Regulation of veterinary point-of-care testing in the European Union, the United States of America and Japan.

Point-of-care testing (POCT) is used to detect diseases and other conditions or to monitor therapeutic procedures. In veterinary medicine, POCT not only helps during the prevention, diagnosis and treatment of animal diseases but it also has a direct impact on human health by safeguarding food supplies and preventing zoonoses. Despite its importance, the regulation of the quality, safety and effectiveness of POCT products is rarely discussed. This review reveals that the level of regulatory surveillance of veterinary POCT products in the European Union (EU), the United States of America and Japan is strikingly different, ranging from no regulation (EU) to comprehensive regulation, which is comparable to the procedures for the regulation of human in vitro medical devices (Japan). Details about the licensing procedures in these three locations, discussion of their strengths and weaknesses, and suggestions for possible future development of the regulation of these products are also provided.

Les tests utilisables sur le lieu d'intervention (POCT) permettent de détecter des maladies ou d'autres affections et d'assurer un suivi des procédures thérapeutiques appliquées. En médecine vétérinaire, les POCT apportent un soutien utile lors de la prévention, du diagnostic et du traitement des maladies animales, en plus d'avoir un impact direct sur la santé humaine en participant à la sécurité de l'approvisionnement alimentaire et à la prévention des zoonoses. La réglementation applicable aux produits utilisés pour les POCT, afin d'en garantir la qualité, la sécurité et l'efficacité, fait rarement l'objet de débats, bien qu'il s'agisse d'une question importante. L'examen présenté par les auteurs révèle que le niveau de la surveillance réglementaire exercée sur les produits vétérinaires utilisés pour les POCT dans l'Union européenne (UE), les états-Unis d'Amérique et le Japon est extrêmement variable, allant d'une absence totale de réglementation (UE) à une réglementation exhaustive comparable aux procédures appliquées pour les dispositifs in vitro utilisés en médecine humaine (Japon). Les auteurs décrivent en détail les procédures d'autorisation de mise sur le marché dans ces trois pays, examinent leurs atouts et points faibles respectifs et font quelques propositions pour faire évoluer la réglementation de ces produits à l'avenir.

Las pruebas realizadas en el punto de consulta, o «pruebas de diagnóstico inmediato¼, sirven para detectar enfermedades y otros trastornos o para seguir de cerca los resultados de procedimientos terapéuticos. En medicina veterinaria, estas pruebas no solo ayudan a prevenir, diagnosticar y tratar enfermedades animales, sino que también tienen una influencia directa en la salud humana, en la medida en que permiten proteger los suministros alimentarios y prevenir zoonosis. A pesar de su importancia, rara vez se aborda el tema de la regulación de la calidad, seguridad y eficacia de los productos que contienen pruebas de diagnóstico inmediato. Los autores revelan aquí llamativas disparidades en los regímenes de vigilancia reglamentaria que aplican a las pruebas de diagnóstico veterinario inmediato la Unión Europea (UE), los Estados Unidos de América (EE.UU.) y el Japón, regímenes que van desde la inexistencia de reglamentos (UE) hasta una regulación exhaustiva comparable a los procedimientos aplicados a los dispositivos médicos in vitro (Japón). Los autores también exponen en detalle los procedimientos de obtención de licencia vigentes en estos tres contextos, examinan sus ventajas e inconvenientes y formulan propuestas para el futuro desarrollo de la reglamentación aplicada a los antedichos productos.

Investigating the antiproliferative activity of quinoline-5,8-diones and styrylquinolinecarboxylic acids on tumor cell lines.

The structure-activity relationships of new quinoline based compounds were investigated. Quinoline-5,8-dione and styrylquinoline scaffolds were used for the design of potentially active compounds. The novel analogues had comparable antiproliferative activity to cisplatin when evaluated in a bioassay against the P388 leukemia cell line. However, these compounds appeared far less efficient against SK-N-MC neuroepithelioma cells. Analogues without the 5,8-dione structure but containing the 8-carboxylic acid group were also found to induce antiproliferative activity. Hydrophobicity as measured by HPLC did not correlate with antiproliferative activity.

Using of HPLC coupled with coulometric detector for the determination of biotin in pharmaceuticals.

