Amel Nouara

Christos Panagiotopoulos
email: christos.panagiotopoulos@mio.osupytheas.fr

Richard Sempéré

Simultaneous determination of neutral sugars, alditols and anhydrosugars using anion-exchange chromatography with pulsed amperometric detection: Application for marine and atmospheric samples

Keywords: Anion-exchange chromatography, carbohydrates, suspended atmospheric particles, marine particulate organic matter, high molecular weight dissolved organic matter

An improved high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) method is described for the simultaneous determination of neutral sugars (hexoses, pentoses and deoxysugars), alditols and anhydrosugars commonly found in atmospheric and marine samples. The method uses a CarboPac MA1 column, at a flow rate of 0.3 mL min -1 , and a NaOH gradient (250-700 mM).

The proposed method applies a temperature gradient (from 25 to 28 °C) to the column for the first 30 min of the analysis, followed by a constant temperature at 28 °C until the end of the analysis. These analytical conditions allowed the separation of 15 out of 17 carbohydrates in 75 min with resolution factors better than 0.5 for the critical pairs levoglucosan/arabitol, galactosan/arabinose, arabinose/mannose and glucose/xylose.

The application of this method to field samples revealed that anhydrosugars represented 53% of total neutral carbohydrates (TCHO) in total suspended atmospheric particles (TSP), whereas they were detected for the first time in marine particulate organic matter (POM) and high-molecular-weight dissolved organic matter (HMWDOM) samples accounting 2% and 3% of TCHO, respectively. Levoglucosan and/or galactosan were the major anhydrosugars in all samples however, their concentrations are undoubtedly underestimated because hydrolysis was applied to the marine samples prior to the HPAEC-PAD analysis. Despite this underestimation, their presence in the marine samples clearly indicates possible terrestrial input most likely via atmospheric deposition as these compounds are considered terrestrial burring biomass tracers. Finally, deoxysugars were also detected for the first time in the TSP sample representing 1% of TCHO, while alditols accounted for 0.4% and 0.3% of TCHO in POM and HMWDOM, respectively.

Introduction

Carbohydrates are among the most abundant organic molecules on the earth because they are detected in all marine and terrestrial ecosystems [START_REF] Giorio | Prospects for reconstructing paleoenvironmental conditions from organic compounds in polar snow and ice[END_REF][START_REF] He | Sources and accumulation of organic carbon in the Pearl River Estuary surface sediment as indicated by elemental, stable carbon isotopic, and carbohydrate compositions[END_REF][START_REF] Mopper | Sugars and uronic acids in sediment and water from the black sea and north sea with emphasis on analytical techniques[END_REF][START_REF] Mopper | The monosaccharide spectra of natural waters[END_REF]Panagiotopoulos and Sempéré, 2005a;Repeta et al., 2015;[START_REF] Theodosi | Sugars in atmospheric aerosols over the Eastern Mediterranean[END_REF]. The determination of carbohydrates at the molecular level has been often used to determine different biogeochemical signatures such as terrestrial inputs in coastal marine environments [START_REF] Cowie | Carbohydrate sources in a coastal marine environment[END_REF][START_REF] Da Cunha | Neutral sugars as biomarkers in the particulate organic matter of a French Mediterranean river[END_REF][START_REF] Panagiotopoulos | Composition and distribution of dissolved carbohydrates in the Beaufort Sea Mackenzie margin (Arctic Ocean)[END_REF], organic matter sources [START_REF] He | Sources and accumulation of organic carbon in the Pearl River Estuary surface sediment as indicated by elemental, stable carbon isotopic, and carbohydrate compositions[END_REF][START_REF] Panagiotopoulos | The composition and flux of particulate and dissolved carbohydrates from the Rhone River into the Mediterranean Sea[END_REF][START_REF] Wicks | Carbohydrate signatures of aquatic macrophytes and their dissolved degradation products as determines by a sensitive highperformance ion chromatography method[END_REF], and decomposition pathways [START_REF] Amon | Linkages among the bioreactivity, chemical composition, and diagenetic state of marine dissolved organic matter[END_REF][START_REF] Giroldo | relative increase of deoxy sugars during microbial degradation of an extracellular polysaccharide released by a tropical freshwater Thalassiosira sp. (Bacillariophyceae)[END_REF][START_REF] Hedges | Origins and processing of organic matter in the Amazon River as indicated by carbohydrates and amino acids[END_REF][START_REF] Opsahl | Characterization of carbohydrates during early diagenesis of five vascular plant tissues[END_REF].

Previous environmental studies on atmospheric suspended particulate matter (PM 2.5-10 ) showed that carbohydrates account for 0.2-3% and 0.7-11% of organic carbon (OC) and water-soluble organic carbon (WSOC), respectively [START_REF] Theodosi | Sugars in atmospheric aerosols over the Eastern Mediterranean[END_REF][START_REF] Tsai | Potential PM2.5 impacts of festivalrelated burning and other inputs on air quality in an urban area of southern Taiwan[END_REF][START_REF] Yang | The chemical composition of inorganic and carbonaceous materials in PM2.5 in Nanjing, China[END_REF]. The dominant carbohydrates reported in aerosols are glucose and sucrose released by terrestrial plants [START_REF] Medeiros | Sugars as source indicators of biogenic organic carbon in aerosols collected above the Howland Experimental Forest, Maine[END_REF]Samaké et al., 2019), levoglucosan, galactosan and mannosan from biomass burning processes [START_REF] Bhattarai | Levoglucosan as a tracer of biomass burning: Recent progress and perspectives[END_REF][START_REF] Simoneit | Sugars -Dominant Water-Soluble Organic Compounds in Soils and Characterization as Tracers in Atmospheric Particulate Matter[END_REF][START_REF] Simoneit | Levoglucosan, a tracer for cellulose in biomass burning and atmospheric particles[END_REF], as well as mannitol and arabitol from airborne fungal spores and/or various vascular plants [START_REF] Bauer | Arabitol and mannitol as tracers for the quantification of airborne fungal spores[END_REF]Samaké et al., 2019). Other minor carbohydrates reported in aerosols include some neutral sugars (e.g., arabinose, fructose, galactose, and mannose) and alditols (e.g., xylitol and sorbitol; [START_REF] Barbaro | Sugars in Antarctic aerosol[END_REF][START_REF] Theodosi | Sugars in atmospheric aerosols over the Eastern Mediterranean[END_REF]. It is worth noting that fucose and rhamnose, two important deoxysugars consistently found in bacterial heteropolysaccharides [START_REF] Giroldo | relative increase of deoxy sugars during microbial degradation of an extracellular polysaccharide released by a tropical freshwater Thalassiosira sp. (Bacillariophyceae)[END_REF], soil organic matter [START_REF] Bock | Sequestration and turnover of plant and microbially derived sugars in a temperate grassland soil during 7 years exposed to elevated atmospheric pCO2[END_REF], freshwater ecosystems [START_REF] Cheng | Simultaneous analyses of neutral carbohydrates and amino sugars in freshwaters with HPLC-PAD[END_REF] as well as vascular plants [START_REF] Opsahl | Characterization of carbohydrates during early diagenesis of five vascular plant tissues[END_REF] have never been reported in atmospheric samples.

