The Improved Method for Determination of Orotic Acid in Milk by Ultra-Fast Liquid Chromatography with Optimized Photodiode Array Detection

Ultra-fast liquid chromatography (UFLC) with a photodiode array detector (DAD) for simple and rapid determination of orotic acid (OAc) in milk of sheep and cows is described. Milk samples are treated with acetonitrile (1:1, <i>v/v</i>) and then centrifuged at 4 °C. To 1 mL of the obtaine...

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	Marian Czauderna, Małgorzata Białek, Edyta Molik, Kamil Zaworski
Formato:	article
Lenguaje:	EN
Publicado:	MDPI AG 2021
Materias:	orotic acid sheep cow reversed-phase chromatography Veterinary medicine SF600-1100 Zoology QL1-991
Acceso en línea:	https://doaj.org/article/0a006baef25d4180b155e86072d92a9b
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:0a006baef25d4180b155e86072d92a9b
record_format	dspace
spelling	oai:doaj.org-article:0a006baef25d4180b155e86072d92a9b2021-11-25T16:18:49ZThe Improved Method for Determination of Orotic Acid in Milk by Ultra-Fast Liquid Chromatography with Optimized Photodiode Array Detection10.3390/ani111131962076-2615https://doaj.org/article/0a006baef25d4180b155e86072d92a9b2021-11-01T00:00:00Zhttps://www.mdpi.com/2076-2615/11/11/3196https://doaj.org/toc/2076-2615Ultra-fast liquid chromatography (UFLC) with a photodiode array detector (DAD) for simple and rapid determination of orotic acid (OAc) in milk of sheep and cows is described. Milk samples are treated with acetonitrile (1:1, <i>v/v</i>) and then centrifuged at 4 °C. To 1 mL of the obtained supernatant 9 mL of ultrapure water was added. Subsequently, 0.5–6 µL of the resulting solution was injected into the UFLC-DAD system. Separation and quantification of OAc in milk samples was achieved using two Kinetex C18 columns (1.7 µm, 150 mm × 2.1 mm, i.d., 100 Å; Phenomenex) fitted with a pre-column of 4 mm × 2 mm, i.d. (Phenomenex) containing C<sub>18</sub> packing material. All separations were performed at a column temperature of 35 °C while the ambient temperature was 21–24 °C. Satisfactory separation of OAc from endogenous species of milk can be achieved using the binary gradient elution program and UV detection at wavelengths 278 nm. Our original procedure resulted in suitable separation and quantification of OAc in milk samples; OAc eluted at 6.44 ± 0.03 min. The total run time of OAc analysis (including re-equilibration) was 27 min. As expected, the OAc peak was absent from the blank when the proposed gradient elution program and UV detection at 278 nm was used. The average recoveries of OAc standards added to milk samples were satisfactory (96.7–105.3%). The low inter-and intra-assay coefficient of variation derived from the measurements of OAc in cow and ovine milk samples (i.e., 0.784%, 1.283% and 0.710%, 1.221%, respectively) and in O-Ac standards (i.e., 0.377% and 0.294%, respectively), as well as high recoveries of OAc added to ovine and cows’ milk (~100%) and the low detection (0.04 ng) and quantification (0.12 ng) limits point to satisfactory accuracy, precision and sensitivity of the reported method. OAc concentrations in ovine milk samples were within the range from 25 to 36 mg/L, while OAc levels in cows’ milk samples was found in the range of 32–36 mg/L. Our original procedure is suitable for routine quantification of OAc in milk of ewes and cows.Marian CzaudernaMałgorzata BiałekEdyta MolikKamil ZaworskiMDPI AGarticleorotic acidsheepcowreversed-phase chromatographyVeterinary medicineSF600-1100ZoologyQL1-991ENAnimals, Vol 11, Iss 3196, p 3196 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	orotic acid sheep cow reversed-phase chromatography Veterinary medicine SF600-1100 Zoology QL1-991
spellingShingle	orotic acid sheep cow reversed-phase chromatography Veterinary medicine SF600-1100 Zoology QL1-991 Marian Czauderna Małgorzata Białek Edyta Molik Kamil Zaworski The Improved Method for Determination of Orotic Acid in Milk by Ultra-Fast Liquid Chromatography with Optimized Photodiode Array Detection
