Impact of contaminant size and density on their incorporation into sea ice

Domain: earth_science
Published: 2025-05-12
Editorial outcome: minor_revision

Abstract
Sea ice accumulates contaminants and redistributes them laterally as the ice drifts and vertically as it melts. Contaminant incorporation into sea ice must be better understood to resolve contaminant cycling and exposure to polar organisms. Here we develop an experimental method that mimics the formation of young sea ice and enables the quantification of model contaminants separately in the ice matrix and brine. Several limitations inherent in field studies are overcome using this approach. Results show that dissolved contaminants (<1 nm) and dispersed colloidal contaminants (1 nm – 1 μm) follow the same behavior as sea salts. When colloids aggregate they follow a similar transport pathway to high-density particulate contaminants (>1 μm). While high-density particles are depleted in sea ice and low-density particles are enriched relative to their initial concentration in seawater, both are engulfed and can travel in wide brine channels. These results can also help to predict the incorporation of natural species in sea ice.

Introduction
There is substantial anthropogenic contamination even in some of the most remote areas on Earth, such as the Arctic Ocean1–3. Important fluxes of contaminants are brought to the Arctic Ocean where they can interact with and be incorporated into sea ice, due to the combination of high anthropogenic activity in the Northern hemisphere, patterns of oceanic circulation4, atmospheric circulation2,5, and riverine inputs to the Arctic Ocean6–8. Once in polar seawater, some contaminants are enriched in sea ice overall9,10, or are locally enriched within the sea ice porosity3,11. For example, microplastic (MPs, 1 μm–5 mm) concentrations were found to be orders of magnitude higher in sea ice compared to the underlying seawater9,10,12. MPs concentrations in Arctic sea ice9,12–14 are in the same order of magnitude as in Arctic sediments10,15–17, demonstrating that sea ice acts as a temporary sink of MPs. Therefore, the role of sea ice in global contaminant cycling must be assessed, since contaminants in sea ice can be redistributed laterally as sea ice drifts and vertically as it melts18,19.

Contaminant incorporation into sea ice and their specific location within sea ice must be understood to predict the risks they pose to polar organisms20,21. In particular, understanding the magnitude of incorporation from seawater to sea ice is necessary since i) substantial volumes of seawater freeze annually due climate change (i.e., the replacement of multi-year ice (MYI) by first-year ice (FYI)) and ii) since the seawater/sea ice interface is an important habitat for ice-associated organisms22,23. Indeed, microalgae which are the base of the polar food web, thrive in the bottom layer of sea ice due to the unique combination of porosity generated by brine pockets and channels, sunlight and nutrients with seawater24,25. This primary productivity could be further enhanced due to climate change, since thin and porous FYI allows for more sunlight and nutrient fluxes22,25.

The incorporation of contaminants into sea ice during freezing will depend on the freezing process and the contaminants’ physicochemical properties11,26. At the onset of freezing, frazil crystals form in the water column and rise to the surface. During this upward movement, they can scavenge particles such as suspended particulate matter27 and microalgae28 from the water column and enrich them in bulk sea ice. In contrast, most of the volume of Arctic sea ice is formed by bottom freezing, that is, the vertical advancement of the freezing front in calm conditions, which forms columnar ice. During bottom freezing, dissolved molecules such as sea salts are depleted from the bulk sea ice and accumulated in brine pockets and channels, a process known as salt segregation or partitioning29–31. This can be followed by gravity drainage, or flushing of the brine which leads to overall desalination of the bulk sea ice31. Likewise, dissolved (<1 nm) contaminants such as mercury as well as chlorinated and fluorinated molecules have also been shown to undergo the same transport pathway3,32,33. However, the effect of bottom freezing on the behavior of colloidal (1 nm to 1 μm) and particulate (>1 μm) contaminants is less understood34. In fact, the accumulation of MPs and nanoplastics (NPs, 1 nm to 1 μm) in bulk sea ice is often attributed to frazil ice without direct evidence35,36. Collectively, this highlights our lack of understanding of contaminant incorporation into sea ice.

Field studies have revealed the burden of contaminants in sea ice1,11. However, understanding the incorporation process of contaminants during freezing is limited by unknown initial concentrations of contaminants in seawater and the unknown conditions in which sea ice has formed. Furthermore, quantifying some contaminants, such as small MPs (<10 μm) and NPs, in environmental matrices is challenging due to the lack of standardized methods. Some persistent challenges, such as differentiating MPs and NPs from natural organic matter (NOM) and avoiding contamination during sampling and analysis require time-intensive methods. Overall, this limits the amount of samples which can be measured20,37,38. For these reasons, we assessed the incorporation of contaminants experimentally under laboratory conditions by generating artificial sea ice in a controlled environment and quantified the incorporation of model contaminants with an array of properties. We hypothesized that the incorporation of contaminants into sea ice would be dictated by their size and, in the case of particulate contaminants, also by their density. On one hand, colloidal species are expected to follow a similar incorporation pathway as dissolved species due to their Brownian motion, which allows them to diffuse away from the freezing front. On the other hand, particulate species with low densities are expected to be enriched in the sea ice relative to the underlying water, due to their positive buoyancy which moves them towards the advancing ice front. Conversely, high-density particles are expected to be depleted by sinking below the advancing ice front. Contrary to dissolved and colloidal species, particles are not expected to be preferentially enriched in the brine but instead may be engulfed by the ice due to the absence of Brownian motion.

To understand the mechanisms of contaminant incorporation into sea ice, we developed an experimental set-up which mimics the growth of columnar sea ice and quantified the incorporation of contaminants into this ice during freezing (Fig. 1). The experimental set-up i) generated environmentally representative artificial sea ice by keeping the salinity of the underlying water constant, and ii) allowed us to quantify a suite of model contaminants within the sea ice. To study the role of contaminant size and density, we used model species with a range of physicochemical properties including a dissolved molecule, two colloidal species (NPs and nanosoot), and four different MPs (two densities and two sizes; Table S1. and Fig. S1). The artificial sea ice formed was harvested to characterize its structure and to quantify species in the underlying liquid, the bulk ice, as well as its two fractions: the solid ice matrix (hereafter called ice) and the brine. This systematic approach allowed us to elucidate how contaminants become incorporated in sea ice. Our results improve our ability to predict how contaminants may be redistributed during sea ice drift and melt and which contaminants organisms living in sea ice will be exposed to.

Results
Experimental conditions resulted in environmentally realistic artificial sea ice
To assess contaminant incorporation into sea ice experimentally, we were careful to generate environmentally realistic sea ice. This was done by adding artificial seawater (ASW) containing a mixture of humic acid salt and sodium alginate to a column that underwent a vertical temperature gradient of −5.1 ± 0.1 °C to 1.0 ± 0.6 °C, for 19 h (Fig. 2a). The artificial sea ice produced was very similar to young columnar ice in terms of bulk salinity, brine salinity, porosity and structure.

At the end of the experiment, the underlying liquid was the dominant fraction accounting for 93.09 ± 1.27% of the total mass while ice and brine comprised 5.60 ± 1.02% and 1.04 ± 0.34% of the total mass, respectively (Fig. S2). This ensured that the salts which were expelled from the ice were strongly diluted in the underlying liquid, allowing ice to grow in realistic water salinities (Fig. 3, panel a). The brine salinity approximately doubled relative to the initial salinity, reaching 58.92 ± 5.75 g L−1. This brine salinity was compared to predictions of brine salinity as a function of temperature29. We used −1.7 °C as the temperature at the bottom of the ice core since it is the freezing temperature of our artificial seawater (31.28 g L−1) and −5.1 °C, as the temperature at the top of the ice core, since it is the temperature of the cooling element. This suggests that the brine salinity should be 57.80 g L−1 which is consistent with our observations. Conversely, the drained ice was strongly desalinated to 6.18 ± 0.95 g L−1 (Fig. 3, panel a). The porosity of the sea ice, which corresponds to the brine volume fraction, was 0.15 ± 0.04. The sea ice bulk salinity (calculated using brine and ice salinity and volumes) was 13.86 ± 2.30 g L−1. Note that since here “brine” refers to brine that is drained by centrifugation and “ice” refers to the residual drained ice, some small brine pockets or thin brine channels which were not effectively drained remained associated to and measured with the ice fraction.

The porosity and salinities of our artificial sea ice were very similar to those of natural young sea ice39. For example, in the Fram Strait, young sea ice had a median brine volume fraction of 0.12 and a brine salinity of 80 g L−140. Furthermore, in a broad assessment of young lab-grown and field-grown sea ice, brine volume fractions varied between 0.07 and 0.23 and bulk salinities ranged from 10 to 20 g L−141. Considering that the ambient temperature was 1 °C and that the temperature of the top of the artificial sea ice was −5.12 ± 0.08 °C, its bulk salinity of 13.86 ± 2.30 g L−1, brine volume fraction of 0.15 ± 0.04 and brine salinity of 58.92 ± 5.75 g L−1 fall within expected values of natural bulk ice grown under similar temperature forcings39,42. For example, Notz and Worster43 observed that sea ice of comparable thickness had a bulk salinity ranging from 15 to 25 g L−144. Our lower bulk salinity may be explained by the fact that brine may be lost when the artificial sea ice is pulled out of the column. The solid mass fraction of their sea ice ranged from 0.5 to 0.744 which is lower than our solid mass fraction of 0.85 ± 0.04. This can again be attributed to some brine loss, as well as some residual brine pockets within the ice that were not connected to the bottom of the ice, and therefore not drained by centrifugation. Despite these experimental limitations, our results agree well with the properties of natural sea ice. It should also be noted that this artificial sea ice produced represents the very early stages of sea ice growth, since young sea ice can reach up to 30 cm depth and can desalinate to 4 to 6 g L−139,44. Finally, the pH of the artificial sea ice cannot be directly compared to natural samples since biotic processes such as photosynthesis will modify pH, however, it is of interest to note that the changes in pH are not directly proportional to salinity (Fig. S3).

The ice cores had similar morphology to young columnar ice, despite variability between each ice core. The 3D rendition of the ice structure shows ice, air (where brine has been drained out) and some thin channels of brine which have not been drained by centrifugation (due to close-off of channels) (Figs. 3 panel b and S4). The top of each ice core had a first layer of short, vertically oriented brine pockets (~1 mm from the cooling element) followed by brine channels which are typical of those observed in natural sea ice41,43. The large channels (in gray) had mean widths of 121.2 ± 30.4 μm, while the thinner channels (in red) had mean widths of 50.5 ± 14.1 μm (Table S2). Some ice cores had a section of horizontally oriented and slightly curved lenticular structures which are not present in natural sea ice. This is likely caused by the fact that the ice started to grow on the freezing element which had a curved shape. This curved shape was used to allow air bubbles to escape as the cooling element was plunged into the water, to enable full contact between cooling the element and the water. Finally, some ice cores clearly showed the last of the ice growth with the parallel alignment of brine channels at the bottom.

Model contaminants had different distributions in the sea ice column after freezing
The various contaminants differentiated strongly in their tendency to be incorporated in the sea ice, depending on whether they were dissolved (<1 nm), colloidal (1 nm to 1 μm) or particulate (>1 μm) (see Table S1. and Fig. S1 for images of model colloidal and particulate contaminants). First, the incorporation of each contaminant was assessed by comparing their concentration (C) in the bulk ice (Cbulk ice which corresponds to brine and ice) normalized by their initial concentration in the column before freezing (C0) (Fig. 4). As mentioned above, the ice bulk salinity was reduced to 0.44 ± 0.07 of the initial salinity. Most other contaminants are depleted from the bulk ice, albeit in varying concentrations. The model dissolved contaminant, Rose Bengal, and the model colloidal contaminant, NP, were depleted in statistically similar proportions as sea salts (with Cbulk ice/C0 = 0.43 ± 0.01 and 0.38 ± 0.09, respectively). This was followed by the other colloidal contaminant, nanosoot, which was depleted to 0.48 ± 0.01 of its initial concentration.

