Work carried out in the Czech Republic - CIPAC · 0,0010 0,0015 0,0020 0,0025 0,0030 0,0035 0,0040...

Work carried out in the Czech Republic

CENTRAL INSTITUTE FOR SUPERVISING AND TESTING IN AGRICULTURE

Olga Nováková

CIPAC Symposium 24th June 2014, Liege, Belgium

NATIONAL REFERENCE LABORATORYDepartment of Testing Plant Protection Product

(version for CIPAC website)

CONTENT

1. Introduction 2. Steps of laboratory control of PPPs in CZ3. Example of analysis of suspicious samples4. Problems with analysis of suspicious samples5. Suggestion to discussion6. Conclusion

CIPAC Symposium 24th June 2014, Liege, BelgiumOlga Nováková 2

1997‐2013: 2 institutes• SPA (State Phytosanitary Administration)

– Postregistration Control Division• Department of Laboratory Testing Pesticide (= NRL for PPP)

• CISTA (Central Institute for Supervising and Testing in Agriculture)– Division: National Reference Laboratory (9 laboratories)

From 1.1.2014: 1 institute• SPA + CISTA = CISTA

– Division: National Reference Laboratory (9+1 laboratories)• Department of Testing Plant Protection Products = former Laboratory Testing Pesticide

Olga Nováková CIPAC Symposium 24th June 2014, Liege, Belgium 3

REORGANIZATION OF FORMER STATE PHYTOSANITARY ADMINISTRATION (SPA)

STRUCTURE OF CENTRAL INSTITUTE FOR SUPERVISING AND TESTING IN AGRICULTURE (CISTA)

Olga Nováková 4CIPAC Symposium 24th June 2014, Liege, Belgium

General Director

Authority Office

Economic and Administration Section

Section of Protection against Harmful Organisms

Section of Plant Production

Section of Agricultural Inspection

Section of Agricultural Inputs

National Reference Laboratory

Division of PPPCoordination

EvaluationPostregistration

3 Divisions

4 Divisions

7 Regional Divisions

Division of Audit and Quality Management

Division of Feed, Fertilisers and Soil

Division of Agricultural Inputs Control

STRUCTURE OF NATIONAL REFERENCE LABORATORY


Director of NRL

Specialized DepartmentsRegional Departments

Brno

Opava

Plzeň

Praha

BrnoDepartment of Testing Plant Protection Products

Planá nad Lužnicí

BrnoDepartment of Proficiency Testing Programes

BrnoDepartment of Microbiology and Biochemistry

LípaDepartment of Special Plant and Feed Analyses

DEPARTMENT OF TESTING PLANT PROTECTION PRODUCTS


Head of Department

Laboratoryof physicochemical

methods

Laboratoryof chromatographic

methods

Post‐registration Control Department ‐ responsibility• annual plan with cooperation of laboratory• sampling original packages ‐ PPP samples are taken directly

in the distributor stores• labeling control• control of sale, storage and use of PPP• control of technical state of application machinery

Laboratory‐ responsibility and competence• Laboratory analysis• laboratory check on physical and chemical properties of plant

protection products and other plant protection preparations and chemical compositions

• report of analysis (+results with evaluation)

Post‐registration Control Department – responsibility• reconvey the rest of samples• evaluation of results from laboratory in accord with act (law)• information of authorization holders • fine


SYSTEM OF POSTREGISTRATION CONTROL –responsibility

Samplingoriginal packages

Laboratory analysis+

report

Evaluation of results in accord

with act

Planned samples • postregistration control according to annual plan

• samples within the process of PPP approvalUnplanned samples• unknown samples• suspicious samples (e.g. counterfeit)Proficiencies tests (AAPCO, AFSCA, …)


LABORATORY CONTROL OF PPPs

• Registration of samples (LIMS)• Laboratory sampling

– Original sample package is higher 1L or 1kg – Original sample package is lower 1L or 1kgPartition of laboratory sample = Analytical sample

• Laboratory analysis • Evaluation of laboratory analysis

– Certificate of analysis (Agreement or disagreement with specification)

– Detailed expert reports


STEPS OF LABORATORY CONTROL

AU

-0,0005

0,0000

0,0005

0,0010

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0,0065

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0,0075

0,0080

Minutes0,00 2,00 4,00 6,00 8,00 10,00 12,00 14,00 16,00 18,00 20,00 22,00 24,00 26,00 28,00 30,00 32,00 34,00 36,00 38,00 40,00 42,00 44,00 46,00 48,00 50,00

Planned samples – postregistration control according to annual planFAO specification or existing national specification

– Identity and content of active substances – Identity and content of relevant impurities– Physical, chemical and technical properties– Storage stability tests

Xylene in EC formulations

Samples from parallel import – chromatographic and FTIR comparison with reference sample.

Methods: CIPAC, OECD or equivalent and the validated methods of producers, which are submitted as part of registration dossiers.

