6. AOACSPIFANMethods-2018Awards

110

6 S alvati et al .: J ournal of AOAC I nternational V ol . 99, N o . 3, 2016

Table 1. Conditions for MS transitions on a Waters TQ-S are given along with retention time windows

Collision energy (V)

Compound

Function No.

Start, min

End, min Molecular ion Fragment ion Cone voltage

Dwell time, s

Nicotinamide a Nicotinamide

1 1 1 1 2 2 2 2 3 3 3 3 4 4 4 4 5 5 5 5 6 6 6 6 7 7 7 7

2.71 2.71 2.71 2.71 0.50 0.50 0.50 0.50 1.76 1.76 1.76 1.76 0.50 0.50 0.50 0.50 2.41 2.41 2.41 2.41 3.01 3.01 3.01 3.01 4.21 4.21 4.21 4.21

3.20 3.20 3.20 3.20 1.70 1.70 1.70 1.70 2.70 2.70 2.70 2.70 1.70 1.70 1.70 1.70 3.00 3.00 3.00 3.00 3.60 3.60 3.60 3.60 5.00 5.00 5.00 5.00

122.9 122.9 127.0 127.0 124.0 124.0 128.0 128.0 168.0 168.0 171.0 171.0 169.0 169.0 172.0 172.0 170.0 170.0 174.0 174.0 265.1 265.1 269.0 269.0 377.0 377.0 383.0 383.0

80.1 96.0 84.0

20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0 20.0

16.0 16.0 16.0 16.0 16.0 16.0 16.0 16.0 22.0 12.0 22.0 12.0 20.0 12.0 20.0 12.0 18.0 12.0 18.0 12.0 30.0 12.0 30.0 12.0 35.0 20.0 35.0 20.0

0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025 0.025

2 H 4 2 H 4

-nicotinamide a -nicotinamide

100.0

Nicotinic acid a Nicotinic acid

80.0

106.0

2 H 4 2 H 4

-nicotinic acid a -nicotinic acid

84.1

109.0

Pyridoxal Pyridoxal a

94.0

150.0

2 H 3 2 H 3

-pyridoxal -pyridoxal a

97.0

153.0 134.0 152.0 136.0 155.0 134.0 152.0 138.0 156.0

Pyridoxamine Pyridoxamine a

2 H 3 2 H 3

-pyridoxamine -pyridoxamine a

Pyridoxine a Pyridoxine

13 C 4 13 C 4

-pyridoxine a -pyridoxine

Thiamine Thiamine a

81.0

122.0

13 C 4 13 C 4

-thiamine -thiamine a

81.0

122.0 172.0 243.0 175.0 249.0

Riboflavin Riboflavin a

13 C 4 13 C 4

, 15 N 2 , 15 N 2

-riboflavin -riboflavin a

0.025 Although the mass transitions are expected to remain the same across instrument platforms, the other parameters may need to be adjusted to maximize sensitivity. a  Indicates primary transition used in quantitation.

concentration of nicotinamide and nicotinic acid are summed to report “Total B 3 ” and concentration of pyridoxal, pyridoxamine, and pyridoxine are summed to report “Total B 6 .” Thiamine and riboflavin do not require this step.

where [Vit] WSx

= vitamin concentration in the working standard = concentration of vitamin in the MWS = volume of the MWS fortified in working

in ng/mL; [Vit] MWS in ng/mL; Vol MWS

standard in μL; and 500 = dilution factor. (d)  Vitamin concentration calculated in product from analytical result:

Validation

Vit [ ]

RW SW PW AS × × ×

500

Vit [ ]

=

Method performance was demonstrated against predefined suitability criteria for these vitamins published in SMPRs (1–4). Although each SMPR is slightly different, methods for B 1 , B 2 , B 3 , and B 6 are required to achieve repeatability of ≤5% RSD, reproducibility of ≤10% RSD, and over-spike recovery of 90–110%. This method met each of these requirements except reproducibility, which was not evaluated. Instead, intermediate precision is given and suggests the reproducibility requirement will be met upon multilaboratory evaluation. Additional measures of method performance are also discussed, including: linearity, specificity, and robustness.

sample

where [Vit] sample

= vitamin concentration in product, μg/kg;

[Vit] AS = vitamin mass in the analytical sample as calculated from calibration curve, ng/mL; RW = reconstitution weight (total), g, for direct weight (liquid) samples RW = 1; SW = analytical sample weight, g; PW = powder weight (for reconstituted samples), g, for liquid samples, this value is 1; and 500 = dilution factor. (e)  For vitamins B 3 and B 6 , the reported concentration of the individual forms is summed to report total. For example,