The fate of particulate contaminants, was assessed by comparing high-density MPs and low-density MPs of two different size-classes resulting in four different MPs. High-density MPs were composed of polyethylene terephthalate (PET-MPs; 1.39 g cm−3) and low-density MPs composed of polypropylene (PP-MPs; 0.91 g cm−3). All MPs were fragments produced by cryo-milling and sieving, resulting in area equivalent diameter of 35.6 ± 14.4 μm and 45.7 ± 20.7 μm for small PET-MPs and small PP-MPs, and 90.0 ± 29.8 μm and 72.3 ± 37.2 μm for large PET-MPs and large PP-MPs, respectively. When inserting PP-MPs, many particles remained at air-liquid interface due to their lightweight and hydrophobicity. These PP-MPs did not interact with the advancing ice front as would be the case during bottom freezing in the environment. Therefore, this fraction of PP-MPs was removed from the evaluation of distribution in the column (Figs. 4, 5 and 6) but still considered when establishing the MPs recovery rate across the experiment (cf: Methods). When considering the portion of PP-MPs in the top layer of bulk ice, strong enrichment is observed as shown in Supplementary Fig. 5 (Cbulk ice/C0 = 7.37 ± 2.53 and 3.22 ± 1.31 for Large and Small PP-MPs, respectively).

Overall, clear density-dependent trends are observed, even when excluding the top layer of ice with PP-MPs that could have been adsorbed to the air/water interface (Fig. 4) The high-density PET-MPs are depleted and the low-density PP-MPs concentrations remain almost constant. Levels of depletion/enrichment increase with particle size (Supplementary Section 1). The Large PET-MPs were significantly more depleted from the bulk ice than Small PET-MPs with Cbulk ice/C0 = 0.06 ± 0.05 and 0.49 ± 0.25, respectively. This is explained by the fact that particle’s settling or rising velocity is directly proportional to the power of its size. Conversely, Large PP-MPs had a higher Cbulk ice/C0 value of 1.21 ± 0.59 than Small PP-MPs (0.86 ± 0.25).

To understand the mechanisms of contaminant incorporation in sea ice, we assessed the distribution of contaminants across the different sea ice fractions. To do so, brine and ice were separated to analyze the distribution of contaminants within the sea ice. The ratio of concentrations between brine and ice was measured as it reflects what happens at the freezing front and allows us to understand contaminants’ incorporation mechanisms. Also, the brine fraction of sea ice is where sympagic organisms thrive due to the availability of nutrients, the space and (sometimes) incoming sunlight22. Consequently, the partitioning of contaminants between brine and ice are an important consideration when assessing exposure. Therefore, we first describe the distribution patterns of solutes, followed by the distribution pattern of particles since they contrasted most. This is followed by a discussion of the distribution of colloidal contaminants, since their distribution was in between those of solutes and particles (Fig. 5).

The model dissolved contaminant, Rose Bengal, followed the same distribution pattern between ice, liquid and brine as sea salts. That is, relative to the initial concentration, it was strongly depleted in ice (C/C0 = 0.200 ± 0.012), enriched in the brine (C/C0 = 1.528 ± 0.033) and only slightly enriched in the underlying water (C/C0 = 1.009 ± 0.004) (Fig. 5). Rose Bengal and sea salt concentrations were not significantly different from each other in either ice or liquid (p = 0.971 and 0.012, respectively). However, Rose Bengal was less enriched in the brine compared to salts which had a C/C0 = 1.881 ± 0.210 (p = <0.001). Note that since brine comprised only ~1% of the total volume, and since the mass balance was constant, even large changes in concentration in the brine fraction only resulted in small concentration changes in the ice and liquid. This similarity in behavior is consistent with prior findings showing that natural or anthropogenic dissolved species (e.g., salts, nutrients and tracer molecules) partition out of the ice and are enriched in the brine and then depleted from bulk ice by gravity drainage42,45. Note that, as for the salts, it is also possible that some Rose Bengal in brine pockets was not centrifugated out of the ice. This would increase their measured concentration in the ice and decrease their measured concentration in the brine.

As demonstrated with the enrichment/depletion trends of MPs in bulk ice, MP’s enrichment/depletion patterns are attributed to i) polymer density, which confer a positive buoyancy (i.e., rising) for the low-density PP-MPs and a negative buoyancy (i.e., settling) for the high-density PET-MPs and to ii) size, which accelerates the enrichment/depletion of MPs (Figs. 4, and S5, and Supplementary Section 1). This is further demonstrated by comparing concentrations in ice and in the underlying liquid. Both small and large high-density PET-MPs were depleted in the ice, with the Large PET-MPs more depleted than the Small PET-MPs (C/C0 = 0.046 ± 0.038 and 0.580 ± 0.322, respectively) (Fig. 5). Conversely, both Large and Small low-density PP-MPs were enriched in the ice (C/C0 = 1.594 ± 0.707 and 1.714 ± 0.620, respectively). No significant differences were observed between Large and Small PP-MPs’ concentrations in ice. This could be due to the removal of the top 0.5 cm ice section from the analysis of particle distribution (due to its affinity for the air water interface, cf.: Methods) which introduced a higher variability in PP-MPs concentrations. PP-MPs’ enrichment was compensated by a depletion in the underlying liquid (C/C0 = 0.308 ± 0.308 and 0.682 ± 0.158 for large and small PP-MPs, respectively). For PET-MP, no significant changes in concentration in the liquid fraction were observed, relative to the initial concentration (C/C0 = 1.027 ± 0.051 and 0.976 ± 0.073, for Large and Small PET-MPs, respectively) since the MPs were depleted from a small volume of ice and transferred into a large volume of underlying water.

In addition to density, MPs’ size also determined the degree of enrichment/depletion in the brine fraction. Surprisingly, the concentrations of MPs in brine were not correlated to their concentrations in ice. Based on the ice morphology, particle size and rising/settling velocities, two effects could be at play. First, the speed of advancement of the freezing front relative to MPs’ settling/rising rate could determine whether MPs have a chance of being located in brine channels and how easily they are drained from the channels by centrifugation. Since PP-MPs rose quickly in the column (Supplementary Section 1), most particles likely entered the ice when it started to form. This would lead them to either be engulfed at the top of the ice and out of the brine channel or to be located at the top of the brine channels, where they would need to travel a longer and/or more tortuous path to be drained out by centrifugation. Second, the size of the particles relative to the brine channel width could explain different MPs depletion/enrichement ratios in brine and ice. Indeed, both small MPs are more depleted in the brine than larger ones (C/C0 = 0.811 ± 1.267 and 0.095 ± 0.023 for Large and Small PP-MPs, respectively 0.167 ± 0.151 and 0.088 ± 0.024 for Large and Small PET-MPs, respectively). This suggests that large MPs are physically trapped in brine channels and unable to settle or rise through them, while small MPs, which are on average smaller than all brine channels, can still move through them.

The colloidal contaminants had a distribution in ice and brine which was in between those of dissolved contaminants and the high-density PET-MPs. The extent of NPs depletion from ice (C/C0 = 0.242 ± 0.124) was not significantly different from salts (p = 0.0098) and was in between the values for Small and Large PET-MPs (Fig. 5). Nanosoot depletion from ice (C/C0 = 0.407 ± 0.021) was significantly less than for salts (p < 0.001) and, again, in between the values for Small and Large PET-MPs. In brine, both NPs and nanosoot were more enriched than any MPs but less enriched compared to salts (p = 0.009 and p < 0.001, for NPs and nanosoot relative to salts, respectively). Indeed, NPs were barely enriched with a C/C0 of 1.324 ± 0.334, while nanosoot was even slightly depleted, with a C/C0 of 0.876 ± 0.044. Finally, concentrations of NPs and nanosoot were relatively stable in the underlying water, again due to the same dilution effects observed for salts and PET-MPs (C/C0 of 0.994 ± 0.023 for NPs and 1.007 ± 0.015 for nanosoot).

Colloidal species’ distribution depends on their aggregation state
The behavior of colloids in sea ice is highly dynamic since freezing can promote colloidal aggregation, as has already been already observed in batch experiments46. The distribution of colloidal contaminants, which is between those of dissolved species and high-density particulate species, can be explained by the co-occurrence of dispersed and aggregated colloids in the artificial sea ice column. On one hand, Brownian motion allows dispersed colloids to diffuse away from the freezing front, and therefore, undergo freeze rejection47,48. On the other hand, colloids aggregate in seawater and these aggregates can be large enough for their transport to be dictated by the Stokes regime rather than Brownian motion49. Indeed, after 19 h aggregates had formed as evidenced by the 5-fold increase in the hydrodynamic diameters of NPs and nanosoot in all fractions, relative to their initial size (Fig. S6). In the case of NPs, the aggregated diameter was directly proportional to the salinity of the dispersion media, whereas nanosoot aggregates were largest in the liquid fraction, followed by brine and ice. Despite the presence of these aggregates, it is anticipated that many individual, dispersed NPs and nanosoot remained in suspension since these distributions of colloidal suspensions are overestimated by dynamic light scattering measurements when aggregates are present50. Microscopy analysis of thawed ice cores showed that nanosoot aggregates were significantly larger than NP aggregates (Fig. S7). This can explain the fact that the distribution of nanosoot between the ice, liquid and brine is more similar to that of high-density MPs than dissolved species (Fig. 6). Furthermore, the larger size of nanosoot aggregates could have led to their settling before dynamic light scattering measurements, which explains the smaller hydrodynamic diameters of nanosoot measured in the ice fraction (Fig. S6).

Species’ distribution between brine and ice is indicative of their incorporation process
To understand the incorporation mechanisms of contaminants, it is useful to specifically consider what happens at the freezing front by comparing the relative concentration of each species in brine and ice for each the replicate experiments separately (Fig. 6). A pattern clearly appears, showing that sea salts are most accumulated in the brine, followed by Rose Bengal and NPs. This highlights that dissolved species and the dispersed colloids are expelled from the ice and accumulated in brine. It also suggests that the decreasing enrichment in brine could be related to the species’ decrease in hydrodynamic diffusion coefficient as molecular weight increases (salt < Rose Bengal < NPs). Such trends have also been observed for per- and poly-fluoroalkyl substances (PFAS), that is, contaminants composed of fluorinated carbon chains of variable lengths. Indeed, short-chain PFAS were more enriched in the brine relative to the solid ice matrix (corresponding here to a high Cbrine/Cice), while long-chain PFAS were enriched in the ice rather than the brine (corresponding here to a low Cbrine/Cice)33. In fact, when modeling these results, there appeared to be a stronger decoupling between concentrations of PFAS and salts in bulk ice as the molecular weight of PFAS increased51. This decoupling was strongly correlated to brine salinity, which suggests that PFAS could also be undergoing salting out, thereby reducing its diffusion coefficient52. Finally, the Cbrine/Cice of MPs clearly showed that smaller MPs were more depleted from brine, presumably because they traveled through it without being physically trapped.

Suitability of lab-scale experiments to study incorporation of contaminants in sea ice
This lab-scale experiment allowed us to generate environmentally representative artificial sea ice by basal growth and to assess the incorporation process of a variety of model contaminants initially dispersed in water. Inevitably, laboratory studies simplify complex environmental processes. To assess how these results translate to the environment, the following section discusses the strengths and limitations of our experimental design.

With respect to the type of ice grown, the advantage of our experimental design is that, by being relatively small, it allows many replicates to be performed since the freezing time and the amount of model contaminants required are reduced. However, this smaller size does not enable the generation of underwater currents, which occur naturally under sea ice. Despite this, the artificial sea ice formed is similar to natural ice. One must also keep in mind that this experimental set-up generated columnar ice, which grows below the layer of frazil ice and, as such, does not interact with contaminants at the air/water interface. However, this columnar ice is most common in the Arctic, which receives more anthropogenic contaminants than the Antarctic.

With respect to the type of observations that can be made, this design does not enable the use of methods that avoid the loss of brine as the sea ice is removed (i.e., by inserting a box under the sea ice as it is pulled out53). Furthermore, the use of centrifugation may not drain the brine pockets that are not connected to the bottom of the ice core. Therefore, the brine content is underestimated and the ice salinity is overestimated, compared to methods which measure brine content in-situ44. Future experiments could incorporate in-situ temperature and salinity measurements to record the ice growth and brine content in real time. However, since microorganisms also thrive in the brine channels that are connected to the underlying water, the brine fraction studied here is the more biologically-active fraction of the sea ice. Finally, this experimental design does not enable in-situ measurement of the adsorption of contaminants onto the ice or of the aggregation state of colloids and particles. Therefore, processes such as particle and colloid adsorption by hydrophobic or van der Waals attraction, or entrapment of aggregates in the brine channels cannot be observed. Future experimental developments that enable in-situ quantification and observation of colloids and particles in ice would allow to better quantify them and assess their surface affinity for ice and their aggregation state in ice.

