William Garat

Stéphane Corn

Nicolas Le Moigne
email: nicolas.le-moigne@mines-ales.fr

Johnny J Beaugrand

Anne Bergeret

Stephane Corn
email: stephane.corn@mines-ales.fr

Analysis of the morphometric variations in natural fibres by automated laser scanning: Towards an efficient and reliable assessment of the cross-sectional area

Keywords: A, Natural fibres B, Optical properties/techniques C, Statistical properties/methods D, Optical microscopy E, Morphometric analysis

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Introduction

Composite industry shows growing interest in the development of biocomposites. Natural fibres of various botanical origins, such as hemp, flax or sisal, have been used as reinforcement in polymer composites [START_REF] Bledzki | Composites reinforced with cellulose based fibres[END_REF][START_REF] Joshi | Are natural fiber composites environmentally superior to glass fiber reinforced composites? Compos Part A[END_REF][START_REF] Mohanty | Natural fiber bioplymers and biocomposites[END_REF][START_REF] Baley | Fibres naturelles de renfort pour matériaux composites[END_REF][START_REF] Goda | Research and development of fully green composites reinforced with natural fibers[END_REF][START_REF] Faruk | Biocomposites reinforced with natural fibers: 2000-2010[END_REF][START_REF] Yan | Flax fibre and its compositesa review[END_REF] in substitution for glass fibres [START_REF] Wambua | Natural fibres: can they replace glass in fibre reinforced plastics?[END_REF]. They offer a solution to current environmental issues, such as the reduction of greenhouse gas emissions and pollution, through the use of renewable and biodegradable resources for the development of lightweight materials [START_REF] Mohanty | Sustainable bio-composites from renewable resources: opportunities and challenges in the green materials world[END_REF]. However, natural fibres do not yet benefit from a sufficient scientific and technological background to offer a controlled and constant level of performances adapted to engineering applications as is the case of synthetic fibres [START_REF] Müssig | Testing methods for measuring physical and mechanical fibre properties (plant and animal fibres)[END_REF][START_REF] Thuault | Effects of the hygrothermal environment on the mechanical properties of flax fibres[END_REF][START_REF] Thomason | Fibre cross-section determination and variability in sisal and flax and its effects on fibre performance characterisation[END_REF][START_REF] Haag | Influence of flax fibre variety and year-to-year variability on composite properties[END_REF]. In this regard, researchers, industrials and natural fibre producers investigate potential solutions to better control, model and predict the mechanical behaviour of biocomposites, by improving the extraction of technical natural fibres, the manufacturing of biocomposites and their conditions of service use [START_REF] Baley | Fibres naturelles de renfort pour matériaux composites[END_REF]. This requires a thorough knowledge of the morphological characteristics of natural fibres, in particular their cross-sectional area (CSA) [START_REF] Bourmaud | Impact of the seeding rate on flax stem stability and the mechanical properties of elementary fibres[END_REF][START_REF] Stamboulis | Effects of environmental conditions on mechanical and physical properties of flax fibers[END_REF][START_REF] Placet | Influence of environmental relative humidity on the tensile and rotational behaviour of hemp fibres[END_REF][START_REF] Rosa | Morphological, thermal and mechanical characterization of okra (Abelmoschus esculentus) fibres as potential reinforcement in polymer composites[END_REF][START_REF] Hidalgo-Reyes | Chemical and morphological characterization of Agave angustifolia bagasse fibers[END_REF][START_REF] Lanzilao | Study of the morphological characteristics and physical properties of Himalayan giant nettle (Girardinia diversifolia L.) fibre in comparison with European nettle (Urtica dioica L.) fibre[END_REF][START_REF] Shah | Why do we observe significant differences between measured and "back-calculated" properties of natural fibres?[END_REF]. Indeed, the accuracy of the CSA measurements is crucial since the evaluation of the mechanical properties of natural fibres directly relies on the collected dimensional data and the data processing methods used to calculate their CSAs [START_REF] Haag | Scatter in tensile properties of flax fibre bundles: influence of determination and calculation of the cross-sectional area[END_REF].

