Henri Sund

Yuan-Yuan Liao

Chantal Andraud

Alain Duperray

Alexei Grichine

Boris Le Guennic

François Riobé

Harri Takalo

Olivier Maury

or two-photon bioprobes

Keywords: Europium, Luminescence, Bioprobe, Two-photon microscopy, Bioimaging 10.1002/cphc.201800557

Polyanionic polydentate europium complexes as Ultrabright one-

Introduction

The design of lanthanide luminescent bioprobes (LLBs) for time resolved fluorimetry immunoassay and one or two photon (1P or 2P, respectively) fluorescence microscopy imaging applications is a very active field of research. [1] In this context, the main challenge is to fulfil simultaneously in a single complex a series of requirements: [2] i) the LLB must be kinetically, chemically and photo-chemically stable in complex biological media (culture media, blood, plasma…); (ii) the LLB has to possess a reactive functional group to allow further covalent labelling of a biomolecule but also hydrosolubilizing fragments avoiding nonspecific inter-actions; [3] (iii) the 1P-LLB excitation wavelength must be as high as possible, ideally above 330 nm (337 nm correspond to the N2 laser frequently used as source) to avoid the use of costly quartz optics and enabling 2P excitation in the 700-900 nm spectral range of the Ti:sapphire laser; (iv) the LLB must present the highest 1P-brigthness defined as the product of the molecular absorptivity by the luminescence quantum yield (B 1 = .), and/or 2P-brigthness (B 2 =   ) where ,   and  represent the molecular the molecular 1P extinction coefficient, 2P-cross-section and quantum yield, respectively. Consequently the structure of the chelating unit must be carefully chosen to rigorously exclude water molecules from the first coordination sphere and the chromophore antenna must be selected to simultaneously ensure an optimal energy transfer to the central lanthanide emitter and optimise the 1P-and 2Pabsorption. [4] During the last decade, charge transfer (CT) antennae became ubiquitous for the design of LLBs because they combined very large extinction coefficient and two-photon cross-section with low energy excitation wavelength due to a direct sensitization mechanism. [5] Using this strategy, extremely bright europium, terbium but also samarium, dysprosium and [a] M.Sc. Sund, Dr. H. Takalo, Radiometer Turku Oy, Biolinja 12, 20750 Turku, Finland. harrijuha.takalo@gmail.com [b] Dr. Accepted Manuscript ARTICLE ytterbium complexes based on the triazacyclononane (TACN) macrocycle have been reported for 1P-or 2P-bioimaging applications. [6] Considering europium, the best antennae are composed of a phenylethynylpyridine chromophore [7] further functionalized by mild electro-donating moieties like alcoxy, thioether or amido ones. [5] Such antennae have been grafted on various macrocycles like bis-bipyridine aza-macrocycle [EuL 1a ] + , [8] TACN [EuL 2b ], [6,9] cyclen [EuL 3b ] + [10] or pyclen [EuL 4b ] [11a] (Figure 1). In particular, excellent brightness has been obtained in water for [EuL 2b ] (B 1 (337) = 15-30000 L.mol - 1 .cm -1 ) and [EuL 4b ] (B 1 (336)= 12300 L.mol -1 .cm -1 ).

In this article, we describe the incorporation of such CT antennae in a polydendate ligand depicted in Figure 1. Two picolinic CT antennas are located at each extremity of the ligand whereas the central pyridine exhibits a thiourea functionality. This fragment comes from a isothiocyanate reactive function which was initially introduced for bioconjugation. To perform the photophysical study, the isothiocyanate was reacted with taurine. [12] The CT antennas are functionalized by one thioether or two alcoxy electrodonating fragments further decorated by carboxylate moieties to ensure hydrosolubility resulting in the formation of tetra, hexa and octa-anionic complexes [EuL 5c ] 4-, [EuL 5d ] 6-to [EuL 5e ] 8-respectively. A combined theoretical and photophysical study is presented to determine the symmetry of these complexes in solution; the linear and nonlinear two-photon absorption properties have been evaluated highlighting the excellent 1P-and 2P brightness of this family. Finally, their potential in 2P-microscopy imaging applications has been evaluated.

Results and Discussion

Synthesis. The synthesis of [Na]6[EuL 5d ] and [Na]8[EuL 5e ] has been previously described. [12] The preparation of [Na]4[EuL 5c ] required a similar procedure which is described in SI (Scheme S1).

Geometry optimization of the complexes. The tridimentionnal structure of this class of complexes remains unknown due to the impossibility to obtain monocrystals suitable for X-ray crystallography. This is the case for the -conjugated antenna functionalized derivatives [EuL 5c ] 4-, [EuL 5d ] 6-to [EuL 5e ] 8-but also for the non-functionalized parent complex [EuL 5 ] -. Consequently, the geometry of the complexes was investigated using density functional theory (DFT, see computational details in Experimental Section) on yttrium models [YL 5 ] -and [YL 5c ] -where the charged carboxylate and sulfonate end-groups were replaced with methyl fragments. The geometry optimisation of [YL 5 ] -indicate the formation of two isomers depending on the wrapping of the polydentate ligand around the central metal ion (Figure 2). In both cases the coordination number is nine and involves 5 nitrogen atoms coming from the central pyridine (Npy), the tertiary amine (2Na) and the picolinate (2Npa) -and 4 oxygen atoms form the acetate (2Oac) and picolinate (2Opa) moieties. However, the geometry of the two isomers differs significantly: for the isomer 1, the two picolinates are distributed on each side of a pseudo-equatorial plane formed by the central pyridine, the two tertiary amines and an additional carboxylate. The second acetate completes the coordination sphere. The second isomer differs by the inversion of the configuration of one of the tertiary amine. The pseudoequatorial plane includes this time one of the picolinate. Thus, the two acetate moities occupy one face of the complex while the second picolinate is in opposition regard to the metal center. The Shape analysis performed on both isomers indicates that resulting coordination polyhedra are close to a Cs symmetry point group (Table S1). Interestingly, isomer 1 was found 0.17 eV (3.98 kcal/mol) more stable than isomer 2. This result is not surprising since the steric hindrance between picolinate fragments is minimized in the case of isomer 1. Consequently, the geometry of the complex [YL 5c ] -was optimized only for Isomer 1. Interestingly, the wrapping of the ligand around the central metallic ion results in an overall apparent threefold symmetry of the -conjugated antenna (Figure 3). Absorption and emission spectroscopy. The photophysical characterization of the compounds has been performed in diluted water and Tris buffer solutions and the results are compiled in Table 1. The absorption spectra (Figure 4) are characteristic of charge transfer antennae [5] with a broad intense transition centred around 340 nm. A slight red shift is observed going from [EuL 5c ] 4-, [EuL 5d ] 6-to [EuL 5e ] 8-( = 6 and 5 nm 10.1002/cphc.201800557

Accepted Manuscript ARTICLE respectively) due to the presence of 1, 2 and 3 electron donor groups in the conjugated antennae reinforcing the charge transfer. Upon excitation in the absorption band, an intense red emission is observed with the characteristic Eu(III) profile assigned to 5 D0 → 7 FJ (J = 0 to 4) transitions (Figure 5). No ligand centred emission remains visible indicating that the sensitization process is very straightforward. Interestingly, due to the broadness of the absorption band (cut-off about 400 nm), it is possible to sensitize the Eu(III) luminescence in the visible range, corresponding to the working range of a wide-used LED (ex = 405 nm). The quantum yields in water and Tris buffer are high (20-25%) and the lifetimes are relatively long (0.7-0.85 ms) (Table 1) similar to other complexes featuring identical antennae but different structure like TACN [EuL 2b ] or Pyclen [EuL 4b ]. Consequently, the one-photon brightness of these complexes is very high 15-20 000 L. Mol -1 .cm -1 comparable to the best Eu(III) LLBs [6,9,11] and making them particularly attractive for further use in fluoroimmunoassay (with possible excitation at 337 nm) or bioimaging experiments.

