NISTIR 7880-7

NIST Micronutrients Measurement 

Quality Assurance Program 

Winter 2009 

Comparability Studies 

Results for Round Robin LXV 

Fat-Soluble Vitamins and Carotenoids in Human Serum 

and Round Robin 30 Ascorbic Acid in Human Serum 

David L. Duewer 

Jeanice B. Thomas 

 

 

 

NISTIR 7880-7
 

NIST Micronutrients Measurement 

Quality Assurance Program 

Winter 2009 

Comparability Studies 

Results for Round Robin LXV 

Fat-Soluble Vitamins and Carotenoids in Human Serum 

and Round Robin 30 Ascorbic Acid in Human Serum 

David L. Duewer 

Jeanice B. Thomas 

Chemical Sciences Division 

Materials Measurement Laboratory 

April, 2013 

U.S. Department of Commerce 

Rebecca Blank, Acting Secretary 

National Institute of Standards and Technology 

Patrick D. Gallagher, Under Secretary of Commerce for Standards and Technology and Director 
(This page intentionally blank) 

ii
 
 

Abstract 

The National Institute of Standards and Technology coordinates the Micronutrients Measurement 

Quality Assurance Program (MMQAP) for laboratories that measure fat- and water-soluble vitamins 

and carotenoids in human serum and plasma. This report describes the design of and results for the 

Winter 2009 MMQAP measurement comparability improvement studies: 1) Round Robin LXV FatSoluble Vitamins and Carotenoids in Human Serum and 2) Round Robin 30 Total Ascorbic Acid in 

Human Serum. The materials for both studies were shipped to participants in December 2008; 

participants were requested to provide their measurement results by March 31, 2009. 

Keywords 

Human Serum
 

Retinol, α-Tocopherol, γ-Tocopherol, Total and Trans-β-Carotene
 

SRM 1950, Total Ascorbic Acid
 

iii 
 

 

 

 

 

 

 

Table of Contents 

Abstract............................................................................................................................................... iii
 

Keywords ............................................................................................................................................ iii
 

Table of Contents ............................................................................................................................... iv
 

Introduction......................................................................................................................................... 1
 

Round Robin LXV: Fat-Soluble Vitamins and Carotenoids in Human Serum .......................... 1
 

Round Robin 30: Vitamin C in Human Serum............................................................................... 2
 

References ............................................................................................................................................ 3
 

Appendix A. Shipping Package Inserts for RR65........................................................................ A1
 

Appendix B. Final Report for RR65 ..............................................................................................B1
 

Appendix C. “All-Lab Report” for RR65 ......................................................................................C1
 

Appendix D. Representative “Individualized Report” for RR65 ............................................... D1
 

Appendix E. Shipping Package Inserts for RR30 ......................................................................... E1
 

Appendix F. Final Report for RR30............................................................................................... F1
 

Appendix G. “All-Lab Report” for RR30..................................................................................... G1
 

Appendix H. Representative “Individualized Report” for RR30............................................... H1
 

iv 
Introduction 

Beginning in 1988, the National Institute of Standards and Technology (NIST) has coordinated the 

Micronutrients Measurement Quality Assurance Program (MMQAP) for laboratories that measure 

fat- and water-soluble vitamins and carotenoids in human serum and plasma. The MMQAP 

provides participants with measurement comparability assessment through use of interlaboratory 

studies, Standard Reference Materials (SRMs) and control materials, and methods development and 

validation. Serum-based samples with assigned values for the target analytes (retinol, alphatocopherol, gamma/beta-tocopherol, trans- and total beta-carotene, and total ascorbic acid) and 

performance-evaluation standards are distributed by NIST to laboratories for analysis. 

Participants use the methodology of their choice to determine analyte content in the control and 

study materials. Participants provide their data to NIST, where it is compiled and evaluated for 

trueness relative to the NIST value, within-laboratory precision, and concordance within the 

participant community. NIST provides the participants with a technical summary report concerning 

their performance for each exercise and suggestions for methods development and refinement. 

Participants who have concerns regarding their laboratory’s performance are encouraged to consult 

with the MMQAP coordinators. 

All MMQAP interlaboratory studies consist of individual units of batch-prepared samples that are 

distributed to each participant. For historical reasons these studies are referred to as “Round 

Robins”. The MMQAP program and the nature of its studies are described elsewhere. [1,2] 

Round Robin LXV: Fat-Soluble Vitamins and Carotenoids in Human Serum 

Participants in the MMQAP Fat-Soluble Vitamins and Carotenoids in Human Serum Round Robin 

LXV comparability study (hereafter referred to as RR65) received one lyophilized and four liquidfrozen human serum test samples for analysis. Unless multiple vials were previously requested, 

participants received one vial of each serum. These sera were shipped on dry ice to participants in 

December 2008. The communication materials included in the sample shipment are provided in 

Appendix A. 

Participants are requested to report values for all fat-soluble vitamin-related analytes that are of 

interest to their organizations. Not all participants report values for the target analytes, and many 

participants report values for non-target analytes. 

The final report delivered to every participant in RR65 consists of three documents: 

•	 A cover letter for the current study, a brief description of the other two documents, and a 

discussion of our analysis of the overall results that may be of broad interest. This cover 

letter is reproduced as Appendix B. 

•	 The “All-Lab Report” that lists all of the reported measurement results, a number of 

consensus statistics for analytes reported by more than one participant, and the mean median 

and pooled SD from any prior distributions of the serum. This report also provides a 

numerical “score card” for each participant’s measurement comparability for the more 

commonly reported analytes. This report is reproduced as Appendix C. 

1
 
 

 

•	 An “Individualized Report” that graphically analyzes each participant’s results for all 

analytes reported by at least five participants. This report also provides a graphical summary 

of their measurement comparability. The graphical tools used in this report are described in 

detail elsewhere [3]. An example “Individualized Report” is reproduced as Appendix D. 

Round Robin 30: Vitamin C in Human Serum 

Participants in the MMQAP Vitamin C in Human Serum Round Robin 30 comparability study 

(hereafter referred to as RR30) received four frozen serum test samples, one frozen control serum, a 

solid ascorbic acid control material, and one optional unknown (SRM 1950 Metabolites in Human 

Serum) for analysis. Unless multiple vials were previously requested, participants received one vial 

of each material. These sample materials were shipped on dry ice to participants in December 2008. 

The communication materials included in the sample shipment are provided in Appendix E. 

The test and control serum materials were prepared by adding equal volumes of 10 % 

metaphosphoric acid (MPA) to human serum that had been spiked with ascorbic acid. While these 

samples contain some dehydroascorbic acid, its content is variable. Therefore, the participants 

report only total ascorbic acid (TAA, ascorbic acid plus dehydroascorbic acid). Participants are also 

encouraged to prepare calibration solutions from the supplied solid control to enable calibrating their 

serum measurements to the same reference standard. 

SRM 1950 is a human plasma pool designed for the evaluation of the measurement of metabolites in 

humans to provide insight into various disease states. The SRM 1950 plasma contains lithium 

heparin as an anticoagulant but does not contain MPA. Participants were asked to analyze SRM 

1950 as an unknown to help evaluate whether TAA could be usefully certified in this material. 

The final report delivered to every participant in RR30 consists of three documents: 

•	 A cover letter for the current study, a brief description of the other two documents, and a 

discussion of our analysis of overall results that may be of broad interest. This cover letter is 

reproduced as Appendix F. 

•	 The “All-Lab Report” that summarizes all of the reported measurement results and provides 

several consensus statistics. This report is reproduced as Appendix G. 

•	 An “Individualized Report” that graphically analyzes each participant’s results for TAA, 

including a graphical summary of their measurement comparability. The graphical tools 

used in this report are described in detail elsewhere [3]. An example “Individualized Report” 

is reproduced as Appendix H. 

2
 
 

References 

1 Duewer DL, Brown Thomas J, Kline MC, MacCrehan WA, Schaffer R, Sharpless KE, May WE, 

Crowell JA. NIST/NCI Micronutrients Measurement Quality Assurance Program: Measurement 

Repeatabilities and Reproducibilities for Fat-Soluble Vitamin-Related Compounds in Human 

Sera. Anal Chem 1997;69(7):1406-1413. 

2 Margolis SA, Duewer DL. Measurement Of Ascorbic Acid in Human Plasma and Serum: 

Stability, Intralaboratory Repeatability, and Interlaboratory Reproducibility. Clin Chem 

1996;42(8):1257-1262. 

3 Duewer DL, Kline MC, Sharpless KE, Brown Thomas J, Gary KT, Sowell AL. Micronutrients 

Measurement Quality Assurance Program: Helping Participants Use Interlaboratory Comparison 

Exercise Results to Improve Their Long-Term Measurement Performance. Anal Chem 

1999;71(9):1870-1878. 

3
 
Appendix A. Shipping Package Inserts for RR65 

The following three items were included in each package shipped to an RR65 participant: 

• Cover letter 

• Datasheet 

• Packing List and Shipment Receipt Confirmation Form 

The cover letter and datasheet were enclosed in a sealed waterproof bag along with the 

samples themselves. The packing list was placed at the top of the shipping box, between 

the cardboard covering and the foam insulation. 

A1
December 3, 2008 

Dear Colleague: 

UNITED STATES DEPARTMENT OF COMMERCE 

National Institute of Standard& and Technology 

Gaithersburg, Maryland 20899-

Enclosed are samples for the first fat-soluble vitamins and carotenoids in serum study (Round Robin 

LXV) for the 2009 NIST Micronutrients Measurement Quality Assurance Program. The set of samples 

(Sera 3 52 - 3 56) consists of one vial of each of four liquid-frozen and one lyophilized serum samples for 

analysis along with a fonn for reporting your results. These samples should be stored in the dark at or 

below -20 °C upon receipt. When reporting your results, please submit one value for each analyte for a 

given serum sample. If a value ·obtained is below your limit of quantification, please indicate this result 

on the fonn by using NQ (Not Quantified). Results are due to NIST by March 31, 2009.· Results 

received more than two weeks after the due date may not be included in the summary report for this round 

robin study. The feedback report concerning the study will be distributed in April 2009. 

Lyophilized samples should be reconstituted with 1.0 mL ofHPLC-grade water or equivalent. Before 

reconstitution, samples should be allowed to stand at room temperature under subdued light until thawed. 

We recommend that dissolution be facilitated with 3 to 5 min agitation in an ultrasonic bath or at least 30 

min at room temperature with intermittent swirling. (CAUTION: Vigorous shaking will cause foaming 

and possibly interfere with accurate measurement. The rubber stopper contains phthalate esters that may 

leach into the sample upon intermittent contact of the liquid sample with the stopper. These esters absorb 

strongly in the UV region and elute near retinol in most LC systems creating analytical problems.) · 

Pipette a known volume of serum from the vial for analysis. The final volume of the reconstituted sample 

is greater than 1.0 mL. Water should not be added to the liquid-frozen samples. 

For consistency; we request that laboratories use the following absorptivities (dL/g ·cm): retinal, 1843 at 

325 nm (ethanol); retinyl palmitate, 975 at 325 nm (ethanol); a.-tocopherol,. 75.8 at 292 nm .(ethanol); ytocopherol, 91.4 at 298 nm (ethanol); a-carotene, 2800 at 444 nm (hexane); 13-carotene, 2560 at 450 run 

(ethanol), 2592 at 452 run (hexane); and lycopene, 3450 at 472 run (hexane). 

Please report your results for Round Robin LXV by e-mail to david.duewer@nist.gov or fax to 301-977-

0685. If you have questions or comments regarding this study, please call me at (301) 975-3120 or e-mail 

me atjbthomas@nist.gov. 

Je nice Brown Thomas 

esearch Chemist 

Analytical Chemistry Division 

Chemical Science and Technology Laboratory 

Enclosures 

N•Si 

A2
 

 

 

Participant #: ________ Date: _____________ 

Round Robin LXV: Human Sera 

NIST Micronutrients Measurement Quality Assurance Program 

Analyte 352 353 354 355 356 Units* 

total retinol 

trans-retinol 

didehydroretinol 

retinyl palmitate 

α-tocopherol 

γ/β-tocopherol 

δ-tocopherol 

total β-carotene 

trans-β-carotene 

total cis-β-carotene 

total α-carotene 

total lycopene 

trans-lycopene 

total β-cryptoxanthin 

total α-cryptoxanthin 

total lutein 

total zeaxanthin 

total lutein&zeaxanthin

 total coenzyme Q10 

ubiquinol (QH2) 

ubiquinone (Qox) 

phylloquinone (K1) 

25-hydroxyvitamin D 

Other measurands? 