Environmental implications
Microalgae living in sea ice may be exposed to high loads of contaminants, adding to other stressors they must contend with (e.g., acidification and warming). Furthermore, the dynamics of contaminant incorporation into and transport with sea ice are changing with global warming, as young sea ice replaces multi-year ice. Therefore, we studied how contaminants present in seawater are incorporated in sea ice to assess exposure levels and understand new patterns of contaminant cycling. Specifically, this study investigated the processes occuring at the seawater/ice interface, which is most biologically active. Our study helps to predict a contaminant’s incoporation into sea ice by taking into consideration its size, density (in the case of particulate species), and aggregation state (in the case of colloids).

Our results suggest that organisms which inhabit the porosity of sea ice will be highly exposed to dissolved contaminants and dispersed colloidal contaminants. While low-density micrometric contaminants were enriched in ice overall, they may only be temporarily bioavailable since they can travel upward through brine channels, and therefore be incorporated at the top of the ice, which is less porous due to brine drainage. Since the properties of sea ice, such as the speed of advancement of the freezing front and the ice porosity, play important roles in contaminant incorporation, measurements of contaminant concentrations in natural sea ice should be accompanied with a description of the sea ice structure and history to understand their transport routes. This can help to understand whether contaminant incorporation occurs during basal ice growth or frazil ice growth.

The processes by which contaminants’ size and density drives their incorporation in sea ice, revealed in this study, are also applicable to natural species. This can help resolve long-standing questions about the incorporation of essential elements such as iron. Indeed, sea ice is an ideal environment for the aggregation of colloids and potentially also for the self-assembly of dissolved species into colloids or particles due to i) the local increase in salt and colloid concentrations in brine, and to ii) the growth of solid ice which can push colloids closer together thereby increasing the possibility of interaction. Therefore, the decoupling between “dissolved” (<0.4 μm) and “particulate” (>0.4 μm) iron observed in sea ice54, as well as the higher enrichement of organic matter <0.7 μm in brine relative to salts55, could be due to the overlooked assembly of macro-molecules and aggregation of colloidal species. In conclusion, the results and methods presented here not only help to understand the fate of contaminants, but more generally sea ice biogeochemistry in the context of climate change.

Methods
Model contaminants
Rose Bengal Sodium salt (CAS: 632-69-9, Sigma-Aldrich) was used as the model dissolved molecule, with an initial conentration of 6.7 ± 0.3 μmol L−1 in the water column. A model carbon-based contaminant, a soot nanoparticle grafted with carboxylic functional groups (hereafter called nanosoot), was used with an initial concentration of 32 mg L−1 (Aqua-Black 162® Tokai Carbon, Tokyo, Japan). The initial hydrodynamic diameter of the nanosoot was 117 ± 3 nm as determined by dynamic light scattering (DLS; Malvern Zeta-sizer). The zeta-potential was measured to be −55.26 ± 1.35 mV, as determined by electrophoretic mobility (Malvern Zeta-sizer).

Model NPs and MPs were composed of polymers that were homogenously enriched with trace-metals, and were quantified as a proxy for the plastic using inductively coupled plasma mass spectrometry (ICP-MS). This approach allowed for quantification of trace concentrations of plastics more easily while also avoiding risks of NPs and MPs contamination within our experimental system. The low metal incorporation did not significantly change the particle density, and there was no leaching of metal over time (as shown in previous studies for NPs56–59 and PET-MPs59, as well as in Supplementary Section 2 for PP-MPs). NPs were composed of a polyacrylonitrile core and a polystyrene (PS) shell with a final dopant concentration of 0.295 ± 0.009 w/w % palladium (Pd). The initial hydrodynamic diameter and zeta-potential of the NPs were 210 ± 19 nm and −52.83 ± 2.23 mV, respectively. High-density (1.39 g cm−3) MPs were composed of polyethylene terephthalate (PET) with 0.18 w/w % Indium (In) and low-density (0.91 g cm−3) MPs were made of polypropylene (PP) with 0.14 w/w % Yttrium (Y). MPs fragments of both PET and PP were produced by cryo-milling and sieving at different mesh sizes. Large MPs were retained between 63 and 125 μm sieves, and Small MPs passed through a 63 μm sieve. The mean area equivalent diameter of Small PET-MPs and PP-MPs were 35.6 ± 14.4 μm and 45.7 ± 20.7 μm, respectively. Large PET-MPs and PP-MPs had a mean area equivalent diameter of 90.0 ± 29.8 μm and 72.3 ± 37.2 μm, respectively.

Artificial sea ice production and processing
Artificial seawater (ASW) with a salinity of 31.28 ± 1.47 g L−1 and pH of 7.96 ± 0.16 was prepared following the method of Kester et al.60. Natural organic matter composed of humic acid salt (solid, Acros Organic, CAS: 1415-93-6) and sodium alginate (solid, Sigma Aldrich, CAS: 9005-38-3) was prepared using the methods described in Pradel et al.61 and added to ASW such that final concentrations were 0.7 mg L−1 and 7 mg L−1 respectively, to mimic conditions below sea ice and to improve the dispersion of particles and colloids.

Artificial sea ice was generated by a temperature gradient in a column under gentle agitation for 19 h (Fig. 2). The cylindrical columns were composed of polymethylpentene with a diameter of 6.55 cm and height of 44.0 cm. After being filled with 1142 ± 2 mL ASW, an aluminum cooling element of 5.9 cm diameter and 4 cm depth was inserted onto the top of the column, in full contact with the water surface, with a slightly rounded shape to allow air bubbles to escape. The cooling elements, cooled by circulation of cooling liquid from a cryothermostat bath, had a bottom of the temperature of −5.1 ± 0.1 °C. The experiments were conducted at an ambient temperature of 1.0 ± 0.6 °C, resulting in a temperature gradient over the column height (Fig. 2, panel a). The bottom of the sea ice column was fitted with a flexible PVC tube with an inner diameter of 3 mm which extended above the level of the column. This allowed the liquid to move up the tube as the ASW froze and expanded, thereby avoiding pressure increases in the system. The columns were set on an orbital shaker (KS15A, Huber Lab) at 50 rpm.

For each experimental condition, three columns with cooling elements were aligned in series (Fig. 2, panel b). Freezing lasted 19 h to ensure that the ice thickness attained a plateau which was typical for sea ice grown under constant temperature forcings (Fig. S8). At the end of the experiment, the ice had a mean depth of 3.32 ± 0.61 cm (Fig. 2, panel c). The ambient temperature of the experimental room was lowered to −2.5 °C and the bulk ice was lifted out of the column with a solid net attached to two nylon strings. Artificial sea ice cores were immediately centrifugated (Sigma 3-16KL) for 10 min at 40 g at −3.5 °C to drain the brine connected to the bottom of the bulk ice core. This temperature was selected since based on the ASW concentration and the temperature gradient, the brine salinity was estimated to be ~60 g L−1 (cf: Figure 12.1 of ref. 40) and 60 g L−1 ASW freezes at −3.5 °C62,63. Therefore, centrifugating above the freezing point of the brine and below the freezing point of pure ice, allows us to preserve the ice core structure. However, sea ice salinity and permeability is complex and nonlinear64, and there is inherent experimental variability in the temperatures. For example, measured temperatures were −2.3 °C at the top and −4.6 °C at the bottom in the centrifugation bucket. Therefore, strictly speaking, the brine obtained represents the brine with freezing points above the actual centrifugation temperature. The mass of centrifugated brine, centrifugated ice and underlying liquid were weighed after centrifugation.

To characterize the structure of the sea ice, several centrifugated ice cores were set aside at −20 °C and scanned by micro-X-ray-tomography (μCT)43. By draining the brine out of the ice prior to scanning, the porosity of the ice was more visible since the contrast between air and ice is higher than between liquid and ice. However, some brine was distinguishable from the ice. Therefore, a 3D rendition of the sea ice structure was created by segmenting into three different levels of absorption to observe ice, drained brine channels, and residual brine. μCT scans were performed on a subsample of 3 cm diameter of the ice core using a μCT 40 by SCANCO Medical AG (Brüttisellen, Switzerland). X-ray tube settings of 70 kVp and 8 W were used. For all samples, at a total exposure time of 600 ms was used with 1000 projections per slice (180° of rotation). The resolution was 11 μm. The images were reconstructed using a proprietary back-projection algorithm. The 16-bit images were filtered by a Gaussian 3D-filter with a size of 4^3 pixels and a standard deviation of 1.2 pixel. Segmentation of the air-ice threshold was 22 permille, of the ice-brine threshold 72 permille. Further visualization was done in ImageJ65.

Quantification of contaminant distribution in artificial sea ice
To evaluate species’ rates of incorporation into sea ice, we performed experiments with one species at a time and assessed their concentrations in thawed ice, liquid and brine at the end of the experiment relative to their initial concentration in the column (C/C0). To quantify the distribution of species at the freezing front, we also assessed the relative concentrations in brine and ice (Cbrine/Cice) in each column separately. Given the physicochemical differences of the model species, different methods were used to initially disperse, recover and quantify them.

Dissolved and colloidal species (Rose Bengal, NPs, and nanosoot) were dispersed into a master batch (i.e., a large volume homogenized by mixing) which was then equally distributed between the three columns. Particulate species were either positively buoyant (PP-MPs) or negatively buoyant (PET-MPs) and consequently could not be homogenously distributed within the columns. To ensure that sufficient quantities of each MPs type would be interacting with the freezing front we either added PP-MPs to the bottom of the column with 100 mL of ASW before completing with the rest of the ASW volume, or we added the ASW and then added the PET-MPs to the top of the column with the last 100 mL. When inserting PP-MPs into the liquid prior to freezing, many PP-MPs remained at air-liquid interface due to their lightweight and hydrophobicity. This resulted in PP-MPs at the top of the sea ice cores which did not interact with the advancing ice front. Therefore, the top 0.5 cm of the ice core was removed after centrifugation and excluded from the evaluation PP-MPs distribution in sea ice but still considered when establishing the PP-MPs recovery rate across the experiment.

Species were recovered and their concentrations measured in each fraction (i.e., brine, ice, liquid) using different analytical approaches. Seawater salts were quantified by measuring electrical conductivity (FiveEasy conductivity meter, Mettler Toledo) and converting electrical conductivity to salinity, using a calibration curve. Rose Bengal and nanosoot were quantified by absorbance spectroscopy (at λ = 548 nm and λ = 400–600 nm, respectively, Cary 60 UV–vis, Agilent). The analytical workflow for plastic contaminants consisted of concentration, digestion, and quantification of their metal content. MPs were recovered on filters (Macherey-Nagel 615, with a cut-off of 4 to 12 µm). NPs were pre-concentrated by flocculation using poly(acrylamide co-acrylic acid) (Sigma-Aldrich) and aluminum nitrate nonahydrate (Sigma-Aldrich). The protocol had a recovery rate of 95 ± 5%, and is further described in Supplementary Section 3. Filters with MPs and NP flocs underwent microwave acid digestion in an ultraWAVE digestor (Milestone Srl, Italy) with either 2.3 mL or 5 mL distilled HNO3 65% (puriss. Sigma-Aldrich), respectively. The temperature and pressure increased from ambient to 220 °C and 160 bar over 30 min and were then maintained at 220 °C and 160 for 30 min. Digestates were then diluted and quantified by ICP-MS (Agilent 7900). For each element measured (106Pd, 115In and 89Y) a calibration curve and quality controls were performed daily. Internal standards of Scandium, Yttrium and Rhodium (10 µg/L) were added to the Pd and In containing samples, while only Sc and Rh (10 µg/L) were used for Y-containing samples. Internal standards were continuously monitored throughout the measurements and results were corrected to account for instrumental drift.

The mass balance for most species was high (ranging from 95.75% to 101.97%), which gave us confidence that the distributions between the different fractions could be used to track the fate of the contaminants in the sea ice column (Table S3). For large PET-MPs, small PP-MPs and large PP-MPs the recoveries were lower (73.60 to 89.86%). For PP-MPs, this can be explained by their hydrophobicity, which caused them to adsorb onto surfaces of the column and the air/water interface. The addition of model contaminants did not significantly affect the mass, pH, salinity or structure of the ice (Table S4 and Figs. S9 and S10).

