OMB Meeting Book_9-11-14

2 concentrated formic acid. ( 4 ) 2.5% Acetonitrile in H O (mobile phase D).— 50 mL acetonitrile diluted to 2000 mL with laboratory water. ( 5 ) 10% Acetonitrile in H O.— 150 mL acetonitrile diluted to 1500 mL with laboratory water. ( 6 ) 25% Acetonitrile in H O SPE elution solvent.— 25 mL 2 2 ( 7 ) 30% Acetonitrile in H O SPE elution solvent.— 30.0 mL acetonitrile diluted to 100 mL with laboratory water. ( 8 ) 50% Acetonitrile in H O, column cleaning and storage solution.— 500 mL acetonitrile diluted to 1000 mL in a volumetric flask. Expiration 6 months. ( 9 ) 25% Ethanol.— 50 mL ethanol diluted to 200 mL with laboratory water. ( 10 ) 0.40% Potassium cyanide.— Dissolve 0.02 g potassium cyanide in and dilute to 5 mL with 0. 125 M sodium acetate buffer. Make fresh immediately before use. ( 11 ) 1% Potassium cyanide.— Dissolve 0.25 g potassium cyanide in and dilute to 25 mL with laboratory water. Prepare fresh immediately before use. Resolution Test Solution: Weigh ~0.005grams riboflavin onto a weigh paper. Transfer to a 100 mL volumetric flask and bring to volume with 10% acetonitrile solution. Stir to dissolve. Mix equal amounts of solution with the highest concentration vitamin B12 working standard. Expiration: 1 day ( 12 ) 0. 251 M sodium acetate buffer.— Dissolve 16.441 g sodium acetate anhydrous or 27.268 g sodium acetate trihydrate in approximately 1800 mL laboratory water. Adjust pH to 4.50 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 immediately 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 stock standard (10000 μ g/L).— Accurately weigh 2 diluted with 750 mL laboratory water and adjust pH to 5–7 with concentrated formic acid. ( 3 ) Mobile phase C.— 4.0 mL TEA and 750 mL acetonitrile diluted with 250 mL laboratory water and adjust pH to 5–7 with acetonitrile diluted to 100 mL with laboratory water. 2

Table 2011.10B. Guidelines for loading sample filtrates onto SPE cartridges a

Vitamin B in RTF

concentration μg/L

Volume of filtrate loaded onto SPE cartridge, mL

Final dilution volume, mL

12

product,

<1

80

5

1–10

70–80

10

11–20

50–60

10

21–50

20–40

10

a Do not load more than 60 mL adult and pediatric nutritionals onto an

Alltech C or C

cartridge.

8

18

( c ) Drierite.— Desiccant, anhydrous calcium sulfate, 8 mesh. ( d ) Ethanol.— Denatured. ( e ) Formic acid.— ACS. ( f ) Laboratory water.—≥ 15 MΩ·cm . ( g ) Potassium cyanide.— 97%ACS reagent. ( h ) Potassium phosphate dibasic.— ACS. Riboflavin - <96% ACS ( i ) Sodium acetate anhydrous or sodium acetate trihydrate .— ACS. ( j ) Taka-diastase.— Accurate Chemical Co. (www. accuratechemical.com) or equivalent. ( k ) Triethylamine (TEA).— HPLC grade.

( l ) Vitamin B

(cyanocobalamin) standard.— USP reference,

12

official lot number (refer to USP catalog for current lot). Store in desiccator protected from white light. ( Note : See standard label for purity.)

D. Solution and Standard Preparation

All solutions can be scaled up or down for convenience provided good laboratory practices are observed. Solutions can be stored at 2–30°C in tight, inert containers unless otherwise noted. ( a ) Solutions.— ( 1 ) Mobile phase A.— 4.0 mL TEA diluted with

1000 mLwater and adjust pH to 5–7 with concentrated formic acid. ( 2 ) Mobile phase B.— 4.0 mL TEA and 250 mL acetonitrile

12

the appropriate amount of vitamin B

USP reference standard to

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