supplemental comments from basf corporation (keller and...

KH® KELLER AND HECKMAN LLP Serving Business through Law and Science®

1001 G Street, N.W. Suite 500 West Washington, D.C. 20001 tel. 202.434.4100 fax 202.434.4646

November 4, 2015

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Division ofDockets Management (HFA-305) Food and Drug Administration 5630 Fishers Lane, Room 1061 Rockville, Maryland 20852

Writer's Direct Access Devon Wm. Hill (202) 434-4279 hill @ khlaw . com

Re: Comments on Natural Resources Defense Council et al.; Filing of Food Additive Petition on Perch/orates; Docket No. FDA-2015-F-0537

Dear Sir or Madam:

On behalf of our client, BASF Corporation, we are writing to provide you with comments regarding the Natural Resources Defense Council (NRDC)1 October 15,2014 petition regarding perchlorates. These Comments are intended to supplement Comments submitted on June 29, 2015.

As the Agency is aware, the NRDC Petition requests, among other items, that the U.S. Food and Drug Administration (FDA) revoke its 2005 approval of Threshold of Regulation (TOR) No. 2005-006.~ The listing for Threshold of Regulation No. 2005-006 currently permits the use of sodium perchlorate monohydrate (CAS Reg. No. 7791-07-3) as a conductivity enhancer in the manufacture of antistatic agents for use in polymeric food packaging, at a level not to exceed 1.2 percent by weight of the finished polymer. The finished polymer may be used in contact with Food Type VIII ("Dry solids with the surface containing no free fat or oil").1

1 The Petition was co-sponsored by the Center for Food Safety (CFS), Breast Cancer Fund, Center for Environmental Health, Environmental Working Group, Improving Kids' Environment, Clean Water Action, Center for Science in the Public Interest and Children's Environmental Health Network.

See http: //www.accessdata.fda.gov/scripts/fdcc/?set=TOR&id=2005-006.

J. As the Agency is aware, the TOR request was originally submitted on behalf ofCiba Specialty Chemicals Corporation on June 17, 2005. BASF later acquired Ciba Specialty Chemicals Company, and the existing TOR, in 2008. The TOR was revised in FDA's online TOR exemptions database on August 17, 2015, to correct certain typographical errors with respect to the intended use limitations. FDA's Food

(continued ... )

Washington, D.C. Brussels San Francisco Shanghai

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KELLER AND HECKMAN LLP

Division of Dockets Management (HFA-305) November 4, 2015

BASF originally submitted Comments to this Docket on June 29,2015 indicating its' belief that the facts supporting the TOR exemption remain valid, and representative of foodcontact materials used in dry food applications. To confirm the validity of these facts, BASF also agreed to conduct analytical work to demonstrate that sodium perchlorate monohydrate does not migrate to dry food.

In a May 18,2015 Pre-Notification Consultation (PNC) with the Agency at its' College Park, Maryland office,1 BASF agreed to demonstrate that the migration of sodium perchlorate would not be detectable at an analytical sensitivity of 10 parts per billion (ppb). The purpose of this analytical work, therefore, was to demonstrate that the potential dietary exposure to sodium perchlorate is safe and below the threshold level where regulation is necessary.

BASF has completed this analysis, and submits a September 4, 2015 Migration Report, Migration of Sodium Perchlorate from Low Density Polyethylene containing 20% lrgastab P 18 into Dry Food Simulant ("Report") in support of the same. The analytical report demonstrates that sodium perchlorate would not, in fact, be detected at an analytical sensitivity of 10 ppb, when used as intended.

If you have any questions regarding these comments, or if we can be of further assistance in any other way, please do not hesitate to contact me.

Devon Wm. Hill

( ... continued) Types and Conditions of Use are available at: http://www.fda.gov/Food/IngredientsPackagingLabeling/ Packagin gFCS/F oodTypesCond itionsofU se/ defau lt.htm.

1 PNC 1642, "Meeting with the Society of the Plastics Industry (SPI) and BASF Corporation to discuss the allowed use of perchlorates in food contact applications."

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Migration Report Migration of Sodium Perchlorate from Low Density

Polyethylene containing 20% Irgastat P18 into Dry Food Simulant

Page 1 of 7 100 Park Avenue Florham Park, NJ 07932

Issued:

Author: _______________________ Date: _____________

Approved: ______________________ Date: ______________




Table of Contents I. Summary ........................................................................................................................................ 3

II. Objectives ....................................................................................................................................... 3

III. Polymer Plaques ............................................................................................................................ 4

IV. Experimental .................................................................................................................................. 4

A. Apparatus and Reagents ............................................................................................................ 4

B. Migration Experiments ................................................................................................................ 4

C. Analysis of Simulants ..................................................................................................................... 5

V. Results............................................................................................................................................ 5

VI. Validation ........................................................................................................................................ 6

VII. Appendices ..................................................................................................................................... 7

A. Analytical method (including calculations) ................................................................................. 7

B. Detailed sample analysis data .................................................................................................... 7

C. Chromatograms .......................................................................................................................... 7

D. Spiking validation at low perchlorate levels ................................................................................ 7




I. Summary

The migration behavior of sodium perchlorate from low density polyethylene (LDPE) loaded with 20%

Irgastab P18 was investigated with Tenax® as the dry food simulant. The polymer was exposed to Tenax®

at 40°C with intervals removed and analyzed at 2h, 1 day, 4 days and 10 days. Perchlorate (as Cl16O4) was

found to be less than 1.5 ng/in2 in the simulant, corresponding to 150 ppt in food.

Table 1 Summary of migration results

Samples Amount in Simulant (ng/in2) Amount in Food (ppt)

2 hrs < 1.5 < 150

1 day < 1.5 < 150

4 days < 1.5 < 150

10 days < 1.5 < 150

Note:

The results were calculated based on the surface area of both sides of the migration plaques (4 in2).

The calculation was based upon the assumption that 1 square inch of surface is in contact with 10 grams of food.

The limit of quantitation (LOQ) of perchlorate was 1.5 ng/in2

Data are reported as concentration of perchlorate ion Cl16O4

II. Objectives

Sodium perchlorate is used as conductivity enhancer in antistatic agents that are used in polymers.

The purpose of current study is to investigate the migration behavior of sodium perchlorate from the

polymer when in contact with dry foods. In this study, low density polyethylene was used as the model

polymer. Antistatic agent Irgastab P18 containing sodium perchlorate was incorporated into LDPE at

an approximate concentration of 20% (w/w). In this study, LC/MS/MS was used for sensitive analysis

of perchlorate ions in the Tenax® at low ppb levels.




