Cargando…
Validation of a Simple HPLC-Based Method for Lysine Quantification for Ruminant Nutrition
Robust and selective quantification methods are required to better analyze feed supplementation effectiveness with specific amino acids. In this work, a reversed-phase high-performance liquid chromatography method with fluorescence detection is proposed and validated for lysine quantification, one o...
Autores principales: | , , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2021
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8304843/ https://www.ncbi.nlm.nih.gov/pubmed/34299448 http://dx.doi.org/10.3390/molecules26144173 |
_version_ | 1783727432493170688 |
---|---|
author | Albuquerque, João Casal, Susana Cruz, Rebeca Van Dorpe, Ingrid Maia, Margarida Rosa Garcez Fonseca, António José Mira Cabrita, Ana Rita Jordão Neves, Ana Rute Reis, Salette |
author_facet | Albuquerque, João Casal, Susana Cruz, Rebeca Van Dorpe, Ingrid Maia, Margarida Rosa Garcez Fonseca, António José Mira Cabrita, Ana Rita Jordão Neves, Ana Rute Reis, Salette |
author_sort | Albuquerque, João |
collection | PubMed |
description | Robust and selective quantification methods are required to better analyze feed supplementation effectiveness with specific amino acids. In this work, a reversed-phase high-performance liquid chromatography method with fluorescence detection is proposed and validated for lysine quantification, one of the most limiting amino acids in ruminant nutrition and essential towards milk production. To assess and widen method applicability, different matrices were considered: namely Li(2)CO(3) buffer (the chosen standard reaction buffer), phosphate buffer solution (to mimic media in cellular studies), and rumen inoculum. The method was validated for all three matrices and found to be selective, accurate (92% ± 2%), and precise at both the inter- and intra-day levels in concentrations up to 225 µM, with detection and quantification limits lower than 1.24 and 4.14 µM, respectively. Sample stability was evaluated when stored at room temperature, 4 °C, and −20 °C, showing consistency for up to 48 h regardless of the matrix. Finally, the developed method was applied in the quantification of lysine on real samples. The results presented indicate that the proposed method can be applied towards free lysine quantification in ruminant feeding studies and potentially be of great benefit to dairy cow nutrition supplementation and optimization. |
format | Online Article Text |
id | pubmed-8304843 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-83048432021-07-25 Validation of a Simple HPLC-Based Method for Lysine Quantification for Ruminant Nutrition Albuquerque, João Casal, Susana Cruz, Rebeca Van Dorpe, Ingrid Maia, Margarida Rosa Garcez Fonseca, António José Mira Cabrita, Ana Rita Jordão Neves, Ana Rute Reis, Salette Molecules Article Robust and selective quantification methods are required to better analyze feed supplementation effectiveness with specific amino acids. In this work, a reversed-phase high-performance liquid chromatography method with fluorescence detection is proposed and validated for lysine quantification, one of the most limiting amino acids in ruminant nutrition and essential towards milk production. To assess and widen method applicability, different matrices were considered: namely Li(2)CO(3) buffer (the chosen standard reaction buffer), phosphate buffer solution (to mimic media in cellular studies), and rumen inoculum. The method was validated for all three matrices and found to be selective, accurate (92% ± 2%), and precise at both the inter- and intra-day levels in concentrations up to 225 µM, with detection and quantification limits lower than 1.24 and 4.14 µM, respectively. Sample stability was evaluated when stored at room temperature, 4 °C, and −20 °C, showing consistency for up to 48 h regardless of the matrix. Finally, the developed method was applied in the quantification of lysine on real samples. The results presented indicate that the proposed method can be applied towards free lysine quantification in ruminant feeding studies and potentially be of great benefit to dairy cow nutrition supplementation and optimization. MDPI 2021-07-09 /pmc/articles/PMC8304843/ /pubmed/34299448 http://dx.doi.org/10.3390/molecules26144173 Text en © 2021 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Albuquerque, João Casal, Susana Cruz, Rebeca Van Dorpe, Ingrid Maia, Margarida Rosa Garcez Fonseca, António José Mira Cabrita, Ana Rita Jordão Neves, Ana Rute Reis, Salette Validation of a Simple HPLC-Based Method for Lysine Quantification for Ruminant Nutrition |
title | Validation of a Simple HPLC-Based Method for Lysine Quantification for Ruminant Nutrition |
title_full | Validation of a Simple HPLC-Based Method for Lysine Quantification for Ruminant Nutrition |
title_fullStr | Validation of a Simple HPLC-Based Method for Lysine Quantification for Ruminant Nutrition |
title_full_unstemmed | Validation of a Simple HPLC-Based Method for Lysine Quantification for Ruminant Nutrition |
title_short | Validation of a Simple HPLC-Based Method for Lysine Quantification for Ruminant Nutrition |
title_sort | validation of a simple hplc-based method for lysine quantification for ruminant nutrition |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8304843/ https://www.ncbi.nlm.nih.gov/pubmed/34299448 http://dx.doi.org/10.3390/molecules26144173 |
work_keys_str_mv | AT albuquerquejoao validationofasimplehplcbasedmethodforlysinequantificationforruminantnutrition AT casalsusana validationofasimplehplcbasedmethodforlysinequantificationforruminantnutrition AT cruzrebeca validationofasimplehplcbasedmethodforlysinequantificationforruminantnutrition AT vandorpeingrid validationofasimplehplcbasedmethodforlysinequantificationforruminantnutrition AT maiamargaridarosagarcez validationofasimplehplcbasedmethodforlysinequantificationforruminantnutrition AT fonsecaantoniojosemira validationofasimplehplcbasedmethodforlysinequantificationforruminantnutrition AT cabritaanaritajordao validationofasimplehplcbasedmethodforlysinequantificationforruminantnutrition AT nevesanarute validationofasimplehplcbasedmethodforlysinequantificationforruminantnutrition AT reissalette validationofasimplehplcbasedmethodforlysinequantificationforruminantnutrition |