CERTIFICATE OF ANALYSIS

Chain of Custody: 308004 Job Name: Task 3 ‐ Analysis of Official Samples Date Su

Client: US Food & Drug Adminitration Job Location: 3rd Group ‐ 2 Samples Date A

Address: Office of Cosmetics & Colors Job Number: CLIN 1 ‐ Task 3 (2 Samples) Rep

4300 River Road PO Number: HHSF223201810337P Date 

College Park, MD 20740 Person Su

Attention: John Gasper 

TEM LOD  TEM LOQ % Tremolite by TEM % Chrysotile by TEM % Total Tremolite &  Chrysotile by TEM

Using ASTM D5756 Mass 

Calculation

Using ASTM D5756 Mass 

Calculation

Using ASTM D5756 Mass 

Calculation

Using ASTM D5756 Mass 

Calculation

Using ASTM D5756 Mass 

Calculation

308004‐1 D‐49 0.00000115% 0.00000946% 0.13214% < 0.00001% 0.13214% ND 12.2% 16.5% 

308004‐1A D‐49 0.00000133% 0.00000532% 0.00018% 0.00002% 0.00020% ND 12.4% 14.7% 

308004‐1B D‐49 0.00000153% 0.00000612% 0.20597% 0.00193% 0.20790% ND 12.5% 14.3% 

LOD = Limit of Detection LOQ = Limit of Quantification ND = Not Detected PLM = Polarized Light Microscopy TEM = Trans

Analytical Method(s):  PLM by Modified NY ELAP 198.6

TEM by Modified NY ELAP 198.4/ASTM D5756

Analyst(s): PLM

TEM

Technical Director: Andreas Sa

All results are to be consideredsigned by the Technical Directo

SUMMARY OF ANALYSIS

% Acid  Soluable 

%  Organics AMA  Sample ID Client  Sample ID %  Asbestos  by PLM

This report applies only to the sample, or samples, investigated and is not necessarily indicative of the quality or condition of apparently identical or similar products.  As a mutual protection to clients, the public, and these Laboratorieexclusive use of the client to whom it is addressed and upon the condition that it is not to be used, in whole or in part, in any advertising or publicity matter nor shall it be reproduced, except in full, without prior written authorization fprotocols are based upon the information provided by the persons submitting them and, unless collected by personnel of these Laboratories, we expressly disclaim any knowledge and liability for the accuracy and completeness of thisdiscarded in accordance with the appropriate regulatory guidelines, unless otherwise requested by the client.  NVLAP accreditation applies only to polarized light microscopy of bulk samples and transmission electron microscopy of AH

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(b) (6)
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Sample Preparation 

Samples were prepared for PLM and TEM bulk analysis by Chon Simpha on July 2, 2019 through July 9, 2019. Sample 

preparation consisted of the following steps:

1) Label and weigh two 8mL glass vials for each sample in the set – one vial for the PLM preparation and one vial 

for the TEM preparation.

2) Weigh out 0.1 to 0.8 grams of material and place in corresponding 8mL glass vial. Record weight.

3) Burn samples at 480° C for at least 12 hours.

4) Record Post-Ash Weight.

5) Treat ashed sample with concentrated hydrochloric acid.

6) Filter acid reduced material onto a pre-weighed 47mm 0.4um PolyCarbonate filter.

7) Place filter into drying oven for 30 minutes and then record Post-Acid Reduced weight. 
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8) Make four PLM slide preparations from the PLM residual ash for each sample in 1.550 dispersion oil. Make

additional preparations in 1.605, 1.625, 1.680 and 1.700 dispersion oil as necessary for particle identification.

9) Weigh a portion of the residue from the TEM residual ash and place it into the corresponding pre-weighed 

100ml jar.

10) Fill the 100ml jar with deionized water

11) Sonicate the jars for approximate 5-minutes.

12) Filter 0.2ml to 1ml of the solution onto a 47mm 0.22um MCE filter.

13) Dry the filter for 10 minutes then collapse, carbon coat, and place on a 3 TEM grids.