Monomeric constituents of carbohydrates after acid hydrolysis (e.g., fucose, rhamnose, arabinose, galactose, glucose, mannose and xylose) have also been detected in the marine environment. As revealed by chromatographic techniques, these monosaccharides account for 10-15%, <10% and <20% in particulate, dissolved and sedimentary organic carbon, respectively (Panagiotopoulos and Sempéré, 2005a and references therein). These monosaccharides have also been reported in marine high-molecular-weight dissolved organic matter (HMWDOM) at equimolar concentrations although 1 H nuclear magnetic resonance spectroscopy (NMR) showed the presence of much broader spectra of carbohydrates including unhydrolyzed polysaccharides and methylated sugars [START_REF] Panagiotopoulos | Characterization of methyl sugars, 3deoxysugars and methyl deoxysugars in marine high molecular weight dissolved organic matter[END_REF]. Other minor carbohydrate classes also reported in marine samples include amino sugars [START_REF] Benner | Abundance of amino sugars and peptidoglycan in marine particulate and dissolved organic matter[END_REF][START_REF] Kaiser | Biochemical composition and size distribution of organic matter at the Pacific and Atlantic time series stations[END_REF] and uronic acids [START_REF] Bergamaschi | Distributions of uronic acids and Omethyl sugars in sinking and sedimentary particles in two coastal marine environments[END_REF][START_REF] Engel | A novel protocol for determining the concentration and composition of sugars in particulate and in high molecular weight dissolved organic matter (HMW-DOM) in seawater[END_REF][START_REF] Hung | Distributions of carbohydrates, including uronic acids, in estuarine waters of Galveston Bay[END_REF].

Surprisingly, alditols and anhydrosugars, two major carbohydrate categories consistently found in aerosols and ice cores [START_REF] Gambaro | Direct determination of levoglucosan at the picogram per milliliter level in antarctic ice by high-performance liquid chromatography/electrospray ionization triple quadrupole mass spectrometry[END_REF][START_REF] Giorio | Prospects for reconstructing paleoenvironmental conditions from organic compounds in polar snow and ice[END_REF], have scarcely been reported in marine samples despite the growing evidence that dry or wet atmospheric deposition is an important process between the atmospheric and marine compartments [START_REF] Pantelaki | Characterization of dissolved organic carbon in rainwater of an urban/coastal site in Mediterranean area[END_REF][START_REF] Zheng | Impacts of springtime biomass burning in the northern Southeast Asia on marine organic aerosols over the Gulf of Tonkin, China[END_REF]. It is worth noting that only [START_REF] Panagiotopoulos | Molecular level characterization of methyl sugars in marine high molecular weight dissolved organic matter[END_REF] have identified anhydrosugars (levoglucosan, mannosan, and galactosan) in surface marine HMWDOM, in addition to the seven neutral sugars described above. This finding raises questions about the possible sources of anhydrosugars in marine water (terrestrial vs marine), which have been recently examined using compound-specific carbon isotopes analysis [START_REF] Nouara | Liquid chromatographic isolation of individual carbohydrates from environmental matrices for stable carbon analysis and radiocarbon dating[END_REF]. Nevertheless, to the best of our knowledge a well-established technique for the measurement of anhydrosugars and alditols does not exist for marine samples. Moreover, the lack of information about deoxysugars in atmospheric samples hinders our understanding of their dynamics and clearly points to the need for a universal method to measure these carbohydrates in any environmental sample.

High-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) is the most employed chromatographic technique for the analysis of carbohydrates in environmental samples [START_REF] Benner | Abundance of amino sugars and peptidoglycan in marine particulate and dissolved organic matter[END_REF][START_REF] Caseiro | Determination of saccharides in atmospheric aerosol using anion-exchange highperformance liquid chromatography and pulsed-amperometric detection[END_REF][START_REF] Iinuma | A highly resolved anion-exchange chromatographic method for determination of saccharidic tracers for biomass combustion and primary bio-particles in atmospheric aerosol[END_REF][START_REF] Mopper | Determination of sugars in unconcentrated seawater and other natural waters by liquid chromatography and pulsed amperometric detection[END_REF]Panagiotopoulos and Sempéré, 2005a;[START_REF] Sempéré | Distribution and bacterial availability of dissolved neutral sugars in the South East Pacific[END_REF]. However, simple carbohydrates (e.g., monosaccharides) are not ionized in the same manner in alkaline media used in the HPAEC mobile phase, and therefore different concentrations of NaOH must be applied to separate these monosaccharide families (e.g., alditols, neutral sugars, and uronic acids). Due to h " m a ", most environmental applications of the HPAEC-PAD technique have focused on one family of carbohydrates (e.g., neutral sugars, or amino sugars) and few attempts have been made to date to separate a much broader spectrum of carbohydrate categories in the same run [START_REF] Engel | A novel protocol for determining the concentration and composition of sugars in particulate and in high molecular weight dissolved organic matter (HMW-DOM) in seawater[END_REF].

The main objective of the present study is to establish a universal HPAEC-PAD method capable of simultaneously identifying several carbohydrate families including neutral sugars, alditols and anhydrosugars commonly found in terrestrial and marine ecosystems. This objective was achieved after the evaluation of the performance of two different anionexchange columns and the optimization of the mobile phase concentration, column temperature and flow rate. The proposed method was validated in three different environmental matrices, namely, total suspended atmospheric particles (TSP), marine particulate organic matter (POM), and marine HMWDOM.

2.

Materials and procedures

Reagents

Carbohydrate standards were purchased from Sigma-Aldrich or Interchim, at their purest available grade (>98% (450 °C, 6 h). A 1-cm 2 section of the filter was extracted with 6 mL ultrapure water in an ultrasonic bath for 1 h and filtered through a Pasteur pipette packed with quartz wool (both precombusted at 450 °C for 6 h) to remove any remaining particles, and then analyzed immediately [START_REF] Theodosi | Sugars in atmospheric aerosols over the Eastern Mediterranean[END_REF]. The recovery yields of the extraction procedure were estimated after the extraction of a precombusted (450 ° C, 6 h) Whatman quartz fiber filter spiked with a standard mixture of neutral carbohydrates spanning a concentration range from 100 to 1000 nM [START_REF] Theodosi | Sugars in atmospheric aerosols over the Eastern Mediterranean[END_REF]. The obtained yields of neutral sugars for all the concentrations tested were between 101 and 119% (n = 5). The abovementioned analytical procedure allowed the extraction and identification only of the free neutral sugars present in the TSP sample and as such the composition of combined sugars was not determined in this study.

Marine POM

Sinking particles (marine POM) were collected over seven-day periods from January 6th to March 3rd, 2013, in the upwelling system located offshore of Lima (Peru) in the Pacific Oc a (12° 02' S -77° 40' W) g m ap (PPS3 ch cap) p y h oxycline/upper oxygen minimum zone (OMZ) layer at a depth of 34 m [START_REF] Bretagnon | Modulation of the vertical particles transfer efficiency in the Oxygen Minimum Zone off Peru[END_REF]. To avoid the bio-degradation of POM, a solution of seawater with 5% formaldehyde was added to the bottom of the collection chamber. After trap recovery, the living and dead swimmers were carefully removed so that only detrital particles remained in the sample.