* we prefer μg/mL 

Were the liquid-frozen samples (353 to 356) frozen when received? Yes | No 

Comments: 

Mail: M2QAP
 

NIST, Stop 8392 Fax: 301-977-0685
 

Gaithersburg, MD 20899-8392 Please return by March 31, 2009 Email: David.Duewer@NIST.gov
 

A3
 

 

 

 

Participant #: ________ Date: _____________ 

Fat-Soluble Vitamins Round Robin LXV
 

NIST Micronutrients Measurement Quality Assurance Program
 

Packing List and Shipment Receipt Confirmation Form 

This box contains: one vial each of the following five FSV M2QAP sera 

Serum Form Reconstitute? Vial/Cap 

#352 Lyophilized Yes (1 ml H2O) 2 mL amber, silver 

#353 Liquid frozen No 2 mL amber, green cap 

#354 Liquid frozen No 2 mL amber, red cap 

#355 Liquid frozen No 2 mL amber, blue cap 

#356 Liquid frozen No 2 mL amber, green cap 

Please 1) Open the pack immediately 

2) Check that it contains all of the above samples 

3) Check if the vials are intact 

4) Store the sera at -20  ̊C or below until analysis 

5) Complete the following information 

6) Fax the completed form to us at 301-977-0685 

(or email requested information to david.duewer@nist.gov) 

1) Date this shipment arrived: ___________________________ 

2) Are all five sera vials intact? Yes | No 

If "No", which one(s) were damaged? 

3) Was there any dry-ice left in cooler? Yes | No 

4) Did the liquid frozen samples arrive frozen? Yes | No 

5) At what temperature are you storing the serum samples? _______  ̊C 

6) When do you anticipate analyzing these samples? ________________________ 

Your prompt return of this information is appreciated. 

The M2QAP Gang 

Mail: M2QAP 

NIST, Stop 8392 Fax: 301-977-0685 

Gaithersburg, MD 20899-8392 Email: David.Duewer@NIST.gov 

A4
 

 

Appendix B. Final Report for RR65 

The following four pages are the final report as provided to all participants: 

•	 Cover letter. 

•	 An information sheet that: 

o	 describes the contents of the “All-Lab” report, 

o	 describes the content of the “Individualized” report, 

o	 describes the nature of the test samples and details their previous distributions, 

if any, and 

o	 summarizes aspects of the study that we believe may be of interest to the 

participants. 

B1
B2
 

 

The NIST M2QAP Round Robin LXV (RR65) report consists of: 

Page 

1-4 

5 

6 

Page 

1 

2-n 

n+1 

Samples. 

Serum

352 

353 

354 

355 

356 

Results 

“All Lab” Report 

A listing of all results and statistics for analytes reported by two or more participants. 

A listing of results reported by only one laboratory and the legend for pages 1-4. 

The text Comparability Summary (“Score Card”) of measurement performance. 

“Individualized” Report 

Your values, the number of labs reporting values, and our assigned values. 

“Four Plot” summaries of your current and past measurement performance, one page for 

each analyte you report that is also reported by at least eight other participants. 

The graphical Comparability Summary (target plot) of measurement performance. 

Five samples were distributed in RR65. 

Description 

Lyophilized 1+1 blend of the pools used to produce 

SRM 968c Levels I and II. 

Fresh-frozen 1+0.03 blend of a normal serum and a 

low-normal serum spiked with trans-β-carotene in a 

lipoprotein carrier. The same materials were used to 

prepare #354 and #355. 

Fresh-frozen 1+0.08 blend of the materials used to 

prepare #353. 

Fresh-frozen 1+0.19 blend of the materials used to 

prepare #353. 

Fresh-frozen 1+0.56 blend of the residual pools used 

to produce SRM 968c Levels I and II. 

Prior Distributions 

#247:RR44-9/98, #317:RR58-9/05 

New, produced Fall 2008 

New, produced Fall 2008 

New, produced Fall 2008 

New, produced Fall 2008 

1)	 Serum #352: This material is a 1+1 mixture of the left-over pools for SRM 968c Level I and Level II 

(where 1+1 means equal volumes of the two pools and left-over means the material left in the carboy 

after stirring became problematic). The material was prepared in 1998, soon after the SRM 968c 

units were packed away, to test how well we could predict measurand levels in a mixture. This is the 

third time this material has been distributed. There has been no statistically significant change in the 

level or variability of any measurand. However, the measured level of total lutein and total lutein 

plus zeaxanthin has somewhat decreased with time. The level of total zeaxanthin does not appear to 

have changed. Given the relatively few measurements available, the apparent decline in lutein is 

mostly a curiosity. We will, however, re-examine the archived data for other materials that have been 

stored for ten or more years. 

B3
 

2)	 Serum #356: This material is also a mixture of the left-over pools for SRM 968c Level I and Level II. 

However, this material was mixed and ampouled in 2009. While there was no surety that results for 

SRM 968c were really applicable to these residual materials, we were again interested in how well we 

can predict the levels in a blend. Table 1 contrasts our predictions for the #352 material (lyophilized, 

stored for 11 years in 2-mL glass vials) with those for #356 (liquid frozen in Teflon for 11 years). 

The “Calc” values are our predictions based upon MMQAP results for the six distributions of the 

SRM 968c Level I and Level II materials over the past ten years, “Obs” are the median results in RR 

LXV, and “%Dif” is 100*(Calc – Obs)/Obs. Note: The “Calc” results of Table 1 define the dark blue 

“expectation” lines for these two SRM 968c-related materials in your “Individualized” Report. 

 Table 1: Calculated and Observed Results for Mixtures of the SRM 968c Level I and II Pools 

Serum #352 Serum #356 

Analyte Calc Obs %Dif Calc Obs %Dif 

Total Retinol 0.67 0.69 -2 0.72 0.76 -5 

α-Tocopherol 12.1 11.4 6 10.8 10.6 2 

γ/β-Tocopherol 2.7 2.6 3 3.0 2.5 24 

Total β-Carotene 0.30 0.31 -4 0.26 0.27 -1 

Total α-Carotene 0.056 0.054 4 0.045 0.044 3 

Total Lycopene 0.36 0.38 -4 0.35 0.37 -7 

trans-Lycopene 0.18 0.17 7 0.18 0.18 0 

Total β-Cryptoxanthin 0.052 0.056 -7 0.058 0.059 -2 

Total Lutein 0.071 0.058 23 0.066 0.055 20 

Total Zeaxanthin 0.027 0.024 16 0.029 0.030 -5 

Total Lutein&Zeaxanthin 0.100 0.091 10 0.097 0.097 0 

Most of the predicted levels are within about 10 % of those observed. Interestingly, total lutein is 

about 20 % less than predicted in both materials. While total zeaxanthin and total lutein plus 

zeaxanthin are somewhat lower than expected in the #352 material but not in the #356, the major 

difference between the two materials is for γ/β-tocopherol, where the level in the #356 material is 

about 25 % lower than expected. We as yet have no defensible explanation for this anomaly. 

3)	 Sera #353, #354, and #355. These three materials are the culmination of an experiment begun in late 

1999 testing whether carotenoids could be successfully augmented. The materials are all mixtures of 

the same β-carotene-deficient but otherwise fairly normal serum and a trans-β-carotene spiking 

solution. Following a number of false-starts, this solution was prepared in the late 1990’s by mixing 

trans-β-carotene and high- and low-density lipoproteins into a low-normal serum followed by 

extensive mixing and filtering. Due to resource constraints, the solution sat unused until this spring. 

Following further mixing and filtering, the three materials were prepared at the same time as #356. 

Serum #353 was spiked with just enough of the solution to raise the level of trans-β-carotene to about 

that of the #352 and #356 materials. Serum #354 was spiked to a reasonably high trans-β-carotene 

level and #355 was spike to an unreasonably high level, just to determine the limits of augmentation. 

Figure 1 displays the results for all of the β-carotene results reported in RR65. Each line represents 

the results for a single participant. The line segments o the left connect results for #352 and #356; the 

lines to the right connect results for #353, #354, and #355. The x-axis spacing for the three spiked 

materials reflects the relative proportion of the spike, so that ideal results will produce a straight line. 

B4
Too low 

Total 

β-Carotene, mg/mL

1.2 

1.0 

0.8 

0.6 

0.4 

0.2 

Too high 

Just Right 

Median±15% 

Mixtures of a low-normal serum 

and a trans -β-Carotene spike 

0.0
 

352356353354355 

Serum 

Figure 1: RR65 Results for Total β-Carotene 

The thick black lines bound the region ± 15 % about the median result. The thin blue lines denote the 

participant results that (mostly) fall within this region. The thick green lines denote the results for 

#355 that are more than 15 % higher than the median; the thick red lines denote the results that are 

(mostly) more than 15 % lower than expected. 

We hypothesize that the “too low” results arise from extraction processes that are not adequate 

enough to completely extract β-carotene. However, there is no correlation between the relative 

location of the “too low” results of the spiked materials and in the two SRM 968c-related mixtures. 

Therefore, the relative under-extraction in the artificial matrix does not imply under-extraction in a 

native matrix. Also, the “too low” results are quite consistently low for all three of the spiked 

materials, thereby indicating that the problem is with the nature of the spike and not its proportion in 

the mixture. 

The “too high” results are less easily explained, since we do not believe that extraction from the 

artificial matrix should be easier than from the native. However, we actually expected the spiked 

levels to be somewhat higher than the median results so it is quite plausible that the “too high” values 

are in fact the “true” values for a really complete extraction. 

Regardless of the causes, the RR65 results strongly suggest that we cannot (yet) successfully augment 

carotenoid levels. We will continue to try to provide you with sera that have interesting levels and 

patterns of analytes, but we are constrained in what we can deliver by the native levels in 

commercially available sera. 

B5
 

 

 

Appendix C. “All-Lab Report” for RR65 

The following six pages are the “All-Lab Report” as provided to all participants, with two 

exceptions: 

• the participant identifiers (Lab) have been altered. 

• the order in which the participant results are listed has been altered. 

The data summary in the “All-Lab Report” has been altered to ensure confidentiality of 

identification codes assigned to laboratories. The only attributed results are those 

reported by NIST. The NIST results are not used in the assessment of the consensus 

summary results of the study. 

C1
 

Round Robin LXV Laboratory Results 

Total Retinol, μg/mL trans-Retinol, μg/mL Retinyl Palmitate, μg/mL α-Tocopherol, μg/mL γ/β-Tocopherol, μ

