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Method Validation Report

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Validation Report for the Determination of Thiamin HCl, Riboflavin, Niacin (Nicotinic Acid and Nicotinamide), Pantothenic acid, Vitamin B6 (Pyridoxine, Pyridoxal and Pyridoxamine) and Biotin in Infant Formula and Adult Nutritionals by LC-MS/MS

Document Title

Table 20B: Ruggedness of Partially hydrolyzed protein infant formula powder

Concentration (mcg/100g)

Vit B6 (as Pyridoxine) Biotin

Pyridoxine

Pyridoxal Pyridoxamine

Day 1 Day 2

376.27

3.98

22.53

402.97

23.25

354.61

3.81

22.01

380.61

21.81

Day 3

386.85

4.37

23.65

415.07

24.34

Day 4

361.07

4.03

23.49

388.78

23.48

Day 5 Day 6 Mean

344.88 367.26 365.16

3.94 3.98 4.02 4.67

22.48 23.54 22.95

371.48 394.96 392.31

22.39 24.41 23.28

% RSD

4.13

3.03

3.98

4.46

Acetance Criteria Accetable (Y/N)

 5%

 5%

 5%

 5%

 5%

Y Y Remark: Vitamin B6 (as Pyridoxine) is sum of Pyridoxine, Pyridoxal and Pyridoxamine. All data have been mass corrected and expressed as Pyridoxine. Y Y Y

9. Accuracy

Analyze NIST SRM 1849a on 3 independent runs over multiple days.

Table 21: Accuracy of SRM 1849a

Result (mcg/100g)

Acceptable

Pantothenic acid 6630-7010

Thiamine Riboflavin Nicotinamide

Pyridoxine Biotin

Analyte

Y/N

1159- 1355

1985- 2089

1253- 1439

186- 212

Expected Value

9800-11800

Run 1 Run 2 Run 3

1283.78 1306.26 1309.12

1999.03 1990.94 2011.72

10280.78 10413.05 10150.15

6881.59 6790.94 6984.88

1357.95 191.47 1285.84 191.87 1309.87 199.05

Y Y Y

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