Assessment of the aggregation state of colloids
Following freezing, the aggregation state of the colloids (nanosoot and NPs) was assessed by measuring the size by DLS and zeta-potential by electrophoretic measurement using the Smoluchowski model (Malvern Zeta-sizer). The larger aggregates were measured by light microscopy using a ZEISS Axiolab microscope at a 10x magnification, an Cam Erc 5s camera and the ZEN lite software. Long exposure times (300 to 1000 ms) were used to increase the visibility of small aggregates. The aggregates were characterized by analyzing the images with the NanoDefine particle sizer plugin66,67 of the ImageJ software65.

Data analysis
Data was plotted either using R Statistical Software68 or Prism software. To compare the concentrations of model contaminants in different fractions, two approaches were employed. First a F-Test for equality of two variances was performed. If variances were unequal at the 0.05 significance level, then a t-test was performed for unequal variances. Otherwise a t-test with equal variances was performed. To quantify the speed of advancement of the ice front, a power equation was fitted by linear regression to the data.

Peer review rounds
[{"round": 1, "reviewer_comments": {"reviewer_1": "The study entitled \" Understanding Contaminant Incorporation in Sea Ice: Impact of Size and Density\" investigates the ice-water distribution patterns of model contaminants during freezing processes. The approach of separating ice and brine based on different freezing points demonstrates a certain degree of innovation. The microtomography has been used to characterize the structure of ice and the distribution of brine channels within it. The paper specifically discusses the influence of size and density on the distribution characteristics of model organic compounds in ice, brine, and water. However, several issues need to be addressed to improve the manuscript. It is recommended to revise and improve the paper from the following aspects:\n1. In lines 389-390, the ice core was extracted using a solid net. The solid net is made of plastic material. Could it release or adsorb microplastics and nanoplastics, thereby affecting the results?\n2. Is this ice core the entirety of the ice that had been frozen for 19 hours? Additionally, a detailed explanation is required regarding how the solid net separated the ice from the water in the mixture of ice and water. During the ice-water separation process, will the ice melt? Do the materials used in the simulation experiment affect the quantitative results? There should be a control experiment.\n3. When considering the separation of brine and pure ice, differentiating them based on different freezing points is an interesting concept; however, several issues arise. The selection of the freezing point is one such issue. The salinity of the branches in different brine channels may not be entirely uniform. Choosing -3.5°C as the freezing point merely indicates that ice with a freezing point above -3.5°C will melt and detach from the ice core. Therefore, strictly speaking, the concentration of the brine obtained represents the concentration of only some brine channels, specifically those with freezing points above -3.5°C. Furthermore, during the separation process within brine channels, microplastics or nanoplastics may not be completely expelled and may partially adhere to the interstitial spaces between ice crystals. Hence, the brine concentration discussed in this context can only be considered as the 'bulk brine' concentration rather than the concentration of all brine. Moreover, this aspect requires supporting citations from relevant literature to enhance credibility, such as references that justify the choice of the freezing point.\n4. In the supplementary materials, the author clarifies the use of microtomography for analyzing and calculating the widths of brine channels, which ranged from 50 to 240 μm. The selected microplastics in the study exhibit varying particle sizes, potentially leading to differing final results. Some microplastics may not be effectively separated from the brine channels, and there may exist brine channels with even smaller widths. These factors have a significant impact on the separation of brine and ice, as well as the ultimate outcomes of the study.\n5. What is the significance of separating brine from ice, considering that in nature, brine and ice exist as an integrated entity? The environmental implications of centrifuging brine are unclear, and the separated brine only represents a portion of the total.\n6. Figure 4 is missing. The text and figures do not match.\n7. Lines 230-232: Microplastics of small particle sizes may not all be able to pass through brine channels, and they may adhere to the edges of brine channels and ice. Additionally, aggregation of small-sized microplastics due to unstable freezing, which leads to an increase in relative particle size, may also trap them within brine channels.\n8. When discussing the occurrence concentrations of different types of microplastics, particle size and density are crucial factors to consider. Additionally, zeta potential and contact angle are also key characteristics for describing the properties of plastics. A more comprehensive discussion of the mechanisms should be included when analyzing the distribution of microplastics in ice, brine, and water.\n9. In Figure 3, the data related to polypropylene (PP) exhibit significant errors. For instance, the data points in brine deviate substantially from the main data cluster, suggesting the possibility of man-made error rather than systematic error. It is advisable to repeat the experiments to eliminate the large man-made error.", "reviewer_2": "Review of: Understanding Contaminant Incorporation in Sea Ice: Impact of Size and Density, by Pradel et al.\nMax Thomas\nPradel et al. present experimental results from a new sea-ice growth facility. A suite of contaminants with a range of physico-chemical properties were added to the experimental system, and their concentrations were measured in the underlying liquid, brine centrifuged from sea-ice samples, and in the sea ice remaining after centrifugation. Pradel et al. then interpret the concentration measurements based on the range of properties in the contaminants.\nThe experiments and measurements look competently done and are presented well. The choice of contaminants seems very well thought out and allows for a detailed and novel analysis of the mechanisms behind the behaviour of these chemicals. Sections 2.2 – 2.4 are particular highlights which have potential to be highly influential on future work. The work is original, and provides important, incremental knowledge gains for the sea-ice contaminant and process modelling community.\nThere is some work to be done on improving clarity, and to explicitly state how (unavoidable) biases and limitations in the measurements impact the conclusions. I was also confused by the lack of any bulk sea ice values being presented (the only major comment, which would require some more data analysis) which would help comparison with other work and might help simplify and strengthen conclusions.\nI recommend the work for publication after the revisions listed below, which are fairly minor.\nMajor comments:\n1) Line 22 – I don’t think this statement is justified: ‘While… particulate contaminants were enriched [or depleted] in sea ice’. Normally, ‘sea ice’ would here refer to the bulk value, which would be the concentration measured in melted ice and brine together. Bulk values aren’t presented. I presume figure 3 is evidence to support this statement but I don’t understand how the results link back to the statement, as a sea ice measurement is not presented. I suggest calculating ‘sea ice’ using the measured concentrations of a contaminant in ice and brine and the measured mass or volume of ice and brine. Possibly evidence is there already for this statement, and if so that needs to be made clear.\nMinor comments:\n1) Line 12 – Sea ice forms near human activity too (Canadian towns, Longyearbyen) so please remove ‘despite being far from human activity’. You may want to add that it transports contaminants as this is important and mentioned prominently in the intro.\n2) Line 14 – Please make clear what aspects are ‘novel’. The suite of chemicals is novel, but the cold plate, growth in tubes, artificial salt solution are not novel. If it is some combination of the design choices that is novel for some specific novel need, that should be made clear and justified. The methods don’t need to be novel for the work to be novel and worthwhile.\n3) Line 18 – What precisely does ‘difficult’ mean? Are you referring to contamination issues?\n4) Line 20 – Are kDa and um related to eachother somehow? I don’t see how a range of values from x to y is useful when x and y have different units.\n5) Line 58 – The authors will need to be more clear about what ‘brine rejection’ is referring to. Generally, ‘brine rejection’ refers to brine moving from the sea ice to the ocean below, which desalinates the sea ice. This happens after sea ice has grown and incorporated everything. There is insignificant segregation at the advancing interface, so everything that was in the water is incorporated into the growing sea ice, then rejected (Notz & Worster, 2009, https://doi.org/10.1029/2008JC004885). What the authors are describing is salt and other dissolved species being excluded from the ice matrix such that they are partitioned to the brine, but remain within the sea ice.\n6) Line 59 – The authors should be careful to use ‘ice’ to mean the frozen, ice matrix of sea ice, and ‘sea ice’ or ‘bulk sea ice’ to mean the collection of ice and brine. Here, ‘bulk ice’ is used, which is confusing.\n7) Line 62 – (see minor comment 4)\n8) Line 77 – ‘Particulate species with low densities are expected to be enriched…’ Enriched relative to what? Where does this hypothesis come from? The other hypotheses follow from the introduction, but this one seems unconnected to the previous introduction material.\n9) Line 83 – This is probably the place to address minor comment 2.\n10) Line 90 – Defining ice and brine earlier could solve previous comments.\n11) Line 92 – ‘Trapped’ probably isn’t the right word here.\n12) Line 92 – ‘Collectively this’ is imprecise (and should be ‘these’ I think). I would suggest replacing this with ‘Our results’ or ‘These knowledge gains’, if that’s what is meant.\n13) ‘4’ is repeated in the figure 1 caption and ‘2’ is missing. Also ‘column’ should be ‘a column’ or ‘columns’. This figure is very clear and informative.\n14) Line 108 – ‘vertical temperature gradient’.\n15) Line 114 – ‘average’ to ‘mean’ (I presume).\n16) Line 129 – ‘brine salinity approximately doubled’. The authors should calculate and report to temperature in Vancoppenolle et al. (2019). This comparison will help the authors think about how representative their centrifuged brine is of the in situ brine.\n17) Lines 133-135 – It is great that the authors have thought deeply about this aspect of the experiment and have noted it here. It then needs to feature in the discussion (see also point 2.4).\n18) Lines 136-143 – These values are reassuring, and suggest the artificial sea ice is a reasonable approximation of the real world. I would note that the sea ice in this study is very young (<5 cm), whereas real world sea ice is considered ‘young’ while less than 30 cm – I would expect 30 cm sea ice to have lower salinity and solid fraction. Also, the values reported by the authors here will be biased by brine loss during sampling (so before centrifuging). This bias is practically unavoidable. These caveats should be noted here. The authors could compare to Notz & Worster (2008) which present in situ measurements of bulk salinity and solid fraction (https://doi.org/10.1029/2007JC004333), and have measurements in <5 cm sea ice. For comparable thicknesses the authors will see the in situ measurements will have a higher bulk salinity and solid fraction.\n19) Line 155 – Why did the freezing element have a curved shape?\n20) Figure 2b – Could the authors please annotate the image? I had to guess what the lenticular structures were. Also, what is ‘black’ in the image?\n21) Line 171 – ‘if’ to ‘is’\n22) Line 229 – Needs an ‘and’ between the reported values.\n23) Line 255 – ‘already’ to ‘already been’\n24) Section 2.4 – As a minimum, I would like to see a line here reminding the reader that Cice in this study includes brine in isolated pockets. It would also be very useful to get the authors take on how the inclusion of isolate brine within Cice impacts their conclusions. I found it difficult to understand how, for example, the exposure of microorganisms to contaminants is impacted by having this imperfect separation of phases in the measurement. The authors should also note bias from brine loss upon sampling and speculate as to impacts on conclusions. Possibly these caveats could form a short paragraph, as this paragraph is already quite complex and well put together.\n25) Figure 5 is excellent.\n26) Line 323 – ‘pressure of crystal growth’? Do the authors mean actual pressure (as in Pa), and if not, what?\n27) Glossary: The statements about this sea ice containing no gas should be softened. Gas is often found in young sea ice, and there could be some here, unless the authors have some evidence to the contrary.\n28) At the end of the paper I realised I don’t understand why the low density MP were so depleted in the liquid. Scanning back through I couldn’t find the answer. Could the authors please explain this, or make the explanation more prominent.\n29) I would also suggest saying ‘: The impact of…’ in the title as it’s plainer English.