IntroducciÃ³n al Foro sobre CirugÃa biliar - Academia Nacional de ...

More documents

Recommendations

Info

International Journal of Research in Pharmaceutical and Biomedical Sciences ISSN: 2229-3701MATERIALS AND METHODMaterialsThe pure drug Diacerein was obtained from OrchidPharmaceutical, Chennai. Celecoxib was obtained asgift sample from Ipca Laboratories, Mumbai. Allother reagents and solvents used were of analyticalgrade.Standard solutionsA stock solution of Diacerein (1mg/ml) was preparedby accurately weighing 0.1g of Diacerein in a 100mlvolumetric flask and dissolved in 25ml of DMSO andmade up with methanol. Standard solutions withconcentration of 50, 100, 200, 500,1000,2000,5000,10000,20000 and 50000ng/ml wereprepared by serial dilution with methanol. Calibrationstandards were prepared by adding 100µl of standardsolution to 1ml of blank plasma.Stock solution of Celecoxib (1µg/ml) was prepared inmethanol.InstrumentsThe liquid chromatography mass spectrometrysystem used consisted of Shimadzu LC-20 AD seriesHPLC system coupled to an API 4000 massspectrometer (MDS SCIEX, Toronto CANADA).Thesamples were injected by a rheodyne injector with20ul loop .All weights were taken on an electronicbalance (Shimadzu,Japan).Chromatographic system and conditionsChromatography was performed on C18, (4.6mm i.d.×50mm) analytical column and operated at 40 0 C.The mobile phase was comprised of acetonitrile:1mM ammonium formate (70:30, v/v) and separationwas carried out isocratically at a flow-rate of400µL/min. The auto sampler temperature was at4°C.Mass Spectrophotometric detection was done atESI Negative mode.Method developmentThe purpose of the present study was to develop andvalidate an LC-MS method for determining Diacereinin rabbit plasma using Celecoxib as the internalstandard. The method involves the extraction of drugform plasma followed by chromatographic separationand detection. Liquid -liquid extraction of Diacereinand internal standard from plasma samples wereperformed with acetonitrile. To 100 µl of plasmaaliquot, internal standard solution (10 µl of workingstock) equivalent to 10ng was added and mixed for30 sec on a cyclomixer followed by extraction with 2ml of acetonitrile. The mixture was vortexed for 2min, followed by centrifugation for 4 min at 3200rpm. The organic layer (1.8 ml) was separated andevaporated to dryness at 50°C using a gentle streamof nitrogen. The residue was reconstituted in 200 µlof the mobile phase and 10 µl were injected onto theanalytical column. Separation was carried outisocratically in C18 analytical column at a flow rateof 400µl/min at 40°C using acetonitrile: 1mMammonium formate (70:30 v/v) as mobile phase.Column effluent was introduced into the massspectrometer.Construction of calibration curveCalibration curve standards were prepared bydissolving appropriate amount of Diacerein in blankrabbit plasma. The plasma standards ranged from 5 -5000ng/ml.To 100 µl of calibration standard, internalstandard solution equivalent to 10µl was added andmixed for 30 sec on a cyclomixer followed byextraction with 2 ml of acetonitrile. The mixture wasvortexed for 2 min, followed by centrifugation for 4min at 3200 rpm. The organic layer (1.8 ml) wasseparated and evaporated to dryness at 50°C using agentle stream of nitrogen. The residue wasreconstituted in 200 µl of the mobile phase and 10 µlwere injected onto the analytical column. Thecalibration curve was acquired by plotting the ratio ofpeak area of Diacerein to that of IS against thenominal concentration of calibration standards. Thefinal concentrations of calibration standards obtainedfor plotting the calibration curve were 5, 10, 20, 50,100, 200, 500, 1000, 2000 and 5000 ng/ml.Determination of diacerein in spiked rabbitplasmaTo 100 µl of plasma spiked with Diacerein, internalstandard