III. Polymer Plaques

Polymer plaques were used as received. They are made of LDPE loaded with approximately 20%

Irgastab P18. Irgastab P18 contains 1.8 % sodium perchlorate.

IV. Experimental

A. Apparatus and Reagents Wheaton™ glass staining dish with screw cap (1.02×1.02×2.7 in) Cat No. 08-813D

Volumetric flasks, class A

Fisher Scientific Isotemp Oven, Model 6841

Balance: Mettler AT261

Acetonitrile, HPLC Grade

Ammonium acetate, 98%

Barnstead Nanopure Diamond ultrapure water system

BÜCHI Heating Bath B-490 and Rotavapor R-205 system

Agilent 1200 High Performance Liquid Chromatograph

Applied Biosystem API 4000 LC/MS/MS

Sodium perchlorate monohydrate, ≥ 98%

18O-labeled sodium perchlorate solution, atom purity 95%, Icon Isotopes

Tenax®, 60-80 mesh, Sigma

B. Migration Experiments

Tenax® was used as a simulant for dry food. The Tenax® was washed upon receiving by overnight soxhlet extraction with acetonitrile (ACN). After thoroughly drying in a vacuum oven, it was used for the migration study.

Wheaton™ glass staining dishes with screw caps were used as migration vessels. One piece of LDPE

plaque with surface area of 4 in2 (double sides) was set upright in the middle of each vessel and 12 g of

Tenax® was added into each vessel such that the plaques were fully covered. The simulant mass to surface

area ratio was 3. The vessels were capped and put in the pre-heated oven for exposure. Each sampling

time point was performed in triplicate. Blank polymer was treated in a similar manner.




The migration study was done at four time points; 2 hours, 1 day, 4 days and 10 days. The vessels were

placed into a preheated oven at 40°C. The temperature was maintained throughout the 10 days test period.

After the specified time intervals, the plaques were removed and all the Tenax® in each vessel was

transferred into a 250 mL beaker for further processing.

Conditions of Exposure Food Simulant

40°C for 2 hrs, 1 day, 4,days and 10 days

Tenax®

C. Analysis of Simulants

After the Tenax® was transferred into the 250 mL beaker, 0.1 mL of internal standard 18O-labeled

perchlorate solution at 1.5 ppm was added, followed by adding 150 mL of ACN. The solution was

stirred for one minute, then allowed to stand for 5 minutes. A 50 mL aliquot of the supernatant was

transferred into a rotary evaporator tube and concentrated to 10 mL. The solution was then filtered

through 0.2 µm PTFE syringe filter for LC/MS/MS analysis. See appendix A for details.

V. Results

At all four time points, perchlorate was detected, but at levels lower than method LOQ of 1.5 ng/in2.

No increase in perchlorate concentration was observed with prolonged exposure time. Additionally,

large RSD values for triplicate analysis, between 33 to 60% (Appendix-D) were obtained for all time

point samples, suggesting that the perchlorate found in the simulant was most likely caused by surface

abrasion.

The results shown below were calculated based on the surface area of both sides of the migration

plaques (4 in2) based on the assumption of 1 in2 of plaque being in contact with 10 grams of food.




Table 2 Results for specified conditions

*, data are reported as concentration of perchlorate ion Cl16O4

VI. Validation

Blank polymer plaques with 10 days exposure to Tenax® was used for the validation. Tenax® was transferred into a beaker followed by spiking with 6 ng of perchlorate (Cl16O4), corresponding to the LOQ level of 1.5 ng/in2 (150 ppt in food). Then 0.1 mL of internal standard 18O-labeled perchlorate (Cl18O4) at 1.5 ppm and 150 mL of ACN was added. The extracts were further processed as stated in section 0 for LC/MS/MS analysis.

Table 3 Results of Validation

Validation replicate

Perchlorate spiked* (ng/in2)

Amount Spike* (ng)

Perchlorate found* (ng/in2)

Amount Found* (ng)

Recovery (%)

1 1.53 6.12 1.57 6.26 102.3

2 1.53 6.12 1.51 6.04 98.6

3 1.53 6.12 1.49 5.97 97.5

AVG 99.4

STD 2.5

RSD (%) 2.5

*, data are reported as perchlorate ion Cl16O4

Samples Amount in Simulant* (ng/in2) Estimated amount in Food * (ppt)

2 hrs < 1.5 < 150

1 day < 1.5 < 150

4 days < 1.5 < 150

10 days < 1.5 < 150




VII. Appendices

A. Analytical method (including calculations)

B. Detailed sample analysis data

C. Chromatograms

D. Spiking validation at low perchlorate levels

Migration Report Migration of Sodium Perchlorate from Low Density Polyethylene containing 20% Irgastat P18 into Food

Simulant.

Appendix-A Analysis Method


Issued:

Author: _______________________ Date: ______________

Approved: ______________________ Date: ______________


Simulant.



Table of Contents I. Principle ................................................................................................................................... 3

II. Apparatus and Reagent .......................................................................................................... 3

III. LC/MS/MS Method .................................................................................................................. 3

IV. Standard preparation and calibration curve ............................................................................ 4

V. Sample processing .................................................................................................................. 6

VI. Calculations ............................................................................................................................. 6

VII. Chromatograms ....................................................................................................................... 8

VIII. Validation ............................................................................................................................... 10

A. Specificity ........................................................................................................................... 10

B. Range ................................................................................................................................. 10

C. Repeatability ...................................................................................................................... 10

D. Linearity .............................................................................................................................. 10

E. Detection limits ................................................................................................................... 10


Simulant.



I. Principle

This method is suitable for quantitative analysis of perchlorate in migration study from low density

polyethylene (LDPE) in to Tenax® as dry food simulant. Perchlorate ion (Cl16O4) was detected by

LC/MS/MS with multiple reaction monitoring (MRM) mode. 18O-labeled perchlorate (Cl18O4) was

used as internal standard for quantitation. Ion transition of m/z 99→83 for Cl16O4 to Cl16O3 and

m/z 107→89 for Cl18O4 to Cl18O3 were monitored for quantitation.

