Assessing carbon-encapsulated iron nanoparticles cytotoxicity in Lewis lung carcinoma cells.

Carbon-encapsulated iron nanoparticles (CEINs) have been considered as attractive candidates for several biomedical applications. In the present study, we synthesized CEINs (the mean diameter 40-80 nm) using a carbon arc route, and the as-synthesized CEINs were characterized (scanning and transmission electron microscopy, dynamic light scattering, turbidimetry, Zeta potential) and further tested as raw and purified nanomaterials containing the carbon surface modified with acidic groups. For cytotoxicity evaluation, we applied a battery of different methods (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, lactate dehydrogenase, calcein AM/propidium iodide, annexin V/propidium iodide, JC-1, cell cycle assay, Zeta potential, TEM and inductively coupled plasma mass spectrometry) to address the strategic cytotoxic endpoints of Lewis lung carcinoma cells due to CEIN (0.0001-100 µg ml(-1) ) exposures in vitro. Our studies evidence that incubation of Lewis lung carcinoma cells with CEINs is accompanied in substantial changes of zeta potential in cells and these effects may result in different internalization profiles. The results show that CEINs increased the mitochondrial and cell membrane cytotoxicity; however, the raw CEIN material (Fe@C/Fe) produced higher toxicities than the rest of the CEINs studied to data. The study showed that non-modified CEINs (Fe@C/Fe and Fe@C) elevated some pro-apoptotic events to a greater extent compared to that of the surface-modified CEINs (Fe@C-COOH and Fe@C-(CH2 )2 COOH). They also diminished the mitochondrial membrane potentials. In contrast to non-modified CEINs, the surface-functionalized nanoparticles caused the concentration- and time-dependent arrest of the S phase in cells. Taken all together, our results shed new light on the rational design of CEINs, as their geometry, hydrodynamic and, in particular, surface characteristics are important features in selecting CEINs as future nanomaterials for nanomedicine applications.

Counterfeit medicines in Poland: opinions of primary healthcare physicians, nurses and lay persons.

AIMS AND OBJECTIVES: To gain information concerning disparities in the understanding of the counterfeit medicines phenomenon between healthcare workers and lay persons. BACKGROUND: Central-eastern Europe is facing significant challenges in combating a multi-billion euro, and often lethal, trade in counterfeit medicines. It is a major challenge especially for primary healthcare workers to expand the understanding of counterfeit medicines to the benefit of patients. DESIGN: Use of questionnaires. Two separate questionnaires were distributed, one for healthcare professionals and the other for lay persons. METHODS: Conducted between September 2009-May 2010. One thousand and seventy-eight primary healthcare professionals and 377 lay persons were surveyed. RESULTS: Findings revealed less awareness among healthcare professionals than lay persons about the danger of purchasing illegal medicines or dietary supplements outside pharmacies. Healthcare professionals have lower levels of awareness about the scale of counterfeit medicines as well as threats of counterfeit medicines to health than lay persons. The majority of medical workers do not know the procedure for reporting suspicious medicine and do not warn their patients against purchasing medicine from unknown sources. CONCLUSIONS: Primary healthcare workers have less awareness of the scale of the counterfeit medicines phenomenon than lay persons. RELEVANCE TO CLINICAL PRACTICE: Nurses and physician need to become aware of the counterfeit medicines phenomenon. Nurses are well positioned to assume the active role in educating patients about the threat of the presence of counterfeit medicines so as to enhance safety for their patients. However, to accomplish that aim, these findings suggest that healthcare professionals need to become better educated about counterfeit medicines and need to be trained in skills to identify counterfeit medicines.

What Polish hospital healthcare workers and lay persons know about counterfeit medicine products?

