the photodegradation quantum yield was found to be 2.7×10⁻³ at pH 5.8 and 1.5×10⁻² at pH 11.5. Forced photodegradation of labetalol solutions under exposure to UVA–UVB radiations (xenon arc lamp) was monitored by reversed-phase liquid chromatography. The main photodegradation products were isolated and characterized by NMR and mass spectrometry; labetalol was found to give 3-amino-1-phenylbutane and salicylamide-4-carboxaldehyde as the main photoproducts. Preliminary phototoxic testings on human keratinocyte cultures were performed evaluating the viability of the cells by the neutral-red uptake assay; mutagenic and photomutagenicity tests were also carried out based on Salmonella typhimurium strains. As a result, labetalol was found to be photolabile,mainly in alkaline medium, but evidences of significant phototoxic and photomutagenic effects by the drug were not observed.     

Titanium dioxide nanoparticles produce phototoxicity in the developing zebrafish

Exposure of humans and other organisms to nanomaterials is increasing exponentially. It is important, but difficult, to predict the biological consequences of these exposures. We hypothesized that the unique chemical properties that make nanoparticles useful might also be the key in predicting their biological impact. To investigate this, we chose titanium dioxide nanoparticles (TiO(2)NPs) and developing zebrafish embryos as model systems. TiO(2)NPs absorb photons to generate electron-hole pairs that react with water and oxygen to form cytotoxic reactive oxygen species (ROS). Here, we show that the exposure of zebrafish embryos to TiO(2)NPs produces malformation and death, but only if the fish are also illuminated. TiO(2)NPs are taken up into the developing fish, but the egg chorion is a barrier to uptake until the embryos hatch. Chemical probes and a transgenic reporter line confirm photo-dependent production of ROS in vivo, and the addition of an ROS scavenger rescues fish embryos from toxicity. To our knowledge, this is the first study to show a photo-dependent toxic response in a whole organism from exposure to TiO(2)NPs. Of further significance, our study highlights the relationship between the property of the material that makes it useful and the biological effect that is produced. This concept should serve as a guide for future nanotoxicological studies aiming to identify potential hazardous effects on organisms.     

噻嗪类利尿药在高血压患者中的应用

利尿药是治疗高血压的重要药物,临床使用已经>50年,但目前临床使用中仍存在许多争议与问题。本文对噻嗪类利尿药治疗高血压的作用机制、药物分型与药代动力学、治疗高血压的循证证据、噻嗪类利尿药治疗高血压的临床地位和指南与共识推荐等进行综述。     

Studies on the in vitro phototoxicity of the antidiabetes drug glipizide

The phototoxic antidiabetes drug glipizide (1) is photolabile under aerobic conditions and UV-B light. Irradiation of a phosphate-buffered solution of 1 under oxygen atmosphere produces 4 photoproducts as well as singlet oxygen, which was detected by trapping it with 2,5-dimethylfuran and by the histidine test. The photochemistry of 1 involves cleavage of the sulfonamine and the sulfonamine-R bonds. Red blood cell lysis, photosensitized by glipizide and the products of its aerobic photolysis were demonstrated. The photohemolysis rate was lower for 1 than for its photoproducts. Inhibition of this process on addition of 1, 4-diazabicyclo[2.2.2]octane (DABCO), reduced glutathione (GSH), Vitamin C, sodium azide, superoxide dismutase, and a-tocopherol confirmed the possibility of singlet oxygen, superoxide ion and free radicals participation. Furthermore, in a lipid-photoperoxidation test with linoleic acid the in vitro phototoxicity of glipizide was also verified. A low decreasing cell viability of lymphocytes and neutrophils was observed.     

Vandetanib-induced phototoxicity in human keratinocytes NCTC-2544

The phototoxicity of the new anticancer drug vandetanib was evaluated using human keratinocyte cell line, NCTC-2544. This study was started since many clinical cases of vandetanib photosensitizing reactions were recently reported in literature. Vandetanib induces a clear drop in human keratinocytes viability after cell irradiation in concentration and UV-A dose dependent mode. Since vandetanib can photolyze with the formation of two main photoproducts after UV-A exposure, the contribution of these new species was also evaluated. These two photoproducts did not have a main role in the phototoxicity of their parent drug. In our opinion, the main hypothesis for the vandetanib phototoxic potential is the formation of a very reactive specie, such as an aryl radical, which can react promptly with different targets inside the cells. In fact, a massive DNA photodamage was detected both in the in vitro DNA photocleavage experiments, and in cells. Moreover, vandetanib was able to photoinduce lipid peroxidation and protein oxidations. Vandetanib photoinduced cell death by apoptosis with the involvement of mitochondria and lysosomes.     

Stability of selected chlorinated thiazide diuretics

In sports, diuretics are used for two main reasons: to flush previously taken prohibited substances with forced diuresis and in sports where weight classes are involved to achieve acute weight loss. A common property observed for thiazides is hydrolysis in aqueous media resulting in the formation of the degradation product aminobenzenedisulphonamide. This degradation product can be observed for several thiazides. Because there is limited information regarding the effect of pH, temperature and light on the stability of thiazides, these parameters were investigated for chlorothiaizide, hydrochlorothiazide and altizide. For all three compounds the degradation product could be detected after incubation at pH 9.5 for 48 h at 60 °C. At lower pH and temperature the degradation product could not be detected for all compounds. When samples were exposed to UV-light altizide and hydrochlorothiazide were photodegraded to chlorothiazide. When the degradation rate between the different compounds was compared for a given temperature and pH, altizide is the most unstable compound. This study confirms that thiazide degradation products can be formed in urine during transport. Hence doping control laboratories shall include them into their routine testing methods as required by WADA.     

