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The BBB is a protective entity that prevents external substances from reaching the CNS but it also hinders the delivery of drugs into the brain when they are needed. The main objective of this work was to improve a previously proposed in vitro cell-based model by using a more physiological cell line (hCMEC/D3) to predict the main pharmacokinetic parameters that describe the access and distribution of drugs in the CNS: Kpuu,brain, fu,plasma, fu,brain and Vu,brain. The hCMEC/D3 permeability of seven drugs was studied in transwell systems under different conditions (standard, modified with albumin and modified with brain homogenate). From the permeability coefficients of those experiments, the parameters mentioned above were calculated and four linear IVIVCs were established. The best ones were those that relate the in vitro and in vivo Vu,brain and fu,brain (r2 = 0.961 and r2 = 0.940) which represent the binding rate of a substance to the brain tissue, evidencing the importance of using brain homogenate to mimic brain tissue when an in vitro brain permeability assay is done. This methodology could be a high-throughput screening tool in drug development to select the CNS promising drugs in three different in vitro BBB models (hCMEC/D3, MDCK and MDCK-MDR1).

Dysphagia is a very common problem suffered by elderly patients. The use of thickeners during administration in these patients helps to prevent difficulties with swallowing larger solid dosage forms. However, there are several indications when the thickeners may influence disintegration and dissolution processes of solid dosage forms, potentially affecting therapeutic efficacy. In this paper the effects of a commonly used thickener on tablet disintegration, dissolution and subsequent absorption of 6 formulated drugs frequently used in elderly patients (Aspirin, Atenolol, Acenocumarol, Candesartan, Ramipril and Valsartan) in two different administration conditions (intact tablet and crushed tablet) are reported. Disintegration times were determined using a modified disintegration test device. The presence of thickener leads to a pseudoplastic behavior with clearly increased viscosity values. The thickener was also shown to significantly affect the release processes (dissolution and disintegration), but not the permeability of the studied drugs. When tablets are crushed the effect of the thickener on drug dissolution is avoided. Consequently, crushing the tablets would be a recommendation for these drugs if the use of a thickener is necessary in patients with dysphagia.

The aim of the present paper is to study the effect of common excipients on the permeability of atenolol (as drug absorbed mainly by passive diffusion) and rhodamine (as P-glycoprotein substrate). The apparent permeability was measured by an in situ perfusion method in Wistar rats using the closed loop Doluisio's method. Permeability values were characterized in the absence and presence of 18 commonly used excipients. Excipient concentrations were selected based on the amounts in oral immediate release dosage forms, which failed the test during the human bioequivalence studies. Atenolol was studied with and without excipients in the whole small intestine, whereas rhodamine was tested in three different intestinal segments to account for the differential expression of P-glycoprotein, and it was further on tested in the ileum, in the presence of excipients. Atenolol presented higher permeability values when it was administered with colloidal silica, croscarmellose, hydroxypropyl methylcellulose (HPMC), magnesium stearate, MgCO3, poly(ethylene glycol) 400, poly(vinylpyrrolidone), sorbitol, starch, and TiO2 rhodamine showed higher permeability values when it was administered with croscarmellose and HPMC. On the one hand, the mechanisms of action were not discernible with the proposed experiments. On the other hand, commercial formulations do not present a single excipient but several, which can counteract their effects. The in situ perfusion technique can be useful for a preliminary screening and risk analysis, while the in vivo pharmacokinetic results would be needed to define conclusive effects.

Controlled release (CR) dosage forms comprise a wide range of technologies, which modify the drug pharmacokinetic (PK) profile by avoiding the immediate release (IR) of the active pharmaceutical ingredient (API). They are particularly of interest in chronic diseases, for narrow therapeutic index drugs or for targeting a particular gastrointestinal tract (GI) segment.Diffusion and dissolution limited controlled release systems are described in terms of release kinetics and formulation strategies with e xamples marketed or under development. Additionally, the physiological variables affecting the release (such as fluid pH, volume and composition, physical forces, and transit times) and the in vitro dissolution techniques currently available are reviewed.Selection of the appropriate release mechanism is not a straightforward process, as it requires a balance based on the desired target, the API properties and the technological challenges of the dosage form structure. Diffusion, dissolution or a combination of both could be adequate without an absolute superiority of one mechanism over the other. The combination of in vivo predictive dissolution systems, with mathematical modeling of the release mechanism and its correlation with formulation composition could help to design prototype candidates, with enhanced probabilities of success in human clinical trials.

One of the main obstacles in neurological disease treatment is the presence of the blood–brain barrier. New predictive high-throughput screening tools are essential to avoid costly failures in the advanced phases of development and to contribute to the 3 Rs policy. The objective of this work was to jointly develop a new in vitro system coupled with a physiological-based pharmacokinetic (PBPK) model able to predict brain concentration levels of different drugs in rats. Data from in vitro tests with three different cells lines (MDCK, MDCK-MDR1 and hCMEC/D3) were used together with PK parameters and three scaling factors for adjusting the model predictions to the brain and plasma profiles of six model drugs. Later, preliminary quantitative structure–property relationships (QSPRs) were constructed between the scaling factors and the lipophilicity of drugs. The predictability of the model was evaluated by internal validation. It was concluded that the PBPK model, incorporating the barrier resistance to transport, the disposition within the brain and the drug–brain binding combined with MDCK data, provided the best predictions for passive diffusion and carrier-mediated transported drugs, while in the other cell lines, active transport influence can bias predictions.

Mesoporous silica microparticles functionalized with lactose for the specific release of essential oil components (EOCs) in the small intestine are presented. In vitro and in vivo intestinal models were applied to validate the microparticles (M41-EOC-L), in which the presence of lactase acts as the triggering stimulus for the controlled release of EOCs. Among the different microdevices prepared (containing thymol, eugenol and cinnamaldehyde), the one loaded with cinnamaldehyde showed the most significant Caco-2 cell viability reduction. On the other hand, interaction of the particles with enterocyte-like monolayers showed a reduction of EOCs permeability when protected into the designed microdevices. Then, a microdevice loaded with cinnamaldehyde was applied in the in vivo model of Wistar rat. The results showed a reduction in cinnamaldehyde plasma levels and an increase in its concentration in the lumen of the gastrointestinal tract (GIT). The absence of payload release in the stomach, the progressive release throughout the intestine and the prolonged stay of the payload in the GIT-lumen increased the bioavailability of the encapsulated compound at the site of the desired action. These innovative results, based on the specific intestinal controlled delivery, suggest that the M41-payload-L could be a potential hybrid microdevice for the protection and administration of bioactive molecules in the small intestine and colon., Ministerio de Ciencia, Innovación y Universidades (Spanish Government) )the Agencia Estatal de Investigación (AEI) and European Union (projects RTI2018-100910-B-C41 and RTI2018-101599-B-C22-AR (MCIU/AEI/ FEDER, EU)), Ministerio de Universidades (Spanish Government) (BG20/00020, A.B. Beatriz Galindo contract), Agencia Estatal de Investigación and European Union through FEDER (Fondo Europeo de Desarrollo Regional, AEI/FEDER EU, project SAF2016-78756), Conselleria de Innovación, Universidades, Ciencia y Sociedad Digital, Generalitat Valenciana (project PROMETEO 2018/024 and E.P-R. predoctoral grant ACIF/2016/023)

Excipients are necessary to develop oral dosage forms of any Active Pharmaceutical Ingredient (API). Traditionally, excipients have been considered inactive and inert substances, but, over the years, numerous studies have contradicted this belief. This review focuses on the effect of excipients on the physiological variables affecting oral absorption along the different segments of the gastrointestinal tract. The effect of excipients on the segmental absorption variables are illustrated with examples to help understand the complexity of predicting their in vivo effects.The effects of excipients on disintegration, solubility and dissolution, transit time, and absorption are analyzed in the context of the different gastrointestinal segments and the physiological factors affecting release and membrane permeation. The experimental techniques used to study excipient effects and their human predictive ability are reviewed.The observed effects of excipient in oral absorption process have been characterized in the past, mainly