AIM: To report on Polish hospital health care workers' and lay persons' knowledge about counterfeit medicine products. MATERIALS AND METHODS: Cross-sectional design was used. Two types of questionnaire survey about counterfeit medicine, separate for health care professionals and lay persons were completed by 201 physicians and nurses, and 450 adult Polish residents between October 2008 and January 2009. RESULTS: Physicians and nurses working in hospitals are more aware of counterfeit medicine than lay persons and more often notice the presence of drugs from unknown sources. Nearly 90% of physicians, 80% of nurses, and more than 40% of representatives of the lay persons had heard about the possibility of importing illegal medicine from Ukraine or China. The majority of medical workers does not know the procedure for reporting suspicious medicine and do not warn their patients against purchasing medicine from unknown sources. CONCLUSIONS: Increase education of nurses and physicians about counterfeit medicine particularly including the procedure of reporting suspicious medicine from unknown sources. In practice, reinforce a role of nurses and physicians in warning their patients against purchasing medicine from unknown sources and educate them about possible health hazards and life risks.

Determination of selected elements in different pharmaceutical forms of some Polish herbal medicinal products.

In the present work, the following microelements and heavy metals were determined in the Willow bark (Cortex Salicis), in the St. John's wort (Herba Hyperici), in the infusions from these raw materials and in the tablets containing these herbs: Cr, Cu, Fe, Ni, Zn and Ba, Cd, Pb. The concentration of the metals were determined by means of the GFAAS and ICP-MS methods. Considering the examined two herbs as a source of toxic metals, the occurrence of high levels (exceeding the European Pharmacopoeia limits) of Cd should be emphasized. The level of cadmium was low in the infusions because of a low extraction efficiency of this metal, so that the infusions can be found as a rather safe form of herbal medicine. In the tablets, the Cd contents were similar to the levels found in the raw materials. Because the total heavy metal content can be accessible, it may cause a potential health risk for human.

Determination of local anesthetics in illegal products using HPLC method with amperometric detection.

An HPLC method with amperometric detection was developed for analysis of two local anesthetics (lidocaine and benzocaine) in products for delaying ejaculation illegally marketed in Polish sex shops. Chromatographic elution on an RP column C18 with mobile phase composed of acetate buffer with acetonitrile, provides an optimal separation not only of active substances but also electroactive preservatives which are occasionally added to cosmetic creams (methylparaben and propylparaben). Application of glassy carbon electrode as a working electrode and a procedure with pulsed potential waveforms enables a sensitive, accurate measurement within a relatively short analysis time (250 s). This method has been successfully employed for the determination of local anesthetics in products under investigation. The obtained results show that most samples contained therapeutic concentrations of lidocaine or benzocaine. According to European law, a sale of products containing lidocaine or benzocaine outside the pharmacy sector is forbidden.

Determination of Gentamicin Sulphate Composition and Related Substances in Pharmaceutical Preparations by LC with Charged Aerosol Detection.

A new, simple and repeatable liquid chromatography method with charged aerosol detection (LC-CAD) for the determination of gentamicin sulphate composition and related substances has been developed. Gentamicin lacks of chromophores, therefore its determination is quite problematic. Using a universal CAD enables to achieve good separation without sample derivatization. Mass spectrometry was employed to confirm the LC-CAD peak profile. The proposed method was validated and applied for the determination of gentamicin sulphate composition and related substances in pharmaceutical preparations.

Determination of total hypericins in St. John's wort and herbal medicinal products.

The work aimed to determine the levels of hypericins expressed as hypericin in the herbal substance of St. John's wort, in capsules and tablets containing the extract of St. John's wort, tablets containing powdered herb and in tincture and juice from fresh St. John's wort, by HPLC method with spectrophotometric detection. In addition, the amount of hypericins in the infusion prepared from St. John's wort was determined by HPLC and spectrophotometry methods. According to traditional indications aqueous infusions from St. John's wort containing mainly hydrophilic components are used in gastrointestinal diseases. On the other hand, ethanolic extracts containing hypericin and hyperforin affect the CNS and are indicated for the treatment of episodes of mild depressive disorders. The results obtained in the work indicate that the daily dose of hypericins taken by a patient as infusions is 0.328 mg on average for herbs in sachets and in bulk form. It can be compared to the daily dose of hypericins contained in tablets and capsules based on the alkoholic extract of St. John's wort and tablets containing powdered St. John's wort herb. For solid dosage forms, this dose ranges from 0.288 mg to 0.636 mg. The assays were performed using consistent analytical methods for all tested pharmaceutical products and consequently it was possible to compare doses taken by patients and their strength of action.