The method for the determination of biotin by high performance liquid chromatography (HPLC) coupled with coulometric detector is presented here. Chromatographic and detection conditions were tested. A LiChrospher 60RP-select B column (250 mm x 4 mm; 5 microm) and the mobile phase containing 0.24 mol/L aqueous solution of acetic acid and acetonitrile in the ratio 85:15 (v/v) were found as the most suitable. The flow rate was 1 mL/min and the injected volume of the sample was 20 microL. The hydrodynamic voltammogram of biotin was measured and according to obtained data the detection parameters were set--channel I 600 mV, channel II 900 mV, sensitivity 1 microA. The developed method has been validated. The calibration curve is linear in the range 15-3600 ng/mL, correlation coefficient is 0.9998, limits of detection and quantification are 5 and 15 ng/mL, respectively. Recovery of the spiked samples was 98.67% with R.S.D. 0.255% on average. The developed method has been successfully applied for determination of biotin in pharmaceutical preparations.

Use of the zirconia-based stationary phase for separation of ibuprofen and its impurities.

A new reversed-phase liquid chromatographic method using zirconia-based stationary phase was developed for determination of ibuprofen, its related compounds and its main degradation products. The chromatographic separation was successfully achieved on the Discovery Zr-PS column (150 mm x 4.6 mm i.d., 5 microm), using a mobile phase methanol-phosphate buffer (pH 4.5; 0.05 M)-tetrahydrofurane (21:74:5, v/v/v) and the flow rate 0.5 ml min(-1). The UV detection was performed in dual wavelength mode (219 and 258 nm) to detect all compounds of interest. The column temperature was set on 60 degrees C to shorten the analysis time and improve the peak symmetry. The method is simple, rapid and cuts down the amount of hazardous waste produced in the analysis. The assay is completed within 22 minutes.

Stability of ramipril in the solvents of different pH.

The stability of ramipril in the buffer solution with different pH and the influence of acid, alkaline and oxidative medium on ramipril stability were studied. The ramipril degradation products were determined by high-performance liquid chromatography (HPLC) method. Acetonitrile:sodium perchlorate was used as the mobile phase, at a flow rate of 1.0 ml/min (linear gradient elution). A Nucleosil 100-S 5 microm C18, 250 mm x 4.6 mm i.d. was utilized as stationary phase. Detection was affected spectrophotometrically at 210 nm. The drug substance was dissolved in the ammonium phosphate buffer (pH 3, 5 and 8) and these solutions were stored at 90 degrees C for 1 h. The other series of test solutions were prepared from stock solution (drug substance dissolved in solvent A of the mobile phase) by dilution in acid (0.1M HCl), alkaline (0.1M NaOH) and oxidative (hydrogen peroxide solution) medium. More then 0.2% of impurity D (ramipril-diketopiperazine) was detected in the buffer of pH 3 and pH 5. In the buffer of pH 8 there was detected more then 1% of impurity E (ramipril-diacid). No peaks for degradation products appeared in the chromatograms above limit of quantification. The alkaline medium has the greatest effect on degradation of ramipril into impurity E (more than 50%).

Lamellar body counts: a consensus on protocol.

Lamellar bodies, concentrically layered "packages" of phospholipid that represent the storage form of surfactant, can be counted in the platelet channel of most electronic cell counters. The lamellar body count has been used for more than a decade and performs as well as traditional phospholipid analysis as an assay for evaluating fetal lung maturity. It is preferable to phospholipid analysis because it is rapid, objective, and inexpensive and can be performed in any hospital laboratory. The current methodologies for specimen preparation vary widely among laboratories, most notably with respect to centrifugation, resulting in differences in maturity cutoffs used. Our goal was to establish a consensus regarding a standardized methodology for the lamellar body count. Institutions that previously had published their results with lamellar body counts were invited to contribute. The consensus of the four participating institutions includes the following: centrifugation is not a necessary step and should be abandoned, maturity is suggested by a count of 50,000/microL or greater, and immaturity is suggested by a count of 15,000/microL or lower. As the lamellar body count gains wider acceptance as a primary assay for assessing fetal lung maturity, the test must be performed uniformly and accurately, given the implications of acting on a falsely negative test resulting from improper methodology.

Amniotic fluid lamellar body count: a rapid and reliable fetal lung maturity test.

OBJECTIVE: To evaluate the lamellar body count as a predictor of fetal lung maturity. METHODS: We conducted a prospective clinical outcome study. Amniocentesis was performed for evaluation of fetal lung maturity status within 72 hours of delivery in 130 patients. A lamellar body count was performed on each specimen, and a lecithin-sphingomyelin ratio and lung phospholipid profile were performed when possible (insufficient sample or contamination in eight cases). Each infant was evaluated for evidence of respiratory distress syndrome (RDS). RESULTS: A lamellar body count exceeding 30,000/microL predicted pulmonary maturity correctly in all cases (negative predictive value 1.00). All 16 cases of RDS had counts of 30,000/microL or less. If the lamellar body count was less than 10,000/microL, the positive predictive value for RDS was 67%, and the likelihood of a mature result from chromatographic phospholipid analysis was low (one of 14, 7%). Values between 10,000-30,000/microL indicated intermediate risk (four of 39, 10%) for developing RDS. Phospholipid analysis indicated fetal lung maturity in 35 of 39 (90%) cases with lamellar body counts in the intermediate risk range. CONCLUSIONS: The lamellar body count compares favorably with traditional phospholipid testing in the prediction of fetal lung maturity. Phospholipid analysis is not needed with lamellar body counts greater than 30,000/microL or less than 10,000/microL, but may be of benefit for values in the intermediate risk range. Advantages of this test include speed, objectivity, small sample volume required, and universal availability of instrumentation.