These detrital particles (marine POM) were stored in the dark at 4 °C in the initial chambers used in the trap. On land, the samples were filtered through 25 mm precombusted (450 °C, 6 h) Whatman GF/F filters, freeze-dried, and subsequently stored in the dark at 4 °C until further analysis. F p (40-60 mg ach) f ach f h f amp b a f m h respective collection chambers of the sediment trap were pooled together, resulting in ~265 mg dry POM powder. To release neutral sugars, the POM powder was hydrolyzed with 1 M HCl at 100 °C for 20 h [START_REF] Panagiotopoulos | Composition and distribution of dissolved carbohydrates in the Beaufort Sea Mackenzie margin (Arctic Ocean)[END_REF]. The acid-soluble fraction recovered after centrifugation (2000 rpm) was then transferred into a precombusted (450 °C, 6 h) glass vial, and the acid was removed from the sample by three successive lyophilizations. The POM recovery (by weight) after acid hydrolysis was 84%. A part of the final dry powder was weighed (25.61 mg), redissolved in 1 mL ultrapure water and filtered through a Pasteur pipette packed with quartz wool (both precombusted at 450 °C for 6 h). The filtrate was diluted 500 times with ultrapure water and filtered again to remove any remaining particles prior to injection into the chromatographic system.

Marine HMWDOM

The surface (15 m) awa amp wa c c f m h Hawa a a (19° 43′ N -156° 03′ W) D cember 2003 and processed at Woods Hole Oceanographic Institution (USA) as described by [START_REF] Panagiotopoulos | Molecular level characterization of methyl sugars in marine high molecular weight dissolved organic matter[END_REF]. Briefly, the sample (8640 L) was filtered through 0.8 µm and 0.2 µm duel cartridge filters, and then ultrafiltered using a cross-flow ultrafiltration system. The concentrated sample (40 L) that included the HMWDOM was further desalted by diafiltration with ultrapure water to a final volume of 2 L and freeze dried to obtain a fluffy white powder. The carbon composition of the HMWDOM collected was 35% by weight, with a carbon/nitrogen ratio of 16. A part of the resultant HMWDOM powder (104 mg) was hydrolyzed to release the neutral sugars with 1 M HCl at 100 °C for 20 h, then the hydrolysis was stopped by placing the sample in an ice bath. The HCl was removed from the sample by lyophilization (3 times; final pH = 7). Then, a portion of the sample (55.83 mg)

was redissolved in 1 mL ultrapure water, filtered through a Pasteur pipette packed with quartz wool (both precombusted at 450 °C for 6 h) and stored frozen at -20 °C. Before analysis, the sample was diluted 3 times with ultrapure water.

Chromatographic system

Carbohydrate analysis was performed using a Thermo-Dionex ICS-3000 anion-exchange chromatograph (HPAEC) equipped with a 250-µL injection loop and a pulsed amperometric detector (PAD). The waveform used for pulsed amperometric detection was the standard quadruple potential for carbohydrate analysis [START_REF] Panagiotopoulos | The composition and flux of particulate and dissolved carbohydrates from the Rhone River into the Mediterranean Sea[END_REF]. The ultrapure water used to prepare the mobile phase was sparged with high purity N 2 for 30 min prior to use. The mobile phase consisted of ultrapure water (eluent A) and NaOH at 1 M (eluent B)

prepared by dilution in degassed ultrapure water. The eluents were kept continuously under pure N 2 pressure to avoid exposure to atmospheric CO 2, which can cause a drastic decrease in column selectivity and loss of sugar resolution. In addition, an on-line degasser (RFIC eluent degasser, Thermo Fisher) was added before the analytical column to decrease interference from dissolved oxygen [START_REF] Cheng | Improved Analysis of Dissolved Carbohydrates in Stream Water with HPLC-PAD[END_REF].

In this study, two anion-exchange analytical columns were tested for sugar separation: a CarboPac MA1 (7.5 µm, 4 × 250 mm; Thermo Fisher) and a CarboPac PA1 (10 µm, 4 × 250 mm; Thermo Fisher), both fitted with their corresponding guard column (4 × 50 mm; Thermo Fisher). The Chromeleon software (Thermo Fisher) was used for processing and data acquisition. The performance of the chromatographic separation (resolution factor, R s )

between the two-carbohydrate species A and B is calculated according to the following formula:

R s = 2 [(t R ) B -(t R ) A ] × [W A + W B ] -1
Where (t R ) A and (t R ) B are the retention times of the two species A and B, respectively, and W A and W B are their corresponding peak widths.

Results and discussion

Method optimization

Although previous investigations described the simultaneous discrimination of some neutral sugars, alditols and anhydrosugars using CarboPac PA1 or CarboPac MA1 columns [START_REF] Caseiro | Determination of saccharides in atmospheric aerosol using anion-exchange highperformance liquid chromatography and pulsed-amperometric detection[END_REF][START_REF] Iinuma | A highly resolved anion-exchange chromatographic method for determination of saccharidic tracers for biomass combustion and primary bio-particles in atmospheric aerosol[END_REF], the analytical HPAEC-PAD conditions used in previous studies were unable to fully resolve the 17-carbohydrate mixture assessed in the present study. Application of the analytical protocol using the CarboPac MA1 column in [START_REF] Iinuma | A highly resolved anion-exchange chromatographic method for determination of saccharidic tracers for biomass combustion and primary bio-particles in atmospheric aerosol[END_REF] revealed that the critical monosaccharide groups that required a further efficient separation were, by elution order: (a) levoglucosan, arabitol, fucose and rhamnose, (b) mannosan and sorbitol, (c) mannose, arabinose and galactosan, and (d) glucose and xylose. The optimization of the method therefore involved evaluating the effects of basic chromatographic parameters (flow rate, mobile phase composition and column temperature)

to achieve optimal separation of the 17-carbohydrate mixture. Charged carbohydrates (e.g., uronic acids and phosphorylated sugars) were not considered in this study because they require different mobile phase conditions compared to neutral carbohydrates using CH 3 COONa as the eluent [START_REF] Engel | A novel protocol for determining the concentration and composition of sugars in particulate and in high molecular weight dissolved organic matter (HMW-DOM) in seawater[END_REF][START_REF] Hu | Quantification of sugar compounds and uronic acids in enzymatic hydrolysates of lignocellulose using high-performance anion exchange chromatography with pulsed amperometric detection[END_REF].

Column selection

Although several types of anion-exchange columns are available for carbohydrate analysis (CarboPac PA10, 20, 100 and 200;Ronkart et al. 2007;[START_REF] Meyer | Improved RP-HPLC and anionexchange chromatography methods for the determination of amino acids and carbohydrates in soil solutions[END_REF]Raessler et al. 2010;[START_REF] Raessler | Sample preparation and current applications of liquid chromatography for the determination of non-structural carbohydrates in plants[END_REF], the most recommended are: (1) the CarboPac PA1 column for analysis of neutral sugars [START_REF] Cheng | Improved Analysis of Dissolved Carbohydrates in Stream Water with HPLC-PAD[END_REF][START_REF] Kerhervé | Neutral monosaccharides in surface sediments of the northwestern Mediterranean Sea[END_REF][START_REF] Panagiotopoulos | The composition and flux of particulate and dissolved carbohydrates from the Rhone River into the Mediterranean Sea[END_REF]Panagiotopoulos and Sempéré, 2005a;[START_REF] Theodosi | Sugars in atmospheric aerosols over the Eastern Mediterranean[END_REF], and (2) the CarboPac MA1 column for analysis of alditols, anhydrosugars and neutral sugars [START_REF] Iinuma | A highly resolved anion-exchange chromatographic method for determination of saccharidic tracers for biomass combustion and primary bio-particles in atmospheric aerosol[END_REF][START_REF] Jung | Quantitative determination of the biomass-burning contribution to atmospheric carbonaceous aerosols in Daejeon, Korea, during the rice-harvest period[END_REF][START_REF] Zhang | Determination of isoprene-derived secondary organic aerosol tracers (2methyltetrols) by HPAEC-PAD: Results from size-resolved aerosols in a tropical rainforest[END_REF].