Laboratory verifies all methods according to standard operation procedure (SOP‐PP‐08‐01).


LABORATORY ANALYSIS

Samples within the process of PPP approval Aim:• Verify input data (technical specification)• Verification or validation analytical methods in CZ conditions (different column…)

• Obtain knowledge of PPP• Focus on chemical composition • Time for discussion of laboratory analysis ‐analytical results with authorization holders

• Input data for postregistration control• Future ‐ Help with detection of counterfeit


LABORATORY ANALYSIS – cont.

Unplanned control PPPs samples• Unknown samples (samples without label, confusion of active ingredient,…)

• Suspicious samples


LABORATORY ANALYSIS – cont.

• Identity and content of active ingredient • Identity and content of relevant impurities• Physical, chemical and technical properties • Chemical composition of sample (co‐formulants,

impurities,…) • Comparison with reference sample (GC, LC, FTIR)

Additional tests for clarification of unregistered sample: GC/MS, FTIR, Particle size distribution (CIPAC MT187), Determination of Sulphated ash (CIPAC MT29), Pour and tap bulk density (USP2/ASTM), Density, Viscosity (CIPAC MT192), Surface tension (OECD 115), TGA method, DSC method


LABORATORY ANALYSIS –SUSPICIOUS SAMPLES

Appearance:


EXAMPLES OF ANALYSIS

Differences in colourbetween reference sample and suspicious sample

Differences of shapegranules between reference sample and suspicious sample

Density:



Density g/ml

Reference PPP 1,170

Suspicious PPP 1,126

Result:Suspicious PPP is probably diluted or hasdifferent chemical composition.

Density:



Density g/ml Dean – Stark (CIPAC MT 30.2) Amount of water

(% w/w)

Amount of active ingredient (%)

Reference PPP 1,170 44,6 30,4

Suspicious PPP 1,126 60,7 21,8

Result:Suspicious sample is diluted.

Confirmation:•determination of water by Dean‐Stark (CIPAC MT 30.2)•determination of amount active ingredients

Sulphated ash (CIPAC MT 29):



Reference PPP (different batches)

year Sulphated ash [% w/w]

2001 1,52

2003 1,30

2006 1,37

2008 1,22

2012 1,26

2013 1,05

average 1,29

Suspicious PPP

Year Sulphated ash [% w/w]

2001 ‐‐‐

2003 15,52

2006 ‐‐‐

2008 19,40

2012 10,52

2013 8,62

2013 6,83

Result: There are difference between inorganic ions in suspicious sample and reference sample ‐ the chemical composition of suspicion sample is different from reference sample.

Surface tension (OECD 115):



Result: Suspicious PPP probably does not contain surface active agent.

PPP Surface tension of 0,01% w/w solution (mN/m)

Surface tension of 0,5% w/w solution

(mN/m)

Surface tension of 1,0 % w/w solution

(mN/m)

Reference PPP 57,0 32,9 32,7

Suspicious PPP 59,4 69,9 71,8

Particle size distribution (CIPAC MT 187) :



Reference PPP Suspicious PPP

Particle size distribution (CIPAC MT 187) :



Result: Particle size distribution of particles which disperse in water in suspicious PPP are different. It means that it can be different technology of production.

Reference PPP Suspicious PPPDiameter at 10% 0,68 μm 0,80 μm

Diameter at 50% 2,45 μm 3,77 μm

Diameter at 90% 6,07 μm 9,68 μm

Mean diameter 2,97 μm 4,56 μm

HPLC screening by HPLC/UV:



Result: Suspicious PPP 1 has probably identical chemical composition with reference PPP, Suspicious PPP 2 has not identical chemical composition with reference PPP.

AU

-0,0005

0,0000

0,0005

0,0010

0,0015

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0,0025

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0,0075

0,0080

Minutes0,00 2,00 4,00 6,00 8,00 10,00 12,00 14,00 16,00 18,00 20,00 22,00 24,00 26,00 28,00 30,00 32,00 34,00 36,00 38,00 40,00 42,00 44,00 46,00 48,00 50,00

Reference PPP (black), Suspicious PPP 1 (blue) Suspicious PPP 2 (green)

GC/FID – identification of co‐formulant:



Reference PPP (blue) Suspicious PPP (red)

min2.5 5 7.5 10 12.5 15 17.5 20

Norm.

10

15

20

25

30

35

FID1 B, (130708\ZK000005.D)

11.

644

FID1 B, (130708\ZK000008.D)

11.

658

GC/FID – identification of co‐formulant:



Result: Suspicious PPP does not contain surface‐active agent which is present in reference PPP.

Reference PPP (blue) Suspicious PPP (red)Surface‐activ agent (green)

min10 11 12 13 14 15 16 17

pA

10

15

20

25

30

35

FID1 B, (130708\ZK000005.D) FID1 B, (130708\ZK000008.D) FID1 B, (130708\ZK000004.D)

11.

