INFOGEST inter-laboratory recommendations for assaying gastric and pancreatic lipases activities prior to in vitro digestion studies
Artikel i vetenskaplig tidskrift, 2021
In vitro digestion studies often use animal digestive enzyme extracts as substitutes of human gastric and pancreatic secretions. Pancreatin from porcine origin is thus commonly used to provide relevant pancreatic enzymes such as proteases, amylase and lipase. Rabbit gastric extracts (RGE) have been recently introduced to provide gastric lipase in addition to pepsin. Before preparing simulated gastric and pancreatic extracts with targeted enzyme activities as described in in vitro digestion protocols, it is important to determine the activities of enzyme preparations using validated methods. The purpose of this inter-laboratory study within the INFOGEST network was to test the repeatability and reproducibility of lipase assays using the pH-stat technique for measuring the activities of gastric and pancreatic lipases from various sources. Twenty-one laboratories having different pH-stat devices received the same protocol with identical batches of RGE and two pancreatin sources. Lipase assays were performed using tributyrin as a substrate and three different amounts (50, 100 and 200 µg) of each enzyme preparation. The repeatability results within individual laboratories were satisfactory with coefficients of variation (CVs) ranging from 4 to 8% regardless of the enzyme amount tested. However, the inter-laboratory variability was high (CV > 15%) compared to existing standards for bioanalytical assays. We identified and weighted the contributions to inter-laboratory variability of several parameters associated with the various pH-stat equipment used in this study (e.g. reaction vessel volume and shape, stirring mode and rate, burette volume for the automated delivery of sodium hydroxide). Based on this, we established recommendations for improving the reproducibility of lipase assays using the pH-stat technique. Defining accurate and complete recommendations on how to correctly quantify activity levels of enzyme preparations is a gateway to promising comparison of in vitro data obtained from different laboratories following the same in vitro digestion protocol.
Enzyme activity
Inhibitor
Lipolysis
Lipases
INFOGEST
Titration method
Författare
Myriam M.L. Grundy
Physiologie, Environnement et Génétique pour l'Animal et les Systèmes d'Elevage (PEGASE)
Evan Abrahamse
Nutricia Research
Wageningen University and Research
Annette Almgren
Chalmers, Biologi och bioteknik, Livsmedelsvetenskap
Marie Alminger
Chalmers, Biologi och bioteknik
Ana Andres
Universitat Politecnica de Valencia (UPV)
Renata M.C. Ariëns
Wageningen University and Research
Shanna Bastiaan-Net
Wageningen University and Research
Claire Bourlieu-Lacanal
Université de Montpellier
Institut national de recherche pour l'agriculture, l'alimentation et l'environnement (INRAE)
A. Brodkorb
Moorepark Food Research Centre
Maria Bronze
Universidade de Lisboa
Universidade NOVA de Lisboa
Instituto de Biologia Experimental e Tecnologica
Irene Comi
Norges miljø- og biovitenskapelige universitet
Leslie Couëdelo
Université de Bordeaux
D. Dupont
Institut national de recherche pour l'agriculture, l'alimentation et l'environnement (INRAE)
Annie Durand
Centre Hospitalier Lyon-Sud
Sedef N. El
Ege Universitesi
Tara Grauwet
KU Leuven
Christine Heerup
Köpenhamns universitet
Ana Heredia
Universitat Politecnica de Valencia (UPV)
Marcos R. Infantes Garcia
KU Leuven
Christian Jungnickel
Politechnika Gdańska
Ilona E. Kłosowska-Chomiczewska
Politechnika Gdańska
Marion Létisse
Centre Hospitalier Lyon-Sud
A. Macierzanka
Politechnika Gdańska
A. Mackie
University of Leeds
D. J. McClements
University of Massachusetts
O. Menard
Institut national de recherche pour l'agriculture, l'alimentation et l'environnement (INRAE)
Anne Meynier
Institut national de recherche pour l'agriculture, l'alimentation et l'environnement (INRAE)
Marie Caroline Michalski
Centre Hospitalier Lyon-Sud
Ana Isabel Mulet-Cabero
Institute of Food Research
Moorepark Food Research Centre
Anette Mullertz
Köpenhamns universitet
Francina M. Payeras Perelló
Fresenius Kabi AG
Irene Peinado
Universitat Politecnica de Valencia (UPV)
Mélina Robert
Université de Montpellier
Sébastien Secouard
Fresenius Kabi AG
Ana T. Serra
Instituto de Biologia Experimental e Tecnologica
Universidade NOVA de Lisboa
Sandra D. Silva
Instituto de Biologia Experimental e Tecnologica
Gabriel Thomassen
Wageningen University and Research
Cecilia Tullberg
Chalmers, Biologi och bioteknik, Livsmedelsvetenskap
Ingrid Undeland
Chalmers, Biologi och bioteknik, Livsmedelsvetenskap
Carole Vaysse
Université de Bordeaux
G. E. Vegarud
Norges miljø- og biovitenskapelige universitet
Sarah H.E. Verkempinck
KU Leuven
Michelle Viau
Institut national de recherche pour l'agriculture, l'alimentation et l'environnement (INRAE)
Mostafa Zahir
Wageningen University and Research
Ruojie Zhang
University of Massachusetts
F. Carriere
Aix-Marseille Université
Journal of Functional Foods
1756-4646 (ISSN)
Vol. 82 104497Ämneskategorier (SSIF 2011)
Biokemi och molekylärbiologi
Analytisk kemi
Annan veterinärmedicin
DOI
10.1016/j.jff.2021.104497