The CarboPac PA1 column is well-suited for the analysis of neutral monosaccharides.

However, when alditols and anhydrosugars are included in the same run, a NaOH gradient is required because these compounds are eluted at lower NaOH concentrations (<1 mM) compared to neutral monosaccharides (>15 mM). However, the use of a NaOH gradient causes disequilibrium of the baseline signal, which further results in a positive [START_REF] Sullivan | Application of high-performance anion-exchange chromatography-pulsed amperometric detection for measuring carbohydrates in routine daily filter samples collected by a national network: 2. Examination of sugar alcohols/polyols, sugars, and anhydrosugars in[END_REF] or a negative peak [START_REF] Caseiro | Determination of saccharides in atmospheric aerosol using anion-exchange highperformance liquid chromatography and pulsed-amperometric detection[END_REF][START_REF] Theodosi | Sugars in atmospheric aerosols over the Eastern Mediterranean[END_REF] that may potentially mask sugar peaks that are eluted within this time window.

The CarboPac MA1 column was introduced much later than the CarboPac PA1 column and, to date, has never been applied to marine samples. Nevertheless, its macroporous stationary phase has a higher anion-exchange capacity over the CarboPac PA1 column (1450 μ q/c m vs 100 μ q/c m ), which further allows the use of gradient conditions without affecting the baseline stability [START_REF] Iinuma | A highly resolved anion-exchange chromatographic method for determination of saccharidic tracers for biomass combustion and primary bio-particles in atmospheric aerosol[END_REF][START_REF] Zhang | Determination of isoprene-derived secondary organic aerosol tracers (2methyltetrols) by HPAEC-PAD: Results from size-resolved aerosols in a tropical rainforest[END_REF]. Moreover, because the CarboPac MA1 column operates at high NaOH elution conditions (>200 mM), the pH values of the mobile phase are >12.5 and as such all the hemiacetal groups of sugars are deprotonated. This further increases the retention of sugars on the column and thus results in a better resolution (Thermo Fisher, Technical Note 20). Finally, the higher baseline stability of the CarboPac MA1 column over the CarboPac PA1 column offers an additional advantage in terms of the retention time of chromatographic peaks, which does not significantly shift over successive injections [START_REF] Gremm | Dissolved carbohydrates in streamwater determined by HPLC and pulsed amperometric detection[END_REF]. Due to these reasons, this study was conducted using the CarboPac MA1 column and different chromatographic parameters (flow rate, etc.) were appropriately adjusted for separation of different classes or sugars.

Optimization of flow rate, mobile phase and column temperature

According to the manufacturer, the CarboPac MA1 column operates at a recommended range of flow rates of 0.2-0.5 mL min -1 , and the flow rate is generally fixed at 0.4 mL min -1 for environmental applications [START_REF] Iinuma | A highly resolved anion-exchange chromatographic method for determination of saccharidic tracers for biomass combustion and primary bio-particles in atmospheric aerosol[END_REF][START_REF] Jung | Quantitative determination of the biomass-burning contribution to atmospheric carbonaceous aerosols in Daejeon, Korea, during the rice-harvest period[END_REF][START_REF] Li | Atmospheric outflow of PM2.5 saccharides from megacity Shanghai to East China Sea: Impact of biological and biomass burning sources[END_REF].

Nevertheless, the latter authors did not report co-elution problems because they did not consider fucose, rhamnose, arabinose, sorbitol and xylose in their standards. To overcome the co-elution problem, we first decreased the flow rate to 0.3 mL min -1 . This resulted in a slight improvement in the separation of the above compounds, and therefore this flow rate was maintained for all further improvements.

Previous investigations employing the CarboPac MA1 column used NaOH concentrations ranging from 300 to 800 mM [START_REF] Andersen | Separation and determination of alditols and sugars by high-pH anion-exchange chromatography with pulsed amperometric detection[END_REF] to elute alditols, anhydrosugars, and some neutral sugars [START_REF] Iinuma | A highly resolved anion-exchange chromatographic method for determination of saccharidic tracers for biomass combustion and primary bio-particles in atmospheric aerosol[END_REF]. Nonetheless, such conditions were not optimal for the 17 carbohydrates considered in this study. Therefore, different NaOH gradient tests were performed at a fixed column temperature of 25 °C and a flow rate of 0.3 mL min -1 . The improved separation of the first seven eluted monosaccharides (xylitol, levoglucosan, arabitol, fucose, rhamnose, sorbitol and mannosan) was achieved by applying an NaOH gradient ranging from 250 mM to 350 mM in the first 30 min (Fig. 1 a, Table 1). Unfortunately, fucose and rhamnose were not resolved (R s = 0) under these conditions despite additional adjustments of the NaOH gradient. The 250 to 350 mM gradient was then followed by a 15 min-NaOH gradient increasing from 350 to 450 mM that allowed an acceptable separation (with resolution factors close to or higher than 1) of mannitol, galactosan, arabinose and mannose.

Following the chromatographic separation after 45 min, the NaOH concentration was increased from 450 to 700 mM and held for 55 min to elute glucose, xylose, galactose and fructose. Under these elution conditions, the xylose/galactose and galactose/fructose pairs were fully resolved (R s > 1.5), while the glucose/xylose pair was partially resolved (R s = 0.3).

Additional tests of decreasing or increasing the final NaOH concentration (700 mM) did not improve the glucose/xylose resolution. Finally, by prolonging the 700 mM NaOH concentration for an additional 20 min, the last two carbohydrates (ribose and sucrose) were eluted with excellent resolution (R s = 8.1). After 75 min, the column was equilibrated for 20 min with the initial NaOH concentration (250 mM). Regeneration and clean-up of the column were not necessary, because the high NaOH concentration (700 mM) reached at the end of the analysis was sufficient to restore column performance.

Because pairs of levoglucosan/arabitol, fucose/rhamnose, galactosan/arabinose, arabinose/mannose and glucose/xylose were completely or partially overlapped under the NaOH gradient conditions optimized above, we also explored the effect of temperature on the resolution of these monosaccharide pairs. Earlier investigations have shown that temperature plays an important role in monosaccharide separation using the CarboPac PA1 column and that sub-ambient temperatures strongly affect the resolution of closely eluting monosaccharide pairs, notably fucose/rhamnose and mannose/xylose [START_REF] Panagiotopoulos | Sub-ambient temperature effects on the separation of monosaccharides by high-performance anion-exchange chromatography with pulse amperometric detection -Application to marine chemistry[END_REF]. The results of this study indicated that glucose co-eluted with xylose at temperatures <20 °C, while arabinose overlapped with mannose at >30 °C. Moreover, fucose and rhamnose always co-eluted under both these temperature conditions. Overall, these results suggest that sub-ambient temperature conditions do not have a significant impact on the separation of the abovementioned carbohydrate pairs on the CarboPac MA1 column.

