1 Dietary fiber and Vitamin analysis Pham Van Hung, PhD Dietary fiber • Dietary fiber is essentially the sum of the nondigestible components of a foodstuff or food product. • Most, but not all, dietary fiber is plant cell-wall material (cellulose, hemicelluloses, lignin) and thus is composed primarily of polysaccharide molecules. • Insoluble dietary fiber: cellulose, lignin, hemicelluloses entrapped in a lignocellulosic matrix, and resistant starch. • Soluble dietary fiber: Hemicelluloses not entrapped in a lignocellulosic matrix, much of the native pectin, and the majority of hydrocolloids/food gums. Dietary fiber analysis • Dietary fiber is often determined gravimetrically. In such a procedure, digestible carbohydrates, lipids, and proteins are selectively solubilized by chemicals or removed by enzyme-catalyzed hydrolysis. • Nonsolubilized and/or nondigested materials are collected by filtration, and the fiber residue is recovered, dried, and weighed. Dietary fiber analysis • Starches and proteins are removed by enzymes • Lipids are removed easily from the sample with organic solvents. • Protein and minerals that are not emoved from the sample during the solubilization steps should be corrected for by Kjeldahl nitrogen analysis and by ashing portions of the fiber residue. 2 Dietary fiber analysis Vitamin • Vitamins are defined as relatively low- molecularweight compounds which humans require small quantities for normal metabolism. • Humans cannot synthesize most vitamins and therefore need to obtain them from food and supplements. • Insufficient levels of vitamins result in deficiency diseases. • The IU is a unit of measurement for the amount of a substance, based on measured biological activity or effect. Vitamin • Fat-soluble vitamins – A: Retinol or β-carotene – D: Cholecalciferol or Ergocalciferol – K: Phylloquinone – E: Tocopherol • Water-soluble vitamins – B complex: Thiamin (B1), Riboflavin (B2), Pyridoxine (B6), Folic acid (B9), Cyanocobalamin (B12) – C: ascorbic acid Vitamin analysis • Extraction Methods – Ascorbic acid: Cold extraction with metaphosphoric acid/acetic acid. – Vitamin B1 and B2: Boiling or autoclaving in acid plus enzyme treatment. – Niacin: Autoclaving in acid (noncereal products) or alkali (cereal products). – Folate: Enzyme extraction with α-amylase, protease and γ-glutamyl hydrolase(conjugase). – Vitamins A, E, or D: Organic solvent extraction, saponification, and re-extraction with organic solvents. For unstable vitamins such as these, antioxidants are routinely added to inhibit oxidation. 3 Vitamin analysis • Bioassay Methods: – vitamins B12 and D. • Microbiological Assays: – water-soluble vitamins. – The growth of a certain microorganism in an extract of a vitamin-containing sample is compared against the growth of this microorganism in the presence of known quantities of that vitamin. • Chemical Methods Vitamin analysis • Vitamin A: The test sample is saponified with ethanolic KOH, vitamin A (retinol) is extracted into organic solvent and then concentrated. Vitamin A isomers – all-trans- retinol and 13-cis-retinol – levels are determined by HPLC on a silica column. • Vitamin E: The sample is saponified under reflux, extracted with hexane, and injected onto a normal phase HPLC column connected to a fluorescence detector. • Vitamin C: L-ascorbic acid is oxidized to L- dehydroascorbic acid by the oxidation–reduction indicator dye, 2,6-dichloroindophenol. At the endpoint, excess unreduced dye appears rosepink in acid solution. • Thiamin (B1): Following extraction, enzymatic hydrolysis of thiamin’s phosphate esters and chromatographic cleanup. • Riboflavin (B2): Following extraction, cleanup, and compensation for the presence of interfering substances, riboflavin is determined fluorometrically. The end! . 1 Dietary fiber and Vitamin analysis Pham Van Hung, PhD Dietary fiber • Dietary fiber is essentially the sum of the. are collected by filtration, and the fiber residue is recovered, dried, and weighed. Dietary fiber analysis • Starches and proteins are removed by enzymes •