title: Size-exclusion HPLC as a sensitive and calibrationless method for complex peptide mixtures quantification.
abstract: This work describes an original methodology to quantify complex peptide mixtures by size-exclusion high-performance liquid chromatography (SE-HPLC). The methodology was first tested on simulated elutions of peptide mixtures. For this set of experiments, a good estimation of the total peptide concentration was observed (error less than 10 %). Then 30 fractions obtained by ultrafiltration of hydrolysates from two different sources were titrated by Kjeldahl or BCA analysis and analysed by SE-HPLC for an experimental validation of the methodology. Very good matchs between methods were obtained. The linear working range depends on the hydrolysate but is generally between 0.2 and 4gL(-1) (i.e. between 10 and 200μg). Moreover, the presence of organic solvents or salts in samples does not impact the accuracy of the methodology contrary to common quantification methods. Hence, the findings of this study show that total concentration of complex peptide mixture can be efficiently determinate by the proposed methodology using simple SE-HPLC analysis.
authors.affiliation: Laboratoire Réaction et Génie des Procédés, UMR-7274, plateforme SVS, 13 rue du bois de la Champelle, F-54500 Vandœuvre-lès-Nancy, France; Université de Lorraine, 2 Avenue de la forêt de Haye, F-54505 Vandœuvre-lès-Nancy, France. Electronic address: alice.bodin@univ-lorraine.fr.
authors.forename: Alice
authors.initials: A
authors.lastname: Bodin
authors.affiliation: Laboratoire Réaction et Génie des Procédés, UMR-7274, plateforme SVS, 13 rue du bois de la Champelle, F-54500 Vandœuvre-lès-Nancy, France; Université de Lorraine, 2 Avenue de la forêt de Haye, F-54505 Vandœuvre-lès-Nancy, France. Electronic address: xavier.framboisier@univ-lorraine.fr.
authors.forename: Xavier
authors.initials: X
authors.lastname: Framboisier
authors.affiliation: Laboratoire Réaction et Génie des Procédés, UMR-7274, ENSIC, 1 rue Grandville, 54001 Nancy, France; Université de Lorraine, 2 Avenue de la forêt de Haye, F-54505 Vandœuvre-lès-Nancy, France. Electronic address: dominique.alonso@univ-lorraine.fr.
authors.forename: Dominique
authors.initials: D
authors.lastname: Alonso
authors.affiliation: Laboratoire Réaction et Génie des Procédés, UMR-7274, plateforme SVS, 13 rue du bois de la Champelle, F-54500 Vandœuvre-lès-Nancy, France; Université de Lorraine, 2 Avenue de la forêt de Haye, F-54505 Vandœuvre-lès-Nancy, France. Electronic address: ivan.marc@univ-lorraine.fr.
authors.forename: Ivan
authors.initials: I
authors.lastname: Marc
authors.affiliation: Laboratoire Réaction et Génie des Procédés, UMR-7274, plateforme SVS, 13 rue du bois de la Champelle, F-54500 Vandœuvre-lès-Nancy, France; Université de Lorraine, 2 Avenue de la forêt de Haye, F-54505 Vandœuvre-lès-Nancy, France. Electronic address: romain.kapel@univ-lorraine.fr.
authors.forename: Romain
authors.initials: R
authors.lastname: Kapel
mesh_terms: D002850:Chromatography, Gel / Q000379:methods*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D016014:Linear Models; D010455:Peptides / Q000032:analysis* / Q000737:chemistry / Q000302:isolation & purification; D011492:Protein Hydrolysates / Q000032:analysis / Q000737:chemistry; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D010455:Peptides; D011492:Protein Hydrolysates
keywords: Protein hydrolysates; Quantification; Size-exclusion HPLC