\n(Just a note to say I thought the section titles were very helpful. The declarative statements made the paper easy to follow.)", "reviewer_3": "Understanding Contaminant Incorporation in Sea Ice: Impact of Size and Density\nPradel et al\nThis manuscript details the experimental set-up and application of an artificial sea ice chamber system that allows the intimate study of solutes and particles, present in sea water, and their interaction with freshly forming ice. The authors are to be applauded for their relatively simple but elegant experimental set up. To my knowledge this small-scale system with multiple plastic polymer columns, an air-side cooling system (representing real Polar conditions over surface seawater) combined with a pragmatic pressure release tube is novel. Given the chemical and physical measurements of the fresh ice plug (with CT imaging) then similarities to initial ice in the Arctic is convincing. Hence this column system does provide a unique system to study the uptake, release and fate of both dissolved and particulate contaminants in ice (e.g. persistent organic chemicals, plastic particles, POC/DOC, heavy metals etc). The authors then demonstrate the utility of the apparatus by examining the fate of selected solutes, colloids and plastic particles (varying density) during a freeze experiment. Enrichment/depletion of these species relative to their occurrence in seawater is then reported with comparisons, where possible, to field and laboratory studies. The study is at the standard for publication however there are several issues that the authors need to comment on or address before publication could proceed.\n1) The columns and their seawater contents were agitated by being placed on an orbital shaker during the freeze period. It is not entirely clear why this was undertaken but does make sense in that particulate matter is likely to remain suspended in the water column, rather than deposited to the vessel floor (for the higher density particles). This agitation however does not represent a sub-ice seawater current though. This could be achieved using a small submerged pump or similar, which is common in larger sea ice chambers, which hence do resemble the surface ocean with this regard. On this note there doesn’t appear to be incorporation of sensors to measure physical parameters of the seawater itself. Most notably the temperature/salinity profile down the water column should be essential. For example, the expulsion of brine with ice aging or the release of low salinity melt water during ice melt, may result in stratified water layers which will affect chemical/material flows and yet are not easily distinguished in the current system.\n2) The freezing process using the aluminium cooling vessel will interfere with air side transfer of particles and vapour/gaseous species. While the focus is clearly on studying the ice itself and the ice-seawater interface, this is nonetheless a significant oversight particularly for studying the transfer of material/chemicals across the ice-air interface. Importantly, the introduction of material to the surface of the ice to simulate ice-rafted snow for example cannot be undertaken with the current set up.\n3) In the Introduction the authors cite those studies that have observed microplastics in Polar sea ice at concentrations orders of magnitude higher than concentrations present in the underlying sea water. This is one of the drivers behind experimental simulations of the seawater/sea ice system like this study. However, in the experimental work for the microplastics the authors discarded the uppermost 0.5cm ice layer on the basis that lower density particles had accumulated in the water-air interface and hence had become artificially rafted onto the ice-surface. Surely, this is the very process which will be occurring to some extent for positively-buoyant plastic particles in the real marine environment! Does this ice-rafting of some types of microplastics occur during early formation of grease/frazil ice?? This is a key question which hopefully this new experiment apparatus could investigate, but which the authors have viewed as an artifact!!\nP1, line 28 contaminantation\nP2, line 72 ….in [a] controlled environment…\nP2, line 94 ….wil be exposed to."}, "author_response": "Answer to reviewers for manuscript NCOMMS-24-86817-T\nReviewer #1 (Remarks to the Author):\nThe study entitled \" Understanding Contaminant Incorporation in Sea Ice: Impact of Size and\nDensity\" investigates the ice-water distribution patterns of model contaminants during freezing\nprocesses. The approach of separating ice and brine based on different freezing points\ndemonstrates a certain degree of innovation. The microtomography has been used to characterize\nthe structure of ice and the distribution of brine channels within it. The paper specifically discusses\nthe influence of size and density on the distribution characteristics of model organic compounds in\nice, brine, and water. However, several issues need to be addressed to improve the manuscript. It\nis recommended to revise and improve the paper from the following aspects:\nWe thank the reviewer for their time to make detailed comments and suggestions to improve our\nmanuscript, which will aid future readers in understanding the rationale of our experimental design\nand important aspects relating to particle physiochemical dynamics.\n1. In lines 389-390, the ice core was extracted using a solid net. The solid net is made of plastic\nmaterial. Could it release or adsorb microplastics and nanoplastics, thereby affecting the results?\nWhile the plastic net could potentially release microplastics (MPs) and nanoplastics (NPs), an\nimportant distinction is that we used model MP and NPs in this experiment which are doped with\ntrace metals. Subsequently, by quantifying the trace metals as a proxy for these model particles\npotential contamination from non-target MP and NP can be excluded. This is also true for potential\nrelease of MPs and NPs from the column, which is also made of plastic.\nThe plastic net (and column) could adsorb MPs and NPs. However, the loss of particles throughout\nthe experimental workflow was minimal, as discussed lines 568 to 573 and shown in\nSupplementary Table 3. For salts and most model contaminants (Rose bengal, NPs, Nanosoot\nand PET) the mass balances is very close to 100%. For Small-PP and Large-PP mass balances\nare lower and more variable : 73.60 +/- 20.38 % and 88.49 +/- 35.28 % respectively. We have\nobserved that some of these particles are retained at the air/water interface and on the column\nand suspect that this is due to their hydrophobicity. Therefore, we suspect that most of the loss\ndoes not occur at the seawater/ice interface, and that it has minimal effect on our conclusions\nobtained from Figures 4, 5 and 6.\n2. Is this ice core the entirety of the ice that had been frozen for 19 hours?\nYes, the entire ice core which has formed over 19 hours has been recovered. We have now clarified\nthis point in the text, see line 125-126.\nAdditionally, a detailed explanation is required regarding how the solid net separated the ice from\nthe water in the mixture of ice and water.\nWe added an explanation on how the ice was separated in both the Results and Materials and\nMethods sections.\nto two strings and were centrifugated (Sigma 3-16KL) to drain the brine connected to\nthe bottom of the ice core.\nLines 494 - 496: The ambient temperature of the experimental room was lowered to -\n2.5 oC and the ice was lifted out of the column with a solid net attached to two nylon\nstrings.\nDuring the ice-water separation process, will the ice melt?\nIt takes less than 2 minutes to pull the ice out of each column and put each ice core in a styrofoam\ncooler at -3.5 oC before centrifugation. Therefore, we do not expect the ice to melt to an extent\nwhich would alter our results under this time frame.\nDo the materials used in the simulation experiment affect the quantitative results?\nWe had conducted experiments without model contaminants to assess the variability of the ice\nproduced. When comparing the mass, salinity and pH of ice and brine for contaminant-laden\nconditions, we did not observe systematic differences with the control.\nFurthermore, to assess whether different contaminants generated different ice structures, micro-\nCT scans were made for different experimental conditions. These included nanosoot (Fig S4.a),\nnanoplastic (Fig S4. b), large PP (Fig S4.c) and large PET (Fig. 3b). As discussed in Lines 177-\n189, the ice cores displayed similar structural features. Collectively, this suggests that the\ncontaminants do not modify the structure of the ice formed under our experimental conditions. This\nwas expected considering the concentration of contaminants was on the order of mg/L while the\nconcentration of artificial sea salts was on the order of g/L.\nWe have now added the controls (i.e., contaminant-free variants) to the supplementary data and\nadded this to the discussion to the Materials and Methods section which presents the production\nof artificial sea ice.\nLines 573 – 574 : The addition of model contaminants did not significantly affect the mass,\npH, salinity or structure of the ice (Table S4 and Fig. S9 and S10).\n3. When considering the separation of brine and pure ice, differentiating them based on different\nfreezing points is an interesting concept; however, several issues arise. The selection of the\nfreezing point is one such issue. The salinity of the branches in different brine channels may not\nbe entirely uniform. Choosing -3.5°C as the freezing point merely indicates that ice with a freezing\npoint above -3.5°C will melt and detach from the ice core. Therefore, strictly speaking, the\nconcentration of the brine obtained represents the concentration of only some brine channels,\nspecifically those with freezing points above -3.5°C. Furthermore, during the separation process\nwithin brine channels, microplastics or nanoplastics may not be completely expelled and may\npartially adhere to the interstitial spaces between ice crystals. Hence, the brine concentration\ndiscussed in this context can only be considered as the 'bulk brine' concentration rather than the\nconcentration of all brine. Moreover, this aspect requires supporting citations from relevant\nliterature to enhance credibility, such as references that justify the choice of the freezing point.\nwhen designing the experiments and interpreting the results. Here, the objective was to separate\nbrine and ice to analyse the distribution of contaminants within the sea ice porosity. The separation\nof brine from the ice core was done at a temperature that minimized changes in the brine content\nof the artificial sea ice. Considering the salinity of our artificial seawater (31.28  1.47 g L-1) and\nthe temperature of the freezing element (-5.1  0.1 oC), brine with a maximum salinity of 62 g L-1\nwas expected to be formed (cf: Figure 12.1 of Thomas, D. N. & Dieckmann, G. S. (2010)). We then\ncalculated the freezing point of seawater at 62 g kg-1, with a sea pressure p (defined as the\nAbsolute Pressure P less the Absolute Pressure of one standard atmosphere) of 0, and an air\nsaturation of 1, using the Gibbs Seawater (GSW) Oceanographic Toolbox. This results in freezing\ntemperature of -3.52 oC. Therefore, we set the temperature of the centrifuge at the freezing\ntemperature to ensure that the brine would not freeze and the ice would not thaw. Please note that\nthis temperature is within the validity domain of the GSW equations, as explained in Vancoppenolle\net al. (2019). We have now explained this reasoning, provided references and discussed limitations\nin the main text :\nLines 498 to 506: This temperature was selected since based on the ASW concentration\nand the temperature gradient, the brine salinity was estimated to be approximately 60 g L-\n1 (cf: Figure 12.1 of ref 39) and 60 g L-1 ASW freezes at -3.5 oC63,64. Therefore,\ncentrifugating above the freezing point of the brine and below the freezing point of pure\nice, allows us to preserve the ice core structure. However, sea ice salinity and permeability\nis complex and nonlinear65, and there is inherent experimental variability in the\ntemperatures. For example, measured temperatures were -2.3 oC at the top and -4.6 oC\nat the bottom in the centrifugation bucket. Therefore, strictly speaking, the brine obtained\nrepresents the brine with freezing points above the actual centrifugation temperature.\nWe agree that that brine concentrations only represent concentrations in the brine that could be\ndrained out and not concentrations in residual brine pockets. We have mentioned this experimental\nlimitation throughout the manuscript:\nLines 165 – 167 : This can be attributed again to some brine loss, as well as some residual\nbrine pockets within the ice that were not connected to the bottom of the ice, and therefore\nnot drained by centrifugation.\nLines 261 – 264: Note that, as for the salts, it is also possible that some Rose Bengal in\nbrine pockets was not centrifugated out of the ice. This would increase their measured\nconcentration in the ice and decrease their measured concentration in the brine.\nWe have also discussed our experimental limitations in section 2.5 entitled “Suitability of lab-scale\nexperiments to study incorporation of contaminants in sea ice”\nLines 387 – 392: With respect to the type observations that can be made, this design does\nnot enable the use of methods that avoid the losss of brine as it the seanice is removed\n(i.e.: by inserting a box under the sea ice as it is pulled out54). Furthermore, the use of\ncentrifugation may not drain the brine pockets that are not connected to the bottom of the\nice core. Therefore, the brine content is underestimated and the ice salinity is\noverestimated, compared to methods which measure brine content in-situ43.\n4. In the supplementary materials, the author clarifies the use of microtomography for analyzing\nand calculating the widths of brine channels, which ranged from 50 to 240 μm. The selected\nmicroplastics in the study exhibit varying particle sizes, potentially leading to differing final results.\nSome microplastics may not be effectively separated from the brine channels, and there may exist\n3\nbrine channels with even smaller widths. These factors have a significant impact on the separation\nof brine and ice, as well as the ultimate outcomes of the study.\nWe agree that MPs may be trapped in brine channels that are narrower than the MP. This is one\nhypothesize that we provide for the reduced concentration of small MPs in brine compared to large\nMPs in lines 294 to 300.