II. Apparatus and Reagent Volumetric flasks, class A

Fisher Scientific Isotemp Oven, Model 6841

Balance: Mettler AT261

Acetonitrile, HPLC Grade

Ammonium acetate, 98%

Barnstead Nanopure Diamond ultrapure water system

BÜCHI Heating Bath B-490 and Rotavapor R-205 system

Agilent 1200 High Performance Liquid Chromatograph

Applied Biosystem API 4000 LC/MS/MS

Sodium perchlorate monohydrate, ≥ 98%

18O-labeled sodium perchlorate solution, atom purity 95%, Icon Isotopes

Tenax®, 60-80 mesh, Sigma

III. LC/MS/MS Method

LC Mode Ion Exchange

Solvents ACN 50%, Water 50%, Ammonium acetate 100 mM

Column Waters IC-Pak Anion HR 4.6 mm i.d.×75 mm

Flow Rate 0.35 mL/min

Gradient Isocratic

Column Temperature

35 °C

Detection MS/MS, ESI(-), MRM mode, monitor ion transition of m/z 99→83 for Cl16O4 to Cl16O3 and monitor ion transition of m/z 107→89 for Cl18O4 to Cl18O3


Simulant.



IV. Standard preparation and calibration curve

Prepare two standard stock solutions A and B at a concentration of 1000 ppm in water using two independent weighings of 35 mg of sodium perchlorate standard into two 25 mL volumetric flasks (stock solutions A and B). Working standards were prepare at the concentration range of 0.2 to 25 ppb with the dilution scheme below. To each standard, internal standard 18O-labeled perchlorate at 7.4 ppb was added. Solution A/Solution B: 18O-labeled sodium perchlorate as internal standard was received as 1 mg

of 18O-labeled sodium perchlorate dissolved in 0.55 g H218O, which equals 1497 ppm of Cl18O4

ion in solution (IS stock solution). A 0.1mL aliquot of IS stock solution was transferred into 10 mL

volumetric flask and diluted to the mark with water to obtain 14.97 ppm (IS Solution A). 0.5 mL

of IS Solution A was transferred into 100 mL volumetric flask. Dilute to mark with water to obtain

75 ppb IS solution B.

Solution C: 5 ppm of perchlorate in ACN. Transfer 0.5 mL of stock solution A to a 100 mL

volumetric flask. Dilute to mark with ACN.

Solution D: 5 ppm of perchlorate in ACN. Transfer 0.5 mL of stock solution B to a 100 mL

volumetric flask. Dilute to mark with ACN.

Solution E: 25 ppb perchlorate in ACN with IS 18O-labeled perchlorate at 7.5 ppb. Transfer 0.5

mL of solution C and 10 mL of IS stock solution B to a 100 mL volumetric flask. Dilute to mark

with ACN.

Solution F: 50 ppb perchlorate in ACN with IS 18O-labeled perchlorate at 7.5 ppb. Transfer 1 mL

of solution D and 10 mL of IS stock solution B to a 100 mL volumetric flask. Dilute to mark with

ACN.

Solution G: 5 ppb perchlorate in ACN with IS 18O-labeled perchlorate at 7.5 ppb. Transfer 2 mL

of solution E and 1 mL of IS stock solution B to a 10 mL volumetric flask. Dilute to mark with

ACN.

Solution H: 10 ppb perchlorate in ACN with IS 18O-labeled perchlorate at 7.5 ppb. Transfer 2

mL of solution F and 1 mL of IS stock solution B to a 10 mL volumetric flask. Dilute to mark with

ACN.


Simulant.



Solution I: 1 ppb perchlorate in ACN with IS 18O-labeled perchlorate at 7.5 ppb. Transfer 2 mL

of solution G and 1 mL of IS stock solution B to a 10 mL volumetric flask. Dilute to mark with

ACN.

Solution J: 2 ppb perchlorate in ACN with IS 18O-labeled perchlorate at 7.5 ppb. Transfer 2 mL

of solution H and 1 mL of IS stock solution B to a 10 mL volumetric flask. Dilute to mark with

ACN.

Solution K: 0.2 ppb perchlorate in ACN with IS 18O-labeled perchlorate at 7.5 ppb. Transfer 2

mL of solution I and 1 mL of IS stock solution B to a 10 mL volumetric flask. Dilute to mark with

ACN.

Solution L: 0.4 ppb perchlorate in ACN with IS 18O-labeled perchlorate at 7.5 ppb. Transfer 2

mL of solution J and 1 mL of IS stock solution B to a 10 mL volumetric flask. Dilute to mark with

ACN.

By using LC/MS/MS method at MRM mode, ion transition of m/z 99→83 for Cl16O4 to Cl16O3 and

m/z 107→89 for Cl18O4 to Cl18O3 was recorded. A calibration curve was established by plotting

the ratio of peak area for Cl16O4 to peak area for Cl18O4 against the ratio of Cl16O4 concentration

to Cl18O4 concentration. A typical calibration curve is shown below.

Concentration of Cl16O4 (ppb)

Concentration of Cl18O4 (ppb)

Ratio of Cl16O4

concentration to

Cl18O4 concentration

Ratio of peak area for Cl16O4 to peak

area for Cl18O4

0.19 7.5 0.03 0.03

0.38 7.5 0.05 0.05

0.96 7.5 0.13 0.13

1.94 7.5 0.26 0.24

4.81 7.5 0.64 0.63

9.70 7.5 1.30 1.21

24.12 7.5 3.22 3.05


Simulant.



V. Sample processing

After each migration exposure interval, the food simulant Tenax® was transferred into a 250 mL

beaker and 0.1 mL of internal standard 18O-labeled perchlorate solution at 1.5 ppm was added,

followed by the addition of 150 mL of ACN. The solution was stirred for one minute and allowed

to stand for 5 minutes. A 50 mL aliquot of the supernatant was then transferred into rotary

evaporator tube for concentration to 10 mL. The solution was then filtered through 0.2 µm PTFE

syringe filter for LC/MS/MS analysis.

VI. Calculations

By using the LC/MS/MS method for sample analysis, the ratio of peak area for Cl16O4 to peak

area for internal standard Cl18O4 was recorded and the concentration of perchlorate in the food

simulant was calculated as following.

• y = mx + b

where x = ratio of Cl16O4 concentration to Cl18O4 concentration, m = slope of the line, y = ratio of

peak area for Cl16O4 to peak area for Cl18O4, and b = y-intercept

y = 0.9464x + 0.0043R² = 0.9999

0.00

1.00

2.00

3.00

4.00

0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50

Pe

ak

are

a o

f C

l16

O4

/Pe

ak

are

a o

f C

l18

O4

[Cl16O4]/[Cl18O4]

Calibration curve


Simulant.



• ppb Cl16O4 in solution = ratio of peak area for Cl16O4 to peak area for Cl18O4 X ppb IS

Cl18O4

• ng analyte/in2 =ppb Cl16O4 x 150 mL / surface area (4 in2)

• ppt analyte in food* = ng analyte/in2 x 100

The calculation is based upon the assumption that 1 square inch of surface is in contact with 10

grams of food.