[EN] The main objective of this investigation was to develop an in vitro-in vivo correlation (IVIVC) for immediate release candesartan cilexetil formulations by designing an in vitro dissolution test to be used as development tool. The IVIVC could be used to reduce failures in future bioequivalence studies. Data from two bioequivalence studies were scaled and combined to obtain the dataset for the IVIVC. Two-step and one-step approaches were used to develop the IVIVC. Experimental solubility and permeability data confirmed candesartan cilexetil. Biopharmaceutic Classification System (BCS) class II candesartan average plasma profiles were deconvoluted by the Loo-Riegelman method to obtain the oral fractions absorbed. Fractions dissolved were obtained in several conditions in USP II and IV apparatus and the results were compared calculating the f(2)similarity factor. Levy plot was constructed to estimate the time scaling factor and to make both processes, dissolution and absorption, superimposable. The in vitro dissolution experiment that reflected more accurately the in vivo behavior of the products of candesartan cilexetil employed the USP IV apparatus and a three-step pH buffer change, from 1.2 to 4.5 and 6.8, with 0.2% of Tween 20. This new model was able to predict the in vivo differences in dissolution and it could be used as a risk-analysis tool for formulation selection in future bioequivalence trials., This research was funded by the Agencia Estatal de Investigacion and the European Union, through FEDER (Fondo Europeo de Desarrollo Regional), grant number SAF2016-78756 (AEI/FEDER, EU). Barbara Sanchez-Dengra received a grant from the Ministry of Science, Innovation and Universities of Spain, grant number FPU17/00530.

[EN] In recent decades, the versatility of mesoporous silica particles and their relevance to develop controlled release systems have been demonstrated. Within them, gated materials able to modulate payload delivery represent great advantages. However, the role played by the porous matrix in this kind of systems is scarce. In this work, different mesoporous silica materials (MCM-41, MCM-48, SBA-15 and UVM-7) are functionalized with oleic acid as a molecular gate. All systems are fully characterized and their ability to confine the entrapped cargo and release it in the presence of bile salts is validated with release assays and in vitro digestion experiments. The cargo release profile of each synthesized support is studied, paying attention to the inorganic scaffold. Obtained release profiles fit to Korsmeyer-Peppas model, which explains the differences among the studied supports. Based on the results, UVM-7 material was the most appropriate system for duodenal delivery and was tested in an in vivo model of the Wistar rat. Payload confinement and its complete release after gastric emptying is achieved, establishing the possible use of mesoporous silica particles as protection and direct release agents into the duodenum and, hence, demonstrating that these systems could serve as an alternative to the administration methods employed until now., This research was funded by the Spanish Government (projects RTI2018-100910-B-C41 and RTI2018-101599-B-C22-AR (MCUI/FEDER, EU)) and the Generalitat Valenciana (project PROMETEO/2018/024 and ACIF/2016/023 grant).

The purpose of this work is to explore the predictive ability of the biopharmaceutics classification system (BCS) biowaiver based on the dissolution methods for two pravastatin test products, where one of them showed bioequivalence (BE) while the other test failed (non-bioequivalence, or NBE), and to explore the reasons for the BE failure. Experimental solubility and permeability data confirmed that pravastatin is a BCS class III compound. The permeability experiments confirmed that the NBE formulation significantly increased pravastatin permeability, and could explain its higher absorption rate and higher Cmax. This finding highlights the relevance of requiring similar excipients for BCS class III drugs. The BCS-based biowaiver dissolution tests at pH 1.2, 4.5, and 6.8, with the paddle apparatus at 50 rpm in 900 mL media, were not able to detect differences in pravastatin products, although the NBE formulation exhibited a more rapid dissolution at earlier sampling times. Dissolution tests conducted in 500 mL did not achieve complete dissolution, and both formulations were dissimilar because the amount dissolved at 15 min was less than 85%. The difference was less than 10% at pH 1.2 and 4.5, while at pH 6.8 f2, results reflected the Cmax rank order.

[EN] Colon targeted drug delivery is highly relevant not only to treat colonic local diseases but also for systemic therapies. Mesoporous silica nanoparticles (MSNs) have been demonstrated as useful systems for controlled drug release given their biocompatibility and the possibility of designing gated systems able to release cargo only upon the presence of certain stimuli. We report herein the preparation of three gated MSNs able to deliver their cargo triggered by different stimuli (redox ambient (S1), enzymatic hydrolysis (S2), and a surfactant or being in contact with cell membrane (S3)) and their performance in solution and in vitro with Caco-2 cells. Safranin O dye was used as a model drug to track cargo fate. Studies of cargo permeability in Caco-2 monolayers demonstrated that intracellular safranin O levels were significantly higher in Caco-2 monolayers when using MSNs compared to those of free dye. Internalization assays indicated that S2 nanoparticles were taken up by cells via endocytosis. S2 nanoparticles were selected for in vivo tests in rats. For in vivo assays, capsules were filled with S2 nanoparticles and coated with Eudragit FS 30 D to target colon. The enteric coated capsule containing the MSNs was able to deliver S2 nanoparticles in colon tissue (first step), and then nanoparticles were able to deliver safranin O inside the colonic cells after the enzymatic stimuli (second step). This resulted in high levels of safranin O in colonic tissue combined with low dye levels in plasma and body tissues. The results suggested that this combination of enzyme-responsive gated MSNs and enteric coated capsules may improve the absorption of drugs in colon to treat local diseases with a reduction of systemic effects., The authors acknowledge the financial support from the Spanish Government (Projects MAT2015-64139-C4-1-R, SAF2016-78756 and AGL2015-70235-C2-2-R) and the Generalitat Valenciana (Project GVA/2014/13).

The purpose of this study was to predict in vivo performance of three oral products of Etoricoxib (Arcoxia® as reference and two generic formulations in development) by conducting in vivo predictive dissolution with GIS (Gastro Intestinal Simulator) and computational analysis. Those predictions were compared with the results from previous bioequivalence (BE) human studies. Product dissolution studies were performed using a computer-controlled multicompartmental dissolution device (GIS) equipped with three dissolution chambers, representing stomach, duodenum, and jejunum, with integrated transit times and secretion rates. The measured dissolved amounts were modelled in each compartment with a set of differential equations representing transit, dissolution, and precipitation processes. The observed drug concentration by in vitro dissolution studies were directly convoluted with permeability and disposition parameters from literature to generate the predicted plasma concentrations. The GIS was able to detect the dissolution differences among reference and generic formulations in the gastric chamber where the drug solubility is high (pH 2) while the USP 2 standard dissolution test at pH 2 did not show any difference. Therefore, the current study confirms the importance of multicompartmental dissolution testing for weak bases as observed for other case examples but also the impact of excipients on duodenal and jejunal in vivo behavior.

[EN] Silica mesoporous microparticles loaded with both rhodamine B fluorophore (S1) or hydrocortisone (S2), and capped with an olsalazine derivative, are prepared and fully characterized. Suspensions of Si and S2 in water at an acidic and a neutral pH show negligible dye/drug release, yet a notable delivery took place when the reducing agent sodium dithionite is added because of hydrolysis of an azo bond in the capping ensemble. Additionally, olsalazine fragmentation induced 5-aminosalicylic acid (5-ASA) release. In vitro digestion models show that S1 and S2 solids are suitable systems to specifically release a pharmaceutical agent in the colon. In vivo pharmacokinetic studies in rats show a preferential rhodamine B release from Si in the colon. Moreover, a model of ulcerative colitis is induced in rats by oral administration of 2,4,6-trinitrobenzenesulfonic acid (TNBS) solutions, which was also used to prove the efficacy of S2 for colitis treatment. The specific delivery of hydrocortisone and 5-ASA from S2 material to the colon tissue in injured rats markedly lowers the colon/body weight ratio and the clinical activity score. Histological studies showed a remarkable reduction in inflammation, as well as an intensive regeneration of the affected tissues., We thank the Generalitat Valenciana (Project PROMETE02018/024) and the Spanish Government (Projects AGL2015-70235-C2-2-R and MAT2015-64139-C4-1-R (MINECO/FEDER)) for support. A.H.T. thanks the Spanish MEC for his FPU fellowship. The authors also thank the support of the Electron Microscopy Service at the UPV. The SCSIE (of the Universitat de Valencia) is also gratefully acknowledged for all the equipment used. NMR spectra were measured at the U26 facility of ICTS "NANBIOSIS" at the Universitat de Valencia.