\n5. What is the significance of separating brine from ice, considering that in nature, brine and ice\nexist as an integrated entity? The environmental implications of centrifuging brine are unclear, and\nthe separated brine only represents a portion of the total.\nIn this study, we wanted to assess the distribution of contaminants across multiple compartments\nin order to i) quantify distribution of contaminants between ice and brine and ii) to assess the\nmechanisms that drive contaminant fate across the system. Therefore, i) the brine and ice were\nseparated to analyze the distribution of contaminants within the sea ice. ii) The ratio of\nconcentrations between brine and ice reflects what happens at the freezing front and allows us to\nunderstand the incorporation mechanisms of contaminants. Also, the brine fraction of sea ice is\nwhere sympagic organisms thrive due to the availability of nutrients, the space and (sometimes)\nincoming sunlight. Consequently, the partitioning of contaminants between the brine and ice\ncompartments are an important consideration when considering exposure. We have explained this\nin the manuscript:\nLines 238 - 245: To understand the mechanisms of contaminant incorporation in sea ice,\nwe assessed the distribution of contaminants across the different sea ice phases. To do\nso, brine and ice were separated to analyse the distribution of contaminants within the sea\nice. The ratio of concentrations between brine and ice was measured as it reflects what\nhappens at the freezing front and allows us to understand contaminants’ incorporation\nmechanisms. Also, the brine fraction of sea ice is where sympagic organisms thrive due to\nthe availability of nutrients, the space and (sometimes) incoming sunlight. Consequently,\nthe partitioning of contaminants between brine and ice are an important consideration\nwhen assessing exposure.\n6. Figure 4 is missing. The text and figures do not match.\nWe thank the reviewer for catching this mistake. Figure 4 was accidently labelled Figure 3, which\nhas now been amended in the revised version of the manuscript.\n7. Lines 230-232: Microplastics of small particle sizes may not all be able to pass through brine\nchannels, and they may adhere to the edges of brine channels and ice. Additionally, aggregation\nof small-sized microplastics due to unstable freezing, which leads to an increase in relative particle\nsize, may also trap them within brine channels.\nWe agree that MPs may be imperfectly drained out of the sea ice due to the brine channels\ntortuosity. We have discussed this as an explicit limitation in section 2.5.\nLines 395 – 401: Finally, this experimental design does not enable in-situ measurement of\nthe adsorption of contaminants onto the ice or the aggregation state of colloids and\nparticles. Therefore, processes such as particle and colloid adsorption by hydrophobic or\nvan der Waals attraction, or entrapment of aggregates in the brine channels cannot be\nobserved. Future experimental developments that enable in-situ quantification and\n4\nobservation of colloids and particles in ice would allow to better quantify them and assess\ntheir surface affinity for ice and their aggregation state in-situ.\n8. When discussing the occurrence concentrations of different types of microplastics, particle size\nand density are crucial factors to consider. Additionally, zeta potential and contact angle are also\nkey characteristics for describing the properties of plastics. A more comprehensive discussion of\nthe mechanisms should be included when analyzing the distribution of microplastics in ice, brine,\nand water.\nWhile our work demonstrates the impact of contaminants’ size and density in their incorporation,\nthe role of contaminant adsorption onto the surface of the ice could not be studied due to the\nmethods used. However, it is not a confounding factor to interpret the effect of size or density, since\nthe smallest species are all hydrophilic (i.e., salts and rose bengal), and for the largest species\n(MPs), both hydrophilic (PET) and hydrophobic particles were used (PP)( see: quantification of\ncontact angle by Schefer et al.4). For each of these polymers, the hydrophobicity does not change\nwith size and the effect of size is dominant, as it controls the sinking/rising speeds (cf: Figures 4,\n5, 6 and Supplementary Figure 5). Furthermore, at this ionic strength ( 600 mmol/L) the effect of\nelectrostatic repulsion is nearly absent for all particles, regardless of their initial surface charge.\nIndeed, the zeta-potential, shown in Figure S6, is higher than -15 mV in brine and liquid.\nWe discuss the inherent limitations in our experimental design in Lines 395 to 401 (see previous\nresponse).\n9. In Figure 3, the data related to polypropylene (PP) exhibit significant errors. For instance, the\ndata points in brine deviate substantially from the main data cluster, suggesting the possibility of\nman-made error rather than systematic error. It is advisable to repeat the experiments to eliminate\nthe large man-made error.\nExperiments were performed by the same operator, and clear records were taken of the\nexperimental and analytical conditions. This leads us to believe there are no fundamental errors\nwhen executing the experiments for PP compared to other contaminants. Rather, we believe that\nthe deviation of the PP-MP represents the inherently more variable behavior of this material. As\nmentioned in lines 275 to 277, the removal of the top 0.5 cm ice section from the analysis of particle\ndistribution (due to its affinity for the air water interface) may have contributed to a higher variability\nin PP-MPs concentrations.\nReviewer #2 (Remarks to the Author):\nReview of: Understanding Contaminant Incorporation in Sea Ice: Impact of Size and Density, by\nPradel et al.\nMax Thomas\nPradel et al. present experimental results from a new sea-ice growth facility. A suite of contaminants\nwith a range of physico-chemical properties were added to the experimental system, and their\nconcentrations were measured in the underlying liquid, brine centrifuged from sea-ice samples,\nand in the sea ice remaining after centrifugation. Pradel et al. then interpret the concentration\nmeasurements based on the range of properties in the contaminants.\nof contaminants seems very well thought out and allows for a detailed and novel analysis of the\nmechanisms behind the behaviour of these chemicals. Sections 2.2 – 2.4 are particular highlights\nwhich have potential to be highly influential on future work. The work is original, and provides\nimportant, incremental knowledge gains for the sea-ice contaminant and process modelling\ncommunity.\nThere is some work to be done on improving clarity, and to explicitly state how (unavoidable) biases\nand limitations in the measurements impact the conclusions. I was also confused by the lack of\nany bulk sea ice values being presented (the only major comment, which would require some more\ndata analysis) which would help comparison with other work and might help simplify and strengthen\nconclusions.\nI recommend the work for publication after the revisions listed below, which are fairly minor.\nWe thank the reviewer for his overall positive comments, especially for the recognition both of our\nnovel experimental design and selection of the contaminant suite which we chose to study. We are\nspecifically grateful that he finds the sections 2.2 – 2.4 to be of high interest and help to steer future\nresearch objectives, and indeed we are hopeful that through our own efforts or those of other\nscientists working in this field further advancements can be made with respect to contaminant fate\nin sea ice. Finally, it is notable that the reviewer has chosen to identify himself in the review (thank\nyou!), which is not often done but helps us to better place the comments and suggestions we\nreceived into better context. We are confident that the revisions suggested have allowed us to\nimprove our manuscript and to have even greater impact for future readers.\nMajor comments:\n1) Line 22 – I don’t think this statement is justified: ‘While… particulate contaminants were enriched\n[or depleted] in sea ice’. Normally, ‘sea ice’ would here refer to the bulk value, which would be the\nconcentration measured in melted ice and brine together. Bulk values aren’t presented. I presume\nfigure 3 is evidence to support this statement but I don’t understand how the results link back to\nthe statement, as a sea ice measurement is not presented. I suggest calculating ‘sea ice’ using the\nmeasured concentrations of a contaminant in ice and brine and the measured mass or volume of\nice and brine. Possibly evidence is there already for this statement, and if so that needs to be made\nclear.\nWe have added the concentration of each contaminant in the bulk ice (ice + brine) to support this\nstatement in Figure 4 and Supplementary Figure 5 along with the corresponding discussion.\nLines 200 - 232: First, the incorporation of each contaminant was assessed by comparing\ntheir concentration (C) in each the bulk ice (C which corresponds to brine and ice)\nbulk ice\nnormalized by their initial concentration in the column before freezing (C ) (Fig. 4). As\n0\nmentioned above, the ice bulk salinity was reduced to 0.44  0.07 of the initial salinity. Most\nother contaminants are depleted from the bulk ice, albeit in varying concentrations. The\nmodel dissolved contaminant, Rose Bengal, and the model colloidal contaminant, NP, were\ndepleted in statistically similar proportions as sea salts (with C /C = 0.43  0.01 and\nbulk ice 0\n0.38  0.09, respectively). This was followed by the other colloidal contaminant, Nanosoot,\nwhich was depleted to 0.48  0.01 of its initial concentration.\nlow-density MPs of two different size-classes resulting in four different MPs. High-density\nMPs were composed of polyethylene terephthalate (PET-MPs; 1.39 g cm-3) and low-\ndensity MPs composed of polypropylene (PP-MPs; 0.91 g cm-3). All MPs were fragments\nproduced by cryo-milling and sieving, resulting in area equivalent diameter of 35.6  14.4\nm and 45.7  20.7 m for small PET-MPs and PP-MPs, and 90.0  29.8 m and 72.3 \n37.2 m for large PET-MPs and PP-MPs, respectively. When inserting PP-MPs, many\nparticles remained at air-liquid interface due to their light weight and hydrophobicity. These\nPP-MPs did not interact with the advancing ice front as would be the case during bottom\nfreezing in the environment. Therefore, this fraction of PP-MPs was removed from the\nevaluation of distribution in the column (Figures 4, 5 and 6) but still considered when\nestablishing the MPs recovery rate across the experiment (cf: Materials and Methods).\nWhen considering the portion of PP-MPs in the top layer of bulk ice, strong enrichment is\nobserved as shown in Supplementary Figure 5 (C /C = 7.37  2.53 and 3.22 1.31\nbulk ice 0\nfor Large and Small PP-MPs, respectively).\nOverall, clear density-dependent trends are observed, even when excluding the top layer\nof ice with PP-MPs that could have been adsorbed to the air/water interface (Fig. 4) The\nhigh-density PET-MPs are depleted and the low-density PP-MPs concentrations remain\nalmost constant. Levels of depletion/enrichment increase with particle size (Supplementary\nSection 1). The Large PET-MPs were significantly more depleted from the bulk ice than\nSmall PET-MPs with C /C = 0.06  0.05 and 0.49  0.25, respectively. This is\nbulk ice 0\nexplained by the fact that particle’s settling or rising velocity is directly proportional to the\npower of its size. Conversely, Large PP-MPs had a higher C /C value of 1.21  0.59\nbulk ice 0\nthan Small PP-MPs (0.86  0.25).\nMinor comments:\n1) Line 12 – Sea ice forms near human activity too (Canadian towns, Longyearbyen) so please\nremove ‘despite being far from human activity’. You may want to add that it transports contaminants\nas this is important and mentioned prominently in the intro.\nYes, we agree with the reviewer and have now removed this statement about remoteness and only\nmention its capacity to transport contaminants.\nLines 12 – 13 : Sea ice accumulates various contaminants and can redistribute them\nlaterally as the ice drifts and vertically as ice melts.\n2) Line 14 – Please make clear what aspects are ‘novel’. The suite of chemicals is novel, but the\ncold plate, growth in tubes, artificial salt solution are not novel. If it is some combination of the\ndesign choices that is novel for some specific novel need, that should be made clear and justified.\nThe methods don’t need to be novel for the work to be novel and worthwhile.\nWe can understand the reviewers concerns that many researchers over emphasize the novelty of\ntheir work, and a combination of factors is in itself not necessarily novel since there are many\nunique research lines which can be explored. There, we have more clearly specified what is novel\nabout this experiment in the main text.\nenvironmentally representative artificial sea ice by keeping the internal pressure and the\nsalinity of the underlying water constant and ii) it allowed us to quantify model contaminants\nwithin the sea ice.\n3) Line 18 – What precisely does ‘difficult’ mean? Are you referring to contamination issues?\nWe have modified the sentence in the abstract to clarify what we mean by difficulties in quantifying\ncertain contaminants:\nLines 20-21: and difficulties quantifying certain contaminants due to the lack of\nstandardized methods.\nWe have also detailed the reasons behind this difficulty in the introduction\nLines 74 - 78 : Furthermore, quantifying some contaminants, such as small MPs (<10 m)\nand NPs, in environmental matrices is challenging due to the lack of standardized methods.\nSome persistent challenges, such as differentiating MPs and NPs from natural organic\nmatter (NOM) and avoiding, contamination during sampling and analysis require time-\nintensive sampling and extraction methods. Overall, this limits the amount of samples\nwhich can be measured.\n4) Line 20 – Are kDa and um related to each other somehow? I don’t see how a range of values\nfrom x to y is useful when x and y have different units.