[Counterfeit phosphodiesterase type 5 inhibitors--growing safety risks for public health]. / Sfalszowane inhibitory fosfodiesterazy typu 5--narastajace zagrozenie dla zdrowia publicznego.

Counterfeit drugs, medical devises and dietary supplements are inherently dangerous and a growing problem. In Europe the growth of the counterfeit medication market is attributable in part to registration of phosphodiesterase type 5 inhibitors (PDE-5) used for the erectile dysfunction. "Viagra, Levitra and Cialis belong to this group. It has been estimated that up to 2.5 million men in Europe are exposed to an illicit sildenafil, suggesting that there may be as many illegal as legal users of sildenafil. In Europe a strong trend is observed towards increasingly professional counterfeits and imitations of Viagra, Cialis and Levitra, with regard to the appearance of tablets, capsules and packaging. The professional presentation will deceive potential consumers into assuming these products are legal, efficacious and safe. Globally, increased obstacles for counterfeiters are necessary to combat pharmaceutical counterfeiting, including fines and penalties. The worldwide nature of the counterfeit problem requires proper coordination between countries to ensure an adequate enforcement. We described the usefulness of the time-of-flight mass spectrometry with the electrospray ionization (LC-ESI-MS-TOF) and the X-ray powder diffraction method (XRPD) for PDE-5 counterfeit screening from the Polish illegal market.

Determination of pipecuronium bromide and its impurities in pharmaceutical preparation by high-performance liquid chromatography with coulometric electrode array detection.

A sensitive and selective HPLC method with coulometric electrode array detection for the determination of pipecuronium bromide and its four impurities has been developed. The coulometric electrode array detection at increasing potentials from +300 to +900mV of the porous graphite electrode versus the palladium reference electrode was used. The limit of detection and quantitation for pipecuronium bromide was 8 and 25ngml(-1), respectively. This elaborate method for the simultaneous analysis of pipecuronium bromide and its impurities proved to be fast, precise, accurate, sensitive, and could be applied to analysis in substances and in pharmaceutical preparations.

Application of high-performance liquid chromatography with amperometric and coulometric detection to the analysis of SZ1677, a new neuromuscular blocking agent, and its two derivatives.

A HPLC method with amperometric and coulometric detection for the determination of SZ1677 and its two derivatives SZ1676 and SZ1823 has been developed. This active substance is under development (clinical trial) and there are no analytical methods published for the determination of SZ1677 thus far. The limit of quantitation for SZ1677 was 25 and 100 ng ml(-1) by the coulometric and amperometric detection, respectively. The elaborate method for the simultaneous analysis of SZ1677 and its two derivatives proved to be fast, precise, accurate and sensitive.

Identification and analysis of drugs in the solid state by 13C CPMAS NMR: suxamethonium chloride and hydrocortisonum (Corhydron).

Cross-polarization (CP) magic angle spinning (MAS) 13C NMR spectroscopy has become a routine tool in pharmacy, employed to identify and characterize drugs in the solid phase. 13C CPMAS NMR spectra were recorded for solid hydrocortisone 21-hemisuccinate and suxamethonium chloride. White crystalline substances, such as these two drugs, can be easily distinguished; and solid-state 13NMR spectra of remarkably good quality are obtained in less than half an hour. 13C CPMAS chemical shifts for solid suxamethonium chloride and hydrocortisone sodium hemisuccinate are given, as well as cross-polarization kinetic parameters for suxamethonium chloride.

The activity of Selol in multidrug-resistant and sensitive human leukemia cells.

Selol is a mixture of selenitetriglycerides synthesized from sunflower oil. As it contains the element selenium in its structure, it is suspected to exhibit chemopreventive and anticancer activity. In this study, the ability of Selol to inhibit cell proliferation and to induce apoptosis was investigated. Three cell lines were used: leukemia HL-60 cell line and multidrug-resistant HL-60/Dox (resistant to doxorubicin) and HL-60/Vinc (resistant to vincristine). Selol was shown to reduce the cell number as a result of treatment with increasing concentrations. For selected concentrations the evidence of apoptosis (changes in mitochondrial potential and caspase activity) was investigated, as well as changes in lysosome distribution. The study has shown that Selol overcame the cell resistance, as doxorubicin-resistant cells were more sensitive towards Selol than sensitive cells.