 (L.mol -1 .cm -1
) Solution structure of the europium complexes. It is well known that europium complexes are particularly sensitive to their local environment and their emission spectra are clearly the signature of the complex symmetry in solution. In this context, two main effects can be explored: the crystal-field splitting (CFS) of each transition and their relative intensity, this latter being frequently used for sensing purposes (ie, titration of an analyte using transition intensity variations). The splitting of each 5 D0 → 7 FJ (J = 0 to 4) transition was studied on the basis of the low temperature (77K) emission spectrum recorded in an organic glass (Figure 6). The resolution is strongly improved due to the reduction of the vibronic broadening of the bands. Importantly, the 5 D0 → 7 F0 transition, insensitive to any CFS effect, can be nicely fitted with a single Gaussian curve indicating the presence of a single emissive species at 77K, in all probability the most stable isomer 1 found by the theoretical calculations. The J=1 and J=4 transitions are now clearly resolved and can be decomposed in three and nine Gaussian curves, respectively (Figure 6). In the hypersensitive J=2 transition the different contributions are merged in 3 main bands. Overall, the splitting of each transition in 2J+1 subcomponents is observed, signature of a low symmetry complex according to the CFS theory [13] and in agreement with the theoretical geometry optimization. In addition, the relative intensity of the 5 D0 → 7 FJ (J = 0 to 4) transition of the different complexes are rather similar (Table S2 and Figure S1). As example [EuL 5c ] 4-presents an emission profile with relative intensity of 0.1, 1, 10.9, 0.5 and 3.1 using the J=1 magnetic dipole transition as internal reference. [13] In order to envisage further applications of these complexes as FRET-donors it is important to minimize the intensity of the J=4 transition, that generally overlaps with the emission of the FRETacceptor dye. For convenience, we defined the r value as the ratio of the intensity of the J=2 over J=4 transition and this parameter is an indicator to compare the symmetry of different complexes. As a general tendency, highly symmetrical 10.1002/cphc.201800557

 [a]   (ms) B 1 [b] (L.mol -1 . cm -1 )  2 (GM)  2 =

Accepted Manuscript

ARTICLE

complexes exhibit very intense J=2 transition and weak J=4 ones hence high r value. [13] In the present cases the three complexes [EuL 5c ] 4-, [EuL 5d ] 6-and [EuL 5e ] 8-present similar r values comprised between 2.8 and 3.5 with an hypersensitive transition representing R 2 = 65-70% of the whole emission spectrum (Table 2). This value is lower than that of the TACN based complex [EuL 2b ] [6,9] (r = 10.9, R 2 = 79%) featuring an almost perfect threefold symmetry but higher than that of other macrocyclic compounds like cyclen [EuL 3b ] + (r = 2.5, R 2 = 56%) [10] or pyclen [EuL 4b ] [11] (r = 3.0, R 2 = 65%) featuring C2v and distorted C3 symmetry, respectively. 4-(black) in organic glass (MeOH/EtOH, 1/4, v/v) under 370 nm excitation with optimal resolution (detection slit 0.2, increment 0.2 nm). (Top) full spectra, in inset is represented the magnification of the J=0 transition emission spectrum with a single Gaussion deconvolution (red). (Middle) Magnification of the J=1 transition and decomposition in three Gaussian curves (dashed red, sum in red). (Bottom) Magnification of the J=4 transition and decomposition in nine Gaussian curves (dashed red, sum in red).

Two-photon absorption properties. The emission of [EuL 5c

] 4-, [EuL 5d ] 6-and [EuL 5e ] 8-is intense enough to measure the twophoton properties of these complexes using the two-photon excited fluorescence (TPEF) method in water. The set-up description is given in the experimental section. Upon 2P- ] 4-at room temperature in water with excitation at 345 and 810 nm, respectively. The spectral resolution difference is du to the different experimental setup (see experimental section for details).

a] r = IJ=2/IJ=4, R 2 = IJ=2/Itot; R 1 = IJ=1/Itot; krad = 1/rad = A(0,1).n 3 /R 1 where A(0,1)
is the spontaneous emission probability in vacuum determined to 14.65 s -1 and n is the refractive index of water equal to 1.333; Eu = /rad = krad/(krad+knr) 10.1002/cphc.201800557

Accepted Manuscript ARTICLE excitation at 810 nm, the europium emission is detected (Figure 7) and its profile looks similar to that obtained under classical 1P-excitation in spite of a lower spectral resolution due to the experimental set-up. Furthermore, the intensity of Eu(III) emission displays a quadratic variation with the incident laser power, unambiguously illustrating the 2P-antenna effect in water (Figure 8). Finally, the 2P-cross sections have been measured in water using coumarin-307 as external standard (Figure 9). Since the complex did not present any inversion center, it is not surprising to observe a perfect correspondence between the 2Pabsoprtion spectrum and the wavelength-doubled 1P-one (Figure 9). This result clearly indicated that the low lying CT state is responsible for the Eu(III) sensitization under 1P-and 2P excitation. The 2P-cross-sections of the three complexes ranged between 50-100 GM which are classical values for this type of antennae. [14] It is worth noting that, due to the spectral limitation of the incident laser source (700-900 nm), the 2P-maximum is not reached and only the red tail of the 2P-spectrum is observed. Nevertheless, one can notice that the 2P-cross-sections at 720 nm increase with the increase of the CT character of the antenna. .

Two-photon microscopy imaging. The complexes [EuL 5c ] 4-, and [EuL 5e ] 8-were then involved in 2P-bioimaging experiments using T24 cells (staining concentration of 5-10 10 -6 mol.L -1 ). The experiment was first performed with living cells (Figure 10). In our hands, it appears clearly that the two complexes did not cross the plasma membrane and remained in the culture medium, the cells appearing as dark areas. Such behaviour is typically observed for anionic complexes and is generally explained by an electrostatic repulsion with the negatively charged membrane surface especially inner leaflet of the lipid bilayer and glycocalix. [15] The cells were then fixed with paraformaldehyde (PFA) and treated with Triton X-100 to increase their permeability. Mild PFA fixation is known to preserve the native shape of biological structures keeping them hydrated. In this case, the complexes are internalized and fixed to some specific area in the cells. This is particularly visible for the [EuL 5e ] 8-complex that clearly accumulates in the nucleus and to less extent the endoplasmic reticulum and mitochondrial network (Figure 11). In particular the brightest spots in the nucleus correspond to nucleoli. This behavior has already been described for related tris anionic trisdipicolinate complexes. [16] The luminescence spectrum of [EuL 5e ] 8-in cellulo was monitored and presented the characteristic Eu(III) emission profile (Figure S2). Finally the luminescence lifetime was measured in fixed cells using the TSLIM method [17] and a rather short lifetime of ca 0.44 ms was found (Figure S3). This value is shorter than that found in water (0.74), in Tris buffer (0.70 ms) or after bioconjugation to IgG (0.69). [12] This lifetime decrease can be explained by an increased fluxionality of the complex in the biological environment containing a complex mixture of salts or biomacromolecules.

