Maria Teixidó

Clément François

Laura Bosch

Claudia Männel

The role of prosody in early speech segmentation and word-referent mapping: Electrophysiological evidence Running title: Prosody in word segmentation and mapping: ERP data

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Introduction

Early in development, infants face the twin problems of segmenting the speech stream to extract linguistically relevant units and mapping these possible word-forms to their referents, the objects and events in the surrounding world. Developmental research has demonstrated the pivotal role of prosody in this process. Prosodic features, such as language rhythm, intonation, and lexical stress patterns, can provide infants with a starting point in early word learning (see [START_REF] Jusczyk | How infants begin extract words from speech[END_REF][START_REF] Goyet | Processing continuous speech in Infancy: From major prosodic units to isolated word forms[END_REF]for reviews). Following the seminal work by [START_REF] Jusczyk | Infants' detection of the sound patterns of words in fluent speech[END_REF], a large number of behavioral studies have revealed infants' gradually refined abilities to exploit prosodic cues, as well as other available cues from the input, to successfully segment the speech stream and build their first vocabulary.

Behavioral research methods involve overt attentional measures from videorecordings or eye-tracking procedures. In contrast, neuroimaging techniques, such as event-related brain potentials (ERPs), can register brain responses underlying early acquisition processes and even reveal infants' abilities where behavioral methods might not be sensitive enough. Following a brief introduction on the ERP methodology, this chapter offers an overview of infant ERP studies on the role of prosody in early word segmentation and word-referent mapping. More specifically, we will focus, in turn, on lexical stress patterns, some properties of infant-directed speech (IDS) such as accentuation and repetition, and melodic information, as cues to early word segmentation. Then, we will review word-referent mapping studies, pointing to the scarcity of ERP data specifically exploring the role of prosody in this domain.

Throughout the sections of this chapter, we will emphasize how ERP data significantly extend our knowledge gathered from behavioral language acquisition research.

The method of event-related brain potentials (ERPs) in language acquisition research Behavior-independent neuroimaging techniques, like the ERP methodology based on electroencephalography (EEG), offer insights into the mechanisms driving infants' behavior and their developmental progression. The ERP methodology has largely contributed to our understanding of early language acquisition processes and continues to be used as a powerful research tool complementing behavioral techniques (for an overview see [START_REF] Conboy | Eventrelated potential studies of early language processing at the phoneme, word, and sentence levels[END_REF][START_REF] Friederici | Neurophysiological markers of early language acquisition: from syllables to sentences[END_REF][START_REF] Friederici | Neural correlates of the development of speech perception and comprehension[END_REF][START_REF] Kuhl | Brain mechanisms in early language acquisition[END_REF][START_REF] Männel | Event-related brain potentials as a window to children's language processing: From syllables to sentences[END_REF].

The human brain constantly produces electrical activity. EEG captures this brain activity by recording ongoing scalp-level voltage fluctuations that originate from neurons' postsynaptic potentials. Electrophysiological research relies on the fact that external stimulation modulates EEG data. However, the effect of a single stimulation represents only a small part of the ongoing brain activity, such that the event-related portion of EEG recordings associated with a stimulus of interest needs to be extracted. Repeated stimulation and subsequent averaging across stimulus-time-locked EEGepochs result in stable ERPs representing average stimulus processing and ruling out non-relevant brain activity.

The schematic ERP waveform in Figure 1 Positive-and negative-going voltage changes are labeled according to their amplitude polarity and latency from stimulus onset (adapted from [START_REF] Männel | The method of event-related brain potentials in the study of cognitive processes. A tutorial[END_REF].

Different ERP paradigms have been developed to study early language acquisition during passive listening. For example, the Mismatch Negativity paradigm tests auditory discrimination of short stimuli, such as tones, phonemes or syllables, and involves the unexpected presentation of deviant stimuli in a series of repeated, standard stimuli.

Familiarization-test paradigms are more suitable when longer or more complex stimuli are of interest, as is the case in word segmentation studies. The latter usually include a familiarization or learning phase, followed by a test phase with familiar and nonfamiliar elements, so that brain responses time-locked to the onset of the target stimuli can reveal familiarity recognition indicating successful word-form segmentation.

Comparing ERPs elicited by different stimuli allows for inferences about the effect of experimental manipulations [START_REF] Kutas | Psycholinguistics electrified II[END_REF]. Differences in amplitude can be the result of variations in processing demands or efficiency across experimental conditions, whereas latency differences can indicate the slowing down or speeding up of cognitive processing.

The ERP methodology, as a non-invasive and highly infant-suitable neuroimaging technique, has been successfully applied in early word segmentation and word-mapping studies. While its spatial resolution is weak, compared to hemodynamic techniques such as functional near-infrared spectroscopy (fNIRS), it has an excellent temporal resolution, comparable to whole-head magneto-encephalography (MEG). Thus, complementing behavioral data, ERP data have largely contributed to a better understanding of the time course of early word segmentation and mapping. The following sections will review available data from ERP studies, with a focus on research specifically exploring the role of prosodic factors on these crucial abilities for language acquisition.

Word segmentation

Finding the words in fluent speech is a fundamental ability in early language acquisition, linked to later vocabulary outcomes [START_REF] Junge | Rapid recognition at 10 months as a predictor of language development[END_REF]Newman et al., 2006;[START_REF] Singh | Infant word segmentation and childhood vocabulary development: a longitudinal analysis[END_REF].

Although single-word utterances are not exceptional in the input to children, most words do not occur in isolation [START_REF] Brent | The role of exposure to isolated words in early vocabulary development[END_REF]. Infants thus need to segment the speech stream to extract the individual word-forms, exploiting any possible cues to word boundaries present in the speech input (see Thorson, this volume; de Carvalho, Dautriche, Millotte, and Christophe, this volume, for reviews). Behavioral studies indicate that word segmentation ability is present at around 7 to 10 months of age [START_REF] Houston | Crosslanguage word segmentation by 9-month-olds[END_REF][START_REF] Jusczyk | The beginnings of word segmentation in English-learning infants[END_REF][START_REF] Polka | Word segmentation in monolingual infants acquiring Canadian English and Canadian French: native language, cross-dialect, and cross-language comparisons[END_REF]. Word segmentation can be observed even earlier, by 6 months of age, for monosyllabic word-forms [START_REF] Höhle | German-learning infants' ability to detect unstressed closed-class elements in continuous speech[END_REF][START_REF] Bosch | Rapid gains in segmenting fluent speech when words match the rhythmic unit: Evidence from infants acquiring syllable-timed languages[END_REF], and for words following an already familiar name or anchor word [START_REF] Bortfeld | Mommy and me. Familiar names help launch babies into speech-stream segmentation[END_REF]. In this context, ERP studies have provided relevant information on the segmentation process, not only by tracking the online processing, but also by offering a direct measure of the factors favoring or constraining early word-form segmentation. In the next sections, we first describe the electrophysiological indicators of word-form segmentation, followed by a review of ERP studies addressing (a) the role of lexical stress patterns in speech segmentation, (b) the effect of specific features of IDS in modulating the developmental onset of this ability, and (c) the contribution of melodic information (song) to the word segmentation process. [START_REF] Kooijman | Electrophysiological evidence for prelinguistic infants' word recognition in continuous speech[END_REF] designed the first ERP study on word segmentation with Dutch-learning 10-month-olds. They adapted the behavioral paradigm from [START_REF] Jusczyk | Infants' detection of the sound patterns of words in fluent speech[END_REF], familiarizing infants with repetitions of strong-weak words and testing them on passages containing either the familiarized or novel words. Infants' ERPs in the familiarization phase showed a negative-going effect, with more negative ERP responses to the last repetitions compared to the first ones.

ERP indicators of word segmentation

This "familiarity" or repetition effect appeared from 200-500 ms post word onset at fronto-central recording sites. At test, a negative ERP effect over the left hemisphere was found from 350-500 ms, with larger amplitudes for familiarized target words than unfamiliarized words. The authors concluded that these negative ERP components (later termed N200-500, see Figure 2) were related to word-form familiarity through repetition, and recognition of the familiarized words, thus, indicating that segmentation of the target word-forms embedded in sentences had occurred. Comparable negative-going ERP responses have been reported in studies on word recognition in older infants, both for known words [START_REF] Mills | acquisition and cerebral specialization in 20-month-old infants[END_REF][START_REF] Mills | Language comprehension and cerebral specialization from 13 to 20 months[END_REF][START_REF] Thierry | Familiar words capture the attention of 11-month-olds in less than 250 ms[END_REF] and for repetitions of pseudowords [START_REF] Von Koss Torkildsen | Brain dynamics of word familiarization in 20month-olds: Effects of productive vocabulary size[END_REF]. The N200-500 component can thus be taken as an ERP indicator of word segmentation. In the next sections, we review ERP evidence of the impact of different prosodic factors on infants' word segmentation abilities.

Language-specific prosodic cues to word segmentation: lexical stress patterns

Behavioral evidence has revealed that differences in the predominant stress pattern of languages may be responsible for the variation in the onset and the robustness of early word segmentation abilities (see Bhatara, Boll-Avetisyan, Höhle, and Nazzi, this volume). For instance, the metrical segmentation strategy, derived from the lexical stress properties of Dutch as the native language, would predict no difficulties in extracting disyllabic word-forms following the predominant stress pattern (e.g., strongweak in Dutch) and less successful segmentation for the less frequent, weak-strong pattern. But behavioral evidence had failed to yield positive evidence of the expected strong-weak segmentation in 7.5-month-old Dutch-learning infants [START_REF] Kuijpers | Using the head-turning technique to explore cross-linguistic performance differences[END_REF]. A follow-up ERP study, however, adopting the same familiarization-test paradigm previously used with 10-month-olds [START_REF] Kooijman | Electrophysiological evidence for prelinguistic infants' word recognition in continuous speech[END_REF], clarified this issue and confirmed successful segmentation of strongweak words in 7-month-olds [START_REF] Kooijman | Predictive brain signals of linguistic development[END_REF]. Here, the N200-500 component emerged during familiarization, and, in addition, two different ERP patterns indicating familiarity recognition appeared at test. Some infants showed a right-frontal positive ERP effect at 350-450 ms to familiarized versus unfamiliarized words, while others showed a left-posterior negative ERP effect at 430-530 ms. When children's language outcomes were followed up at 3 years, those previously showing a negative-going ERP response obtained higher scores in vocabulary measures. Thus, polarity differences in the ERP could be taken to reflect different strategies underlying infants' word segmentation, linked to maturational variation and related to later vocabulary outcomes.

