Quantitative microanalysis of oligosaccharides by high-performance liquid chromatography
ABSTRACT The rapid separation and quantitative determination of per-O-benzoyl oligosaccharides were obtained using high performance, liquid chromatography. Oligosaccharides were completely O-benzoylated without concomitant N-benzoylation of acetamidodeoxyhexoses. Benzoylation prior to analysis allowed a quantitative determination of picomolar amounts because the absorbance at 230 nm of these derivatives is directly proportional to the number of benzoyl groups present. Separation by normal and reversed-phase chromatography was demonstrated, and the best resolution was obtained on an Ultrasphere octyl column. Excellent separations of oligosaccharides containing up to 10 sugar residues present in mannosidosis urine and of malto-oligosaccharides containing up to 15 sugar residues present in Karo syrup were achieved within an analysis time of 30 min. Anomers of maltose, maltotriose, and maltotetraose were separated; for this reason, reduction of complex samples prior to analysis is advisable. The effect of linkage configuration on retention time was tested with reduced, α-linked di- and tri-glucopyranosides. The presence of an acetamidodeoxyhexose residue in an oligosaccharide significantly reduced its retention time, whereas branching had the opposite effect. A linear response was obtained for the injection of 1–600 pmol of raffinose, and the detection limit was 0.5 pmol. Derivatization and analysis of raffinose was shown to yield reproducible results within the range 0.01–1 μmol, and, with special precautions to minimize losses, as little as 100 pmol could be analyzed successfully.
SourceAvailable from: ecn.www.ecn.purdue.edu[Show abstract] [Hide abstract]
ABSTRACT: The analysis of water-soluble cellodextrins using liquid chromatography is readily achieved with a variety of packings. Direct injection of enzyme incubation mixtures allows quantitation of 10 mM cellodextrins in hydrolysis mixtures, resulting in a method which is useful for kinetic studies. Reported here are operating procedures for a 4% cross-linked, styrene-divinyl benzene cation exchanger (Aminex 50W-X4 (Bio Rad Lab., Griffin, CA, USA), 20–30 μm particle size) in the Ca++ form, packed in a column of dimensions 6 mm i.d. × 60 cm long. Using this column, resolution of the cellodextrins, celloheptaose through cellobiose and glucose was possible with 91 mM H2SO4 as the eluent. Requirements of the separation system included use of a pulsation free syringe pump to minimize baseline fluctuations, the use of Ca++ as the counterion to give a column operational life of 500–1000 injections, and injection of sample volumes of up to 25 μL. cellodextrins were quantified at sub-microgram (nmole) levels using a differential refractometer as the detector. Examples of this technique for analysis of the acid hydrolysis of cellodextrins and enzymatic hydrolysis of cellodextrins and carboxymethylcellulose are described.Biomass 01/1990; 23(4):307-317. DOI:10.1016/0144-4565(90)90039-M
[Show abstract] [Hide abstract]
ABSTRACT: The chemistry of the acid stage of the synthesis of carbohydrate-based adhesives has been investigated. The sulfuric acid-catalyzed reactions of d-glucose with urea in a phenol-water solution provided both N-β-d-glucopyranosylurea and N,N′-di-β-d-glucopyranosylurea. These compounds have been isolated from the resin as perbenzoylated derivatives, and compared spectroscopically with authentic samples. The d-glucosylureas are transient products whose combined yields approach 35% of the carbohydrate fraction during the initial stages of resin synthesis. Only traces of 5-(hydroxymethyl)-2-furaldehyde, levulinic acid, and formic acid were detected in the resin mixture, suggesting that the classical dehydration pathway is not a major route for the disappearance of d-glucose.Carbohydrate Research 06/1989; 189:103–112. DOI:10.1016/0008-6215(89)84089-3 · 1.97 Impact Factor
[Show abstract] [Hide abstract]
ABSTRACT: Little is known regarding the impact of elevated [CO2] on the chemical composition of rice grains. A field experiment was conducted using open-top chambers with rice (Oryza sativa L. cv. Ariete) grown at two levels of atmospheric CO2 (375 and 550 µmol/mol), and their effects monitored on the proximate composition and carbohydrate contents of the grains. Following exposure to elevated [CO2], soluble dietary fibers increased by 136%, 82%, and 77% in the brown rice, white rice, and bran, respectively. Increases of a lower magnitude (8%) were observed for insoluble dietary fibers in the bran and brown rice. For all ten sugars identified, there was a trend for increasing their content. For example, increases of 135% were recorded for glucose in the white rice. In all rice milling fractions, elevated [CO2] reduced the protein (4%–15%) and amylose (6%–16%) contents, with no effect on the ash, starch and gross energy contents. The fat content was increased by elevated [CO2] in the white rice (23%) and tended to decrease in the bran (9%). It is concluded that besides yield, increased dietary fibers might be another positive effect of high levels of atmospheric CO2 expected by the middle of the current century.Cereal Chemistry 01/2014; 91:293-299. DOI:10.1094/CCHEM-09-13-0180-R · 1.06 Impact Factor