in situ 3D characterization of bidisperse cakes using confocal laserscanning microscopy

Ines Ben Hassan a,b,c,d, Christine Lafforgue a,b,c, Abdelmoneim Ayadi d, Philippe Schmitz a,b,c,n

a Université de Toulouse; INSA, UPS, INP; LISBP, 135 Avenue de Rangueil, F-31077 Toulouse, Franceb INRA, UMR792 Ingénierie des Systèmes Biologiques et des Procédés, F-31400 Toulouse, Francec CNRS, UMR5504, F-31400 Toulouse, Franced LRAE (Laboratoire Radio analyses et Environnement), Université de Sfax, Sfax, Tunisia

a r t i c l e i n f o

Article history:Received 8 February 2014Received in revised form8 April 2014Accepted 21 April 2014Available online 26 April 2014

Keywords:FiltrationBidisperse particlesMicrosieveCakeCLSM

a b s t r a c t

In situ 3D characterization of a filtration cake obtained with a model bidisperse suspension containingtwo sizes of fluorescent particles (1 mm and 4.8 mm) has been performed using Confocal Laser ScanningMicroscopy (CLSM). The suspensions were dead-end filtered through 0.8 mm pore diameters siliconnitride microsieves, under a constant flow rate. The membrane was put in a specific filtration chamberpreviously designed [1], allowing direct on-line microscopic observations of particles deposition. Thefinal position of the first particles arriving on the membrane was investigated. Then, the onlinedeposition of the following particles was recorded and, thus, the cake build-up could be analyzed layerby layer. In addition, to analyze the protective effect of a layer of large particles, 1 mm particles werefiltrated over a previously deposited cake formed with the 4.8 mm microspheres.

& 2014 Elsevier B.V. All rights reserved.

1. Introduction

Microfiltration membrane processes are widely encountered inindustry in order to clarify the products retaining micro-sizedparticle (biotechnology, water and wastewater treatment, food andpharmaceutics) [2]. Usually, membranes with pores much smallerthan the size of the particles to be retained are selected; thus, apredominant external fouling is expected [3]. However, theretained particles lead to membrane fouling that reduces produc-tivity and changes membranes selectivity, which makes theprocess less efficient [4] and increases operating cost [5]. Indeed,the cake layer build-up on the membrane surface is the mostimportant drawback of these processes. In most cases, suspensionscontain a mixture of particles of different sizes (fermentationbroth, wastewater, juices, alcoholic beverages, etc.) [6]. Althoughthe mechanisms leading to fouling during filtration are complexand not yet fully understood, macroscopic laws have been devel-oped assuming basic mechanisms taking place such as poreclogging and cake formation [7]. Indeed, cake characteristics(mass, thickness, porosity, particle size distribution, etc.) that playa key role in the fouling effect on filtration [8] have been poorlystudied at particle or pore scale. Interactions between the diffe-rent feed components as well as particle organization on the

membrane surface are not accessible by the classic fouling studytechniques. ex situ techniques like scanning electron microscopy(SEM) and atomic force microscopy (AFM) are relevant techniquesfor autopsying the structure of the fouled layer with high resolu-tion levels [5]. However, they require that the cake and themembrane be removed from the filtration chamber and some-times treated before observation, which could modify its organi-zation. Several optical techniques have been proposed to performan accurate in situ characterization of cake local properties, such asthe use of a laser beam [9] and direct visualization (DO) methods [8].These optical techniques provide accurately in situ sensitive real-time observation [10]. However, their main drawback is that theobservation is limited to the focal plane and/or the membranesurface on the side view [10]. Recently, direct observation techni-que of filtration cake using a confocal laser scanning microscope(CLSM) has been proposed. In this in situ non-invasive fluores-cence imaging method, selected planes at given depths areobserved and planes out of focus are excluded [5]. A stack of thesuccessive images recorded at consecutive depths permits the3-dimensional image reconstruction of the whole sample [11].In the case of filtration cakes, the use of fluorescent dyes permitsone to differentiate membrane and cake, and allows the particledistribution on the membrane surface to be studied. In addition, inthe case of bidispersed suspensions, the observation of the cake atdifferent depths associated with particles dyed with differentfluorochromes (different fluorescent excitation and emissionwavelengths) could allow the distinction between the different

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journal homepage: www.elsevier.com/locate/memsci

Journal of Membrane Science

http://dx.doi.org/10.1016/j.memsci.2014.04.0410376-7388/& 2014 Elsevier B.V. All rights reserved.

n Corresponding author.E-mail address: schmitz@insa-toulouse.fr (P. Schmitz).

Journal of Membrane Science 466 (2014) 103–113

species [11]. However, laser penetration in the cake is limited anddepends on the concentration and transparency of the particle [5],to scattering and absorption of the excitation and emission light[12]. The decrease in fluorescence as a function of cake depthcould make it difficult to analyze thick fouling layers [5]. Anoriginal strategy for analysis of cake build-up, proceeding step-by-step examination of the result of the addition of successivelayers upon the cake, has been previously described and applied tothe filtration of a monodisperse suspension of fluorescent particles[13]. In this work the effect of the relative sizes of particle and poreon membrane fouling has been studied, thanks to the observationby CLSM of the cake organization.