Lab 352 353 354 355 356 

C2

FSV-BA 0.666 0.318 0.320 0.292 0.781 

FSV-BB 0.750 0.326 0.333 0.300 0.799 

FSV-BC 0.683 0.335 0.331 0.301 0.804 

FSV-BD 0.593 0.310 0.312 0.289 0.778 

FSV-BE 0.525 0.239 0.250 0.233 0.589 

FSV-BF 0.790 0.340 0.350 0.310 0.800 

FSV-BG 0.700 0.325 0.324 0.297 0.774 

FSV-BH 0.545 0.247 0.246 0.215 0.647 

FSV-BJ 0.698 0.315 0.319 0.293 0.749 

FSV-BK 

FSV-BL 0.690 0.320 0.320 0.290 0.740 

FSV-BM 0.690 0.360 0.360 0.350 0.770 

FSV-BN 0.574 0.246 0.245 0.235 0.624 

FSV-BO 0.673 0.327 0.347 0.295 0.768 

FSV-BP 0.689 0.339 0.335 0.304 0.680 

FSV-BQ 0.740 0.330 0.320 0.290 0.800 

FSV-BR ≥0.650 ≥0.290 ≥0.290 ≥0.260 ≥0.620

FSV-BS ≥0.698 ≥0.356 ≥0.353 ≥0.326 ≥0.745

FSV-BU 0.650 0.264 0.313 0.294 0.702 

FSV-BV 0.727 0.317 0.312 0.261 0.776 

FSV-BW 0.728 0.338 0.342 0.316 0.805 

FSV-CC 0.565 0.310 0.314 0.340 0.806 

FSV-CD 0.620 0.280 0.290 0.260 0.670 

FSV-CE 0.668 0.264 0.286 0.267 0.599 

FSV-CF 0.843 0.349 0.399 0.291 0.862 

FSV-CG 0.803 0.337 0.334 0.314 0.737 

FSV-CI 0.818 0.321 0.323 0.302 0.812 

FSV-CP 

FSV-CW 0.697 0.268 0.285 0.241 0.752 

FSV-CZ 0.664 0.309 0.397 0.293 0.584 

FSV-DD 0.590 0.270 0.280 0.260 0.690 

FSV-DV ≥0.638 ≥0.259 ≥0.258 ≥0.230 ≥0.594

FSV-EE 0.612 0.304 0.294 0.279 0.643 

FSV-EZ ≥0.572 ≥0.259 ≥0.251 ≥0.233 ≥0.708

352 353 354 355 356 

0.650 0.290 0.290 0.260 0.620 

0.698 0.356 0.353 0.326 0.745 

0.529 0.310 0.314 0.340 0.806 

0.638 0.259 0.258 0.230 0.594 

0.572 0.259 0.251 0.233 0.708 

352 353 354 355 356 

0.058 0.066 0.072 0.096 0.053 

0.049 0.017 0.021 0.031 0.040 

0.054 0.037 0.036 0.036 0.051 

0.061 nq nq nq nq 

0.040 0.001 0.002 0.001 0.034 

0.058 0.010 0.010 0.012 0.077 

0.040 0.010 0.020 0.010 0.040 

0.071 0.039 0.051 0.034 0.056 

0.034 0.006 0.008 0.006 0.036 

0.049 <0.02 <0.02 <0.02 0.045 

352 353 354 355 356 

10.86 7.73 7.69 6.73 10.24 

11.96 8.03 8.13 7.13 10.55 

12.20 8.80 8.40 7.30 10.70 

10.57 6.83 6.79 6.25 9.58 

11.20 7.70 7.70 6.80 11.10 

12.28 8.54 8.37 7.38 11.44 

10.85 7.59 7.91 6.74 10.44 

12.55 8.48 8.51 7.47 11.07 

13.44 8.27 7.97 7.32 10.35 

12.06 8.18 8.18 7.32 10.77 

11.90 8.80 9.10 8.10 11.70 

9.25 6.07 5.98 5.49 8.20 

9.50 7.80 7.70 6.90 10.20 

10.67 9.03 8.45 7.25 9.66 

11.40 6.90 7.80 6.90 10.70 

12.54 8.40 8.20 6.20 9.70 

11.32 7.58 7.38 6.79 10.26 

13.17 8.98 9.11 7.30 11.74 

12.69 8.55 8.53 7.67 11.05 

9.37 7.85 8.26 7.56 10.70 

13.91 9.27 6.67 8.51 12.40 

10.07 7.70 8.04 6.91 7.80 

12.90 8.10 8.70 6.90 10.80 

10.67 7.37 7.03 6.22 9.52 

11.94 7.70 7.70 6.93 10.57 

9.58 6.41 6.26 5.32 8.24 

12.07 8.27 7.97 7.13 10.57 

12.64 6.98 7.25 6.25 9.95 

10.90 7.40 7.20 6.40 9.70 

9.70 7.80 7.40 6.60 9.60 

11.43 6.89 6.98 6.58 11.48 

352 353 354 3

2.80 3.50 3.49 3

2.78 3.23 3.27 2

2.00 2.41 2.42 2

2.75 3.54 3.51 3

3.00 3.67 3.67 3

2.73 3.43 3.49 3

2.06 2.39 2.34 2

2.40 3.30 3.30 2

2.28 2.96 2.82 2

2.58 3.26 3.28 2

2.95 3.65 3.65 3

2.66 3.06 3.19 2

2.63 3.17 3.08 2

3.24 3.80 3.77 3

2.55 2.68 2.87 2

2.62 3.15 3.08 2

2.73 3.15 3.27 2

2.41 3.00 3.09 2

N 28 28 28 

Min 0.525 0.239 0.245 

Median 0.686 0.318 0.320 

Max 0.843 0.360 0.399 

SD 0.071 0.027 0.027 

CV 10 9 9 

Npast 41 0 0 

Medianpast 0.673 

SDpast 0.051 

NIST 0.683 0.309 0.308 

NNIST 2 2 2 

Mean 0.683 0.309 0.308 

Srep 0.006 0.004 0.010 

Shet 0.017 0.011 0.007 

SNIST 0.018 0.012 0.012 

NAV 0.686 0.313 0.314 

NAU 0.059 0.027 0.028 

28 

0.215 

0.293 

0.350 

0.019 

6 

0 

0.276 

2 

0.276 

0.011 

0.002 

0.011 

0.285 

0.027 

28 

0.584 

0.760 

0.862 

0.066 

9 

0 

0.722 

2 

0.722 

0.027 

0.013 

0.030 

0.745 

0.068 

5 5 5 5 5 

0.529 0.259 0.251 0.230 0.594 

0.638 0.290 0.290 0.260 0.708 

0.698 0.356 0.353 0.340 0.806 

0.089 0.046 0.047 0.044 0.130 

14 16 16 17 18 

6 0 0 0 0 

0.681 

0.059 

0.638 0.290 0.290 0.260 0.708 

0.089 0.046 0.047 0.044 0.130 

10 8 8 8 9 

0.034 0.001 0.002 0.001 0.034 

0.052 0.014 0.021 0.022 0.045 

0.071 0.066 0.072 0.096 0.077 

0.012 0.015 0.021 0.020 0.012 

23 112 103 94 26 

13 0 0 0 0 

0.052 

0.015 

0.052 0.014 0.021 0.022 0.045 

0.016 0.015 0.021 0.020 0.015 

31 31 31 31 31 

9.25 6.07 5.98 5.32 7.80 

11.43 7.80 7.91 6.90 10.55 

13.91 9.27 9.11 8.51 12.40 

1.260 0.697 0.755 0.593 0.812 

11 9 10 9 8 

41 0 0 0 0 

11.619 

0.993 

11.65 7.87 8.09 6.93 10.71 

2 2 2 2 2 

11.653 7.872 8.094 6.927 10.715 

0.601 0.429 0.110 0.026 0.284 

0.270 0.398 0.109 0.201 0.227 

0.659 0.585 0.155 0.203 0.363 

11.542 7.836 8.002 6.914 10.633 

1.270 0.699 0.766 0.593 0.827 

18 18 18 

2.00 2.39 2.34 2

2.65 3.20 3.27 2

3.24 3.80 3.77 3

0.212 0.351 0.328 0.3

8 11 10 

23 0 0 

2.730 

0.183 

2.61 2.95 3.11 2

2 2 2 

2.608 2.954 3.106 2.8

0.059 0.013 0.086 0.0

0.123 0.058 0.110 0.0

0.136 0.060 0.139 0.

2.627 3.076 3.187 2.7

0.262 0.391 0.348 0.3

All Lab Report 
 

Round Robin LXV Laboratory Results 

Total β-Carotene, μg/mL trans-β-Carotene, μg/mL Total cis-β-Carotene, μg/mL Total α-Carotene, μg/mL Total Lycopene, μ

Lab 352 353 354 355 356 

C3

FSV-BA 0.292 0.299 0.448 0.868 0.256 

FSV-BB 0.294 0.295 0.453 0.896 0.253 

FSV-BC 

FSV-BD 

FSV-BE 0.311 0.211 0.247 0.476 0.266 

FSV-BF 0.267 0.306 0.495 1.050 0.275 

FSV-BG 0.356 0.380 0.540 1.186 0.307 

FSV-BH 0.299 0.336 0.482 0.976 0.259 

FSV-BJ 0.319 0.280 0.368 0.843 0.295 

FSV-BK 

FSV-BL 

FSV-BM 

FSV-BN 0.340 0.229 0.329 0.629 0.269 

FSV-BO 0.326 0.309 0.470 0.898 0.263 

FSV-BP 0.315 0.298 0.475 1.254 0.322 

FSV-BQ 

FSV-BR 

FSV-BS ≥0.290 ≥0.333 ≥0.523 ≥0.870 ≥0.255

FSV-BU 0.326 0.371 0.531 0.936 0.261 

FSV-BV 0.367 0.399 0.639 1.170 0.314 

FSV-BW 0.355 0.323 0.425 0.879 0.354 

FSV-CC 

FSV-CD 0.340 0.330 0.480 1.030 0.280 

FSV-CE 0.300 0.359 0.612 1.153 0.103 

FSV-CF 

FSV-CG 0.299 0.328 0.491 0.964 0.255 

FSV-CI 0.268 0.270 0.329 0.604 0.205 

FSV-CP 0.253 0.257 0.362 0.743 0.202 

FSV-CW 0.319 0.332 0.512 0.925 0.277 

FSV-CZ 0.395 0.271 0.373 0.552 0.308 

FSV-DD 

FSV-DV 

FSV-EE 

FSV-EZ ≥0.266 ≥0.345 ≥0.529 ≥1.223 ≥0.294

352 353 354 355 356 

0.274 0.285 0.428 0.833 0.243 

0.274 0.281 0.433 0.855 0.239 

0.283 0.322 0.462 0.933 0.248 

0.304 0.210 0.301 0.569 0.252 

0.290 0.333 0.523 0.870 0.255 

0.274 0.303 0.455 0.901 0.233 

0.266 0.345 0.529 1.223 0.294 

352 353 354 355 356 

0.019 0.014 0.020 0.036 0.013 

0.020 0.014 0.021 0.041 0.013 

0.016 0.014 0.020 0.043 0.011 

0.036 0.019 0.028 0.060 0.017 

0.025 0.024 0.036 0.063 0.021 

352 353 354 355 356 

0.049 0.032 0.032 0.030 0.043 

0.048 0.028 0.029 0.024 0.042 

0.056 0.038 0.034 0.033 0.051 

0.043 nq nq nq 0.040 

0.058 0.032 0.034 0.031 0.057 

0.056 0.025 0.025 0.024 0.044 

0.024 0.026 0.026 0.027 0.044 

0.057 0.021 0.047 0.038 0.065 

0.050 0.037 0.039 0.040 0.040 

0.056 0.040 0.034 0.027 0.056 

0.071 0.040 0.043 0.033 0.063 

0.047 0.019 0.020 0.021 0.039 

0.050 0.030 0.030 0.040 0.040 

0.066 0.049 0.048 0.045 0.060 

0.082 0.062 0.058 0.059 0.062 

0.052 0.030 0.033 0.028 0.042 

0.054 0.028 0.027 0.026 0.049 

352 353 354 3

0.416 0.581 0.597 0.

0.376 0.538 0.533 0.

0.375 0.519 0.497 0.

0.367 0.518 0.515 0.

0.416 0.533 0.572 0.

0.331 0.437 0.464 0.

0.262 0.443 0.441 0.

0.396 0.321 0.408 0.

0.404 0.531 0.544 0.

0.381 0.566 0.543 0.

0.442 0.584 0.627 0.

0.448 0.644 0.628 0.

0.320 0.380 0.430 0.

0.311 0.506 0.507 0.

0.280 0.365 0.364 0.

N 20 20 20 20 20 7 7 7 7 7 5 5 5 5 5 17 16 16 16 17 15 15 15 

Min 0.253 0.211 0.247 0.476 0.103 0.266 0.210 0.301 0.569 0.233 0.016 0.014 0.020 0.036 0.011 0.024 0.019 0.020 0.021 0.039 0.262 0.321 0.364 0.

Median 0.317 0.307 0.472 0.912 0.268 0.274 0.303 0.455 0.870 0.248 0.020 0.014 0.021 0.043 0.013 0.054 0.031 0.033 0.031 0.044 0.376 0.519 0.515 0.

Max 0.395 0.399 0.639 1.254 0.354 0.304 0.345 0.529 1.223 0.294 0.036 0.024 0.036 0.063 0.021 0.082 0.062 0.058 0.059 0.065 0.448 0.644 0.628 0.

SD 0.034 0.042 0.079 0.190 0.020 0.012 0.033 0.040 0.055 0.010 0.006 0.000 0.001 0.010 0.003 0.006 0.009 0.009 0.008 0.007 0.059 0.092 0.085 0.

CV 11 14 17 21 7 4 11 9 6 4 30 0 7 24 23 11 29 26 26 16 16 18 16 

Npast 28 0 0 0 0 13 0 0 0 0 9 0 0 0 0 25 0 0 0 0 24 0 0 

Medianpast 0.320 0.301 0.024 0.055 0.370 

SDpast 0.043 0.034 0.008 0.012 0.078 

NIST 0.326 0.303 0.482 0.885 0.264 0.048 0.029 0.031 0.031 0.050 

NNIST 2 2 2 2 2 2 2 2 2 2 

Mean 0.326 0.303 0.482 0.885 0.264 0.048 0.029 0.031 0.031 0.050 

Srep 0.021 0.007 0.020 0.030 0.020 0.002 0.002 0.004 0.004 0.002 

Shet 0.006 0.011 0.009 0.035 0.001 0.001 0.001 0.000 0.001 0.003 

SNIST 0.022 0.013 0.022 0.046 0.020 0.002 0.002 0.004 0.005 0.003 

NAV 0.321 0.305 0.477 0.898 0.266 0.274 0.303 0.455 0.870 0.248 0.020 0.014 0.021 0.043 0.013 0.051 0.030 0.032 0.031 0.047 0.376 0.519 0.515 0.