Lamellar body number density and the prediction of respiratory distress.

The determination of amniotic fluid lamellar body number density (LBND) has recently been shown to correlate well with other established indicators of fetal lung maturity. The authors have compared the LBND with a fetal lung phospholipid profile in predicting the clinical outcome in 52 well-documented cases. If a cutoff of 30,000/microL was used to indicate fetal lung maturity, there were no false-negative results for the LBND whereas there was one for the fetal lung profile. On the other hand, this cutoff resulted in 22 false-positive results for the LBND, whereas there were only 7 false-positive results by the fetal lung profile. The number of false-positive results by the LBND can be decreased by using a separate cutoff of less than 10,000/microL to indicate high risk for development of respiratory distress, while leaving the cutoff for predicting mature lung at 30,000/microL. This resulted in only four false-positive results for the LBND; each of these were from the same patients who also had false-positive results by the fetal lung profile. Care must be taken to ensure that the particle counter used is properly calibrated and that the appropriate cutoffs for both lung maturity and immaturity are used.

[Determination of mefenoxalone in substances, tablets and biological materials]. / Stanovení mefenoxalonu v substanci, tabletách a biologickém materiálu.

For the determination of mefenoxalon in substance, tablets and biological material (blood plasma), four variants were worked out. The first of them is based on the substitution of the benzene ring with bromine with the use of an excess of bromine, which is determined ionometrically. The second variant is based on direct titration of the hydrolytic product mefenoxalon, 3-(2-methoxyphenoxy)-2-hydroxypropylamine, with the use of the ion-selective electrode of the coated-wire type with a volumetric solution of tetraphenyl-borsodium. The third variant utilizes direct measurements of UV spectra of mefenoxalon in a solution of dichlorethane. The fourth variant consists in its fluorimetric measurement in a solution of dichlorethane, or ethylacetate. A detailed discussion of advantages and disadvantages of all above-mentioned variants of determination is presented.

[Determination of sulbactam in human serum using capillary isotachophoresis]. / Stanovení sulbaktamu v lidském séru kapilární izotachoforézou.

A method of determination of sulbaktam in human serum by capillary isotachophoresis with the use of a conductivity detector was worked out. Prior to the proper analysis, a pretreatment of the sample of serum was carried out by extracting sulbaktam to butyl acetate. The total yield of the proposed analytical procedure with an extraction first stage approaches 94%. The smallest determinable amount of the sample corresponds to 1 micrograms of sulbaktam in 1 ml of serum. The reported method was employed to evaluate the samples of sera of volunteers after intravenous administration of the dosage form sulbaktam-ampicillin (VUFB) and the foreign pharmaceutical preparation Unasyn (Pfizer). Statistically insignificant differences in pharmacokinetic parameters have confirmed that the preparations are highly bio-equivalent.

Distribution of microfilariae of Onchocerca lienalis and Onchocerca gutturosa in the skin of cattle in Germany and their development in Simulium ornatum and Culicoides nubeculosus following artificial infestation.

Onchocerca microfilariae were isolated form the umbilicus and neck of 438 cow hides at the abattoir in Tübingen, F.R.G. The overall Onchocerca infection rate was 40.4%. The presence of Onchocerca lienalis and O. gutturosa microfilariae, which are difficult to distinguish by morphological criteria, was retrospectively demonstrated after artifically infesting Simulium ornatum and Culicoides nubeculosus and identifying the infective larvae recovered. Nine of 16 samples of umbilical microfilariae fed to C. nubeculosus through a latex membrane developed to O. gutturosa third stage larvae (L3). Six of seven umbilical samples injected into the thorax of S. ornatum yielded O. lienalis L3. In six infestation trials in which microfilariae were introduced both into S. ornatum and C. nubeculosus, O. lienalis L3 were recovered exclusively from simuliids, while O. gutturosa L3 developed only in midges. Of six umbilical skins tested by cross-infestation, one contained exclusively O. gutturosa microfilariae, four only O. lienalis microfilariae and one was infected with both species. Developmental success of O. lienalis microfilariae to L3 in S. ornatum following intrathoracic injection was 22% of the mean inoculum. O. gutturosa microfilariae, ingested by C. nubeculosus through a latex membrane, developed to L3 at a rate of 2.3% of the mean microfilarial uptake.