Simultaneous determination of rocuronium and its eight impurities in pharmaceutical preparation using high-performance liquid chromatography with amperometric detection.

A sensitive and selective HPLC method with amperometric detection (HPLC-ED) for the determination of rocuronium bromide and its eight impurities has been developed. The analysis was performed on Hypersil 100 Silica column 5 microm (250 mm x 4.6 mm; Thermo Electron). The mobile phase consisting of 4.53 g l(-1) solution of tetramethylammonium hydroxide adjusted to pH 7.4 with 85% phosphoric acid:acetonitrile (1:9), was found the best for the separation and determination of the studied compounds. The chromatograms were recorded over 10 min using the amperometric detection at a potential +0.9 V of the glassy carbon electrode versus the reference electrode Ag/AgCl. The limit of quantitation was 45 ng ml(-1) for rocuronium and from 25 to 750 ng ml(-1) for the examined impurities. The proposed HPLC-ED method was successfully applied to the analysis of rocuronium and its impurities in Esmeron solution for injection.

The usefulness of simple X-ray powder diffraction analysis for counterfeit control--the Viagra example.

Counterfeit and illegally manufactured drugs become a very important problem all over the world, hence, a search for new fast, easy, reliable and not expensive methods of drugs screening is essential. We describe the use of X-ray powder diffraction method for the fast screening of tablets for counterfeit medicines identification. The original Viagra tablets and some counterfeit and/or imitations of Viagra were used as example. We demonstrate the application of diffraction method for discrimination of tablets coatings and for identification of differences in drug composition. The X-ray diffraction method turns out to be very fast and reliable for detecting counterfeits and imitation, and it correctly predicts the presence or absence of active substance and/or particular excipients.

Determination of selected microelements in polish herbs and their infusions.

Ba, Cd, Cr, Cu, Fe, Ni, Pb and Zn were determined in birch leaves (Folium Betulae), dandelion roots (Radix Taraxacae), hawthorn blossom (Inflorescentia Crataegi) and their infusions by graphite furnace atomic absorption spectrometry (GFAAS) after microwave digestion of plant samples. Infusions were made from herbs according to prescription for patients, provided by the producer of medicine on the package. The results obtained were compared with daily requirements for each element. Results show high content of cadmium in the medicinal plants analyzed. The highest level in infusions was observed for Ni and Zn (over 90% of the total element concentration for Ni and in most cases over 50% for Zn), and the lowest for Cd and Pb. The calculated daily intake of majority of the analyzed elements was very low (under 1% of daily requirements).

Internalization and cytotoxicity effects of carbon-encapsulated iron nanoparticles in murine endothelial cells: Studies on internal dosages due to loaded mass agglomerates.

Carbon-encapsulated iron nanoparticles (CEINs) qualified as metal-inorganic hybrid nanomaterials offer a potential scope for an increasing number of biomedical applications. In this study, we have focused on the investigation of cellular fate and resulting cytotoxic effects of CEINs synthesized using a carbon arc route and studied in murine endothelial (HECa-10) cells. The CEIN samples were characterized as pristine (the mean diameter between 47 and 56nm) and hydrodynamic (the mean diameter between 270 and 460nm) forms and tested using a battery of methods to determine the cell internalization extent and cytotoxicity effects upon to the exposures (0.0001-100µg/ml) in HECa-10 cells. Our studies evidenced that the incubation with CEINs for 24h is accompanied with substantial changes of Zeta potential in cells which can be considered as a key factor for affecting the membrane transport, cellular distribution and cytotoxicity of these nanoparticles. The results demonstrate that CEINs have entered the endothelial cell through the endocytic pathway rather than by passive diffusion and they were mainly loaded as agglomerates on the cell membrane and throughout the cytoplasm, mitochondria and nucleus. The studies show that CEINs induce the mitochondrial and cell membrane cytotoxicities in a dose-dependent manner resulting from the internal dosages due to CEIN agglomerates. Our results highlight the importance of the physicochemical characterization of CEINs in studying the magnetic nanoparticle-endothelial cell interactions because the CEIN mass agglomerates can sediment more or less rapidly in culture models.

