BACKGROUND: Assessing vitamin A status in populations remains a high public health priority for low- and middle-income countries. However, analytical difficulties with serum retinol measurements persist in international laboratories. Nearly all participants in a Centers for Disease Control and Prevention external quality assessment program use HPLC to measure serum retinol, but round-to-round results failing to meet acceptable criteria suggest the need to provide a straightforward stable HPLC ultraviolet (UV) method that can be adopted by these laboratories to improve performance. We present a protein precipitation HPLC-UV method that measures serum retinol below the deficiency cutoff value (<0.7 μmol/L or 20 μg/dL) that is suitable for low- and middle-income countries and uses commercially available materials. METHODS: Serum (25 μL) added to retinyl acetate was precipitated with acetonitrile (125 μL) to extract retinol. Solvent-based calibration solutions required no extraction. Calibration used either single-point (50 μg/dL) or multipoint solutions (0.52-100 μg/dL). C18 column (4.6 × 100 mm) and acetonitrile with 0.1% triethylamine/water (83/17, v/v) as isocratic mobile phase (1.1 mL/min), achieved baseline separation (7 minutes). RESULTS: With only 25 μL of serum, the limit of detection was 0.52 μg/dL. Single- and multipoint calibration generated equivalent results. Over several years, between-run imprecision was ≤7.1% in multiple quality-control materials. Overall mean (CV) method bias for NIST-certified reference materials (e-series) was -0.2% (5.8%). Maximally, 180 samples were processed within 24 h. CONCLUSIONS: This method was robust and stable over years and accurately measured serum retinol with low-volume samples. Thus, it may be of interest to low- and middle-income countries and to pediatric and finger stick applications.
BACKGROUND: Assessing vitamin A status in populations remains a high public health priority for low- and middle-income countries. However, analytical difficulties with serum retinol measurements persist in international laboratories. Nearly all participants in a Centers for Disease Control and Prevention external quality assessment program use HPLC to measure serum retinol, but round-to-round results failing to meet acceptable criteria suggest the need to provide a straightforward stable HPLC ultraviolet (UV) method that can be adopted by these laboratories to improve performance. We present a protein precipitation HPLC-UV method that measures serum retinol below the deficiency cutoff value (<0.7 μmol/L or 20 μg/dL) that is suitable for low- and middle-income countries and uses commercially available materials. METHODS: Serum (25 μL) added to retinyl acetate was precipitated with acetonitrile (125 μL) to extract retinol. Solvent-based calibration solutions required no extraction. Calibration used either single-point (50 μg/dL) or multipoint solutions (0.52-100 μg/dL). C18 column (4.6 × 100 mm) and acetonitrile with 0.1% triethylamine/water (83/17, v/v) as isocratic mobile phase (1.1 mL/min), achieved baseline separation (7 minutes). RESULTS: With only 25 μL of serum, the limit of detection was 0.52 μg/dL. Single- and multipoint calibration generated equivalent results. Over several years, between-run imprecision was ≤7.1% in multiple quality-control materials. Overall mean (CV) method bias for NIST-certified reference materials (e-series) was -0.2% (5.8%). Maximally, 180 samples were processed within 24 h. CONCLUSIONS: This method was robust and stable over years and accurately measured serum retinol with low-volume samples. Thus, it may be of interest to low- and middle-income countries and to pediatric and finger stick applications.
Authors: Georg Lietz; Harold C Furr; Bryan M Gannon; Michael H Green; Marjorie Haskell; Veronica Lopez-Teros; Janet A Novotny; Amanda C Palmer; Robert M Russell; Sherry A Tanumihardjo; Carolien A Van Loo-Bouwman Journal: Food Nutr Bull Date: 2016-04-06 Impact factor: 2.069
Authors: Pavlos F Chatzimichalakis; Victoria F Samanidou; Ioannis N Papadoyannis Journal: J Chromatogr B Analyt Technol Biomed Life Sci Date: 2004-06-15 Impact factor: 3.205
Authors: Sherry A Tanumihardjo; Robert M Russell; Charles B Stephensen; Bryan M Gannon; Neal E Craft; Marjorie J Haskell; Georg Lietz; Kerry Schulze; Daniel J Raiten Journal: J Nutr Date: 2016-08-10 Impact factor: 4.798
Authors: Gretchen A Stevens; James E Bennett; Quentin Hennocq; Yuan Lu; Luz Maria De-Regil; Lisa Rogers; Goodarz Danaei; Guangquan Li; Richard A White; Seth R Flaxman; Sean-Patrick Oehrle; Mariel M Finucane; Ramiro Guerrero; Zulfiqar A Bhutta; Amarilis Then-Paulino; Wafaie Fawzi; Robert E Black; Majid Ezzati Journal: Lancet Glob Health Date: 2015-09 Impact factor: 26.763
Authors: Elise F Talsma; Hans Verhoef; Inge D Brouwer; Anne S Mburu-de Wagt; Paul J M Hulshof; Alida Melse-Boonstra Journal: BMC Med Date: 2015-02-11 Impact factor: 8.775