Loading…

An optimized protocol to assess trypsin activity in biological samples

Trypsin is an enzyme that facilitates digestion. It is found in the small intestine and may also be synthesized by bacteria, plants, and fungi. However, it is mainly synthesized for commercial use from cattle pancreases. Serum trypsin determination seems to be a specific diagnostic for acute pancrea...

Full description

Saved in:
Bibliographic Details
Published in:Monatshefte für Chemie 2023-02, Vol.154 (2), p.267-277
Main Authors: Hadwan, Mahmoud Hussein, Al-Obaidy, Saba S. M., Al-Kawaz, Hawraa Saad, Almashhedy, Lamia A., Kadhum, Mohammed A., Khudhair, Dunia Abbas, Hadwan, Asad M., Hadwan, Muntadher M.
Format: Article
Language:English
Subjects:
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
cited_by cdi_FETCH-LOGICAL-c319t-fbd58e0efe05a1c5e5af523f3706cd95c5ef52a80290c3e42b102540207179cd3
cites cdi_FETCH-LOGICAL-c319t-fbd58e0efe05a1c5e5af523f3706cd95c5ef52a80290c3e42b102540207179cd3
container_end_page 277
container_issue 2
container_start_page 267
container_title Monatshefte für Chemie
container_volume 154
creator Hadwan, Mahmoud Hussein
Al-Obaidy, Saba S. M.
Al-Kawaz, Hawraa Saad
Almashhedy, Lamia A.
Kadhum, Mohammed A.
Khudhair, Dunia Abbas
Hadwan, Asad M.
Hadwan, Muntadher M.
description Trypsin is an enzyme that facilitates digestion. It is found in the small intestine and may also be synthesized by bacteria, plants, and fungi. However, it is mainly synthesized for commercial use from cattle pancreases. Serum trypsin determination seems to be a specific diagnostic for acute pancreatitis. Therefore, the development of improved trypsin activity protocols is essential for clinical pathology and pharmaceutical research. In this study, a simple spectrophotometric procedure for trypsin, a pancreatic protease, was developed. The current protocol utilized a previously known peroxidase-like reaction to estimate the trypsin activity. An additional acidification step is employed to improve the efficiency of the protocol. Trypsin has the ability to preferentially cleave the natural compound cytochrome c into heme-peptide fragments. In our study, the resulting peroxidase-like activity was catalyzed by the oxidation of 3,3ʹ,5,5ʹ-tetramethylbenzidine in the presence of H 2 O 2 . Sulfuric acid was added to stop the enzymatic reaction before recording the absorbance at 450 nm. To optimize the formation of the end product, we used the response surface methodology to apply the Box–Behnken design to assess the assay's precision. The reliability of this new method was compared to a Bland–Altman plot analysis of trypsin activity in matched samples using the standard procedure. The protocol allowed for trypsin investigations in the 5–1500 ng/cm 3 range, with a detection limit of 0.691 ng/cm 3 . The protocol demonstrated higher accuracy in the measurement of 500 ng/cm 3 trypsin solution, with a relative standard percentage error of 1.4–2.2%. This new protocol was verified against a Bland–Altman plot analysis of trypsin activity in matched samples using the Thiamine–Trypsin test, confirming its potential for application in pharmaceutical development and disease treatment. Our study demonstrated a simple, rapid, low-cost, sensitive, and selective method for assessment of the trypsin enzyme, which can be used to study the clinical importance and pharmaceutical significance of the trypsin enzyme. Graphical abstract
doi_str_mv 10.1007/s00706-022-03028-1
format article