Intravitreal administration of known phototoxicants in the rabbit fails to produce phototoxicity: implications for phototoxicity testing of intravitreally administered small molecule therapeutics

Context: Intravitreal (ITV) dosing has become a clinically important route of administration for the treatment of uveitis, endophthalmitis, retinal vein occlusion, diabetic macular edema and age-related macular degeneration. Despite this, there are no validated non-clinical models of phototoxicity for ITV products.

Objective: The objective of this study was to develop an ITV rabbit model of phototoxicity for use in assessing the photosafety of small molecules therapeutics.

Materials and methods: Dutch Belted rabbits were intravitreally injected bilaterally with four known phototoxicants: 8-methoxypsoralen, lomefloxacin, doxycycline and stannsoporfrin. Triescence®, a non-phototoxic triamcinolone acetonide steroid formulation designed for ITV administration, was used as a negative control. One eye was then irradiated with solar-simulated ultraviolet radiation for 30?min, 1?h after dosing, while the other eye was occluded, serving as a non-irradiated control.

Results: Despite the direct administration of known phototoxicants into the vitreous, no evidence of ocular phototoxicity was observed in any dose group. Direct (non-phototoxic) retinal toxicity was observed in the doxycycline dose group only.

Conclusion: These data suggest that the posterior segment of the rabbit eye is protected against phototoxicity by anatomical and/or physiological mechanisms, and is not a useful model for the assessment of phototoxicity of intravitreally administered molecules.     

Sympatho-adrenal mechanisms and the antihypertensive response to thiazide diuretics

Abstract: Thazide diuretics lower the blood pressure during long-term treatment by reducing the peripheral vascular resistance. This seems to be related to the natriuresis rather than to a direct vasodilating effect of the thiazides. There is evidence that the volume depletion caused by thiazide treatment initially triggers an increase in sympathetic nerve activity, which may account for initial increases in vascular resistance. Although both responses subside during long-term treatment, there is no evidence showing a causal relationship between these changes. The long-term hemodynamic adaptation to thiazide treatment may be related to altered cardiovascular reflexes. Changes in sympathetic nerve activity and reduced vascular sensitivity to noradrenaline may contribute to the adaptation. An analysis of the relative importance of these mechanisms requires further studies comparing the time courses for various effects in respondcrs and non-responders to the treament.     

Evaluation of the skin phototoxicity of systemically administered pharmaceuticals in Sprague-Dawley rats

探讨3种光毒性条件下细胞内p38和Erk1/2的表达变化,同时对3种检测方法进行比较,并深入研究其分子机制。

使用3种测试方法评估氯丙嗪(chlorpromazine,CPZ)和噻洛芬酸(tiaprofenic acid,TA)的光毒性。随后收集3种测试方法所涉及的生物样本的全细胞裂解物,并利用蛋白质印迹法评估细胞内p38和Erk1/2的表达变化。

这3种测试方法均能准确判断CPZ和TA作为光毒性阳性化合物。在3T3 NRU光毒性试验中,与空白组相比,TA和CPZ在光毒性剂量下显著提高了p38的表达水平(P<0.05),这一现象在其他测试方法中未观察到。在豚鼠和人工皮肤模型的光毒性试验中,p38和Erk1/2表达模式的变化相似。

p38和Erk1/2在3T3 NRU光毒性试验中具有明显的剂量依赖性和高度敏感性,可被视为评估3T3 NRU光毒性的潜在分子标志物,但在豚鼠试验和EpiKutis人工皮肤试验中尚未可行。豚鼠和EpiKutis人工皮肤表现出更相似的p38和Erk1/2表达变化,提示EpiKutis人工皮肤模型试验方法可能比3T3 NRU方法能更好地替代动物试验。

     

An evaluation of chemical photoreactivity and the relationship to phototoxicity

The existing regulatory guidance for photosafety testing of new drug products states that studies are warranted for those chemicals that both absorb light in the range of 290–700 nm, and that are either applied locally/topically, or “reach” (EMEA)/“significantly partition” (FDA) to the skin or eyes. The initial in vitro study recommended for the assessment of phototoxic potential is the 3T3 Neutral Red Uptake (NRU) Assay. The current study was undertaken to establish superior triggers for the initiation of biological photosafety testing. In this study, photophysical and photochemical parameters for 40 drug or drug-like molecules were studied. Principal Component Analysis (PCA), Partial Least Squares-Discriminant Analysis (PLS-DA), and a fivefold cross-validation PLS algorithm were used to evaluate the relationship between subsets of photophysical and photochemical parameters with the 3T3 NRU PIF/MPE (Photo Irritation Factor/Mean Photo Effect) results. The parameters most indicative of a 3T3 NRU positive PIF or MPE score were the extent of degradation in solution, the quantum yield of formation of singlet oxygen and the relative formation of superoxide anion. The results demonstrate that while absorption of light is critical to the induction of a light-induced process, it is the resultant events that may be used to predict the 3T3 NRU assay result. It is therefore proposed that the trigger for photosafety testing be revised to include a molecular basis for photoreactivity. From this limited investigation, estimated thresholds leading to 3T3 NRU positive results due to photodegradation, formation of singlet oxygen quantum yield or a relative superoxide anion formation value are proposed.