AOAC SPIFAN ERP & Working Group Method Reviews (July 10, 2019)

( 8 ) 25% Ethanol.— 50 mL ethanol diluted to 200 mL with laboratory water. Expiration 1 year in tightly stoppered container. ( 9 ) 0.40% Potassium cyanide for samples with 5 mL final dilution volume.— Dissolve 0.02 g potassium cyanide in and dilute to 5 mL with 0.25 M sodium acetate buffer. Make fresh daily before use. ( 10 ) 1% Potassium cyanide.— Dissolve 0.25 g potassium cyanide in and dilute to 25 mL with laboratory water. Prepare fresh daily before use. ( 11 )  Resolution test solution for determining appropriate gradient conditions when a new analytical column is installed .— Weigh about 0.005 g riboflavin onto a weigh paper. Transfer to a 100 mL volumetric flask and dilute to volume with 10% acetonitrile solution. Stir to dissolve. Mix equal amounts of riboflavin solution with the highest concentration vitamin B 12 working standard. Expiration: 1 week. ( 12 ) 0.25Msodiumacetate buffer.— Dissolve 41 g sodiumacetate anhydrous or 68 g sodium acetate trihydrate in approximately 1800 mL laboratory water. Adjust pH to 4.5 with concentrated acetic acid. Dilute to 2000 mL with laboratory water. Expiration 3 months. ( 13 ) 6% Taka-diastase.— Dissolve 0.6 g taka-diastase in 10 mL water. Prepare fresh daily before use. ( b ) Standards.— Prepare all standards under UV shielded fluorescent lights and store at 2–8°C in tightly stoppered volumetric flasks. ( 1 ) Vitamin B 12 stock standard (10000 μ g/L).— Accurately weigh the appropriate amount of vitamin B 12 USP reference standard to give a stock standard concentration of 10000 m g/L. Dissolve in and dilute to 100 mL with 25% ethanol. Expiration 6 months. Use the following equation to calculate the amount of vitamin B 12 reference standard that should be weighed: S w = 10000 × 0.1 × 1/P where S w = amount of vitamin B 12 standard to be weighed in

Table 2011.10D. Guidelines for loading sample filtrates onto SPE cartridges a Vitamin B 12 concentration in RTF product, μg/kg Volume of filtrate loaded onto SPE cartridge, mL Final dilution volume, mL

<1

80

5

1–10

70–80 50–60 20–40

10 10

11–20 21–50

10 a  Do not load more than 60 mL adult and pediatric nutritionals onto an Alltech C 8 or C 18 cartridge.

( a ) Solutions.— ( 1 ) Mobile phase A.— 4.0 mL TEA diluted with 1000 mL water and pH adjusted to 5–7 with about 1.25 mL concentrated formic acid. Expiration 1 week. ( 2 ) Mobile phase B.— 4.0 mL TEA and 250 mL acetonitrile diluted with 750 mL laboratory water and pH adjusted to 5–7 with about 1.25 mL concentrated formic acid. Expiration 1 week in tightly stoppered container. ( 3 ) Mobile phase C.— 4.0 mL TEA and 750 mL acetonitrile diluted with 250 mL laboratory water and pH adjusted to 5–7 with about 1.25 mL concentrated formic acid. Expiration 1 week in tightly stoppered container. ( 4 ) 2.5% Acetonitrile in H 2 O (mobile phase D).— 50 mL acetonitrile diluted to 2000 mL with laboratory water. Expiration 1 week in tightly stoppered container. ( 5 ) 10% Acetonitrile in H 2 O.— 150 mL acetonitrile diluted to 1500 mL with laboratory water. Expiration 1 month in tightly stoppered container. ( 6 ) 30% Acetonitrile in H 2 O SPE elution solvent.— 30.0 mL acetonitrile diluted to 100 mL with laboratory water. Expiration 1 month in tightly stoppered container. ( 7 ) 50% Acetonitrile in H 2 O, column cleaning and storage solution.— 500 mL acetonitrile diluted to 1000 mL in a volumetric flask. Expiration 6 months.

Figure 2011.10A. System setup and configuration: Configuration 1.

Figure 2011.10B. System setup and configuration: Configuration 2.

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