10.1002/cphc.201800557

Accepted Manuscript

ARTICLE

Conclusions

In conclusion, this family of europium (III) complexes based on polydentate CT functionalized ligand presents excellent 1P-and 2P-brigthness in water and buffer making them suitable for bioimaging experiments. A careful photophysical study suggests that the wrapping of the ligand around the central rare-earth ion results in an overall threefold symmetry with about 65-70% of the emission intensity concentrated in the hypersensitive 5 D0 → 7 F2 transition at 615 nm. These remarkable linear and nonlinear photophysical properties, combined with the presence of a reactive function in the central pyridine moieties make these complexes particularly attractive for further use as LLBs in fluoroimmunoassay.

Experimental Section

Photophysical measurements in solution. Absorption spectra were recorded on a JASCO V-650 spectrophotometer as solutions in spectrophotometric-grade methanol or water (ca. 10 -5 or 10 -6 mol.L -1 ). Emission spectra were measured using a Horiba-Jobin-Yvon Fluorolog-3 fluorimeter. Spectra were corrected for both excitation-source lightintensity variation and emission spectral responses. Luminescence lifetimes were obtained by pulsed excitation with a FL-1040 UP xenon lamp. Luminescence quantum yields, Φ, were measured with dilute solutions in water or organic solvents with an absorbance of less than 0.1 by using equation ( 1):

𝛷 𝑥 𝛷 𝑟 = 𝐴 𝑟 (𝜆) 𝐴 𝑥 (𝜆) 𝑛 𝑥 ² 𝑛 𝑟 ² 𝐷 𝑥 𝐷 𝑟 (1)
in which A is the absorbance at the excitation wavelength (λ), n is the refractive index, and D is the integrated luminescence intensity; r and x stand for reference and sample, respectively The reference is quinine bisulfate in a 1N aqueous solution of sulfuric acid (Φr = 0.546). Excitation of reference and sample compounds was performed at the same wavelength. Practically, for each sample, series of measurements are performed with different absorbance ranging from 0.1 to 0.01. The plot of the integrated luminescence intensity vs. absorbance gives straight line with excellent correlation coefficients and the slope S can be determined. Equation ( 1) becomes ( 2):

𝛷 𝑥 𝛷 𝑟 = 𝑆 𝑥 (𝜆) 𝑆 𝑟 (𝜆) 𝑛 𝑥 ² 𝑛 𝑟 ² (2)
Two-photon excited luminescence measurements. The TPA crosssection spectrum was obtained by up-conversion luminescence using a Ti:sapphire fs laser in the range 700-900 nm. The excitation beam (5 mm diameter) is focalized with a lens (focal length 10 cm) at the middle of the 10-mm cell. Emitted light was collected at 90° and was focused into an optical fiber (diameter 600 m) connected to an Ocean Optics S2000 spectrometer. The incident beam intensity was adjusted to 50 mW in order to ensure an intensity-squared dependence of the luminescence over the whole spectral range. The detector integration time was fixed to 1s. Calibration of the spectra was performed by comparison with the published 700-900 nm Coumarin-307 two-photon absorption spectrum [18] (luminescence quantum yield = 0.56 in ethanol). The measurements were done at room temperature in dichloromethane and at a concentration of 10 -4 M.

Cell culturing and treatment. We used the T24 human epithelial bladder cancer cell line (ATCC no. HBT-4). In our experiments, T24 cells were cultured in 25 cm 2 tissue-culture flasks (T25) at 37°C in a humidified atmosphere with 5% CO2. They were incubated in RPMI 1640 supplemented with 100 UmL -1 penicillin, 100 µg.mL -1 streptomycin, and 10% fetal calf serum (complete medium). Cells were grown to near confluence in the culture flasks and then suspended with a solution of 0.05% trypsin-ethylenediaminetetraacetic acid (EDTA; Sigma). Cells were placed on a LabTek I chambered cover glass (Nunc) at low cell density in complete culture medium 24 h before experiments. After being washed with phosphate-buffered saline (PBS), cells were fixed with PFA (3% in PBS) for 10 min, permeabilized with PBS containing 0.5% Triton X100 for 10 min, and then washed with PBS. The staining is performed by addition of a concentrated solution of complex (10 -4 mol.L -1 ) in PBS buffer to the cell culture medium to reach a final concentration of 5-10 10 -6 mol.L -1 .

ARTICLE

Entry for the

Figure 2 .

 2 Figure 2. Representation of the DFT optimized geometry of the two isomers of [YL 5 ] -. Donor atoms belonging to the pseudo-equatorial plane represented in light colours with black bonds to the Yttrium (light blue). Other coordination bonds are depicted in green.

Figure 3 .

 3 Figure 3. Representation of the DFT optimized geometry of the isomers 1 of [YL 5c ] -.

Figure 5 .

 5 Figure 5. Representative emission spectrum of [EuL 5c ] 4-(black), in water

Figure 6 .

 6 Figure 6. 77K emission spectra of [EuL 5c ]4-(black) in organic glass (MeOH/EtOH, 1/4, v/v) under 370 nm excitation with optimal resolution (detection slit 0.2, increment 0.2 nm). (Top) full spectra, in inset is represented the magnification of the J=0 transition emission spectrum with a single Gaussion deconvolution (red). (Middle) Magnification of the J=1 transition and decomposition in three Gaussian curves (dashed red, sum in red). (Bottom) Magnification of the J=4 transition and decomposition in nine Gaussian curves (dashed red, sum in red).

Figure 7 .

 7 Figure 7. 1P-(red) and 2P-(blue) excited luminescence spectra of [EuL 5c

Figure 8 .

 8 Figure 8. Variation of the Eu(III) luminescence intensity with the laser excitation of [EuL 5c ] 4-in water (2P-excitation ex = 760 nm, •). The red line represents the fitting of the experimental points by the polynomial function indicated in inset.

Figure 10 .Figure 11 .

 1011 Figure 10. 2P imaging of living T24-cells stained with [EuL 5c ] 4-(b) and [EuL 5e ] 8-(d). (λexc = 780 nm). In (a) and (c) the corresponding transmitted light DIC images are represented. Complexes are added in the cell medium in phosphate buffer solution to reach a final concentration of 5-10 10 -6 mol.L -1.

  H. Sund, Y.-Y. Liao, C. Andraud, A. Duperray, A. Grichine, B. Le Guennic, F. Riobé, H. Takalo*, O. Maury* Page No. -Page No. Polyanionic polydentate europium complexes as bright one-or twophoton bioprobe 10.1002/cphc.201800557 Accepted Manuscript

Table 1 .

 1 Photophysical Data of europium complexes in water or Tris buffer solutions at room temperature.

max (nm)

Table 2 .