[EN] Hydrogels are widely studied as drug delivery system. In this work we propose the employment of tetrakis (hydroxymethyl)phosphonium chloride as crosslinking agent to obtain covalent hydrogels based on chitosan. These hydrogels are obtained by Mannich reaction between the amino groups of chitosan with the hydroxymethyl groups of the crosslinker molecule. They show a pH sensitive second order swelling kinetic, have low toxicity, are biocompatible, mucoadhesive and allow a modified release of the encapsulated drug, camptothecin, for 48 h. This antitumor drug has been studied as a drug of interest to develop oral chemotherapy administration strategies. According to the obtained results, oral administration of camptothecin through hydrogels would provide low concentrations of drug at the absorption site, avoiding carrier saturation and reducing its intestinal toxicity., Authors acknowledge partial financial support to project SAF2016-78756 from MINECO (Spanish Ministry of economy, industry and competitivity). Mayte Martinez-Martínez received a grant from the Ministry of Education and Science of Spain (FPU13-01105).

[EN] Nanohydrogels based on natural polymers, such as polysaccharides, are gaining interest as vehicles for therapeutic agents, as they can modify the pharmacokinetics and pharmacodynamics of the carried drugs. In this work, hyaluronan-riboflavin nanohydrogels were tested in vivo in healthy rats highlighting their lack of toxicity, even at high doses, and their different biodistribution with respect to that of native hyaluronan. They were also exploited as carriers of a hydrophobic model drug, the anti-inflammatory piroxicam, that was physically embedded within the nanohydrogels by an autoclave treatment. The nanoformulation was tested by intravenous administration showing an improvement of the pharmacokinetic parameters of the molecule. The obtained results indicate that hyaluronan-based self-assembled nanohydrogels are suitable systems for low-soluble drug administration, by increasing the dose as well as the circulation time of poorly available therapeutic agents., Financial support from University Sapienza Progetti di Ricerca: grant RP116154C2EF9AC8 and grant RM11715C1743EE89 are acknowledged. Isabel Gonzalez-Alvarez, Marta Gonzalez-Alvarez and Marival Bermejo acknowledge partial financial support to project SAF2016-78756 from MINECO (Spanish Ministry of economy, industry and competitivity). Mayte Martinez-Martínez received a grant from the Ministry of Education and Science of Spain (FPU13-01105).

Seven novel Copper(II) complexes, namely [Cu(Am4DHotaz)(H2O)2](ClO4) (1), [Cu(Am4DHotaz)(NO3)(MeOH)]·H2O (2), [Cu(Am4Motaz)2(H2O)](ClO4)2·0.83H2O (3), [Cu(Am4Motaz)2(NO3)]NO3·MeOH (4), [Cu(Am4Eotaz)2(NO3)]3(NO3)3·2H2O (5), [Cu(Am4Eotaz)2(ClO4)](ClO4) (6) and [Cu(Am4Eotaz)(ClO4)(H2O)](ClO4) (6a) (HAm4DHotaz = N′-(4-oxothiazolidin-2-ylidene)pyridine-2-carbohydrazonamide, Am4Motaz = N′-(3-methyl-4-oxothiazolidin-2-ylidene)pyridine-2-carbohydrazonamide and Am4Eotaz = N′-(3-ethyl-4-oxothiazolidin-2-ylidene)pyridine-2-carbohydrazonamide), have been successfully synthesized and characterized by several physicochemical techniques and, for 1–6 complexes, single crystal X-ray diffraction. Having the structural data as a base, complexes 1, 2 and 3 exhibited square pyramidal to square pyramidal slightly distorted geometry, whereas 4, 5 and 6 an intermediate between square pyramidal and trigonal bipyramidal. The ability of complexes 1–6 to cleave DNA was assayed with the aid of gel electrophoresis on supercoiled pUC18-DNA. Except for [Cu(Am4Motaz)2(H2O)](ClO4)2·0.83H2O (3), the compounds were not able to perform DNA cleavage (data not shown). Since 3 has been shown to behave as a nuclease, its interaction with DNA was studied by means of thermal denaturation and viscosimetry measurements.

Atenolol is a drug widely used for the treatment of hypertension. However, the great drawback it presents is a low bioavailability after oral administration. To obtain formulations that allow to improve the bioavailability of this drug is a challenge for the pharmaceutical technology. The objective of this work was to increase the rate and extent of intestinal absorption of atenolol as model of a low permeability drug, developing a double technology strategy. To increase atenolol permeability an ion pair with brilliant blue was designed and the sustained release achieved through encapsulation in polymeric nanoparticles (NPs). The in vitro release studies showed a pH-dependent release from NPs, (particle size 437.30 ± 8.92) with a suitable release profile of drug (atenolol) and counter ion (brilliant blue) under intestinal conditions. Moreover, with the in vivo assays, a significant increase (2-fold) of atenolol bioavailability after administering the ion-pair NPs by oral route was observed. In conclusion, the combination of ion-pair plus polymeric NPs have proved to be a simple and very useful approach to achieve a controlled release and to increase the bioavailability of a low permeability charged drugs.

The purpose of this work was to describe the closed loop in situ perfusion method in rats and to compare the difficulties and advantages with other methods proposed by regulatory agencies for BCS classification and finally to illustrate its application to evaluate the permeability of digoxin at relevant clinical concentrations. Digoxin was evaluated at two concentration levels: 1.0 μg/ml (with and without sodium azide 65.0 μg/ml) and 6.0 μg/ml. These concentrations correspond to the ratio of the highest dose strength (0.25 mg) and the highest single dose administered (1.5 mg) and the 250 ml of water. In situ closed loop perfusion studies in rats were performed in the whole small intestine and also in duodenum, jejunum and ileum segments to evaluate the relevance of P-gp secretion in the overall permeability. A kinetic modelling approach involving passive permeation and efflux transport mechanism allowed the estimation of the passive diffusional component and the Michaelis-menten parameters. The estimated Km value demonstrated that at clinical luminal concentrations the efflux process is not saturated and then it could be inhibited by other drugs, excipients or food components leading to the already reported clinical drug-drug and drug-food interations. The present data confirms from a mechanistic point of view these interactions.

Oral controlled release (CR) formulations have many benefits and have become a valuable resource for the local and systemic administration of drugs. The most important characteristic of these pharmaceutical products is that drug absorption occurs mainly in the colon. Therefore, this review analyses the physiological and physicochemical features that may affect an orally administered CR product, as well as the different strategies to develop a CR dosage form and the methods used to evaluate the formulation efficacy. The models available to study the intestinal permeability and their applicability to colonic permeability determinations are also discussed.

The purpose of this investigation was to develop an exploratory two-step level A IVIVC for three telmisartan oral immediate release formulations, the reference product Micardis, and two generic formulations (X1 and X2). Correlation was validated with a third test formulation, Y1. Experimental solubility and permeability data were obtained to confirm that telmisartan is a class II compound under the Biopharmaceutic Classification System. Bioequivalence (BE) studies plasma profiles were combined using a previously published reference scaling procedure. X2 demonstrated in vivo BE, while X1 and Y1 failed to show BE due to the lower boundary of the 90% confidence interval for Cmax being outside the acceptance limits. Average plasma profiles were deconvoluted by the Loo-Riegelman method to obtain the oral fractions absorbed ( fa). Fractions dissolved ( fdiss) were obtained in several conditions in USP II and USP IV apparatus, and later, the results were compared in order to find the most biopredictive model, calculating the f2 similarity factor. The apparatus and conditions showing the same rank order than in vivo data were selected for further refinement of conditions. A Levy plot was constructed to estimate the time scaling factor and to make both processes, dissolution and absorption, superimposable. The in vitro dissolution experiment that reflected more accurately the in vivo behavior of the different formulations of telmisartan employed the USP IV dissolution apparatus and a dissolution environment with a flow rate of 8 mL/min and a three-step pH change, from 1.2 to 4.5 and 6.8, with a 0.05% of Tween 80. Thus, these conditions gave rise to a biopredictive dissolution test. This new model is able to predict the formulation differences in dissolution that were previously observed in vivo, which could be used as a risk-analysis tool for formulation selection in future bioequivalence trials.

