Review of fiber methods and applicability to fortified foods and supplements: choosing the correct method and interpreting results.

Fiber is known to be an important part of our nutrition and has many positive health benefits, including weight management and maintaining heart health. In recent years, a number of new ingredients have been manufactured or isolated that are being used to increase the health benefits of a product. Some are used as prebiotics that stimulate the growth of the beneficial bacteria in the gut, or are used as replacements for sugars, starch, or fat in manufactured foods. Fiber supplements have also been produced that can be taken to provide additional fiber to the diet. The term "fiber" does not relate to a single analyte or entity, but instead relates to a multitude of components. This adds to the complexity of analytical testing as there are a number of AOAC International and AACC International official methods which have been validated and can be used. Although methods have been developed for specific fiber ingredients, a number of methods have also been developed to capture just "fiber". These "fiber" methods will capture differing degrees of the different fiber ingredients, so knowledge of the fiber sources is critical. The net result is that a variety of testing approaches may be used, but caution must be exercised in order to ensure that the total fiber result is accurately determined. A critical review of available fiber methodology and possible testing approaches is presented, along with how to accurately interpret and understand results.

A comparative study of monosaccharide composition analysis as a carbohydrate test for biopharmaceuticals.

The various monosaccharide composition analysis methods were evaluated as monosaccharide test for glycoprotein-based pharmaceuticals. Neutral and amino sugars were released by hydrolysis with 4-7N trifluoroacetic acid. The monosaccharides were N-acetylated if necessary, and analyzed by high-performance liquid chromatography (HPLC) with fluorometric or UV detection after derivatization with 2-aminopyridine, ethyl 4-aminobenzoate, 2-aminobenzoic acid or 1-phenyl-3-methyl-5-pyrazolone, or high pH anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD). Sialic acids were released by mild acid hydrolysis or sialidase digestion, and analyzed by HPLC with fluorometric detection after derivatization with 1,2-diamino-4,5-methylenedioxybenzene, or HPAEC-PAD. These methods were verified for resolution, linearity, repeatability, and accuracy using a monosaccharide standard solution, a mixture of epoetin alfa and beta, and alteplase as models. It was confirmed that those methods were useful for ensuring the consistency of glycosylation. It is considered essential that the analytical conditions including desalting, selection of internal standards, release of monosaccharides, and gradient time course should be determined carefully to eliminate interference of sample matrix. Various HPLC-based monosaccharide analysis methods were evaluated as a carbohydrate test for glycoprotein pharmaceuticals by an inter-laboratory study.

Determination of saccharide content in pneumococcal polysaccharides and conjugate vaccines by GC-MSD.

A simple and sensitive gas chromatographic method was designed for quantitative analysis of Streptococcus pneumoniae capsular polysaccharides, activated polysaccharides, and polysaccharide conjugates. Pneumococcal serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19A, 19F, and 23F polysaccharide or conjugate were subjected to methanolysis in 3N hydrochloric acid in methanol followed by re-N-acetylation and trimethylsilylation. Derivatized samples were chromatographed and detected using gas chromatography with mass selective detector. Gas chromatographic results were compared with colorimetric values with agreement of 92 to 123% over the range of all samples tested. Monosaccharides released during methanolysis included hexoses, uronic acids, 6-deoxy-hexoses, amino sugars, and alditols. Quantitative recovery of monosaccharides was achieved for all serotypes by the use of a single methanolysis, derivatization, and chromatography procedure. Response factors generated from authentic monosaccharide standards were used for quantitation of pneumococcal polysaccharides and conjugates with confirmation of peak assignments by retention time and mass spectral analysis. This method allows saccharide quantitation in multivalent pneumococcal vaccine intermediates and final drug products with low-level detection (10 pg) and peak purity.