2015.14 Vitamin B (July 2018)

2015.14 (JULY 2018) BVit-02 Total B_AOAC_2015.14_MLT_Final

FOR ERP USE ONLY DO NOT DISTRIBUTE

c) Pyridoxal Stock Solution (140 ug/mL) - Accurately weigh 17.0 ± 0.5 mg pyridoxal dihydrochloride standard. Quantitatively transfer to a 100 mL volumetric flask. Add lab water to a total volume of about 70 mL and swirl until completely dissolved. Bring to volume with lab water. Mix well. Expiration: 3 months. d) Pyridoxamine Stock Solution (160 ug/mL) - Accurately weigh 23.0 ± 0.5 mg pyridoxamine hydrochloride standard. Quantitatively transfer to a 100 mL volumetric flask. Add lab water to a total volume of about 70 mL and swirl until completely dissolved. Bring to volume with lab water. Mix well. Expiration: 3 months. 9. Mixed Working Standard (MWS) - Combine 500 μL VSSM, 25 μL pyridoxamine stock, 25 μL pyridoxal stock, and 65 μL nicotinic acid stock solutions in a 10 mL volumetric flask containing approximately 5 mL of 50 mM ammonium formate. Bring to volume with 50 mM ammonium formate and mix well. Prepare fresh daily. 10. Working Standard Solution Preparation a) WS1 – Add 20 μL MWS and 980 μL of 50 mM ammonium formate to a 50 mL centrifuge tube. Add 100 μL of ISSM, and vortex to mix. Prepare fresh daily. b) WS2 – Add 50 μL MWS and 950 μL of 50 mM ammonium formate to a 50 mL centrifuge tube. Add 100 μL of ISSM, and vortex to mix. Prepare fresh daily. c) WS3 – Add 100 μL MWS and 900 μL of 50 mM ammonium formate to a 50 mL centrifuge tube. Add 100 μL of ISSM, and vortex to mix. Prepare fresh daily. d) WS4 – Add 200 μL MWS and 800 μL of 50 mM ammonium formate to a 50 mL centrifuge tube. Add 100 μL of ISSM, and vortex to mix. Prepare fresh daily. e) WS5 – Add 500 μL MWS and 500 μL of 50 mM ammonium formate to a 50 mL centrifuge tube. Add 100 μL of ISSM, and vortex to mix. Prepare fresh daily. f) WS6 – Add 1000 μL MWS to a 50 mL centrifuge tube. Add 100 μL of ISSM, and vortex to mix. Prepare fresh daily. E. Procedure 1. Sample preparation a) For powdered products, using a tarred beaker or LDPE cup, weigh 10.0 ± 0.3 g of sample. Record the weight to at least 4 significant figures. This is the powder weight. Add room temperature laboratory water to bring the total reconstituted sample weight (to include the product weight) to 100 ± 2 g. Record the weight to at least 4 significant figures. This is the reconstitution weight. Carefully add a stir bar so as not to splash the liquid from the beaker/cup and place it onto a stir plate. Set the stir plate to stir the sample as fast as possible without causing the sample to splatter or froth. Powder samples should stir for at least 10 minutes but not more than 30 minutes. b) For reconstituted powders and liquid products - using a tarred, 50 mL centrifuge tube, weigh the appropriate sample amount (1.000 ± 0.100 g for infant formula, 0.500 ± 0.050 g for pediatric formulas and the NIST SRM, and 0.250 ± 0.050 g for adult nutritionals). Record the weight to 0.0001 g. This is the sample weight. Add 100 µL of the internal standard mixture (ISSM) via positive-displacement pipette. Vortex to mix.