fullrecord <record><control><sourceid>proquest_cross</sourceid><recordid>TN_cdi_proquest_journals_2776071429</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><sourcerecordid>2776071429</sourcerecordid><originalsourceid>FETCH-LOGICAL-c319t-fbd58e0efe05a1c5e5af523f3706cd95c5ef52a80290c3e42b102540207179cd3</originalsourceid><addsrcrecordid>eNp9UE1LAzEQDaJgrf4BTwHP0clXt3ssxVqh4EXPIc1mS8p2s2ZSof56oyt48zIzb3jvzfAIueVwzwGqBywFZgyEYCBBzBk_IxOupGJKVfqcTADEjNVCVZfkCnEPBSuQE7Ja9DQOORzCp2_okGKOLnY0R2oRPSLN6TRg6Kl1OXyEfKJl3obYxV1wtqNoD0Pn8ZpctLZDf_Pbp-Rt9fi6XLPNy9PzcrFhTvI6s3bb6LkH33rQljvttW21kK0sz7um1mVTsJ2DqMFJr8SWg9AKBFS8ql0jp-Ru9C2fvh89ZrOPx9SXk0ZU1azQlKgLS4wslyJi8q0ZUjjYdDIczHdeZszLlLzMT16GF5EcRVjI_c6nP-t_VF959G2R</addsrcrecordid><sourcetype>Aggregation Database</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>2776071429</pqid></control><display><type>article</type><title>An optimized protocol to assess trypsin activity in biological samples</title><source>Springer Nature</source><creator>Hadwan, Mahmoud Hussein ; Al-Obaidy, Saba S. M. ; Al-Kawaz, Hawraa Saad ; Almashhedy, Lamia A. ; Kadhum, Mohammed A. ; Khudhair, Dunia Abbas ; Hadwan, Asad M. ; Hadwan, Muntadher M.</creator><creatorcontrib>Hadwan, Mahmoud Hussein ; Al-Obaidy, Saba S. M. ; Al-Kawaz, Hawraa Saad ; Almashhedy, Lamia A. ; Kadhum, Mohammed A. ; Khudhair, Dunia Abbas ; Hadwan, Asad M. ; Hadwan, Muntadher M.</creatorcontrib><description>Trypsin is an enzyme that facilitates digestion. It is found in the small intestine and may also be synthesized by bacteria, plants, and fungi. However, it is mainly synthesized for commercial use from cattle pancreases. Serum trypsin determination seems to be a specific diagnostic for acute pancreatitis. Therefore, the development of improved trypsin activity protocols is essential for clinical pathology and pharmaceutical research. In this study, a simple spectrophotometric procedure for trypsin, a pancreatic protease, was developed. The current protocol utilized a previously known peroxidase-like reaction to estimate the trypsin activity. An additional acidification step is employed to improve the efficiency of the protocol. Trypsin has the ability to preferentially cleave the natural compound cytochrome c into heme-peptide fragments. In our study, the resulting peroxidase-like activity was catalyzed by the oxidation of 3,3ʹ,5,5ʹ-tetramethylbenzidine in the presence of H 2 O 2 . Sulfuric acid was added to stop the enzymatic reaction before recording the absorbance at 450 nm. To optimize the formation of the end product, we used the response surface methodology to apply the Box–Behnken design to assess the assay's precision. The reliability of this new method was compared to a Bland–Altman plot analysis of trypsin activity in matched samples using the standard procedure. The protocol allowed for trypsin investigations in the 5–1500 ng/cm 3 range, with a detection limit of 0.691 ng/cm 3 . The protocol demonstrated higher accuracy in the measurement of 500 ng/cm 3 trypsin solution, with a relative standard percentage error of 1.4–2.2%. This new protocol was verified against a Bland–Altman plot analysis of trypsin activity in matched samples using the Thiamine–Trypsin test, confirming its potential for application in pharmaceutical development and disease treatment. Our study demonstrated a simple, rapid, low-cost, sensitive, and selective method for assessment of the trypsin enzyme, which can be used to study the clinical importance and pharmaceutical significance of the trypsin enzyme. Graphical abstract</description><identifier>ISSN: 0026-9247</identifier><identifier>EISSN: 1434-4475</identifier><identifier>DOI: 10.1007/s00706-022-03028-1</identifier><language>eng</language><publisher>Vienna: Springer Vienna</publisher><subject>Acidification ; Analytical Chemistry ; Biological activity ; Biological properties ; Chemistry ; Chemistry and Materials Science ; Chemistry/Food Science ; Cytochromes ; Enzymes ; Hydrogen peroxide ; Inorganic Chemistry ; Intestine ; Organic Chemistry ; Original Paper ; Oxidation ; Peroxidase ; Pharmaceuticals ; Physical Chemistry ; Reliability analysis ; Response surface methodology ; Spectrophotometry ; Sulfuric acid ; Synthesis ; Theoretical and Computational Chemistry ; Thiamine ; Trypsin</subject><ispartof>Monatshefte für Chemie, 2023-02, Vol.154 (2), p.267-277</ispartof><rights>Springer-Verlag GmbH Austria, part of Springer Nature 2023. Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c319t-fbd58e0efe05a1c5e5af523f3706cd95c5ef52a80290c3e42b102540207179cd3</citedby><cites>FETCH-LOGICAL-c319t-fbd58e0efe05a1c5e5af523f3706cd95c5ef52a80290c3e42b102540207179cd3</cites><orcidid>0000-0003-1958-7764</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,27924,27925</link.rule.ids></links><search><creatorcontrib>Hadwan, Mahmoud Hussein</creatorcontrib><creatorcontrib>Al-Obaidy, Saba S. M.</creatorcontrib><creatorcontrib>Al-Kawaz, Hawraa Saad</creatorcontrib><creatorcontrib>Almashhedy, Lamia A.</creatorcontrib><creatorcontrib>Kadhum, Mohammed A.</creatorcontrib><creatorcontrib>Khudhair, Dunia Abbas</creatorcontrib><creatorcontrib>Hadwan, Asad M.</creatorcontrib><creatorcontrib>Hadwan, Muntadher M.</creatorcontrib><title>An optimized protocol to assess trypsin activity in biological samples</title><title>Monatshefte für Chemie</title><addtitle>Monatsh Chem</addtitle><description>Trypsin is an enzyme that facilitates digestion. It is found in the small intestine and may also be synthesized by bacteria, plants, and fungi. However, it is mainly synthesized for commercial use from cattle pancreases. Serum trypsin determination seems to be a specific diagnostic for acute pancreatitis. Therefore, the development of improved trypsin activity protocols is essential for clinical pathology and pharmaceutical research. In this study, a simple spectrophotometric procedure for trypsin, a pancreatic protease, was developed. The current protocol utilized a previously known peroxidase-like reaction to estimate the trypsin activity. An additional acidification step is employed to improve the efficiency of the protocol. Trypsin has the ability to preferentially cleave the natural compound cytochrome c into heme-peptide fragments. In our study, the resulting peroxidase-like activity was catalyzed by the oxidation of 3,3ʹ,5,5ʹ-tetramethylbenzidine in the presence of H 2 O 2 . Sulfuric acid was added to stop the enzymatic reaction before recording the absorbance at 450 nm. To optimize the formation of the end product, we used the response surface methodology to apply the Box–Behnken design to assess the assay's precision. The reliability of this new method was compared to a Bland–Altman plot analysis of trypsin activity in matched samples using the standard procedure. The protocol allowed for trypsin investigations in the 5–1500 ng/cm 3 range, with a detection limit of 0.691 ng/cm 3 . The protocol demonstrated higher accuracy in the measurement of 500 ng/cm 3 trypsin solution, with a relative standard percentage error of 1.4–2.2%. This new protocol was verified against a Bland–Altman plot analysis of trypsin activity in matched samples using the Thiamine–Trypsin test, confirming its potential for application in pharmaceutical development and disease treatment. Our study demonstrated a simple, rapid, low-cost, sensitive, and selective method for assessment of the trypsin enzyme, which can be used to study the clinical importance and pharmaceutical significance of the trypsin enzyme. Graphical abstract</description><subject>Acidification</subject><subject>Analytical Chemistry</subject><subject>Biological activity</subject><subject>Biological properties</subject><subject>Chemistry</subject><subject>Chemistry and Materials Science</subject><subject>Chemistry/Food Science</subject><subject>Cytochromes</subject><subject>Enzymes</subject><subject>Hydrogen peroxide</subject><subject>Inorganic Chemistry</subject><subject>Intestine</subject><subject>Organic Chemistry</subject><subject>Original Paper</subject><subject>Oxidation</subject><subject>Peroxidase</subject><subject>Pharmaceuticals</subject><subject>Physical Chemistry</subject><subject>Reliability analysis</subject><subject>Response