Dear DSQAP participants, 

Please find a copy of the report from Exercise C enclosed, we are happy to answer any 

questions you may have about the report and the exercise. We would like to thank the 

laboratories that sent additional method information; we are using the data to plan 

discussions for the next DSQAP workshop. 

Do you have any suggestions for Exercise D? Based on the results from Exercise C, we 

would recommend repeating some water soluble vitamins, possibly the phytosterols and 

fatty acids. Do you see any problem areas in the report? Are there other analyte/matrix 

pairs that would help you? If so, please let us know in an e-mail to dsqap@nist.gov. 

Best regards, 

DSQAP Team 

1 
Dietary Supplement Quality Assurance Program 

Exercise C 

Final Data Report 

Your laboratory code for this exercise: NIST 

This report consists of several parts: 

• Overview: a general description of the statistical treatment of the data, and how 

to read the plots. 

• Data Table:, a table with your laboratory’s individual results, the community 

results, and the NIST results. 

• Graphs: a section that includes graphical representation of the data for the 

analytes tested in this exercise, points to consider when examining the data, and 

when appropriate, recommendations from exercise C. 

Arsenic

Cadmium

Calcium

Phosphorus

Sodium

Zinc

Retinol

Niacin

Vitamin B6

Campesterol in Solution

β-sitosterol in Solution

Stigmasterol in Solution 

Campesterol in Serenoa repens Fruit

β-sitosterol in Serenoa repens Fruit

Stigmasterol in Serenoa repens Fruit

Linoleic Acid

α-Linolenic Acid

γ-Linolenic Acid

As always, if you have any questions, please contact us. 

2 
OVERVIEW 

STATISTICS: 

Your individual data table and graphs contain information about your performance 

relative to the rest of the participants and relative to a target around the expected result. 

INDIVIDUAL DATA TABLE 

Section 1 of the data table contains your results, including your mean and standard 

deviation. Please check these and make sure that you agree with the data in the table. 

Section 2 of the data table contains the community results, including the median value 

for each analyte, the MADe (a robust estimate of the standard deviation), and the 

minimum/maximum values reported for the analyte. 

Section 3 of the data table contains the NIST results. In most cases, the value and the 

U95 confidence interval have been determined with two independent analytical methods. 

At least six samples have been tested with each of the methods and duplicate sample 

preparations from the sample package have been included allowing for the U95 to 

encompass homogeneity within and between packages. 

GRAPHS 

Two graphs are provided for each analyte in each sample, one which plots lab results for 

the sample vs. the participating labs (View 1), the second plots lab results for the sample 

vs. lab results for the control (View 2). Both views include the consensus values and the 

target values. 

View 1 

Individual laboratory data are plotted with the individual laboratory standard deviation. 

The black solid line represents the consensus median and the black dotted lines represent 

the consensus variability. The center of the region with the red hash marks represents our 

view of the “correct answer”, it is bounded (+/-) two times the pooled standard deviation 

of the participants or two times the NIST uncertainty (whichever is larger), thus creating 

a target zone. With this view, it is relatively easy to determine if a laboratory falls within 

the target zone, it is also possible to compare where the target zone lies relative to the 

consensus values. In most cases, the target zone falls within the consensus values which 

is the expected result. One program goal is to bring the consensus values closer together 

and clustered around the target value. 

Return to Introduction

3 
View 2 

In this view the results reported for the sample are plotted vs. the results for the control. 

The red box represents the target values for the control (x-axis) and the sample (y-axis) 

and the black dotted lines represent the analogous information for the consensus values. 

This view provides additional information to complement view 1. For example, if your 

values are low for both the control and sample (or high for both), you may have 

calibration issues. If your laboratory falls into this category, you may want to investigate 

how your calibrants are prepared as well as the purity of your calibrant material. 

