AOAC Final Action Methods in 2018

Figure 2015.09B. Chromatogram of vitamin K 1 standard.

they must be wrapped tightly in foil. All standard solutions must be prepared using Class A volumetric glassware. ( a ) 10% Isopropanol. —Add about 70 mL iso-octane to a 100 mL volumetric flask. Add 10.0 mL isopropanol to the volumetric flask and dilute to volume with iso-octane. Expiration 6 months. ( b ) Mobile phase.— Add about 900 mL iso-octane to a 1000 mL volumetric flask. Add 3–4 mL 10% isopropanol to the volumetric flask and dilute to volume with iso-octane. ( Note : The isopropanol concentration in the mobile phase can be adjusted slightly until baseline resolution of cis and trans vitamin K 1 from other peaks present in some samples is achieved. See Figures 2015.09B–D ). ( c ) Post-column electrolyte solution.— Transfer 0.5 (±0.05) g zinc chloride and 0.20 (±0.02) g sodium acetate anhydrous to a 1000 mL volumetric flask with reagent alcohol. Add 150 (±15) µL concentrated acetic acid and dilute to volume with reagent alcohol. Add a stir bar and stir solution for about 30 min or until solution is clear and all salts are dissolved. Expiration 6 months. ( d )( 1 ) Vitamin K 1 (phytonadione) stock standard solution.— Accurately weigh to 0.00001 g about 0.05500 g vitamin K 1 (phytonadione) into a 250 mL volumetric flask. Dissolve oil and

dilute to volume with iso-octane. Store refrigerated in a tightly stoppered container protected from light. Expiration 6 months. ( 2 ) Vitamin K 1 stock standard concentration (for non-USP standard).— To determine the stock standard concentration, evaporate 1.0 mL vitamin K 1 stock standard under a stream of nitrogen and redissolve residue in 25.0 mL hexane. Measure absorbance of this solution in a 1 cm cell against a reference of hexane at a wavelength of 248 nm with a spectrophotometer. Calculate the vitamin K 1 stock standard concentration, C stock , in mg/L using the following equation: C stock = ( A 248 × 10000 × 25)/419 where A 248 is the absorbance of the solution at 248 nm; 419 is the A 1% 1 cm value of vitamin K 1 in hexane at 248 nm; 10000 is the conversion of A 1% 1 cm to mg/L; and 25 is the dilution volume of the stock standard. ( e ) Vitamin K 1 (phytonadione) intermediate I standard solution.— Dilute 1.0 mL vitamin K 1 stock standard to 100 mLwith iso-octane. Prepare fresh each time new working standards are made.

Figure 2015.09C. Chromatogram of SRM 1849a.

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