Evaluating the effect of lexical stress on word segmentation, [START_REF] Kooijman | Prosodic structure in early word segmentation: ERP evidence from Dutch ten-month-olds[END_REF] examined whether Dutch-learning 10-month-olds would also be able to segment words that did not follow the predominant stress pattern, namely weakstrong words. ERPs of the familiarization revealed similar N200-500 effects as reported by [START_REF] Kooijman | Electrophysiological evidence for prelinguistic infants' word recognition in continuous speech[END_REF], a familiarity effect that started while infants were still hearing the weak (first) syllable. Results of the test phase revealed a more variable pattern linked to the strong syllables of the target words.

While infants were able to recognize repetitions of whole words occurring in isolation, they still relied on the strong syllable in more complex sentence contexts, failing to correctly segment the weak-strong words. These results thus differ from the ones obtained by [START_REF] Kooijman | Electrophysiological evidence for prelinguistic infants' word recognition in continuous speech[END_REF], showing that 10-month-old infants recognized the whole strong-weak word and not only the strong syllable. Together, these findings highlight the important role of the predominant lexical-stress pattern of the native language in facilitating word segmentation.

We now turn to cross-linguistic differences in word segmentation, looking at French as a syllable-timed language that does not show word-initial stress, but phrase-final lengthening. [START_REF] Goyet | Words and syllables in fluent speech segmentation by French-learning infants: An ERP study[END_REF] ran an ERP study to further evaluate the delayed disyllabic weak-strong word segmentation reported in a previous behavioral study with French 12-month-olds [START_REF] Nazzi | Early segmentation of fluent speech by infants acquiring French: Emerging evidence for crosslinguistic differences[END_REF]. Again, 12-month-old infants were familiarized with weak-strong words and then tested on sentences containing familiarized and novel words. Results from the familiarization phase showed an enhanced N200-500 occurring at 300-450 ms post word onset for later repetitions.

Similar results were found at test, with the N200-500 effects of word-form familiarity resembling the ones previously reported by [START_REF] Kooijman | Electrophysiological evidence for prelinguistic infants' word recognition in continuous speech[END_REF].

Importantly, however, here they indicate successful word segmentation of weak-strong words by French-learning infants, while the 10-month-old Dutch-learning infants in [START_REF] Kooijman | Prosodic structure in early word segmentation: ERP evidence from Dutch ten-month-olds[END_REF] had failed at segmenting words with this specific stress pattern. While confirming French-learning infants' ability to segment disyllabic weak-strong words at the tested age of 12 months, it remains to be shown whether similar results (i.e., whole-word disyllabic segmentation and no independent finalsyllable segmentation) can be found at an earlier age. Moreover, ERP data from infants acquiring other syllable-timed languages with variable lexical stress patterns (e.g., Catalan and Spanish) are still missing, limiting the overall conclusions about the function of cross-linguistically different prosodic features in infants' emerging segmentation abilities. In summary, the described ERP studies show that word-level prosody affects early word segmentation in several significant ways. First, the fact that the language of interest has a more or less predominant word-stress structure seems to affect the onset of word segmentation. Second, words following the predominant pattern of a given language are segmented earlier than words that do not follow this pattern. Third, crosslinguistic rhythm differences can set some constraints on infants' ability to segment words longer than the smallest, monosyllabic elements. Beyond any cross-linguistic differences reported in the literature on infant word segmentation, it is important to highlight that 10-month-old infants show stable ERP responses indicating successful word segmentation; the N200-500 component, which has been previously reported for word-form familiarity. This finding suggests that infants at this early age are able to build up word-form representations from only a few word repetitions in the speech input. Moreover, ERP studies have identified a developmental transition between 7 and 10 months of age, from a positive-going to a negative-going familiarity effect in the ERP, suggesting different underlying mechanisms modulated by advancing brain maturation and sustained language exposure.

Properties of infant-directed speech (IDS) supporting word segmentation: Prosodic emphasis on sentence-embedded words

Across languages, adults have been shown to use infant-directed speech (IDS) when communicating with infants [START_REF] Fernald | A cross-language study of prosodic modifications in mothers' and fathers' speech to preverbal infants[END_REF]. IDS is described as a speech register with especially salient prosodic features, such as more exaggerated pitch contours and emphasis on specific words, produced with higher pitch and intensity levels and longer duration. Moreover, IDS features a slower speech rate, shorter mean length of utterance, more prosodic repetition, higher frequency of specific words and syntactic patterns, and simpler grammatical structures [START_REF] Fernald | Human maternal vocalizations to infants as biologically relevant signals: An evolutionary perspective[END_REF]. The specific acoustic features of IDS have been found to facilitate language acquisition, and, importantly, word segmentation [START_REF] Thiessen | Infant-directed speech facilitates word segmentation[END_REF]; see also [START_REF] Cristia | Input to language: The phonetics and perception of infant-directed speech[END_REF]Soderstrom, 2007, for reviews).

While infants show an initial strong preference for IDS over ADS, this preference for prosodically enhanced speech seems to reduce during the first year of life [START_REF] Newman | Changes in preference for infant-directed speech in low and moderate noise by 4.5-to 13-month-olds[END_REF]. Moreover, parents have been shown to adjust their IDS register to infants' age, such that IDS features are especially pronounced during infants' first months of life, but become less salient later on [START_REF] Kitamura | Pitch and communicative intent in mother's speech: Adjustments for age and sex in the first year[END_REF][START_REF] Stern | The prosody of maternal speech: Infant age and context related changes[END_REF]). Thus, it seems that both infants' use of IDS in language learning and IDS features provided in the speech input change as a function of infants' age. In this context, [START_REF] Männel | The method of event-related brain potentials in the study of cognitive processes. A tutorial[END_REF]Friederici (2013, 2010), using electrophysiological measures, systematically explored the age-dependent role of different speech cues that are typically used by caregivers in word-learning situations: prosodic accentuation and repetition (see [START_REF] Aslin | Models of word segmentation in fluent maternal speech to infants[END_REF][START_REF] Bernstein-Ratner | From Signal to syntax: But what is the nature of the signal[END_REF][START_REF] Fernald | Prosody and focus in speech to infants and adults[END_REF][START_REF] Fernald | Common themes and cultural variations in Japanese and American mothers' speech to infants[END_REF]. The authors tested German-learning infants at 6, 9 and 12 months of age in a familiarization-test paradigm and manipulated the degree of prosodic modulation and number of repetitions of sentence-embedded target words. Infants were first familiarized with different sentences containing the same target word, presented with or without an additional, naturally produced prosodic emphasis (for acoustic analyses, see [START_REF] Männel | Accentuate or repeat? Brain signatures of developmental periods in infant word recognition[END_REF]. The test phase consisted of tokens of the familiar target word and an unfamiliar control word. ERP responses to sentence-embedded targets collected during familiarization differed depending on infants' age. At 6 months, infants showed a fronto-central positive shift at 200-400 ms post word onset for accentuated versus unaccentuated words, but no effect of repetition. At 9 months, infants also showed an early occurring positive-going ERP effect of prosodic realization and, in addition, an N200-500 word-repetition effect. Finally, 12-month-old infants did not show an effect of prosodic realization on target word processing, but a sustained N200-500 as an effect of repetition. These results suggest that at younger ages, prosodic input cues drive infants' word processing, while with increasing age repetition takes more effect. When infants were subsequently tested on their word recognition, ERP responses to familiarized and unfamiliarized words were compared as a function of prosodic realization during familiarization. As displayed in Figure 3, test results revealed that 6-month-olds were only able to recognize words that had received an additional prosodic emphasis during familiarization, showing a positive-going familiarity response in the ERP, starting at around 500 ms post word onset. Ninemonth-old infants showed two familiarity effects in the ERP: an N200-500 to familiarized versus unfamiliarized words independently of previous prosodic realization; and a late negativity, only for words that were accentuated in the familiarization phase. Finally, 12-month-old infants also showed a sustained N200-500 effect that occurred for all familiarized compared to unfamiliarized words, independently of accentuation. Taken together, these data suggest that infants recognize familiar words, relying on different speech input cues provided by the speaker when teaching infants new words. While younger infants primarily benefit from the additional prosodic emphasis on sentence-embedded words, older infants primarily rely on word repetition, and recognize familiar words independently of their prosodic realization during the learning situation. The ERP study by [START_REF] Männel | Accentuate or repeat? Brain signatures of developmental periods in infant word recognition[END_REF] further contributes to the idea that prosodic information in the speech input enables infants' initial word segmentation, and confirms previous behavioral research showing that infants at 6 months of age make use of prosodic cues aligned with word boundaries to segment disyllabic words [START_REF] Shukla | Prosody guides the rapid mapping of auditory word forms onto visual objects in 6-mo-old infants[END_REF]. Moreover, [START_REF] Männel | Accentuate or repeat? Brain signatures of developmental periods in infant word recognition[END_REF] observed a developmental positivity-negativity transition in the ERP responses indicating word recognition, with the latter indicating a more mature process (see also [START_REF] Kooijman | Electrophysiological evidence for prelinguistic infants' word recognition in continuous speech[END_REF][START_REF] Kooijman | Predictive brain signals of linguistic development[END_REF]. This transition in the ERP markers of infant word segmentation can be attributed to infants' growing linguistic experience and parallel brain maturation. More specifically, at around 6 months of age, myelination and neurogenesis of the auditory cortex undergo tremendous changes [START_REF] Moore | Cytoarchitectural and axonal maturation in human auditory cortex[END_REF][START_REF] Moore | The human auditory system: a timeline of development[END_REF] that can influence the latency and polarity of early sensory ERP components (see [START_REF] Eggermont | Morphological and functional development of the auditory nervous system[END_REF].

These maturational changes may also affect the polarity of infants' word recognition responses. Thus, the current example demonstrates how ERP studies additionally deliver information about the underlying brain mechanisms that trigger infants' word segmentation abilities.

From IDS to song: How early can melodic cues contribute to word segmentation?