Use of cyclodextrins in isotachophoresis. VII. Resolution of structurally related and chiral phenothiazines.

Commercially available phenothiazine derivatives were used for the study of cyclodextrin complex formation by cationic isotachophoresis with alpha-, beta- and gamma-cyclodextrin and methylated analogues of beta-cyclodextrin as leading electrolyte additives. The relationships between the type of solute substituent in the 10- and/or 2-position and the stability of the created cyclodextrin complex were studied and the results were utilized for the optimization of isotachophoretic conditions suitable for the resolution of the studied phenothiazine derivatives. Successful resolution of three racemic solutes was achieved.

Determination of sulphated glycosaminoglycans by automated potentiometric titration with simple coated-wire electrodes.

A method for the determination of sulphated glycosaminoglycans is based on their precipitation with (1-ethoxycarbonyl)pentadecyltrimethylammonium bromide (Septonex), the excess of which is back-titrated with sodium tetraphenylborate. The titration is monitored by a simple coated-wire ion-selective electrode with a plasticized poly(vinyl chloride) membrane on aluminium wire. Under certain conditions the results are almost independent of the relative molecular mass of glycosaminoglycans. The method has been applied to the determination of the active ingredient in the pharmaceutical preparation, heparon injection.

Use of cyclodextrins in isotachophoresis. VI. Cyclodextrins as leading electrolyte additives for the separation of bile acids.

Cyclodextrins (CDs) and some of their methyl derivatives have been used for the optimization of the isotachophoretic separation of bile acids in aqueous electrolyte systems. The addition of heptakis(2,3,6-tri-O-methyl)-beta-cyclodextrin to the leading electrolyte proved useful for both the solubilization and the structural differentiation of the solutes studied and led to the successful separation of their mixtures. Other CDs tested, even if they gave a satisfactory solubilization effect, did not support the resolution of bile acid mixtures.

Functional phospholipid/sphingomyelin ratio: an easy to interpret indicator of fetal lung maturity.

Data are presented which show that the functional phospholipid/sphingomyelin (FPL/S) ratio is a sensitive and specific prenatal indicator of fetal respiratory status. Phospholipids included in the ratio are acetone precipitable lecithin(L), phosphatidylglycerol(PG), phosphatidylethanolamine(PE), and sphingomyelin(S). These lipids have been shown to contribute to either the lowering of surface tension or to changes in microviscosity. Both of these properties contribute independently to the physiologic effectiveness of fetal lung surfactant. Although other phospholipid profiles have been described in the literature, the FPL/S ratio accounts for the major phospholipids which contribute to lung surfactant function with the added advantage of presenting the data in the form of a single easy to interpret number. Clinical correlates are also presented for the L/S ratio, fluorescence polarization assay (using 1,6-diphenyl-3,5-hexatriene), and absorbance at 650 nanometers.

Degree of unsaturation of the fatty acid chains of phospholipids influences the fluorescence polarization: implications for evaluating fetal lung maturity.

To study the effect of fatty acid chain saturation on the fluorescence polarization assay as a measure of fetal lung maturity, we used purified phospholipids isolated from human amniotic fluid and various commercial phospholipids. We found that the fluorescence polarization value decreased as the concentration of unsaturated fatty acids increased. In contrast, the lecithin/sphingomyelin ratio increases with increasing amounts of saturated lecithin, produced as the fetal lung matures. Since only saturated lecithins are surface active, the two indices of fetal respiratory status must reflect different properties of lung surfactant.

The community deals with the child who has a handicap.

Exclusion messages, however subtle, are interwoven into the community of the child who is handicapped. The subsystems of family, religion, neighborhood, education, health care, and financial assistance agencies have good intentions but frequently communicate poorly with the child and the parents. What is meant as a help becomes a hindrance for the child who must adapt to a limitation while continuing to move toward self-esteem, self-sufficiency, and skills that will enhance productivity and employability. No one negative message will destroy a handicapped child: it is the "history of learned inferiority" that cripples the child who is handicapped. Only when able-bodied individuals within the subsystems recognize the cumulative effect of these messages will the community be responsive to the real needs of the child who has a handicap. Nurses, schooled in sensitivity for the person, should resolve to be in the vanguard in this movement, becoming ever more sensitive to the needs of the handicapped. Such a giant step will begin a fresh and long-needed approach toward understanding those needs central to the well-being of the child who resides in the community and is also handicapped.