Several measurement methods based on Scanning Electron Microscopy (SEM) observations, optical microscopy and scanner measurements and observations of polished cross-section have been used to assess the dimensions of natural fibres [START_REF] Navarro | Microstructural characterization of natural fibers: etlingera elatior, costus comosus, and heliconia bihai[END_REF][START_REF] Romão | Single filament mechanical characterisation of hemp fibres for reinforcing composite materials[END_REF][START_REF] Ilczyszyn | Effect of hemp fibre morphology on the mechanical properties of vegetal fibre composite material[END_REF][START_REF] Virk | Numerical models for natural fibre composites with stochastic properties[END_REF][START_REF] Thuault | Morphologie, biocomposition et comportement mécanique des fibres de lin[END_REF][START_REF] Grishanov | A method of estimating the single flax fibre fineness using data from the LaserScan system[END_REF][START_REF] Mwaikambo | Mechanical properties of alkali treated plant fibres and their potential as reinforcement materials. I. hemp fibres[END_REF][START_REF] Mwaikambo | The effect of chemical treatment on the properties of hemp, sisal, jute and kapok for composite reinforcement[END_REF][START_REF] Fidelis | The effect of fiber morphology on the tensile strength of natural fibers[END_REF][START_REF] Almeida | Evaluation of the cross-section of lignocellulosic fibers using digital microscopy and image analysis[END_REF]. Some authors [START_REF] Müssig | Testing methods for measuring physical and mechanical fibre properties (plant and animal fibres)[END_REF][START_REF] Thomason | Fibre cross-section determination and variability in sisal and flax and its effects on fibre performance characterisation[END_REF][START_REF] Haag | Scatter in tensile properties of flax fibre bundles: influence of determination and calculation of the cross-sectional area[END_REF][START_REF] Charlet | Multi-scale morphological characterisation of flax: from the stem to the fibrils[END_REF][START_REF] Thomason | Natural fibre cross sectional area, its variability and effects on the determination of fibre properties[END_REF] also related the measurement of the length, width and CSA of various natural fibre species with their mechanical properties. They evidenced wide variations both in terms of morphometric characteristics and resulting mechanical properties. However, several of these studies are based on simplified assumptions of the cross-sectional shape that could be valid for synthetic fibres but not for the specific shape and dimensional irregularities of natural fibres [START_REF] Müssig | Testing methods for measuring physical and mechanical fibre properties (plant and animal fibres)[END_REF]. Indeed, in many cases, an assumption considering cross-section of natural fibres as circular and homogeneous is used, though natural fibres, whatever their botanical origin and the scale considered (i.e. elementary fibre cells or fibre bundles), show wide variations in their dimensional and morphometric features. Besides, their cross-sectional shape can vary from fibre to fibre and along their length [START_REF] Thomason | Fibre cross-section determination and variability in sisal and flax and its effects on fibre performance characterisation[END_REF][START_REF] Thomason | Natural fibre cross sectional area, its variability and effects on the determination of fibre properties[END_REF][START_REF] Munawar | Characterization of the morphological, physical, and mechanical properties of seven nonwood plant fiber bundles[END_REF][START_REF] Da Costa | Diameter dependence of tensile strength by Weibull analysis: Part I bamboo fiber[END_REF][START_REF] Bevitori | Diameter dependence of tensile sectional shapes encountered in natural fibres. This filtering method can be implemented in routine as an automated post-processing of the data obtained from the laser scanning method. strength by Weibull analysis: Part II jute fiber[END_REF][START_REF] Inacio | Diameter dependence of tensile strength by Weibull analysis: Part III sisal fiber[END_REF][START_REF] Charlet | Characteristics of Hermès flax fibres as a function of their location in the stem and properties of the derived unidirectional composites[END_REF]. This partly explains the large scattering in fibre mechanical properties reported in literature since many years [START_REF] Bledzki | Composites reinforced with cellulose based fibres[END_REF][START_REF] Faruk | Biocomposites reinforced with natural fibers: 2000-2010[END_REF][START_REF] Müssig | Testing methods for measuring physical and mechanical fibre properties (plant and animal fibres)[END_REF][START_REF] Shah | Why do we observe significant differences between measured and "back-calculated" properties of natural fibres?[END_REF]. Studying the tensile properties of flax 2.2. Methods for the measurement of fibre bundle cross-section

Automated laser scanning measurements

For the automated laser scanning measurement, twelve fibre bundles of each plant species were cut to a length of roughly 3 mm. Then, each fibre bundle was positioned in a plastic tab before being fixed quickly (approximately 5 s) with a photo-curing glue (DYMAX, Wiesbaden, Germany). This step prevents fibre bundles from slipping out of the support due to gentle tension applied during measurements. Before automated laser scanning measurements, fibre bundles were conditioned at 23 °C and 50% RH overnight (12 h).

The automated laser scanning method used the Fibre Dimensional Analysis System (FDAS) device controlled by UvWin 3.60® software (Diastron Ltd., Hampshire, UK). This apparatus allows measuring apparent transverse dimensions of the fibre bundles with a resolution of 0.01 µm, thanks to a high accuracy non-contact laser measuring system (LSM 500S, Mitutoyo, Japan). Fibre bundles fixed on plastic tabs are maintained by a pneumatic system which allows 360°rotative scanning all along their length (Fig. 2a). For each scan position with a pitch of 40 µm along the fibre, up to 600 apparent diameters were measured on the rotating fibre bundle. This leads to more than 45.000 values of apparent diameter for a fibre bundle of 3 mm length. Then, apparent transverse dimensions such as maximum, minimum and average diameters can be deduced for each cross-section and along the fibre bundle (Fig. 2b). It should be pointed out that considering the measurement principle which is based on laser ombroscopy; concave surfaces cannot be measured with this device.