Very early in life, infants are exposed to both language and music [START_REF] Brandt | Music and early language acquisition[END_REF], and both domains share important similarities in terms of structural information and involved cerebral resources [START_REF] Heffner | Prosodic structure as a parallel to musical structure[END_REF][START_REF] Patel | Science & music: talk of the tone[END_REF]. But, is music, and specifically song, relevant for speech segmentation and word learning? Song and IDS registers exhibit several similar acoustic and structural characteristics [START_REF] Fernald | Human maternal vocalizations to infants as biologically relevant signals: An evolutionary perspective[END_REF][START_REF] Papousek | The naturalistic vocal environment of young infants: On the significance of homogeneity and variability in parental speech[END_REF]. For example, many of the IDS features, such as exaggerated pitch contours, emphasis on specific words, higher pitch and intensity levels, longer duration, slower rate, shorter utterances and repetition, are also found in songs for infants. Infants show a clear preference for ID songs over AD songs [START_REF] Unyk | Lullabies and simplicity: A cross-cultural perspective[END_REF], and ID melodic information has been shown to specifically benefit speech processing as well as language acquisition [START_REF] Gervain | Prosody cues word order in 7-month-old bilingual infants[END_REF][START_REF] Lebedeva | Sing that tune: Infants' perception of melody and lyrics and the facilitation of phonetic recognition in songs[END_REF][START_REF] Schön | Songs as an aid for language acquisition[END_REF][START_REF] Thiessen | How the melody facilitates the message and vice versa in infant learning and memory[END_REF].

Assessing a beneficial effect of song on word segmentation, Schön and colleagues (2008) compared segmentation of a spoken, flat-contour stream and a sung stream in adults. While the level of performance was at chance after exposure to the spoken stream, successful word recognition was observed following exposure to the sung stream. These results suggest that redundancy in statistical musical and linguistic structures benefits segmentation. Based on these findings in adults [START_REF] Schön | Songs as an aid for language acquisition[END_REF] and evidence showing that prosodic cues facilitate segmentation in 7-month-old infants [START_REF] Thiessen | Infant-directed speech facilitates word segmentation[END_REF] and even neonates [START_REF] Bosseler | Infantdirected speech enhances statistical learning in newborn infants: An ERP study[END_REF], it has been recently tested whether melodic information of ID song may benefit speech segmentation in newborns [START_REF] François | Enhanced neonatal brain responses to sung streams predict vocabulary outcomes by age 18 months[END_REF]. The authors recorded EEG in a group of 2-day-old sleeping neonates during a familiarization-test paradigm with continuous artificial speech streams that were either spoken or sung. In the spoken condition, the continuous streams consisted of 100 repetitions of four pseudowords, with transitional probabilities between adjacent syllables as the only cues to word boundaries. In the sung condition, the language streams were identical except that each syllable had a unique pitch from the diatonic scale. Therefore, the resulting sung streams contained two types of overlapping cues signaling word boundaries: the statistical cues (drop in transitional probabilities between adjacent syllables) together with the pitch interval changes that systematically occurred at word boundaries only. In each condition, spoken and sung, newborns were first exposed to a 3.5 min familiarization stream followed by a test phase in which test items that violated the statistically-defined syllabic order of the words were inserted at random positions allowing for an evaluation of the implicit detection of structural violations.

During familiarization, a comparison between the ERPs of the first and the second half revealed the presence of a fronto-central negative deflection at 400-600 ms post word onset in both spoken and sung conditions. However, while in the spoken condition the fronto-central negativity reached maximum amplitude during the second half, its amplitude was already at maximum during the first half in the sung condition, suggesting a faster segmentation process. To determine whether familiarization was sufficient to establish pseudoword representations and detect violations of syllabic order in the test phase, ERPs to the newly learned pseudowords were compared to those elicited by the violation items. Evidence of successful detection of structural violations was found in the sung condition, but not in the spoken condition. Compared to the newly learned pseudowords, test items violating the syllabic structure elicited an early positivity in the 250-400 ms time window. This positivity resembled the mismatch response consistently observed in infants for speech and non-speech stimuli (see [START_REF] Cheour-Luhtanen | Mismatch negativity indicates vowel discrimination in newborns[END_REF][START_REF] Mueller | Auditory perception at the root of language learning[END_REF][START_REF] Stefanics | Newborn infants process pitch intervals[END_REF]. Thus, the test results suggest that sung streams facilitated the building of robust word-form memory traces and triggered strong enough expectations to correctly detect online structural violations. In addition, these results confirm the importance of the fastemerging fronto-central negativity during the learning phase for subsequent word-form recognition, as neonates were sensitive to structural violations in the sung condition, but not in the spoken one. These results provide direct evidence of the benefit of songs during the initial steps of language acquisition, thus highlighting the relevance of both speech prosody and melodic information to break the speech code and begin identifying basic linguistic units. Moreover, the benefit of song on speech segmentation supports the idea that musical interventions might be a powerful tool to facilitate language acquisition [START_REF] Flaugnacco | Music training increases phonological awareness and reading skills in developmental dyslexia: a randomized control trial[END_REF][START_REF] François | Musical training as an alternative and effective method for neuro-education and neurorehabilitation[END_REF][START_REF] Zhao | Musical intervention enhances infants' neural processing of temporal structure in music and speech[END_REF].

Word-referent mapping

So far, we have focused on research evaluating how infants extract word-forms from fluent speech, greatly aided by prosodic cues. Word learning, however, goes beyond word segmentation, because newly segmented word-forms will eventually become labels for objects, people, and events in children's environment. Thus, establishing word-referent associations and storing these as memory representations are crucial steps children have to accomplish in building a first receptive vocabulary.

Behavioral studies have shown that starting at 6-8 months of age, infants are able to successfully establish word-to-world mappings in laboratory settings [START_REF] Gogate | Attention to maternal multimodal naming by 6-to 8-month-old infants and learning of word-object relations[END_REF][START_REF] Matatyaho | Type of maternal object motion during synchronous preverbal infants' learning of word-object relations[END_REF], and even at 2 months of age when presented with distinct syllable-object motion pairings [START_REF] Gogate | Two-month-old infants' sensitivity to changes in arbitrary syllable-object pairings: The role of temporal synchrony[END_REF]. Regarding semantic representations, [START_REF] Bergelson | At 6-9 months, human infants know the meanings of many common nouns[END_REF] even found that 6-month-old infants already know the meaning of very frequent words, having established some word-object associations from their daily experience. These abilities have also been explored with the ERP method in infants at different ages, offering further insight into the underlying brain mechanisms and their developmental progression. In the following sections, we will first review ERP evidence of infants' early word knowledge and their ability to establish novel word-object associations. We will then turn to the discussion of the role of prosody in infants' word-referent mapping skills, revealing the scarcity of ERP studies in this field and the need for further neuroimaging research.

ERP indicators of word-referent mapping

Friedrich and [START_REF] Friedrich | N400-like semantic incongruity effect in 19month-olds: Processing known words in picture contexts[END_REF] were the first to investigate infants' brain mechanisms underlying the processing of words in meaningful contexts. To tackle the link between words and their referents by means of ERPs, the authors used a crossmodal picture-word priming paradigm with German-speaking 19-month-olds and adults (see also Friedrich and Friederici 2005a;2005b;2008;2011;2015; for review, see [START_REF] Männel | The method of event-related brain potentials in the study of cognitive processes. A tutorial[END_REF]. Participants were presented with pictures of objects and accompanying IDS words that correctly or incorrectly labeled the objects. Crucially, the objects' presentation preceded the label onset, such that the picture content could prime subsequent word processing. Thus, if participants knew the objects (from previous experimental familiarization or existing memory representations), this knowledge could trigger the expectation of the corresponding word and thus ease phonological wordform processing and lexical-semantic processing upon the label's presentation.

The adult ERP data collected in the picture-word priming paradigm revealed two processes indicating successful word processing by priming from the preceding picture context [START_REF] Friedrich | N400-like semantic incongruity effect in 19month-olds: Processing known words in picture contexts[END_REF]): First, facilitated phonological word-form processing as reflected in a frontal N200-500 effect (i.e., word-form familiarity) for picture-matching words compared to non-matching ones, proving established wordobject associations. And second, successful lexical-semantic word comprehension as reflected in an N400 effect (i.e., semantic integration efforts) for words not matching the picture compared to matching ones, due to semantic expectancy violations given referential connections between words and the semantic picture context. The N400 describes a negativity that occurs at around 400 ms relative to stimulus onset and is more pronounced the more semantically unfamiliar, unexpected, or non-matching an event is, given the current semantic context or semantic knowledge (see [START_REF] Holcomb | Semantic priming and stimulus degradation: Implications for the role of the N400 in language processing[END_REF]. Evidence of the N400 in infancy research would thus be a valid indicator of infants' emerging ability to understand the referential meaning of words. It is important to note that although both N200-500 and N400 effects are partly overlapping in their time of occurrence, they can still be differentiated based on their different scalp distributions: The N200-500 typically occurs fronto-centrally, while the N400 occurs broadly distributed over the scalp, with a centro-parietal focus. Crucially, using the same cross-modal picture-word priming paradigm in 19-montholds, [START_REF] Friedrich | N400-like semantic incongruity effect in 19month-olds: Processing known words in picture contexts[END_REF] observed comparable ERP effects in infants as reported for adults. Infants did not only show ERP indicators of word-form familiarity from cross-modal phonological priming (N200-500), but also the establishment of referential meaning (N400) (Figure 4). When analyzing these ERP effects in light of children's lexical knowledge (as reported by their parents), there was no correlation of children's receptive vocabulary and the N200-500 effect, but a correlation with the N400-like effect. In other words, only infants with higher comprehension abilities showed an N400 similar to the one found in adults, confirming the interpretation of this infant ERP component as indicating referential processes.

Following this seminal work, Friedrich and colleagues conducted a series of ERP experiments in German exploring the developmental progression of the emerging ability to form referential connections during early infancy. In these follow-up ERP studies, the authors were able to show that even 14-month-olds displayed recognition of word-forms (N200-500) and were able to form referential connections between objects and labels (N400) similarly to older infants and adults (Friedrich and Friederici, 2005b;2008; Figure 4). However, younger infants at 12 and 6 months of age, while initiating word-object associations both phonologically (N200-500) and semantically (N400) during the familiarization phase, only showed phonologically-driven associations (N200-500) when tested later, suggesting memory limitations preventing full mapping representations (Friedrich and Friederici, 2005a;2011;Figure 4). Finally, 3-month-old infants were found to generate word-form representations during familiarization (N200-500), as well as an early perceptually-driven form of association between objects and words [START_REF] Friedrich | The origins of word learning: Brain responses of three-month-olds indicate their rapid association of objects and words[END_REF]. The latter ERP effect appeared as centro-parietal negativity at 500-1000 ms post word onset to matching versus nonmatching picture-word pairs. This late centro-parietal negativity might indicate a precursor learning mechanism that enables infants to establish first connections between the perceptual representations of objects and words, forming their first proto-words (see [START_REF] Nazzi | Before and after the vocabulary spurt: two modes of word acquisition[END_REF]. However, 3-month-olds did not reveal any sign of retention of these associations when tested one day later, making evident the difficulties infants at this age have in retaining this information beyond the immediate encoding stage. Together, these results suggest that the neural mechanisms underlying the building of a first vocabulary are present from early on, while the capacity for creating full referential connections and their successful retention is still highly constrained below 14 months of age.

