AOAC 2018 Methods

( q )  Vortex mixer .—VWR, or equivalent. ( r )  Ultrasonic bath .—Model 5800 (Branson, Danbury, CT, USA), or equivalent. ( s )  Centrifuge .—Model Allegra 6 with 20 and 30 mm diameter tube adaptors (Beckman Coulter, Brea, CA, USA), or equivalent; capable of 1500× g . ( t )  Nitrogen evaporation system with heating capabilities .— Model N-Evap 112 (Organimation, Berlin, MA, USA), or equivalent. ( u )  Analytical balances .—Accurate to 0.00001 g, Model CPA225D (Sartorius, Göttingen, Germany), and 0.01 g, Model GF3000 (A&D, Tokyo, Japan), or equivalent. ( v )  Magnetic stir plate .—Model Variomag Poly (Thermo Fisher Scientific), or equivalent. ( w )  Air displacement pipets .—Various sizes (Eppendorf, Hamburg, Germany), or equivalent. ( x )  Electronic repeater pipet .—Various sizes (Eppendorf), or equivalent. ( y )  4-Dram vials/caps .—Borosilicate glass/Teflon-lined (VWR), or equivalent. ( z )  Bottle-top dispenser .—100±0.6 mL, adjustable with graduations (Thermo Fisher Scientific), or equivalent. ( aa )  Mobile phase containers .—2 L polypropylene (Thermo Fisher Scientific), or equivalent. ( bb )  Syringe filters .—0.45 µm, 13 mm, GHP (Pall, Plano, TX, USA), or equivalent. ( cc )  Disposable syringes .—1 mL (BD Biosciences, Franklin Lakes, NJ, USA) or equivalent. ( dd )  Magnetic stir bars .—Various sizes, Teflon-coated (VWR), or equivalent. ( ee )  Autosampler vials/caps .—1.5 mL snap cap with preslit caps (Thermo Fisher Scientific), or equivalent. ( ff )  Resistance tubing .—5 ft of 0.007 in. id PEEK (Thermo Fisher Scientific), or equivalent. ( gg )  High-pressure mixing tee .—PEEK (IDEX, Lake Forest, IL, USA), or equivalent. C. Chemicals and Reagents ( a )  Water .—≥18 MΩ-cm, dispensed by filtration unit (MilliporeSigma), or equivalent. ( b )  Ethanol .—190 proof, American Chemical Society grade (Pharmco-AAPER, Brookfield, CT, USA), or equivalent. ( c )  Sodium hydroxide .—50% (w/w) and 1 N (Thermo Fisher Scientific), or equivalent. ( d )  Celite .—Acid-washed (IMERYS, Paris, France), or equivalent. ( e )  Carrez I solution .—3.6% (w/v), Part No. SC910116 (Thermo Fisher Scientific), or equivalent. ( f )  Carrez II solution .—7.2% (w/v), Part No. SC910216 (Thermo Fisher Scientific), or equivalent. ( g )  Reference standards .—D-(–)-arabinose (≥99%), D-(+)- galactose (≥99%), D-(+)-glucose (≥99%), D-(–)-fructose (≥99%), sucrose (≥99%), lactose (≥99%), D-(+)-maltose monohydrate (≥99%), and palatinose hydrate (≥99%; MilliporeSigma), or equivalent. ( h )  Sodium azide .—(MilliporeSigma), or equivalent. Note : All use of water must be ≥18 MΩ-cm, and sodium hydroxide must be low-carbonate. Do not use sodium hydroxide pellets, as they contain high amounts of carbonate.

AOAC Official Method 2018.16 Sugar Profile in Food, Dietary Supplements, Pet Food, and Animal Feeds High-Performance Anion-Exchange Chromatography with Pulsed Amperometric Detection First Action 2018 [Applicable for determination of sugar profile in food, dietary supplements, pet food, and animal feed.] Caution : Refer to the material safety data sheets of chemicals prior to use, and follow the use of suggested personal protective equipment. Sodium azide is toxic and should be handled with care and disposed of according to local regulations. Carrez I reagent should be kept away from strong acids, as toxic cyanide gas can form. A. Principle The HPAEC–PAD method allows quantification of the sugar profile in food, dietary supplements, pet food, and animal feeds. Sugars are extracted with a mixture of equal parts water and ethanol, with the addition of an internal standard. Extracts may be treated with Carrez reagents to precipitate proteins. Test solutions are filtered through a molecular weight (MW) filter, dried, and reconstituted to optimize chromatography and then diluted within the calibration range and filtered prior to analysis. Sugars are eluted from an anion-exchange column with a low concentration of sodium hydroxide, mixed with postcolumn addition of a higher concentration of sodium hydroxide, and detected by PAD. B. Apparatus ( a )  HPAEC–PAD .—ICS-5000 (Thermo Fisher Scientific, Waltham, MA, USA) or equivalent. ( b )  Working electrode .—ICS-5000 ED conventional Au (1 mm; Thermo Fisher Scientific), or equivalent. ( c )  Reference electrode .—ICS-5000 Ag/AgCl (Thermo Fisher Scientific), or equivalent. ( d )  Analytical column .—CarboPac PA20, 3×150 mm (Thermo Fisher Scientific), or equivalent. ( e )  Guard column .—CarboPac PA20, 3×30 mm (Thermo Fisher Scientific), or equivalent. ( f )  Borate trap .—4×50 mm (Thermo Fisher Scientific), or equivalent. ( g )  Anion trap .—ATC-3, 9×24 mm (Thermo Fisher Scientific) or equivalent. ( h )  Volumetric flasks .—ClassAglass (Thermo Fisher Scientific), or equivalent. ( i )  Volumetric pipets .—ClassAglass (Thermo Fisher Scientific), or equivalent. ( j )  Erlenmeyer flasks .—500 mL glass with stoppers (Pyrex; Corning, NY, USA), or equivalent. ( k )  MW filter .—Amicon Ultra-4 10K (MilliporeSigma, Burlington, MA, USA), or equivalent. ( l )  Plastic filter funnels .—VWR (West Chester, PA, USA), or equivalent. ( m )  Filter paper .—Qualitative P8, medium porosity (Thermo Fisher Scientific), or equivalent. ( n )  Cups/beakers .—Plastic, disposable (VWR), or equivalent. ( o )  Serological pipets .—Plastic, disposable (VWR), or equivalent. ( p )  Dessicator .—Model Dry Keeper (Sanplatec, Osaka, Japan), or equivalent.

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