Optical microscopy measurements

Direct optical microscopy observations of the fibre bundles crosssections were performed with a Leica Laborlux 12 POL S optical microscope in reflection mode equipped with a 1600 × 1200 pixels mono-CDD Sony digital camera. This method was used by Virk [START_REF] Virk | Numerical models for natural fibre composites with stochastic properties[END_REF] for jute fibres, by Müssig et al. [START_REF] Müssig | Testing methods for measuring physical and mechanical fibre properties (plant and animal fibres)[END_REF] and Thomason et al. [START_REF] Thomason | Natural fibre cross sectional area, its variability and effects on the determination of fibre properties[END_REF] for flax and sisal fibres and by Navarro et al. [START_REF] Navarro | Microstructural characterization of natural fibers: etlingera elatior, costus comosus, and heliconia bihai[END_REF] for less common plant fibre bundles. The fibre bundles analysed by the automated laser scanning technique beforehand were then embedded vertically in an epoxy resin-pad. The Geofix® resin (ESCIL, Chassieu, France) was used. It is a transparent resin specially formulated to impregnate samples (high fluidity and low shrinkage) with a curing time at room temperature of 24 h. Fibre fibre bundles, Haag and Müssig [START_REF] Haag | Scatter in tensile properties of flax fibre bundles: influence of determination and calculation of the cross-sectional area[END_REF] proposed an advanced measurement technique of the CSA based on an automated laser scanning that has the advantage to measure the cross-sections dimensions all along the fibre and to be highly reproducible and not influenced by user. Nevertheless, the authors pointed out that errors can occur during laser measurements if partly spliced fibre bundles are measured instead of the whole element. They assume that filtering procedures can be necessary to exclude falsified data from the evaluation.

The purpose of this study is to develop a methodology to better assess the dimensional and morphometric characteristics of natural fibres from various botanical origin. Cross-sectional measurements obtained from optical microscopy observations are correlated with measurements recorded with an automated laser scanning technique. Based on these measurements, data processing approaches adapted to the morphometric characteristics of each plant fibre species are proposed for the reliable calculation of natural fibres CSAs.

Experimental section

Materials

The dimensional and morphometric analysis of natural fibres was conducted on fibre bundles from five different plant species, three of them belonging to Eudicots and are commonly called 'bast fiber', poorly lignified, with a support function of the stem in planta: flax (Linum usitatissimum) and hemp (Canabis sativa) as annual fibre crop (after retting step) and nettle (Urtica dioica) as an perennial herbaceous plant. The other two species are from perennial Monocots, highly lignified, with a dominant conducting tissues function in planta: sisal (Agave sisalana) coming from the leaf of agave and Palm (Phoenix dactylifera) are mats of leaf sheath surrounding the palm tree stems.

Due to either their various botanical origins and functions in plant (strength versus conducting), these fibre bundles are likely to show contrasted dimensional and morphometric characteristics as described in literature [START_REF] Baley | Fibres naturelles de renfort pour matériaux composites[END_REF][START_REF] Müssig | Testing methods for measuring physical and mechanical fibre properties (plant and animal fibres)[END_REF][START_REF] Bourmaud | Exploring the potential of waste leaf sheath date palm fibres for composite reinforcement through a structural and mechanical analysis[END_REF] and observed in Fig. 1. Wide variations in their cross-sectional dimensions as well as variations in their cross-sectional shape in terms of ellipticity and concavity are clearly observed. bundles axial position within the epoxy resin-pad was precisely determined so that the cross-sections measured by optical microscopy can be directly compared one to one with those measured by the automated laser scanning technique. The resin-pads containing the fibre bundles were gradually sanded with a pitch of roughly 300 µm. A ten-polishing sequences procedure using different fine polishing papers (600, 2400, 4000 grade) wetted with ethanol was achieved to ensure optimum observation quality (Fig. 3a). After each sanding and polishing step, the cross-section of each fibre bundle was observed under the optical microscope and their cross-sectional areas (CSA OM ) were measured with the image acquisition and processing software (Archimede®, Microvision Instruments, France). As shown in Fig. 3b andc, the outline of the fibre bundle cross-section was cropped manually to calculate its area. With 12 fibre bundle samples and 10 successive sanding and polishing sequences, this leads to approximately 120 CSA OM measurements for each of the five plant species.

Results and discussion

Lengthwise morphometric variations in fibre bundles from various plant species

Based on the FDAS measurements, two main morphotypes among the five plant species were observed. Palm and sisal fibre bundles appear relatively uniform and regular with low intra-fibre variability, i.e. the minimum, maximum and average diameters are relatively constant along the fibre bundles (Fig. 4a). Moreover, the average diameter and maximum and minimum diameters are relatively close, suggesting that their cross-sectional shape is almost circular (Fig. 4b andc). Similar results were obtained for each fibre bundles samples from the same plant species, suggesting that, for the tested batch of sisal and palm fibre bundles, low inter-fibre bundles variability occurred. It should be pointed out that the cross-sections of sisal fibre bundles always showed a kidney shape which should be taken into account as the concave surfaces cannot be measured with the FDAS device (see Section 2.2.1.). Besides, palm and sisal fibre bundles are composed of many strongly packed elementary fibres, reported as highly lignified, with large lumen for some of them.

Flax, hemp and nettle fibre bundles are much more irregular with wide variations in the minimum, maximum and average diameters along their length, i.e. high intra-fibre variability (Fig. 5a). The large difference measured between their maximum and minimum diameters confirms their non-circularity, as can be evidently concluded from optical microscopy observations (Fig. 5b-d). Besides, the analysis of the whole data set available for flax, hemp and nettle fibre bundles showed that there is more inter-fibre bundles variability for these plant species. This has to be related to their botanical origin but also their harvesting and extraction conditions. In particular, the presence of elementary fibres detached from the main bundle was observed for retted flax fibres due to their lower intercellular cohesion that favoured fibre bundles splicing.