In summary, the existing ERP studies on word-referent mapping across infancy have reliably revealed two ERP components: First, a fronto-centrally distributed N200-500 that indicates word-form familiarity and evidences word-object associations at a phonological level. Second, a centro-parietal N400 that indicates semantic integration efforts and evidences word-object mapping at a semantic level. Importantly, the association and reference processes underlying these components seem to work semiindependently, and show a different developmental onset. Thus, the reported findings impressively illustrate how the use of electrophysiological measures at early ages not only enables us to successfully delineate the developmental onset and progression of infants' mapping abilities, but also to differentiate the underlying processes.

The contribution of prosody to word-referent mapping: From behavioral to ERP studies

Comparable to behavioral research exploring the role of prosodic features in infants' successful word segmentation, there are an impressive number of behavioral studies addressing the effect of prosody in word-object mapping. Factors such as accentuation [START_REF] Grassmann | Two-year-olds use primary sentence accent to learn new words[END_REF][START_REF] Kuhl | Brain mechanisms in early language acquisition[END_REF], lexical stress [START_REF] Curtin | Mapping novel labels to actions: How the rhythm of words guides infants' learning[END_REF][START_REF] Graf-Estes | Learning about sounds contributes to learning about words: Effects of prosody and phonotactics on infant word learning[END_REF], specific pitch accents [START_REF] Thorson | Acoustic correlates of information structure in child and adult speech[END_REF], and the role of IDS [START_REF] Graf-Estes | Infant-directed prosody helps infants map sounds to meanings[END_REF][START_REF] Ma | Word learning in infant-and adult-directed speech[END_REF] have been analyzed and connected to word mapping abilities (see Thorson, this volume). For example, Graf-Estes and Hurley (2013) examined whether the pronounced prosodic features of IDS would enhance word-object associations in 17.5-month-olds. The authors habituated infants to pairs of novel objects and novel words, either in IDS or ADS. Looking-time measures revealed that infants were only able to map words onto objects for labels produced in IDS, but not in ADS.

Crucially, the advantage of IDS only took effect when the realization of IDS labels involved prosodic variability. Focusing on a different prosodic property, [START_REF] Curtin | Mapping novel labels to actions: How the rhythm of words guides infants' learning[END_REF] found that lexical stress cues can also guide labelreferent associations. In their study, English-learning 16-month-olds were already showing a bias to associate weak-strong patterns with action labels, a language-specific bias consistent with the stress pattern properties of their native language. Thus, behavioral research has delivered significant indications of prosodic cues also driving infants' initial word mapping skills. However, a systematic examination of how prosodic input features affect referential processes by means of electrophysiological measures is still missing. As exemplified above, ERP studies on infants' word-referent mapping have used IDS material [START_REF] Friedrich | N400-like semantic incongruity effect in 19month-olds: Processing known words in picture contexts[END_REF]2005a;2005b;2006;2008;2011). Yet, these studies have never directly contrasted the effect of IDS and ADS, or manipulated other prosodic speech input features, despite promising behavioral evidence. Thus, ERP studies aiming at exploring the impact of prosody on infants' developing mapping skills are clearly needed.

Another line of ERP research that needs to be strengthened is the investigation of word segmentation and mapping as a joint process, also assessing the impact of speech input features. Naturally, when infants start to segment word-forms from fluent speech, they eventually conceive them as labels to referents in their surrounding world. Recent behavioral research has shown that from an early age, infants can simultaneously segment and map simple, statistically defined word-forms, but only if prosody favors this process [START_REF] Shukla | Prosody guides the rapid mapping of auditory word forms onto visual objects in 6-mo-old infants[END_REF]. Investigating simultaneous segmentation and mapping in a more ecological learning situation, [START_REF] Teixidó | Early segmentation and word mapping in a natural language: evidence from 4-, 6-and 9-months-olds[END_REF] tested 4-, 6-and 9-month-old infants on natural language stimuli. Interestingly, the authors observed successful word segmentation and mapping only for infants at 9 months, but not at younger ages. Here, a follow-up ERP study has set out to explore the underlying neural mechanisms driving the success (or failure) of infants' segmentation and mapping skills at young ages [START_REF] Männel | Sentence prosody cues object category learning at 6 months[END_REF].

Importantly, this study manipulated the prosodic realization of target words by placing them at different sentence positions. Preliminary ERP results indicate that while positioning did not affect infants' segmentation ability, it impacted on infants' mapping success, because only targets at prosodically salient edge positions enabled mapping.

This ERP study represents a relevant first research step, yet there are clearly more studies needed that assess the neural mechanisms supporting infants' emerging mapping skills and the role played by prosody at different developmental stages.

Conclusions

In this chapter, we have reviewed the role of prosody in infants' word learning, evaluating their emerging abilities of word segmentation and word-referent mapping.

Here we have focused on electrophysiological studies that capture the underlying neural mechanisms of infants' emerging abilities, accessible from an early age, when behavioral assessments can be challenging. Current evidence from ERP studies suggests that there are at least two different electrophysiological indicators related to infant word learning. First, an N200-500 component, that is, a word-form familiarity effect characterized by a fronto-lateral negativity in the ERP at 200-500 ms post word onset, elicited when a given phonological sequence is familiar. Second, an N400 component, that is, a semantic (violation) effect characterized by a centro-parietal negativity at around 400 ms post word onset, elicited when the meaning of a word does not match the semantic (picture) context. Importantly, the developmental onset and the amplitude of these two ERP components can predict later language development, particularly children's vocabulary. Crucial for developmental research, both ERP components can be utilized as indicators of infants' emerging word learning abilities to draw a more fined-grained picture of the progression of word-form segmentation and the establishment of referential meaning across infancy. Over and above the illustration of infants' advancing word learning skills by means of ERPs, the data presented here have highlighted the idea that prosody is a key factor in getting infants started on detecting word forms in fluent speech and conceiving them as referents to their surrounding world.

  displays a sequence of positive-and negative-going stimulus-triggered voltage changes. The labeling of the different ERP components indicates their polarity and the latency of maximum amplitude relative to the onset of the stimulation. The N1(00) component, for example, refers to a negativity that can be observed around 100 ms after stimulus onset. In general, ERP components occurring early after stimulus onset (around 50-200 ms) are mainly modulated by sensory input features, while those occurring later (after 300 ms) reflect cognitive processes to a much larger degree.

Figure 1 .

 1 Figure 1. Schematic ERP waveform representing the average response to a class of stimuli.

Figure 2 .

 2 Figure 2. ERP responses of 12-month-olds to familiarized and unfamiliarized test words in a familiarization-test paradigm (modified from Männel and Friederici, 2013). L/RF = left/right frontal region, L/RC = left/right central region, L/RP = left/right posterior region.

Figure 3 .

 3 Figure 3. Developmental changes of ERP responses indicating word-form recognition as a function of prosodic manipulation during familiarization (adapted from Männel and Friederici, 2013). RF = right frontal region.

Figure 4 .

 4 Figure 4. ERP responses for word processing in a cross-modal picture-word priming paradigm at electrodes F7 and PZ (adapted from Friederici, 2005; Friedrich and Friederici 2004; Friedrich and Friederici, 2005a, b).

Acknowledgments

The present chapter was supported by the Spanish Ministerio de Ciencia e Innovación (projects PSI-2014-55105-P to L.B. and PSI-2015-69132-P to C.F.), the German Research Foundation (project MA 6897/2-1 to C.M.) and the Max Planck Society (C.M.). We would like to thank the editors and two anonymous reviewers for their helpful comments and editorial improvements.


Aurélie Marion
email: aurelie.marion@etu.univ-amu.fr

Julien Morin

Adrien Gandolfo

Elena Ormeño

Barbara D'anna

Henri Wortham

Nitrous acid formation on Zea mays leaves by heterogeneous reaction of nitrogen dioxide in the laboratory

Keywords: HONO sources, hydroxyl radicals, atmospheric chemistry, solid/gas reactivity, leaf surface

Nitrous acid (HONO) is of considerable interest because it is an important precursor of hydroxyl radicals (OH), a key species in atmospheric chemistry. HONO sources are still not well understood, and air quality models fail to predict OH as well as HONO mixing ratios. As there is little knowledge about the potential contribution of plant surfaces to HONO emission, this laboratory work investigated HONO formation by heterogeneous reaction of NO2 on Zea mays.

Experiments were carried out in a flow tube reactor; HONO, NO2 and NO were measured online with a Long Path Absorption Photometer (LOPAP) and a NOx analyzer. Tests were performed on leaves under different conditions of relative humidity (5 to 58 %), NO2 mixing ratio representing suburban to urban areas (10 to 80 ppbv), spectral irradiance (0 to 20 W m -2 ) and temperature (288 to 313 K). Additional tests on plant wax extracts from Zea mays leaves showed that this component can contribute to the observed HONO formation. Temperature and NO2 mixing ratios were the two environmental parameters that showed substantially increased HONO emissions from Zea mays leaves. The highest HONO emission rates on Zea mays leaves were observed at 313 K for 40 ppbv of NO2 and 40% RH and reached values of (5.6 ± 0.8) × 10 9 molecules cm -2 s -1 . Assuming a mixing layer of 300 m, the HONO flux from Zea mays leaves was estimated to be 171 ± 23 pptv h -1 during summertime, which is comparable to what has been reported for soil surfaces.

Introduction

Since its first atmospheric measurement in 1979 by [START_REF] Perner | Detection of nitrous acid in the atmosphere by differential optical absorption[END_REF], nitrous acid (HONO) has become of considerable interest in atmospheric chemistry, as its photolysis (R1) is an important source of hydroxyl radicals (OH), a key species in atmospheric reactivity and photooxidative pollution episodes [START_REF] Acker | Strong daytime production of OH from HNO2 at a rural mountain site[END_REF][START_REF] Alicke | Impact of nitrous acid photolysis on the total hydroxyl radical budget during the limitation of oxidant production/Pianura Padana Produzione di Ozono study in Milan[END_REF][START_REF] Kleffmann | Daytime formation of nitrous acid: A major source of OH radicals in a forest[END_REF].