NAU 0.046 0.045 0.079 0.191 0.039 0.030 0.033 0.048 0.090 0.027 0.007 0.005 0.007 0.017 0.005 0.017 0.010 0.011 0.010 0.015 0.082 0.107 0.106 0.

All Lab Report 
 

Round Robin LXV Laboratory Results 

Total β-Cryptoxanthin, μg/mL Total α-Cryptoxanthin, μg/mL Total Lutein, μg/mL Total Zeaxanthin, μg/mL Total Lutein&Zeaxanthin, μg/

Lab 352 353 354 355 356 

C4

FSV-BA 0.056 0.097 0.098 0.090 0.061 

FSV-BB 0.056 0.093 0.096 0.084 0.061 

FSV-BC 

FSV-BD 

FSV-BE 

FSV-BF 

FSV-BG 0.058 0.100 0.097 0.088 0.061 

FSV-BH 0.060 0.100 0.102 0.095 0.066 

FSV-BJ 0.056 0.089 0.092 0.082 0.058 

FSV-BK 

FSV-BL 

FSV-BM 

FSV-BN 0.051 0.088 0.088 0.081 0.054 

FSV-BO 0.070 0.077 0.078 0.074 0.050 

FSV-BP 0.056 0.043 0.044 0.036 0.052 

FSV-BQ 

FSV-BR 

FSV-BS 0.046 0.074 0.081 0.076 0.049 

FSV-BU 0.075 0.110 0.131 0.111 0.080 

FSV-BV 0.050 0.089 0.093 0.076 0.057 

FSV-BW 0.042 0.090 0.089 0.075 0.046 

FSV-CC 

FSV-CD 0.070 0.110 0.120 0.130 0.070 

FSV-CE 

FSV-CF 

FSV-CG 0.061 0.132 0.135 0.134 0.082 

FSV-CI 

FSV-CP 0.054 0.091 0.090 0.079 0.060 

FSV-CW 0.046 0.080 0.075 0.070 0.050 

FSV-CZ 

FSV-DD 

FSV-DV 

FSV-EE 

FSV-EZ 

352 353 354 355 356 

0.020 0.034 0.035 0.031 0.020 

0.017 0.032 0.033 0.028 0.020 

0.007 0.019 0.022 0.017 0.007 

0.028 0.041 0.039 0.028 0.026 

0.060 0.100 0.110 0.100 0.060 

352 353 354 355 356 

0.067 0.130 0.132 0.113 0.063 

0.052 0.101 0.096 0.087 0.050 

0.069 0.137 0.139 0.125 0.068 

0.054 0.107 0.109 0.100 0.054 

0.058 0.120 0.117 0.108 0.056 

0.047 0.092 0.090 0.075 0.042 

0.057 0.119 0.101 0.093 0.052 

0.073 0.133 0.139 0.122 0.074 

352 353 354 355 356 

0.040 0.068 0.066 0.050 0.040 

0.027 0.060 0.057 0.050 0.030 

0.020 0.047 0.047 0.043 0.023 

nd 0.086 0.071 0.071 0.045 

0.019 0.045 0.044 0.031 0.021 

0.013 0.026 0.024 0.023 0.014 

0.039 0.077 0.083 0.076 0.044 

352 353 354 355 

0.082 0.171 0.172 0.154 

0.107 0.199 0.197 0.163 

0.098 0.174 0.162 0.142 

0.079 0.161 0.153 0.137 

0.074 0.151 0.152 0.140 

0.058 0.206 0.188 0.179 

0.094 0.088 0.090 0.102 

0.066 0.137 0.134 0.106 

0.103 0.196 0.212 0.181 

0.107 0.234 0.232 0.187 

0.082 0.204 0.203 0.169 

0.230 0.360 0.400 0.360 

0.122 0.251 0.252 0.224 

0.070 0.145 0.125 0.116 

0.083 0.168 0.165 0.150 

0.113 0.209 0.221 0.199 

N 16 16 16 16 16 5 5 5 5 5 8 8 8 8 8 6 7 7 7 7 16 16 16 16 

Min 0.042 0.043 0.044 0.036 0.046 0.007 0.019 0.022 0.017 0.007 0.047 0.092 0.090 0.075 0.042 0.013 0.026 0.024 0.023 0.014 0.058 0.088 0.090 0.102 

Median 0.056 0.091 0.093 0.082 0.059 0.020 0.034 0.035 0.028 0.020 0.058 0.120 0.113 0.104 0.055 0.024 0.060 0.057 0.050 0.030 0.088 0.185 0.180 0.159 

Max 0.075 0.132 0.135 0.134 0.082 0.060 0.100 0.110 0.100 0.060 0.073 0.137 0.139 0.125 0.074 0.040 0.086 0.083 0.076 0.045 0.230 0.360 0.400 0.360 

SD 0.007 0.014 0.011 0.011 0.010 0.012 0.010 0.006 0.004 0.009 0.011 0.019 0.027 0.021 0.010 0.011 0.022 0.019 0.028 0.015 0.024 0.036 0.044 0.033 

CV 13 15 12 13 18 59 31 17 16 44 19 16 24 20 17 48 37 34 56 49 28 19 25 21 

Npast 26 0 0 0 0 6 0 0 0 0 16 0 0 0 0 15 0 0 0 0 24 0 0 0 

Medianpast 0.057 0.019 0.068 0.027 0.100 

SDpast 0.011 0.002 0.015 0.009 0.025 

NIST 0.058 0.100 0.101 0.083 0.060 0.059 0.121 0.118 0.101 0.061 0.034 0.062 0.062 0.048 0.034 0.093 0.183 0.179 0.149 

NNIST 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 

Mean 0.058 0.100 0.101 0.083 0.060 0.059 0.121 0.118 0.101 0.061 0.034 0.062 0.062 0.048 0.034 0.093 0.183 0.179 0.149 

Srep 0.002 0.002 0.003 0.001 0.001 0.001 0.002 0.002 0.001 0.002 0.005 0.003 0.002 0.004 0.002 0.005 0.005 0.003 0.005 

Shet 0.001 0.003 0.000 0.001 0.004 0.004 0.005 0.005 0.011 0.003 0.001 0.001 0.002 0.003 0.004 0.005 0.004 0.007 0.014 

SNIST 0.002 0.004 0.003 0.002 0.004 0.004 0.006 0.005 0.011 0.004 0.005 0.003 0.003 0.005 0.004 0.007 0.006 0.007 0.015 

NAV 0.057 0.095 0.097 0.082 0.059 0.020 0.034 0.035 0.028 0.020 0.058 0.120 0.115 0.103 0.058 0.029 0.061 0.059 0.049 0.032 0.091 0.184 0.180 0.154 

NAU 0.014 0.022 0.022 0.019 0.014 0.012 0.010 0.006 0.004 0.009 0.012 0.022 0.027 0.021 0.012 0.013 0.022 0.020 0.028 0.015 0.025 0.039 0.044 0.034 

All Lab Report 
Round Robin LXV Laboratory Results 

Phylloquinone (K1), ng/mL 25-hydroxyvitamin D, μg/mL 

C5

Lab 352 353 354 355 356 352 353 354 355 356 

FSV-BA 

FSV-BB 0.013 0.008 0.009 0.010 0.014 

FSV-BC 

FSV-BD 

FSV-BE 1.188 0.587 0.578 0.605 1.132 

FSV-BF 

FSV-BG 

FSV-BH 0.012 0.009 0.010 0.010 0.012 

FSV-BJ 

FSV-BK 

FSV-BL 

FSV-BM 

FSV-BN 

FSV-BO 

FSV-BP 

FSV-BQ 

FSV-BR 

FSV-BS 

FSV-BU 

FSV-BV 

FSV-BW 

FSV-CC 

FSV-CD 

FSV-CE 

FSV-CF 

FSV-CG 

FSV-CI 1.837 0.820 0.714 <0.03 1.253 

FSV-CP 

FSV-CW 

FSV-CZ 

FSV-DD 

FSV-DV 

FSV-EE 

FSV-EZ 

N 

Min 

Median 

Max 

SD 

CV 

Npast 

Medianpast 

SDpast 

NIST 

NNIST 

Mean 

Srep 

Shet 

SNIST 

NAV 

NAU 

2 2 2 1 2 2 2 2 2 2 

1.188 0.587 0.578 1.132 0.012 0.008 0.009 0.010 0.012 

1.513 0.704 0.646 0.605 1.193 0.013 0.008 0.009 0.010 0.013 

1.837 0.820 0.714 1.253 0.013 0.009 0.010 0.010 0.014 

0 0 0 0 0 0 0 0 0 0 

All Lab Report 
 

 

 

 

 

 

 

Round Robin LXV Laboratory Results 

Analytes Reported By One Laboratory 

Term 

N Number of (non-NIST) quantitative values reported for this analyte

Min Minimum (non-NIST) quantitative value reported

Median Median (non-NIST) quantitative value reported

Max Maximum (non-NIST) quantitative value reported

SD Standard deviation for (non-NIST) results: 0.741*(3rd Quartile - 1st Quartile)

CV Coefficient of Variation for (non-NIST) results: 100*SD/Median

Npast Mean of N(s) from past RR(s)

Medianpast Mean of Median(s) from past RR(s)

SDpast Pooled SD from past RR(s)

MeanNIST Mean of NIST results

Srep NIST's within-vial pooled standard deviation

Shet NIST's among-vial pooled standard deviation 

SNIST Combined standard deviation for NIST analyses: √(Srep 2

 + Shet 2)

NAV NIST Assigned Value

 = (Median + MeanNIST)/2 for analytes reported by NIST analyst(s) 

= Median for analytes reported by ≥ 5 labs but not NIST

NAU NIST Assigned Uncertainty: √(S 2

 + Sbtw 2)

 S is the maximum of (0.05*NAV, SD, SNIST, eSD) and Sbtw is the standard

 deviation between Median and MeanNIST. The expected long-term SD, eSD,

 is defined in: Duewer et al., Anal Chem 1997;69(7):1406-1413.

nd Not detected (i.e., no detectable peak for analyte)

nq Detected but not quantitatively determined 

≥x Concentration greater than or equal to x

italics Not explicitly reported but calculated by NIST from reported values 

Analyte Code 352 353 354 355 356 

Ubiquinol FSV-BW 1.283 1.265 1.543 1.207 0.555 

Ubiquinone FSV-BW 0.066 0.320 0.274 0.126 0.528 

Legend

All Lab Report 

C6

Page 5 / 5 
 

 

 

 

 

 

 

 

Lab 

FSV-BA 

FSV-BB 

FSV-BC 

FSV-BD 

FSV-BE 

FSV-BF 

FSV-BG 

FSV-BH 

FSV-BJ 

FSV-BK 

FSV-BL 

FSV-BM 

FSV-BN 

FSV-BO 

FSV-BP 

FSV-BQ 

FSV-BR 

FSV-BS 

FSV-BU 

FSV-BV 

FSV-BW 

FSV-CC 

FSV-CD 

FSV-CE 

FSV-CF 

FSV-CG 

FSV-CI 

FSV-CP 

FSV-CW 

FSV-CZ 

FSV-DD 

FSV-DV 

FSV-EE 

FSV-EZ 

NIST 

n 

% 1 

% 2 

% 3 

% 4 

Round Robin LXV Laboratory Results
 

Comparability Summary
 

TR aT g/bT bC tbC aC TLy TbX TLu TZ L&Z 

1 1 1 1 1 1 1 1 1 

1 1 1 1 1 1 1 1 1 1 1 

1 

1 1 

3 2 3 2 

2 1 1 

1 1 1 2 1 1 1 1 

3 1 2 1 1 1 1 1 1 1 1 

1 1 1 1 1 1 1 1 

1 

1 1 

2 2 

3 3 3 2 3 1 1 1 1 1 1 

1 1 1 1 1 1 1 1 1 1 

1 2 2 2 2 2 2 

1 1 

2 2 

2 1 1 1 1 1 2 1 2 

2 1 1 1 1 1 2 1 

1 2 1 2 1 1 1 1 

1 1 2 2 1 2 1 1 

2 2 

2 3 1 1 1 1 2 4 

2 2 3 

3 1 

2 2 1 1 1 2 1 2 2 

2 1 2 2 3 1 2 2 

3 2 2 1 2 1 1 

2 1 1 1 1 1 2 2 1 

2 2 1 2 

2 

3 1 

2 1 

3 2 1 2 3 

1 1 1 1 1 1 1 1 1 

33 32 19 23 7 18 15 17 9 8 17 

TR 

39 

42 

18 

0 

aT 

59 

31 

9 

0 

g/bT 

68 

21 

11 

0 

bC 

52 

43 

4 

0 

tbC 

71 

0 

29 

0 

aC 

83 

11 

6 

0 

TLy TbX TLu 

80 76 78 

20 24 22 

0 0 0 

0 0 0 

TZ 

75 

25 

0 

0 

L&Z 

71 

24 

0 

6 

Label

Lab 

TR 

aT 

g/bT

bC 

tbC 

aC 

TLy 

TbX 

TLu 

TZ 

L&Z

n 

% 1 

% 2 

% 3 

% 4 

 Definition 

 Participant code 

 Total Retinol 

α-Tocopherol 

 γ/β-Tocopherol 

 Total β-Carotene 

 trans-β-Carotene 

 Total α-Carotene 

 Total Lycopene 

Total β-Cryptoxanthin 

 Total Lutein 

 Total Zeaxanthin 

 Total Lutein & Zeaxanthin 

 number of participants providing quantitative data 

 Percent of CS = 1 (within 1 SD of medians) 