The current work aims to study more realistic situations such asthose encountered in various applications where suspensions con-taining different sizes of particle are filtered. The objective is to betterunderstand the mechanisms of cake build-up in the case of abidisperse suspension. A multilayered cake is built layer by layer ona microsieve, by deposing successively small amounts of particlesand observed in situ by CLSM. The description of the characteristics ofthe cake (porosity, mass, homogeneity, etc.) is combined withmacroscopic measured parameters. Different combinations of largeand small size particles in the suspension have been undertaken.

2. Materials and methods

The experimental methods used in the current work are thesame as the ones previously described [13].

2.1. Microsieve and model particles

The silicon nitride microsieves (25 10�6 m2 total area) (Aqua-marijns) were considered as the best compromise in terms ofmembrane area minimization and permeate flow rate maximiza-tion [14]. Membranes with 0.8 mm pores diameter, 1.6 mm porepitch and 22.7% porosity were used (Fig. 1). Each microsieveconsists of 14 identical filtrating stripes (6.3 10�6 m2

filtrationarea). Before experiments, the microsieves were plasma treatedwith a Plasma Cleaner (Harrick Plasma, Belgium) and wetted withisopropanol.

After filtration experiments, water fluxes were expressed aspermeability reduction LnP:

LnP ¼LcPL0P

ð1Þ

where Lcp is the water permeability at a given time of the filtrationexperiment and L0p the water permeability of the clean microsieve.The fluorescent polystyrene microspheres, 1 mm and 4.8 mm dia-meter (Table 1), (1.05�103 kg/cm3), were provided from Thermo

Scientific (USA). Their characteristics have been detailed in aprevious work [13].

2.2. Filtration procedure with direct observation system and in situCLSM analysis

The design of the filtration chamber used to perform in situ,layer by layer, analysis of cake building was the same as describedin a previous work [7]. This apparatus was directly set under aCLSM microscope (LEICA SP2, DMRXA2 and TCS AOBS) duringfiltration experiments, allowing a direct microscopic observationof particle deposition and cake formation. Images were recordedon-line with the Leica CLSM software. The volume of injectedsuspension and the permeate flow rate were measured by weigh-ing the liquid collected after the pump during a few minutes(triplicates measurements). The transmembrane pressure duringfiltration was monitored (vacuum meter Thyrcont, Germany).

In order to overcome CLSM limitation due to insufficient laserpenetration for compact and/or opaque deposit, the cake wasformed layer by layer, thanks to successive injections of theparticle suspension and images were recorded after each injection.

The clean microsieve was first observed in bright light mode todetermine the zero position of the z-axis and to verify its flatness.A selected volume of the suspension of fluorescent particles at aknown concentration was, then, flown to the membrane. Observa-tions were done in both bright light and fluorescent modes. Theparticles deposition on the microsieves was recorded with a 10�objective (x, y, t). The fouled membrane was, then, analyzed withthe 63� objective at two magnifications, zoom 1 and 6. CSLMimages consist of successive thin slices parallel to the microsieve(xy-plane) along the z-axis. Two scans of every image wereperformed to improve resolution. The z-series or stacks of imagestaken at a z-axis were distanced at 0.32 mm. After all imagerecording, a new suspension volume was injected and the sameprocedure was repeated.

2.3. Image analysis

Images were processed using Image J (1.45s downloaded fromhttp://rsb.info.nih.gov/ij/). The optical sections captured in the

Fig. 1. CLSM images of a silicon nitride microsieve surface (A) (objective 10� ) filtrating stripes and details of pores on the stripes (Objective 63� ), zoom 1 (B) and 6 (C).

Table 1Spectral properties of fluorescent microspheres.

Microspheresdiameter

Specifiedcolor

Excitation maximawave length (nm)

Emission maximawave length (nm)

1 mm Red 542 6124.8 mm Green 468 508

I. Ben Hassan et al. / Journal of Membrane Science 466 (2014) 103–113104

reflection and fluorescence modes could be visualized simulta-neously to localize the particles and microsieve. Images wereautomatically calibrated from the voxel sizes (depth, width andlength) and converted to grey-scale images. They were, then,segmented into background and foreground by setting a threshold.

The non-uniformity of the microsieves due to the succession offiltering stripes separated by non-filtering stripes (Fig. 1) wastaken into account in the calculation of the apparent fraction offiltration area covered by particles, αc:

αc ¼ ScSm

ð3Þ

Sc and Sm are, respectively, the surfaces occupied by the cake andthe area of the filtrating stripes. They are obtained from the imagestacks captured with the 63� objectives (zoom 1) in fluorescencemode, projected to get the top down view of the cake, and fromthe bright field mode image for the filtrating stripes, respectively.