Simulant.



VII. Chromatograms

Figure 1. Standard of Cl16O4 at 0.2 ppb (upper) with internal standard Cl18O4 at 7.5 ppb (lower)

XIC of -MRM (4 pairs): 99.0/83.0 amu from Sample 42 (std 0.2ppb) of MJ150034.wiff (Turbo Spray) Max. 416.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

50

100

150

200

250

300

350

400

In

te

ns

it

y,

c

ps

10.45

10.59

10.86

13.5011.64 16.875.42 7.78 17.621.37 14.217.474.65 9.836.06 15.163.26

XIC of -MRM (4 pairs): 107.0/89.0 amu from Sample 42 (std 0.2ppb) of MJ150034.wiff (Turbo Spray) Max. 1.3e4 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.0

2000.0

4000.0

6000.0

8000.0

1.0e4

1.2e4

1.3e4

In

te

ns

it

y,

c

ps

10.44


Simulant.



Figure 2. Sample analysis (2 h exposure) with Cl16O4 (upper) and internal standard Cl18O4 (lower)

XIC of -MRM (4 pairs): 99.0/83.0 amu from Sample 83 (2h sample-3) of MJ150034.wiff (Turbo Spray) Max. 216.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

120

140

160

180

200

216

In

te

ns

it

y,

c

ps

10.38

2.50

10.062.0611.170.19 6.43 16.366.165.46 8.391.20 7.74 15.81 18.5613.637.24 17.87


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

8000

9000

In

te

ns

it

y,

c

ps

10.41

11.50 14.20


Simulant.



VIII. Validation

The method was validated by its specificity, linear range, repeatability and detection limits.

A. Specificity

LC/MS/MS at MRM mode monitors ion transition of m/z 99→83 for Cl16O4 to Cl16O3 and monitor ion

transition of m/z 107→89 for Cl18O4 to Cl18O3 for quantitation. Ion transition of m/z 101→85 for 37Cl16O4 to 37Cl16O3 is used for confimation of perchlorate identity.

B. Range

0.2 to 25 ppb for perchlorate in solution for LC/MS/MS analysis.

C. Repeatability

Six individual injections of perchlorate at 0.2 ppb was done for assessing repeatability. RSD

obtained was 4.5%.

D. Linearity

Linear regression coefficient R2 is better than 0.999.

E. Detection limits

LC/MS/MS method limit of detection is 0.1 ppb and limit of quantitation is 0.2 ppb for perchlorate

in solution. For analysis of perchlorate migration into Tenax® as food simulant, limit of detection

is 0.4 ng/in2 and limit of quantitation is 1.5 ng/in2 (Appendix-D).


Simulant.

Appendix-B Detailed Sample Analysis Data


Issued:

Author: ___________________ Date: ______________

Approved: ______________________ Date: ______________


Simulant.



Table of Contents I. Detailed sample analysis data ....................................................................................................... 3


Simulant.



I. Detailed sample analysis data

Table 1 Detailed data analysis - Perchlorate levels in food simulant

Sample replicate

2 hrs 1 day 4 days 10 days Validation with blank polymer

at LOQ

Ratio* ng/in2** Ratio* ng/in2** Ratio* ng/in2** Ratio* ng/in2** Ratio* ng/in2**

1 0.017 0.51 0.027 0.96 0.024 0.82 0.013 0.31 0.040 1.6

2 0.011 0.30 0.016 0.54 0.020 0.66 0.021 0.63 0.039 1.5

3 0.019 0.59 0.015 0.53 0.007 0.18 0.013 0.31 0.038 1.5

AVG

0.47

0.68

0.56

0.42

1.52

STD 0.15 0.25 0.33 0.18 0.04

RSD (%) 33 36 60 44 3

*, ratio of peak area for Cl16O4 to peak area for Cl18O4

**, the numbers were estimated by extrapolating the calibration curve to lower than LOQ level. Numbers

are reported as perchlorate ion Cl16O4

With the method stated in Appendix-A, Analysis method, the ratio of peak area for Cl16O4 to peak

area for Cl18O4 was calculated from LC/MS/MS analysis chromatograms and with the calibration

curve listed in Appendix-A, Section IV Standard Preparation and calibration curve, the ratio of

Cl16O4 concentration to Cl18O4 concentration can be calculated. Knowing the concentration of

internal standard Cl18O4, the perchlorate concentration in the solution for LC/MS/MS injection can

thus be obtained. As can be seen from Table 1 Detailed data analysis the ratio of peak area for

Cl16O4 to peak area for Cl18O4 for all time point are smaller than method LOQ. Additionally,

although the RSD values are high for all four time points between 33 to 60%, the levels of

perchlorate found in the simulant are less than 1 ng/in2. This suggests that instead of migrating

from the plaque into Tenax®, perchlorate found in Tenax® is more likely due to physical contact

and scraping during sample handling.


Simulant.

Appendix-C Chromatograms


Issued:

Author: ___________________ Date: ______________

Approved: ______________________ Date: ______________


Simulant.



Table of Contents I. Figure 1 Standard of Cl16O4 at 0.2 ppb (upper) with internal standard Cl18O4 at 7.5 ppb (lower) .. 4

II. Figure 2 Standard of Cl16O4 at 0.4 ppb (upper) with internal standard Cl18O4 at 7.5 ppb (lower) .. 5

III. Figure 3 Standard of Cl16O4 at 1 ppb (upper) with internal standard Cl18O4 at 7.5 ppb (lower) ..... 6

IV. Figure 4 Standard of Cl16O4 at 2 ppb (upper) with internal standard Cl18O4 at 7.5 ppb (lower) ..... 7

V. Figure 5 Standard of Cl16O4 at 5 ppb (upper) with internal standard Cl18O4 at 7.5 ppb (lower) ..... 8

VI. Figure 6 Standard of Cl16O4 at 10 ppb (upper) with internal standard Cl18O4 at 7.5 ppb (lower) ... 9

VII. Figure 7 Standard of Cl16O4 at 25 ppb (upper) with internal standard Cl18O4 at 7.5 ppb (lower) . 10

VIII. Figure 8. 2 Hrs exposure blank-1 with Cl16O4 (upper) and internal standard Cl18O4 (lower) ...... 11

IX. Figure 9. 2 Hrs exposure blank-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower) ........ 12

X. Figure 10. 2 Hrs exposure blank-3 with Cl16O4 (upper) and internal standard Cl18O4 (lower) ...... 13

XI. Figure 11. 2 Hrs exposure sample-1 with Cl16O4 (upper) and internal standard Cl18O4 (lower) ... 14

XII. Figure 12. 2 Hrs exposure sample-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower) ... 15

XIII. Figure 13. 2 Hrs exposure sample-3 with Cl16O4 (upper) and internal standard Cl18O4 (lower) . 16

XIV. Figure 14. 1 Day exposure blank-1 with Cl16O4 (upper) and internal standard Cl18O4 (lower) ... 17

XV. Figure 15. 1 Day exposure blank-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower) ..... 18

XVI. Figure 16. 1 Day exposure blank-3 with Cl16O4 (upper) and internal standard Cl18O4 (lower) ... 19

XVII.Figure 17. 1 Day exposure sample-1 with Cl16O4 (upper) and internal standard Cl18O4 (lower) 20

