Directeurs de Thèse : Dr. S. MOUNIER Université du Sud Toulon Var – PROTEE

1

Directeurs de Thèse :

Dr. S. MOUNIER Université du Sud Toulon Var – PROTEE(PROcessus de Transferts et d'Echanges dans l'Environnement)

Dr. D. OMANOVIĆ Institut Ruđer Bošković – LPCT(Laboratory for Physical Chemistry of Traces)

« Mise au point d’une systématique de caractérisation des interactions Matière Organique Naturelle Dissoute

(MOND) – Contaminants métalliques »

Université du Sud Toulon Var 21 novembre 2008

Thèse de Doctorat soutenue par:

M. Yoann LOUIS

En vue d’obtenir le titre de Docteur de l’Université du Sud Toulon-Var

Subvention N° 03/1214910/TMatière Organique NAturelle en miLIeu SAlé

2

1.Introduction

2.Analytical protocol improvements

3.Concentrated Marine DNOM study

4.Natural Estuarine ecosystem study

5.Conclusions & perspectives

SUMMARY

Introduction – Analytical protocol – Marine DNOM study – Estuarine DNOM Study - Conclusion

3

I. Introduction


4

• Trace metals in the environment


Metals

Metals

Metals

Natural origin

Anthropogenic origin

MetalsAQUATIC ENVIRONMENT

(Coastal and estuarine system)SOILS

WATER

ATMOSPHERE

SEDIMENTS

5

Metals


“Oligoelements”:necessary for metabolismCu, Fe, F, Mg, Mn, Zn, …

Toxic metals:not neededPb, Hg, Cd, …

Concentra

tion in

crease

Toxicity ≠ Total concentration

When metal became toxic ? depend on its speciation

• Trace metals in the environment

6

Generally toxic for biota

MM

M

Micro-organisms (bacteria, virus,…)

Organic and Inorganic Particules

M n+

Inorganic Ligands

Cl-, NO3-, SO4

2- …

OH-

Filtration (0.45µm)

Particulate > 0.45 µm Dissolved < 0.45 µm

M

MM

M

Water column

Dissolved metal speciation

Organic Ligands

EDTA, DNOM …

Metal trap: Less toxic

• Trace metals speciation in the water column


MétalMétal

“Not” or lessbioavailable

bioavailable

[MTOTAL] ≠ [MTOXIC]

“Not” or less bioavailable

Could be bioavailable

7

• DNOM Origin?


Anthropogenic activities

PhotosynthesisBacterial activitydegradation

Phytoplankton activity

Humification&Polymerization DNOM modifications

Heterogeneous origins heterogeneous and complex structure

Plants, animals, µorganisms decomposition River input

Representation of a simplified NOM

8

• DNOM speciation


“Analytical speciation” “Mechanistical speciation”

“structural” determination

separation and analysis

Dialysis, UF GFAASCFFFF, HPSEC ICP-MSHPLC, GC CV-AFSC18, Chelex Voltammetry… ...

Specific components determination

Less usable for metal behavior prediction

Interactions characterization

ISEVoltammetry

Fluorescence Quenching…

Results usable in speciation codes for prediction

(for example MOCO from IFREMER)

No Functional characterization

9

KHthermo

KMthermo

kM1

kM-1

• DNOM speciation


DNOM-Metal interaction study

•Continuous model: NICA-Donnan•Discrete model: WHAM

M+

Used to describe the DNOM reactivity

Assuming a kinetic of 1st order

LDNOM

M

H++

H+

+ Comp KCompthermo

Comp

LDNOM

For 1 DNOM: All K and [LiT] determined = “Chimiotype”

For Metal-DNOM interaction study: Need a technique to measure only M

orM

LDNOMMM

LDNOM LLDNOM

10

3 electrodes system:Counter electrode (Pt)Reference electrode (Ag/AgCl/KClsat)Working electrode (Hg)

StirrerPurging (N2)

Metal additionOxydation step

Escan (V)

-0.5 -0.4 -0.3 -0.2 -0.1 0.0 0.1

Inte

nsi

ty (

A)

0.0

2.0e-9

4.0e-9

6.0e-9

8.0e-9

1.0e-8

1.2e-8

1.4e-8

1.6e-8

• Analytical tool used to measure trace metal: DPASV

M

DNOM

L Metal-Ligand Complex

DNOM

L

DNOM

L

M

DNOM

L

DNOM

L

DNOM

L

DNOM

L

DNOM

L

DNOM

L

M

M

M

M

Reduction Step

Edep

e-

MM

M

M

M

MM

M

MM

Edep

After tdep = 5 minEscan

Voltamogram

I=f([M])