 Percent of CS = 2 (within 2 SD of medians) 

 Percent of CS = 3 (within 3 SD of medians) 

 Percent of CS = 4 (3 or more SD from medians) 

“Comparability Score” 

The Comparability Score (CS) of summarizes your measurement 

performance for a given measurand, relative to the consensus 

medians. CS is the average distance, in standard deviation units, 

that your measurement performance characteristics are from the 

consensus performance. CS is calculated when the number of 

quantitative values you reported for a measurand, Nyou, is at least 

two and the measurand has been reported by 10 or more 

participants. 

CS = MIN(4, INT(1 + C2 + AP 2 )) 

N yo u You − Median C = Concordanc e = i i ∑ Ny ou NAU i i 

N yo u  You i − Median i 

2 

AP = Apparent Precision = ∑  (Ny ou − 1)  NAU  i  i  

NAU = NIST Assigned Uncertainty, our estimate of the overall 

measurement standard deviation for each sample. The 

estimate includes serum heterogeneity, analytical 

repeatability, and among-participant reproducibility variance 

components. 

For further details, please see: Duewer DL, Kline MC, Sharpless 

KE, Brown Thomas J, Gary KT. Micronutrients Measurement 

Quality Assurance Program: Helping participants use 

interlaboratory comparison exercise results to improve their longterm measurement performance. Anal Chem 1999;71(9):1870-8. 

All Lab Report C7 Page 6 / 6 
 

 

Appendix D. Representative “Individualized Report” for RR65 

Each participant in RR65 received an “Individualized Report” reflecting their reported 

results. Each report included a detailed analysis for analytes that were assayed by at least 

five participants. The following analytes met this criterion in RR65: 

• Total Retinol 

• trans-Retinol 

• Retinyl Palmitate 

• α-Tocopherol 

• γ/β-Tocopherol 

• Total β-Carotene 

• trans-β-Carotene 

• Total cis-β-Carotene 

• Total α-Carotene 

• Total Lycopene 

• trans-Lycopene 

• Total β-Cryptoxanthin 

• Total α-Cryptoxanthin 

• Total Lutein 

• Total Zeaxanthin 

• Total Lutein & Zeaxanthin 

• Coenzyme Q10 

The following thirteen pages are the “Individualized Report” for the analytes evaluated 

by participant FSV-BA. 

D1
 

 Set 1 of 38 

Individualized Round Robin LXV Report: FSV-BA 

Summary 

Serum 352 Serum 353 Serum 354 Serum 355 S

D2

Analyte You NAV n You NAV n You NAV n You NAV n Yo

Total Retinol 0.666 0.685 28 0.318 0.313 28 0.320 0.314 28 0.292 0.284 28 0.7

Retinyl Palmitate 0.06 0.05 10 0.1 0.0 8 0.1 0.0 8 0.10 0.02 8 0.0

α-Tocopherol 10.86 11.54 31 7.73 7.84 31 7.69 8.00 31 6.73 6.91 31 10.2

γ/β-Tocopherol 2.795 2.627 18 3.495 3.076 18 3.485 3.187 18 3.064 2.794 18 2.7

δ-Tocopherol 0.338 0.330 4 0.104 0.092 4 0.106 0.084 4 0.098 0.061 4 0.2

Total β-Carotene 0.292 0.319 20 0.299 0.305 20 0.448 0.477 20 0.868 0.898 20 0.2

trans-β-Carotene 0.274 0.274 7 0.285 0.303 7 0.428 0.455 7 0.833 0.870 7 0.2

Total cis-β-Carotene 0.019 0.020 4 0.014 0.014 4 0.020 0.021 4 0.036 0.043 5 0.0

Total α-Carotene 0.049 0.051 17 0.032 0.030 16 0.032 0.032 16 0.030 0.031 16 0.0

Total Lycopene 0.416 0.376 15 0.581 0.519 15 0.597 0.515 15 0.571 0.467 15 0.4

trans-Lycopene 0.218 0.186 7 0.322 0.227 7 0.330 0.228 7 0.310 0.230 7 0.2

Total β-Cryptoxanthin 0.056 0.057 16 0.097 0.095 16 0.098 0.097 16 0.090 0.082 16 0.0

Total α-Cryptoxanthin 0.020 0.020 5 0.034 0.034 5 0.035 0.035 5 0.031 0.028 5 0.0

Total Lutein&Zeaxanthin 0.082 0.092 16 0.171 0.186 16 0.172 0.181 16 0.154 0.154 16 0.0

You : Your reported values for the listed analytes (micrograms/milliliter) 

NAV : NIST Assigned Values, here equal to this RR's median 

n : Number of non-NIST laboratories reporting quantitative values for this analyte in this serum 

Please check our records against your records. Send corrections and/or updates to... 

Micronutrients Measurement Quality Assurance Program 

National Institute of Standards and Technology Tel: 

100 Bureau Drive Stop 8392 Fax: 

Gaithersburg, MD 20899-8392 USA Email: david.du

Individualized Report 
NIST, this RR

D3

 Set 1 of 38 

Individualized RR LXV Report: FSV-BA

Total Retinol, μg/mL 

Apparent Precision, SD [Analyte], μg/mL

247317352356353354355 

Your [Analyte], μg/mL 

Apparent Precision, SD 

Concordance, SD 

3rd Quartile (75%) You, this RR You, ≥x, this RR 

Median [Analyte], μg/mL 

Long-term Precision, SD 

Median (50%) You, past RRs You, ≥x, past RRs Others, this RR 

1st Quartile (25%) Expectation 

For details of the construction and interpretation of these plots, see: 

Duewer, Kline, Sharpless, Brown Thomas, Gary, Sowell. Anal Chem 1999;71(9):1870-8. 

Serum Comments History 

#352 1+1 mixture of multi-donor pools #248 and #249 44:#247, 58:#317 

#356 1+0.56 mixture of multi-donor pools #248 and #249 New 

#353 126+4 mixture of single-donor #353 with spike pool New 

#354 120+10 mixture of single-donor #353 with spike pool New 

#355 105+25 mixture of single-donor #353 with spike pool New 

Individualized Report Page 2 / 13 
NIST, this RR

D4

 Set 1 of 38 

Individualized RR LXV Report: FSV-BA

Retinyl Palmitate, μg/mL 

Apparent Precision, SD [Analyte], μg/mL

247317352356353354355 

Your [Analyte], μg/mL 

Apparent Precision, SD 

Concordance, SD 

3rd Quartile (75%) You, this RR You, ≥x, this RR 

Median [Analyte], μg/mL 

Long-term Precision, SD 

Median (50%) You, past RRs You, ≥x, past RRs Others, this RR 

1st Quartile (25%) Expectation 

For details of the construction and interpretation of these plots, see: 

Duewer, Kline, Sharpless, Brown Thomas, Gary, Sowell. Anal Chem 1999;71(9):1870-8. 

Serum Comments History 

#352 1+1 mixture of multi-donor pools #248 and #249 44:#247, 58:#317 

#356 1+0.56 mixture of multi-donor pools #248 and #249 New 

#353 126+4 mixture of single-donor #353 with spike pool New 

#354 120+10 mixture of single-donor #353 with spike pool New 

#355 105+25 mixture of single-donor #353 with spike pool New 

Individualized Report Page 3 / 13 
NIST, this RR

D5

 Set 1 of 38 

Individualized RR LXV Report: FSV-BA

α-Tocopherol, μg/mL 

Apparent Precision, SD [Analyte], μg/mL

247317352356353354355 

Your [Analyte], μg/mL 

Apparent Precision, SD 

Concordance, SD 

3rd Quartile (75%) You, this RR You, ≥x, this RR 

Median [Analyte], μg/mL 

Long-term Precision, SD 

Median (50%) You, past RRs You, ≥x, past RRs Others, this RR 

1st Quartile (25%) Expectation 

For details of the construction and interpretation of these plots, see: 

Duewer, Kline, Sharpless, Brown Thomas, Gary, Sowell. Anal Chem 1999;71(9):1870-8. 

Serum Comments History 

#352 1+1 mixture of multi-donor pools #248 and #249 44:#247, 58:#317 

#356 1+0.56 mixture of multi-donor pools #248 and #249 New 

#353 126+4 mixture of single-donor #353 with spike pool New 

#354 120+10 mixture of single-donor #353 with spike pool New 

#355 105+25 mixture of single-donor #353 with spike pool New 

Individualized Report Page 4 / 13 
NIST, this RR

D6

 Set 1 of 38 

Individualized RR LXV Report: FSV-BA

γ/β-Tocopherol, μg/mL 

Apparent Precision, SD [Analyte], μg/mL

247317352356353354355 

Your [Analyte], μg/mL 

Apparent Precision, SD 

Concordance, SD 

3rd Quartile (75%) You, this RR You, ≥x, this RR 

Median [Analyte], μg/mL 

Long-term Precision, SD 

Median (50%) You, past RRs You, ≥x, past RRs Others, this RR 

1st Quartile (25%) Expectation 

For details of the construction and interpretation of these plots, see: 

Duewer, Kline, Sharpless, Brown Thomas, Gary, Sowell. Anal Chem 1999;71(9):1870-8. 

Serum Comments History 

#352 1+1 mixture of multi-donor pools #248 and #249 44:#247, 58:#317 

#356 1+0.56 mixture of multi-donor pools #248 and #249 New 

#353 126+4 mixture of single-donor #353 with spike pool New 

#354 120+10 mixture of single-donor #353 with spike pool New 

#355 105+25 mixture of single-donor #353 with spike pool New 

Individualized Report Page 5 / 13 
NIST, this RR

D7

 Set 1 of 38 

Individualized RR LXV Report: FSV-BA

Total β-Carotene, μg/mL 

Apparent Precision, SD [Analyte], μg/mL

247317352356353354355 

Your [Analyte], μg/mL 

Apparent Precision, SD 

Concordance, SD 

3rd Quartile (75%) You, this RR You, ≥x, this RR 

Median [Analyte], μg/mL 

Long-term Precision, SD 

Median (50%) You, past RRs You, ≥x, past RRs Others, this RR 

1st Quartile (25%) Expectation 

For details of the construction and interpretation of these plots, see: 

Duewer, Kline, Sharpless, Brown Thomas, Gary, Sowell. Anal Chem 1999;71(9):1870-8. 

Serum Comments History 

#352 1+1 mixture of multi-donor pools #248 and #249 44:#247, 58:#317 

#356 1+0.56 mixture of multi-donor pools #248 and #249 New 

#353 126+4 mixture of single-donor #353 with spike pool New 

#354 120+10 mixture of single-donor #353 with spike pool New 

#355 105+25 mixture of single-donor #353 with spike pool New 

Individualized Report Page 6 / 13 
NIST, this RR

D8

 Set 1 of 38 

Individualized RR LXV Report: FSV-BA

trans-β-Carotene, μg/mL 

Apparent Precision, SD [Analyte], μg/mL

247317352356353354355 

Your [Analyte], μg/mL 

Apparent Precision, SD 

Concordance, SD 

3rd Quartile (75%) You, this RR You, ≥x, this RR 

Median [Analyte], μg/mL 

Long-term Precision, SD 

Median (50%) You, past RRs You, ≥x, past RRs Others, this RR 

1st Quartile (25%) Expectation 

For details of the construction and interpretation of these plots, see: 

Duewer, Kline, Sharpless, Brown Thomas, Gary, Sowell. Anal Chem 1999;71(9):1870-8. 