Based on these first observations, it is obvious that the simplifying geometrical assumption that considers natural fibre bundle cross-sections as being purely circular and uniform is not appropriate. In the following, a correlation between the data obtained from optical microscopy measurements and those measured by the automated laser scanning is carried on.

Morphometric indicators of the (non-)circularity of natural fibre crosssection

Fig. 6 correlates the apparent median diameter D median measured with the FDAS for each cross-sections of each fibre bundle sample and the CSA OM of the corresponding cross-section (as determined by optical microscopy measurements). A data table gives the minimum, maximum and median apparent diameters and CSA OM of the tested fibre bundles for the five plant species. For each plant species, the set of data pairs, i.e. CSA OM from optical microscopy and corresponding D median from FDAS, was fitted with a circular-like model by least squares method according to Equation 1:

= × CSA k D OM median 2 (1)
Where, k is an adjustable parameter that equals to π/4 (i.e. 0.785) for a perfect circular shape. The typical curve that should be obtained for a perfect circular shape was superimposed.

As seen in Table 1, a good correlation to the circular model was obtained for palm and sisal fibre bundles with k parameters equal to 0.778 and 0.702 respectively, very close to that of a perfect circular shape (0.785), and good coefficients of determination R 2 of 0.94 and 0.93 respectively. For the other plant species, k parameters were much lower, 0.605, 0.660 and 0.485 for hemp, flax and nettle fibre bundles, respectively, suggesting a strong deviation to the circular shape for these fibre bundles. Larger scattering to the best fit obtained with k was also observed with coefficients of determination R 2 of 0.88, 0.68 and 0.79, respectively. It should be pointed out that deviations to the circular model do not seem to be diameter dependent because there is substantial overlapping of apparent median diameter values between the different plant species considered in this study.

To complete this first approach and determine if morphometric indicators can be attributed to each plant fibre species, shape factors α were calculated based on the ratio of the maximum to the minimum diameters obtained by FDAS measurement for each fibre bundle crosssection of the different plant species (Equation 2):

= α D D max min (2)
Exhibiting median shape factors of 1.17 and 1.24, respectively, cross-sectional shapes of palm and sisal fibre bundles appear to be close to a circular shape (Fig. 7). In contrast, hemp, flax and nettle fibre bundles present median shape factors of 2.35, 2.58 and 2.74, respectively, with a wider dispersion. These results are in agreement with previous observations and conclusions made from the results presented in Fig. 5 and Fig. 6, and suggest that a geometrical model based on the assumption of an elliptical cross-sectional shape would be more suitable for the calculation of CSAs for hemp, flax and nettle fibre bundles. Furthermore, the determination of shape factors from FDAS measurements appears as an easy-to-calculate and robust morphometric indicator to assess the circularity of cross-sections in natural fibre bundles, whatever the diameter considered (Fig. 6).

Geometrical models and filtering method for the determination of CSAs in natural fibre bundles

Several studies have been conducted on natural fibre dimensional measurements, i.e. diameters and sections. Many studies considered natural fibres as circular and regular objects [START_REF] Virk | Numerical models for natural fibre composites with stochastic properties[END_REF][START_REF] Wang | An improved fibril angle measurement method for wood fibres[END_REF]. This hypothesis does not take into account the non-circularity of the fibre cross-section, neither does the variability of the fibre cross-section along their length [START_REF] Thomason | Fibre cross-section determination and variability in sisal and flax and its effects on fibre performance characterisation[END_REF]. Other authors took into account the irregularity of the section and considered the cross-section as elliptic [START_REF] Thomason | Fibre cross-section determination and variability in sisal and flax and its effects on fibre performance characterisation[END_REF][START_REF] Haag | Scatter in tensile properties of flax fibre bundles: influence of determination and calculation of the cross-sectional area[END_REF], hexagonal [START_REF] Nitta | Il. Cross-sectional area evaluation and tensile properties of alkali-treated kenaf fibres[END_REF] or as a convex hull [START_REF] Virk | Physical characterization of jute technical fibers: fiber dimensions[END_REF], to better assess the real cross-sectional shape of natural fibres. It should be pointed out that all these assumptions on the cross-sectional shape of the fibres directly influence the evaluation of their mechanical properties and can contribute to the large scattering found in literature for tensile strength and elastic modulus of natural fibres.

Because of the considerable number of apparent diameter values that can be collected in a reasonable time, the use of an automated laser scanning device appears as a relevant and robust method which should allow taking into account apparent dimensional irregularities of natural fibre bundles. Based on the morphometric analysis, two morphotypes have been identified: (i) palm and sisal fibre bundles which present a lengthwise regular cross-sectional shape and low shape factors close to 1, and (ii) hemp, flax and nettle fibre bundles which present wide lengthwise dimensional variations as well as irregular cross-sectional shapes with high shape factors up to 2.74. According to these morphometric features, different approaches of data processing and geometrical models were applied to FDAS data for CSA calculation as shown in Table 2.

