Method Modification for Liquid Chromatographic Determination of Thiamine, Riboflavin, and Pyridoxine in Medical Foods

Abstract

A reversed-phase ion pair liquid chromatographic method developed for the simultaneous determination of thiamine (B1), riboflavin (B2), and pyridoxine (B6) in perchloric acid extracts of infant formulas was modified to include medical foods. UV detection of B1 and B2 was replaced by fluorescence detection, which resulted in improved sensitivity and specificity. B1 was detected by fluorescence after conversion to thiochrome by a postcolumn reaction with sodium hydroxide and potassium ferricyanide. The method uses a mobile phase of water, acetonitrile, hexanesulfonic acid sodium salt, ammonium hydroxide, and phosphoric acid adjusted to pH 3.6. The column is a 300 × 3.9 mm Nova Pak C18. Limits of detection were 0.05 μg/mL for B1 and B2 and 0.01 μg/mL for B6 by fluorescence detection. The system reproducibility was evaluated by completing 10 repetitive determinations on a medical food that gave a coefficient of variation of 5.9, 6.0, and 10.7% for B1, B2, and B6, respectively. Mean recoveries (n = 10) were 111,96.3, and 113% for B1, B2, and B6, respectively. The results compared favorably with those by AOAC Official Methods 942.23, 940.33, and 961.15 for B1, B2, and B6, respectively.

Ion pair liquid chromatography method for the determination of thiamine (vitamin B1) homeostasis

“…A multitude of HPLC methods for the determination of thiamine in food products (Amin and Reusch, 1987;Ang and Moseley, 1980;Barna and Dworschak, 1994;Baumgartner et al, 1997;Chase et al, 1993;Gehring et al, 1995;Hurst et al, 1983;Kirchmeier and Upton, 1978;Mauro and Wetzel, 1984;Nicolas and Pfender, 1990;Ohta et al, 1984;Sims and Shoemaker, 1993;Valls et al, 1999;Walker et al, 1981;Wimalasiri and Wills, 1985) and biological matrices (Bailey et al, 1994;Botticher and Botticher, 1987;Floridi et al, 1984;Kimura et al, 1982;Mascher and Kikuta, 1993;Roser et al, 1978;Schrijver et al, 1982;Warnock, 1982;Weber and Kewitz, 1985;Wielders and Mink, 1983) have been introduced over the years. However, only a handful of methods capable of simultaneous measurement of thiamine and its phosphate esters exist.…”

Determination of Thiamine and Its Esters in Beers and Raw Materials Used for Their Manufacture by Liquid Chromatography with Postcolumn Derivatization

“…Then, when it is necessary to distinguish between all of the naturally occurring species of thiamine, liquid chromatography (LC) is usually the method of choice (). Procedures for the determination of thiamine in foods using reversed-phase ( − ) or ion pair chromatography ( − ) have been proposed, and several reviews have also been published ( − ). UV spectrophotometry ( 11 , 13−15 , 18 , 21 , 24 , 25 ) has been used for samples that contain sufficient amounts of thiamine, while a fluorescence derivatization reaction based on the oxidation of thiamine to thiochrome, which shows strong fluorescence ( 5−7 , 10 , 12 , 16−20 , 23 , 26 , 28 ), is used for small concentrations.…”

“…The procedure could be applied to the determination of thiamine and its esters in beer and the raw materials used in its manufacture. Separation using the amide-based column is advantageous with respect to other existing methods ( − ) because the peaks are much narrower and column life is longer due to the simplicity of the mobile phase. Moreover, the use of the fluorescence detector allowed lower detection limits than those obtained using the UV spectrophotometer ().…”

Effect of gamma irradiation on beef quality

“…Both irradiated and nonirradiated beef samples were analyzed for its proximate chemical analysis according to (Ronald and Ronald, 1991). Thiamin and riboflavin content according to (Chase et al, 1993). Moreover, deterioration criteria (pH value, thiobarbituric acid value and total volatile bases nitrogen) were assessed according to (FAO 1986).…”