Return to Introduction

4 
DATA TABLE 

Return to Introduction

5 
ARSENIC

Arsenic View 1 

C101C102

--

C104

C105

C106C107

C108C109C112C114

C116

--

C118

--

C121

NIST

200

300

400

500

600

 Individual Lab Mean (±2SD)

 Consensus Median

 Consensus Variability

 Target

> 1000

Arsenic in Bitter Orange Extract, ng/g

> 2800

Return to Introduction

6 
Arsenic View 2

Return to Introduction

7 
CADMIUM 

Cadmium View 1 

--

C102C103C104C105C106C107C108C109

C112C114

--

C118C119

C121

NIST

0

10

20

30

40 > 70

Cadmium in Bitter Orange Extract, ng/g

Laboratory

> 120

Return to Introduction

8 
Cadmium View 2 

Return to Introduction

CALCIUM 

9 
Calcium View 1 

C101C102C103C104C105C107C108C109C112C114

C117C118C121

NIST

0.30

0.35

0.40

0.45

0.50

0.55

0.60

0.65

Calcium in Fortified Milk Powder, % mass fraction

Return to Introduction

10 
Calcium View 2 

Return to Introduction

11 
PHOSPHORUS 

Phosphorus View 1 

C101

--

C103

----

C107

C108

C109C112C114C117C118

C121

NIST

0.20

0.25

0.30

0.35

0.40

0.45

0.50 > 2.2

Phosphorus in Fortified Milk Powder, % mass fraction

Return to Introduction

12 
Phosphorus View 2 

Return to Introduction

13 
SODIUM 

Sodium View 1 

C101

--

C103

--

C105C107

C108C109C112

C114C117C118

C121

NIST

1000

2000

3000

4000

5000

6000

7000

Sodium in Fortified Milk Powder, mg/kg

Laboratory

Return to Introduction

14 
Sodium View 2 

Return to Introduction

15 
ZINC 

Zinc View 1 

C101C102C103C104C105C107

C10

C109C112C114

C117C118C121

NIST

100

125

150

175

200

8

 Individual Lab Mean (±2SD)

 Consensus Median

 Consensus Variability

 Target

Zinc in Fortified Milk Powder, mg/kg

Laboratory

Return to Introduction

16 
Zinc View 2 

Return to Introduction

17 
RETINOL 

The fortified milk powders used for this exercise were fortified with fat-soluble vitamins, 

water soluble vitamins, elements, and fatty acids. The control and sample were related 

but not identical. 

• Overall good results-only a few labs are outliers 

• Several labs have higher standard deviations than expected (based on a pooled 

standard deviation) 

• View 2 shows some evidence of calibration issues. Consider how the calibrants 

are made: 

• Are they traceable to a gravimetric number or a spectroscopic number? 

• Was the purity of the calibrant evaluated? 

• Did you see 13-cis-retinol and how was it factored in to your retinol 

calculation? 

Recommendations: 

• Check calibration method including traceability 

• If your individual laboratory results had a large RSD, consider the addition of an 

internal standard 

• Would you like to measure tocopherols in this material? 

Retinol View 1 

C101C104C107C108C110

111

C112C115C117C121

NIST

0

5

10

15

20

25

30

C

 Individual Lab Mean (±2SD)

 Consensus Median

 Consensus Variability

 Target

Retinol in Fortified Milk Powder, mg/kg

Laboratory

Return to Introduction

18 
Retinol View 2 

Return to Introduction

19 
NIACIN AS NIACINAMIDE 

The fortified milk powder used for this exercise was fortified with fat-soluble vitamins, 

water soluble vitamins, elements, and fatty acids. The control was SRM 3244 Ephedra 

Containing Protein Powder. The level of Niacinamide in the control is approximately 2.5 

times the level of niacinamide in the fortified milk powder. 

• The results are significantly more scattered than we had expected and the scatter 

appears to be random (not related to method) 

• Please check to be certain that you reported niacinamide. If you did not and you 

would like to submit amended data we are happy to accept it. 

Recommendations: 

• Check form of Niacin reported 

• We suggest that we repeat this exercise with a more closely related 

control/unknown sample pair. 