Escan


Direct measurement of free & inoganic copper fraction

= electrolabile fraction

(= bioavailable fraction)

11

-9.50

-9.00

-8.50

-8.00

-7.50

-7.00

-6.50

-6.00

-5.50

-5.00

-9.50 -9.00 -8.50 -8.00 -7.50 -7.00 -6.50 -6.00 -5.50 -5.00

pCuT

pCu

Lab

[Metal added]: From nM to µM

5.8

6.2

6.6

7.0

7.4

7.8

8.2

8.6

5.86.26.67.07.47.88.2pCuT

pCul

ab

1:1Sal11 expSal11 calc

Metal complexed by DNOM

Data at equilibrium Kequilibrium, [LT]

0.0E+00

2.0E-09

4.0E-09

6.0E-09

8.0E-09

1.0E-08

1.2E-08

1.4E-08

1.6E-08

0 50 100 150t (mn)

Cu

(M)

CuT

Culabile

Kinetic k1, k-1, [LT]

For each point:2h of equilibriumMeasurements every 6 min.

Discrete fitting of experimental data with

PROSECE program(Speciation calculus + optimization)

Determination of Kequilibrium, [LT]

Determination of k1, k-1, [LT] New characterization of the DNOM: reactivity

• Metal logarithmic scale titration (Garnier et al., 2004, Env. Technol. 25, 589-599)


12

0

2

4

6

8

10

12

14

16

-0.65-0.60

-0.55-0.50

-0.45-1.2

-1.0-0.8

-0.6-0.4

stru

ja /

A

potencijal / Vpotencijal akumulacije / V

potencijal akumulacije / V

-1.4 -1.3 -1.2 -1.1 -1.0 -0.9 -0.8 -0.7 -0.6 -0.5 -0.4

stru

ja /

A

0

2

4

6

8

10

12

14

16

0

2

4

6

8

10

12

14

16

-0.65-0.60

-0.55-0.50

-0.45-1.2

-1.0-0.8

-0.6-0.4

stru

ja /

A



-1.4 -1.3 -1.2 -1.1 -1.0 -0.9 -0.8 -0.7 -0.6 -0.5 -0.4

stru

ja /

A

0

2

4

6

8

10

12

14

16

0

2

4

6

8

10

12

14

16

-0.65-0.60

-0.55-0.50

-0.45-1.2

-1.0-0.8

-0.6-0.4

stru

ja /

A



-1.4 -1.3 -1.2 -1.1 -1.0 -0.9 -0.8 -0.7 -0.6 -0.5 -0.4

stru

ja /

A

0

2

4

6

8

10

12

14

16

0

2

4

6

8

10

12

14

16

-0.65-0.60

-0.55-0.50

-0.45-1.2

-1.0-0.8

-0.6-0.4

stru

ja /

A



-1.4 -1.3 -1.2 -1.1 -1.0 -0.9 -0.8 -0.7 -0.6 -0.5 -0.4

stru

ja /

A

0

2

4

6

8

10

12

14

16

0

2

4

6

8

10

12

14

16

-0.65-0.60

-0.55-0.50

-0.45-1.2

-1.0-0.8

-0.6-0.4

stru

ja /

A



-1.4 -1.3 -1.2 -1.1 -1.0 -0.9 -0.8 -0.7 -0.6 -0.5 -0.4

stru

ja /

A

0

2

4

6

8

10

12

14

16

0

2

4

6

8

10

12

14

16

-0.65-0.60

-0.55-0.50

-0.45-1.2

-1.0-0.8

-0.6-0.4

stru

ja /

A



-1.4 -1.3 -1.2 -1.1 -1.0 -0.9 -0.8 -0.7 -0.6 -0.5 -0.4

stru

ja /

A

0

2

4

6

8

10

12

14

16

Voltammograms Pseudopolarogram

• Pseudopolarography measurements (Nicolau et al., 2008, ACA 618, 35-42)


Deposition potential (V)Deposition potential (V)Scanning potential (V)

Inte

nsi

ty (

nA

)

Inte

nsi

ty (

nA

)