[EN] Magnetic mesoporous silica microparticles were loaded with safranin O (S1) and with hydrocortisone (S2) and the outer surface functionalized with a bulky azo derivative bearing urea moieties. Aqueous suspensions of both solids at pH 7.4 showed negligible payload release whereas a marked delivery was observed in the presence of sodium dithionite due to the rupture of the azo bonds. Besides, a moderate cargo release was observed at acidic pH due to the hydrolysis of the urea bonds that linked the azo derivative onto the external surface of the inorganic scaffolds. In vitro digestion models showed that S1 and S2 microparticles could be used for the controlled release of payload in the reducing colon environment (in which azoreductase enzymes are present). On the other hand, in vivo pharmacokinetic studies in rats showed that safranine O release from S1 microparticles was concentrated in colon. The performance of S2 microparticles for the treatment of colitis in rats (induced by oral administration of a 2,4,6-trinitrobenzenesulfonic acid solution) was tested. The controlled release of hydrocortisone from S2 in the colon of injured rats induced marked reduction in colon/body weight ratio and in clinical activity score. Also, histological studies showed a marked decrease in inflammation followed by intensive regeneration and almost normal mucosal structure of the individuals treated with S2. Besides, the use of a magnetic belt increased the therapeutic performances of S2 due to an enhanced retention time of the particles in the colon., We thank the Spanish Government (projects MAT2015-64139-C4-1-R and AGL2015-70235-C2-2-R (MINECO/FEDER)) and the Generalitat Valenciana (project PROMETEOII/2014/047) for support. AHT thanks to the Spanish MEC for his FPU grant. The authors also thank the Electron Microscopy Service at the Universitat Politecnica de Valencia for support. SCSIE (Universitat de Valencia) is also gratefully acknowledged for all the equipment employed. NMR was registered at the U26 facility of ICTS "NANBIOSIS" at the Universitat de Valencia. The authors thanks Dr. L. A. Villaescusa for his helpful discussion about the 1H NMR analysis of the composition of loaded and functionalized supports.

[EN] In this work, 6-phosphogluconic trisodium salt (6-PG(-)Na(+)) is introduced as a new aqueous and nontoxic cross-linking agent to obtain ionic hydrogels. Here, it is shown the formation of hydrogels based on chitosan cross-linked with 6-PG(-)Na(+). This formulation is obtained by ionic interaction of cationic groups of polymer with anionic groups of the cross linker. These hydrogels are nontoxic, do not cause dermal irritation, are easy to extend, and have an adequate adhesion force to be applied as polymeric film over the skin. This AWN formulation exhibits a first order release kinetic and can be applied as drug vehicle for topical administration or as wound dressing for wound healing. The primary goal of this communication is to report the identification and utility of 6-phosphogluconic trisodium salt (6-PG(-)Na(+)) as a nontoxic cross-linker applicable for cationic polymers., The authors acknowledge partial financial support to project SAF2016-78756 from MINECO (Spanish Ministry of economy, industry and competitiveness). Maria Teresa Martinez Martinez received a grant from the Ministry of Education and Science of Spain (FPU13-01105). The product was patented in Spain in 2016 by authors of this paper. Patent application 201631463.

[EN] Magnetic micro-sized mesoporous silica particles were used for the preparation of a gated material able to release an entrapped cargo in the presence of an azo-reducing agent and, to some extent, at acidic pH. The magnetic mesoporous microparticles were loaded with safranin O and the external surface was functionalized with an azo derivative 1 (bearing a carbamate linkage) yielding solid S1. Aqueous suspensions of S1 at pH 7.4 showed negligible safranin O release due to the presence of the bulky azo derivative attached onto the external surface of the inorganic scaffold. However, in the presence of sodium dithionite (azoreductive agent), a remarkable safranin O delivery was observed. At acidic pH, a certain safranin O release from S1 was also found. The pH-triggered safranin O delivery was ascribed to the acid-induced hydrolysis of the carbamate moiety that linked the bulky azo derivatives onto the mesoporous inorganic magnetic support. The controlled release behavior of S1 was also tested using a model that simulated the gastro intestinal tract., We thank the Spanish Government (projects MAT2015-64139-C4-1-R and AGL2015-70235C2-2-R (MINECO/FEDER)) and the Generalitat Valenciana (project PROMETEOII/2014/047) for support. AHT thanks to the Spanish MEC for his FPU grant. The authors also thank the Electron Microscopy Service at the Universitat Politecnica de Valencia for support. SCSIE (Universitat de Valencia) is also gratefully acknowledged for all the equipment employed. NMR was registered at the U26 facility of ICTS "NANBIOSIS" at the Universitat de Valencia. The authors thanks L.A. Villaescusa for his helpful discussion about the 1H-NMR analysis of the composition of loaded and functionalized supports.

The effective rat intestinal permeability (P eff ) was deconvolved using a biophysical model based on parameterized paracellular, aqueous boundary layer, transcellular permeabilities, and the villus-fold surface area expansion factor. Four types of rat intestinal perfusion data were considered: single-pass intestinal perfusion (SPIP) in the jejunum (n = 40), and colon (n = 15), closed-loop (Doluisio type) in the small intestine (n = 78), and colon (n = 74). Moreover, in vitro Caco-2 permeability values were used to predict rat in vivo values in the rat data studied. Comparable number of molecules permeate via paracellular water channels as by the lipoidal transcellular route in the SPIP method, although in the closed-loop method, the paracellular route appears dominant in the colon. The aqueous boundary layer thickness in the small intestine is comparable to that found in unstirred in vitro monolayer assays; it is thinner in the colon. The mucosal surface area in anaesthetized rats is 0.96–1.4 times the smooth cylinder calculated value in the colon, and it is 3.1–3.6 times in the small intestine. The paracellular permeability of the intestine appeared to be greater in rat than human, with the colon showing more leakiness (higher P para ) than the small intestine. Based on log intrinsic permeability values, the correlations between the in vitro and in vivo models ranged from r2 0.82 to 0.92. The SPIP-Doluisio method comparison indicated identical log permeability selectivity trend with negligible bias.

The purpose of this work is to investigate the discriminatory power of the Biopharmaceutics Classification System (BCS)-biowaiver in vitro methodology, i.e., to investigate if a BCS-biowaiver approach would have detected the Cmax differences observed between two zolpidem tablets and to identify the cause of the in vivo difference. Several dissolution conditions were tested with three zolpidem formulations: the reference (Stilnox), a bioequivalent formulation (BE), and a nonbioequivalent formulation (N-BE). Zolpidem is highly soluble at pH 1.2, 4.5, and 6.8. Its permeability in Caco-2 cells is higher than that of metoprolol and its transport mechanism is passive diffusion. None of the excipients (alone or in combination) showed any effect on permeability. All formulations dissolved more than 85% in 15 min in the paddle apparatus at 50 rpm in all dissolution media. However, at 30 rpm the nonbioequivalent formulation exhibited a slower dissolution rate. A slower gastric emptying rate was also observed in rats for the nonbioequivalent formulation. A slower disintegration and dissolution or a delay in gastric emptying might explain the Cmax infra-bioavailability for a highly permeable drug with short half-life. The BCS-biowaiver approach would have declared bioequivalence, although the in vivo study was not conclusive but detected a 14% mean difference in Cmax that precluded the bioequivalence demonstration. Nonetheless, these findings suggest that a slower dissolution rate is more discriminatory and that rotation speeds higher than 50 rpm should not be used in BCS-biowaivers, even if a coning effect occurs.

New organometallic gold(III) and platinum(II) complexes containing iminophosphorane ligands are described. Most of them are more cytotoxic to a number of human cancer cell lines than cisplatin. Cationic Pt(II) derivatives 4 and 5, which differ only in the anion, Hg2Cl6(2-) or PF6(-) respectively, display almost identical IC50 values in the sub-micromolar range (25-335-fold more active than cisplatin on these cell lines). The gold compounds induced mainly caspase-independent cell death, as previously reported for related cycloaurated compounds containing IM ligands. Cycloplatinated compounds 3, 4, and 5 can also activate alternative caspase-independent mechanisms of death. However, at short incubation times cell death seems to be mainly caspase dependent, suggesting that the main mechanism of cell death for these compounds is apoptosis. Mercury-free compound 5 does not interact with plasmid (pBR322) DNA or with calf thymus DNA. Permeability studies of 5 by two different assays, in vitro Caco-2 monolayers and a rat perfusion model, have revealed a high permeability profile for this compound (comparable to that of metoprolol or caffeine) and an estimated oral fraction absorbed of 100%, which potentially makes it a good candidate for oral administration.