The simplest approach is the circular model (Eq. ( 3)) which considers the apparent median diameter of each cross-section, determined from 600 diameter values, to calculate the CSA. It is valid for fibre bundles having a nearly circular cross-sectional shape. The elliptic model (Eq. ( 4)) uses the maximum and minimum apparent diameters of each cross-section, extracted from 600 values, to calculate the CSA. The filtered elliptic model (Eq. ( 5)) has been developed in this work to exclude wrong data obtained with FDAS device, due to partly spliced fibre bundles with detached elementary fibres (Fig. 8a) that can greatly mislead the determination of the maximum and minimum diameters. This model is based on a statistical approach that considers the occurrence frequency of apparent diameters during the data acquisition. As shown in Fig. 8 for the scan of a flax fibre bundle cross-section, the presence of detached elementary fibres can be identified by the appearance of peaks of low frequency (< 0.5%), generally located at small diameter range of the order of the elementary fibre cell (Fig. 8b).

It is then possible to identify, for each cross-section scan, the wrong values and exclude them from the statistical dataset. Filtered minimum ′ D min and maximum ′ D max diameters can be identified and used for the calculation of the CSA (Eq. ( 5)).

The three methods of data processing described in Table 2 to determine CSA from FDAS measurements were applied to the five plant fibre species, and the resulting CSA FDAS were compared to the CSA OM measured by optical microscopy.

Natural fibres with regular cross-sectional shape and low shape factor

Fig. 9 shows a radar graph of the apparent diameters obtained from FDAS measurements for one specific cross-section of a palm (a) and sisal (b) fibre bundle. For palm, the radar pattern appears relatively circular, with a low deviation to the three models, and CSA OM is very close to CSAs obtained from the models (23,460 µm 2 versus 25,522 µm 2 and 23,297 µm 2 for circular and elliptic models, respectively). For sisal, the deviation is slightly increased because of its particular kidney shape cross-section (Fig. 1a).

Considering now the whole data set, a good correlation was found between CSA OM and CSA FDAS values for palm fibre bundles (Fig. 10a), whatever the model used, with coefficients of determination higher than 0.95. Moreover, the correction factor C defined by = × CSA C CSA FDAS OM was close to 1, attesting for a very good estimation of the CSA with the automated laser scanning method.

For sisal fibre bundles (Fig. 10b), only slight differences between the three models are observed. Coefficients of determination are roughly

Table 2

Geometrical models and data processing approaches used to calculate the CSA from FDAS measurements. (Eq. ( 5)) 0.9, and the analysis of correction factors C showed that the circular model slightly overestimates CSA OM with a C value of 1.02, whereas both elliptical models slightly underestimate CSA OM with a C value of 0.97. The kidney shape of sisal fibre bundles and the resulting concave surfaces do not seem to hamper the reliable and accurate determination of their CSA.

Method

Our results thus show that CSAs of regular palm and sisal fibre bundles can be satisfactorily approached from the FDAS measurements using a circular model with minimal data processing. Indeed, the filtered elliptic model does not enhance accuracy since the intercellular cohesion within the fibre bundles is sufficient to avoid any fibre bundle splicing. It should be pointed out that no fibre sample pre-treatments aiming at fibre individualization were used for sisal or palm fibre bundles. Also, both bundles are reported to be highly cohesive thanks to a high lignin amount, arguably in their middle lamellas.

Natural fibres with irregular cross-sectional shape and high shape factor

For hemp, flax and nettle fibre bundles, an important deviation from circularity characterised by high shape factors was found (Fig. 7). Moreover, large lengthwise morphometric variations were highlighted. In this regard, the use of an elliptical model appears more appropriate to determine the CSA.

Fig. 11 shows the radar of the apparent diameter obtained from FDAS measurements for one flax cross-section with fibre bundle splicing and one without. In both cases, the radar patterns are close to that of an ellipse. In the absence of measurement defects, the elliptic model is appropriate with CSA of 9057 µm 2 compared to CSA OM of 8950 µm 2 (Fig. 11b). On the other hand, the use of a filtered elliptic model is mandatory to identify and eliminate wrong apparent diameter values, responsible for the large deviation of the simple elliptic model as evidenced in Fig. 11a with a large error in the CSA determination (1389 µm 2 versus 8179 µm 2 for the CSA OM ). Considering the whole data set for hemp and flax fibre bundles (Fig. 12a andb), the circular model gives higher coefficients of determination. However, this model significantly overestimates the CSA. For nettle fibre bundles (Fig. 12c), the elliptic model is better with a slightly higher coefficient of determination and a correction factor C of 1.01.

For all these fibre bundles, the filtered elliptic model appears to greatly enhance the quality of the correlation with CSA OM . Coefficients of determination are indeed greatly increased and correction factors are closer to 1. This is explained by the numerous splicing of fibre bundles related to lower intercellular cohesion, that produce wrong data for these plant fibres, especially low apparent diameter values originating from detached elementary fibres. These wrong data must be excluded from the analysis, and the filtered elliptic model developed in this study greatly enhances the determination of the CSA for the irregular cross-