 2 Photophysical parameters determined from the complexes room temperature emission spectra in water.

				[a]
		r	R (%) 2	R (%) 1	rad (ms)	Eu	krad (s -1 )	knr (s -1 )
	[EuL 5c ] 4-	3.5	69.8	0.06	1.99	0.43	502.9	673.6
	[EuL 5d ] 6-	2.8	65.5	0.07	2.13	0.40	468.7	694.1
	[EuL 5e ] 8-	3.5	69.6	0.06	2.01	0.37	498.7	852.6
	[

  Table of Contents (Please choose one layout)

	Layout 1:
	ARTICLE
	A family of europium (III) complexes
	based	on		polydentate	ligand
	functionalized by charge transfer
	antennae presents remarkable one
	and two photon photophysical
	properties	in	water	excellent
	brightness suitable for bio-imaging
	applications.

Acknowledgements

B.L.G. thanks the French GENCI-CINES centre for high-performance computing resources. Confocal and 2P facility of the IAB platform was co-funded thanks to grants of "Association pour la Recherche sur le Cancer", Ligue Contre le Cancer, and the CPER program.

Confocal and two-photon microscopy. All confocal experiments were performed using a LSM710 NLO (Carl Zeiss) confocal laser scanning microscope based on the inverted motorized stand (AxioObserver, Zeiss). The excitation was provided by either 561 nm DPSS cw laser or Ti:Sa femtosecond tuneable laser (Chameleon, Ultra II, Coherent) for 2P excitation at 750 nm in descanned detection mode. In the former case the pinhole was closed to 1 Airy Unit and in the latter one it was fully open. Spectral imaging was realized using an internal Quasar detector in the range 577-723 nm with the resolution of 9.7 nm.

Computational details. DFT geometry optimizations were carried out with the Gaussian 09 (Revision D.01) [19] package employing the PBE0 hybrid functional [20] and tightening self-consistent field convergence thresholds (10-10 a.u.) and geometry optimization (10-5 a.u.) convergence thresholds. The "Stuttgart/Dresden" basis sets and effective core potentials were used to describe the yttrium atom, [21] whereas all other atoms were described with the SVP basis sets. [22] The analysis of the calculated symmetry was performed with the Shape program. [23]


Sylvie Lhospitalier

Philippe Bourges

Alexandre Bert

Jean Quesada

M Lambertin

Temperature measurement inside and near the weld pool during laser welding

Keywords: CO 2 laser welding, temperature measurement, austenitic stainless steel

come

I. INTRODUCTION

To better understand thermomechanical phenomena that occur during laser welding, it is necessary to know the temperatures inside the weld pool and near it. Many numerical simulations can be found in the literature, [START_REF] Metzbower | On the temperature and size of the keyhole[END_REF][START_REF] Strauss | Determination of temperature distribution in laser welded heat exchangers with the aid of finite element method[END_REF][START_REF] Kou | Simulation of heat flow during the welding of thin plates[END_REF] but little experimental data concerning thermal cycles inside the weld pool are available. The great difficulty of such experiments is the very high temperatures that are reached by the treated material, especially steels. In this article, we describe the experimental method used to determine the thermal field around laser impact during welding of austenitic stainless steel. The study is conducted on 8 mm thick sheets. Two characterization methods have been used. The temperature inside the material is determined with thermocouples, and that on its surface is determined by use of a charge coupled device ͑CCD͒ camera equipped with infrared ͑IR͒ filters. The results obtained by these two methods may be used as a database for numerical simulation.

II. PRESENTATION OF THE EXPERIMENTAL PARAMETERS

The material used is austenitic stainless steel UNS N08904 ͑CLI URB6™͒, which contains low phosphorus and sulfur levels. The actual composition is given in Table I.

The sample size was chosen so that the temperature at the border remains approximately constant ͑i.e., room tem-perature͒. To control this, eight thermocouples were placed along the sample, and the border temperatures during welding were recorded. These measurements gave us a sample size of 215 mmϫ80 mm. Moreover, to reproduce the mechanical behavior of a massive piece, the samples were bridled on a steel block with six screws. Finally, to avoid gap problems between the two sheets, fusion lines ͑keyhole welds͒ were realized, instead of real welding of the two separate sheets ͑Fig. 1͒.

III. THERMAL MEASUREMENTS WITH THERMOCOUPLES

The temperatures measured ranged between 1000 and 1800 °C ͑the melting temperature of UNS N08904 is 1380 °C͒. That is why tungsten ͑W5 type͒ thermocouples were used. They have a good resistance to high temperatures, and their small diameter ͑0.2 mm͒ results in a small temperature lag time. The diameter of the entire thermocouple was 1.2 mm ͑including the tungsten wires and sleeving͒. The thermocouples were placed inside the samples, 4 mm under the surface, into 1.4 mm diam holes made by electroerosion, inclined 20°from horizontal to disturb the thermal fields as little as possible ͑Fig. 2͒. Eight samples were machined, then two thermal cycles were characterized ͑four measurements for each thermal cycle͒. Because thermal cycles are very short during laser welding, it was necessary to choose a sufficiently high acquisition rate. The maximum rate authorized by the system used is 100 measurements per second providing that there is only one thermocouple at a time working. Then, we had to put one thermocouple per sample in place and perform as many weldings as necessary. The process parameters chosen for the two thermal cycles were very close: 5 kW, 0.5 m/min and 6 kW, 0.5 m/min. The results obtained are similar for these two cycles, that is why we shall present just the latter one. The widths of the melted zones are approximatively 7 mm on the surface of the sheet and 2 mm inside the material ͑Fig. 3͒.

The curves in Fig. 4 show the evolution of the thermal gradients inside the samples with time. Time 0 indicates that the laser beam is situated on a given point ͑the temperature maximum in the center of the weld͒, and abscissa 0 represents the center of the weld ͑the X axis being perpendicular to the joint͒.

Upon arrival of the laser beam, there is a sudden rise in temperature in the center of the weld, whereas the rest of the sample remains cold. Very quickly, heat is transmitted by conduction to the rest of the sheet. The result is that the zones most distant from the center of the weld start to heat up while the nearest zones start to cool down. This is shown by the progressive slope of the curves. At the end, these curves become parallel, and the entire sample cools down. This phenomenon, so called differential heating, is very important in explaining deformations during welding. [START_REF] Lhospitalier | Approche thermome ´canique de la fissuration a `chaud dans un acier inoxydable auste ´nitique; cas du soudage laser[END_REF] Figure 5 gives a different concept of heating differences according to the distance of the point measured from the center of the weld. For instance, points which are 2 mm from the center of the weld receive maximum thermal flux 5 s after the beginning of the recording.

IV. SURFACE TEMPERATURE MEASUREMENTS WITH AN IR-CCD CAMERA

In order to complete these observations, we have measured surface temperatures using a CCD camera equipped with infrared filters. Two additional cycles were completed: 5 kW, 0.4 m/min and 6 kW, 0.5 m/min.

Infrared imagery consists of recording an image of light intensity emitted by a fixed wavelength target in the infrared domain. Usually this method is conducted with thermal cameras equipped with light intensity sensors sensitive to wavelengths that are often far ͑several microns͒ from the visible. The construction of infrared cameras is complex, and special materials are needed for optical sensors, an optomechanical scanning system ͑used to read images͒, temperature balance of the equalization system ͑used to avoid measure drift due to camera heating͒, rigorous calibration etc. [START_REF] Pajani | Thermographie infrarouge[END_REF][START_REF] Pajani | Mesure par thermographie infrarouge[END_REF] This method is not very easy to use.