HONO + hν → NO + OH (R1)

To model and predict OH formation, it is essential to identify and estimate HONO emission sources in the atmosphere, as it can contribute up to 55% of the atmospheric OH radicals during the morning hours [START_REF] Elshorbany | Oxidation capacity of the city air of Santiago, Chile[END_REF]. Highly contrasting HONO mixing ratios were observed between rural environments with values of a few pptv [START_REF] Honrath | Vertical fluxes of NOx, HONO, and HNO3 above the snowpack at Summit, Greenland[END_REF][START_REF] Li | Exploring the atmospheric chemistry of nitrous acid (HONO) at a rural site in Southern China[END_REF][START_REF] Liao | Observations of HONO by laser-induced fluorescence at the South Pole during ANTCI 2003[END_REF][START_REF] Zhou | Snowpack photochemical production of HONO: A major source of OH in the Arctic boundary layer in springtime[END_REF] and polluted urban areas with values of several ppbv at night [START_REF] Huang | Concentration and sources of atmospheric nitrous acid (HONO) at an urban site in Western China[END_REF][START_REF] Li | Impacts of HONO sources on the photochemistry in Mexico City during the MCMA-2006/MILAGO Campaign[END_REF][START_REF] Michoud | Study of the unknown HONO daytime source at a European suburban site during the MEGAPOLI summer and winter field campaigns[END_REF]. The major HONO sources currently considered in atmospheric models include heterogeneous processes (the main pathway for HONO formation), homogeneous gas-phase reactions, and direct emissions (e.g., on-road transport, biomass burning) [START_REF] Aumont | Contribution of HONO sources to the NOx/HOx/O3 chemistry in the polluted boundary layer[END_REF][START_REF] Michoud | Study of the unknown HONO daytime source at a European suburban site during the MEGAPOLI summer and winter field campaigns[END_REF][START_REF] Vogel | Measured and simulated vertical profiles of nitrous acid-Part II. Model simulations and indications for a photolytic source[END_REF].

Modeling studies have shown discrepancies between predicted HONO mixing ratios and ambient measurements, indicating missing sources, estimated to account for 2.6 ppbv h -1 of HONO [START_REF] Spataro | Occurrence of atmospheric nitrous acid in the urban area of Beijing (China)[END_REF]. This unknown source correlates with NO2 mixing ratios and reaches a maximum value at, approximately, midday [START_REF] Acker | Strong daytime production of OH from HNO2 at a rural mountain site[END_REF][START_REF] Kleffmann | Daytime formation of nitrous acid: A major source of OH radicals in a forest[END_REF][START_REF] Li | Exploring the atmospheric chemistry of nitrous acid (HONO) at a rural site in Southern China[END_REF][START_REF] Michoud | Study of the unknown HONO daytime source at a European suburban site during the MEGAPOLI summer and winter field campaigns[END_REF][START_REF] Spataro | Occurrence of atmospheric nitrous acid in the urban area of Beijing (China)[END_REF][START_REF] Villena | Nitrous acid (HONO) during polar spring in Barrow, Alaska: A net source of OH radicals[END_REF][START_REF] Zhou | Summertime nitrous acid chemistry in the atmospheric boundary layer at a rural site in New York State: summertime nitrous acid chemistry[END_REF]. As a result, additional sources were proposed, such as heterogeneous conversion of HNO3 [START_REF] Yang | Significant HONO formation by the photolysis of nitrates in the presence of humic acids[END_REF] and orthonitrophenol photolysis [START_REF] Bejan | The photolysis of ortho-nitrophenols: a new gas phase source of HONO[END_REF], but both processes were discredited. Indeed, for realistic values of HNO3 concentrations and relative humidity, the proposed reaction was too low and could not explain the observed HONO [START_REF] Kleffmann | Heterogeneous reaction of nitric acid with nitric oxide on glass surfaces under simulated atmospheric conditions[END_REF]. Moreover, considering the low ambient concentrations of nitrophenols, this source is unlikely to produce the amount of HONO detected [START_REF] Li | Impacts of HONO sources on the photochemistry in Mexico City during the MCMA-2006/MILAGO Campaign[END_REF]. [START_REF] Zhou | Nitric acid photolysis on surfaces in low-NOx environments: Significant atmospheric implications: nitric acid photolysis on surface[END_REF] suggested nitric acid (HNO3) photolysis as a possible HONO source, and a modeling study in the United States confirmed that this reaction pathway is important and could represent up to 32% of HONO diurnal levels [START_REF] Sarwar | A comparison of CMAQ HONO predictions with observations from the Northeast Oxidant and Particle Study[END_REF]. When adding the gas phase reaction between excited NO2 and water [START_REF] Li | Atmospheric hydroxyl radical production from electronically excited NO2 and H2O[END_REF] into the model, only slight improvements were achieved because 25% of the observed HONO was missing. [START_REF] Li | Impacts of HONO sources on the photochemistry in Mexico City during the MCMA-2006/MILAGO Campaign[END_REF] added in their model the NO2 reaction on semivolatile organics adsorbed on atmospheric particles, and this implementation enabled them to correctly evaluate HONO production in Mexico city. More recently, abiotic release from soil [START_REF] Donaldson | Soil surface acidity plays a determining role in the atmospheric-terrestrial exchange of nitrous acid[END_REF], as well as biological processes such as nitrification or denitrification, were proposed to be this unknown HONO source [START_REF] Oswald | HONO emissions from soil bacteria as a major source of atmospheric reactive nitrogen[END_REF][START_REF] Scharko | Combined flux chamber and genomics approach links nitrous acid emissions to ammonia oxidizing bacteria and archaea in urban and agricultural soil[END_REF][START_REF] Wu | Soil HONO emissions at high moisture content are driven by microbial nitrate reduction to nitrite: tackling the HONO puzzle[END_REF]. HONO fluxes were measured above a maize field by [START_REF] Laufs | Diurnal fluxes of HONO above a crop rotation[END_REF], who then proposed the heterogeneous reaction of NO2 on soil as a major HONO source. Nevertheless, field measurements in Finland reported higher HONO mixing ratios above the canopy than close to the ground [START_REF] Oswald | A comparison of HONO budgets for two measurement heights at a field station within the boreal forest in Finland[END_REF], suggesting that vegetative surfaces could play a role in HONO formation through the heterogeneous reduction of NO2.

It is generally assumed that HONO formation on various surfaces is driven by NO2 hydrolysis (R2), according to the following equation [START_REF] Jenkin | Laboratory studies of the kinetics of formation of nitrous acid from the thermal reaction of nitrogen dioxide and water vapour[END_REF][START_REF] Kleffmann | Heterogeneous NO2 conversion processes on acid surfaces: possible atmospheric implications[END_REF][START_REF] Ramazan | The photochemical production of HONO during the heterogeneous hydrolysis of NO2[END_REF][START_REF] Svensson | Kinetics of the reaction between nitrogen dioxide and water vapour[END_REF]:

2 NO2 + H2O → HONO + HNO3 (R2)
Several studies demonstrated that HONO could also be formed by photoinduced processes through the generation of intermediate species that act as efficient electron donors [START_REF] Cazoir | Heterogeneous photochemistry of gaseous NO2 on solid fluoranthene films: A source of gaseous nitrous acid (HONO) in the urban environment[END_REF][START_REF] George | Photoenhanced uptake of gaseous NO 2 on solid organic compounds: a photochemical source of HONO?[END_REF][START_REF] Stemmler | Light induced conversion of nitrogen dioxide into nitrous acid on submicron humic acid aerosol[END_REF].

In France, agricultural surfaces represent approximately 33 million hectares, while wooded areas account for 19 million ha [START_REF] Moreau | L'occupation des sols en France : progression plus modérée de l'artificialisation entre 2006 et 2012[END_REF]. Depending on the season, the foliage in rural or cultivated suburban areas can be very dense, implying a considerably higher available surface than that provided by the soil. If foliage surfaces were involved in the heterogeneous conversion of NO2 to HONO, it would be important to quantify this emission and to evaluate its impact on the OH budget in agricultural or rural areas.

The main aim of this study was to quantify HONO formation on Zea mays leaves by investigating the influence of environmental factors such as relative humidity, NO2 mixing ratio, reaction temperature, and photon flux on the emission rate. Zea mays leaves were chosen as a model for cultivated vegetation because they are one of the most common cultures in the world and represent a cultivated surface of more than 6 million ha in Europe (Eurostat, 2019;[START_REF] Wrigley | Encyclopedia of Food Grains[END_REF]. Then, HONO fluxes from leaves were estimated to evaluate the potential contribution of vegetation to the missing HONO sources. To our knowledge, this is the first work reporting a systematic investigation of HONO formation on plant leaves under simulated environmental conditions.

Materials and methods

Chemical reagents

Synthetic air (≥ 99.999%, Linde gas) and NO2 (100 ppmv in helium, Linde gas) were used in Teflon tubes (perfluoroalkoxy). The air flow was humidified with Milli-Q water (18 MΩ cm).

Long path absorption photometer (LOPAP) solutions were prepared with sulfanilamide (≥ 99%, Sigma Aldrich), N-(1-naphthyl) ethylene di-amine dihydrochloride (≥ 98%, Sigma Aldrich), sodium hydroxide pellets (≥ 98%, Sigma Aldrich) and hydrochloric acid (37%, Fisher). A nitrite standard solution (NaNO2 in H2O, 1000 mg L -1 , Merck) was used for LOPAP calibration.

Surface proxies were prepared using chlorophyll a (United States Pharmacopeia (USP)

Reference Standard, Sigma Aldrich), diethyl ether (≥ 99.5%, Sigma Aldrich), acetone (HPLC grade, Carlo Erba), and n-hexane (99%, Carlo Erba).

Experimental setup

Experiments were carried out in a cylindrical flow tube reactor (V = 130 cm 3 ) made of borosilicate glass. Gas flows, hereafter referred to as Fi, were controlled using mass flow controllers (Brooks SLA Series, accuracy ± 1%). The experimental setup is depicted in Figure 1 and Figure S2. A 3D sketch of the system was previously presented [START_REF] Gandolfo | The influence of wall temperature on NO2 removal and HONO levels released by indoor photocatalytic paints[END_REF], and a photo of the flow tube reactor is shown in Figure S1. Synthetic air was continuously injected in the reactor as a sheath flow (F4; 990 mL min -1 ), while NO2 gas was introduced via a movable injector (F2; 10 mL min -1 ). The total flow (F2 + F4 = 1 L min -1 ) was chosen to ensure laminar flow in the reactor. Before introduction into the movable injector, NO2 gas (F1) was diluted in synthetic air (F3) with variable flows to adjust the desired NO2 concentration. The total flow in the reactor was kept constant, and the contact time between the reactant gas and the leaf surface was controlled by varying the injector position.