By narrowing the temperature range between 25 and 30 °C, we found that these monosaccharide pairs were better resolved (except the fucose/rhamnose pair which again coeluted), as shown by their corresponding resolution factors (Fig. 2). The results showed that except for the arabinose/mannose (R s = 0.4) and fucose/rhamnose pairs (R s = 0), the resolution factors for the levoglucosan/arabitol (R s = 0.7), galactosan/arabinose (R s = 1.3), and glucose/xylose (R s = 0.6) pairs were significantly improved at a temperature of 28 °C.

Therefore, a temperature gradient was applied between 25 and 28 °C for the first 30 min of analysis. The latter temperature was maintained for the next 45 min to distinguish the galactosan/arabinose, arabinose/mannose and glucose/xylose pairs (Table 1,Table A 1 until the next injection.

Linearity, detection limit, precision and blanks

The optimized method showed a linear response for all monosaccharides (R 2 >0.999) over a concentration range of 50 nM to 10 µM, which is typical for environmental monosaccharide concentrations. The detection limits calculated at a signal-to-noise ratio (S/N) of three were 4-51 nM for neutral sugars, 2-15 nM for alditols and 8-32 M for anhydrosugars (Table A 1), in agreement with previous studies using different elution conditions and anion-exchange columns for monosaccharide determination [START_REF] Caseiro | Determination of saccharides in atmospheric aerosol using anion-exchange highperformance liquid chromatography and pulsed-amperometric detection[END_REF][START_REF] Iinuma | A highly resolved anion-exchange chromatographic method for determination of saccharidic tracers for biomass combustion and primary bio-particles in atmospheric aerosol[END_REF][START_REF] Panagiotopoulos | Sub-ambient temperature effects on the separation of monosaccharides by high-performance anion-exchange chromatography with pulse amperometric detection -Application to marine chemistry[END_REF][START_REF] Theodosi | Sugars in atmospheric aerosols over the Eastern Mediterranean[END_REF]. The high detection limit obtained for fructose (51 nM), also observed in a previous study [START_REF] Iinuma | A highly resolved anion-exchange chromatographic method for determination of saccharidic tracers for biomass combustion and primary bio-particles in atmospheric aerosol[END_REF], is probably due to the low detector response toward this compound. The precision of the method was calculated by performing six identical runs with the standard mixture of the 17 carbohydrates (50 nM each). The relative standard deviation (RSD%) ranged from 1.8 to 16.6% for the peak area and from 0.2 to 1.7% for the retention time (Table A 1). The procedural blank for the marine POM consisted of a 25-mm Whatman GF/F filter (precombusted at 450 °C for 6 h), hydrolyzed in the same manner as the samples. Similarly, the procedural blank for the TSP consisted of a Whatman quartz fiber filter (precombusted at 450 °C for 6 h) extracted with the ultrapure water. The results showed no detectable sugars in blanks (Fig. S 1).

Comparison with the CarboPac PA1 column

The final optimized MA1-HPAEC-PAD method (Table 1, Fig. 1 a) was compared with a different method employing the CarboPac PA1 column [START_REF] Caseiro | Determination of saccharides in atmospheric aerosol using anion-exchange highperformance liquid chromatography and pulsed-amperometric detection[END_REF][START_REF] Theodosi | Sugars in atmospheric aerosols over the Eastern Mediterranean[END_REF]. The chromatogram obtained with the latter column (Fig. 1

Application to environmental samples

The method established in this study was tested on three different environmental matrices including TSP, marine POM and HMWDOM. The chromatograms of these matrices are presented in Fig. 3 and indicated the presence of large quantities of monosaccharides (Table 2).

TSP

The major carbohydrate classes identified in the TSP sample included anhydrosugars and disaccharides, followed by neutral sugars and alditols. The sum of their concentration was 563 ng m -3 , representing the total neutral carbohydrate (TCHO) content recorded in the TSP sample. The results of this study showed that levoglucosan (261 ng m -3 ) and sucrose (178 ng m -3 ) were among the major neutral carbohydrates (Table 2; Fig. 3 a), which is in agreement with other studies of European urban atmospheres [START_REF] Yttri | Ambient aerosol concentrations of sugars and sugaralcohols at four different sites in Norway[END_REF]. Anhydrosugars represented 53% of TCHO in the TSP sample and were dominated by levoglucosan and mannosan, which accounted for 87% and 10% of the anhydrosugar pool, respectively (Fig. 4 a). Galactosan was also detected in the TSP sample but in lower concentrations (Table 2).

Neutral sugars accounted for 11% of the TCHO pool (Fig. 4 a) with fructose and glucose exhibiting the highest concentrations (Fig. 4 a, Table 2). The presence of these neutral sugars in the TSP sample strongly supports the influence of biogenic emission, which is typical of plants growth during spring [START_REF] Fu | Seasonal variations of sugars in atmospheric particulate matter from Gosan, Jeju Island: Significant contributions of airborne pollen and Asian dust in spring[END_REF][START_REF] Medeiros | Sugars as source indicators of biogenic organic carbon in aerosols collected above the Howland Experimental Forest, Maine[END_REF]. Other minor neutral sugars found in the TSP sample included the arabinose, mannose and xylose (Table 2).

Finally, alditols represented the least abundant carbohydrate class, accounting for 4% of TCHO in the TSP sample (Fig. 4 a). The alditols decreased in abundance in the order of xylitol, mannitol, arabitol and sorbitol (Table 2), and their presence in the sample indicates inputs from fungal spores and/or microbial activities [START_REF] Bauer | Arabitol and mannitol as tracers for the quantification of airborne fungal spores[END_REF][START_REF] Dahlman | Carbon and nitrogen distribution in the green algal lichens Hypogymnia physodes and Platismatia glauca in relation to nutrient supply[END_REF][START_REF] Loos | Sorbitol promotes growth of zymomonas-mobilis in environments with high-concentrations of sugar -evidence for a physiological-function of glucose-fructose oxidoreductase in osmoprotection[END_REF].

Although fucose and rhamnose co-eluted under the current HPAEC-PAD conditions, they were also identified and quantified in the aerosol sample for the first time and accounted for 1% of TCHO and 8% of neutral sugars. Fucose and rhamnose may have multiple sources in atmospheric particles including soil organic matter [START_REF] Gunina | Sugars in soil and sweets for microorganisms: Review of origin, content, composition and fate[END_REF][START_REF] Ogner | Analysis of the carbohydrates of fulvic and humic acids as their partially methylated alditol acetates[END_REF][START_REF] Simoneit | Sugars -Dominant Water-Soluble Organic Compounds in Soils and Characterization as Tracers in Atmospheric Particulate Matter[END_REF], vascular plants [START_REF] Bianchi | Biogeochemistry of estuaries[END_REF][START_REF] Popper | 3-O-methylrhamnose in lower land plant primary cell walls[END_REF][START_REF] Schädel | Quantification and monosaccharide composition of hemicelluloses from different plant functional types[END_REF], microorganisms and bacteria [START_REF] Cowie | Carbohydrate sources in a coastal marine environment[END_REF][START_REF] Fox | Determination of carbohydrate profiles of Bacillus anthracis and Bacillus cereus including identification of O-methyl methylpentose by using gas chromatography-mass spectrometry[END_REF][START_REF] Petit | Involvement of a Bacterial Microcompartment in the Metabolism of Fucose and Rhamnose by Clostridium phytofermentans[END_REF], while recent studies indicated that marine sources via water/air exchange may also be an important contributor (Rastelli et al., 2017).