Niacin View 1 

C101

C103C104C107

--

C109C110

C111

C112C115

C117

C118

C120C121

NIST

0

100

200

300

400

500

600

 Individual Lab Mean (±2SD)

 Consensus Median

 Consensus Variability

 Target

Niacin in Fortified Milk Powder, mg/kg

Laboratory

Return to Introduction

20 
Niacin View 2 

Return to Introduction

21 
VITAMIN B6 AS PYRIDOXINE HCL 

The fortified milk powder used for this exercise was fortified with fat-soluble vitamins, 

water soluble vitamins, elements, and fatty acids. The control was SRM 3244 Ephedra 

Containing Protein Powder. The level of Niacinamide in the control is approximately 2.5 

times the level of niacinamide in the fortified milk powder. 

• The results are significantly scattered, as with the niacinamide, the scatter does 

not appear to be related to the analytical measurement technique. 

• Please check to be certain that you reported pyridoxine HCl. If you did not and 

you would like to submit amended data we are happy to accept it. 

Recommendations: 

• Check form of Pyridoxine HCl reported 

• We suggest that we repeat this exercise with a more closely related 

control/unknown sample pair.

Vitamin B6 View 1 

Return to Introduction

C101C103C104C107C108

C109C110

11

C112

C115C117

C118C120C121

NIST

0

5

10

15

20

25

30

35

40

45

50

Vitamin B6 in Fortified Milk Powder, mg/kg

C1

22 
Vitamin B6 View 2 

Return to Introduction

23 
PHYTOSTEROL SOLUTIONS 

The phytosterols solution was gravimetrically prepared mixture of campesterol, βsitosterol, and stigmasterol (plus some impurities) in chloroform. It was made at a 

concentration that would be appropriate for a calibrant and required only derivatization. 

The average measurements of the sterol solution appeared to be close to the expected 

value; however the standard deviations were larger than expected. Most laboratories 

“hydrolyzed” the solution; it appears that this step introduces uncertainty into the 

measurement. Most labs report adding an internal standard immediately prior to the 

sample derivatization step, adding the internal standard at the earliest opportunity (i.e. 

prior to extraction and hydrolysis) would almost likely reduce the uncertainty introduced 

with the extended sample handling. One lab mentioned problems weighing the solution 

reproducibly, did others experience this? 

• Results for phytosterols berries are significantly low (view 1) indicating: 

• Incomplete extraction 

• Incomplete hydrolysis 

• However, view 2 shows better results for the control indicating: 

• Hydrolysis is relatively complete 

• Extraction is the issue yielding significantly low results 

Campesterol in Phytosterol Solution View 1 

C101C107

C109

C114C118

NIST

0.0

0.1

0.2

0.3

0.4

0.5

0.6

Campesterol in Solution, mg/g

C110

Return to Introduction

24 
Campesterol in Phytosterol Solution View 2 

Return to Introduction

25 
β-SITOSTEROL IN SOLUTION 

A discussion of the measurement of phytosterols in Serenoa repens extract and Serenoa 

repens fruit is included with the campesterol in solution graphs. 

β-sitosterol in Solution View 1 

C101C107C109

C110C114

C118

NIST

0.0

0.5

1.0

1.5

2.0

2.5

3.0

β-sitosterol in Solution, mg/g

Return to Introduction

26 
β-sitosterol in Solution View 2 

Return to Introduction

27 
STIGMASTEROL IN SOLUTION 

A discussion of the measurement of phytosterols in Serenoa repens extract and Serenoa 

repens fruit is included with the campesterol in solution graphs. 

Stigmasterol in Solution View 1 

C101C107C109C110C114C118

NIST

0.00

0.05

0.10

0.15

0.20

0.25

0.30

Stigmasterol in Solution, mg/g

Laboratory

Return to Introduction

28 
Stigmasterol in Solution View 2 

Return to Introduction

29 
CAMPESTEROL IN SERENOA REPENS FRUIT 

A discussion of the measurement of phytosterols in Serenoa repens extract and Serenoa 

repens fruit is included with the campesterol in solution graphs. 