Upstream of the reactor, the sheath flow (F4) was split into two fluxes controlled by two needle valves. The first one consisted of dry air gas, while the second one was humidified by bubbling in deionized water. Mixing these two flows at different ratios generated a sheath flow at controlled RH. Downstream of the reactor, a hygrometer "Hygrolog NT2" (Rotronic) with a "HygroClip SC04" probe was used to measure the resulting relative humidity (accuracy = 1.5% RH). For these experiments, the RH ranged between 5 and 58%. The flow tube reactor was thermostated using a circulating water bath (Lauda RC6 refrigerated bath with RCS thermostat, temperature accuracy ± 0.1 K at 263 K) connected to the double wall of the reactor.

The reactor was placed in a stainless-steel box containing six UV fluorescent lamps (Philips TL-D18W, 340-400 nm, λmax = 368 nm) that could be switched on/off individually. The integrated irradiance of the six lamps in the wavelength region 340-400 nm was 20 W m -2 , which is of the same order of magnitude as the solar irradiance during a spring sunny day in southern France.

At the reactor outlet, two analyzers (LOPAP and NOx monitor) were connected for online detection of HONO and NOx (NOx = NO + NO2). The LOPAP analyzer (QUMA) has been previously described in detail elsewhere [START_REF] Heland | A new instrument to measure gaseous nitrous acid (HONO) in the atmosphere[END_REF][START_REF] Heland | A new instrument to measure gaseous nitrous acid (HONO) in the atmosphere[END_REF]. Briefly, gaseous HONO is dissolved in a hydrochloric acid solution and then converted into an azo dye, detected by long path absorption using a mini spectrometer equipped with a diode array detector. The detection limit (twice the noise) was approximately 2 pptv, and the total accuracy was ± 10%. The instrument was calibrated with a nitrite standard solution following the manufacturer's protocol. The NOx analyzer was an Eco Physics model (CLD 88p) associated with an Eco

Physics photolytic (metal-halide lamp; 180 W) converter (PLC 860) used to alternatively monitor NO and NOx mixing ratios. NO was first analyzed by chemiluminescence (reaction of NO with ozone). Then, NOx was measured after NO2 photoconversion to NO and reaction with ozone. Finally, NO2 was calculated from the difference between the NOx and NO mixing ratios.

The detection limit was 50 pptv, and the relative uncertainty was approximately 1%.

Downstream of the reactor, an air flow (F5) of 1 L min -1 was added to achieve the flow rate needed for the two analyzers.

Experimental design

The experiments were carried out on fully expanded leaves of Zea mays plants (hybrid MB866)

provided by INREA (Institut National de recherche pour l'agriculture, l'alimentation et l'environment) and grown in the greenhouse facilities of Aix-Marseille University. Leaves were harvested at mid-height 10 to 20 weeks after planting at stage V6. A cut leaf was introduced in the reactor and adapted to cover a glass flat surface (2 cm width and 30 cm length) within the reactor. To assure physical integrity of the reactive surface, a mature cut leaf was used every day. Since HONO production from glass surfaces has been previously reported [START_REF] Finlayson-Pitts | The heterogeneous hydrolysis of NO2 in laboratory systems and in outdoor and indoor atmospheres: an integrated mechanism[END_REF][START_REF] Jenkin | Laboratory studies of the kinetics of formation of nitrous acid from the thermal reaction of nitrogen dioxide and water vapour[END_REF][START_REF] Ramazan | The photochemical production of HONO during the heterogeneous hydrolysis of NO2[END_REF], blank experiments were carried out in the reactor replacing the leaf with a glass plate. As a result, hereafter, all the graphs present blank and Zea mays leaf experiments.

Leaf exposure to UV light promoted large emissions of water molecules for several hours.

Consequently, a new protocol was implemented where the leaves were cut 24 hours before the experiment and irradiated for approximately 8 h. Water release from plant leaves is a wellknown phenomenon that results from passive transpiration induced by hydropassive stomatal opening (Franks et al., 1998(Franks et al., , 1995;;Raschke, 1970). After this treatment, the plant leaves were used for kinetic investigations.

Additionally, experiments were carried out using commercial chlorophyll a and wax extracts from Zea mays leaves as reactive surfaces to better understand the origin of HONO emission on the studied leaves. The chlorophyll a film was prepared by dissolving chlorophyll a in acetone. Then, 4 mL of this solution was immediately deposited on a glass plate (less than 2 min after solution preparation) and placed in the dark under a fume cupboard until complete solvent evaporation. The experiments were carried out at three chlorophyll a concentrations (0.6, 1.6, and 2.3 µg cm -2 ). Concerning the plant wax experiments, 6 g of fresh Zea mays leaves were mixed with 20 mL of a mixture of hexane/diethyl ether 90/10 (v/v) for 20 seconds as proposed by [START_REF] Loneman | A robust and efficient method for the extraction of plant extracellular surface lipids as applied to the analysis of silks and seedling leaves of maize[END_REF]. Then, 6 mL of the resulting solution was deposited on two different glass plates, and solvents were evaporated as previously described for chlorophyll a experiments. In both cases, blank experiments were carried out with pure acetone or hexane/diethyl ether on glass plates.

To ensure the statistical significance of our results, three replicates were conducted on leaf and wax plate experiments, while four replicates were carried out on chlorophyll a experiments.

The published values represent the calculated means and standard deviations. Fitting curves presented in this work and their corresponding errors were determined using a least squared fitting procedure (Igor Pro software). The associated errors correspond to one standard deviation (1) (Table S1).

Experimental methodology and data analysis

First, the movable injector was pushed to the end of the reactor (position P0) to avoid any contact between the gaseous NO2 and the leaf surface. The NO2 mixing ratio in the injector was adjusted to the chosen value, and light was switched on. NO2 loss as well as HONO and NO formation were measured moving the injector and successively exposing 14, 29 and 44 cm 2 of leaf surface corresponding to injector positions P1, P2 and P3, respectively. After each measurement, the injector was pushed back to position P0 to control the initial signal level. Each position was held for 30 minutes for signal stabilization and to have enough measurement points. The injector positions enable us to vary the contact time between the gaseous NO2 and the leaf surface and allow the calculation of the reaction kinetics. HONO and NO emission rates as well as NO2 losses are defined as the average formation/consumption rates per unit of area calculated for each injector position. The influence of air relative humidity (5 to 58%), NO2 mixing ratio (10 to 80 ppbv), light intensity (0 to 20 W m -2 ) and temperature leaf surface (288 to 313 K) was investigated.

HONO emission rates (HONOER) were calculated according to the following equation:

HONOER = [HONO] × V R t × S
where [HONO] is the concentration of HONO (molecules cm -3 ) measured at a defined residence time (Rt in s), V is the volume of the reactor (cm 3 ) and S is the reactive surface (cm 2 ).

The uptake coefficient of the leaf surface was determined by integrating the "drop" in the NO2 signal as a function of the contact time -according to the following equation:

𝛾 = 4𝑘 𝑁𝑂2 𝑉 𝜈 𝑁𝑂2 𝑆
where the injector distance (cm), the mass flow rate (cm 3 min -1 ) and the reactor inner diameter (cm) are known, 𝑘 𝑁𝑂 2 is the pseudofirst-order rate for the reaction of NO2 with the surface,

𝜈 𝑁𝑂 2 is the diffusion coefficient determined as √ 8𝑅𝑇 𝜋𝑀
, with R being the universal gas constant (J mol -1 K -1 ), T being the temperature (K), M being the molar mass (g mol -1 ), V being the reactor volume (cm 3 ) and S being the reactive surface (cm 2 ).

The pseudofirst-order reaction rate of NO2 is defined as ln(

[𝑁𝑂 2 ] 𝑡 [𝑁𝑂 2 ] 0 ) = -𝑘 𝑁𝑂 2 t, with [𝑁𝑂 2 ] 𝑡 being
the measured NO2 mixing ratio versus exposure time, [𝑁𝑂 2 ] 0 being the initial NO2 mixing ratio at P0 and t being the contact time between NO2 and the leaf surface. Under irradiation conditions, the consumption rate of NO2 is lower than the sum of the emission rates of HONO, NO, and NO3 -/HNO3 (assumed to be equal to the HONO emission rate according to equation R2). This phenomenon is due to the photolysis of NO2 in the injector (up to a 10% NO2 mixing ratio). It would be interesting to calculate this photolysis frequency, but unfortunately, the actinic flux in the movable injector cannot be measured because of its low inner diameter (approximately 2 mm). Under the experimental conditions used in this study, HONO and HNO3 photolysis were negligible in the system.

Finally, HONO production on vegetation under field conditions was estimated by knowing HONO production on glass plates, on Zea mays leaves and the LAI of Zea mays plants (up to 6 m 2 m -2 ). As suggested in the literature, ground-level HONO can be assimilated to a production rate in the mixing layer by considering a homogeneous HONO mixing ratio throughout the air column [START_REF] Meusel | Emission of nitrous acid from soil and biological soil crusts represents an important source of HONO in the remote atmosphere in Cyprus[END_REF][START_REF] Stemmler | Photosensitized reduction of nitrogen dioxide on humic acid as a source of nitrous acid[END_REF][START_REF] Xue | Development and application of a twin open-top chambers method to measure soil HONO emission in the North China Plain[END_REF]. As explained by [START_REF] Meusel | Emission of nitrous acid from soil and biological soil crusts represents an important source of HONO in the remote atmosphere in Cyprus[END_REF], this calculation mode does not take into account the HONO vertical gradient but enables estimation of the order of magnitude of the gap between the calculated emission rate and the HONO missing source [START_REF] Stemmler | Photosensitized reduction of nitrogen dioxide on humic acid as a source of nitrous acid[END_REF][START_REF] Xue | Development and application of a twin open-top chambers method to measure soil HONO emission in the North China Plain[END_REF]. As a result, the HONO maximal emission rate from Zea mays leaves was estimated per unit of ground surface at several temperatures, and approximations of the HONO production rate in the mixing layer were calculated with and without blank subtraction. [START_REF] Meusel | Emission of nitrous acid from soil and biological soil crusts represents an important source of HONO in the remote atmosphere in Cyprus[END_REF] and [START_REF] Xue | Development and application of a twin open-top chambers method to measure soil HONO emission in the North China Plain[END_REF] evaluated the contribution of HONO emissions from soil using an averaged mixing layer height of 300 m. To enable a direct comparison with these previous studies, the same mixing layer height was chosen in this work.