XVIII.Figure 18. 1 Day exposure sample-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower)21

XIX. Figure 19. 1 Day exposure sample-3 with Cl16O4 (upper) and internal standard Cl18O4 (lower) 22

XX. Figure 20. 4 Days exposure blank-1 with Cl16O4 (upper) and internal standard Cl18O4 (lower) ... 23

XXI. Figure 21. 4 Days exposure blank-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower) .. 24

XXII.Figure 22. 4 Days exposure blank-3 with Cl16O4 (upper) and internal standard Cl18O4 (lower) . 25

XXIII.Figure 23. 4 Days exposure sample-1 with Cl16O4 (upper) and internal standard Cl18O4 (lower) .

.............................................................................................................................................. 26

XXIV.Figure 24. 4 Days exposure sample-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower) .

.............................................................................................................................................. 27


Simulant.



XXV.Figure 25. 4 Days exposure sample-3 with Cl16O4 (upper) and internal standard Cl18O4 (lower) ..

.............................................................................................................................................. 28

XXVI.Figure 26. 10 Days exposure blank-1 with Cl16O4 (upper) and internal standard Cl18O4 (lower) ..

.............................................................................................................................................. 29

XXVII.Figure 27. 10 Days exposure blank-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower) .

.............................................................................................................................................. 30

XXVIII.Figure 28. 10 Days exposure blank-3 with Cl16O4 (upper) and internal standard Cl18O4 (lower)

.............................................................................................................................................. 31

XXIX.Figure 29. 10 Days exposure sample-1 with Cl16O4 (upper) and internal standard Cl18O4

(lower) ............................................................................................................................................ 32

XXX.Figure 30. 10 Days exposure sample-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower)

.............................................................................................................................................. 33

XXXI.Figure 31. 10 Days exposure sample-3 with Cl16O4 (upper) and internal standard Cl18O4

(lower) ............................................................................................................................................ 34

XXXII.Figure 32. Validation with 10 days blank-1. Cl16O4 (upper) was spiked at the level of 150 ppt in

food and internal standard Cl18O4 (lower) ..................................................................................... 35

XXXIII.Figure 33. Validation with 10 days blank-2. Cl16O4 (upper) was spiked at the level of 150 ppt

in food and internal standard Cl18O4 (lower).................................................................................. 36

XXXIV.Figure 35. Validation with 10 days blank-3. Cl16O4 (upper) was spiked at the level of 150 ppt

in food and internal standard Cl18O4 (lower).................................................................................. 37


Simulant.



I. Figure 1 Standard of Cl16O4 at 0.2 ppb (upper) with internal standard Cl18O4 at 7.5 ppb (lower)

XIC of -MRM (4 pairs): 99.0/83.0 amu from Sample 42 (std 0.2ppb) of MJ150034.wiff (Turbo Spray) Max. 416.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

50

100

150

200

250

300

350

400

In

te

ns

it

y,

c

ps

10.45

10.59

10.86

13.5011.64 16.875.42 7.78 17.621.37 14.217.474.65 9.836.06 15.163.26

XIC of -MRM (4 pairs): 107.0/89.0 amu from Sample 42 (std 0.2ppb) of MJ150034.wiff (Turbo Spray) Max. 1.3e4 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.0

2000.0

4000.0

6000.0

8000.0

1.0e4

1.2e4

1.3e4

In

te

ns

it

y,

c

ps

10.44


Simulant.



II. Figure 2 Standard of Cl16O4 at 0.4 ppb (upper) with internal standard Cl18O4 at 7.5 ppb (lower)

XIC of -MRM (4 pairs): 99.0/83.0 amu from Sample 43 (std 0.4 ppb) of MJ150034.wiff (Turbo Spray) Max. 726.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

100

200

300

400

500

600

700

In

te

ns

it

y,

c

ps

10.44

2.08 11.34 17.37 19.1216.407.284.85 13.24 15.720.50 6.37 8.92 15.390.70

XIC of -MRM (4 pairs): 107.0/89.0 amu from Sample 43 (std 0.4 ppb) of MJ150034.wiff (Turbo Spray) Max. 1.3e4 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.0

2000.0

4000.0

6000.0

8000.0

1.0e4

1.2e4

1.3e4

In

te

ns

it

y,

c

ps

10.46


Simulant.



III. Figure 3 Standard of Cl16O4 at 1 ppb (upper) with internal standard Cl18O4 at 7.5 ppb (lower)

XIC of -MRM (4 pairs): 99.0/83.0 amu from Sample 44 (std 1ppb) of MJ150034.wiff (Turbo Spray) Max. 1796.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

200

400

600

800

1000

1200

1400

1600

1796

In

te

ns

it

y,

c

ps

10.49

3.03 15.585.93 12.366.66 17.9416.689.533.67

XIC of -MRM (4 pairs): 107.0/89.0 amu from Sample 44 (std 1ppb) of MJ150034.wiff (Turbo Spray) Max. 1.3e4 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.0

2000.0

4000.0

6000.0

8000.0

1.0e4

1.2e4

1.3e4

In

te

ns

it

y,

c

ps

10.43


Simulant.



IV. Figure 4 Standard of Cl16O4 at 2 ppb (upper) with internal standard Cl18O4 at 7.5 ppb (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

500

1000

1500

2000

2500

3000

3500

In

te

ns

it

y,

c

ps

10.45


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.0

2000.0

4000.0

6000.0

8000.0

1.0e4

1.2e4

1.3e4

In

te

ns

it

y,

c

ps

10.42


Simulant.