\n1 nm is roughly equivalent to 1 kDa. Daltons are the more scientifically accurate unit to assess the\nsize of small colloids, since they are malleable and their dimensions will depend on solution\nchemistry. However, we agree that using two different units is confusing and so we will use the\ndefinition of colloids as 1nm to 1m as per the definition of Lead and Wilkinson (2007). Changes\nwere made throughout the text and the reference was added line 66.\n5) Line 58 – The authors will need to be more clear about what ‘brine rejection’ is referring to.\nGenerally, ‘brine rejection’ refers to brine moving from the sea ice to the ocean below, which\ndesalinates the sea ice. This happens after sea ice has grown and incorporated everything. There\nis insignificant segregation at the advancing interface, so everything that was in the water is\nincorporated into the growing sea ice, then rejected (Notz & Worster,\n2009, https://doi.org/10.1029/2008JC004885). What the authors are describing is salt and other\ndissolved species being excluded from the ice matrix such that they are partitioned to the brine,\nbut remain within the sea ice.\nThank you for bringing this mistake to our attention. We have modified this sentence and added\nanother one, as well as the Notz and Worster reference, for further clarity.\nLines 60- 63: During bottom freezing, dissolved molecules such as sea salts are depleted\nfrom the bulk ice and accumulated in brine pockets and channels, a process known as salt\nsegregation or partitioning. This can be followed by gravity drainage, or flushing of the brine\nin the sea ice which leads to overall desalination of the bulk sea ice.\n6) Line 59 – The authors should be careful to use ‘ice’ to mean the frozen, ice matrix of sea ice,\nand ‘sea ice’ or ‘bulk sea ice’ to mean the collection of ice and brine. Here, ‘bulk ice’ is used, which\nis confusing.\nWe have made the correction here and throughout the text.\n7) Line 62 – (see minor comment 4)\nWe have made the correction.\nLines 66 and 67: dissolved (< 1nm) […] colloidal (1nm to 1 μm)\n8) Line 77 – ‘Particulate species with low densities are expected to be enriched…’ Enriched relative\nto what? Where does this hypothesis come from? The other hypotheses follow from the\nintroduction, but this one seems unconnected to the previous introduction material.\nWe have added some clarifications to this part :\nLines 85 – 88 : On the other hand, particulate species with low densities are expected to\nbe enriched in the ice relative to the underlying water due to their positive buoyancy which\nmoves them towards the advancing ice front. Conversely high-density particles, due to\ntheir negative buoyancy, are expected to be depleted by sinking below the advancing ice\nfront.\n9) Line 83 – This is probably the place to address minor comment 2.\nWe have addressed this comment :\nLines 94 to 97: The experimental set-up was a novel development since i) it generated\nenvironmentally representative artificial sea ice by keeping the internal pressure and the\nsalinity of the underlying water constant and ii) it allowed us to quantify model contaminants\nwithin the sea ice.\n10) Line 90 – Defining ice and brine earlier could solve previous comments.\nWe have added the distinction between ice and bulk ice, lines 58 and 61. We have also added\nsome clarity to this sentence :\nLines 100 – 102 : The artificial sea ice formed was harvested to characterize the artificial\nsea ice structure and to quantify species in the underlying liquid, the bulk ice, as well as its\ntwo fractions: the solid ice matrix (hereafter called ice) and the brine.\n11) Line 92 – ‘Trapped’ probably isn’t the right word here.\nWe have changed the term to “incorporated” (line 103)\n12) Line 92 – ‘Collectively this’ is imprecise (and should be ‘these’ I think). I would suggest replacing\nthis with ‘Our results’ or ‘These knowledge gains’, if that’s what is meant.\nThank you for your suggestion. We have made the suggested changes.\nLine 103-104 : Our results improve our ability to […].\n9\n13) ‘4’ is repeated in the figure 1 caption and ‘2’ is missing. Also ‘column’ should be ‘a column’ or\n‘columns’. This figure is very clear and informative.\nWe made the appropriate corrections in the caption of Figure 1.\n14) Line 108 – ‘vertical temperature gradient’.\nWe added “vertical” to “temperature gradient” in line 119.\n15) Line 114 – ‘average’ to ‘mean’ (I presume).\nWe have made corrections to “mean” throughout the text.\n16) Line 129 – ‘brine salinity approximately doubled’. The authors should calculate and report to\ntemperature in Vancoppenolle et al. (2019). This comparison will help the authors think about how\nrepresentative their centrifuged brine is of the in situ brine.\nWe have added this comparison to our discussion\nLines 142 to 146 : This brine salinity was compared to predictions of brine salinity as a\nfunction of temperature5. We used -1.7 oC as the temperature at the bottom of the ice core\nsince it is the freezing temperature of our artificial seawater (31.28 g L-1) and the\ntemperature of the cooling element, -5.1 oC, as the temperature at the top of the ice core.\nThis suggests that the brine salinity should be 57.8 g L-1 which is consistent with our\nobservations.\n17) Lines 133-135 – It is great that the authors have thought deeply about this aspect of the\nexperiment and have noted it here. It then needs to feature in the discussion (see also point 2.4).\nThis aspect is now further discussed in lines 196 to 199. We have further elaborated on the\nlimitations involved in separating brine by centrifugation throughout the manuscript\nLines 165 - 167: This can again be attributed to some brine loss, as well as some residual\nbrine pockets within the ice that were not connected to the bottom of the ice, and therefore\nnot drained by centrifugation. Despite these experimental limitations, our results agree well\nwith the properties of natural sea ice.\nLines 261 to 264 : Note that, as for the salts, it is also possible that some Rose Bengal in\nbrine pockets was not centrifugated out of the ice. This would increase their measured\nconcentration in the ice and decrease their measured concentration in the brine.\n18) Lines 136-143 – These values are reassuring, and suggest the artificial sea ice is a reasonable\napproximation of the real world. I would note that the sea ice in this study is very young (<5 cm),\nwhereas real world sea ice is considered ‘young’ while less than 30 cm – I would expect 30 cm\nsea ice to have lower salinity and solid fraction. Also, the values reported by the authors here will\nbe biased by brine loss during sampling (so before centrifuging). This bias is practically\nunavoidable. These caveats should be noted here. The authors could compare to Notz & Worster\n(2008) which present in situ measurements of bulk salinity and solid fraction\n(https://doi.org/10.1029/2007JC004333), and have measurements in <5 cm sea ice. For\n10\ncomparable thicknesses the authors will see the in situ measurements will have a higher bulk\nsalinity and solid fraction.\nThank you for bringing this to our attention. We have further discussed these aspects in the main\nmanuscript.\nLines 161 to 170 : For example, Notz and Worster (2008) observed that sea ice of\ncomparable thickness had a bulk salinity ranging from 15 to 25 g L-1 6. Our lower bulk\nsalinity may be explained by the fact that brine may be lost when the artificial sea ice is\npulled out of the column. The solid mass fraction of their sea ice ranged from 0.5 to 0.76\nwhich is lower than our solid mass fraction of 0.85  0.04. This can be again attributed to\nsome brine loss, as well as some residual brine pockets within the ice that were not\nconnected to the bottom of the ice, and therefore not drained by centrifugation. Despite\nthese experimental limitations, our results agree well with the properties of natural sea ice.\nIt should also be noted that this artificial sea ice produced represents the very early stages\nof sea ice growth, since young sea ice can reach up to 30 cm depth and can desalinate to\n4 to 6 g L-1 1,6.\n19) Line 155 – Why did the freezing element have a curved shape?\nTo avoid any entrapment of air bubbles between the freezing element and the surface of water, the\nfreezing elements was designed with a curved shape. This allowed the air to escape when the\nfreezing element was inserted.\nLines 184 - 186 : This curved shape was used to allow air bubbles to escape as the cooling\nelement was plunged into the water, to enable full contact between cooling the element\nand the water.\n20) Figure 2b – Could the authors please annotate the image? I had to guess what the lenticular\nstructures were. Also, what is ‘black’ in the image?\nWe have now annotated the image in order to make the areas of interest more immediately visible.\nWhat appears “black” is simply the edges of the grey zones which delimit the ice. Indeed, in the\nrendering the grey area has darker edges at the border with the white areas which show the “air”\n(i.e.: drained brine).\n21) Line 171 – ‘if’ to ‘is’\nWe have made the change in Line 230.\n22) Line 229 – Needs an ‘and’ between the reported values.\nWe have made the change in Line 289.\n23) Line 255 – ‘already’ to ‘already been’\nWe have made the change in Line 314.\n24) Section 2.4 – As a minimum, I would like to see a line here reminding the reader that Cice in\nthis study includes brine in isolated pockets. It would also be very useful to get the authors take on\n11\nhow the inclusion of isolate brine within Cice impacts their conclusions. I found it difficult to\nunderstand how, for example, the exposure of microorganisms to contaminants is impacted by\nhaving this imperfect separation of phases in the measurement. The authors should also note bias\nfrom brine loss upon sampling and speculate as to impacts on conclusions. Possibly these caveats\ncould form a short paragraph, as this paragraph is already quite complex and well put together.\nWe have discussed the limitations of our experiments and how these affect our interpretations of\nin section 2.5 called “2.5. Suitability of lab-scale experiments to study incorporation of\ncontaminants in sea ice”. Specifically, this point was addressed in the following way:\nLines 387 to 395: With respect to the type observations that can be made, this design does\nnot enable the use of methods that avoid the loss of brine as it the sea ice is removed (i.e.:\nby inserting a box under the sea ice as it is pulled out54). Furthermore, the use of\ncentrifugation may not drain the brine pockets that are not connected to the bottom of the\nice core. Therefore, the brine content is underestimated and the ice salinity is\noverestimated, compared to methods which measure brine content in-situ43. Future\nexperiments could incorporate in-situ temperature and salinity measurements to record the\nice growth and brine content in real time. However, since microorganisms also thrive in the\nbrine channels that are connected to the underlying water, the brine fraction studied here\nis the more biologically-active fraction of the sea ice.\n25) Figure 5 is excellent.\nThank you !\n26) Line 323 – ‘pressure of crystal growth’? Do the authors mean actual pressure (as in Pa), and\nif not, what?\nWe have clarified this sentence to mean the fact that ice solidification can move colloids and push\nthem towards each other which increases the possibility of interaction.\nLines 427- 431: Indeed, sea ice is an ideal environment for the aggregation of colloids and\npotentially also for the self-assembly of dissolved species into colloidal or particulate\nspecies due to i) the local increase in salt and colloid concentrations in brine, and to ii) the\ngrowth of solid ice which can push colloids closer together thereby increasing the possibility\nof interaction.\n27) Glossary: The statements about this sea ice containing no gas should be softened. Gas is\noften found in young sea ice, and there could be some here, unless the authors have some\nevidence to the contrary.\nWe have made the changes accordingly:\nLines 597 - 598: The artificial sea ice created here contains solid and liquid fractions (i.e.,\nice and brine, respectively). Gas bubbles were not observed by -CT.\n28) At the end of the paper I realised I don’t understand why the low density MP were so depleted\nin the liquid. Scanning back through I couldn’t find the answer. Could the authors please explain\nthis, or make the explanation more prominent.\nWe have made this explanation more prominent:\n12\n1) The first part of section 2.2 (lines 210 to 232) discusses the incorporation of MPs in bulk\nice, and more clearly demonstrates the impact of MP density\n2) The paragraph which compares MP concentrations in ice and liquid states more\nprominently the role of particle density.\nLines 266 to 281: As demonstrated with the enrichment/depletion trends of MPs in bulk ice,\nMP’s enrichment/depletion patterns are attributed to i) polymer density, which confer a\npositive buoyancy (i.e., rising) for the low-density PP-MPs and a negative buoyancy (i.e.,\nsettling) for the high-density PET-MPs and to ii) size, which accelerates the\nenrichment/depletion of MPs (Fig. 4, Fig. S5, and Supplementary Section 1). This is further\ndemonstrated by comparing concentrations in ice and in the underlying liquid. […] PP-MPs’\nenrichment was compensated by a depletion in the underlying liquid for the PP-MPs (C/C\n0\n= 0.308 ± 0.308 and 0.682 ± 0.158 for large and small PP-MPs, respectively).\n29) I would also suggest saying ‘: The impact of…’ in the title as it’s plainer English.\n(Just a note to say I thought the section titles were very helpful. The declarative statements made\nthe paper easy to follow.)