To perform our experiments, a classical CCD camera was transformed into a pseudothermal camera. The CCD sensor of the camera has to be sensitive to infrared radiation ͑usually the nearest infrared͒. This related technique permits an initial approach to temperatures inside the weld pool and near it with sufficient accuracy for our purpose.

The camera chosen has good features in the infrared domain ͑seen in Fig. 6͒. CCD classical cameras are sold with a ''cold filter,'' which ''cuts'' the infrared radiation for common application or use. In our case, it is necessary to take this away. The interaction between the laser beam and the treated material gives high intensity radiation because of the melted pool and the surface plasma. It is necessary to do a filtration of the major part of this radiation. It is known that plasma radiation takes place preferentially in low wavelengths and in the ultraviolet zone. [START_REF] Sokolovsky | Spectral plasma diagnostics in laser welding with CO 2 lasers[END_REF] So it is interesting to observe the weld pool in the infrared domain since the residual plasma radiation will be lower. However, the experimentation wavelength bandwidth is limited by the camera sensor technology. Figure 6 shows that total infrared radiation is cut at 1200 nm. The working wavelength is chosen as 1100 nm. To avoid the camera sensor from being saturated with light, and in order to be able to consider the material as a graybody, the wavelength bandwidth must be very narrow. The previous conditions imposed the use of an interferential filter, 1100 nm centered and 10 nm width. It is a standard filter that is a good compromise between cost and technical needs. And it is useful to indicate a problem due to the filter. Because of the technical properties of the filter, on the recorded images we can observe interference rings, which exist for a large range of optical adjustments. The result is that the image obtained is an overlap of the original image and the interference rings ͑see Fig. 7͒. In order to solve this optical problem, it is necessary to increase the size of the lens aper-ture of the camera, bring the camera as close as possible from the measured point, and increase the wavelength bandwidth of the filter used.

In practice, none of these solutions was applicable to the experiment because of the space required on the laser machine and because of an increase in the total installation cost. The remaining solution is a posttreatment computing, which will be explained later.

Analysis of infrared radiation is dependent on the emissivity of the material. The material's emissivity is related to numerous parameters: direction, surface quality, physicochemical state, temperature etc. [START_REF] Ujihara | Reflectivity of metal at high temperatures[END_REF][START_REF] Herve | Influence de l'e ´tat de surface sur le rayonnement thermique des mate ´riaux solides[END_REF][START_REF] Chapman | Heat Transfer[END_REF] First, it is necessary to choose a good angle of observation, taking into consideration spectral emissivity, minimization of geometrical aberrations, and the space required for the entire device. The maximum observation angle obtained was 70°. Gaseous protection is realized with helium in a glove box. Images are transmitted to the camera by two convergent standard BK7 lenses ͑Fig. 8͒. Braces are placed between the objective and the camera's CCD sensor to correctly magnify the zone being observed. But, there was a big problem: vibrations of the machine tool. To eliminate them, it was necessary to put the entire optical device on an antivibrating carpet.

Images are stored using a video recorder ͑25 images per second͒. This is a sufficient speed in this case because the welding speed is too low ͑0.5 m/min͒ as are the temperature changes associated with it. ''Key'' images are then located precisely and digitalized before analyzing.

Before analyzing the images, it is necessary to do a calibration of the entire optical device, i.e., the camera ϩ the interferential filter ϩ the lenses associated to the test configuration ͑observation angle, distance to the sample, etc.͒ in order to relate the gray levels to the surface temperatures. Several methods are possible depending on the temperature range. In our case, we used direct calibration, which means that the variable emissivity and other phenomena like plasma are directly integrated. To do that, a sufficient surface area of the material is uniformally heated with a defocused laser beam; the image ͑gray level͒ is recorded with the same optical device as that described previously, while the temperature is measured simultaneously with a thermocouple placed just under the heated surface ͑Fig. 9͒. A comparison of the two results ͑thermocouple data and recorded images͒ allows one to relate the gray level recorded to a given temperature. This test is repeated for several temperatures, and it gives a calibration curve ͑Fig. 10͒. The great advantage of this calibration technique is that the emissivity and other unnecessary effects ͑like plasma radiation, for example͒ are totally integrated into the measurements. It is then certain that a given gray level is associated to the real temperature.

It appears that the CCD camera sensor equipped with the infrared filter was sensitive in the 1000-1800 °C temperature range. According to the shape of the curve ͑Fig. 10͒, it appears that if the gray level error is considered as being constant, the error in the corresponding temperature may vary, and is more important in the curve extremities. The major source of error concerning the real values of the temperature is the variation of the emissivity of the material during welding ͑the steel becomes brighter after solidification of the melted zone͒. According to data in the literature, [START_REF] Huetz-Aubert | Rayonnement thermique des mate ´riaux opaques[END_REF] the emissivity of rough stainless steel can be estimated as approximately 0.7 and its emissivity when polished is approximately 0.5. A rapid calculation using blackbody and graybody laws reveals that the error made in the temperatures is less than 8% at 3000 °C ͑2% at 1800 °C͒. According to the experimental conditions, a final maximum error of 10% can reasonably be expected.

The problem of interference rings observed during the experiments is solved by averaging the gray levels for a given temperature range. An algorithm was developed, which associates a temperature range to a gray level range according to the calibration curve. Using the algorithm, images in the gray levels are automatically converted into thermal cards showing isothermal domains. Figure 11͑a͒ shows an example of a gray level primary image, and Fig. 11͑b͒ shows the same image after computing the analysis.

After welding, the image of the joint was recorded, and this allows one to determine the limit of the melted zone ͓Fig. 12͑b͔͒. When compared with the temperature distribution, it can be observed that the temperature range at that limit ͑1300-1405 °C͒ is in quite good agreement with the melting point of the steel ͑1380 °C͒.

The maximum temperatures detected are higher than 1770 °C. A saturated zone appears ͑Ͼ1770 °C͒, which probably corresponds to the keyhole location.

One can also observe in the images weld pool asymmetry, which can easily be explained by plasma displacement due to protection gas. Moreover, for temperatures below the melting temperature, there seems to be an optical aberration: isothermal lines are distorted, which could be due to the radiation induced by laser-material interaction on the surface of the steel. But there is also another explanation of this phenomenon: as previously mentioned, the plasma is blown by the protection gas, and this can create additional heating of the zone close to the keyhole.

V. CONCLUSION

Four different thermal cycles that occur during welding have been discussed in this study. The method which was used gives satisfactory results: good sensitivity to experimental parameters was exhibited. Many differences can be revealed between very close thermal cycles. For instance, the weld pool is wider for a 6 kW 0.5 m/min cycle than for a 6 kW 0.7 m/min one. So this experimental method is well adapted to characterize thermal surface phenomena inside and near the weld pool. The temperatures reached are difficult to access by any other method. The most original part of this work is the camera sensor calibration, which takes the total environment ͑emissivity, plasma interference, operating conditions, etc.͒ into account. In order to do so our technique needs one calibration per material and welding configuration.

A great number of images have been realized, and they will allow us to calibrate a finite element simulation of laser welding. Moreover, similar experiments will be soon conducted on other materials ͑duplex stainless steels, for in-stance͒ and for other welding processes ͑particularly GTAW͒.