Serum Comments History 

#352 1+1 mixture of multi-donor pools #248 and #249 44:#247, 58:#317 

#356 1+0.56 mixture of multi-donor pools #248 and #249 New 

#353 126+4 mixture of single-donor #353 with spike pool New 

#354 120+10 mixture of single-donor #353 with spike pool New 

#355 105+25 mixture of single-donor #353 with spike pool New 

Individualized Report Page 7 / 13 
NIST, this RR

D9

 Set 1 of 38 

Individualized RR LXV Report: FSV-BA

Total α-Carotene, μg/mL 

Apparent Precision, SD [Analyte], μg/mL

247317352356353354355 

Your [Analyte], μg/mL 

Apparent Precision, SD 

Concordance, SD 

3rd Quartile (75%) You, this RR You, ≥x, this RR 

Median [Analyte], μg/mL 

Long-term Precision, SD 

Median (50%) You, past RRs You, ≥x, past RRs Others, this RR 

1st Quartile (25%) Expectation 

For details of the construction and interpretation of these plots, see: 

Duewer, Kline, Sharpless, Brown Thomas, Gary, Sowell. Anal Chem 1999;71(9):1870-8. 

Serum Comments History 

#352 1+1 mixture of multi-donor pools #248 and #249 44:#247, 58:#317 

#356 1+0.56 mixture of multi-donor pools #248 and #249 New 

#353 126+4 mixture of single-donor #353 with spike pool New 

#354 120+10 mixture of single-donor #353 with spike pool New 

#355 105+25 mixture of single-donor #353 with spike pool New 

Individualized Report Page 8 / 13 
NIST, this RR

D10

 Set 1 of 38 

Individualized RR LXV Report: FSV-BA

Total Lycopene, μg/mL 

Apparent Precision, SD [Analyte], μg/mL

247317352356353354355 

Your [Analyte], μg/mL 

Apparent Precision, SD 

Concordance, SD 

3rd Quartile (75%) You, this RR You, ≥x, this RR 

Median [Analyte], μg/mL 

Long-term Precision, SD 

Median (50%) You, past RRs You, ≥x, past RRs Others, this RR 

1st Quartile (25%) Expectation 

For details of the construction and interpretation of these plots, see: 

Duewer, Kline, Sharpless, Brown Thomas, Gary, Sowell. Anal Chem 1999;71(9):1870-8. 

Serum Comments History 

#352 1+1 mixture of multi-donor pools #248 and #249 44:#247, 58:#317 

#356 1+0.56 mixture of multi-donor pools #248 and #249 New 

#353 126+4 mixture of single-donor #353 with spike pool New 

#354 120+10 mixture of single-donor #353 with spike pool New 

#355 105+25 mixture of single-donor #353 with spike pool New 

Individualized Report Page 9 / 13 
NIST, this RR

D11

 Set 1 of 38 

Individualized RR LXV Report: FSV-BA

trans-Lycopene, μg/mL 

Apparent Precision, SD [Analyte], μg/mL

247317352356353354355 

Your [Analyte], μg/mL 

Apparent Precision, SD 

Concordance, SD 

3rd Quartile (75%) You, this RR You, ≥x, this RR 

Median [Analyte], μg/mL 

Long-term Precision, SD 

Median (50%) You, past RRs You, ≥x, past RRs Others, this RR 

1st Quartile (25%) Expectation 

For details of the construction and interpretation of these plots, see: 

Duewer, Kline, Sharpless, Brown Thomas, Gary, Sowell. Anal Chem 1999;71(9):1870-8. 

Serum Comments History 

#352 1+1 mixture of multi-donor pools #248 and #249 44:#247, 58:#317 

#356 1+0.56 mixture of multi-donor pools #248 and #249 New 

#353 126+4 mixture of single-donor #353 with spike pool New 

#354 120+10 mixture of single-donor #353 with spike pool New 

#355 105+25 mixture of single-donor #353 with spike pool New 

Individualized Report Page 10 / 13 
NIST, this RR

D12

 Set 1 of 38 

Individualized RR LXV Report: FSV-BA

Total β-Cryptoxanthin, μg/mL 

Apparent Precision, SD [Analyte], μg/mL

247317352356353354355 

Your [Analyte], μg/mL 

Apparent Precision, SD 

Concordance, SD 

3rd Quartile (75%) You, this RR You, ≥x, this RR 

Median [Analyte], μg/mL 

Long-term Precision, SD 

Median (50%) You, past RRs You, ≥x, past RRs Others, this RR 

1st Quartile (25%) Expectation 

For details of the construction and interpretation of these plots, see: 

Duewer, Kline, Sharpless, Brown Thomas, Gary, Sowell. Anal Chem 1999;71(9):1870-8. 

Serum Comments History 

#352 1+1 mixture of multi-donor pools #248 and #249 44:#247, 58:#317 

#356 1+0.56 mixture of multi-donor pools #248 and #249 New 

#353 126+4 mixture of single-donor #353 with spike pool New 

#354 120+10 mixture of single-donor #353 with spike pool New 

#355 105+25 mixture of single-donor #353 with spike pool New 

Individualized Report Page 11 / 13 
NIST, this RR

D13

 Set 1 of 38 

Individualized RR LXV Report: FSV-BA

Total Lutein&Zeaxanthin, μg/mL 

Apparent Precision, SD [Analyte], μg/mL

247317352356353354355 

Your [Analyte], μg/mL 

Apparent Precision, SD 

Concordance, SD 

3rd Quartile (75%) You, this RR You, ≥x, this RR 

Median [Analyte], μg/mL 

Long-term Precision, SD 

Median (50%) You, past RRs You, ≥x, past RRs Others, this RR 

1st Quartile (25%) Expectation 

For details of the construction and interpretation of these plots, see: 

Duewer, Kline, Sharpless, Brown Thomas, Gary, Sowell. Anal Chem 1999;71(9):1870-8. 

Serum Comments History 

#352 1+1 mixture of multi-donor pools #248 and #249 44:#247, 58:#317 

#356 1+0.56 mixture of multi-donor pools #248 and #249 New 

#353 126+4 mixture of single-donor #353 with spike pool New 

#354 120+10 mixture of single-donor #353 with spike pool New 

#355 105+25 mixture of single-donor #353 with spike pool New 

Individualized Report Page 12 / 13 
D14

 Set 1 of 38 

Individualized Round Robin LXV Report: FSV-BA 

Graphical Comparability Summary
 

Total Retinol α-Tocopherol Total α-Carotene T

trans-Retinol γ/β-Tocopherol Total Lycopene Tot

Retinyl Palmitate Total β-Carotene trans-Lycopene Total L

trans-β-Carotene Total β-Cryptoxanthin Co

Individualized Report 
 

 

Appendix E. Shipping Package Inserts for RR30 

The following five items were included in each package shipped to an RR30 participant: 

•	 Cover letter 

•	 Protocol for Preparation and Analysis of the Ascorbic Acid Solid Control
 

Material
 

•	 Preparation and Validation of Ascorbic Acid Solid Control Material Datasheet 

•	 Analysis of Control Materials and Test Samples Datasheet 

•	 Packing List and Shipment Receipt Confirmation Form 

The cover letter, preparation protocol, and the two datasheets were enclosed in a sealed 

waterproof bag along with the samples themselves. The packing list was placed at the 

top of the shipping box, between the cardboard covering and the foam insulation. 

E1
December 3, 2008 

Dear Colleague: 

UNITED STATES DEPARTMENT OF COMMERCE 

National Institute of Standards and Technology 

Gaithersburg, Maryland 20899-

The samples within this package constitute Vitamin C Round Robin 30 (RR30) of the 2009 Micronutrients 

Measurement Quality Assurance Program. 

RR30 consists of four vials of frozen serum test samples (#27, #57, #117, and #119), one vial of frozen control 

serum (CS #1), and one vial of ascorbic acid solid control material (Control). Please follow the attached protocols 

when y'ou prepare and analyze these samples. If you cannot prepare the solid control solutions gravimetrically, 

please prepare equivalent solutions volumetrically and report the exact volumes used. (Routine 0.5 g gravimetric 

measurements are generally 10-fold more accurate than routine 0.5 mL volumetric measurements.) 

Also included for analysis is one vial ofSRM 1950 Metabolites in Human Serum. We would appreciate your 

assistance in the value-assignment of ascorbic acid in this material. SRM 1950 is a human plasma pool that was 

collected from healthy male and female donors. It contains lithium heparin as an anticoagulant. Please note: 

SRM 1950 does not contain metaphosphoric acid. Therefore, please prepare this'sample as you would a routine 

plasma sample and provide us with any preparation and dilution details you consider relevant. 

Please use the control serum to validate the performance of your measurement system before you analyze the test 

samples. The target value and ::::95% confidence interval for CS #1 is 8.4 ± 0.7 μmol/L of sample. 

The report for RR29 was e-mailed November 25, 2008. If you find your results for RR29 unsatisfactory, we 

recommend that you obtain Standard Reference Material (SRM) 970 Ascorbic Acid in Serum to validate your 

methodology and value assign in-house control materials. This SRM may be purchased from the Standard 

Materials Reference Program at NIST (Tel: 301-975-6776, Fax: 301-948-3730, or e-mail: srminfo@nist.gov). 

Please be aware that sample contact with any oxidant-contaminated surface (vials, glassware, etc.) may degrade 

your measurement system's performance (SA Margolis and E Park, "Stability of Ascorbic Acid in Solutions in 

Autosampler Vials", Clinical Chemistry 2001, 47(8), 1463-1464). You should suspect such degradation if you 

observe unusually large variation in replicate analyses. 

If you have any questions or concerns about the Vitamin C Micronutrients Measurement Quality Assurance 

Program please contact Jeanice Brown Thomas at tel: 301-975-3120, fax: 301-977-0685, or 

e-mail: jbthomas@nist.gov. 

We ask tlfat you return your results for these RR30 samples by March 31, 2009. We would appreciate receiving 

your results as soon as they become available. Please use the attached form. Your results will be kept confi.dential. 

lytical Chemistry Division 

Chemical Science and Technology Laboratory 

Enclosures: Protocols, Preparation and Analysis of Control Materials and Analysis of Test Samples 

RR30 Report Form for Ascorbic Acid Solid Control Material Preparation 

RR30 Report Form for Control Material and Test Sample Analyses 

N•Si 

E2
 

 

 

 

 

Micronutrient Measurement Quality Assurance Program for
 

Vitamin C
 

Please Read Through Completely BEFORE Analyzing Samples 

Protocol for Preparation and Analysis of the Ascorbic Acid Solid Control Material 

The ascorbic acid solid control material (in the amber vial) should be prepared and used in the 

following manner: 

1)	 Prepare at least 500 mL of 5% mass fraction metaphosphoric acid (MPA) in distilled water. This 

solution will be referred to as the “Diluent” below. 

2)	 Weigh 0.20 to 0.22 g of the ascorbic acid solid control material to 0.0001 g (if possible), dissolve 

it in the Diluent in a 100 mL volumetric flask, and dilute with the Diluent to the 100 mL mark. 

Weigh the amount of Diluent added to 0.1 g. Record the weights. The resulting material will be 

referred to as the “Stock Solution” below. 

3)	 Prepare three dilute solutions of the Stock Solution as follows: 

Dilute Solution 1: Weigh 0.500 mL of the Stock Solution to 0.0001 g into a 100 mL volumetric 

flask; dilute with Diluent to the 100 mL mark. Record the weight. 

Dilute Solution 2: Weigh 0.250 mL of the Stock Solution to 0.0001 g into a 100 mL volumetric 

flask; dilute with Diluent to the 100 mL mark. Record the weight. 

Dilute Solution 3: Weigh 0.125 mL of the Stock Solution to 0.0001 g into a 100 mL volumetric 

flask; dilute with Diluent to the 100 mL mark. Record the weight. 

4)	 Calculate and record the total ascorbic acid concentrations, [TAA], in these Dilute Solutions. 