The aim of the present study was to develop a method for water flux reabsorption measurement in Doluisio's Perfusion Technique based on the use of phenol red as a non-absorbable marker and to validate it by comparison with gravimetric procedure. The compounds selected for the study were metoprolol, atenolol, cimetidine and cefadroxil in order to include low, intermediate and high permeability drugs absorbed by passive diffusion and by carrier mediated mechanism. The intestinal permeabilities (P-eff) of the drugs were obtained in male and female Wistar rats and calculated using both methods of water flux correction. The absorption rate coefficients of all the assayed compounds did not show statistically significant differences between male and female rats consequently all the individual values were combined to compare between reabsorption methods. The absorption rate coefficients and permeability values did not show statistically significant differences between the two strategies of concentration correction. The apparent zero order water absorption coefficients were also similar in both correction procedures. In conclusion gravimetric and phenol red method for water reabsorption correction are accurate and interchangeable for permeability estimation in closed loop perfusion method. (C) 2014 Elsevier B.V. All rights reserved.

The oral controlled release (CR) formulations have become more important in recent years. Among them, the polymeric nanoparticles have been thoroughly studied during the last decades, consequently they are extensively employed for a broad range of applications and drugs. The objective of this research was to develop polymeric nanoparticles (NPs) of ibuprofen with poly(lactic-co-glycolic) acid (PLGA) as polymer, and to test their applicability for oral CR formulations development. Different proportions of drug/polymer were employed to develop the ibuprofen NPs and their in vitro release profiles were analysed. The in situ segmental permeability of ibuprofen was tested in Wistar rat and demonstrated the high permeability of ibuprofen in rat colon. In addition, in vivo assays were performed to study the plasma concentration-time profiles of encapsulated versus non-encapsulated ibuprofen. The results showed that ibuprofen release from the NPs was pH-dependent and consequently higher at colonic pH. Moreover, the plasma concentration-time profiles reveal a controlled release from the ibuprofen NP. Therefore, the ibuprofen PLGA-NPs will be a good CR formulation to achieve a controlled release targeted to the colon, where the release rate of the drug from the NPs will be the limiting factor for the absorption process.

Nowadays, alternative methods have been developed to predict intestinal permeability values in human as in vitro, in situ or ex vivo methods. They were developed by the necessity to avoid the problems of the human permeability experiments. However, determination of human permeability is needed to properly validate the alternative methods. For this reason, recently, Dahlgren et al. published an indirect method based on a deconvolution technique to estimate the human permeability in different gastrointestinal segments (jejunum, ileum and colon). Therefore, the objective of this research was to demonstrate that Doluisio technique is a useful method to predict the human permeability in different gastrointestinal segments. To achieve this goal, the rat permeability in different segments, of the same drugs studied by Dahlgren et al. (atenolol, metoprolol and ketoprofen), have been compared with the human data obtained by the deconvolution method. The results obtained in this work show that the Doluisio method is a reliable tool to predict segmental human permeability. Consequently, the deconvolution method can be employed to build an extensive database of human permeability, overall from ileum and colon, because there is a lack of human permeability data of these distal segments. Once there are enough human data available, the Doluisio technique will be a valuable alternative method to predict the permeability of new molecules with therapeutic activity without the requirement of human experiments.

The relevant parameters for predicting rate and extent of access across the blood-brain barrier (BBB) are fu,plasma (unbound fraction in plasma), Vu,brain (distribution volume in brain) and Kp,uu,brain (ratio of free concentrations in plasma and brain). Their estimation still requires animal studies and in vitro low throughput experiments which make difficult the screening of new CNS candidates. The aim of the present work was to develop a new whole in vitro high throughput method to predict drug rate and extent of access across the BBB. The system permits estimation of fu,plasma, Vu,brain and Kp,uu,brain in a single experimental system, using in vitro cell monolayers in different conditions. From the ratios of the apparent permeability values (Papp) with the adequate mathematical analysis the relevant parameters can be estimated. Papp of ten model compounds has been obtained in MDCKII and MDCK-Mdr1cell monolayers in the absence and presence of albumin and brain homogenate. The ratio of Papp in the absence and presence of albumin allows estimation of in vitro fu,plasma. Papp in the presence of brain homogenate is used to estimate fu,brain and Vu,brain. Kp,uu,brain is estimated from the apical to basal versus basal to apical clearances. The BBB parameters obtained with the new method were predictive of the in vivo behavior of candidates. In vitro fu,plasma, Kp,uu,brain and Vu,brain (calculated with Papp from MDCKII cell line) presented a good correlation with in vivo fu,plasma, Kp,uu,CSF and Vu,brain published values (r=0.92; r=0.85; and r=0.99 respectively). Despite its simplicity the predictive performance is fairly good considering the reduced number of tested compounds with different physicochemical and transport properties. Further experimental modifications could be checked to optimize the method, but the present data support its feasibility. As other in vitro cell culture models, the system is suitable for miniaturization and robotization to allow high throughput screening of CNS candidates.

Our aim was to evaluate the in vitro efficacy of a series of N-benzenesulfonamides of amine substituted aromatic rings, sulfonamides 1-6, against Trypanosoma cruzi and Leishmania spp. and to compare their trypanocidal and leishmanicidal profile. In order to elucidate the probable mechanism of action, the interaction of selected sulfonamides with pUC18 plasmid DNA was investigated by nuclease activity assays. In addition, the cellular targets of these sulfonamides in treated parasites were also searched by transmission and scanning electron microscopy. The most active compounds 4-nitro-N-pyrimidin-2-ylbenzenesulfonamide 1a and 4-chloro-N-5-methyl-thiazol-2-yl-benzenesulfonamide 2d displayed significant in vitro activity against Leishmania spp. promastigotes, without toxicity to J774 macrophages. Selected sulfonamides 1a, 4-nitro-N-pyrazin-2-yl-benzenesulfonamide 1n and 2d were also active against Leishmania infantum intracellular amastigotes. Compounds 1n and 2d showed nuclease activity in the presence of copper salt analogous to our previous results with sulfonamide 1a. Mechanistic data reveal the involvement of a redox process. Evidence for the formation of reactive oxygen species (ROS) responsible for DNA strand scission is provided for sulfonamides 1a, 1n and 2d. Transmission electron microscopic (TEM) analysis of L. infantum promastigotes treated with compounds 1a, 1n and 2d shows an overall cellular disorganization effects which are mainly addressed to DNA bearing structures such as the nucleus, mitochondria and kinetoplast. Disruption of double nuclear membrane and loss of cellular integrity along with accumulation of cytoplasmic electrodense bodies were also frequently observed.

β-Lapachone is a naphthoquinone with high and diverse biological activity. It is useful to treat several pathologies, in particular cancer due to its ability to induce selective apoptosis in tumoral cells. Despite its advantages, β-lapachone’s clinical applications are limited by its low solubility in biological fluids and its low specific distribution pattern. In the last decade various formulation strategies have been designed and developed in order to overcome these limitations and to make feasible its wide therapeutic use. The most relevant strategies for oral administration have been focused on the improvement of its solubility in order to increase β-lapachone oral bioavailability. The use of cyclodextrins or the inclusion of the active substance in micelles and microparticles has demonstrated to be successful approaches. For topical and local application temperature-sensitive hydrogels have been developed to achieve high target concentrations. Polymeric micelles administrated subcutaneously have proved to be a long-lasting circulating system and to increase accumulation in tumor tissue making them an interesting formulation strategy.