Edep for CC measurements

Labile fraction

Direct ML complex reduction

Labile fraction = Free + inorganic fraction : bioavailableDissociable organic fraction: Probably not bioavailableNot measured fraction = electroinactive in the Edep range used

Measured if: UV, pH=2,

Edep << Edep for CC

13

8. Goals of the study


Analytical protocol determinationadapted to low [DOC] and [Metal]

Improvements:

•Technical •Analytical •Mathematical “Model DNOM” definition

Based on the concentrated sample from GDR MONALISA

Real complex natural ecosystem study

14

II. Analytical Protocol determination


15Introduction – Analytical protocol – Marine DNOM study – Estuarine DNOM Study - Conclusion

• Technical and mathematical improvements

pCuT6.06.57.07.58.0

pC

ula

bile

6.0

6.5

7.0

7.5

8.0

8.5

9.0

E (V)-0.5 -0.4 -0.3 -0.2 -0.1 0.0

i (A

)

0

1e-9

2e-9

3e-9

4e-9

5e-9 experimentals pointsfitted curvepeak 1 peak 2baseline

•Limit adsorption (Teflon use)•Precise metal additions (automatic pumps 500µl)•Avoid pollution with additions (tubing separation)•Avoid evaporation (N2 wet purging)•Mathematical baseline and peak definition •Multi-PROSECE (more optimization loop & confidence interval calculus)

16

• Analytical improvement: Surface Active Substances (SAS) interferences (Louis et al., 2008, ACA 606, 37-44)


M M

M

e-

Edep

SAS

Escan

e-

Escan

SAS

M M

M

I [Cu]meas [LT]

-0.45V without SAS

-0.45V with SAS

Distorded shape

17

Analytical process to get rid of SAS interferences during the stripping step

Additional experiment A.C Voltametry (Phase angle: 90° measure of capacitive current)

Max. Ads.At pzc

Only 1% of the total deposition time (297s)


Edep = -0.6V

Edep = -1.6V

Edep = -0.6V + 3s at -1.6V

Classical used tdep

Classical Edep used for Cu

ΔI ↑ = Itcap- It0

cap = SAS quantity ↑

High influence of SAS at tdep = 300 s and Edep ≈ - 0.5 V

- +Eacc= -0.45 V + 3sec at -1.6V Eacc= -1.6 V

Eacc= -0.45 V

Full circles Edep=60 sDotted circles Edep=60 s + 1s at -1.6VTriangles Edep =60s (After UV)

18

Influence of these SAS on the apparent [LT]

Without 3sec

[LT]= 335 nM

logK=6.17

With 3sec

[LT]= 160 nM

logK=6.47

[LT] change from 335 nM to 160 nM Artificially « Hidden Metal » by SAS bad speciation determination toxicity


Ruzić linearization

19

Filtration at 0.45µm

DOC

Total Metal

UV at pH2

Salinity or majors ions by Ionic

Chromatography

Log addition window determination :

Add1= 10% MiniFinal conc.:

1mgC/L 1µM10 mgC/L 10µM

Pseudo Edep

(Optional)

H+ , Ca2+ competition(After Chelex)

Log additions at Edep, Kinetic experiment

PROSECE

Raw sample Potentiometry(Chelex)

(1)

(2)

(4)

(3)

(5)

(6)

(7)

(8)

(9)

(10)

(11)

• Analytical protocol for DNOM study


For concentrated samples

“Chimiotype”

20

III. Study of a natural seawater sample (MONALISA project)

(Article submitted to Marine Environmental Research)


21

Military port

1. Sampling site: Balaguier Bay


1000L seawater sampling (online filtration and nanofiltration and reverse osmosis concentration by GDR MONALISA, ISM-LPTC: E. Parlanti, PhD of Arnaud Huguet). Concentrated from 1000 L to ~10 L, [DOC]final= 30.4 mg/L

Site interest: Coastal Semi-Closed Area under

anthropogenic influencesGoal: Give standard DNOM usable in metal

speciation/transport program

> 100 nM

~ 15 nM

~ 5 nM

22

2

4

6

8

10

12

0 0.02 0.04 0.06 0.08 0.1 0.12 0.14nOH

-ajouté (mmol)

pH

-5

-4

-3

-2

-1

0

1

2

3

4

5

Err

eur

(%)

PROSECE Fitting for 4 types of acidic sites (DOC=1.2mmolC.L-1).