Results

NO2 uptake and HONO surface emission rates

Figure 2 depicts the HONO surface emission rates for a set of experimental conditions (temperature, irradiation flux, NO2 mixing ratio and relative humidity), while Table 2 lists a summary of HONO emission rates reported in the literature. The impact of light intensity (from 0 to 20 W m -2 ) on the HONO emission rate was investigated (Figure 2A). Depending on the experimental conditions, the values ranged from (5.4 ± 2.0) × 10 8 to (1.4 ± 0.2) × 10 9 molecules cm -2 s -1 on the reactor surfaces (blank experiments) and from (2.2 ± 0.5) × 10 9 to (3.3 ± 2.0) × 10 9 molecules cm -2 s -1 on Zea mays leaves. Based on experimental uncertainties, it can be concluded that there was no clear influence of UV-A light on the HONO emission rate from Zea mays leaves. The NO2 uptake on leaves was calculated in the dark under steady-state conditions at 303 K, 40 ppb of NO2 and 40% RH after approximately 20 minutes of exposure, and the average was (6.5 ± 0.2) × 10 -7 . Under these experimental conditions, uptake on the glass plate was negligible.

The effect of variable humidity from 5 to 58% was also tested (Figure 2B). HONO emission rates ranged from (5.1 ± 1.4) × 10 8 to (1.5 ± 0.5) × 10 9 molecules cm -2 s -1 in the Pyrex reactor (blank experiment) and from (2.0 ± 0.5) × 10 9 to (2.9 ± 0.4) × 10 9 molecules cm -2 s -1 on Zea mays leaves. The HONO emission rate linearly increased with RH on both substrates.

A linear correlation between HONO emission rates and NO2 mixing ratios was highlighted from 10 to 80 ppbv (Figure 2C). The observed HONO formation ranged from (9.0 ± 2.5) × 10 8 to (2.2 ± 0.6) × 10 9 molecules cm -2 s -1 on reactor Pyrex surfaces and from (6.7 ± 0.5) × 10 8 to (4.2 ± 1.2) × 10 9 molecules cm -2 s -1 on Zea mays leaves.

The temperature dependence of HONO formation on Zea mays leaves was tested by varying the experimental temperature from 288 and 313 K (Figure 2D). The emission rates varied by a factor of six, ranging from (1.2 ± 0.4) × 10 9 to (6.8 ± 0.8) × 10 9 molecules cm -2 s -1 . No temperature dependence was observed on the reactor surface, which showed an average formation rate of (1.3 ± 0.9) × 10 9 molecules cm -2 s -1 . From the experimental data on Zea mays leaves, an overall activation energy of 56 ± 10 kJ mol -1 could be derived.

Table 3 presents an estimation of HONO emissions under environmental conditions for Zea Mays, which possesses an LAI of 6 m 2 m -2 . Zea mays leaf emissions correspond to emission rates from (5.8 ± 1.2) × 10 17 molecules m -2 h -1 at 303 K to (1.2 ± 0.2) × 10 18 at 313 K.

HONO formation on chlorophyll a and wax surfaces

HONO emission rates on chlorophyll a and wax surfaces are presented in Figure 4. Chlorophyll a experiments showed a high decrease in the HONO signal over time, suggesting a very fast depletion of the reactive sites. As a result, only the first position of the injector was used to calculate HONO emission rates for chlorophyll thin films. However, the chlorophyll a concentrations deposited on the glass plate were lower than those measured inside the Zea mays leaves, which normally range from 24 to 30 µg cm -2 [START_REF] Zhao | Corn (Zea mays L.) growth, leaf pigment concentration, photosynthesis and leaf hyperspectral reflectance properties as affected by nitrogen supply[END_REF]. Despite these low concentrations, the HONO formation rate on chlorophyll a films was over a factor ten higher than the values observed on the leaf surface.

To evaluate the repeatability of the measurements on wax films, two different plates (wax plates 1 and 2, Figure 4) covered by aliquots of the same wax extract were tested. The results showed similar HONO emissions on both plates: (4.1 ± 0.5) × 10 9 molecules cm -2 s -1 and (4.8 ± 0.9) × 10 9 molecules cm -2 s -1 . The HONO emission from the wax plates was clearly closer to the values calculated on leaves than those measured on chlorophyll a. To test the wax reactivity over time, two experiments were realized on the same wax plate film, one after the other. The results in Figure 4 show average emission rates of (4.6 ± 0.8) × 10 9 molecules cm - 2 s -1 from wax plates 1 and 2 and (2.3 ± 0.3) × 10 9 molecules cm -2 s -1 from wax plate 1bis, suggesting a fast depletion of the reactive sites on this type of thin film.

Discussion

This study highlighted the efficient conversion of NO2 to HONO on Zea mays leaves and elucidated the dependence of this heterogeneous reaction on several environmental parameters.

A steady-state uptake of (6.5 ± 0.2) × 10 -7 was measured in this work. As this is the first time that NO2 uptake has been reported on a leaf surface, uptake on other organic surfaces is presented for comparison in Table 1. Humic acid films are an important chemical system because they are part of the soil composition and could be a good representation of soil reactivity [START_REF] Stemmler | Photosensitized reduction of nitrogen dioxide on humic acid as a source of nitrous acid[END_REF]. The uptakes observed on humic acids and at the leaf surface are similar (on the order of 10 -7 ), suggesting that the heterogeneous reactivity of NO2 on both surfaces is highly comparable in dark conditions. Films of anthrarobin, catechol, and anthracene showed larger uptakes up to 10 -6 , suggesting faster electron transfer from substrates containing monoaromatic compounds and PAHs [START_REF] Arens | The reaction of NO2 with solid anthrarobin (1,2,10-trihydroxy-anthracene)[END_REF][START_REF] George | Photoenhanced uptake of gaseous NO 2 on solid organic compounds: a photochemical source of HONO?[END_REF].

The observed linear dependence of HONO formation on the NO2 mixing ratio suggests that the leaf surface sites were not saturated under the experimental conditions. A similar increase was previously reported on several organic surfaces, but most of the time, HONO formation ended in a plateau for high NO2 mixing ratios, probably due to saturation of the adsorption sites [START_REF] Arens | The reaction of NO2 with solid anthrarobin (1,2,10-trihydroxy-anthracene)[END_REF][START_REF] Han | Heterogeneous Photochemical Conversion of NO 2 to HONO on the Humic Acid Surface under Simulated Sunlight[END_REF][START_REF] Stemmler | Light induced conversion of nitrogen dioxide into nitrous acid on submicron humic acid aerosol[END_REF]. The differences regarding the concentration at which the saturation effect appears can be explained by the number of sites at the surface and the accessibility of NO2 to the inner layer of the substrate. Thin films or suspended particles of humic acids presented saturation effects for NO2 concentrations above 50-60 ppbv [START_REF] Stemmler | Light induced conversion of nitrogen dioxide into nitrous acid on submicron humic acid aerosol[END_REF], while other surfaces with multiple layers or porous surfaces showed saturation at higher NO2 mixing ratios [START_REF] Arens | The reaction of NO2 with solid anthrarobin (1,2,10-trihydroxy-anthracene)[END_REF][START_REF] Han | Heterogeneous Photochemical Conversion of NO 2 to HONO on the Humic Acid Surface under Simulated Sunlight[END_REF]. Han et al.

(2016) stated that the whole thickness of their humic acid film was both accessible and reactive towards NO2, leading to saturation for NO2 concentrations higher than 160 ppbv. Similarly, anthrarobin films presented high surface roughness and were composed of 1 × 10 3 monolayers, explaining why no saturation effects were observed until 120 ppbv [START_REF] Arens | The reaction of NO2 with solid anthrarobin (1,2,10-trihydroxy-anthracene)[END_REF]. The Zea mays leaf surfaces present a certain roughness, as shown by electron microscopy images [START_REF] Januszkiewicz | Effect of surfactants and leaf surface morphology on the evaporation time and coverage area of ZnIDHA droplets[END_REF], which implies a higher number of adsorption sites than a thin film and probably explains why saturation was not observed for the NO2 mixing ratio of 80 ppbv, the highest value tested.

The influence of humidity on HONO emissions on the blank and Zea mays leaves resulted in two parallel linear fits (Figure 4B). This behavior suggests that the observed increment is due to the contribution from the reactor walls rather than from the leaf surface. [START_REF] Finlayson-Pitts | The heterogeneous hydrolysis of NO2 in laboratory systems and in outdoor and indoor atmospheres: an integrated mechanism[END_REF] previously observed a similar behavior on borosilicate glass. The experiments were carried out by varying the humidity from 10 to 60%, which corresponded to an increase from one to five water layers adsorbed on the glass surface. They interpreted the results of HONO formation on the glass plate as correlated to the number of water molecules adsorbed on the surface rather than the water vapor molecules in the gas phase colliding with the substrate surface [START_REF] Finlayson-Pitts | The heterogeneous hydrolysis of NO2 in laboratory systems and in outdoor and indoor atmospheres: an integrated mechanism[END_REF]. The number of water layers on plant leaves has been previously determined using electric conductivity measurements, and it was estimated that a water monolayer can be formed for RH above 50% [START_REF] Lammel | Formation of nitrous acid: parameterisation and comparison with observations[END_REF]. The lack of dependence of HONO emissions on ambient humidity observed in this work could be an indication that even at the highest humidity (58%) used in our experiments, a water monolayer was not formed.

Furthermore, [START_REF] Arens | Heterogeneous reaction of NO2 on diesel soot particles[END_REF] showed that HONO formation on soot particles needed water molecules but was independent of humidity in the range of 4 to 77%. Some experiments using D2O molecules showed the formation of DONO, assessing the important role of water molecules in the heterogeneous mechanism [START_REF] Kleffmann | Heterogeneous conversion of NO2 on carbonaceous surfaces[END_REF][START_REF] Longfellow | Reactive uptake on hydrocarbon soot: Focus on NO 2[END_REF].

When using H2 18 O in the gas phase, only HONO was formed, suggesting that the oxygen of the water molecules did not participate in the reaction [START_REF] Longfellow | Reactive uptake on hydrocarbon soot: Focus on NO 2[END_REF]. The authors also demonstrated that an efficient conversion of NO2 to HONO on soot surfaces could take place at a low humidity of 4% [START_REF] Arens | Heterogeneous reaction of NO2 on diesel soot particles[END_REF][START_REF] Kleffmann | Heterogeneous conversion of NO2 on carbonaceous surfaces[END_REF]. It was then concluded that the overall reaction rate of HONO formation must be limited by another chemical process [START_REF] Arens | Heterogeneous reaction of NO2 on diesel soot particles[END_REF][START_REF] Kleffmann | Heterogeneous conversion of NO2 on carbonaceous surfaces[END_REF].