To ensure the optimal performance of oral controlled release formulations, drug colon permeability is one of the critical parameters. Consequently developing this kind of formulations for low permeability molecules requires strategies to increase their ability to cross the colonic membrane. The objective of this work is to show if an ion-pair formation can improve the colon permeability of atenolol as a low permeability drug model. Two counter ions have been tested: brilliant blue and bromophenol blue. The Distribution coefficients at pH 7.00 (DpH7) of atenolol, atenolol + brilliant blue and atenolol + bromophenol blue were experimentally determined in n-octanol. Moreover, the colonic permeability was determined in rat colon using in situ closed loop perfusion method based in Doluisio's Technique. To check the potential effects of the counter ions on the membrane integrity, a histological assessment of colonic tissue was done. The results of the partitioning studies were inconclusive about ion-pair formation; nevertheless colon permeability was significantly increased by both counter ions (from 0.232 ± 0.021 cm/s to 0.508 ± 0.038 cm/s in the presence of brilliant blue and to 0.405 ± 0.044 cm/s in the presence of bromophenol blue). Neither damage on the membrane was observed on the histological studies, nor any change on paracellular permeability suggesting that the permeability enhancement could be attributed to the ion-pair formation.

β-Lapachone (βLAP) is a promising, poorly soluble, antitumoral drug. βLAP combination with cyclodextrins (CDs) improves its solubility and dissolution but there is not enough information about the impact of cyclodextrins on βLAP intestinal permeability. The objectives of this work were to characterize βLAP intestinal permeability and to elucidate cyclodextrins effect on the dissolution properties and on the intestinal permeability. The final goal was to evaluate CDs influence on the oral absorption of βLAP. Binary systems (physical mixtures and inclusion complexes) including βLAP and CDs (β-cyclodextrin: βCD, random-methyl-β-cyclodextrin: RMβCD and sulfobutylether-β-cyclodextrin: SBEβCD) have been prepared and analysed by differential scanning calorimetry. βLAP (and its combinations with CDs) absorption rate coefficients and effective permeability values have been determined in vitro in MDCK or MDCK-Mdr1 monolayers and in situ in rat by a closed loop perfusion technique. DSC results confirmed the formation of the inclusion complexes. βLAP–CDs inclusion complexes improve drug solubility and dissolution rate in comparison with physical mixtures. βLAP presented a high permeability value which can provide complete oral absorption. Its oral absorption is limited by its low solubility and dissolution rate. Cyclodextrin (both as physical mixtures and inclusion complexes) showed a positive effect on the intestinal permeability of βLAP. Complexation with CDs does not reduce βLAP intestinal permeability in spite of the potential negative effect of the reduction in free fraction of the drug. The use of RMβCD or SBEβCD inclusion complexes could benefit βLAP oral absorption by enhancing its solubility, dissolution rate and permeability.

The interaction of the Cu(II) drugs CuL(NO(3)) and CuL'(NO(3)) (HL is pyridine-2-carbaldehyde thiosemicarbazone and HL' is pyridine-2-carbaldehyde 4N-methylthiosemicarbazone, in water named [CuL](+) and [CuL'](+)) with [poly(dA-dT)](2), [poly(dG-dC)](2), and calf thymus (CT) DNA has been probed in aqueous solution at pH 6.0, I = 0.1 M, and T = 25 degrees C by absorbance, fluorescence, circular dichroism, and viscosity measurements. The results reveal that these drugs act as groove binders with [poly(dA-dT)](2), with a site size n = 6-7, whereas they act as external binders with [poly(dG-dC)](2) and/or CT-DNA, thus establishing overall electrostatic interaction with n = 1. The binding constants with [CuL'](+) were slightly larger than with [CuL](+). The title compounds display some cleavage activity in the presence of thiols, bringing about the rupture of the DNA strands by the reactive oxygen species formed by reoxidation of Cu(I) to Cu(II); this feature was not observed in the absence of thiols. Mutagenic assays performed both in the presence and in the absence of S9 mix, probed by the Ames test on TA 98, TA 100, and TA 102, were negative. Weak genotoxic activity was detected for [CuL](+) and [CuL'](+), with a significative dose-response effect for [CuL'](+), which was shown to be more cytotoxic in the Ames test and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide cell proliferation assays. Methylation of the terminal NH(2) group enhances the antiproliferative activity of the pyridine-2-carbaldehyde thiosemicarbazones.

We report herein the synthesis of a novel tetracationic tris(2,2′-bipyridine) ligand 4 . We show that this ligand metalated with copper(II), and in the presence of ascorbate as a reducing agent, strongly damages pUC18 plasmid DNA. Copper complex formation was demonstrated by ESI-MS (electrospray ionization-mass spectrum) at a 1:3 ligand to metal ratio. Binding of both 4 and its copper(II) complex to CT-DNA (calf thymus DNA) was characterized by viscosimetry, thermal denaturation and fluorescence-based competition assays. The viscosimetric data indicated that 4 and its copper(II) complex bind DNA through partial intercalation and thermal denaturation studies revealed a significant increase of duplex DNA stability in the presence of these species (Δ T m = 16.4 and 18.3 °C, respectively). Moreover, 4 and its copper(II) complex were found to effectively compete with ethidium bromide for the intercalative binding sites of DNA. Overall, the copper(II)– 4 complex constitutes a very efficient DNA cleaving agent in the presence of ascorbate. Experiments with scavengers further suggest that the generation of Cu(I), hydrogen peroxide, superoxide, hydroxyl radical and singlet oxygen-like species contributes to the DNA breakage induced by the Cu(II) complex of 4 .

The complex [Cu(N9-ABS)(phen) 2 ]·3.6H 2 O, H 2 N9-ABS = N -(9 H -purin-6-yl)benzenesulfonamide and phen = 1,10-phenanthroline, has been synthesized and then characterized with the aid of X-ray diffraction, analytical, and spectroscopic techniques. The geometry of Cu(II) is distorted square pyramidal with the equatorial positions occupied by three N atoms from two phenantroline molecules and one N atom from the adenine ring of the sulfonamide ligand. The interaction of the complex with DNA was studied by means of viscosity measurements and fluorescence spectroscopy. The results pointed to a classic intercalation of the complex between the DNA base pairs. The complex was found to be a very efficient agent of plasmid DNA cleavage in the presence of ascorbate. Both the kinetics and the mechanism of the cleavage reaction were studied. In addition, the cytotoxic properties of the complex were evaluated in human Jurkat T and Caco-2 cell lines. The cytotoxicity of the compound was higher than that of the reference ([Cu(phen) 2 ] 2+ ). The mechanism and type of cell death induced by the compound was determined by flow cytometry and Hoechst dye staining. The compound demonstrated a significant ability to induce cell death by apoptosis. The apoptosis induced by [Cu(N9-ABS)(phen) 2 ]·3.6H 2 O was associated with an increase in p53 protein levels while those of Bcl-2 were reduced.

Three doubly-bridged, trinuclear copper(II) compounds with hydroxo and carboxylato bridges, ∞ 1 [Cu 3 (L1) 2 (μ-OH) 2 (μ-propionato) 2 ] ∞ 1 [ Cu 3 ( L 1 ) 2 ( μ - OH ) 2 ( μ - propionato ) 2 ] (1) , [Cu 3 (L2) 2 (μ-OH) 2 (μ-propionato) 2 (DMF) 2 ] (2) and ∞ 1 {[Cu 3 (L3) 2 (μ-OH) 2 (μ-propionato) 2 ]} ∞ 1 { [ Cu 3 ( L 3 ) 2 ( μ - OH ) 2 ( μ - propionato ) 2 ] } [Cu 3 (L3) 2 (μ-OH) 2 (μ-propionato) 2 (DMF) 2 ]} (3) [HL1 = N -(pyrid-2-ylmethyl)benzenesulfonylamide, HL2 = N -(pyrid-2-ylmethyl)toluenesulfonylamide, HL3 = N -(pyrid-2-ylmethyl)naphthalenesulfonylamide], have been synthesized and characterized. 1 is built from [Cu 3 (L1) 2 (μ-OH) 2 (μ-propionato) 2 ] clusters. Each unit contains three copper(II) with two different coordination environments: the terminal centers are square-base pyramidal whereas the central copper is square planar. 2 presents a similar square-base pyramidal geometry in the terminal centers, but the central copper is six-coordinate. 3 shows an unusual 1D coordination polymer comprised of two distinct building blocks: one similar to that found in 1 and the other similar to that found in 2 . The magnetic susceptibility measurements (2–300 K) reveal a ferromagnetic interaction between the Cu(II) ions with J values of 76.0, 55.0, and 48.0 cm −1 for 1 , 2 , and 3 , respectively. Emission spectroscopy, thermal denaturation, viscosimetry and cyclic voltammetry show an interaction of the complexes with DNA through the sugar–phosphate backbone. All three Cu(II) complexes were found to be very efficient agents of plasmid DNA cleavage in the presence of ascorbato or mercaptopropionic acid. Both the kinetics and the mechanism of the cleavage reaction have also been examined.

