A colorimetric method to quantify endo-polygalacturonase activity

Autores
Torres, Sebastián; Sayago, Jorge Esteban; Ordóñez, Roxana Mabel; Isla, Maria Ines
Año de publicación
2011
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
We report a new colorimetric assay to quantify endo-polygalacturonase activity, which hydrolyzes polygalacturonic acid to produce smaller chains of galacturonate. Some of the reported polygalacturonase assays measure the activity by detecting the appearance of reducing ends such as the Somogyi–Nelson method. As a result of being general towards reducing groups, the Somogyi–Nelson method is not appropriate when studying polygalacturonase and polygalacturonase inhibitors in plant crude extracts, which often have a strong reducing power. Ruthenium Red is an inorganic dye that binds polygalacturonic acid and causes its precipitation. In the presence of polygalacturonase, polygalacturonic acid is hydrolyzed bringing about a corresponding gain in soluble Ruthenium Red. The described assay utilizes Ruthenium Red as the detection reagent which has been used previously in plate-based assays but not in liquid medium reactions. The new method measures the disappearance of the substrate polygalacturonic acid and is compared to the Somogyi–Nelson assay. The experimental results using lemon peel, a fern fronds and castor leaf crude extracts demonstrate that the new method provides a way to the quickly screening of polygalacturonase activity and polygalacturonase inhibitors in plant crude extracts containing high amounts of reducing power. On the other hand, the Ruthenium Red assay is not able to determine the activity of an exo-polygalacturonase as initial velocity and thus would allow the differentiation between endo- and exo-polygalacturonase activities.
Fil: Torres, Sebastián. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto de Química del Noroeste. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia. Instituto de Química del Noroeste; Argentina
Fil: Sayago, Jorge Esteban. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia; Argentina. Universidad Nacional de Tucumán. Facultad de Ciencias Naturales e Instituto Miguel Lillo; Argentina
Fil: Ordóñez, Roxana Mabel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto de Química del Noroeste. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia. Instituto de Química del Noroeste; Argentina. Universidad Nacional de Tucumán. Facultad de Ciencias Naturales e Instituto Miguel Lillo; Argentina
Fil: Isla, Maria Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto de Química del Noroeste. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia. Instituto de Química del Noroeste; Argentina. Universidad Nacional de Tucumán. Facultad de Ciencias Naturales e Instituto Miguel Lillo; Argentina
Materia
Endo Polygalacturonase Activity
Colorimetric Detection
Ruthenium Red
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/51930
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/51930
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling A colorimetric method to quantify endo-polygalacturonase activityTorres, SebastiánSayago, Jorge EstebanOrdóñez, Roxana MabelIsla, Maria InesEndo Polygalacturonase ActivityColorimetric DetectionRuthenium Redhttps://purl.org/becyt/ford/1.4https://purl.org/becyt/ford/1We report a new colorimetric assay to quantify endo-polygalacturonase activity, which hydrolyzes polygalacturonic acid to produce smaller chains of galacturonate. Some of the reported polygalacturonase assays measure the activity by detecting the appearance of reducing ends such as the Somogyi–Nelson method. As a result of being general towards reducing groups, the Somogyi–Nelson method is not appropriate when studying polygalacturonase and polygalacturonase inhibitors in plant crude extracts, which often have a strong reducing power. Ruthenium Red is an inorganic dye that binds polygalacturonic acid and causes its precipitation. In the presence of polygalacturonase, polygalacturonic acid is hydrolyzed bringing about a corresponding gain in soluble Ruthenium Red. The described assay utilizes Ruthenium Red as the detection reagent which has been used previously in plate-based assays but not in liquid medium reactions. The new method measures the disappearance of the substrate polygalacturonic acid and is compared to the Somogyi–Nelson assay. The experimental results using lemon peel, a fern fronds and castor leaf crude extracts demonstrate that the new method provides a way to the quickly screening of polygalacturonase activity and polygalacturonase inhibitors in plant crude extracts containing high amounts of reducing power. On the other hand, the Ruthenium Red assay is not able to determine the activity of an exo-polygalacturonase as initial velocity and thus would allow the differentiation between endo- and exo-polygalacturonase activities.Fil: Torres, Sebastián. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto de Química del Noroeste. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia. Instituto de Química del Noroeste; ArgentinaFil: Sayago, Jorge Esteban. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia; Argentina. Universidad Nacional de Tucumán. Facultad de Ciencias Naturales e Instituto Miguel Lillo; ArgentinaFil: Ordóñez, Roxana Mabel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto de Química del Noroeste. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia. Instituto de Química del Noroeste; Argentina. Universidad Nacional de Tucumán. Facultad de Ciencias Naturales e Instituto Miguel Lillo; ArgentinaFil: Isla, Maria Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto de Química del Noroeste. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia. Instituto de Química del Noroeste; Argentina. Universidad Nacional de Tucumán. Facultad de Ciencias Naturales e Instituto Miguel Lillo; ArgentinaElsevier Science Inc2011-02info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/51930Torres, Sebastián; Sayago, Jorge Esteban; Ordóñez, Roxana Mabel; Isla, Maria Ines; A colorimetric method to quantify endo-polygalacturonase activity; Elsevier Science Inc; Enzyme and Microbial Technology; 48; 2; 2-2011; 123-1280141-0229CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.1016/j.enzmictec.2010.04.004info:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/pii/S0141022910000712info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T10:20:39Zoai:ri.conicet.gov.ar:11336/51930instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-29 10:20:39.389CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv A colorimetric method to quantify endo-polygalacturonase activity