Temperature variation produced the most remarkable increase in HONO emissions (a factor of 28 between 293 and 313 K). Two possible effects were invoked to explain this observation. The first is that temperature affects partitioning between the gas and solid phases, favoring desorption of molecules from the surface [START_REF] Liu | Temperature Dependence of the Heterogeneous Reaction of Carbonyl Sulfide on Magnesium Oxide[END_REF][START_REF] Tlili | Humidity and temperature dependences of the adsorption and desorption rates for acetone and xylene on silicon wafer[END_REF]. Mainly, the leaf temperature increase enhanced HONO desorption kinetics, leaving access to reactive sites for the adsorption of a new H2O and/or NO2 molecule. A second hypothesis is that the conversion kinetics of NO2 to HONO on the leaf surface can be accelerated by the temperature increase [START_REF] Arens | The reaction of NO2 with solid anthrarobin (1,2,10-trihydroxy-anthracene)[END_REF]. Unfortunately, the relative contribution of each of these two individual processes can neither be demonstrated nor computed separately. From the temperature dependence measurements, an overall activation energy of 56 ± 10 kJ mol -1 was calculated.

This overall activation energy represents the sum of the activation energy and the reaction enthalpies of all the heterogeneous reaction steps. Values of the same order of magnitude of 47.1 kJ mol -1 were reported for soot particles [START_REF] Shrivastava | Kinetics of Soot Oxidation by NO 2[END_REF], while anthrarobin films showed a slightly lower activation energy of 39 kJ mol -1 . The reaction of NO2 therefore requires more energy on leaves and soot than on anthrarobins. This may be tentatively explained by the fact that their surfaces are a complex medium and contain less efficient electron donors than anthrarobin films, which are composed of pure aromatic compounds.

Apparently, NO2 to HONO conversion on Zea mays leaves did not show dependence on the photon flux, and the emission rates observed were comparable in the darkness and under irradiation up to a light intensity of 20 W m -2 . The NO2 to HONO conversion requires an electron donor available on the reactive surface. Under the leaf surface, many pigments are available, including chlorophyll a, which is well known as an efficient electron donor.

However, chlorophyll a is not directly available to oxidants, as it is protected by a cuticular wax layer and epidermal cells (which do not possess chlorophyll). Waxes are recognized as a major component of the leaf surface [START_REF] Barthlott | Classification and terminology of plant epicuticular waxes[END_REF][START_REF] Croft | Leaf Pigment Content[END_REF], forming a protective interface towards the atmosphere and preventing water loss from the leaf [START_REF] Barthlott | Plant Surfaces: Structures and Functions for Biomimetic Innovations[END_REF]. Maize wax investigations revealed that plant wax contains hydrocarbons, fatty acids, aldehydes, esters and alcohol [START_REF] Javelle | Overexpression of the Epidermis-Specific Homeodomain-Leucine Zipper IV Transcription Factor outer cell layer in Maize Identifies Target Genes Involved in Lipid Metabolism and Cuticle Biosynthesis[END_REF][START_REF] Loneman | A robust and efficient method for the extraction of plant extracellular surface lipids as applied to the analysis of silks and seedling leaves of maize[END_REF]. Several functional groups, such as carbonyls and/or unsaturated groups, can potentially act as electron donors and therefore contribute to HONO formation [START_REF] Kunst | Biosynthesis and secretion of plant cuticular wax[END_REF][START_REF] Prasad | Epicuticular waxes from leaves of Quercus robur[END_REF][START_REF] Wen | Composition of secondary alcohols, ketones, alkanediols, and ketols in Arabidopsis thaliana cuticular waxes[END_REF]. Since waxes are present on the leaf surface and HONO formation on these films was quite similar to the values observed on Zea mays leaves, it is more likely that waxes are partly responsible for the HONO measured. Many structures and wax compositions are reported in the literature, suggesting that different plants could emit different amounts of HONO [START_REF] Barthlott | Classification and terminology of plant epicuticular waxes[END_REF][START_REF] Holloway | Structural determination of secondary alcohols from plant epicuticular waxes[END_REF][START_REF] Koch | Diversity of structure, morphology and wetting of plant surfaces[END_REF][START_REF] Reynhardt | Structures and molecular dynamics of plant waxes: II. Cuticular waxes from leaves of Fagus sylvatica L. and Hordeum vulgare L[END_REF].

Our experiments on wax extracts deposited on thin films also showed a rapid depletion of the reactive sites not observed on Zea mays leaves. It is difficult to say if this is due to the different surfaces (one flat and the other with a certain roughness) or if other chemical components of the leaf surface could additionally contribute to HONO formation (for example, components of the cutin that contain both unsaturated carboxylic acids or diacids).

Table 2 presents several surfaces onto which the formation of HONO has been reported.

Recently, soil was proposed as a possible contributor to missing HONO sources. HONO production in soil is highly variable depending on soil composition, moisture, and the presence of fertilizers [START_REF] Bhattarai | Emissions of atmospherically important nitrous acid (HONO) gas from northern grassland soil increases in the presence of nitrite (NO2)[END_REF][START_REF] Donaldson | Soil surface acidity plays a determining role in the atmospheric-terrestrial exchange of nitrous acid[END_REF][START_REF] Meusel | Emission of nitrous acid from soil and biological soil crusts represents an important source of HONO in the remote atmosphere in Cyprus[END_REF][START_REF] Scharko | Combined flux chamber and genomics approach links nitrous acid emissions to ammonia oxidizing bacteria and archaea in urban and agricultural soil[END_REF][START_REF] Wu | Soil HONO emissions at high moisture content are driven by microbial nitrate reduction to nitrite: tackling the HONO puzzle[END_REF]. [START_REF] Bhattarai | Emissions of atmospherically important nitrous acid (HONO) gas from northern grassland soil increases in the presence of nitrite (NO2)[END_REF] and [START_REF] Wu | Soil HONO emissions at high moisture content are driven by microbial nitrate reduction to nitrite: tackling the HONO puzzle[END_REF] evaluated the HONO emission rate from soil ranging from 9.6 × 10 7 to 8.2 × 10 11 molecules cm -2 s -1 . On a thin film of humic acid exposed to 17 ppbv of NO2 at 20% RH, the HONO emission rate was approximately 2.5 × 10 10 molecules cm -2 s -1 [START_REF] Stemmler | Photosensitized reduction of nitrogen dioxide on humic acid as a source of nitrous acid[END_REF]; a similar value was reported for soot surfaces [START_REF] Arens | Heterogeneous reaction of NO2 on diesel soot particles[END_REF][START_REF] Han | Influences of O2 and O3 on the heterogeneous photochemical reaction of NO2 with humic acids[END_REF]. Most of these surfaces produce at least 3 times more HONO than the tested Zea mays leaf surface. However, to assess which surface may mostly contribute to the total HONO formation in the environment, the total available surface of plant leaves should be considered. Considering HONO emission rates from soil calculated from previous work [START_REF] Bhattarai | Emissions of atmospherically important nitrous acid (HONO) gas from northern grassland soil increases in the presence of nitrite (NO2)[END_REF][START_REF] Meusel | Emission of nitrous acid from soil and biological soil crusts represents an important source of HONO in the remote atmosphere in Cyprus[END_REF][START_REF] Wu | Soil HONO emissions at high moisture content are driven by microbial nitrate reduction to nitrite: tackling the HONO puzzle[END_REF][START_REF] Xue | Development and application of a twin open-top chambers method to measure soil HONO emission in the North China Plain[END_REF], HONO fluxes ranged from 0.5 pptv h -1 to 400 pptv h -1 depending on the soil properties. Estimations of HONO fluxes from Zea mays leaf surfaces were negligible below 303 K and ranged from 79 ± 17 pptv h -1 at 303 K to 171 ± 23 pptv h -1 at 313 K. Therefore, for warm environments in the presence of 40 ppbv of NO2, leaf contribution to HONO emission can compete with production from soil. In the literature, the so-called "HONO missing sources"

were estimated to range from 0.3 -0.6 ppbv h -1 in a rural mountain site in Germany [START_REF] Acker | Strong daytime production of OH from HNO2 at a rural mountain site[END_REF], approximately 0.22 ppbv h -1 in a rural site in New York state, USA [START_REF] Zhou | Summertime nitrous acid chemistry in the atmospheric boundary layer at a rural site in New York State: summertime nitrous acid chemistry[END_REF], and 0.05 -1.3 ppbv h -1 in a suburban site in the Paris region, France [START_REF] Michoud | Study of the unknown HONO daytime source at a European suburban site during the MEGAPOLI summer and winter field campaigns[END_REF]. As a result, HONO fluxes from Zea mays leaves may partially contribute to missing HONO sources.

Conclusions

For the first time, a systematic investigation of NO2 to HONO conversion on Zea mays leaf surfaces is reported in the literature. HONO formation was highly enhanced by increasing the temperature and NO2 mixing ratios, while RH (5-58%) and UVA irradiation (0 to 20 W m -2 between 340 and 400 nm) showed no effect. The highest emission rate values reached

(5.6 ± 0.8) × 10 9 molecules cm -2 s -1 at 313 K, 40 ppbv of NO2, 20 W m -2 , and 40% RH.

Experiments on synthetic surfaces prepared using wax extract from the leaves suggest that waxes could play a role in HONO formation. Estimations of HONO fluxes under environmental conditions showed that leaves can contribute up to 171 pptv h -1 at 313 K. Based on our results and considering the high foliage surface present in natural environments, HONO emission on leaves could represent a relevant source of HONO in the boundary layer. As HONO formation depends on environmental conditions, the importance of this source is expected to exhibit both daily and seasonal variations.

Figure 1 :

 1 Figure 1: Experimental setup for the determination of HONO formation on Zea mays leaves

Figure 3 :

 3 Figure 3: Arrhenius plot of HONO formation rates on Zea mays leaves at 40 ppbv of NO2, 40% relative humidity and 20 W m -2 . Points represent the average and error bars are the standard

Figure 4 :

 4 Figure 4: HONO emission rates (molecule cm -2 s -1 ) for chlorophyll a (Chl a), wax and leaf surfaces at 313 K, 40 ppbv of NO2, 40% relative humidity and 20 W m -2 (Experiments wax

Acknowledgements

We thank ADEME for funding the project HONO-CORN, included in PRIMEQUAL program.

We also thank Sylvie Dupouyet for the growth of Zea mays plants and INRA (Gif sur Yvette) for providing the Zea mays seeds.

Funding

This work was supported by ADEME who funding the project HONO-CORN, included in PRIMEQUAL program.

Author Surfaces Temperature (K) RH (%) [NO2] (ppb) Uptake NO2

This study Zea mays leaves 303 40 40 (6.5 ± 0.2) × 10 -7 [START_REF] Han | Heterogeneous Photochemical Conversion of NO 2 to HONO on the Humic Acid Surface under Simulated Sunlight[END_REF] Humic acid 298 22 30 ~1.9 × 10 -7