surface methodology</subject><subject>Spectrophotometry</subject><subject>Sulfuric acid</subject><subject>Synthesis</subject><subject>Theoretical and Computational Chemistry</subject><subject>Thiamine</subject><subject>Trypsin</subject><issn>0026-9247</issn><issn>1434-4475</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2023</creationdate><recordtype>article</recordtype><recordid>eNp9UE1LAzEQDaJgrf4BTwHP0clXt3ssxVqh4EXPIc1mS8p2s2ZSof56oyt48zIzb3jvzfAIueVwzwGqBywFZgyEYCBBzBk_IxOupGJKVfqcTADEjNVCVZfkCnEPBSuQE7Ja9DQOORzCp2_okGKOLnY0R2oRPSLN6TRg6Kl1OXyEfKJl3obYxV1wtqNoD0Pn8ZpctLZDf_Pbp-Rt9fi6XLPNy9PzcrFhTvI6s3bb6LkH33rQljvttW21kK0sz7um1mVTsJ2DqMFJr8SWg9AKBFS8ql0jp-Ru9C2fvh89ZrOPx9SXk0ZU1azQlKgLS4wslyJi8q0ZUjjYdDIczHdeZszLlLzMT16GF5EcRVjI_c6nP-t_VF959G2R</recordid><startdate>20230201</startdate><enddate>20230201</enddate><creator>Hadwan, Mahmoud Hussein</creator><creator>Al-Obaidy, Saba S. M.</creator><creator>Al-Kawaz, Hawraa Saad</creator><creator>Almashhedy, Lamia A.</creator><creator>Kadhum, Mohammed A.</creator><creator>Khudhair, Dunia Abbas</creator><creator>Hadwan, Asad M.</creator><creator>Hadwan, Muntadher M.</creator><general>Springer Vienna</general><general>Springer Nature B.V</general><scope>AAYXX</scope><scope>CITATION</scope><orcidid>https://orcid.org/0000-0003-1958-7764</orcidid></search><sort><creationdate>20230201</creationdate><title>An optimized protocol to assess trypsin activity in biological samples</title><author>Hadwan, Mahmoud Hussein ; Al-Obaidy, Saba S. M. ; Al-Kawaz, Hawraa Saad ; Almashhedy, Lamia A. ; Kadhum, Mohammed A. ; Khudhair, Dunia Abbas ; Hadwan, Asad M. ; Hadwan, Muntadher M.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c319t-fbd58e0efe05a1c5e5af523f3706cd95c5ef52a80290c3e42b102540207179cd3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2023</creationdate><topic>Acidification</topic><topic>Analytical Chemistry</topic><topic>Biological activity</topic><topic>Biological properties</topic><topic>Chemistry</topic><topic>Chemistry and Materials Science</topic><topic>Chemistry/Food Science</topic><topic>Cytochromes</topic><topic>Enzymes</topic><topic>Hydrogen peroxide</topic><topic>Inorganic Chemistry</topic><topic>Intestine</topic><topic>Organic Chemistry</topic><topic>Original Paper</topic><topic>Oxidation</topic><topic>Peroxidase</topic><topic>Pharmaceuticals</topic><topic>Physical Chemistry</topic><topic>Reliability analysis</topic><topic>Response surface methodology</topic><topic>Spectrophotometry</topic><topic>Sulfuric acid</topic><topic>Synthesis</topic><topic>Theoretical and Computational Chemistry</topic><topic>Thiamine</topic><topic>Trypsin</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Hadwan, Mahmoud Hussein</creatorcontrib><creatorcontrib>Al-Obaidy, Saba S. M.</creatorcontrib><creatorcontrib>Al-Kawaz, Hawraa Saad</creatorcontrib><creatorcontrib>Almashhedy, Lamia A.</creatorcontrib><creatorcontrib>Kadhum, Mohammed A.</creatorcontrib><creatorcontrib>Khudhair, Dunia Abbas</creatorcontrib><creatorcontrib>Hadwan, Asad M.</creatorcontrib><creatorcontrib>Hadwan, Muntadher M.</creatorcontrib><collection>CrossRef</collection><jtitle>Monatshefte für Chemie</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Hadwan, Mahmoud Hussein</au><au>Al-Obaidy, Saba S. M.</au><au>Al-Kawaz, Hawraa Saad</au><au>Almashhedy, Lamia A.</au><au>Kadhum, Mohammed A.</au><au>Khudhair, Dunia Abbas</au><au>Hadwan, Asad M.</au><au>Hadwan, Muntadher M.