Nevertheless, field measurements using gas chromatography [START_REF] Alves | Emission of trace gases and organic components in smoke particles from a wildfire in a mixed-evergreen forest in Portugal[END_REF] or nuclear magnetic resonance spectroscopy [START_REF] Chalbot | NMR Analysis of the Water-Soluble Fraction of Airborne Pollen Particles[END_REF] identified only fucose or rhamnose but never simultaneously reported both sugars in atmospheric samples. On the other hand, studies using HPAEC-PAD have never reported fucose and rhamnose in atmospheric particles, probably because of the co-elution of these deoxysugars with other monosaccharides (fucose with galactosan and rhamnose with arabinose after analysis with the CarboPac PA1 column; Fig. 1 b). Therefore, the presence of fucose and rhamnose in the sample has never been thoroughly assessed to date.

The lack of data on deoxysugars in atmospheric samples may also be related to their original structure. As deoxysugars are mostly found in bacterial lipopolysaccharides [START_REF] Perry | Characterization of the Opolysaccharide structure of lipopolysaccharide from Actinobacillus actinomycetemcomitans serotype b[END_REF][START_REF] Weckesser | The identification of 3-O-methyl-L-rhamnose (Lacofriose) as constituent of the lipopolysaccharide of Rhodopseudomonas capsulata[END_REF][START_REF] Zdorovenko | Composition, structure, and biological properties of lipopolysaccharides from different strains of Pseudomonas syringae pv[END_REF] and/or plant structural polysaccharides [START_REF] Colombini | Ion chromatography characterization of polysaccharides in ancient wall paintings[END_REF][START_REF] Grössl | Characterisation of natural polysaccharides (plant gums) used as binding media for artistic and historic works by capillary zone electrophoresis[END_REF][START_REF] Popper | 3-O-methylrhamnose in lower land plant primary cell walls[END_REF][START_REF] Schädel | Quantification and monosaccharide composition of hemicelluloses from different plant functional types[END_REF], they are either not efficiently recovered after water extraction (case of lipopolysaccharides) or are recovered in the water phase but cannot be released from carbohydrate polymer (case of structural polysaccharides) as no hydrolysis is applied.

Marine POM

The major carbohydrate classes identified in the marine POM sample included the neutral sugars followed by anhydrosugars and alditols. The sum of their concentration was 16.2 mg g -1 . Neutral sugars accounted for 98% of the TCHO pool (Fig. 4 b) and were dominated by galactose (5.7 mg g -1 ) and glucose (3.8 mg g -1 ) (Table 2; Fig. 3 b). These monosaccharides were followed by fructose (2.0 mg g -1 ) and fucose/rhamnose (co-eluted; 1.8 mg g -1 ) while xylose, mannose and arabinose exhibited concentrations <1 mg g -1 . These results are in good agreement with previously reported monosaccharide patterns for marine POM in several oceanic regimes [START_REF] Hernes | Neutral carbohydrate geochemistry of particulate material in the central equatorial Pacific[END_REF]Panagiotopoulos and Sempéré, 2005b;[START_REF] Skoog | A a z f ac f ma ga c ma : Implications for carbon cycling[END_REF]. Fructose and ribose, occasionally reported in marine POM samples due to their low recovery after acid hydrolysis [START_REF] Borch | Concentration and composition of dissolved combined neutral sugars (polysaccharides) in seawater determined by HPLC-PAD[END_REF]Panagiotopoulos and Sempéré, 2005a), were also identified in this sample (Table 2). This finding most likely indicates the presence of storage polysaccharides (fructans) and nonstructural labile compounds such as RNA or nucleotides originating from marine organisms [START_REF] Cowie | Carbohydrate sources in a coastal marine environment[END_REF][START_REF] Haug | Polysaccharides of marine diatoms with special reference to Chaetoceros species[END_REF][START_REF] Hicks | Deposition, resuspension, and decomposition of particulate organic matter in the sediments of Lake Itasca, Minnesota, USA[END_REF]Panagiotopoulos and Sempéré, 2005b). Alternatively, fructose may originate from sucrose. As sucrose was not detected in the marine POM sample (the same holds for the HMWDOM sample), this most likely implies its complete hydrolysis under the acid conditions used for its extraction, which results in two monosaccharides (fructose and glucose).

The proposed analytical method allowed the identification and quantification of alditols (0.4% of TCHO) and anhydrosugars (2% of TCHO), the two monosaccharide families that have been largely overlooked to date in marine environmental studies (Fig. 4 b). The results

showed that xylitol was the major alditol type detected in the POM sample, followed by sorbitol, while arabitol and mannitol were not detected in the sample (Table 2). Alditols are generally reported as terrestrial tracers derived from fungal spores and/or microbial activities [START_REF] Dahlman | Carbon and nitrogen distribution in the green algal lichens Hypogymnia physodes and Platismatia glauca in relation to nutrient supply[END_REF][START_REF] Vandeska | Effects of environmental conditions on production of xylitol by Candida boidinii[END_REF], therefore, their presence in the marine POM sample may suggest possible terrestrial input [START_REF] Van Pinxteren | Chemical characterization of dissolved organic compounds from coastal sea surface microlayers (Baltic Sea, Germany)[END_REF]. Alternatively, alditols may have marine origin. As carbohydrates are an important fraction of the organic matter produced by marine primary production or released into the environment via degradation processes, it is highly possible that alditols are also part of the carbohydrate pool.

Nevertheless, additional measurements using compound-specific 13 C analysis are warranted to test this hypothesis and constrain the sources of alditols in the marine waters.

Compositionally, anhydrosugars were dominated by galactosan followed by levoglucosan and mannosan, which represented for 63%, 25% and 13% of total anhydrosugars, respectively (Fig. 4 b). The dominance of galactosan over levoglucosan has also been reported for sea surface microlayer samples in Baltic sea [START_REF] Van Pinxteren | Chemical characterization of dissolved organic compounds from coastal sea surface microlayers (Baltic Sea, Germany)[END_REF] during different seasons of the year however, more data are warranted in the same time from both atmospheric and marine compartments to validate/confirm these findings.

However, the anhydrosugar concentrations found in this sample (and in the HMWDOM sample) are most likely underestimated because these compounds are converted to their respective hexoses after acid hydrolysis [START_REF] Blanco | Production of Glucose from the Acid Hydrolysis of Anhydrosugars[END_REF]. Indeed, in an additional set of experiments to investigate whether the hydrolysis conditions affect the stability of monosaccharides, we performed acid hydrolysis (1 M HCl, 100 °C, 20 h) on a standard monosaccharide mixture containing neutral sugars, alditols and anhydrosugars. The results showed that levoglucosan, mannosan and galactosan were completely converted to glucose, mannose and galactose, respectively, while minimal loss was observed for neutral sugars and alditols (Fig. S 2), which is consistent with previous studies [START_REF] Skoog | A a z f ac f ma ga c ma : Implications for carbon cycling[END_REF][START_REF] Wang | Influences of acidic reaction and hydrolytic conditions on monosaccharide composition analysis of acidic, neutral and basic polysaccharides[END_REF].

Overall, these results revealed that the concentration of anhydrosugars is biased in environmental samples if these sugars are acid-extracted (POM and HMWDOM sample).

