AOAC SPIFAN Review Team Meeting Book-July 22, 2015

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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

Slope

(weighting: 1/x)

0.07177 0.00845

0.07257 0.00601

0.07348 0.005490

Y-Intercept Correlation Coefficient Acceptance Criteria

0.99902

0.99964

0.99950

Correlation Coefficient (r) ≥ 0.995

Acceptable Y/N

Y

Y

Y

3. Instrument Precision

Six replicates of each of the lowest, middle and highest standard concentrations were analyzed in a single analytical run. Table 9: Instrument Precision of Thiamine, Riboflavin and Nicotinic acid Thiamine HCl Riboflavin Nicotinic acid Area Ratio Area Ratio Area Ratio

Lowest Standard

Middle Standard

Highest Standard

Lowest Standard

Middle Standard

Highest Standard

Lowest Standard

Middle Standard

Highest Standard

Replicate 1

0.1902

1.1830

3.6200

0.1419

0.9851

2.8740

0.2246

0.4955

1.3260

Replicate 2

0.1923

1.1850

3.6270

0.1420

0.9466

2.8570

0.2227

0.4991

1.3060

Replicate 3

0.1923

1.1900

3.6620

0.1408

0.9431

2.9600

0.2240

0.4913

1.3260

Replicate 4

0.1922

1.1780

3.6440

0.1405

0.9573

2.9150

0.2236

0.4924

1.3330

Replicate 5

0.1930

1.1660

3.6310

0.1418

0.9625

2.8580

0.2141

0.5011

1.2940

Replicate 6

0.1946

1.1660

3.6340

0.1416

0.9945

2.8220

0.2166

0.4954

1.3280

Mean

0.1924 0.0014

1.1780 0.0101

3.6363 0.0149

0.1414 0.0006

0.9649 0.0208

2.8810 0.0491

0.2209 0.0044

0.4958 0.0038

1.3188 0.0153

Std. Dev.

% RSD

0.74

0.85

0.41

0.44

2.15

1.70

2.01

0.76

1.16

Acceptance Criteria %RSD Acceptable (Y/N)

RSD ≤ 5%

RSD ≤ 3%

RSD ≤ 3%

RSD ≤ 5%

RSD ≤ 3%

RSD ≤ 3%

RSD ≤ 5%

RSD ≤ 3%

RSD ≤ 3%

Y

Y

Y

Y

Y

Y

Y

Y

Y

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