V. Figure 5 Standard of Cl16O4 at 5 ppb (upper) with internal standard Cl18O4 at 7.5 ppb (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

8000

In

te

ns

it

y,

c

ps

10.47

18.15


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.0

2000.0

4000.0

6000.0

8000.0

1.0e4

1.2e4

1.3e4

In

te

ns

it

y,

c

ps

10.44

11.74 12.56


Simulant.



VI. Figure 6 Standard of Cl16O4 at 10 ppb (upper) with internal standard Cl18O4 at 7.5 ppb (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.0

2000.0

4000.0

6000.0

8000.0

1.0e4

1.2e4

1.4e4

1.6e4

In

te

ns

it

y,

c

ps

10.50


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.00

2000.00

4000.00

6000.00

8000.00

1.00e4

1.20e4

In

te

ns

it

y,

c

ps

10.41


Simulant.



VII. Figure 7 Standard of Cl16O4 at 25 ppb (upper) with internal standard Cl18O4 at 7.5 ppb (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.0

5000.0

1.0e4

1.5e4

2.0e4

2.5e4

3.0e4

3.5e4

3.7e4

In

te

ns

it

y,

c

ps

10.48


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.00

2000.00

4000.00

6000.00

8000.00

1.00e4

1.20e4

In

te

ns

it

y,

c

ps

10.47

11.51


Simulant.



VIII. Figure 8. 2 Hrs exposure blank-1 with Cl16O4 (upper) and internal standard Cl18O4 (lower)

XIC of -MRM (4 pairs): 99.0/83.0 amu from Sample 17 (2h blk-1) of MJ150035.wiff (Turbo Spray) Max. 112.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

112

In

te

ns

it

y,

c

ps

3.151.95

9.2911.04

6.32 11.695.164.01 18.2215.875.391.15 7.274.89 10.64 17.41 19.468.44 13.40 15.041.31 7.9214.08

4.13 10.20


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

8000

9000

In

te

ns

it

y,

c

ps

10.41


Simulant.



IX. Figure 9. 2 Hrs exposure blank-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

10

20

30

40

50

60

70

80

90

100

In

te

ns

it

y,

c

ps

2.04

6.87

3.07 15.4510.27 16.460.58 6.62 9.973.74 5.28 11.41

16.974.93 8.18 12.97 13.84 18.381.4510.448.75


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

7786

In

te

ns

it

y,

c

ps

10.40

11.58 12.98 14.24


Simulant.



X. Figure 10. 2 Hrs exposure blank-3 with Cl16O4 (upper) and internal standard Cl18O4 (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

10

20

30

40

50

60

70

80

90

96

In

te

ns

it

y,

c

ps

2.609.232.06 9.81 18.11

2.9310.475.424.10

11.28 18.4917.6016.856.09 8.227.300.35 15.2513.365.21


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

7564

In

te

ns

it

y,

c

ps

10.48


Simulant.



XI. Figure 11. 2 Hrs exposure sample-1 with Cl16O4 (upper) and internal standard Cl18O4 (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

50

100

150

200

250

In

te

ns

it

y,

c

ps

6.43

10.33

2.962.73

11.558.79 14.743.83 16.906.204.67 17.80 18.8515.650.16 13.106.90


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

8000

9000

9596

In

te

ns

it

y,

c

ps

10.40

14.00


Simulant.



XII. Figure 12. 2 Hrs exposure sample-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

120

138

In

te

ns

it

y,

c

ps

10.38

2.6918.59

16.4015.45

9.39 19.090.53 7.570.70 14.215.75

6.19 18.085.55 13.52 17.8212.73


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

7632

In

te

ns

it

y,

c

ps

10.41

11.91


Simulant.



XIII. Figure 13. 2 Hrs exposure sample-3 with Cl16O4 (upper) and internal standard Cl18O4 (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

120

140

160

180

200

216

In

te

ns

it

y,

c

ps

10.38

2.50

10.062.0611.170.19 6.43 16.366.165.46 8.391.20 7.74 15.81 18.5613.637.24 17.87


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

8000

9000

In

te

ns

it

y,

c

ps

10.41

11.50 14.20


Simulant.



XIV. Figure 14. 1 Day exposure blank-1 with Cl16O4 (upper) and internal standard Cl18O4 (lower)

XIC of -MRM (4 pairs): 99.0/83.0 amu from Sample 72 (1 day blank-1) of MJ150034.wiff (Turbo Spray) Max. 118.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

118

In

te

ns

it

y,

c

ps

2.12

2.63

15.79

10.674.113.17 12.298.557.34 14.58 16.911.08 11.78

5.12 7.16 16.729.76 14.9218.6917.528.91 13.600.33


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

8000

9000

9880

In

te

ns

it

y,

c

ps

10.43

12.76


Simulant.



XV. Figure 15. 1 Day exposure blank-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

100

200

300

400

500

574

In

te

ns

it

y,

c

ps

14.48

14.9810.770.60

12.79


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

8000

9000

In

te

ns

it

y,

c

ps

10.40


Simulant.



XVI. Figure 16. 1 Day exposure blank-3 with Cl16O4 (upper) and internal standard Cl18O4 (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

120

136

In

te

ns

it

y,

c

ps

2.02

2.75

13.0713.77 16.59

7.2415.08

0.84 12.379.38 18.9818.3111.013.33 4.386.131.01 8.78 10.745.53


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

8000

8556

In

te

ns

it

y,

c

ps

10.42

11.30


Simulant.



XVII. Figure 17. 1 Day exposure sample-1 with Cl16O4 (upper) and internal standard Cl18O4 (lower)

XIC of -MRM (4 pairs): 99.0/83.0 amu from Sample 36 (1day-sample-1) of MJ150034.wiff (Turbo Spray) Max. 304.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

50

100

150

200

250

300

In

te

ns

it

y,

c

ps

10.44

2.59

10.03 11.1816.028.28


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

500

1000

1500

2000

2500

3000

3500

4000

4500

5000

In

te

ns

it

y,

c

ps

10.36

11.74 12.17


Simulant.



XVIII. Figure 18. 1 Day exposure sample-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

120

140

160

In

te

ns

it

y,

c

ps

10.40

10.56

10.712.62

10.072.0018.1115.324.54 7.52 14.549.216.464.81 11.25 13.165.92 18.6616.971.22

8.14


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

500

1000

1500

2000

2500

3000

3500

4000

4500

5000

In

te

ns

it

y,

c

ps

10.38

11.65 14.71


Simulant.



