RESUMEN
An anti-infective bilayer implant coating with selectively activatable properties was developed to prevent biofilm formation and to support the treatment of periprosthetic infection as a local adjunct to current treatment concepts. In a first step, Ti6Al4V discs were coated with a permanent layer of Poly(l-lactide) (PLLA) including silver ions. The PLLA could be optionally released by the application of extracorporeal shock waves. In a second step, a resorbable layer of triglyceride (TAG) with incorporated antibiotics was applied. The second layer is designed for resorption within weeks. Prior to approval and clinical application, a comprehensive evaluation process to determine mechanical/physical and microbiological properties is obligate. To date, none of the existing test standards covers both drug-releasing and activatable coatings for orthopedic implants. Therefore, a comprehensive test concept was developed to characterize the new coating in a pilot series. The coatings were homogeneously applied on the Ti6Al4V substrate, resulting in an adhesion strength sufficient for non-articulating surfaces for PLLA. Proof of the extracorporeal shockwave activation of PLLA was demonstrated both mechanically and microbiologically, with a simultaneous increase of biocompatibility compared to standard electroplated silver coating. Wettability was significantly reduced for both layers in comparison to the Ti6Al4V substrate. Thus, potentially inhibiting biofilm formation. Furthermore, the TAG coating promoted cell proliferation and bacterial eradication. In conclusion, the testing concept is applicable for similar biopolymer coating systems. Furthermore, the extracorporeal activation could represent a completely new supportive approach for the treatment of periprosthetic joint infections.
Asunto(s)
Materiales Biocompatibles Revestidos , Plata , Biopolímeros/farmacología , Materiales Biocompatibles Revestidos/farmacología , Ensayo de Materiales , Prótesis e Implantes , Plata/farmacologíaRESUMEN
BACKGROUND: Simultaneous measurement of testosterone (T) and 5α-dihydrotestosterone (DHT) is important for diagnosing androgen deficiency states and hyperandrogenism in males and females, respectively. However, immunoassays used for T and DHT determination suffer from inadequate specificity and sensitivity, while tandem mass spectrometry is expensive and demanding in use. METHODS AND RESULTS: We developed a selective gas chromatography-mass spectrometry (GC-MS) method for parallel T and DHT measurement. The assay showed a linear response up to 46.5nmol/L, intra- and interassay imprecision and inaccuracy <15% and recoveries in spiked samples >90% for both analytes. The limit of quantitation was 0.117nmol/L for T and 0.168nmol/L for DHT. Comparison with immunoassays revealed good agreement for T in males, but a bias in favour of immunoassays at low concentrations for T in females and DHT in both sexes. We established reference ranges for T and DHT and suggest interval partitioning for T according to age in men and menstrual cycle in women. Assay validation in a clinical setting suggests that measuring DHT or T/DHT ratio may help identify patients with polycystic ovary syndrome. CONCLUSION: We developed a selective, simple and inexpensive GC-MS method for parallel measurement of T and DHT with potential use in the clinical laboratory.
Asunto(s)
Dihidrotestosterona/sangre , Testosterona/sangre , Adulto , Femenino , Cromatografía de Gases y Espectrometría de Masas , Humanos , Masculino , Adulto JovenRESUMEN
Niemann-Pick type C (NPC) is a neurological disease caused by an intracellular cholesterol accumulation. Cholesterol oxidation product cholestane-3ß,5α,6ß-triol (C-triol) serves as diagnostic biomarker for NPC, but its measurement in the routine laboratory remains difficult. We developed an isotope dilution gas chromatography-mass spectrometry (GC-MS) method permitting screening for NPC in plasma. 1440 plasma samples obtained from clinically suspicious patients were subjected to alkaline saponification. C-triol was extracted with carbon tetrachloride, transformed into the trimethylsilylethers, separated on a fused silica capillary column with a nonpolar silicone stationary phase, and analyzed by GC-MS. NPC diagnosis was confirmed by DNA sequencing. The method was linear over a concentration range of 0.03-200ng/mL with a mean recovery rate of 98.6%. The intra- and inter-day variation coefficients assessed at two concentrations were below 15%. Limits of quantification (LOQ) and detection (LOD) were 0.03ng/mL and 0.01ng/mL, respectively. Receiver operating characteristic (ROC) analysis estimated that the area under curve was 0.997 implying a significant discriminatory power to identify subjects with NPC. Nevertheless, 13 NPC patients and 29 control subjects confirmed by sequencing showed false negative or positive results, respectively. Two patients with cerebrotendinous xanthomatosis showed a 5-10-fold increase in C-triol levels. We developed a quick and sensitive GC-MS method for determination of C-triol, which may serve as a simple and inexpensive diagnostic tool aiding NPC diagnosis in a routine hospital laboratory. As C-triol elevation is not limited to NPC, the NPC diagnosis has to be confirmed by DNA sequencing.
Asunto(s)
Colestanoles/sangre , Enfermedad de Niemann-Pick Tipo C/sangre , Enfermedad de Niemann-Pick Tipo C/diagnóstico , Área Bajo la Curva , Biomarcadores/sangre , Calibración , Relación Dosis-Respuesta a Droga , Cromatografía de Gases y Espectrometría de Masas , Humanos , Límite de Detección , Modelos Lineales , Curva ROC , Reproducibilidad de los Resultados , Análisis de Secuencia de ADNRESUMEN
In current analytical practice, simple methods are required for assessing the quality of a deep-frying fat after some time of use. Therefore, a new procedure was developed for the fast and selective determination of higher carbonyl compounds, i.e., those triglycerides resulting from the oxidation and/or oxidative cleavage of double bonds in unsaturated fatty acids. For analysis, a fat sample is dissolved in a 1-butanol/toluene mixture containing 6-undecanone as an internal standard. Aldehydes and ketones present are allowed to react with 2,4-dinitrophenylhydrazine (DNPH) in acidic solution for at least 1 h at room temperature. The mixture is injected directly onto a reversed phase HPLC column that is eluted with a very steep gradient between methanol and tert-butylmethyl ether (TBME) and is fitted with a UV detector set at 370 nm. In this way, the DNPH derivatives of all higher carbonyl compounds are eluted as one single peak. The result is calculated as milliequivalents of carbonyls per kg of fat (meq kg(-1)). The method takes a minimum of time and reagents and only requires the usual equipment.