Therefore, an alternative extraction method should be considered (e.g., extraction with water or MeOH). Despite the possible underestimation of the amount of the anhydrosugars found in the marine POM, their presence in this sample clearly indicates an external input, most likely via atmospheric deposition, as shown by the isotopic signature (δ 13 C) of levoglucosan and mannosan pointing to a terrestrial origin [START_REF] Nouara | Liquid chromatographic isolation of individual carbohydrates from environmental matrices for stable carbon analysis and radiocarbon dating[END_REF].

Marine HMWDOM

The major carbohydrate classes identified in the HMWDOM sample included the neutral sugars followed by anhydrosugars and alditols. The sum of their concentration was 766 µg g -1 . Neutral sugars accounted for 97% of the TCHO pool in the HMWDOM (Fig. 4 c). The results of this study revealed that fucose & rhamnose (300 µg g -1 ), glucose (122 µg g -1 ), mannose (103 µg g -1 ), galactose (83 µg g -1 ), xylose (74 µg g -1 ), and arabinose (34 µg g -1 )

were among the major monosaccharides, which agrees with the neutral sugar pattern reported in previous studies [START_REF] Aluwihare | A major biopolymeric component to dissolved organic carbon in surface sea water[END_REF]Repeta et al., 2002) (Table 2; Fig. 3c). Fructose and ribose were also quantified in the HMWDOM sample, and similar to POM, they most likely originate from storage polysaccharide or RNA.

Anhydrosugars accounted for 3% of TCHO and were dominated by levoglucosan, galactosan and mannosan, which represented 61%, 22% and 17% of total anhydrosugars, respectively (Fig. 4 c). The lower concentration of anhydrosugars in the HMWDOM sample (the same holds for the POM sample) compared to the TSP sample maybe due to the high dilution occurring at sea and also to photodegradation processes by OH * radicals which take place in the atmosphere for these compounds [START_REF] Hoffmann | Atmospheric stability of levoglucosan: a detailed laboratory and modeling study[END_REF][START_REF] Lai | Degradation kinetics of levoglucosan initiated by hydroxyl radical under different environmental conditions[END_REF]. In addition, as suggested for marine POM, the concentration of anhydrosugars in HMWDOM is most likely biased due to the acid extraction procedure. The presence of the anhydrosugars in the sample may indicate possible input of terrestrial organic matter (burning biomass tracers), although additional studies using  13 C are warranted to confirm this hypothesis. Finally, the least abundant carbohydrate class was the alditols, which accounted 0.3% of TCHO and represented by mannitol and xylitol (Fig. 4 c).

Conclusions

The results of this study indicated that under the optimized analytical conditions, neutral sugars, alditols, and anhydrosugars commonly found in most environmental samples can be analyzed with a single run in 75 min with satisfactory resolution except for the fucose/rhamnose pair that completely co-eluted. These deoxysugars were identified for the first time in the TSP sample by this improved HPAEC-PAD technique, which will facilitate future investigations by shedding light on bacterial and/or microorganism activities on atmospheric samples. However, additional deconvolution of these deoxysugars signals using the traditional HPAEC-PAD method for neutral sugar determination [START_REF] Panagiotopoulos | Sub-ambient temperature effects on the separation of monosaccharides by high-performance anion-exchange chromatography with pulse amperometric detection -Application to marine chemistry[END_REF] might also be useful.

The proposed improved HPAEC-PAD method allowed, for the first time, the identification and quantification of anhydrosugars and alditols in marine samples (marine POM and HMWDOM). These two carbohydrate families have largely been overlooked in the dissolved free monosaccharide pool (DFMS), and, to the best of our knowledge, they have never been reported in marine DOM. Indeed, anhydrosugars and alditols can easily be extracted from seawater using cation and anion-exchange resins [START_REF] Kirchman | Glucose fluxes and concentrations of dissolved combined neutral sugars (polysaccharides) in the Ross Sea and Polar Front Zone, Antarctica[END_REF][START_REF] Mopper | Determination of sugars in unconcentrated seawater and other natural waters by liquid chromatography and pulsed amperometric detection[END_REF][START_REF] Sempéré | Distribution and bacterial availability of dissolved neutral sugars in the South East Pacific[END_REF][START_REF] Skoog | A a z f ac f ma ga c ma : Implications for carbon cycling[END_REF][START_REF] Van Pinxteren | Chemical characterization of dissolved organic compounds from coastal sea surface microlayers (Baltic Sea, Germany)[END_REF] in the same manner as neutral sugars. However, because the subsequent PA1-HPAEC-PAD analysis has always been performed at >15 mM NaOH to elute the neutral sugars; anhydrosugars and alditols always co-elute at time window of 2-5 min, therefore, they have never been identified in the DFMS pool.

On the other hand, dissolved combined monosaccharides (DCHO) or polysaccharides in marine samples are generally measured as their monomeric constituents that are released after acid hydrolysis. Thus, if anhydrosugars are present in the sample, they will be counted in the hexose pool as the sample is hydrolyzed. Therefore, we recommend that DCHO analysis always be preceded or accompanied by the analysis of DFMS to evaluate the contribution of anhydrosugarsif presentin the sample. This strategy is particularly critical for samples from coastal areas that receive significant terrestrial inputs, particularly in wintertime, and are characterized by strong influence from wood burning.

The proposed HPAEC method provides an acceptable resolution for most of the carbohydrates examined, and therefore its coupling with mass spectrometry may further help with the structural elucidation and confirmation of the analyzed carbohydrates. Moreover, its high potential may further be explored in other environmental matrices, including sediments, seawater, and marine biota, which will provide a more complete profile of carbohydrates in the environment. This complete profile, in turn, may be helpful to improve our understanding on carbohydrate dynamics in the marine and terrestrial ecosystems and potentially better evaluate the impact of biomass burning processes on marine waters.

Table of Contents

Table S 1. HPAEC-PAD performance after optimization (Table 1). The average retention time (t R ), and resolution factor (R s ) of the carbohydrate standards is given at 1 µM level (n = 3). The relative standard deviations (RSD) of peak area and retention time of the same standards is calculated at 50 nM level (n = 6), except for fructose (1 µM level; n = 6). The detection limit (DL) was calculated at signal to noise ratio of 3…………………………………………………..

  ; Fig.1a). After 75 min, the temperature was reset to 25 °C during column re-equilibration (20 min)

Fig. 1 .

 1 Fig. 1. HPAEC-PAD chromatograms of a standard mixture of 17 monosaccharides (1 µM

Fig

  Fig. 2. h f g c a /a ab (♦) f c / ham (×), galactosan/arabinose

Fig. 3 .Fig. 4 .

 34 Fig. 3. HPAEC-PAD chromatograms according to Table 1 of (a) total suspended atmospheric

  FIGURE 1

Fig S 1 .Fig S 2 .

 12 Fig S 1. Example of HPAEC-PAD chromatograms of procedural blanks (a) ultrapure water (b) TSP blank and (c) POM blank. Blanks were prepared in triplicate for each sample (TSP and marine POM) and run according to the method established ab 1…………………………………………..46

Fig. S 2 .