Campesterol in Serenoa repens Fruit View 1 

C101C107C109C110C114C118

NIST

0.00

0.05

0.10

0.15

0.20

Campesterol in Serenoa repens Fruit, mg/g

Return to Introduction

30 
Campesterol in Serenoa repens Fruit View 2 

Return to Introduction

31 
β-SITOSTEROL IN SERENOA REPENS FRUIT 

A discussion of the measurement of phytosterols in Serenoa repens extract and Serenoa 

repens fruit is included with the campesterol in solution graphs. 

β-sitosterol in Serenoa repens Fruit View 1 

C101C107

C109C110

C114

C118

NIST

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

β-sitosterol in Serenoa repens Fruit, mg/g

Return to Introduction

32 
β-sitosterol in Serenoa repens Fruit View 2 

Return to Introduction

33 
STIGMASTEROL IN SERENOA REPENS FRUIT 

A discussion of the measurement of phytosterols in Serenoa repens extract and Serenoa 

repens fruit is included with the campesterol in solution graphs. 

Stigmasterol in Serenoa repens Fruit View 1 

C101C107C109C110C114C118

NIST

0.00

0.01

0.02

0.03

0.04

0.05

0.06

0.07

 Individual Lab Mean (±2SD)

 Consensus Median

 Consensus Variability

 Target

Stigmasterol in Serenoa repens Fruit, mg/g

Laboratory

Return to Introduction

34 
Stigmasterol in Serenoa repens Fruit View 2 

Return to Introduction

35 
OMEGA-3 AND OMEGA-6 FATTY ACIDS IN BORAGE OIL 

Fatty Acids 

• View 1 indicates that in general the results for linoleic acid and γ-linoleic acid 

agree well with each other and the target 

• View 1 indicates that the participants agree well on α-linoleic acid and are 

significantly different than the target values-this is under further investigation (we 

suspect that there is a stability issue). 

• View 2 shows a correlation between results for the control and the unknown 

sample-in this case this cannot be definitively diagnosed as calibration issues; it 

could also be due to the derivatization of the sample. Would you like to 

investigate this further? 

Recommendation: 

• Measure Omega-3 and Omega-6 fatty acids in oils with more dissimilar ratios 

than the pair used in exercise C 

• Perhaps include a solution to determine if the correlation between the control and 

unknown sample results are due to the sample preparation or calibration. 

Linoleic Acid in Borage Oil View 1 

C101C104C105C107

C108

C109

C110

C113C114

C117

C121

NIST

200

250

300

350

400

450

500

550

 Individual Lab Mean (±2SD)

 Consensus Median

 Consensus Variability

 Target

Linoleic Acid in Borage Oil, mg/g

Return to Introduction

36 
Linoleic Acid in Borage Oil View 2 

Return to Introduction

37 
α-LINOLENIC ACID 

A discussion of the measurement of omega-3 and omega-6 fatty acids in botanical oils is 

included with the graphs of linoleic acid. 

 

α-Linolenic Acid in Borage Oil View 1 

C101C104

--

C107C108

C109

C110C113C114

C117

C121

NIST

0

1

2

3

4

5

α-Linolenic Acid in Borage Oil, mg/g

Return to Introduction

38 
α-Linolenic Acid in Borage Oil View 2 

Return to Introduction

39 
γ-LINOLENIC ACID 

A discussion of the measurement of omega-3 and omega-6 fatty acids in botanical oils is 

included with the graphs of linoleic acid. 

γ-Linolenic Acid in Borage Oil View 1 

C101C104C105C107C108C109C110C113

C114C117C121

NIST

150

200

250

300

350

γ-Linoleic Acid in Borage Oil, mg/g

Laboratory

Return to Introduction

40 
γ-Linolenic Acid in Borage Oil View 2 

Return to Introduction

41