FIG. 1 .

 1 FIG. 1. Geometry of the samples.

FIG. 5 .FIG. 6 .

 56 FIG. 5. Thermal cycles of different points inside the samples.

FIG. 8 .

 8 FIG. 8. Schematic of the experimental device.

TABLE I .

 I Composition of the stainless steel used.

	Elements C	Mn Si	S	P	Ni	Cr Mo	N	Cu
	wt % 0.01 1.461 0.2 0.001 0.025 24.78 19.95 4.11 0.082 1.357

This article is copyrighted as indicated in the article. Reuse of AIP content is subject to the terms at: http://scitation.aip.org/termsconditions. Downloaded to IP: 193.50.222.52 On: Wed, 05 Feb 2014 15:09:23

ACKNOWLEDGMENTS

The authors are very grateful to R. M'Saoubi, J. Outeiro ͑LM3, ENSAM Paris͒, C. Le Calvez and M. Gauthier ͑CLI͒ for their contribution to this work, and the Regional Council of Burgundy for its financial support.


Florent Colombet
email: florent.colombet-renexter@renault.com

Zhou Fang
email: zhou.fang@renault.com

Andras Kemeny
email: andras.kemeny@renault.com

Pitch tilt rendering for an 8-DOF driving simulator

Keywords: Driving simulation, Motion cueing algorithm, Tilt coordination, Pitch acceleration threshold, Vestibular system

Tilt coordination technique is used for reproducing a sustained linear acceleration in driving simulation by tilting the simulator cabin. Thus a part of gravity is interpreted as a horizontal acceleration. Between the slow rotational motion limited by the rotational perception threshold of semi-circular canals of vestibular system and the fast change of acceleration to render, the design of coupling motions of rotation and translation plays a critical role in the realism of driving simulator. We present here an experiment focusing on longitudinal accelerations rendering with pitch tilt coordination technique. Eight drivers experienced the same accelerations and decelerations on a straight double-lane motorway. During the driving, we varied the pitch rate (4, 5, 6 and 7 °/s) and the pitch acceleration (8 and 30 °/s²) used to render these accelerations with the tilt coordination technique. We have found an acceptability threshold of about 6°/s, similar to the value found in our previous work on roll tilt rendering. We have also shown a strong role of pitch rotational acceleration on acceptability, involving that it should be taken into account to properly render accelerations with the tilt coordination technique.

Introduction

The role played in automotive industry by driving simulators is increasingly important. During the design phase, they allow testing new advanced driver assistance systems (ADAS) such as ACC, AEBA, EPS, LKA etc. by studying driver's behaviour. They also allow testing the car's handling, ride comfort, drivability, behaviour, performance or fuel consumption without having to build a physical prototype. In order to have a driver's behaviour as close as possible as the one he/she could have in a real driving environment, driving simulators should provide the driver with the necessary cues. Especially for tasks like accelerating and braking, which are one of the most common basic tasks a driver can perform, driving simulators should provide the driver with inertial cues allowing him/her to regulate his/her action [Kem03]. For ADAS and AV (Autonomous Vehicle) experimentations at Renault VR and Immersive Simulation Center, being able to perform accurate tests on a driving simulator is a crucial point, because of the large number of interests depending on it in terms of cost and delay reduction. Unfortunately, the level of longitudinal acceleration rendered in simulation during accelerations and decelerations is sometimes not strong or long enough according to professional test pilots who interpret the driving simulator feeling tightly connected to the driving commands, including the vehicle speed. Actually, the available effective X-Y rails strokes (5.2 meters) of the ULTIMATE simulator appear to not be enough to render the needed acceleration level during the complete duration of accelerations or brakings for some of the most demanding vehicle applications. This is why we are focusing our research on the implementation of the tilt coordination technique in the motion cueing algorithm of the ULTIMATE simulator. After presenting the tilt coordination technique and the difficulties generally encountered when using it in driving simulation, we will see that some thresholds needed for its implementation on a motion cueing algorithm remain unclear. Our experiment aims at comparing eight parameters configurations in terms of rotational rate and acceleration to see the acceptance of drivers and then determine what acceleration levels can be reached with the tilt coordination technique.

Tilt coordination technique

Human body perceives accelerations among others thanks to the inner ear [Gra06] which is composed of two parts. The first, called the otholitic system, is sensitive to the linear accelerations while the second, called the semi-circular canals, is sensitive to angular accelerations. Nevertheless the otholitic system presents a perception ambiguity: it cannot differentiate a horizontal acceleration from an inclination around a horizontal axis because of the horizontal component of gravity. In driving simulation, this gravito-inertial equivalence is thus used in motion cueing strategies to render a part of the vehicle accelerations. It is known as tilt coordination technique (see Figure 1). However the inclination during the tilt coordination technique has to be done under certain conditions to remain unnoticed by the driver. First the driver must not be able to visually perceive the tilting.

Visual rendering has then to be compensated if the display screen is not fixed to the cabin. Secondly, the rotation has to remain under the semi-circular canals detection thresholds. Knowledge about rotational motion perception appears thus as primordial for the use of the tilt coordination technique, especially the detection thresholds. According to the particular dynamics of canals' model, Guedry [Gue74] has traced a relationship between the rotational acceleration and the time exposed to the excitation.

Motion cueing algorithm

Renault ULTIMATE simulator

We intend to conduct our experiment on the highperformance dynamic ULTIMATE simulator [Dag06] at Renault Virtual Reality and Immersive Simulation Centre (VRISC) (Fig. 2). First developed in 2001, this simulator has been renewed in 2011 [Sch12] and consists now of a closed cabin based on a Renault Twingo 2 car which has been lightened and instrumented. Inside the cab, transmission is carried out using a manual gearbox, and a software of sound synthesis is used to reproduce engine noise and the audio environment for an interactive vehicle. Sound is rendered thanks to active noisecancelling headphones Bose QuietComfort 25. The cabin is mounted on a large X-Y table and a hexapod motion system to render physical accelerations and rotations. Table 1 presents the physical capabilities of the motion system.

MPC-based motion cueing algorithm

The motion cueing algorithm is in charge of computing the physical displacements of the simulator cabin as a function of the simulated vehicle motion. It has to realize a compromise between rendering the vehicle accelerations (in terms of driver perception) and keeping the simulator within its physical limits. The algorithm used on the ULTIMATE simulator is a MPC-based (Model Predictive Control) motion cueing algorithm as described by Fang [Fan14]. Compared with classical or LQR optimal filters' approaches, the MPC integrates directly the system constraints into its optimization process, and then gives a real optimal solution and hardly needs the tuning process to check the workspace limits and the driver's perception thresholds.

In the motion cueing process, acceleration rendering with the tilt coordination technique has been added and is performed as a priority. The equivalent acceleration thus rendered is then subtracted from the vehicle acceleration before being rendered with the rails. Tilt rotation thresholds (in terms of amplitude, rate and acceleration) are explicitly taken into account in the optimization process of the algorithm. Different configuration sets can be used and the possibility to switch online from one to another has been implemented. In this case, a transition phase between the two configurations is performed during 5 seconds.

We can also specify that rotation motions are rendered around the driver's head centre. Both vehicle and tilt coordination rotations are computed around this particular point. Specific modules are in charge of realizing the change of coordinates from the rotation point of the hexapod to the driver's head by adding linear motions (on the hexapod and not on the XY rails).

