Retinol analysis in dried blood spots by HPLC

Neal E. Craft, Tom Haitema, Lisa K. Brindle, Sedghen Yamini, Jean Hawes Humphrey, Keith West

Research output: Contribution to journalArticle

Abstract

There are many advantages to measuring vitamin A in dried blood spots (DBS) from a finger prick as compared to plasma collected by venipuncture. The advantages include easier collection, transport and storage; accessibility to younger and more remote populations; and decreased risk of disease transmission. We describe a method for the extraction of retinol from DBS for analysis by HPLC and initial comparison to plasma retinol. The effects of various buffers, detergents, antioxidants and chelators were evaluated to establish the most effective approach to elute the retinol: retinol binding protein (holo-RBP) complex from the blood collection cards. The process involves ultrasonic agitation to elute holo-RBP into a phosphate buffer containing an antioxidant and metal chelator. The holo-RBP complex was denatured by the addition of ethanol containing additional antioxidants permitting the extraction of free retinol into hexane. Following solvent evaporation, the extract was dissolved in methanol for HPLC analysis. The initial measured retinol levels in freshly collected DBS declined for 6-10 d whether stored at 25, 4 or -20°C, but remained consistent thereafter (homeostatic). By incorporating a 'recovery/volume adjustment' factor, measured retinol values in homeostatic DBS were adjusted to the equivalent of plasma retinol. For 17 normal adults, the correlation coefficient was 0.90 between plasma retinol and adjusted DBS retinol in samples that had been stored at -70°C for <9 mo. The use of this new sample matrix for vitamin A assessment will allow access to previously unavailable populations.

Original languageEnglish (US)
Pages (from-to)882-885
Number of pages4
JournalJournal of Nutrition
Volume130
Issue number4
StatePublished - 2000

Fingerprint

Dried Blood Spot Testing
Vitamin A
vitamin A
High Pressure Liquid Chromatography
blood
Antioxidants
Chelating Agents
chelating agents
antioxidants
Buffers
buffers
Retinol-Binding Proteins
retinol-binding protein
Phlebotomy
risk reduction
agitation
Hexanes
disease transmission
Ultrasonics
detergents

Keywords

  • Dried blood spot
  • Filter paper
  • HPLC
  • Nutritional assessment
  • Vitamin A

ASJC Scopus subject areas

  • Medicine (miscellaneous)
  • Food Science

Cite this

Craft, N. E., Haitema, T., Brindle, L. K., Yamini, S., Humphrey, J. H., & West, K. (2000). Retinol analysis in dried blood spots by HPLC. Journal of Nutrition, 130(4), 882-885.

Retinol analysis in dried blood spots by HPLC. / Craft, Neal E.; Haitema, Tom; Brindle, Lisa K.; Yamini, Sedghen; Humphrey, Jean Hawes; West, Keith.

In: Journal of Nutrition, Vol. 130, No. 4, 2000, p. 882-885.

Research output: Contribution to journalArticle

Craft, NE, Haitema, T, Brindle, LK, Yamini, S, Humphrey, JH & West, K 2000, 'Retinol analysis in dried blood spots by HPLC', Journal of Nutrition, vol. 130, no. 4, pp. 882-885.
Craft NE, Haitema T, Brindle LK, Yamini S, Humphrey JH, West K. Retinol analysis in dried blood spots by HPLC. Journal of Nutrition. 2000;130(4):882-885.
Craft, Neal E. ; Haitema, Tom ; Brindle, Lisa K. ; Yamini, Sedghen ; Humphrey, Jean Hawes ; West, Keith. / Retinol analysis in dried blood spots by HPLC. In: Journal of Nutrition. 2000 ; Vol. 130, No. 4. pp. 882-885.
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