Conclusions

The morphometric variations occurring in natural fibres have been investigated. Various plant fibre species (sisal, palm, flax, hemp, and nettle) having contrasted morphometric characteristics were analysed by optical microscopy and automated laser scanning (FDAS) measurements. Two main morphotypes have been identified: (i) monocots palm and sisal fibre bundles from leaves which presented a lengthwise regular cross-sectional shape and low shape factor close to 1, and in contrast (ii) eudicots hemp, flax and nettle fibre bundles from bast fibres which presented wide lengthwise dimensional variations as well as irregular cross-sectional shape and high shape factors up to 2.74. Based on these observations, geometrical models and a filtering method were applied to the FDAS data so as to determine the best approach to calculate the cross-sectional area (CSA) of each plant fibre species. Our results show that CSAs of regular palm and sisal fibre bundles can be satisfactorily assessed from the FDAS measurements using a circular model with minimal data processing. For more irregular cross-sectional shape encountered in natural fibres such as hemp, flax and nettle, the filtered elliptic model developed in the present study greatly enhances the determination of the CSA. This filtering method can be implemented in routine as an automated post-processing of the data obtained from the FDAS. Furthermore, based on the CSA measured by optical microscopy, CSA correction factors were determined for each plant fibre species and the applied geometrical models so that an accurate CSA can be calculated from the automated laser scanning data. The methodology developed in this study appears as a fast, efficient and reliable approach for the assessment of the cross-sectional area in natural fibres. This opens interesting perspectives for a better assessment of natural fibre tensile properties both for research and quality control purposes in industry.

Fig. 1 .

 1 Fig. 1. Typical cross-sections of (a) sisal Agave sisalana, (b) palm Phoenix dactylifera, (c) flax Linium usitatissimum, (d) nettle Urtica dioica, and (e) hemp Canabis sativa drawn from optical microscopy observations.

Fig. 2 .

 2 Fig. 2. (a) FDAS device principle and (b) minimum, maximum and average diameters along a palm fibre bundle as measured with the FDAS device.

Fig. 3 .

 3 Fig. 3. (a) Scheme of an embedded fibre bundle in an epoxy resin-pad; Manual outline of the fibre bundle perimeter with the image processing software (Archimede®, Microvision Instruments, France) for (b) sisal and (c) nettle.

Fig. 4 .

 4 Fig. 4. (a) Lengthwise morphometric variations for palm and sisal fibre bundles from the FDAS, and typical cross-section of (b) palm and (c) sisal fibre bundles embedded in epoxy resinpad as observed by optical microscopy.

Fig. 5 .

 5 Fig. 5. Lengthwise morphometric variations (minimum, maximum and average diameters) for (a) hemp, (b) flax and (c) nettle fibre bundles, and typical cross-section of (d) hemp, (e) flax and (f) nettle fibre bundles embedded in epoxy resin-pad as observed by optical microscopy.

Fig. 6 .

 6 Fig. 6. Correlation between CSA OM measured by optical microscopy and corresponding apparent median diameters (D ) median measured by FDAS for sisal, palm, hemp, nettle and flax fibre bundles. (Full line corresponds to the curve obtained for a perfect circular shape).

Fig. 7 .

 7 Fig. 7. Boxplots of fibre bundles shape factors α for the different plant species (palm, sisal, hemp, flax, nettle).

Fig. 8 .

 8 Fig. 8. (a) Optical micrograph of a flax fibre bundle with a detached elementary fibre, (b) corresponding occurrence frequency of diameters (interval unit: 1 µm) for one cross-section scan.

Fig. 9 .Fig. 10 .

 910 Fig. 9. Radar of the apparent diameters from FDAS and corresponding modelled crosssections for one palm (a) and sisal (b) fibre bundle cross-section.

Fig. 11 .

 11 Fig. 11. Radar of the apparent diameters from FDAS and corresponding modelled crosssections for one flax fibre bundle cross-section, (a) with fibre bundle splicing, and (b) without fibre bundle splicing.

Fig. 12 .

 12 Fig. 12. Correlation between CSA FDAS and CSA OM for circular, elliptic and filtered elliptic models for (a) hemp, (b) flax and (c) nettle fibre bundles.

Table 1

 1 Fibre cross-sectional shape analysis based on a circular model.

	Fibre bundles	Number of data pairs (CSA OM -D median )	k	R 2
	Perfect cylinder	-	0.785	1
	Palm	140	0.778	0.94
	Sisal	138	0.702	0.93
	Hemp	125	0.605	0.88
	Flax	130	0.660	0.68
	Nettle	120	0.485	0.79

Acknowledgements

William Garat thanks Montpellier SupAgro for funding its thesis scholarship. The authors thank Alain Diaz (IMT Mines Ales, C2MA) for technical support on the preparation of the samples, Arnaud Day (Fibres Recherche Développement, Troyes, France) for providing the nettle fibres and Yann Leray and Steve Bucknell (Diastron Ltd., Hampshire, UK) for technical support on FDAS device.


Dr Arun-Libertsen Lawzer
email: alawzer@ichf.edu.pl

Dr Thomas Custer
email: thomas.custer@ichf.edu.pl

Dr Jean-Claude Guillemin
email: jean-claude.guillemin@ensc-rennes.fr

Dr Robert Kołos
email: rkolos@ichf.edu.pl

An efficient photochemical route towards triplet ethynylphosphinidene, HCCP

Keywords: phosphinidene, rearrangement, phosphapropyne, photochemistry, HCCP, matrix isolation, phosphapropadiene 10

While the archetypal free phosphinidene, H-P, has been studied for over a century, reports on uncomplexed, univalent phosphorus compounds are very sparse. Here we demonstrate production of HCCP in solid argon through the UV-induced rearrangement and subsequent dehydrogenation of phosphapropyne, CH 3 CP. Migration of H atoms along the CCP backbone of CH 3 CP resulted in production of the previously unobserved species 1phosphapropadiene, CH 2 =C=PH, followed by ethynylphosphine, HCCPH 2 .