XIX. Figure 19. 1 Day exposure sample-3 with Cl16O4 (upper) and internal standard Cl18O4 (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

10

20

30

40

50

60

70

80

90

100

108

In

te

ns

it

y,

c

ps

10.56

2.12

2.532.87

3.637.17 9.974.45

9.45 19.704.953.37 9.19 12.59 16.091.78 17.6214.751.600.23 7.03 14.188.28


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

500

1000

1500

2000

2500

3000

3500

4000

4500

5000

In

te

ns

it

y,

c

ps

10.40

11.24


Simulant.



XX. Figure 20. 4 Days exposure blank-1 with Cl16O4 (upper) and internal standard Cl18O4 (lower)

XIC of -MRM (4 pairs): 99.0/83.0 amu from Sample 47 (4day blk-1) of MJ150034.wiff (Turbo Spray) Max. 132.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

120

132

In

te

ns

it

y,

c

ps

11.680.43 5.961.98

9.190.60 13.0310.817.40 15.653.52 7.58 19.546.66 17.978.91 17.544.72

12.15

XIC of -MRM (4 pairs): 107.0/89.0 amu from Sample 47 (4day blk-1) of MJ150034.wiff (Turbo Spray) Max. 1.0e4 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.00

1000.00

2000.00

3000.00

4000.00

5000.00

6000.00

7000.00

8000.00

9000.00

1.00e4

In

te

ns

it

y,

c

ps

10.38

11.75 12.2312.86 15.55


Simulant.



XXI. Figure 21. 4 Days exposure blank-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

120

140

160

180

200

In

te

ns

it

y,

c

ps

0.61

2.68

2.262.88

3.5312.005.42 8.956.394.65 10.857.24 17.0316.469.89

13.94


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

In

te

ns

it

y,

c

ps

10.40


Simulant.



XXII. Figure 22. 4 Days exposure blank-3 with Cl16O4 (upper) and internal standard Cl18O4 (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

120

140

In

te

ns

it

y,

c

ps

2.03

2.45

5.323.63

17.509.296.00 18.9312.6010.1118.687.88 15.450.89

14.13


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

In

te

ns

it

y,

c

ps

10.42

11.92


Simulant.



XXIII. Figure 23. 4 Days exposure sample-1 with Cl16O4 (upper) and internal standard Cl18O4 (lower)

XIC of -MRM (4 pairs): 99.0/83.0 amu from Sample 52 (S4 day sample-1) of MJ150034.wiff (Turbo Spray) Max. 212.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

120

140

160

180

200212

In

te

ns

it

y,

c

ps

10.39

10.54

10.25

2.73

2.08 7.576.4319.5913.9011.337.91 9.125.45 15.221.44 13.679.626.98 17.9517.54

XIC of -MRM (4 pairs): 107.0/89.0 amu from Sample 52 (S4 day sample-1) of MJ150034.wiff (Turbo Spray) Max. 7374.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

In

te

ns

it

y,

c

ps

10.41

11.85


Simulant.



XXIV. Figure 24. 4 Days exposure sample-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower)

XIC of -MRM (4 pairs): 99.0/83.0 amu from Sample 54 (4 day sample-2) of MJ150034.wiff (Turbo Spray) Max. 184.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

120

140

160

180

In

te

ns

it

y,

c

ps

10.31

2.15

2.665.821.95 18.52

16.87 19.6014.313.67 13.705.48 6.564.370.21 16.4912.399.997.14 7.505.33

XIC of -MRM (4 pairs): 107.0/89.0 amu from Sample 54 (4 day sample-2) of MJ150034.wiff (Turbo Spray) Max. 7858.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

7858

In

te

ns

it

y,

c

ps

10.42

12.02


Simulant.



XXV. Figure 25. 4 Days exposure sample-3 with Cl16O4 (upper) and internal standard Cl18O4 (lower)

XIC of -MRM (4 pairs): 99.0/83.0 amu from Sample 56 (4 day sample -3) of MJ150034.wiff (Turbo Spray) Max. 142.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

120

140

In

te

ns

it

y,

c

ps

10.41

2.86

14.44

2.421.5511.240.10

3.2018.52

7.808.86 16.369.94 17.704.34 15.7613.704.51 7.425.09 13.29

8.016.576.3319.759.24

XIC of -MRM (4 pairs): 107.0/89.0 amu from Sample 56 (4 day sample -3) of MJ150034.wiff (Turbo Spray) Max. 7596.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

1000

2000

3000

4000

5000

6000

7000

7596

In

te

ns

it

y,

c

ps

10.40


Simulant.



XXVI. Figure 26. 10 Days exposure blank-1 with Cl16O4 (upper) and internal standard Cl18O4 (lower)

XIC of -MRM (4 pairs): 99.0/83.0 amu from Sample 15 (10day blank-1) of MJ150036.wiff (Turbo Spray) Max. 140.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

120

140

In

te

ns

it

y,

c

ps

2.66

2.14 3.035.69

9.49 17.0717.2315.99 18.397.06

8.3513.80 14.311.42 13.083.54

17.85

XIC of -MRM (4 pairs): 107.0/89.0 amu from Sample 15 (10day blank-1) of MJ150036.wiff (Turbo Spray) Max. 1.1e4 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.00

1000.00

2000.00

3000.00

4000.00

5000.00

6000.00

7000.00

8000.00

9000.00

1.00e4

1.10e4

In

te

ns

it

y,

c

ps

10.52

12.02


Simulant.



XXVII. Figure 27. 10 Days exposure blank-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

120

130

In

te

ns

it

y,

c

ps

2.29

2.63

1.991.24

16.507.64 11.7510.908.82 14.110.23 6.06 12.794.68 7.91 14.711.6418.686.36

17.10


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.00

1000.00

2000.00

3000.00

4000.00

5000.00

6000.00

7000.00

8000.00

9000.00

1.00e4

1.10e4

In

te

ns

it

y,

c

ps

10.51

13.12 13.77


Simulant.



XXVIII. Figure 28. 10 Days exposure blank-3 with Cl16O4 (upper) and internal standard Cl18O4 (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

10

20

30

40

50

60

70

80

90

100

In

te

ns

it

y,

c

ps

11.053.70

2.592.32 13.63

5.28

16.19 17.880.90 14.441.90 3.00 9.16 14.8510.004.75 16.8011.858.427.71


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.00

1000.00

2000.00

3000.00

4000.00

5000.00

6000.00

7000.00

8000.00

9000.00

1.00e4

In

te

ns

it

y,

c

ps

10.48

14.07


Simulant.