In this study, a new binary complex [Cu(HCip) 2 ](NO 3 ) 2 · 6H 2 O ( 1 ) has been synthesized and then characterized by X-ray structure analyses. In this compound, each ciprofloxacin acts as a bidentate ligand resulting in a crystallographically planar configuration; the nitrate anions are located in apical positions with an axial distance significantly larger than the equatorial distances, which would be consistent with a very weak metal ion interaction due to the Jahn–Teller effect. In addition, both the synthesis and characterization of two new ternary complexes of ciprofloxacin–copper(II)–1,10-phenanthroline, [Cu(phen)(Cip)](NO 3 ) · 4H 2 O ( 2 ) and Cu(phen)(HCip)(NO 3 ) 2 · H 2 O ( 3 ) , have been accomplished. We have also undertaken the single crystal structural determination of 2 , in which the ciprofloxacin acts as tridentate bridging ligand; the complex exhibits a five-coordinated motif in a distorted square pyramidal environment around the metal center. The chemical nuclease activity of compounds 2 and 3 has also been studied, revealing that both compounds behave as efficient chemical nucleases in the presence of ascorbate. Mechanistic studies, with various radical oxygen scavengers, indicate that the DNA cleavage reaction is mediated by hydroxyl radicals, singlet oxygen, and the superoxide anion.

The work presented here describes the synthesis of new copper(II) complexes derived from 2-amino-5-(pyridin-2-yl)-1,3,4-oxadiazole (L1) and 2-methylamino-5-(pyridin-2-yl)-1,3,4-oxadiazole (L2) which also incorporate azido (N3 -), thiocyanato (NCS-), cyanato (NCO-), dicyanamido [N(CN)2 -, dca] and malonato (C3H2O4 2-, mal) coligands. Structures of the [Cu(L2)(mal)(H2O)]·2H 2O(1), [{Cu(L2)(mal)}2] (2), [Cu(L 2)2-(NCS)2] (3), [Cu(L1)(NCS) 2]n (4) and [{Cu(L2)}2(dca) 2](ClO4)2·2H2O (5) compounds show the dependence of the dimensionality on parameters such as the type of oxadiazole and coligand utilised, solvents or reaction times. In this sense, 1 and 3 are mononuclear complexes, 2 contains centrosymmetric dinuclear entities, 4 is a 1D compound in which thiocyanato groups act as end-to-end bridges and 5 shows a 2D arrangement of dinuclear motifs linked through dicyanamido spacers. The copper(II) ions acquire distorted octahedral (3 and 4), square-pyramidal (1 and 2) and trigonal bipyramidal (5) geometries. Stacking π-π interactions and hydrogen bonds stabilise the crystal frameworks. Water tetramers are present in 1. Spectroscopic (infrared, UV/Vis absorption and electron spin resonance) studies have also been developed for 1-5 and for the Cu(L2)(dca) 2(H2O) (6), Cu(L2)(NCO)2 (7) and Cu(L2)2(N3)2(H2O) 2 (8) complexes. The studies suggest the coordination of oxadiazole and coligands in all cases. Magnetic measurements show antiferromagnetic coupling for compound 5, ferromagnetic interactions for compound 2 and paramagnetic behaviour in compound 4. The susceptibility data were fitted by using the Bleaney-Bowers' equation for the copper(II) dimers derived from H = -2JS1S2. The resultant J/k values are +2.15 and -38.3 K for compounds 2 and 5, respectively. Magneto-structural correlations are discussed and extended to more than 30 published dinuclear Cu(NN) 2Cu′ diazole-only bridged compounds. All these systems show J values close to or greater than those given by the expression J ≈ -90 + 3 Δβ (where Δβ is the difference between the Cu-N-N and Cu-N′-N′ angles). The artificial nuclease activity of 1, 2, 3 and 5 in the presence of mercaptopropionic acid or glutathione has also been analysed and compared with that exhibited by [Cu(L2)2(H 2O)2](NO3)2 and CuSO4. © Wiley-VCH Verlag GmbH & Co. KGaA, 2009., We thank Dr. J. J. Delgado (SCAI, Universidad de Burgos, Spain) for the elemental analyses and Prof. R. Olazcuaga and Dr. F. Guillen (CNRS, Pessac, France) for the reflectance measurements. This work was financially supported by the Junta de Castilla y Leon (BU033A06) and Ministerio de Ciencia y Tecnologia (MCYT) [CTQ2007-63690 (ERDF(EU)-DGI-MCYT)]. P. G.-S. thanks the Universidad de Burgos for a Doctoral Fellowship.

The reaction of 2,4,6-tris(pyridyl)-1,3,5-triazine (ptz) and copper(II) salts in DMF–water (4:1) resulted in the hydrolysis of ptz, giving rise to the bis(2-pyridylcarbonyl)amide anion ( ptO 2 - ) and affording the complexes [Cu(ptO 2 )(2-Clbenz)(H 2 O)] ( 1 ), [Cu(ptO 2 )(3-Clbenz)] ( 2 ), and [Cu(ptO 2 )(4-Clbenz)(DMF)] ( 3 ). This report includes the chemical and spectroscopic characterization of all these compounds along with the crystal structures of the Cu(II) complexes thus formed. The coordination spheres of Cu(II) in 1 and 3 are best described as distorted square based pyramidal, while for 2 the coordination sphere is distorted square planar.

Three new coordination compounds have been prepared by reacting copper salts with sulfonamides derived from amino acids. Their molecular structures have been determined with the aid of single crystal X-ray diffraction. The copper cations are four-coordinated in all cases and the local environment is almost planar. Compound [Cu(ts-gly)(NH3)2]·H2O (1) crystallizes in monoclinic space group P21/c (no 14) with Z = 4 and with unit cell parameters a = 6.0490(12) A, b = 24.719(5) A, c = 9.159(2) A, α = γ = 90°, and β = 94.69(3)°. Compound [Cu(ts-isoleu)(NH3)2] (2) crystallizes in monoclinic space group C2/c (no 8) with Z = 15 and with unit cell parameters a = 30.800(6) A, b = 6.5510(13) A, c = 15.830(3) A, α = γ = 90°, and β = 100.19(3)°. Compound Na[Cu(ts-gly)(gly)]·2H2O (3) crystallizes in triclinic space group P-1 (no 2) with Z = 2 and with unit cell parameters a = 5.6220(11) A, b = 7.775(2) A, c = 18.704(4) A, α = 94.61(3)°, β = 98.32(3)°, and γ = 98.29(3)°. The study of nuclease activity with plasmid pUC18 shows that all three compounds act as chemical nucleases, although complex Na[Cu(ts-gly)(gly)]·2H2O is more efficient in splitting the DNA chains than the other two compounds.

Three new binary Cu(II) complexes of norfloxacin have been synthesized and characterized. We also report the synthesis, characterization and X-ray crystallographic structures of a new binary compound, [Cu(HNor)(2)]Cl(2).2H(2)O (2) and two new ternary complexes norfloxacin-copper(II)-phen, [Cu(Nor)(phen)(H(2)O)](NO(3)).3H(2)O (4), and [Cu(HNor)(phen)(NO(3))](NO(3)).3H(2)O (5). The structure of 2 consists of two crystallographically independent cationic monomeric units of [Cu(HNor)(2)](2+), chloride anions, and uncoordinated water molecules. The Cu(II) ion is placed at a center of symmetry and is coordinated to two norfloxacin ligands which are related through the inversion center. The structures of 4 and 5 consist of cationic units ([Cu(Nor)(phen)(H(2)O)](+) for 4 and [Cu(HNor)(phen)(NO(3))](+) for 5), nitrate counteranions, and lattice water molecules that provide crystalline stability through a network of hydrogen-bond interactions. The complexes exhibit a five coordinated motif in a square pyramidal environment around the metal center. The ability of compounds 4 and 5 to cleave DNA has also been studied. Mechanistic studies with different inhibiting reagents reveal that hydroxyl radicals, singlet oxygen, and superoxide radicals are all involved in the DNA scission process mediated by these compounds.