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>An optimized protocol to assess trypsin activity in biological samples</atitle><jtitle>Monatshefte für Chemie</jtitle><stitle>Monatsh Chem</stitle><date>2023-02-01</date><risdate>2023</risdate><volume>154</volume><issue>2</issue><spage>267</spage><epage>277</epage><pages>267-277</pages><issn>0026-9247</issn><eissn>1434-4475</eissn><abstract>Trypsin is an enzyme that facilitates digestion. It is found in the small intestine and may also be synthesized by bacteria, plants, and fungi. However, it is mainly synthesized for commercial use from cattle pancreases. Serum trypsin determination seems to be a specific diagnostic for acute pancreatitis. Therefore, the development of improved trypsin activity protocols is essential for clinical pathology and pharmaceutical research. In this study, a simple spectrophotometric procedure for trypsin, a pancreatic protease, was developed. The current protocol utilized a previously known peroxidase-like reaction to estimate the trypsin activity. An additional acidification step is employed to improve the efficiency of the protocol. Trypsin has the ability to preferentially cleave the natural compound cytochrome c into heme-peptide fragments. In our study, the resulting peroxidase-like activity was catalyzed by the oxidation of 3,3ʹ,5,5ʹ-tetramethylbenzidine in the presence of H 2 O 2 . Sulfuric acid was added to stop the enzymatic reaction before recording the absorbance at 450 nm. To optimize the formation of the end product, we used the response surface methodology to apply the Box–Behnken design to assess the assay's precision. The reliability of this new method was compared to a Bland–Altman plot analysis of trypsin activity in matched samples using the standard procedure. The protocol allowed for trypsin investigations in the 5–1500 ng/cm 3 range, with a detection limit of 0.691 ng/cm 3 . The protocol demonstrated higher accuracy in the measurement of 500 ng/cm 3 trypsin solution, with a relative standard percentage error of 1.4–2.2%. This new protocol was verified against a Bland–Altman plot analysis of trypsin activity in matched samples using the Thiamine–Trypsin test, confirming its potential for application in pharmaceutical development and disease treatment. Our study demonstrated a simple, rapid, low-cost, sensitive, and selective method for assessment of the trypsin enzyme, which can be used to study the clinical importance and pharmaceutical significance of the trypsin enzyme. Graphical abstract</abstract><cop>Vienna</cop><pub>Springer Vienna</pub><doi>10.1007/s00706-022-03028-1</doi><tpages>11</tpages><orcidid>https://orcid.org/0000-0003-1958-7764</orcidid></addata></record>
fulltext fulltext
identifier ISSN: 0026-9247
ispartof Monatshefte für Chemie, 2023-02, Vol.154 (2), p.267-277
issn 0026-9247
1434-4475
language eng
recordid cdi_proquest_journals_2776071429
source Springer Nature
subjects Acidification
Analytical Chemistry
Biological activity
Biological properties
Chemistry
Chemistry and Materials Science
Chemistry/Food Science
Cytochromes
Enzymes
Hydrogen peroxide
Inorganic Chemistry
Intestine
Organic Chemistry
Original Paper
Oxidation
Peroxidase
Pharmaceuticals
Physical Chemistry
Reliability analysis
Response surface methodology
Spectrophotometry
Sulfuric acid
Synthesis
Theoretical and Computational Chemistry
Thiamine
Trypsin
title An optimized protocol to assess trypsin activity in biological samples
url http://sfxeu10.hosted.exlibrisgroup.com/loughborough?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2024-12-25T04%3A39%3A30IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-proquest_cross&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=An%20optimized%20protocol%20to%20assess%20trypsin%20activity%20in%20biological%20samples&rft.jtitle=Monatshefte%20f%C3%BCr%20Chemie&rft.au=Hadwan,%20Mahmoud%20Hussein&rft.date=2023-02-01&rft.volume=154&rft.issue=2&rft.spage=267&rft.epage=277&rft.pages=267-277&rft.issn=0026-9247&rft.eissn=1434-4475&rft_id=info:doi/10.1007/s00706-022-03028-1&rft_dat=%3Cproquest_cross%3E2776071429%3C/proquest_cross%3E%3Cgrp_id%3Ecdi_FETCH-LOGICAL-c319t-fbd58e0efe05a1c5e5af523f3706cd95c5ef52a80290c3e42b102540207179cd3%3C/grp_id%3E%3Coa%3E%3C/oa%3E%3Curl%3E%3C/url%3E&rft_id=info:oai/&rft_pqid=2776071429&rft_id=info:pmid/&rfr_iscdi=true