Use of compounds naturally labeled with stable isotopes for the study of the metabolism of glycoprotein neutral sugars by gas-liquid chromatography-isotope-ratio mass spectrometry. Technical validation in the rat.
In order to develop an alternative method to radioactive labeling for the study of the glycoprotein sugar metabolism in man, the possible use of stable isotopes provided by naturally, 13C-enriched dietary compounds has been explored in rat intestine and serum. Rats were fed a semisynthetic diet containing 67% wheat starch (containing 1.08692 13C atom/100 carbon atoms) for a week, and then the same diet containing corn starch (1.10042% 13C) for a week. Neutral sugars were prepared from delipidated, trichloroacetic acid-precipitable material from 200-400 mg of intestinal mucosa or 1 mL of serum, separated, and analyzed as alditol acetates by gas-liquid chromatography coupled to isotope-ratio mass spectrometry. This technique allowed the determination, in a single experiment, of the amount and 13C abundance of six neutral sugars (including xylose used as internal standard). Despite the low abundance of 13C in natural products, the sensitivity of the technique was found to be sufficient to detect isotopic enrichment as low as 0.001% with good accuracy and reproducibility in 2 micrograms of each glycoprotein neutral sugar. As an example, the pattern of labeling by dietary D-glucose from corn starch appears to be very different for fucose, ribose, mannose, galactose, and glucose of intestine or serum macromolecules.
Rambal
C
C
Department of General and Medical Biochemistry, INSERM-CNRS U189, Faculté de Médecine Lyon-Sud, Oullins, France.
Pachiaudi
C
C
Normand
S
S
Riou
J P
JP
Louisot
P
P
Martin
A
A
D000818:Animals; D050260:Carbohydrate Metabolism; D002241:Carbohydrates / Q000737:chemistry; D002247:Carbon Isotopes; D002849:Chromatography, Gas; D006023:Glycoproteins / Q000097:blood / Q000737:chemistry / Q000378:metabolism*; D007413:Intestinal Mucosa / Q000378:metabolism*; D013058:Mass Spectrometry / Q000379:methods; D051381:Rats; D017207:Rats, Sprague-Dawley; D015203:Reproducibility of Results
D002241:Carbohydrates; D002247:Carbon Isotopes; D006023:Glycoproteins