Fluorescent based assessment of trypsin activity vs total proteases in the queen triggerfish, Balistes vetula

dc.contributor.advisor Uscian, John M. Puerta-Martínez, Francisco College of Arts and Sciences - Sciences en_US
dc.contributor.committee Williams, Lucy B.
dc.contributor.committee Montalvo, Rafael
dc.contributor.committee Kubaryk, John
dc.contributor.department Department of Biology en_US
dc.contributor.representative Grove, Kurt 2018-05-16T16:11:37Z 2018-05-16T16:11:37Z 2007
dc.description.abstract Amino acids that comprise proteins constitute essential nutrients for any animal organism. Indeed they are directly related to its growth and maintenance. Protein assimilation depends on the catalytic properties of proteases in the alimentary canal. Such protein-degredative enzymes include trypsin, chymotrypsin and elastase, with trypsin being the most important and abundant. The main objective of this research project was to use a highly sensitive fluorometric technique to assess both total protease and trypsin activities from intestines of the queen triggerfish, Balistes vetula, and determine if these were affected by fish size. To this end, 32 west coast and 4 east coast intestinal samples were obtained. Each sample contained from 2 – 10 total intestines. The weight (wet weight) was obtained and recorded for each sample. Samples were then individually homogenized in distilled water to produce a supernatant. Filtered supernatant was assayed for the following three properties: 1) protein content; 2) total protease activity; 3) trypsin activity. The remaining supernatant was pooled and subjected to size exclusion chromatographic procedure in order to obtain a partial purification of proteins conferring trypsin activity. The kinetic analysis of this trypsin activity was assessed as per the following four variables: 1) protein concentration; 2) pH; 3) temperature; and 4) presence of soybean trypsin inhibitor. The data are considered with respect to both the utility of the fluorometric technique in this analysis and the significance of trypsin with respect to the biochemistry/physiology of the queen triggerfish. en_US
dc.description.abstract Los aminoácidos contenidos en las proteínas son nutrientes esenciales para todo organismo animal y su asimilación depende de proteasas presentes en el tracto digestivo, tales como tripsina, quimotripsina y elastasas, siendo tripsina la más abundantes e importante. El objetivo principal de esta investigación, fue determinar mediante un sensible método fluorométrico, la actividad total de proteasas y la actividad de tripsina en el tracto intestinal del queen triggerfish, Balistes vetula y determinar si dichas actividades son afectadas por la edad de los peces. Con propósito se obtuvieron 32 muestras intestinales de la costa oeste y 4 muestras de la costa este de Puerto Rico; cada muestra conteniendo de 2 a 10 intestinos. El peso húmedo de cada muestra fue determinado y registrado. Las muestras fueron después individualmente homogenizadas en agua destilada para producir sobrenadantes (homogenatos crudos). A los sobrenadantes filtrados se les determinó 1) Contenido de proteína, 2) actividad total de proteasas y 3) actividad de tripsina. El homogenato crudo fue además sometido a cromatografía de exclusión por peso molecular, con el objetivo de obtener la purificación parcial de tripsina. La tripsina parcialmente purificada fue caracterizada cinéticamente de acuerdo a los siguientes 4 parámetros 1) actividad de tripsina, 2) pH, 3) temperatura y 4) inhibición usando el inhibidor de soya de Kunitz. Los resultados de este estudio son útiles para evaluar la utilidad del método fluorométrico usado y la importancia de tripsina con respecto a la fisiología digestiva del queen triggerfish. en_US
dc.description.graduationYear 2007 en_US
dc.description.sponsorship University of Puerto Rico and to the Biology Department (financial support to complete master’s studies) en_US
dc.language.iso en en_US
dc.rights.holder (c) 2007 Francisco Puerta-Martínez en_US
dc.rights.license All rights reserved en_US
dc.subject Amino acids en_US
dc.subject Protein assimilation en_US
dc.subject Catalytic properties of proteases en_US
dc.subject Trypsin en_US
dc.subject Chymotrypsin en_US
dc.subject Elastase en_US
dc.subject.lcsh Trypsin--Analysis. en_US
dc.subject.lcsh Digestive anzymes. en_US
dc.subject.lcsh Proteolytic enzymes. en_US
dc.subject.lcsh Trypsin inhibitors. en_US
dc.subject.lcsh Fluorimetry. en_US
dc.subject.lcsh Balistidae--Growth. en_US
dc.title Fluorescent based assessment of trypsin activity vs total proteases in the queen triggerfish, Balistes vetula en_US
dc.type Thesis en_US
dspace.entity.type Publication Biology en_US M.S. en_US
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