If you follow the above gravimetric preparation directions, the [TAA] in μmol/L is calculated: 

(g Stock Solution in Dilute Solution 	 ) (⋅ g AA in Stock Solution )⋅(56785 mol/g μ ⋅ L) [TAA] DS = (g AA in Stock Solution ) + (g Diluent in Stock Solution)

For example, if you prepared the Stock Solution with 0.2000 g of solid ascorbic acid and 103.0 g 

of Diluent, then 0.5 mL of the Stock Solution should weigh (0.2+103)/200 = 0.52 g and 

[TAA]DS1 = (0.52 g)(0.2 g)∙(56785 μmol/g·L)/(0.2 + 103 g) = 57.2 μmol/L. Likewise, 0.25 mL 

of the Stock Solution should weigh 0.26 g and [TAA]DS2 = 29.4 μmol/L and 0.125 mL should 

weigh 0.13 g and [TAA]DS3 = 14.2 μmol/L. 

5)	 Measure the ultraviolet absorbance spectrum of Dilute Solution 1 against the Diluent as the blank 

using paired 1 cm path length cuvettes. Record the absorbance at 242, 243, 244, and 245 nm. 

Record the maximum absorbance (Amax) within this region. Record the wavelength (λmax) at 

which this maximum occurs. 

The extinction coefficient (E1%) of ascorbic acid at λmax (using a cell with a 1 cm path length) of 

Dilute Solution #1 can be calculated: 

E3
 

 

 

 

 

1% dL (A max )⋅	 ((g AA in Stock Solution ) + (g Diluent in Stock Solution )) E ( ) = g ⋅ cm (g Stock Solution in Dilute Solution 1)⋅(g AA in Stock Solution)

If your spectrophotometer is properly calibrated, λmax should be between 243 and 244 nm and 

E1% should be 550 ± 30 dL/g·cm. If they are not, you should recalibrate the wavelength and/or 

absorbance axes of your spectrophotometer and repeat the measurements. 

6)	 Measure and record the concentration of total ascorbic acid in all three dilute solutions and in the 

5% MPA Diluent in duplicate using exactly the same method that you will use for the serum 

control materials and test samples, including any enzymatic treatment. We recommend that you 

analyze these solutions in the following order: Diluent, Dilute Solution 1, Dilute Solution 2, 

Dilute Solution 3, Dilute Solution 3, Dilute Solution 2, Dilute Solution 1, Diluent. 

a)	 Compare the values of the duplicate measurements. Are you satisfied that your measurement 

precision is adequate? 

b)	 Compare the measured with the calculated [TAA] values. This is most conveniently done by 

plotting the measured values on the y-axis of a scatterplot against the calculated values on the 

x-axis. The line through the four {calculated, measured} data pairs should go through the 

origin with a slope of 1.0. Are you satisfied with the agreement between the measured and 

calculated values? 

Do not analyze the serum control materials or test samples until you are satisfied that your 

system is performing properly! 

7)	 Once you have confirmed that your system is properly calibrated, analyze the serum control 

CS #1 (see protocol below). The target values for this materials is 8.4 ±0.7 μmol/L of sample. 

If your measured values are not close to this value, please review your sample preparation 

procedure and whether you followed exactly the same measurement protocol the solutions 

prepared from the solid control material as you used for these serum controls. If the protocols 

differ, please repeat from Step 6 using the proper protocol. If the proper protocol was used, your 

measurement system may not be suitable for MPA-preserved samples; please contact us at 301­

975-3120 or jbthomas@NIST.gov. 

Do not analyze the test samples until you are satisfied that your system is performing properly 

and is suitable for the analysis of MPA-preserved serum! 

Protocol for Analysis of the Serum Control Materials and Test Samples 

The serum control material and test samples are in sealed ampoules. They were prepared by adding 

equal volumes of 10% MPA to spiked human serum. We have checked the samples for stability and 

homogeneity. Only the total ascorbic acid is stable. While these samples contain some 

dehydroascorbic acid, its content is variable. Therefore, only total ascorbic acid should be reported. 

The serum control material and test samples should be defrosted by warming at 20 °C for not more 

than 10 min otherwise some irreversible degradation may occur. 

Each serum test sample contains between 0.0 and 80.0 μmol of total ascorbic acid/L of solution. The 

total ascorbic acid in each ampoule should be measured in duplicate. Please report your results in 

μmol/(L of the sample solution) rather than μmol/(L of serum NIST used to prepare the sample). 

E4
 

 

Participant #: Date: 

Vitamin C Round Robin 30 

NIST Micronutrient Measurement Quality Assurance Program 

Preparation and Validation of Ascorbic Acid Solid Control Material 

STOCK SOLUTION 

Mass of ascorbic acid in the Stock Solution ..................................................__________ g 

Mass of 5% MPA Diluent added to the 100 mL volumetric flask .................__________ g 

DILUTE SOLUTION 1 

Mass of added stock solution (0.5 mL) ..........................................................__________ g 

Mass of 5% MPA Diluent added to the 100 mL volumetric flask .................__________ g 

Absorbance of Dilute Solution 1 at 242 nm...................................................__________ AU
 

Absorbance of Dilute Solution 1 at 243 nm...................................................__________ AU
 

Absorbance of Dilute Solution 1 at 244 nm...................................................__________ AU
 

Absorbance of Dilute Solution 1 at 245 nm...................................................__________ AU
 

Absorbance of Dilute Solution absorbance maximum ..................................__________ AU
 

Wavelength of maximum absorbance............................................................__________ nm
 

Calculated E1% ...............................................................................................__________ dL/g·cm
 

Calculated [TAA]DS1......................................................................................__________ μmol/L
	

DILUTE SOLUTION 2 

Mass of added stock solution (0.25 mL) ........................................................__________ g 

Mass of 5% MPA Diluent added to the 100 mL volumetric flask .................__________ g 

Calculated [TAA]DS2......................................................................................__________ μmol/L
	

DILUTE SOLUTION 3 

Mass of added stock solution (0.125 mL) ......................................................__________ g 

Mass of 5% MPA Diluent added to the 100 mL volumetric flask .................__________ g 

Calculated [TAA]DS3......................................................................................__________ μmol/L
	

Please return by March 31, 2009 

MMQAP Fax: 301-977-0685 

100 Bureau Drive, Stop 8392 Email: david.duewer@nist.gov 

Gaithersburg, MD 20899-8392 

E5
 

 

 

 

 

 

 

 

 

 

 

 

Participant #: Date: 

Vitamin C Round Robin 30 

NIST Micronutrient Measurement Quality Assurance Program
 

Analysis of Control Materials and Test Samples
 

Sample Replicate 1 Replicate 2 Units
 

Dilute Solution 1 μmol/L of Dilute Solution 

Dilute Solution 2 μmol/L of Dilute Solution 

Dilute Solution 3 μmol/L of Dilute Solution 

5% MPA Diluent μmol/L of Diluent 

μmol/L of Sample CS #1 

Target: 8.4 ±0.7 μmol/L 

Serum Test Sample #27 μmol/L of Sample 

Serum Test Sample #57 μmol/L of Sample 

Serum Test Sample #117 μmol/L of Sample 

Serum Test Sample #119 μmol/L of Sample 

SRM 1950 μmol/L of Plasma 

Preparation details you 

consider relevant to our 

interpretation of these 

results: 

Were samples frozen upon receipt? Yes | No 

Analysis method: HPLC-EC | HPLC-Fluor DAB | HPLC-OPD | HPLC-UV | AO-OPD | Other 

If “Other”, please describe: 

COMMENTS: 

Please return by March 31, 2009 

MMQAP Fax: 301-977-0685 

100 Bureau Drive, Stop 8392 Email: david.duewer@nist.gov 

Gaithersburg, MD 20899-8392 

E6
 

 

 

 

Participant #: ________ Date: _____________ 

Vitamin C Round Robin 30
 

NIST Micronutrients Measurement Quality Assurance Program
 

Packing List and Shipment Receipt Confirmation Form
 

This box contains one vial each of the following seven VitC M2QAP samples:
 

Label Form
 

VitC #27 Liquid frozen (1:1 serum:10% MPA) 

VitC #57 Liquid frozen (1:1 serum:10% MPA) 

VitC #117 Liquid frozen (1:1 serum:10% MPA) 

VitC #119 Liquid frozen (1:1 serum:10% MPA) 

CS #1 Liquid frozen (1:1 serum:10% MPA) 

Control Solid AA 

SRM 1950 Liquid frozen (plasma) 

Please 1) Open the pack immediately 

2) Check that it contains one vial each of the above samples 

3) Check if the samples arrived frozen 

4) Store the samples at -20  ̊C or below until analysis 

5) Complete the following information 

6) Fax the completed form to us at 301-977-0685 

(or email requested information to david.duewer@nist.gov) 

1) Date this shipment arrived: ___________________________ 

2) Are all of the vials intact? Yes | No 

If "No", which one(s) were damaged? 

3) Was there any dry-ice left in cooler? Yes | No 

4) Did the samples arrive frozen? Yes | No 

5) At what temperature are you storing the samples? _______  ̊C 

6) When do you anticipate analyzing these samples? ________________________ 

Your prompt return of this information is appreciated. 

The M2QAP Gang 

Mail: M2QAP 

NIST, Stop 8392 Fax: 301-977-0685 

Gaithersburg, MD 20899-8392 Email: David.Duewer@NIST.gov 

E7
 

 

Appendix F. Final Report for RR30 

The following three pages are the final report as provided to all participants: 

•	 Cover letter. 

•	 An information sheet that: 

o	 describes the contents of the “All-Lab” report, 

o	 describes the content of the “Individualized” report, 

o	 describes the nature of the test samples and details their previous distributions, 

if any, and 

o	 summarizes aspects of the study that we believe may be of interest to the 

participants. 

F1
F2
The NIST M2

QAP summary report for vitamin C intercomparison study RR30 consists of: 

Page “Individualized” Report 

1 Summarizes your reported values for the nominal 55 mmol/L solution you prepared from the 

ascorbic acid solid control sample, the serum control sample, and the four serum test 

samples. 

2 Graphical summary of your RR30 sample measurements. 

Page “All Lab” Report 

1 A tabulation of results and summary statistics for Total Ascorbic Acid [TAA] in the RR30 

samples and control/calibration solutions. 

Serum-based Samples. One serum control, four test unknowns, and an optional unknown (SRM 1950) 

were distributed in RR30. 

CS#1 SRM 970 level 1, ampouled in mid-1998. 

S30:1 VitC #27, ampouled in late 2001, previously distributed as sample S17:1 (RR17, Fall 02), 

S19:2 (RR19, Fall 03), S21:2 (RR21, Fall 04), S22:1 (RR24, Spring 05), S24:1 (RR24, 

Spring 06), and S28:1 (Spring 08). 

S30:2 VitC #57. ampouled in late 2001, previously distributed as sample S16:3 (RR16, Spring 02), 

S17:3 (RR17, Fall 02), S20:2 (RR20, Spring 04), S21:4 (RR21, Fall 04), S23:3 (RR23, Fall 

05), S27:2 (RR27, Fall 07). 

S30:3 VitC #117, ampouled in 1995, previously distributed sample 682a=S08:3 (RR8, 1996), 

682a=S10:2 (RR10, 1997), S25:4 (RR25, Fall 06), and S28:3 (Spring 08). (Unfortunately, 

two participants received miss-labeled units of a different serum. We apologize for the error.) 

S30:4 VitC #119, ampouled in 1995, previously distributed sample 688b=S07:2 (RR7, Spring 

1995), 688b=S08:2 (RR8, 1997), S24:4 (RR24, Spring 06), and S28:4 (Spring 08) 

SRM 1950 Metabolites in Human Serum: A human plasma pool that was prepared in the Spring 07 

from healthy male and female donors. This sample was designed for the evaluation of the 

measurement of metabolites in humans to provide insight into various disease states. 

Results. 

1)	 All participants who prepared the four 5% MPA control/calibration solutions (the three “Dilute 

Solutions” and the “Diluent”) did so correctly. The criteria used to evaluate this success are: the 

density of the 5% MPA (≈1.03 gm/mL), the observed wavelength maximum of “Dilute Solution 

#1”(≈244 nm), the observed absorbance at that maximum (≈0.58 OD), the calculated E1% #1”(≈560 

dL/g·cm). 

2)	 The Measured = a+b*Gravimetric calibration parameters for the control/calibration solutions 

(columns 10 to 13 of the All Lab Report) indicate that the measurement systems for most participants 

are linear (R2

 close to 1 and RMS close to 0.0) and reasonably well calibrated (intercepts close to zero 

and slopes close to 1). However, one participant’s RMS is larger than expected and “re-calibration” 

to the gravimetric values degrades their results for the unknowns. This suggests that their 

measurement process may be non-linear and that it is sensitive to the sample matrix (5 % MPA vs. a 

1+1 blend of serum and 10 % MPA). 