LH1 LH2 LH3 LH4Carboxylic

like Phenolic

like

Total acidic Sites

LHiT

(meq/molC)210 ± 10.8 54 ± 2.4 80.4 ± 1.2 100.8 ± 1.2 265.2 181.2 446.4

pKa 3.6 ± 0.1 4.8 ± 0.1 8.6 ± 0.1 12.0 ± 0.4

60% 40%

Lu and Allen (2002) : Suwanee River(also concentrated by RO)

30%70%

Carboxylic-like Phenolic-like

165.3

/2.7


• Potentiometry on Concentrated DNOM (Garnier et al., 2004, Water Research, 38, 3685-3692)

(Letizia and Gnudi, 1999)

23

pCuT

45678

Cu

T (

%)

0

20

40

60

80

100

5,6 4,6

1st site saturation

2nd site saturationEdep = -0.5V

Estimation of a [1st site]: 90% x 2.5µM = 2.25 µM (= 1.87 meq/molC)

Estimation of a [2nd site]: 50% x 25µM - [1st site]: = 10.25 µM (= 8.54 meq/molC)

• Exploratory experiment: Pseudopolarography coupled with logarithmic addition


24

• Log addition and Cu2+ competition with H+ and Ca2+

[Cu]T = 12.5µM, pH = 8.2 [Cu]T = 4µM.

LM1 LM2

LMiT (meq/molC) 1.72 ±0.13 10.25 ±2.7

logKCuL 9.9 ±0.1 6.9 ±0.1

logKCaL 2.5 ±0.4 5.5 ±0.6

pKa 8.6 ±0.1 8.2 ±0.3

Complexing parameters determined after simultaneous fitting by PROSECE of the 3 experiments

Strong affinity of copper for the studied DNOM

Total metal binding site density LMT

12

Ca additions

% Cu lab

≈ 2µM of Cu bound to specific sites

% Cu lab

pH

Strong affinity toward proton phenolic-like sites

Phenolic-like sites

Total acidic sites density

446

~3% of

(= Buffle, 1988)

Strong complexing site specific to copperHight calcium competition effect


Edep = -0.5V, pH = 8.2, DOC = 1.2mmolC.L-1.

[L M1] =

0.03 meq/mol C

Comparison of 2 different Edep (-0.5V and -1.5V).

Closed to values estimated with pseudo coupled with log add.

Model Marine DNOM complexing parameters = DNOM “Chimiotype”(Comparable to standards OM used in NICA-Donnan /WHAM models, obtained for soil/river extracted OM)

LH1 LH2 LH3 LH4

LHiT

(meq/molC)210 ± 10.8 54 ± 2.4 80.4 ± 1.2 100.8 ± 1.2

pKa 3.6 ± 0.1 4.8 ± 0.1 8.6 ± 0.1 12.0 ± 0.4

25

seawater sample treated with Chelex (DOC = 0.09 mmolC.L-1); pH = 8.2, Salinity 37.

Experimental pointsDNOM simulated by Mineql adjusting only [DOC]Difference between modeled DNOM and experimental points

<<5%

• Correct simulation validating the characterization protocol• DNOM reactivity is not strongly modified by concentration stepModel DNOM determined usable


• MINEQL simulation of natural DNOM according to determined model marine DNOM

26

7.5 8.3N

atur

al m

arin

e w

ater

cond

itio

ns

80% of total copper complexed as organic forms >90% found in several paper: Influence of SAS ? specific behavior of the studied DNOM and high copper content

Condition: Majors ions for salinity of 38, DOC = 0.09 mmolC.L-1, Cutot = 14.8nM


• Simulation of copper speciation for the studied marine environment

27

IV. Estuarine DNOM Study (Article submitted to Marine Chemistry)


28

• Sampling in the water column: gradient of salinity

-9

-8

-7

-6

-5

-4

-3

-2

-1

0

0 10 20 30 40

Salinity

Dep

th (

m)

FSI layerBrackish

Seawater

1. Sampling site: Krka, Croatia (2007&2008)

•Pristine watershed•Potential anthropogenic inputs in estuary•On site measurements in nearby laboratory


•Low tide on Adriatic sea stratified estuary

•Challenge is to give data on speciation and kinetic in this natural area

29

Filtration at 0.45µm

DOC

Total Metal

UV at pH2

Salinity or majors ions by Ionic

Chromatography

Log addition window determination :

Add1= 10% MiniFinal conc.:

1mgC/L 1µM10 mgC/L 10µM

Pseudo Edep

(Optional)

H+ , Ca2+ competition(After Chelex)

Log additions at Edep, Kinetic experiment

PROSECE

Raw sample Potentiometry(Chelex)

(1)

(2)

(4)

(3)

(5)

(6)

(7)

(8)

(9)

(10)

(11)

• Simplified protocol used


No concentration step and no use of Chelex

30

Salinity

0 10 20 30 40

Dep

th (

m)

-8

-6

-4

-2

0

DOC (µMC)

40 50 60 70 80

CuT (nM)2 4 6 8 10 12

Salinity0 10 20 30 40

Cu

T

2

4

6

8

10

12

Salinity0 10 20 30 40

DO

C

40

50

60

70

80

A B C

• Same curve shape for 2007 & 2008 • Oligotrophic freshwater Very few carbon content, DOC est. < DOC sea low anthrop. inputs• Non conservative behavior: Bigger amount of metal & DOC in the FSI “special layer”• Additional source of DOC in the FSI: can be due to an exacerbated biological activity (Svensen et al, 2006)

• Increase of copper in the FSI: particulate/dissolved metal exchange due to salinity increase


• Salinity, DOC and Copper profiles(Elbaz-Poulichet F. et al., 1991) 1.78 in may 1988

31

0.0E+00

2.0E-08

4.0E-08

6.0E-08

8.0E-08

1.0E-07

1.2E-07

1.4E-07

6.97.17.37.57.77.9pCuT

LT

(mol

/L)

2

4

6

8

10

logk

1 &

logK

th

5.5

6.0

6.5

7.0

7.5

8.0

8.5

9.0

5.56.06.57.07.58.0 pCuT

pCul

ab

-10

-5

0

5

10

erre

ur (

%)

5.5

6.0

6.5

7.0

7.5

8.0

8.5

9.0

5.56.06.57.07.58.0 pCuT

pCul

ab

-10

-5

0

5

10

erre

ur (

%)

5.5

6.0

6.5

7.0

7.5

8.0

8.5

9.0

5.56.06.57.07.58.0 pCuT

pC

ula

b

-10

-5

0

5

10

erre

ur

(%)

Fitting at EquilibriumData at Equilibrium

0.0E+00

5.0E-09

1.0E-08

1.5E-08

2.0E-08

2.5E-08

3.0E-08

3.5E-08

4.0E-08

4.5E-08

5.0E-08

0 1000 2000 3000 4000 5000 6000 7000 8000t (s)

Cul

abil

e (M

)

0.0E+00

5.0E-09

1.0E-08

1.5E-08

2.0E-08

2.5E-08

3.0E-08

3.5E-08

4.0E-08

4.5E-08

5.0E-08

0 1000 2000 3000 4000 5000 6000 7000 8000t (s)

Cul

abil

e (M

)

Kinetic dataFitted Kinetic data

Log K kinetic

LT kinetic

Log k1 kinetic

0.0E+00

2.0E-08

4.0E-08

6.0E-08

8.0E-08

1.0E-07

1.2E-07

1.4E-07

6.97.17.37.57.77.9pCuT

LT

(mol

/L)

2

4

6

8

10

logk

1 &

logK

th

0.0E+00

2.0E-08

4.0E-08

6.0E-08

8.0E-08

1.0E-07

1.2E-07

1.4E-07

6.97.17.37.57.77.9pCuT

LT

(mol

/L)

2

4

6

8

10

logk

1 &

logK

th

Log K at Equilibrium

LT at Equilibrium

Average of Log K kinetic

Average of Log k1 kinetic

Average of LT kinetic

Good agreement between the constants obtained at equilibrium

and with the kinetic approach

Data obtained for only 1 sample: Example from Salinity 11, April 2007


4. Comparison of the kinetic and at Equilibrium approach

32

Time (s)0 2000 4000 6000 8000

Cu

lab

ile (

M)

0

1e-8

2e-8

3e-8

4e-8

pCuT6.06.57.07.58.0

pC

ula

bile

6.0

6.5

7.0

7.5

8.0

8.5

9.0

A B

k1

logKM Li at equilibrium9 10 11 12 13 14

log

KM

Li K

inet

ic9

10

11

12

13

14

LiT at equilibrium (nM)5 6 7 8 60 90 120 150

LiT

Kin

etic

(n

M)

5

6

7

8

60

90

120

150 A B


Good agreement between constants determined at equilibrium or kinetic

Apparent overestimation of Kinetically determined logK (or underestimation of the at equ. approach) due to:

• kinetic first point estimation of Culab at t0

• Is the solution at equilibrium with the at equ. Approach•Both approaches are complementary

At equ: Higher M/L ratio better [L] determination

Kinet.: more points after each addition less equilibrium dependent, kinetic parameters determined

4. Comparison of the kinetic and at Equilibrium approach

k-1

2007 (,) and 2008 (,).