solution equivalent to 10µl was added andmixed for 30 sec on a cyclomixer followed byextraction with 2 ml of acetonitrile. The mixture wasvortexed for 2 min, followed by centrifugation for 4min at 3200 rpm. The organic layer (1.8 ml) wasseparated and evaporated to dryness at 50°C using agentle stream of nitrogen. The residue wasreconstituted in 200 µl of the mobile phase and 10 µlwere injected onto the analytical column. Linearregression equation was used for quantitation.Application to bioavailabilitystudyNormal healthy rabbits (2.0±0.2kg) were used in thisstudy. Food was withdrawn during the entire periodof experimentation, however water was available adlibitum at all times during the experiment. Rabbit wasadministered with Diacerein 50mg/kg body weightorally, followed by distilled water to ensure correctdose of administration .Venous blood sample 1mlwere collected from the marginal ear vein intoheparinised centrifuge tubes before drugadministration and at 1, 2, 3, 4, 5 and 6 hours afterdrug administration. All blood samples wereVol. 3 (4) Oct – Dec 2012 www.ijrpbsonline.com 1739
Foro Razetti: Cirugía biliar. Introducciónde las vías biliares que le daría colangiografía, pero al ligar y seccionarla arteria cística y el cístico, siguió extirpando la vesícula con latécnica retrógrada, pero se dio cuenta que había seccionado un hepáticoderecho accesorio de suficiente calibre como para no ligarlo y quedesembocaba en el cuerpo de la vesícula biliar. Ante la sorpresa de losvisitantes, les demostró su gran experiencia en esta cirugía y con todala tranquilidad que el caso ameritaba, practicó una anastomosis términolateral del hepático derecho accesorio seccionado al hepático comúny dejo un tubo en “T”. Relato esto porque lo que vimos demostraba laveracidad de mi recomendación en la publicación de mi trabajo sobreaccidentes operatorios en cirugías de vías biliares y sus solucionesinmediatas (6), donde pido a los colegas cirujanos de vías biliares, noproceder a extirpar la vesícula, si no han visto las placas radiográficasde la colangiografía, salvo que estén utilizando un equipo portátil derayos X con intensificador de imágenes, si quieren evitar lo relatado.Como ustedes comprenderán, éste fue uno de los mejores aprendizajes,imborrables, del curso que realizábamos. Por supuesto, que hoy en díacon los avances y experiencia de la colangiografía endoscópicaretrógrada preoperatoria, no disponible en todos los sitios donde seopera cirugía biliar, se puede conocer, previamente a la operación, laanatomía de todo el árbol biliar y algunas patologías del mismo, perono olvidar que la colangiografía de Mirizzi, todavía sigue siendo útil.Se comentó en esta misma ocasión (3), lo que decía el profesorMallet-Guy sobre la cirugía biliar “El cirujano que no ha cometidoaccidentes operatorios en cirugía biliar, es porque no ha operadosuficientes casos todavía”.Esto es muy cierto, pero debemos recordar lo que opinaban otroscirujanos eminentes de vías biliares, sobre la necesidad de nocometerlos y se expresaban en la siguiente forma:Mouchet y col. (9): “Herir la vía biliar principal es un pecadovenial, herirla y no repararla es un pecado mortal”.Finochietto y Markman (10): “Herida de la vía biliar principalrepararla enseguida. Herida ignorada, muerte asegurada”.Lasala y Molmenti (11): “Quien hiere la vía biliar principal y noEspinoza León L 431
Page 4 and 5: Colección Razetti. Volumen I.la re
Page 6 and 7: Colección Razetti. Volumen I.Figur
Page 8 and 9: Colección Razetti. Volumen I.papil
Page 14: Colección Razetti. Volumen I.REFER

IntroducciÃ³n al Foro sobre CirugÃ­a biliar - Academia Nacional de ...

You also want an ePaper? Increase the reach of your titles

Delete template?

Save as template?

IntroducciÃ³n al Foro sobre CirugÃa biliar - Academia Nacional de ...