Analysis of Sugars 1 Definition Nutritional labelling of food products requires listing of sugars content. Sugars means all mono-saccharides and di-saccharides present in food*. *Codex Guidelines on Nutrition Labelling, CAC/GL 2-1985 2 Mono- and di-saccharides HO OH OH OH OH OH OH O OH O O O etc HO HO OH HO OH HO OH HO OH HO OH OH Fructose Glucose Galactose Mannose OH O OH HO OH OH HO OH O HO HO OH O O HO HO O OH HO O O O OH HO OH OH OH O HO OH OH HO OH Sucrose Lactose Maltose OH OH O O HO O etc HO HO OH OH OH Cellobiose 3 Common mono- and di-saccharides HO OH OH OH OH O OH O O HO OH HO OH HO OH HO OH OH Fructose Glucose Galactose OH O OH HO OH OH HO OH O HO HO OH O O HO HO O OH HO O O O OH HO OH OH OH O HO OH OH HO OH Sucrose Lactose Maltose 4 Common mono- and di-saccharides HO OH OH OH OH O OH O O HO OH HO OH HO OH HO OH OH Fructose Glucose Galactose OH O OH HO OH OH HO OH O HO HO OH O O HO HO O OH HO O O O OH HO OH OH OH O HO OH OH HO OH Sucrose Lactose Maltose 5 Common sugars in foods Mono-saccharides: Fructose, glucose, galactose Di-saccharides: Lactose, maltose, sucrose 6 Properties of sugars (LC Analysis) OH Trimethylsilyl Primary amine chloride Glycosylamine O Methylsilyl HO ether (LC Analysis) HO OH (GC Analysis) OH Ac2O Acetate (GC Analysis) 7 Properties of sugars Water soluble. Low thermal stability. Do not have properties of fluorescence emission. 8 Sugar Analysis LC Methods with: Refractive Index Detector Evaporative Light Scattering Detector Pulsed Amperometric Detector 9 Current AOAC LC Methods 977.20 Honey Fructose, glucose, sucrose 980.13 Milk chocolate Fructose, glucose, lactose, maltose, sucrose 982.14 Presweetened cereals Glucose, fructose, sucrose, maltose 984.22 Purity of lactose Lactose 996.04 Cane & beet final Glucose, fructose, sucrose, molassess lactose 2000.17 Raw cane sugar Glucose, fructose 10 Current GB LC Method GB-T 22221-2008 食品中果糖、葡萄糖、蔗糖、麦芽糖、乳糖的测 定 高效液相色谱法 Determination of fructose, glucose, sucrose, maltose, lactose in foods – high performance liquid chromatography 11 Mono- and di-saccharides analysis Amine column Acetonitrile/water (80:20) Refractive index detector 2 mL/min at 30 oC 12 Mono- and di-saccharides analysis Amine column may not be able to resolve all the peaks. Alternative column may be used, e.g., Shodex sugar column. 13 Mono- and di-saccharides analysis Shodex column Water Refractive index detector 0.5 mL/min at 80 oC 14 Mono- and di-saccharides analysis 15 Analysis of reducing sugars AOAC methods: 920.183 Sugars (Reducing) in Honey 920.190 Sugars (Reducing) in Maple Products 945.66 Total Reducing Sugars etc. 16 Analysis of reducing sugars Fehling solution is used in the methods It relies on chemical reaction between the sugars and the Fehling solution. O O CuO +Cu2O R H R OH OH OH O OH HO HO HO OH HO O OH OH 17 Limitation of analyzing reducing sugar The best-known and most significant non-reducing sugar in foods is sucrose! (The most important OH O low molecular weight HO HO carbohydrate of animal diet). HO HO O O OH NOT appropriate for testing general food!! HO OH 18 Typical analysis SugarsSugars inin Preparation/Preparation/ LCLC samplesample CleanClean upup 19 Mono- and di-saccharides analysis Extract the sugars from sample using a mixture of ethanol and water 20 Mono- and di-saccharides analysis Centrifuge the mixture after the extraction 21 Mono- and di-saccharides analysis Clean up the extract by SPE 22 Mono- and di-saccharides analysis Clean up the extract by SPE 23 Mono- and di-saccharides analysis Analyse the extract using LC with refractive index detector 24 Proficiency test FAPAS LGC AOAC 25 CRM BCR NIST LGC 26 Points to note Sugar alcohols (polyols) are not “sugars” Not all monosaccharides and disaccharides are reducing sugars. Test method for reducing sugar may not be suitable for the analysis of sugars. 27 Points to note Definition of “0” Sugars ≤0.5 g/100 g Limit of detection of total sugars should be less than or equal to 0.5 g/100 g. 28 Thank You 29.