The reaction of 2,4,6-tri(pyridyl)-1,3,5-triazine (ptz) and copper(II) salts in dmf/water (1:1) results in the hydrolysis of ptz and formation of the anions bis(2-pyridylcarbonyl)amide (ptO2–) and bis(2-pyridylamine)amide (ptN2–), which are found in the complexes [Cu(ptN2)(OAc)]·3H2O (1), [Cu(ptO2)(OAc)(H2O)]·H2O (2), [Cu(ptN2)(for)]·3H2O (3) (for = formate), [Cu(ptO2)(for)(H2O)] (4), [Cu(ptO2)(benz)]·H2O (5) (benz = benzoate), and [Cu(ptO2)F(H2O)]2·3H2O (6). This report includes the chemical and spectroscopic characterization of all these complexes along with the crystal structures of 4–6. The coordination spheres of CuII in 4 and 5 are best described as distorted tetragonal square pyramidal for the former and distorted square planar for the latter. The crystal structure of 6 shows the presence of two discrete monomeric [Cu(ptO2)F(H2O)] entities in the crystallographic asymmetric unit in which both copper(II) ions have a distorted square-pyramidal coordination geometry. The binding of the complexes to DNA has been investigated with the aid of viscosity and thermal denaturation studies, both of which indicate that the interaction is probably due to the outer-sphere mechanism. The ability of the compounds to cleave DNA has also been tested. Efficient oxidative cleavage was observed in the presence of a mild reducing agent (ascorbate) and dioxygen. Mechanistic studies with reactive oxygen species (ROS) scavengers confirm that hydrogen peroxide, the hydroxyl radical, singlet oxygen-like species, and the superoxide anion are necessary diffusible intermediates in the scission process. A mechanism involving either the Fenton or theHaber–Weiss reaction plus the formation of copper oxene species is proposed for the DNA cleavage mediated by these compounds.(© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2007)

Predictive animal models of intestinal drug absorption are essential tools in drug development to identify compounds with promising biopharmaceutical properties. In situ perfusion absorption studies are routinely used in the preclinical setting to screen drug candidates. The objective of this work is to explore the differences in magnitude and variability on intestinal absorption associated with rat strain and gender. Metoprolol and Verapamil absorption rate coefficients were determined using the in situ closed loop perfusion model in four strains of rats and in both genders. Strains used were Sprague-Dawley, Wistar-Han, Wistar-Unilever, Long-Evans and CD∗IGS. In the case of Metoprolol only CD∗IGS and Wistar Unilever showed differences between males and females. For Verapamil, Wistar Han and Sprague-Dawley strains do not show differences between male and female rats. That means that in these strains permeability data from male and female could be combined. In male rats, which are commonly used for permeability estimation, there were differences for Metoprolol permeability between Sprague-Dawley (with lower permeability values) and the other strains, while for Verapamil Sprague-Dawley and Wistar-Han showed the lower permeability values. In conclusion, the selection of rat's strain and gender for intestinal absorption experiments is a relevant element during study design and data from different strains may not be always comparable.

Four ternary complexes [Cu(L)2(phen)] where L is an N(benzothiazol-2-yl)sulfonamide derivative have been prepared and their ability to cleave DNA has been studied. The complexes were structurally characterized with the aid of single-crystal X-ray crystallography. Whereas the molecular structure of the [Cu(L1)2(phen)] (1) [HL1 = N-(6-chlorobenzothiazol-2-yl)benzenesulfonamide] and [Cu(L3)2(phen)] (3) [HL3 = N-(benzothiazol-2-yl)benzenesulfonamide] complexes can best be described as having a distorted squareplanar geometry, that of the [Cu(L4)2(phen)] (4) [HL4 = N(benzothiazol-2-yl)toluenesulfonamide] complex shows a strictly square-planar geometry. The [Cu(L2)2(phen)MeOH] (2) [HL2 = N-(6-chlorobenzothiazol-2-yl)toluenesulfonamide] complex displays an axially elongated square-pyramidal coordination geometry in which the phen ligand binds at the

Reaction between 2-picolylamine (2-aminomethylpyridine) with 2-mesitylenesulfonyl and 4- tert -butylbenzene sulfonyl chlorides leads to the formation of 2,4,6-trimethyl- N -[pyridin-2-ilmethyl]benzenesulfonamide (Hpmesa) and 4-ter-butyl- N -[pyridin-2-ilmethyl]benzenesulfonamide (Hptbsa). These compounds react with Cu(II) salts to yield coordination compounds with CuL 2 stoichiometry. The immediate environment of the metal ion is a highly distorted tetrahedron, the sulfonamide ligands acting in a bidentate fashion. Compound Hpmesa crystallizes in monoclinic space group P 2 1 /c (number 14), with Z = 4; complex [Cu(pmesa) 2 ] crystallizes in monoclinic space group P 1 ¯ (number 2), with Z = 4 and complex [Cu(ptbsa) 2 ] crystallizes in monoclinic space group Pca 2 1 (number 29), with Z = 4. Both complexes have been characterized by spectroscopic methods. EPR measurements show rhombic spectra with parameters indicating that the unpaired electron is in d x 2 -y 2 d x 2 - y 2 orbital. The two complexes act as chemical nucleases in the presence of ascorbate/H 2 O 2 , the reactive oxygen species being the hydroxyl and the singlet oxygen-like species.

A new dinuclear copper(II) complex has been synthesised and structurally characterised: [Cu2(tz-ben)4] (Htz-ben = N-thiazol-2-yl-benzenesulfonamide). Its crystal structure, magnetic properties and electronic paramagnetic resonance (EPR) spectra were studied in detail. In the compound the metal centres are bridged by four non-linear triatomic NCN groups. The coordination geometry of the copper ions in the dinuclear entity is distorted square pyramidal (4+1). Two thiazole N and two sulfonamido N atoms occupy the equatorial positions and one sulfonamido O atom is in the axial position. Magnetic susceptibility data show a strong antiferromagnetic coupling, -2J = 114.1 cm(-1). The EPR spectra of a polycrystalline sample of compound has been obtained at the X- and Q-band frequencies at different temperatures. Above 20K the spectra are characteristic of S = 1 species with a zero field splitting parameter D = 0.4 cm(-1). The EPR parameters are discussed in terms of the known binuclear structures. The chemical nuclease ability of the title complex and that of the related [Cu2(tz-tol)4] compound (Htz-tol = N-thiazol-2-yl-toluenesulfonamide) is reported. The participation of hydroxyl radicals and a singlet oxygen-like entity in the DNA cleavage reaction has been deduced from the assays with radical oxygen scavengers.

Copper(II) complexes of N-benzothiazolesulfonamides (HL1=N-2-(4-methylphenylsulfamoyl)-6-nitro-benzothiazole, HL2=N-2-(phenylsulfamoyl)-6-chloro-benzothiazole, and HL3=N-2-(4-methylphenylsulfamoyl)-6-chloro-benzothiazole) with ammonia have been synthesized and characterized. The crystal structures of the [Cu(L1)2(NH3)2].2MeOH, [Cu(L2)2(NH3)2], and [Cu(L3)2(NH3)2] compounds have been determined. Compounds and present a distorted square planar geometry. In both compounds the metal ion is coordinated by two benzothiazole N atoms from two sulfonamidate anions and two NH3 molecules. Complex is distorted square-pyramidal. The Cu(II) ion is linked to the benzothiazole N and sulfonamidate O atoms of one of the ligands, the benzothiazole N of another sulfonamidate anion, and two ammonia N atoms. We have tested the superoxide dismutase (SOD)-like activity of the compounds and compared it with that of two dinuclear compounds [Cu2(L4)2(OCH3)2(NH3)2] and [Cu2(L4)2(OCH3)2(dmso)2] (HL4=N-2-(phenylsulfamoyl)-4-methyl-benzothiazole). In vitro indirect assays show that the dimeric complexes are better SOD mimics than the monomeric ones. We have also assayed the protective action provided by the compounds against reactive oxygen species over Deltasod1 mutant of Saccharomyces cerevisiae. In contrast to the in vitro results, the mononuclear compounds were more protective to SOD-deficient S. cerevisiae strains than the dinuclear complexes.