\nWe have made the suggested change to the title \nReviewer #3 (Remarks to the Author):\nUnderstanding Contaminant Incorporation in Sea Ice: Impact of Size and Density\nPradel et al\nThis manuscript details the experimental set-up and application of an artificial sea ice chamber\nsystem that allows the intimate study of solutes and particles, present in sea water, and their\ninteraction with freshly forming ice. The authors are to be applauded for their relatively simple but\nelegant experimental set up. To my knowledge this small-scale system with multiple plastic polymer\ncolumns, an air-side cooling system (representing real Polar conditions over surface seawater)\ncombined with a pragmatic pressure release tube is novel. Given the chemical and physical\nmeasurements of the fresh ice plug (with CT imaging) then similarities to initial ice in the Arctic is\nconvincing. Hence this column system does provide a unique system to study the uptake, release\nand fate of both dissolved and particulate contaminants in ice (e.g. persistent organic chemicals,\nplastic particles, POC/DOC, heavy metals etc). The authors then demonstrate the utility of the\napparatus by examining the fate of selected solutes, colloids and plastic particles (varying density)\nduring a freeze experiment. Enrichment/depletion of these species relative to their occurrence in\nseawater is then reported with comparisons, where possible, to field and laboratory studies. The\nstudy is at the standard for publication however there are several issues that the authors need to\ncomment on or address before publication could proceed.\nWe thank the reviewer for their enthusiastic and positive comments for our work, and we hope that\nthis experimental design can also be implemented by others in the future in order to accurately\nassesses various processes and contaminant pathways within sea ice. With the additional changes\n13\nsuggested by this and other reviewers, we believe that we have further refined the manuscript for\nfuture readers to give the work its full potential.\n1) The columns and their seawater contents were agitated by being placed on an orbital shaker\nduring the freeze period. It is not entirely clear why this was undertaken but does make sense in\nthat particulate matter is likely to remain suspended in the water column, rather than deposited to\nthe vessel floor (for the higher density particles). This agitation however does not represent a sub-\nice seawater current though. This could be achieved using a small submerged pump or similar,\nwhich is common in larger sea ice chambers, which hence do resemble the surface ocean with\nthis regard. On this note there doesn’t appear to be incorporation of sensors to measure physical\nparameters of the seawater itself. Most notably the temperature/salinity profile down the water\ncolumn should be essential. For example, the expulsion of brine with ice aging or the release of\nlow salinity melt water during ice melt, may result in stratified water layers which will affect\nchemical/material flows and yet are not easily distinguished in the current system.\nAs the reviewer has already identified, the purpose of placing the columns on orbital shakers was\nto aid in homogenously dispersing the contaminants, particularly the MPs. We agree that circular\nagitation motion is not representative of underwater currents, but despite this limitation, we were\nable to achieve and structure in the sea ice which is comparable to very young natural sea ice. It\nis also notable that the dissipation energy generated through the orbital shaker is on the order of\n10-6 m2 s-3 which is a median value for the open ocean, according to Arnott et al (2021)7.\nTo generate currents that are more environmentally representative and to measure the\ntemperature and salinity profiles of the ice and of the underlying seawater would require making a\nlarger experimental set-up. Indeed, a lateral current cannot easily be generated in a column of 6.5\ncm diameter by 44 cm height. Also, we had put temperature sensors throughout the sides of one\ncolumn but the temperatures were too strongly influenced by the ambient temperature since the\nsensors could not be entirely plunged in the column due to space constraints. So, small sensors\nplunged into a larger volume of ice would enable in-situ measurements of temperature and salinity\nwhile minimizing disturbance.\nWe have addressed these experimental limitations in Section 2.5 “ 3. Suitability of lab-scale\nexperiments to study sea ice processes.”\nLines 378 – 382 : With respect to the type of ice grown, the advantage of our experimental\ndesign is that, by being relatively small it allows many replicates to be performed since the\nfreezing time and the amount of model contaminants required are reduced. However, this\nsmaller size does not enable the generation of underwater currents which occur naturally\nunder sea ice. Despite this, the artificial sea ice formed is similar to natural ice.\nLines 392 – 393 : Future experiments could incorporate in-situ temperature and salinity\nmeasurements to record the ice growth and brine content in real time.\n2) The freezing process using the aluminium cooling vessel will interfere with air side transfer of\nparticles and vapour/gaseous species. While the focus is clearly on studying the ice itself and the\nice-seawater interface, this is nonetheless a significant oversight particularly for studying the\ntransfer of material/chemicals across the ice-air interface. Importantly, the introduction of material\nto the surface of the ice to simulate ice-rafted snow for example cannot be undertaken with the\ncurrent set up.\nfocuses solely on processes occurring at the water/ice interface. This is due to our choice of looking\nspecifically at bottom freezing, which, as indicated in the introduction, forms most of the volume of\nsea ice (lines 58 to 60). As noted by Hall et al. (2023)8, experimental designs which can generate\nboth frazil and columnar ice generally require larger volumes. However, this also has limitations\nsuch as reduced capacity for replicate experiments and reduced control on temperature gradients.\nIn sum, since no experimental design is perfect, we have discussed the limitations and how they\nimpacts our results in section 2.5.\nLines 382 to 385: One must also keep in mind that this experimental set-up generated\ncolumnar ice, which grows below the layer of frazil ice and, as such, does not interact with\ncontaminants at the air/water interface. However, this columnar ice is most common in the\nArctic, which receives more anthropogenic contaminants than the Antarctic.\nLines 422 – 423: This can help to understand whether contaminant incorporation occurs\nduring basal ice growth or frazil ice growth.\n3) In the Introduction the authors cite those studies that have observed microplastics in Polar sea\nice at concentrations orders of magnitude higher than concentrations present in the underlying sea\nwater. This is one of the drivers behind experimental simulations of the seawater/sea ice system\nlike this study. However, in the experimental work for the microplastics the authors discarded the\nuppermost 0.5cm ice layer on the basis that lower density particles had accumulated in the water-\nair interface and hence had become artificially rafted onto the ice-surface. Surely, this is the very\nprocess which will be occurring to some extent for positively-buoyant plastic particles in the real\nmarine environment! Does this ice-rafting of some types of microplastics occur during early\nformation of grease/frazil ice?? This is a key question which hopefully this new experiment\napparatus could investigate, but which the authors have viewed as an artifact!!\nWe agree that the early stages of sea ice growth (i.e., grease/frazil ice) may incorporate MPs that\nare present at the seawater/air. However, the type of artificial sea ice produced here is not grease\nor frazil ice, but columnar ice that grows by bottom freezing and does not interact with the air. This\nis why we have discarded the MPs stuck at the air/ice interface (cf: previous response). We would\nlike to point out that we have another study, currently under review, which uses a similar\nexperimental set-up to generate frazil ice and to investigate this specific process. So stay tuned\nfor more information on this topic in (hopefully!) the near future! \nP1, line 28 contaminantation\nLine 31 : Contamination\nP2, line 72 ….in [a] controlled environment…\nLine 83 : in a controlled\nP2, line 94 ….wil be exposed to.\nLine 110 : will be exposed to\n1. Thomas, D. N. & Dieckmann, G. S. Sea Ice. (Blackwell Publishing Ltd, 2010).\n2. IOC, SCOR & IAPSO. The International Thermodynamic Equation of Seawater – 2010:\nCalculation and Use of Thermodynamic Properties. 196 https://www.teos-10.org/pubs/TEOS-\n10_Manual.pdf (2010).\n3. McDougall, T. J. & Barker, P. M. Getting Started with TEOS-10 and the Gibbs Seawater\n(GSW) Oceanographic Toolbox. (SCOR/IAPSO WG127, 2011).\n4. Schefer, R. B., Armanious, A. & Mitrano, D. M. Eco-Corona Formation on Plastics:\nAdsorption of Dissolved Organic Matter to Pristine and Photochemically Weathered Polymer\nSurfaces. Environ. Sci. Technol. 57, 14707–14716 (2023).\n5. Vancoppenolle, M., Madec, G., Thomas, M. & McDougall, T. J. Thermodynamics of Sea Ice\nPhase Composition Revisited. J. Geophys. Res. Oceans 124, 615–634 (2019).\n6. Notz, D. & Worster, M. G. In situ measurements of the evolution of young sea ice. J.\nGeophys. Res. 113, 2007JC004333 (2008).\n7. Arnott, R. N., Cherif, M., Bryant, L. D. & Wain, D. J. Artificially generated turbulence: a review\nof phycological nanocosm, microcosm, and mesocosm experiments. Hydrobiologia 848,\n961–991 (2021).\n8. Hall, B., Johnson, S., Thomas, M. & Rampai, T. Review of the design considerations for the\nlaboratory growth of sea ice. J. Glaciol. 1–13 (2023) doi:10.1017/jog.2022.115.\n16"}, {"round": 2, "reviewer_comments": {"reviewer_1": "As the authors have repeatedly stated, the simulation experiments indeed have many limitations, but they have made the greatest possible effort to address my comments. I recommend that this version of the manuscript can be published.", "reviewer_2": "The authors have thoroughly responded to my first review, and I am happy that the manuscript is fit for publication.\nThere are three further points I suggest they address:\n1) Comment 2, new lines 94-97: There are existing systems which keep pressure from building, so this aspect is not novel (Thomas et al. (2020) for example). The chemical suite should be mentioned here as that is novel. The curved cooling plate and net are also new to the sea-ice literature I think.\n2) Figures 4 and 5 should be combined as they share axes and this would aid comparing them.\n3) On lines 24-25 in the abstract, where the enrichment/depletion is discussed, it should be made clear that this is relative to salt.", "reviewer_3": "I have now reviewed the authors'responses to the comments made on the manuscript and they have satisfactorily addressed the points that I raised and as well the points raised by the other reviewers. Importantly the revised manuscript reflects this with substantial additions/changes made to the Results & Discussion. These additions improve the manuscript markedly and now highlight the limitations and/or biases that occur in the experimental ice columns relative to the real environment. The rationale for excluding or removing the ice rafted plastic particles is now clearly articulated. Overall I recommend that this manuscript can now be published.\nThere are a couple of typos in the following lines...\n377 \"With respect to the type [OF] observations that can be made, this design does not enable\n388 of methods that avoid the loss of brine as [it] the sea ice is removed\""}, "author_response": "Answer to reviewers\nReviewer #1 (Remarks to the Author):\nAs the authors have repeatedly stated, the simulation experiments indeed have many limitations,\nbut they have made the greatest possible effort to address my comments. I recommend that this\nversion of the manuscript can be published.\nThank you for your time reviewing our work.\nReviewer #2 (Remarks to the Author):\nThe authors have thoroughly responded to my first review, and I am happy that the manuscript is\nfit for publication.\nThere are three further points I suggest they address:\n1) Comment 2, new lines 94-97: There are existing systems which keep pressure from building,\nso this aspect is not novel (Thomas et al. (2020) for example). The chemical suite should be\nmentioned here as that is novel. The curved cooling plate and net are also new to the sea-ice\nliterature I think.\nWe have made the changes according to your feedback :\nLines 94-97: The experimental set-up was a novel development since i) it generated\nenvironmentally representaBve arBficial sea ice by keeping the salinity of the underlying\nwater constant, and ii) it allowed us to quanBfy a suite of model contaminants within the sea\nice.\n2) Figures 4 and 5 should be combined as they share axes and this would aid comparing them.\nThank you for your suggesBon. However, we find that combining the two figures would\nmake them too data-heavy and less clear to read. Furthermore, the disBncBon between bulk\nice and (brine-free) ice might be blurred by combining them. Therefore, for clarity, we would\nprefer to keep them separate.\n3) On lines 24-25 in the abstract, where the enrichment/depletion is discussed, it should be made\nclear that this is relative to salt.\nThis is relative to the initial concentrations in seawater. We have specified this :\nLines 24-26 : While high-density parBculate contaminants were depleted in sea ice and low-\ndensity parBculate contaminants were enriched relaBve to their iniBal concentraBon in\nseawater, both were engulfed and traveled in the wider brine channels.\nReviewer #3 (Remarks to the Author):\nI have now reviewed the authors'responses to the comments made on the manuscript and they\nhave satisfactorily addressed the points that I raised and as well the points raised by the other\nreviewers. Importantly the revised manuscript reflects this with substantial additions/changes\nmade to the Results & Discussion. These additions improve the manuscript markedly and now\nhighlight the limitations and/or biases that occur in the experimental ice columns relative to the\nreal environment. The rationale for excluding or removing the ice rafted plastic particles is now\nclearly articulated. Overall I recommend that this manuscript can now be published.\nThank you for your time reviewing our work.\nThere are a couple of typos in the following lines...\n377 \"With respect to the type [OF] observations that can be made, this design does not enable\nthe use\n388 of methods that avoid the loss of brine as [it] the sea ice is removed\"\nWe have corrected both typos."}]

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