644

Overlay chromatograms of reference PPP and suspicious PPP and Surface‐active agent, extract 10‐18min

GC/MSD identification of co‐formulants:



Reference PPP (green), Suspicious PPP (black)

GC/MSD chromatographic profile of PPP (split 100:1, extract 2‐4min)

2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.800

100000

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Time-->

Abundance

TIC: 11101210.DTIC: 11101211.D (*)

N,N‐dimethylformamide

Ethylene glycol

MSD spectrum of ethylene glycol

20 40 60 80 100 120 140 160 180 200 220 240 260 2800

20000

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m/z-->

Abundance

Scan 27 (2.206 min): 11102502.D31

43

6284 207133 28198

MSD spectrum of N,N‐dimethylformamid

20 30 40 50 60 70 80 90 1001101201301401501601701801902002100

200000

400000

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1000000

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m/z-->

Abundance

Scan 127 (2.671 min): 11113005.D73

44

30 58

20721 91 191





GC/MSD chromatographic profile of PPP (split 10:1, extract 4‐8min)MSD spectrum of naphthalene

4.00 4.50 5.00 5.50 6.00 6.50 7.00 7.500

50000

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Time-->

Abundance

TIC: 11101205.DTIC: 11101206.D (*)

naphthalene20 40 60 80 100 120 140 160 180 200 220 240 260 280

0

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Abundance

Scan 770 (5.660 min): 11102604.D128

1026451 75 873928 113 207 281





GC/MSD chromatographic profile of PPP (split 10:1,extract 8‐11,6 min)

MSD spectrum of N‐octyl‐2‐pyrrolidone

8.00 8.50 9.00 9.50 10.00 10.50 11.00 11.500

50000

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Abundance

TIC: 11101205.DTIC: 11101206.D (*)

N‐octyl‐2‐pyrrolidone

20 40 60 80 100 120 140 160 180 200 220 240 260 2800

20000

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m/z-->

Abundance

Scan 1341 (8.315 min): 11113009.D98

19770 126

112

41

86 14028 55 154168180 281209





GC/MSD chromatographic profile of EW formulation PPP ‐ active ingredient (extract 11,6‐12,6 min)

11.60 11.70 11.80 11.90 12.00 12.10 12.20 12.30 12.40 12.50 12.60

500000

1000000

1500000

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Time-->

Abundance

TIC: 11101210.DTIC: 11101211.D (*)

active ingredient




Result: The chemical composition of suspicious PPP are different from reference sample.

Summary:

Identified components Reference PPP Suspicious PPP Content % w/w

N,N‐dimethylformamide No Yes 16,78

N‐octyl‐2‐pyrrolidone No Yes 0,306

Ethylene glycol No Yes 1,525

Naphthalene No Yes 0,018

UV‐VIS spectroscopy:



Reference PPP Suspicious PPPVIS spectrum:

Sample λ max v DI water [ nm]

Reference sample 628 nm

Suspicious sample 620 and 657 nm

Result: The suspicious PPP does not contain the same dye as reference sample.

FTIR spectroscopy :



Reference PPP (blue), Suspicious PPP (red)

FTIR spectroscopy :



FTIR spectrum of Insoluble residues of PPP in CH2Cl2

Reference PPP (blue), Suspicious PPP (red)

FTIR spectroscopy :



Result: The suspicious PPP contains kaolin = the chemical composition is different from reference sample.

FTIR spectrum of Insoluble residues of suspicious PPP in CH2Cl2 and kaolin CAS No. 1332‐58‐7

Suspicious PPP (blue), kaolin CAS No. 1332‐58‐7(red)

Differential Scanning Calorimetry (DSC):




Result: The chemical composition of suspicious sample is different from reference sample.

Thermogravimetric analysis (TGA):




Result: Different chemical composition of volatile compounds present in PPP or/and other compounds in PPP.

• Active ingredients and relevant impurities in suspicious PPP mostly agree with specification.

• We usually find differences in chromatography profile and/or in FTIR spectra between reference and suspicious samples:

– we use the other technics and methods for identification.

• Necessity of cooperation between laboratories of national authority and producers (in interpretation of raw data, support of analytical method, standards…)

• Necessity of clear legislation.


CONCLUSION

THANK YOU FOR THE ATTENTION!

CENTRAL INSTITUTE FOR SUPERVISING AND TESTING IN AGRICULTURE

NATIONAL REFERENCE LABORATORYDepartment of Testing Plant Protection Product

Name: Olga Novákováe-mail: [email protected]: Department of Testing Plant Protection ProductAddress: Zemědělská 1a, 613 00 Brno, Czech Republicweb: www.ukzuz.cz

Work carried out in the Czech Republic - CIPAC · 0,0010 0,0015 0,0020 0,0025 0,0030 0,0035 0,0040...

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