Experimental Protocol

General Purpose

We aim at determining the acceptable pitch tilt coordination parameters for longitudinal acceleration and deceleration rendering. Table 2 gives the details of the 8 compared tilt configurations in terms of maximum rotational rate and acceleration. Maximum tilt angle is 8 ° for every configuration. We varied only the maximum tilt rate from 4 to 7 °/s and the maximum tilt acceleration from 8 to 30 °/s². Task description

After starting the vehicle and setting up the vehicle speed at 60 km/h, subjects were asked to perform longitudinal accelerations and decelerations respectively from 60 to 100 km/h and from 100 to 60 km/h. These accelerations and decelerations are long enough (nearly 8 seconds) so they cannot be fully rendered with rails only. Thus the use of pitch tilt coordination rendering does make sense in these cases.

On one hand, we wanted the driver to be as much active as possible, while having an active driving has an impact on driver's perception threshold [Pre14]. On the other hand, allowing the driver to behave freely can induce a bias while comparing the different tilt configurations. The solution we used was to perform pre-scripted accelerations and decelerations when the driver pressed the gas pedal or the brake pedal. Thus the driver was free to accelerate or brake when he wanted but accelerations and decelerations had the same duration and intensity so the comparison of the different tilt configurations was not biased. Figure 3 presents an example of the crenel of speed which was repeated during the experiment. Acceleration and deceleration last 8 seconds and 15 seconds are left between in order to let enough time to the platform to preposition itself in the simulation room. For the same reason during experiment and also to let enough time to switch motion cueing configuration, speed crenels were spaced between them of at least 15 seconds. In order to help drivers to keep a constant speed between accelerations and decelerations, a modified cruise control has been implemented in the vehicle. Its working is quite simple: it maintains the current vehicle speed when the driver releases gas and brake pedals.

Road description

The road used for this experiment was a straight portion of a double-lane motorway (see Figure 4). This portion is visually realistic and there was no traffic. Orange cones were dispatched on the road so the drivers knew where and when to accelerate and to brake.

Protocol

After presenting them the simulator and the purpose of the experiment, subjects were proposed to perform a familiarization driving in which they could perform 4 speed crenels with alternatively no tilt (rails only) and tilt rendering.

For the experiment, there were 16 speed crenels to perform in total. The crenels were paired (1A/1B, 2A/2B, 3A/3B, ...). For each "A" crenel, no tilt coordination was done: accelerations/decelerations were rendered with rails only. For each "B" crenel, one of the 8 configurations (see Table 2) was used for tilt rendering.

Subjects were asked to accelerate/brake when they reached a pair of orange cones as explained in previous subsection. These cones were spaced in order to let enough time to the simulator cabin to preposition itself in the simulation room (nearly 15 seconds) and also to the motion cueing algorithm to switch configuration. They were disposed as shown in Figure 5.

Questions asked

After each pair of speed crenels, three questions were asked to the drivers. They had nearly 1 minute to answer these three questions before reaching the next pair of orange cones and beginning the following pair of speed crenels (see Figure 5). 1. "During accelerations/decelerations, how would you quote the visual stability of virtual environment?"  Quote between 1 (perfectly stable) and 6 (not stable at all). 2. "During accelerations/decelerations, did you experienced ocular discomfort?"  Quote between 1 (no discomfort) and 6 (strong discomfort). 3. "How would you quote the acceptability of the acceleration/deceleration rendering with tilt coordination?"  Quote between 1 (not acceptable at all) and 6 (perfectly acceptable).

Subjects and configurations order

Eight volunteer expert subjects have participated to the experiment. Table 3 presents the configurations order for the 8 participants. We used a counterbalanced diagram latin square in order to avoid rank effects (a given configuration is only once in a particular position) and report effects (any configuration is followed or preceded only once by each of the 7 other configurations).

Results

Acceleration rendered profiles

Figure 6 presents a comparison of longitudinal acceleration rendering between using rails only (no tilt) and using tilt coordination technique (configuration 5: 6 °/s and 8 °/s²). The second graph of this figure shows the corresponding longitudinal displacement of the cabin in the simulation room.

Quotation results

Tables 4 to 6 present the three quotation results: visual stability of the virtual environment, ocular discomfort and overall acceptability of the tilt configuration. Table 7 presents the "binarized" quotation of acceptability: "0" if subject quoted "1", "2" or "3" and "1" if subject quoted "4", "5" or "6". These "binarized" data are plotted on Figure 7, with a reminder of the tilt parameters to which they correspond.

# Subject 1 1 1 3 1 1 1 3 3 2 2 1 1 2 1 3 1 1 3 1 1 2 2 2 3 2 3 4 4 4 5 5 5 4 4 4 5 2 2 2 2 2 2 2 2 6 2 3 2 2 2 2 3 4 7 1 1 1 1 1 2 3 3 8 2 2 2 3 2 3 4 3 Mean
1.9 1.9 2.3 2.3 2 2.5 2.8 2.9

# Subject 1 1 1 1 1 1 1 1 1 2 1 0 1 0 1 0 0 1 3 1 1 1 1 0 0 0 0 4 0 1 0 1 1 0 1 0 5 1 1 1 1 1 1 1 0 6 1 1 1 1 1 1 1 0 7 1 1 1 0 0 0 0 0 8

Discussion

For the accelerations and decelerations we wanted to render in this experiment, we see on Figure 6 the effect of the tilt coordination technique. On this figure, the first acceleration and deceleration are rendered with rails only (no tilt) and it appears clearly that nothing is rendered after nearly 2 seconds because the longitudinal displacement of the cabin has reached the maximum possible excursion on the rails. On the other acceleration and deceleration showed in figure 6, the tilt coordination technique is used and we see that it allows rendering a sustained longitudinal acceleration / deceleration during all the virtual vehicle acceleration / deceleration. However, the drawback is that it needs to preposition the cabin in order to benefit from the maximum possible rail stroke. In fact the motion cueing algorithm uses the time during which the cabin accelerates with the rails to slowly tilt the cabin under the determined pitch rate and acceleration thresholds. The quotation results (Tables 4 to Table 6) are also interesting. First we can see that their results are congruent: when pitch rate and acceleration are increased, the visual stability quotation decreases (drivers find the visual environment less stable), the ocular discomfort quotation increases, and the acceptability quotation decreases. An 2 way ANOVA analysis shows that the effect of rotational speed on the level of acceptability is significant (F(2,14) = 5.58, p=0.017). Concerning the level of acceptability, results presented on Table 6 shows that we can split our configurations into 2 groups: configurations 1 to 5 with a mean acceptance level beyond 4, and configurations 6 to 8 with a mean acceptance level near 3. It leads us to think that the maximum acceptable pitch rate is 6 °/s if the pitch acceleration is 8 °/s². For this particular rate, there is a significant difference of mean acceptance between 8 °/s² (cfg #5) and 30 °/s² (cfg #6). A 2 way ANOVA analysis on the data of configurations 3 to 8 (pitch rate >= 5 °/s) shows that the impact of rotational acceleration is significant (F(1,7) = 9,33, p=0.018). This impact of rotational acceleration on the driver's acceptability of the tilt coordination technique is more visible with the "binarized" data (Table 7 and Figure 7). Once again we can split configurations in the 2 same groups: configurations 1 to 5 with a mean acceptance greater than 60 % and configurations 6 to 8 with a mean acceptance below 50 %. In addition to showing the decrease of mean acceptance when the pitch rate increases, Figure 7 shows also the impact of rotational acceleration. For every pitch rate greater than 4 °/s, the mean acceptance level is 30 % lower with a rotational acceleration of 30 °/s² than with a rotational acceleration of 8 °/s². As semi-circular canals are sensitive to accelerations, this can be easily understandable. As for the configurations 1 and 2 (4 °/s), we can explain the equivalent result between the 2 levels of rotational acceleration by the fact that these rotational accelerations are very quick with this rate. The rotational speed of 4 °/s is quickly reached and as Guedry [Gue74] showed, the quicker the stimulus, the higher the threshold. Finally, quotation of the visual stability of the environment reveals also an interesting fact. Some of the drivers have reported that during accelerations and decelerations they felt like the road is going respectively up and down. This feeling was the main cause of their quotation. It can be explained with the vestibulo-occular reflex, activated by the tilting motion and trying to visually compensate an image that doesn't need to and inducing the previously described effect.