Phosphinidenes, the phosphorus analogues of nitrenes, have long been known as ligands in transition metal complexes, [1] but only a handful of the free species have been experimentally characterized. The hydride (HP), [2] cyanophosphinidene (PCN), [3] and ethynylphosphinidene (HCCP) [4] were detected as inseparable gaseous components produced through electrical discharge or high energy irradiation. Mesitylphosphinidene was observed in a solvent following the photolysis of mesitylphosphirane. [5] Phenylphosphinidene was later produced [6] in a cryogenic argon matrix (just like mesitylphosphinidene) [7] from the corresponding phosphirane. Reaction of silicon atom with PH 3 , also in an argon matrix, showed the presence of H 3 SiP (silylphosphinidene). [8] Recently, a singlet phosphinophosphinidene, stable at room temperature, was synthesized. [9] All other known phosphinidenes have triplet electronic ground states.

Given the presence of HCCN [10] and the CCP radical [11] in the interstellar medium, HCCP has been theorized as a potentially detectable species of astrochemical interest. [12] The rotational spectrum [4] and theoretical studies [13] of HCCP indicated a linear, allenic structure, H-C ̇=C= P ̇, with a significant contribution from the H-C≡C-P ̇• electronic configuration (Tab. S4). For HCCN, [14] the prevailing arrangement, H-C ̇=C=N ̇ , is accompanied by a significant carbenic admixture, H-.C ̇-C≡N .A theoretical (coupled cluster) study [13a] did not reveal any potential energy minimum that would correspond to an analogous phosphorus-bearing carbene, H-.C ̇-C≡P. Although both HCCX species have a triplet ground state, the singlet-triplet energy gap, very small for HCCN (3 kJ/mol), [15] is 75 kJ/mol for HCCP. [13a] HCCN has been formed through photolysis of diazoacetonitrile (N 2 CHCN) and tentatively through VUV photolysis of acetonitrile (CH 3 CN). [16] The synthesis of the phosphorus analogue of diazoacetonitrile remains a challenge while that of phosphapropyne [17] (CH 3 CP, 1) is known. No photodegradation studies have been carried out for any C 2 H 3 Pstoichiometry compound.

Scheme 1. Isomerisation of phosphapropyne 1 followed by dehydrogenation of ethynylphosphine 2.

Previous studies of Guillemin et al. [18] have shown that ethynylphosphine (HC≡CPH 2 , 3) rearranges to 1 in a reaction catalyzed by a strong Lewis base or a weak Bronsted base at high temperature in a vacuum gas-solid reaction. The authors proposed an intermediate species, phosphapropadiene (CH 2 =C=PH, 2), although it has never been observed experimentally. Herein, we report the UV-induced rearrangement 10.1002/anie.202016052

Accepted Manuscript

Angewandte Chemie International Edition Theory (B3LYP/aug-cc-pVTZ) b Experiment (Argon matrix)

Freq. c (cm -1 ) Intensity (km mol -1 ) Isotopic shifts (cm -1 ) Freq.

(cm -1 ) Relative intensity (%)

Isotopic shifts (cm -1 ) [a] Five bands of the highest DFT-predicted IR intensity (listed here) were experimentally observed. [b] Table S2 collects all fundamental modes and Table S3 shows DFT-predicted geometry of 2.

[c] Scaling factor 0.96.

and dehydrogenation of phosphaalkyne 1 isolated in a cryogenic argon matrix, finally leading to HCCP, with species 2 and 3 being formed en route (Scheme 1).

Precursor 1 was isolated in solid argon at a molar ratio of 1:1000 (T=6 K) and exposed to Hg-lamp radiation with peak emission at 254 nm. The progress of photoreactions was monitored using FTIR and UV/visible absorption spectroscopy. At early stages of irradiation, the strongest product bands are directly attributable to the phosphine 3 based on their perfect match with a reference spectrum of a freshly synthesized sample made from diethyl ethynylphosphonate [19] (see Fig. S2 andTab. S1 for the relevant IR-spectroscopic data). Closer analysis reveals the presence of the allenic species 2, hypothesized in the work of Guillemin et al. [18] Its identification is based on DFT computations assisted by photochemistry of isotopically labeled precursors, 13 CH 3 CP and CD 3 CP (Tab. 1, Tab. S2, Fig. S3). The maximum concentration of 2 is reached prior to that of 3 (Fig. 1), suggesting a stepwise hydrogen migration from methyl group towards the phosphorus atom. The decay of all C 2 H 3 P species was observed upon prolonged photolysis (Fig. 1), and dehydrogenation turned out to be the main process. The final product, HCCP (ethynyl phosphinidene), was unambiguously identified using isotopic labelling, use of different photolysis wavelengths to identify related bands, and comparison with DFT-predicted IR spectra and their isotopic shifts as illustrated in Figs. 2 andS1. Observed experimental IR absorption data along with theoretical values, including isotopic shifts, are listed in Table 2.