description	Ultra-fast liquid chromatography (UFLC) with a photodiode array detector (DAD) for simple and rapid determination of orotic acid (OAc) in milk of sheep and cows is described. Milk samples are treated with acetonitrile (1:1, <i>v/v</i>) and then centrifuged at 4 °C. To 1 mL of the obtained supernatant 9 mL of ultrapure water was added. Subsequently, 0.5–6 µL of the resulting solution was injected into the UFLC-DAD system. Separation and quantification of OAc in milk samples was achieved using two Kinetex C18 columns (1.7 µm, 150 mm × 2.1 mm, i.d., 100 Å; Phenomenex) fitted with a pre-column of 4 mm × 2 mm, i.d. (Phenomenex) containing C<sub>18</sub> packing material. All separations were performed at a column temperature of 35 °C while the ambient temperature was 21–24 °C. Satisfactory separation of OAc from endogenous species of milk can be achieved using the binary gradient elution program and UV detection at wavelengths 278 nm. Our original procedure resulted in suitable separation and quantification of OAc in milk samples; OAc eluted at 6.44 ± 0.03 min. The total run time of OAc analysis (including re-equilibration) was 27 min. As expected, the OAc peak was absent from the blank when the proposed gradient elution program and UV detection at 278 nm was used. The average recoveries of OAc standards added to milk samples were satisfactory (96.7–105.3%). The low inter-and intra-assay coefficient of variation derived from the measurements of OAc in cow and ovine milk samples (i.e., 0.784%, 1.283% and 0.710%, 1.221%, respectively) and in O-Ac standards (i.e., 0.377% and 0.294%, respectively), as well as high recoveries of OAc added to ovine and cows’ milk (~100%) and the low detection (0.04 ng) and quantification (0.12 ng) limits point to satisfactory accuracy, precision and sensitivity of the reported method. OAc concentrations in ovine milk samples were within the range from 25 to 36 mg/L, while OAc levels in cows’ milk samples was found in the range of 32–36 mg/L. Our original procedure is suitable for routine quantification of OAc in milk of ewes and cows.
format	article
author	Marian Czauderna Małgorzata Białek Edyta Molik Kamil Zaworski
author_facet	Marian Czauderna Małgorzata Białek Edyta Molik Kamil Zaworski
author_sort	Marian Czauderna
title	The Improved Method for Determination of Orotic Acid in Milk by Ultra-Fast Liquid Chromatography with Optimized Photodiode Array Detection
title_short	The Improved Method for Determination of Orotic Acid in Milk by Ultra-Fast Liquid Chromatography with Optimized Photodiode Array Detection
title_full	The Improved Method for Determination of Orotic Acid in Milk by Ultra-Fast Liquid Chromatography with Optimized Photodiode Array Detection
title_fullStr	The Improved Method for Determination of Orotic Acid in Milk by Ultra-Fast Liquid Chromatography with Optimized Photodiode Array Detection
title_full_unstemmed	The Improved Method for Determination of Orotic Acid in Milk by Ultra-Fast Liquid Chromatography with Optimized Photodiode Array Detection
title_sort	improved method for determination of orotic acid in milk by ultra-fast liquid chromatography with optimized photodiode array detection
publisher	MDPI AG
publishDate	2021
url	https://doaj.org/article/0a006baef25d4180b155e86072d92a9b
work_keys_str_mv	AT marianczauderna theimprovedmethodfordeterminationoforoticacidinmilkbyultrafastliquidchromatographywithoptimizedphotodiodearraydetection AT małgorzatabiałek theimprovedmethodfordeterminationoforoticacidinmilkbyultrafastliquidchromatographywithoptimizedphotodiodearraydetection AT edytamolik theimprovedmethodfordeterminationoforoticacidinmilkbyultrafastliquidchromatographywithoptimizedphotodiodearraydetection AT kamilzaworski theimprovedmethodfordeterminationoforoticacidinmilkbyultrafastliquidchromatographywithoptimizedphotodiodearraydetection AT marianczauderna improvedmethodfordeterminationoforoticacidinmilkbyultrafastliquidchromatographywithoptimizedphotodiodearraydetection AT małgorzatabiałek improvedmethodfordeterminationoforoticacidinmilkbyultrafastliquidchromatographywithoptimizedphotodiodearraydetection AT edytamolik improvedmethodfordeterminationoforoticacidinmilkbyultrafastliquidchromatographywithoptimizedphotodiodearraydetection AT kamilzaworski improvedmethodfordeterminationoforoticacidinmilkbyultrafastliquidchromatographywithoptimizedphotodiodearraydetection
_version_	1718413238880698368

The Improved Method for Determination of Orotic Acid in Milk by Ultra-Fast Liquid Chromatography with Optimized Photodiode Array Detection

Ejemplares similares