Conclusion

We have presented in this paper an experiment aiming at determining the pitch rate and acceleration thresholds for the acceptability of the tilt coordination technique. We compared 8 configurations, resulting of the combination of 4 rotational rates (4, 5, 6 and 7 °/s) and 2 rotational accelerations (8 and 30 °/s²). We have found an acceptability rate threshold of 6 °/s, close to the value of 5 °/s we found on our previous work on roll tilt rendering. We also found a strong influence of rotational acceleration on the driver's acceptability if the rotational rate used is greater than 5 °/s. Although most studies focus on rotational rate, we've found that, for the same rotational speed, having a rotational acceleration of 30 °/s² instead of 8 °/s² decreased the acceptability of at least 30%. We thus recommend taking this acceleration value into account into motion cueing algorithm using the tilt coordination technique. These pitch tilt thresholds of 6 °/s and 8 °/s² obtained as acceptable values for 6 out of our 8 subjects will be used with Renault's 8-DOF Ultimate high performance driving simulator for a more efficient use of the available workspace for forthcoming industrial applications using driving simulation.

Fig. 1 .

 1 Fig. 1. Illustration of the tilt coordination technique. By tilting the simulator cabin of an angle , the component g*sin of the gravity may be interpreted by the driver as an horizontal acceleration.

  Active steering force feedback is computed by a proprietary model and reproduced by a SENSO-Wheel system. The SCANeR© Studio 1.4 software package is used with a real-time version of the MADA (Advanced Modelling of Vehicle Dynamics) vehicle dynamics software, developed by RENAULT. The visual environment is displayed on a cylindrical screen (radius 1.9 m) thanks to five single-chip DLP projectors (Projection Design F12), each with a resolution of 1980 x 1080. The system covers a horizontal field of view of 210°.

Fig. 2 .

 2 Fig. 2. Renault ULTIMATE driving simulator at Virtual Reality and Immersive Simulation Centre.

Fig. 3 .

 3 Fig. 3. Pre-scripted crenel of speed used for the experiment.

Fig. 4 .

 4 Fig. 4. Screenshot of the double-lane motorway.

Fig. 5 .

 5 Fig. 5. Disposition of the orange cones on the road.

Fig. 6 .

 6 Fig. 6. Comparison of longitudinal acceleration rendering between using rails only (no tilt) between 530s and 580s and using tilt coordination technique (configuration 5: 6 °/s and 8°/s²) between 580s and 630s. For the same vehicle acceleration, the acceleration rendered is closer to the input in the second case.

  Fig. 7. Mean "binarized" acceptance in percent for each configuration

Table 1 . Physical capabilities of Renault ULTIMATE simulator

 1

		Stroke	Speed	Accel.
	X Rail	± 2.6 m	± 2.0 m/s	± 5.0 m/s²
	Y Rail	± 2.6 m	± 3.0 m/s	± 5.0 m/s²
	X Axis	± 0.28 m	± 0.7 m/s	± 7.5 m/s²
	Y Axis	± m	± 0.7 m/s	7.5 m/s²
	Z Axis	± 0.20 m	± 0.7 m/s	± 5.0 m/s²
	H Axis	± 15 °	± 40 °/s	± 300 °/s²
	P Axis	± 15 °	± 40 °/s	± 300 °/s²
	R Axis	± 15 °	± 60 °/s	± 600 °/s²

Table 2 . Compared tilt configurations # CFG Max tilt rate [°/s] Max tilt acceleration [°/s²

 2

	]

Table 3 . Configurations order for the 8 participants

 3

	Subject			Configuration order
	# 1	1	8	2	7	3	6	4	5
	# 2	2	1	3	8	4	7	5	6
	# 3	3	2	4	1	5	8	6	7
	# 4	4	3	5	2	6	1	7	8
	# 5	5	4	6	3	7	2	8	1
	# 6	6	5	7	4	8	3	1	2
	# 7	7	6	8	5	1	4	2	3
	# 8	8	7	1	6	2	5	3	4

Table 4 . Quotation of visual stability of virtual environment during accelerations and decelerations (1 = perfectly stable, 6 = not stable at all)

 4

					Configuration
			1	2	3	4	5	6	7	8
	max (°/s)	4		5		6		7
	max (°/s²) 8 30	8	30	8	30	8	30
		1	6	6	4	6	6	6	4	4
		2	3	5	5	5	6	4	6	6
	# Subject	3 4 5 6	5 3 5 5	5 3 5 4	4 2 5 5	4 2 5 5	3 2 5 5	2 3 5 4	3 4 5 4	3 3 5 2
		7	5	5	5	4	2	2	2	2
		8	4	4	5	4	4	3	3	3
		Mean	4.5 4.6 4.4 4.4 4.1 3.6 3.9 3.5

Table 5 . Quotation of ocular discomfort during accelerations and decelerations (1 = no discomfort, 6 = strong discomfort)

 5

			Configuration
	1	2	3	4	5	6	7	8
	max (°/s)	4	5		6		7
	max (°/s²) 8	30	8	30	8	30	8	30

Table 6 . Quotation of the acceptability of acceleration rendering with given tilt configuration (1 = not acceptable at all, 6 = perfectly acceptable)

 6

					Configuration
			1	2	3	4	5	6	7	8
	max (°/s)	4		5		6		7
	max (°/s²) 8	30	8	30	8	30	8	30
		1	6	6	4	5	6	5	4	4
		2	4	3	4	3	5	3	3	5
	# Subject	3 4 5 6	5 3 5 5	5 4 5 4	4 3 5 5	4 4 5 5	3 4 4 5	2 3 5 4	3 4 4 4	3 3 3 2
		7	4	5	4	3	3	2	1	2
		8	4	4	4	3	4	3	2	3
		Mean	4.5 4.5 4.1 4 4.3 3.4 3.1 3.1

Table 7 . "Binarized" quotation of the acceptability of acceleration rendering with given tilt configuration : : "0" if subject quoted "1", "2" or "3" and "1" if subject quoted "4", "5" or "6".

 7

			Configuration
	1	2	3	4	5	6	7	8
	max (°/s)	4	5		6		7
	max (°/s²) 8	30	8	30	8	30	8	30