3)	 One participant reported that they were unable to obtain a reasonable value for the serum control 

while results for the gravimetrically prepared solutions were as expected. On evaluation, they found 

F3
that an enzymatic reagent was insufficiently active, giving rise to plausible but incorrect results for 

the test unknown. This emphasizes the importance of validating your methods with a wellcharacterized control material having composition similar to that of your samples. 

4)	 There is no evidence of sample degradation in any of the materials. 

5)	 We appreciate your efforts to help us value-assign ascorbic acid in SRM 1950. With one exception, 

all of the reported results were at or below the detection limits for your methods. Since SRM 1950 

plasma is not stabilized to oxidation/reduction, this was the expected result. 

F4
Appendix G. “All-Lab Report” for RR30 

The following single page is the “All-Lab Report” as provided to all participants, with 

two exceptions: 

• the participant identifiers (Lab) have been altered. 

• the order in which the participant results are listed has been altered. 

The data summary in the “All-Lab Report” has been altered to ensure confidentiality of 

identification codes assigned to laboratories. 

G1
Micronutrients Measurement Quality Assurance Program for Total Ascorbic Acid 

"Round Robin" 30 - March 2009 

Control / Calibration Samples MPA Dilute Solution 1 Samples 

G2

Density 

Lab Date Dil:1 Dil:2 Dil:3 Dil:1 Dil:2 Dil:3 MPA Inter Slope R2 RMS g/mL λmax Amax E1% CS#1 S30:1 S30:2 S30:3 S30:4 Inter Slope R2 RM

VC-MA 03/03/09 58.1 29.6 14.5 60.0 31.3 15.1 0.0 0.19 1.03 1.000 0.4 1.037 242. 0.5690 555.6 8.1 9.4 50.8 44.6 28.6 -0.16 1.07 1.000 0

VC-MB 17/12/08 59.5 29.8 14.7 58.3 27.9 13.4 0.0 -0.68 0.98 0.999 0.8 1.031 243. 0.5880 561.2 9.0 8.8 48.0 43.9 28.7 0.16 1.02 0.998 1

VC-MC 09/03/09 58.2 29.2 14.7 61.2 30.3 14.2 0.0 -0.57 1.06 1.000 0.7 1.024 243. 0.5496 536.3 8.4 8.5 45.8 42.4 26.9 0.07 0.98 0.998 0

VC-ME 10/02/09 56.6 28.4 14.3 57.0 28.2 13.8 0.0 -0.33 1.01 1.000 0.4 1.031 243. 0.5804 582.4 11.0 10.8 59.5 54.1 34.9 -0.06 1.27 0.999 0

VC-MG 17/03/09 60.9 31.4 16.8 65.1 32.8 16.2 0.0 -0.89 1.08 0.999 1.0 1.029 243.6 0.5850 545.8 7.9 7.6 50.4 47.4 28.4 -2.33 1.14 0.997 1

VC-MH 20/03/09 62.8 31.5 15.4 63.9 32.2 15.7 0.0 0.05 1.02 1.000 0.1 1.030 244. 0.6117 552.7 8.4 9.2 42.0 41.6 26.7 1.92 0.89 0.986 2

VC-MI 10/02/09 57.0 28.3 14.2 60.1 28.8 14.4 0.0 -0.43 1.06 1.000 0.6 1.030 8.3 6.3 47.8 b 26.9 -2.94 1.08 0.999 1

VC-MJ 27/03/09 55.7 27.8 13.4 60.5 29.7 14.9 0.4 0.31 1.08 1.000 0.5 1.018 253a 0.473a 482.5a na 13.4 49.9 42.5 27.0 3.83 0.93 0.989 2

VC-MK 26/03/09 62.6 30.7 15.2 63.1 34.6 16.8 8.5 6.41 0.90 0.991 2.8 1.030 244. 0.5810 526.8 8.4 10.8 49.8 41.1 23.4 -0.07 1.00 0.975 3

VC-MN 24/09/09 63.0 31.6 15.8 62.5 30.8 14.3 0.0 -0.64 1.00 0.999 0.7 1.029 243.7 0.6178 556.7 na 10.7 44.9 b 25.3 2.25 0.89 0.998 1

VC-MP 30/03/09 8.5 8.7 46.8 42.4 26.9 -0.05 0.99 1.000 0

VC-MU 06/03/09 6.3 7.4 42.0 b 23.9 -0.56 0.90 1.000 0

VC-NE 11/02/09 9.0 9.2 47.2 41.4 27.0 

VC-NF 17/03/09 60.5 29.5 14.9 61.2 30.7 14.9 0.2 0.16 1.01 1.000 0.6 1.031 243. 0.5930 556.9 8.9 8.7 44.5 41.0 26.8 0.80 0.94 0.998 0

11 11 11 11 11 11 11 11 9 9 9 12 14 14 11 14 

59.5 29.8 14.9 61.2 30.6 14.9 0.8 1.029 243.3 0.5862 552.7 8.5 9.3 47.8 43.8 27.2 

2.6 1.3 0.9 2.4 2.0 1.0 2.6 0.005 0.6 0.0206 15.7 1.1 1.8 4.4 3.9 2.7 

55.7 27.83 13.4 57.0 27.87 13.4 0.0 1.018 242.0 0.5496 526.8 6.3 6.3 42.0 41.0 23.4 

57.6 28.78 14.4 60.0 29.28 14.3 0.0 1.029 243.0 0.5804 545.8 8.3 8.6 45.1 41.5 26.7 

59.5 29.63 14.7 61.2 30.72 14.9 0.0 1.030 243.0 0.5850 555.6 8.4 9.0 47.5 42.4 26.9 

61.7 31.05 15.3 62.8 31.70 15.4 0.1 1.031 243.7 0.5930 556.9 8.9 10.4 49.8 44.3 28.0 

63.0 31.58 16.8 65.1 34.56 16.8 8.5 1.037 244.0 0.6178 582.4 11.0 13.4 59.5 54.1 34.9 

3.7 1.8 0.7 1.9 2.1 1.0 0.0 0.0 0.9 0.0119 8.3 0.5 1.3 3.7 2.0 1.2 

6 6 5 3 7 6 0.15 0.37 2.0 1.5 6 15 8 5 5 

MADe eSD 

CV CV 

Median Median 

%75 %75 

Max Max 

Min Min 

%25 %25 

N N 

Average Average 

SD SD 

Grav, μmol/L Measured, μmol/L Measured = a + b*Grav Spectrophotometry Measured, μmol/L Measured = p+q*Media

a ) 5% Trichloroacetic acid solution 

b ) Mislabeled sample 

All Lab Report 
Appendix H. Representative “Individualized Report” for RR30 

Each participant in RR30 received an “Individualized Report” reflecting their reported 

results. The following two pages are the “Individualized Report” for participant 

“VC-MA”. 

H1
 Set 1 of 17 

Vitamin C "Round Robin" 30 Report: Participant VC-MA
 

MPA Dilute Solution 1 Control/Calibration Solutions 

Density Spectrophotometry Ymeas = Inter + Slope* Xgrav 

Date RR Method 

08/28/06 25 HPLC-EC 

03/20/07 26 HPLC-EC 

10/05/07 27 HPLC-EC 

03/04/08 28 HPLC-EC 

08/11/08 29 HPLC-EC 

03/03/09 30 HPLC-EC 

g/mL 

1.039 

1.033 

1.032 

1.035 

1.037 

1.037 

λmax Amax E1% 

242.0 0.555 557.4 

244.0 0.573 554.3 

242.0 0.561 557.2 

243.0 0.572 562.2 

243.0 0.567 553.2 

242.0 0.569 555.6 

Inter Slope R2 SEE 

0.8 0.95 0.999 0.92 

0.3 1.00 1.000 0.31 

-0.1 0.99 1.000 0.14 

0.7 1.03 0.999 0.99 

0.3 1.03 1.000 0.64 

0.2 1.03 1.000 0.40 

Mean 1.036 242.7 0.57 556.6 Pooled SEE 0.65 

SD 0.002 0.8 0.01 3.2 

CV 0.24 0.34 1.2 0.6 

[TAA] mmol/Lsample 

7 9.5 0.2 0.5 

7 48.9 1.4 1.5 

3 42.5 0.5 2.3 

Date RR Sample 

08/28/06 25 CS#1 

03/20/07 26 CS#1 

03/04/08 28 CS#1 

03/03/09 30 CS#1 

12/12/02 17 S17:1 

11/13/03 19 S19:2 

09/13/04 21 S21:2 

03/08/05 22 S22:1 

03/09/06 24 S24:1 

03/04/08 28 S28:1 

03/03/09 30 S30:1 

11/18/02 16 S16:3 

12/12/02 17 S17:3 

02/23/04 20 S20:2 

09/13/04 21 S21:4 

10/17/05 23 S23:3 

10/05/07 27 S27:2 

03/03/09 30 S30:2 

ND 08 

08/20/97 10 S10:2 

08/28/06 25 S25:4 

03/04/08 28 S28:3 

03/03/09 30 S30:3 

05/23/95 07 S07:2 

Rep1 Rep2 Fadj Mean SDdup 

8.3 8.6 1.0 8.4 0.2 

8.6 8.3 1.0 8.5 0.2 

9.1 9.0 1.0 9.0 0.1 

8.3 8.0 1.0 8.1 0.2 

N Mean SDrepeat SDreprod 

4 8.5 0.2 0.4 

9.9 9.1 1.0 9.5 0.6 

9.2 9.1 1.0 9.2 0.1 

8.8 8.7 1.0 8.7 0.1 

9.6 9.6 1.0 9.6 0.0 

9.8 9.6 1.0 9.7 0.2 

10.4 10.3 1.0 10.4 0.1 

9.5 9.4 1.0 9.4 0.1 

49.9 44.9 1.0 47.4 3.5 

49.7 49.1 1.0 49.4 0.4 

50.6 50.0 1.0 50.3 0.4 

47.1 47.0 1.0 47.0 0.0 

49.8 48.8 1.0 49.3 0.7 

48.6 47.6 1.0 48.1 0.8 

51.2 50.4 1.0 50.8 0.6 

81.5 80.6 0.5 40.5 0.3 

42.0 42.2 1.0 42.1 0.2 

45.6 44.4 1.0 45.0 0.9 

44.8 44.5 1.0 44.6 0.2 

51.1 49.8 0.5 25.2 0.5 

29.0 29.5 1.0 29.2 0.4 

29.5 28.7 1.0 29.1 0.5 

28.5 28.6 1.0 28.6 0.0 

3 27.8 0.5 2.3 

ND 08 

03/09/06 24 S24:4 

03/04/08 28 S28:4 

03/03/09 30 S30:4 

Please check our records against your records. Send corrections and/or updates to... 

Micronutrients Measurement Quality Assurance Program 

National Institute of Standards and Technology 

100 Bureau Drive Stop 8392 Fax: (301) 977-0685 

Gaithersburg, MD 20899-8392 USA Email: david.duewer@nist.gov 

Individualized Report 1 / 2 

H2
 

NIST, RR 22

H3

 Set 1 of 17 

Vitamin C "Round Robin" 30 Report: Participant VC-MA 

Total Ascorbic Acid, μmol/mL 

[Total Ascorbic Acid], μmol/L 

S17:1S19:2 S21:2S22:1 S24:1 S28:1 S30:1 S16:3 

Apparent Precision, SD S17:3 S20:2 

S21:4S23:3 S27:2S30:2S08:3S10:2S25:4S28:3S30:3S07:2 S08:1S24:4S28:4 S30:4 

Your [Total Ascorbic Acid], μmol/L 

Median [Total Ascorbic Acid], μmol/L 

Concordance, SD 

3rd Quartile (75%) You, this RR Median (50%) You, pat RRs Others, this RR 1st Quartile (25%) 

For details of the construction and interpretation of these plots, see: 

Duewer, Kline, Sharpless, Brown Thomas, Gary, Sowell. Anal Chem 1999;71(9):1870-8. 

Sample Comments 

S30:1 VitC #27 previously distributed in RRs 17, 19, 21, 22, 24, and 28 

S30:2 VitC #57, previously distributed in RRs 16, 17, 20, 21, 23, and 27 

S30:3 VitC #117, previously distributed in RRs 8, 10, 25, and 28 

S30:4 VitC #119, previously distributed in RR 7, 8, 24, and 28 

Individualized Report 2 / 2