PLM Analysis

Analysis was performed in accordance with NY ELAP 198.6 protocols. The analysis was conducted using an Olympus BH2 polarized light microscope (PLM) equipped with a dispersion staining objective. All four slide preparations for each 

aliquot were examined. 400-point count was performed for those samples on which asbestos or a regulated amphibole 

was observed. If no asbestos was detected on any of the slides, the percentage of fibrous components was determined 

by visual estimation. The results of this analysis are detailed below in the Discussion and Interpretation of Analytical 

Findings section for each individual sample.

TEM Analysis

Analysis was performed in accordance with modified NY ELAP Method 198.4 protocols. The analysis was performed 

using a JEOL JEM-100CX II transmission electron microscope (TEM), equipped with a Thermo Fisher Quest Energy 

Dispersive X-Ray Analyzer (EDXA), at magnifications of 19,000x. Two grids for each aliquot were examined. Twenty (20)

grid openings were examined per sample.

Modifications to the NY ELAP 198.4 Method were:

1) The residue was not placed in alcohol and prepared using the quick drop method. To obtain a more uniform 

preparation, the residue was placed in a jar and filled with 100ml of deionized water. The jar was sonicated, and 

a portion of the solution was filtered onto a 47mm 0.22um MCE filter.

2) The tremolite and chrysotile were not visually estimated. The length and width of the observed particles were 

measured and the mass of each particle was calculated using the ASTM D5756 method. All particles identified as 

tremolite were included with the counts/concentrations, regardless of size and aspect ratio.

The results of this analysis are detailed below in the Discussion and Interpretation of Analytical Findings section for each 

individual sample.

Calculations 

ASTM D5756 Mass

M = π/4 L * W2 * D * 10-12

M = mass

L = length

W = width

D = density

Percent Calculation

EFA(mm2) * 100ml * MA(g) * RW(g)

 VF(ml) * IW(g) * AA(mm2) * RJ(g) 

The calculated value is then multiplied by 100 to convert it to percent.

EFA – Effective filter area

MA – Mass of asbestos

RW – Weight of residue

VF – Volume filtered
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IW – Initial weight of the sample

AA – Area analyzed

RJ – Weight of residue placed into the jar

Limit of Detection and Quantification

We used the mass of a 0.5 x 0.04-micron chrysotile fiber as the basis for our calculations. Limit of detection was defined 

as 1 fiber and limit of quantification was defined as 4 fibers.

Some aliquots of samples D-49 contained very small amounts of asbestos that were either at or below our 4-fiber limit 

of quantification. For these samples we defined our limit of quantification as follows:

308004-1: mass of the single observed chrysotile fiber plus the mass of three tremolite fibers measuring 

 0.5 x 0.04 microns

Discussion and Interpretation of Analytical Findings:

PLM

All three aliquots of sample D-49 were analyzed by Peerawut Chaikeenee on July 25, 2019. No asbestos or nonasbestos amphibole variants were detected the samples. The results were calculated using the equations detailed in the 

calculations section. 

308004-1 NAD

308004-1A NAD

308004-1B NAD

 

TEM

Michael Greenberg analyzed sample 1 on July 29, 2019, 1A on July 30, 2019 and 1B on August 1, 2019. The sample 

consisted of a mix of talc and mica particles, with a few talc fibers, mica fibers and titanium fibers/particles. Chrysotile 

and tremolite were observed on all three aliquots. The results were calculated using the equations detailed in the 

calculations section.

308004-1 0.13214%

308004-1A 0.00020%

308004-1B 0.20790%

The following charts plot aspect ratio vs. length, aspect ratio vs. width, and length vs. width for all the tremolite particles 

counted over all three aliquots.
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0.00

1.00

2.00

3.00

4.00

5.00

6.00

7.00

0 2 4 6 8 10 12

Aspect Ratio

Length microns

Tremolite Length vs Aspect Ratio

0.00

1.00

2.00

3.00

4.00

5.00

6.00

7.00

0 0.5 1 1.5 2 2.5 3 3.5 4 4.5

Aspect Ratio

Length microns

Tremolite Width vs Aspect Ratio
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Below are representative pictures, diffraction patterns, and chemistry from the observed particles. The unidentified 

peaks in chemistry spectra are copper, zinc, and carbon. Those peaks are from the TEM specimen holder and specimen 

grid.