33

Cu (%)0 10 20 80 90 100

LiT (nM)

5 20 50 20010 100D

epth

(m

)

-10

-8

-6

-4

-2

0

Salinity0 10 20 30 40

log Kiequ

8.5 9.5 10.5 11.5 12.5 13.5

A B C

Cu2+(M)1e-12 1e-11 1e-10

Dep

th (

m)

-10

-8

-6

-4

-2

0

strong (,) and weak (,) ligands, 2007 (,) and 2008 (,), In dotted line in inset: toxicity limit of 10 pM (Sunda et al., 1987).

• Complexing parameters results

Expected variation with salinity

Observed variation

Difference Autochthonous DNOM production in the estuaryHigher affinity for ligands from seawater originIn the FSI: Higher inorganic and free copper content (up to 20pM) in spite of [ligands] increase

Organic Cu

90 to 99%

83%


& In MINEQL:

34

CuT (nM)

0 2 4 6 8 10 12 14

Dep

th (

m)

-8

-6

-4

-2

0

Cu2+ (M)1e-12 1e-11 1e-10

Reaction time (min)0 20 40 100 200 300

A B C

2008 2008 2006 t50% t99%

2006

Omanović et al., 2006

• Simulation of DNOM reactivity under a Cu contamination

Used for prediction

•Higher [Metal] in summer due to traffic of touristic boats•Calculated free copper concentration potentially toxic for µorganisms at the surface in summer•Lower reactivity of the FSI DNOM•Compared to hydrodynamics variations tequ. are quite long system probably not at equilibrium in the estuary

≈ 2h30

≈ 4h30


time

35

• Comparison of the measured DNOM vs. the model DNOM

dep

th (

m)

dep

th (

m)

simulated from model marine DNOM

Determined with the simplified protocol

Higher free [Cu] with the use of model DNOM > toxicity limitBigger difference for marine sample ≠ DNOM behavior between Toulon & ŠibenikModel DNOM not sufficient, even if DNOM is from same origin This ≠ show the necessity to study samples representatives of the studied system


36

V. Conclusion


37

• NEW use of “3sec” for DNOM analysis remove SAS interferences

•Determined protocol NEW direct analysis of coastal natural samples at low [DOC] and [M] complexing parameters determination + NEW Kinetic parameters (reactivity prediction) model DNOM usable in environmental contaminant speciation/transport programs

•Standard DNOM hardly usable to model DNOM behavior of a complex environmentUse of the determined protocol for specific ecosystem understanding

•Main improvement needed: Voltammograms automatic fitting

•Deeper analysis of pseudopolarograms,

•On site measurements (DGT) and comparison of data

•Actually protocol applied on a depth profile from Dycomed (Dyfamed site) …


Conclusions and perspectives

38

[Cu]Free

1e-12 1e-11 1e-9 1e-8

Dep

th

-2000

-1500

-1000

-500

0

[Cu]Tot


Application of the method to an “oceanic” depth profile

First results shows:

•At natural [Cu]: Cufree under toxicity limit until simulated total [Cu] up to 5nM•Surface DNOM is less complexant•Still analyzing samples (Dycomed 15) and need to treat all kinetics data…• Need to make a connection with on site measurments (Chlorophyll, COT, fluorescence … )

5 nM

39

Martinska (Šibenik, Croatia)Balaguier Bay (Toulon, France)

Merci à tous de votre attention !Special thanks to my Directors:

Dr. Mounier S.Dr. Omanović D.

To the Jury’s members: Prof. Marmier N. Prof. Riso R.

D.R. Elbaz-Poulichet F. D.R. Cossa D. Dr. Garnier C.

And members from PROTEE (USTV) and LPCT (RBI) labs

Documents

Directeurs de Thèse : Dr. S. MOUNIER Université du Sud Toulon Var – PROTEE