Determination of local anaesthetics and their impurities in pharmaceutical preparations using HPLC method with amperometric detection.

A method for the determination of local anaesthetics and their impurities--2,6-dimethylaniline and o-toluidine--by high-performance liquid chromatographic method with amperometric detection has been developed. The analysis was performed in an isocratic mode on a reversed phase Luna column 5 microm C-18 (100 mm x 4.6 mm). A mobile phase [0.01 mol l(-1) Tris buffer of pH 7.9:acetonitrile (45:55)] was selected for the separation and determination of studied anaesthetics and their impurities. Chromatograms were recorded for 500 s by means of an amperometric detector at a potential of +1.0 V of the glassy carbon electrode versus the reference electrode Ag/AgCl. The proposed liquid chromatographic method was successfully applied to the analysis of commercially available pharmaceutical preparations. The limit of the detection for 2,6-dimethylaniline and o-toluidine was 0.8 ng ml(-1). The limit of quantitation, considering a signal to noise ratio was 1.5 ng ml(-1). The method developed in this study is sensitive and selective and can be applied to routine studies of pharmaceuticals in the form of cream and injection.

Determination of chromium and selected elements in multimineral and multivitamin preparations and in pharmaceutical raw material.

The content of elements in pharmaceutical preparations is one of the indispensable factors of the evaluation of their quality. In the present work, the following macro- and microelements Ca, Cr, Cu, Fe, Mg, Mn, Mo, P, Se and Zn were determined in multimineral and multivitamin preparations and in pharmaceutical raw material. Inductively coupled plasma mass spectrometry (ICP-MS) and electrothermal atomic absorption spectrometry (ET AAS) were used throughout the study. The examined samples were dissolved in a high-pressure microwave system using concentrated nitric acid. The effect of the carbon residue in the digest solution on the determination result was eliminated by introducing an equation correcting the ArC+ interference with 52Cr.

Voltammetric and electrochemical quartz crystal microbalance studies of sulforaphane and its Zn(II) complexes.

Cyclic voltammetry (CV) and electrochemical quartz crystal microbalance (EQCM) methods were used to study the behavior of sulforaphane (SFR). To established electrochemical properties of SFR methyl-, ethyl-, 2-chloroethyl-, propyl-, butyl- and tert-butyl-isothiocyanates were examined at gold electrode. Optimal measurement parameters were established and stock solutions developed. The electrode activity of the studied compounds (cathodic peak E(p)=-0.6 V) was found to be primarily due to the oxidation of the isothiocyanate group. By the EQCM method it was found that during the reduction of SFR the mass of the electrode systematically increased in successive scans by ca. 120 ng per scan and in the mixture of SFR and Zn(2+) ions ca. 350 ng. Formation of SFR complexes with Zn(2+) ions was confirmed by EQCM and UV methods. Cyclic voltammetric method has been elaborated for SFR determination and compared with UV method.

Determination of selected trace elements in herbs and their infusions.

The following macro- and microelements were determined in the leaf of peppermint (Mentha piperitae folium) and nettle (Urticae folium) (as tea bags) and in their infusions: As, Ba, Ca, Cd, Co, Cr, Cu, Fe, I, Li, Mg, Mn, Ni, Pb, Se, Sn, Sr, Ti, V and Zn. The determinations were performed using inductively coupled plasma mass spectrometry (ICP-MS) and atomic absorption spectrometry (AAS). From all the determined microelements the highest content found was that of iron, 244 and 107 mg/kg in the leaf of peppermint and nettle, respectively. However, the lowest content found was that of cobalt, 0.10 and 0.08 mg/kg for the leaf of mint and nettle, respectively. The most readily water eluting elements were strontium, selenium and iodine, the most difficult ones - barium and iron.