XXIX. Figure 29. 10 Days exposure sample-1 with Cl16O4 (upper) and internal standard Cl18O4 (lower)

XIC of -MRM (4 pairs): 99.0/83.0 amu from Sample 9 (10day sample-1) of MJ150036.wiff (Turbo Spray) Max. 208.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

20

40

60

80

100

120

140

160

180

200

In

te

ns

it

y,

c

ps

7.2710.42

11.82

2.1515.32

2.67 2.8916.4910.00

5.75 14.541.780.91

12.663.26 4.14 17.65

XIC of -MRM (4 pairs): 107.0/89.0 amu from Sample 9 (10day sample-1) of MJ150036.wiff (Turbo Spray) Max. 1.1e4 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.00

1000.00

2000.00

3000.00

4000.00

5000.00

6000.00

7000.00

8000.00

9000.00

1.00e4

1.07e4

In

te

ns

it

y,

c

ps

10.46

11.61


Simulant.



XXX. Figure 30. 10 Days exposure sample-2 with Cl16O4 (upper) and internal standard Cl18O4 (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

50

100

150

200

250

300

350

400

In

te

ns

it

y,

c

ps

10.49

2.2317.615.59 7.043.87 6.56 18.727.75 9.69 16.2911.66 14.495.38 12.990.16 1.39 15.048.85


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.00

1000.00

2000.00

3000.00

4000.00

5000.00

6000.00

7000.00

8000.00

9000.00

1.00e4

1.10e4

In

te

ns

it

y,

c

ps

10.47

13.26 14.03


Simulant.



XXXI. Figure 31. 10 Days exposure sample-3 with Cl16O4 (upper) and internal standard Cl18O4 (lower)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

50

100

150

200

224

In

te

ns

it

y,

c

ps

7.58

10.38

2.25

15.65 18.620.09 17.84 19.059.408.52 11.96


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.00

1000.00

2000.00

3000.00

4000.00

5000.00

6000.00

7000.00

8000.00

9000.00

1.00e4

1.10e4

In

te

ns

it

y,

c

ps

10.47

11.64


Simulant.



XXXII. Figure 32. Validation with 10 days blank-1. Cl16O4 (upper) was spiked at the level of 150 ppt in

food and internal standard Cl18O4 (lower)

XIC of -MRM (4 pairs): 99.0/83.0 amu from Sample 50 (validation-1) of MJ150037.wiff (Turbo Spray) Max. 654.0 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

100

200

300

400

500

600

654

In

te

ns

it

y,

c

ps

10.51

1.950.10 3.13 18.4811.87 15.29 19.907.63 8.25 9.350.94 13.334.34 15.62

XIC of -MRM (4 pairs): 107.0/89.0 amu from Sample 50 (validation-1) of MJ150037.wiff (Turbo Spray) Max. 1.3e4 cps.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.0

2000.0

4000.0

6000.0

8000.0

1.0e4

1.2e4

1.3e4

In

te

ns

it

y,

c

ps

10.50

11.55


Simulant.



XXXIII. Figure 33. Validation with 10 days blank-2. Cl16O4 (upper) was spiked at the level of 150 ppt in



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

100

200

300

400

500

600

662

In

te

ns

it

y,

c

ps

10.52

2.25 3.800.10 2.074.28 6.97 12.93 14.886.42 17.748.923.28 13.719.555.92 18.75 19.53


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.0

2000.0

4000.0

6000.0

8000.0

1.0e4

1.2e4

1.4e4

1.5e4

In

te

ns

it

y,

c

ps

10.52


Simulant.



XXXIV. Figure 35. Validation with 10 days blank-3. Cl16O4 (upper) was spiked at the level of 150 ppt in



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0

100

200

300

400

500

600

700

782

In

te

ns

it

y,

c

ps

10.63

10.83

2.455.35 12.7712.324.780.23 13.669.16 15.598.481.62 6.29 17.14 18.203.74


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19Time, min

0.0

2000.0

4000.0

6000.0

8000.0

1.0e4

1.2e4

1.4e4

1.6e4

In

te

ns

it

y,

c

ps

10.54


Simulant.

Appendix-D Spiking Validation at Low Perchlorate Concentrations


Issued:

Author: ___________________ Date: ______________

Approved: ______________________ Date: ______________


Simulant.



Table of Contents I. Spiking validation with 10 days blank migration samples .............................................................. 3

II. Spiking validation with Tenax at low perchlorate concentrations .................................................. 3


Simulant.



I. Spiking validation with 10 days blank migration samples

Blank polymer plaque with 10 days exposure was used for validation. Tenax® was transferred into a beaker followed by spiking with 6 ng of perchlorate Cl16O4, corresponding to the LOQ level of 1.5 ng/in2 (150 ppt in food). Then 0.1 mL of internal standard 18O-labeled perchlorate Cl18O4 at 1.5 ppm and 150 mL of ACN was added. The solution was stirred for one minute and sit still for 5 min. 50 mL of the supernatant was then transferred into rotary evaporator tube for drying down to approximately 10 mL. The solution was then filtered through 0.2 µm PTFE syringe filter for LC/MS/MS analysis.

Table 1 Validation with 10 days blank migration samples

Validation replicate

Perchlorate spiked* (ng/in2)

Amount Spike* (ng)


Amount Found* (ng)

Recovery (%)

1 1.53 6.12 1.57 6.26 102.3

2 1.53 6.12 1.51 6.04 98.6

3 1.53 6.12 1.49 5.97 97.5 AVG 99.4 STD 2.5

RSD (%) 2.5


II. Spiking validation with Tenax® at low perchlorate

concentrations

Spiking experiments were also carried out with perchlorate spiked directly into Tenax® at one fourth

and half of the method LOQ level at 0.4 and 0.8 ng/in2. Except for the exposure procedure, all

sample preparation procedures were the same as those for analysis of migration samples. Results

are shown in Table 2 Spiking at 0.4 ng/in2and Table 3 spiking at 0.8 ng/in2


Simulant.



Table 2 Spiking at 0.4 ng/in2

Validation replicate Perchlorate spiked* (ng/in2)

Amount Spike* (ng)


Amount Found* (ng)

Recovery (%)

1 0.4 1.5 0.5 1.9 125

2 0.4 1.5 0.3 1.4 91

3 0.4 1.5 0.4 1.8 116

AVG 111

STD 18

RSD (%) 16


Table 3 spiking at 0.8 ng/in2

Validation replicate Perchlorate spiked* (ng/in2)

Amount Spike* (ng)


Amount Found* (ng)

Recovery (%)

1 0.8 3.1 0.8 3.3 113

2 0.8 3.1 1.0 4.1 142

3 0.8 3.1 0.9 3.7 126

AVG 127

STD 15

RSD (%) 12


supplemental comments from basf corporation (keller and...

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