In addition to aiding the identification of photoproducts experiments involving precursor 1 with 13 C-substituted methyl group also shed light on the isomerization mechanism. Photolysis 10.1002/anie.202016052

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Angewandte Chemie International Edition Table 2. IR-spectroscopic identification of ethynylphosphinidene, HCCP.

Mode

Theory (B3LYP/aug-cc-pVTZ) a Experiment (Argon matrix)

Freq.

(cm -1 )

Intensity (km mol -1 )
Isotopic shifts (cm -1 ) Freq.

(cm -1 ) Intensity (km mol -1 ) Isotopic shifts (cm -1 )

H 13 of 13 CH 3 CP gave rise, as anticipated, to the 13 CH 2 =C=PH isotopomer of 2. However, the formation of both H 13 CCPH 2 and HC 13 CPH 2 (Tab. S1 and Fig. S2), with a ratio of approx. 1:1, was observed. This suggests that the rearrangement from 2 to 3 proceeds through a phosphacyclopropene-like species (Scheme 2). Either one of the two CP bonds may break within a ring, leading to the two observed 13 C-isotopomers of 3. This scrambling is conserved upon extended photolysis where the final products H 13 CCP and HC 13 CP appear in equal amounts. The cyclic intermediate may correspond either to a saddle point or a local energy minimum of the ground or excited state (singlet or triplet) potential energy surface. Its structure may resemble 1Hphosphirene (Scheme 2). Pertinent quantum chemical computations are under way to identify the nature of this species. The lack of any preference for H 13 CCP over HC 13 CP among the products of 13 CH 3 CP photolysis implies that ethynylphosphinidene is not produced through direct dehydrogenation of either 1 or 2 but rather by loss of H 2 from 3. In a separate experiment involving Hg-lamp photolysis of pure 3 in solid argon, HCCP was the final product. However, species 1 and 2 were also generated (Fig. S5), demonstrating the reversibility of photoisomerization reactions 1->2 and 2->3.

The course of photolysis of CH3CP was also monitored using electronic absorption spectroscopy (Fig. S6), providing first insights into the electronic spectra of species 1-3 and HCCP. As photolysis progresses, the characteristic spectrum of CH 3 CP is gradually eclipsed by two absorption maxima at 243 and 237 nm by 40 minutes of photolysis. These features, tentatively assigned to species 2, disappear upon extensive irradiation while new absorption features appear, including strong bands at 277 and 259 nm as well as weaker features between 350 and 290 nm, at least partially due to HCCP. None of these features can be assigned to 3 whose spectrum is featureless.

In summary, photolysis of phosphapropyne (CH 3 CP, 1) in a cryogenic argon matrix with mid-UV radiation results in formation of triplet HCCP. Dehydrogenation is preceded by the appearance of the C 2 H 3 P isomers CH 2 =C=PH (2) and HC≡CPH 2 (3). Isotopic labelling experiments suggest passage through a 3-membered ring in the 2 -> 3 isomerization. This photochemical route for production of matrix isolated HCCP paves the way for the experimental exploration of its reactivity and electronic structure.

Figure 1 .

 1 Figure 1. Evolution of concentrations over the course of 254 nm photolysis of CH3CP (1) in solid Ar, scaled with respect to the initial concentration of the precursor. Integrated band intensities measured at 1556 cm-1(1), 1733 cm -1 (2; CH2=C=PH), 3311 cm -1 (3; HC≡CPH2), and 3284 cm -1 (HCCP) were used in conjunction with the respective B3LYP/aug-cc-pVTZ absolute IR band strengths of 43 km/mol, 79 km/mol, 66 km/mol, and 76 km/mol. Common ordinate scale for the species 1, 3, and HCCP; 10-fold vertical expansion for species 2.

Figure 2 .

 2 Figure 2. Identification of HCCP. Top: IR spectrum (bands pointing downwards) theoretically derived (B3LYP/aug-cc-pVTZ, harmonic approximation, scaling factor 0.96) for the ground triplet electronic state of HCCP. Bottom: spectrum resulting from prolonged 254 nm irradiation of phosphapropyne (1) in solid Ar; ν1 -ν4 refer to the identified bands of HCCP. Bands due to 1 and 3 (ethynylphosphine) are labeled accordingly (see Fig S4, for the IR spectrum of matrix-isolated 1). Asterisk indicates an artefact feature (736 cm -1 ) originating in the CsI substrate window.

Table 1 .

 1 IR spectroscopic identification of phosphapropadiene (CH2=C=PH, 2) produced upon CH3CP photolysis

	Mode a

Acknowledgements J. C. G. thanks the Centre National d'Etudes Spatiales (CNES) for a grant and the Programme National Physique et Chimie du Milieu Interstellaire" (PCMI) of CNRS/INSU with INC/INP cofunded by CEA and CNES.

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Entry for the Table of Contents

Triplet HCCP is produced as the final product of the UV induced rearrangement and subsequent dehydrogenation of phosphapropyne, CH 3 C≡P, in solid Ar. Phosphaallene CH 2 =C=PH, and ethynylphosphine, HC≡CPH 2 are produced as H atoms are transferred along the CCP skeleton prior to dehydrogenation.