title A colorimetric method to quantify endo-polygalacturonase activity
spellingShingle A colorimetric method to quantify endo-polygalacturonase activity
Torres, Sebastián
Endo Polygalacturonase Activity
Colorimetric Detection
Ruthenium Red
title_short A colorimetric method to quantify endo-polygalacturonase activity
title_full A colorimetric method to quantify endo-polygalacturonase activity
title_fullStr A colorimetric method to quantify endo-polygalacturonase activity
title_full_unstemmed A colorimetric method to quantify endo-polygalacturonase activity
title_sort A colorimetric method to quantify endo-polygalacturonase activity
dc.creator.none.fl_str_mv Torres, Sebastián
Sayago, Jorge Esteban
Ordóñez, Roxana Mabel
Isla, Maria Ines
author Torres, Sebastián
author_facet Torres, Sebastián
Sayago, Jorge Esteban
Ordóñez, Roxana Mabel
Isla, Maria Ines
author_role author
author2 Sayago, Jorge Esteban
Ordóñez, Roxana Mabel
Isla, Maria Ines
author2_role author
author
author
dc.subject.none.fl_str_mv Endo Polygalacturonase Activity
Colorimetric Detection
Ruthenium Red
topic Endo Polygalacturonase Activity
Colorimetric Detection
Ruthenium Red
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.4
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv We report a new colorimetric assay to quantify endo-polygalacturonase activity, which hydrolyzes polygalacturonic acid to produce smaller chains of galacturonate. Some of the reported polygalacturonase assays measure the activity by detecting the appearance of reducing ends such as the Somogyi–Nelson method. As a result of being general towards reducing groups, the Somogyi–Nelson method is not appropriate when studying polygalacturonase and polygalacturonase inhibitors in plant crude extracts, which often have a strong reducing power. Ruthenium Red is an inorganic dye that binds polygalacturonic acid and causes its precipitation. In the presence of polygalacturonase, polygalacturonic acid is hydrolyzed bringing about a corresponding gain in soluble Ruthenium Red. The described assay utilizes Ruthenium Red as the detection reagent which has been used previously in plate-based assays but not in liquid medium reactions. The new method measures the disappearance of the substrate polygalacturonic acid and is compared to the Somogyi–Nelson assay. The experimental results using lemon peel, a fern fronds and castor leaf crude extracts demonstrate that the new method provides a way to the quickly screening of polygalacturonase activity and polygalacturonase inhibitors in plant crude extracts containing high amounts of reducing power. On the other hand, the Ruthenium Red assay is not able to determine the activity of an exo-polygalacturonase as initial velocity and thus would allow the differentiation between endo- and exo-polygalacturonase activities.
Fil: Torres, Sebastián. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto de Química del Noroeste. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia. Instituto de Química del Noroeste; Argentina
Fil: Sayago, Jorge Esteban. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia; Argentina. Universidad Nacional de Tucumán. Facultad de Ciencias Naturales e Instituto Miguel Lillo; Argentina
Fil: Ordóñez, Roxana Mabel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto de Química del Noroeste. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia. Instituto de Química del Noroeste; Argentina. Universidad Nacional de Tucumán. Facultad de Ciencias Naturales e Instituto Miguel Lillo; Argentina
Fil: Isla, Maria Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto de Química del Noroeste. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia. Instituto de Química del Noroeste; Argentina. Universidad Nacional de Tucumán. Facultad de Ciencias Naturales e Instituto Miguel Lillo; Argentina
description We report a new colorimetric assay to quantify endo-polygalacturonase activity, which hydrolyzes polygalacturonic acid to produce smaller chains of galacturonate. Some of the reported polygalacturonase assays measure the activity by detecting the appearance of reducing ends such as the Somogyi–Nelson method. As a result of being general towards reducing groups, the Somogyi–Nelson method is not appropriate when studying polygalacturonase and polygalacturonase inhibitors in plant crude extracts, which often have a strong reducing power. Ruthenium Red is an inorganic dye that binds polygalacturonic acid and causes its precipitation. In the presence of polygalacturonase, polygalacturonic acid is hydrolyzed bringing about a corresponding gain in soluble Ruthenium Red. The described assay utilizes Ruthenium Red as the detection reagent which has been used previously in plate-based assays but not in liquid medium reactions. The new method measures the disappearance of the substrate polygalacturonic acid and is compared to the Somogyi–Nelson assay. The experimental results using lemon peel, a fern fronds and castor leaf crude extracts demonstrate that the new method provides a way to the quickly screening of polygalacturonase activity and polygalacturonase inhibitors in plant crude extracts containing high amounts of reducing power. On the other hand, the Ruthenium Red assay is not able to determine the activity of an exo-polygalacturonase as initial velocity and thus would allow the differentiation between endo- and exo-polygalacturonase activities.
publishDate 2011
dc.date.none.fl_str_mv 2011-02
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/51930
Torres, Sebastián; Sayago, Jorge Esteban; Ordóñez, Roxana Mabel; Isla, Maria Ines; A colorimetric method to quantify endo-polygalacturonase activity; Elsevier Science Inc; Enzyme and Microbial Technology; 48; 2; 2-2011; 123-128
0141-0229
CONICET Digital
CONICET
url http://hdl.handle.net/11336/51930
identifier_str_mv Torres, Sebastián; Sayago, Jorge Esteban; Ordóñez, Roxana Mabel; Isla, Maria Ines; A colorimetric method to quantify endo-polygalacturonase activity; Elsevier Science Inc; Enzyme and Microbial Technology; 48; 2; 2-2011; 123-128
0141-0229
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.1016/j.enzmictec.2010.04.004
info:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/pii/S0141022910000712
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
application/pdf
application/pdf
dc.publisher.none.fl_str_mv Elsevier Science Inc
publisher.none.fl_str_mv Elsevier Science Inc
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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score 13.070432

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