Tremolite Particle from 308004-1

0

0.5

1

1.5

2

2.5

3

3.5

4

4.5

0 2 4 6 8 10 12

Width

Length microns

Tremolite Length vs Width
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Zone-Axis Diffraction Pattern from the Tremolite Particle pictured above

Diffraction Pattern from the Tremolite Particle pictured above
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Chemistry from the Tremolite Particle pictured above

Tremolite Particle from 308004-1B
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Diffraction Pattern from Tremolite Particle pictured above

Chemistry from Tremolite Particle pictured above
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Chrysotile Fiber from 308004-1A

Diffraction Pattern from Chrysotile Fiber pictured above
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Chemistry from Chrysotile Fiber pictured above

Talc Particle from 308004-1
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Diffraction Pattern from the Talc Particle pictured above

Chemistry from the Talc Particle pictured above
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Mica Particle from 308004-1

Diffraction Pattern from the Mica Particle pictured above
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Chemistry from the Mica Particle pictured above

Talc Fiber from 308004-1
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Diffraction Pattern from the Talc Fiber pictured above

Chemistry from the Talc Fiber pictured above
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Talc Ribbon from 308004-1

Diffraction Pattern from the Talc Ribbon pictured above
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Chemistry from the Talc Ribbon pictured above

Titanium Fiber from 308004-1
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Diffraction Pattern from the Titanium Fiber pictured above

Chemistry from the Titanium Fiber pictured above
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Particle coated with Titanium from 308004-1

Diffraction Pattern from the Titanium Coated particle pictured above
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Chemistry from the Titanium Coated Particle pictured above

Elongated Mica Particle from 308004-1
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Diffraction Pattern from the Elongated Mica Particle pictured above

Chemistry from the Elongated Mica Particle pictured above
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Tremolite Particle from 308004-1

Diffraction Pattern from the Tremolite Particle pictured above
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Diffraction Pattern from the Tremolite Particle pictured above

Chemistry from the Tremolite Particle pictured above
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Tremolite Particle from 308004-1

Diffraction Pattern from the Tremolite Particle pictured above
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Diffraction Pattern from the Tremolite Particle pictured above

Chemistry from the Tremolite Particle pictured above
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Tremolite Particle from 308004-1A

Diffraction Pattern from the Tremolite Particle pictured above
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Chemistry from the Tremolite Particle pictured above

Tremolite Particle from 308004-1B
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Diffraction Pattern from the Tremolite Particle pictured above

Diffraction Pattern from the Tremolite Particle pictured above
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Chemistry from the Tremolite Particle pictured above

Chrysotile Fiber from 308004-1
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Diffraction Pattern from the Chrysotile Fiber pictured above

Diffraction Pattern from the Chrysotile Fiber pictured above
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Chrysotile Structure from 308004-1B

Diffraction Pattern from the Chrysotile Structure pictured above
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Chrysotile Structure from 308004-1B

Diffraction Pattern from the Chrysotile Structure pictured above
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QC Discussion:

During preparation, one blank control sample and one reference control sample were prepared. These samples were 

prepared alongside the customer samples. The blank sample was prepared using Sigma-Aldrich Talc Powder, <10 

micron. No asbestos was detected on the blank sample. The reference sample was made from the same Sigma-Aldrich 

talc powder spiked with 1% Chrysotile. The reference sample was analyzed by and found to be within acceptable limits.

Our LIMS randomly selects samples for additional replicate and duplicate QC. 308004-1, 1A, and 1B/D-49 were not 

selected for any additional QC analysis.

Attachments:

The following items are attached to this case narrative for your reference:

1) Sample Log-In Sheet

2) Daily PLM Scope Calibration Log

3) Refractive Index Oil Calibration Log

4) Daily TEM Scope Calibration Log

5) QC Results Summary

6) Replicate and Duplicate QC Charts for Peerawut Chaikeenee for samples analyzed between 1/1/2019 & 

8/8/2019

7) Replicate and Duplicate QC Charts for Michael Greenberg for samples analyzed between 1/1/2019 & 8/8/2019

8) Raw Data Sheets

a. Gravimetric Data

b. Filtration Worksheets

c. PLM Analysis

d. TEM Analysis

e. QC Samples

I certify that all information contained in this report pertaining to laboratory events, procedures, and protocols is true 

and accurately describes the handling of this project by AMA Analytical Services, Inc. and its personnel.

8/14/2019

Andreas Saldivar Date

Laboratory Director