 2 Fig. S 2. High-performance liquid chromatography with refractive index detection (HPLC-RI) chromatograms of a monosaccharide mixture (1 mM each) before hydrolysis (black solid line) and after hydrolysis (red dotted line) with 1 M HCl under 100 °C for 20 h. Peak identification : 1 = glucose, 2 = xylose, 3 = galactose & rhamnose, 4 = galactosan, 5 = arabinose & fucose, 6 = mannose, 7 = fructose, 8 = mannitol, 9 = levoglucosan, 10 = xylitol, 11 = sorbitol, 12 = mannosan. The analytical HPLC-RI conditions are given in Nouara et al. (2019) and are as follows: cation exchange column in Pb 2+ form, isocratic elution with ultrapure water at 0.6 mL min -1 , column temperature 75 °C.

  ). A mixture of standard solutions at concentrations ranging from 50 nM to 10 µM was prepared by dilution with ultrapure water of a standard stock solution (1 mM) of neutral sugars (fucose, rhamnose, arabinose, galactose, glucose, mannose, xylose,

	fructose and ribose), alditols (xylitol, arabitol, sorbitol and mannitol), anhydrosugars
	(levoglucosan, mannosan and galactosan), and one disaccharide (sucrose). All the prepared
	solutions including the stock solution were stored	h a k a -25 °C	. Despite that
	no degradation was observed after melting and refrozen, h a a	(50 M-10
	µM) were renewed after 2 weeks. The HCl solution (37%, Sigma-Aldrich) diluted with
	ultrapure water to 1 M was used for sample hydrolysis, while the NaOH solution (50% w/v in
	H

2 O, low carbonate; PanReac AppliChem ITW Reagents) diluted with ultrapure water was used for the mobile phase of the HPAEC-PAD chromatography. The ultrapure water used in this work was produced by a Millipore Milli-Q system (Molsheim, France).

2.2.

Sampling and extraction of carbohydrates

2.2.1. TSP

The aerosol sample was collected on a precombusted (450 °C, 6 h) weighed Whatman quartz fiber filter (20.3 cm × 25.4 cm, nominal retention size, 0.7 µm) using an automatic sampler (Tisch Environmental USA; flow rate 85 m 3 h -1 ). The sample was collected from March 10th to March 17th of 2016, from the rooftop of the Endoume Marine Station (Marseille; 43° 16' N -5° 21' E). After collection, the sample was dried for 24 h in a desiccator, weighed, and then a f z a -25 °C in precombusted aluminum foil

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	for		higher	accuracy	of	data.	Anal.	Methods	2,	532-538.
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	to the special issue on the Biogeochemistry of Dissolved Organic Matter. Mar. Chem.
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	Paquot, M., 2007. Isolation and identification of inulooligosaccharides resulting from
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Le Meur, S., Descheemaecker, M., Chretien, E., Uzu, G., 2019. Polyols and glucose particulate species as tracers of primary biogenic organic aerosols at 28 french sites.

Table and Figure captions Table 1 :

 andcaptions1 Optimal analytical conditions for the simultaneous analysis of neutral sugars, alditols, anhydrosugars and sucrose

Table 2 .

 2 Carbohydrate composition in atmospheric TSP, marine POM and marine HMWDOM samples. The carbohydrate concentrations presented in this table are not corrected for losses during the hydrolysis or the extraction procedure (see materials and methods), and therefore they should be considered as minimum concentrations found in the samples.

Table 1

 1 Optimal analytical conditions for the simultaneous analysis of neutral sugars, alditols, anhydrosugars and sucrose.

	Parameters		Conditions
	Column		CarboPac MA1 (250 × 4 mm I.D.), Thermo Fisher
	Eluent flow rate (mL min	-1 )	0.3
	Detector temperature (°C)	25
	Sample loop size (µL)		250
	Injection volume (µL)		230
	Column oven temperature (°C)	25-28	(0-30 m ); c	5
			28	(30-75 m )
			25	(75-95 m )
	NaOH gradient (mM)		250-350 (0-30 m ) ; curve 5
			350-450 (30-45 m ) ; curve 5
			450-700 (45-55 m ) ; curve 5
			700	(55-75 m )
			250	(75-95 m )

Table 2

 2 Carbohydrate composition in atmospheric TSP, marine POM and marine HMWDOM samples. The carbohydrate concentrations presented in this table are not corrected for losses during the hydrolysis or the extraction procedure (see materials and methods), and therefore they should be considered as minimum concentrations found in the samples. Xylitol concentration was outside the calibration range (50-1000 nM) but ~6 times higher than its detection limit (Table A 1), thus it was quantified with an uncertainty of ~24%.

	Carbohydrate	TSP (ng m -3 )	POM (mg g -1 )	HMWDOM (µg g -1 )
	Xylitol	11	0.05	0.13*
	Levoglucosan	261	0.08	12
	Arabitol	3.4	ND	ND
	Fucose & Rhamnose	5.0	1.8	300
	Sorbitol	1.9	0.02	ND
	Mannosan	30	0.04	3.4
	Mannitol	7.8	ND	2.3
	Galactosan	9.4	0.2	4.4
	Arabinose	2.1	0.1	34
	Mannose	1.2	0.8	103
	Glucose	15	3.8	122
	Xylose	1.2	0.9	74
	Galactose	ND	5.7	83
	Fructose	36	2.0	11
	Ribose	ND	0.7	19
	Sucrose	178	ND	ND
	ND = Not detected

*

Table S 1

 S .HPAEC-PAD performance after optimization (Table1). The average retention time (t R ), and resolution factor (R s ) of the carbohydrate standards is given at 1 µM level (n = 3). The relative standard deviations (RSD) of peak area and retention time of the same standards is calculated at 50 nM level (n = 6), except for fructose (1 µM level; n = 6). The detection limit (DL) was calculated at signal to noise ratio of 3.

	Carbohydrate	t R (min)	R s	Area RSD (%)	t R RSD (%)	DL (nM)
	Xylitol	19.9	3.1	5.6	0.2	2
	Levoglucosan	23.8	0.6	4.0	0.5	8
	Arabitol	24.4	1.3	6.7	0.4	13
	Fucose & Rhamnose	26.0	1.3	6.5	0.3	17
	Sorbitol	27.8	1.2	5.0	0.3	15
	Mannosan	29.3	3.2	1.9	0.3	18
	Mannitol	33.3	5.1	2.7	0.4	7
	Galactosan	40.2	0.8	9.3	0.4	32
	Arabinose	41.4	0.7	4.9	0.4	7
	Mannose	42.6	2.4	1.8	0.3	6
	Glucose	46.7	0.5	4.4	0.3	9
	Xylose	47.7	1.6	6.7	0.3	24
	Galactose	50.5	1.7	5.2	0.6	4
	Fructose	53.1	3.0	16.6	1.7	51
	Ribose	57.2	8.1	6.0	0.4	13
	Sucrose	68.9	-	5.8	0.7	13

Acknowledgments

This research was supported by the funding from the projects AIOLOS (Labex OT-Med; ANR-11-LABX-0.061) and MANDARINE (grant No 2008-10372; Région Provence Alpes Côte d'Azur). The authors thank D. Navarro for analytical assistance to conduct a part of this research at the INRA laboratory of Fungal Biodiversity and Biotechnology UMR 1163 (13009 Marseille; France). The authors also acknowledge A. Paulmier and D.J. Repeta for providing the POM sample and HMWDOM powder, respectively as well as the two anonymous reviewers for valuable comments and fruitful discussions.

TSP blank and (c) POM blank. Blanks were prepared in triplicate for each sample (TSP and marine POM) and run according to the method established in Table 1.