According to Meng et al., the cake porosity could change greatlyalong the cake thickness [5]. In order to analyze the internal cakeconstruction, different parameters were determined based on theoptical sectional views obtained with the 63� objective (zoom 6).After binarization of gray-scale images fixing a threshold, the areacoated by the particles in each image along the z axis αs was

measured:

αsðzÞ ¼ SðzÞSf6

ð4Þ

S(z) is the area of the surface occupied by the particles at each imageof the stack and Sf6 is the area of the filtrating area at zoom 6.

3. Results and discussion

The formation and the morphology of the cakes obtained bydead-end filtration of bidispersed polystyrene fluorescent micro-spheres are studied according to the procedure detailed in aprevious paper [13], i.e. each sequence of filtration is divided intosuccessive filtration steps.

Four experiments are performed (Table 2) in order to analyzethe collaborative or protective effects of particles of different sizeson membrane fouling and cake characteristics when they aremixed together:

– Experiment E1: a single filtration sequence of a monodispersedsuspension of 1 mm particles.

– Experiment E2: a first filtration sequence of a monodispersedsuspension of 4.8 mm particles followed by a second filtration

Table 2Particle concentrations in the suspensions filtrated in the experiments.

Experiment Particle number ratio Feed concentration (g.L�1)

1 mm particles 4.8 mm particles

E1 3.34�10�4 –

E2 3.34�10�4 16.1�10�4

E3 1.67�10�4 8�10�4

E4 4�10�4 4�10�4

I. Ben Hassan et al. / Journal of Membrane Science 466 (2014) 103–113 105

sequence of a monodispersed suspension of 1 mm particles. Thevolume ratio of 4.8 mm particles to 1 mm particles is fixed to 5,thus the number of 1 mm particles is about 25 times higherthan the number of 4.8 mm ones.

– Experiment E3: a single filtration sequence of bidispersedsuspension. The suspension is obtained by mixing a volumeof 1 mm particles five times lower than the volume of 4.8 mmparticles. The number of smaller particles is, thus, the same asthe one in experiment E2.

– Experiment E4: a single filtration sequence of a bidispersedsuspension obtained by mixing an equal volume of 4.8 mm and1 mm particles. Here, the number of smaller particles is about125 times higher than the number of the larger ones.

As discussed in a previous paper [7], the membrane regionobserved at high magnification (say 63� , zoom 6) is assumed tobe representative of the whole membrane. The cake can beconsidered roughly homogenous at the membrane scale. Nonoticeable difference is observed along the filtration stripes.Additionally, the particles seem to deposit on all the stripes anda careful analysis of the filtration field shows that the particles aredeposited all over it, reflecting a uniform membrane substructure[15]. Moreover, the continuous recording of cake formation atmagnification 10� (record not shown) confirms that particlesarrive one by one to the microsieve filtration surface, no aggregateis observed as it was expected since the suspensions concentra-tions were very low. Furthermore, no rearrangement of theparticles is observed, which will be confirmed at a higher magni-fication level (63� ). These first observations confirm that themorphology of the cake characterized in a sample region of themicrosieve at the microscopic scale, i.e. the particle scale, can beconsidered as representative of all the cake over the membrane.

3.1. Permeability reduction

The decrease of permeability when particles accumulate at themembrane surface is expressed in terms of permeability reductionLnP versus mass deposited (Fig. 2). As observed in a previous work[13], the smaller size particles always have a drastic effect onpermeability reduction. It is especially obvious during experimentE2 since LnP sharply decreases from about 45% when 44 mg of4.8 mm particles are deposited, to 5% once a very small amount of2 mg of 1 mm particles are additionally deposited. We can also seethat the different initial rates of permeability reduction dLnP=dMT

obtained for experiments E1, E2 and E3 (straight lines in Fig. 3) areconsistent with the number of 1 mm particles deposited for a giventotal mass filtrated MT. The number of 1 mm particles which arelikely to be deposited are represented by N1, N3 and N4 forexperiments E1, E3 and E4 and, for the sake of simplification, weassume that the diameter ratio between big and small particles isabout 5. Then, according to Table 2, for the same total mass ofparticle filtrated MT, assuming that the particle density is about1 (for mass to volume conversion), the number of 1 mm particlesdeposited in experiments E3 and E4 should be equal to N1/6 andN1=2, respectively, and the corresponding mass of 1 mm particlesshould be MT=6 and MT=2, respectively. It means that N3oN4oN1.

Accordingly, we should obtain jdLnP=dMT j3o jdLnP=dMT j4o jdLnP=dMT j1. This is what can be observed by comparing the beginning ofthe experiments Fig. 4. This result demonstrates that smallerparticles mainly prevail in pore blockage and, thus, in permeabilityreduction. On the contrary, the blockage of one pore by biggerparticles results in the protection of neighboring pores that remainopen [13]. This particular mechanism was previously observedduring the CLSM analysis of filtration of monodispersed particles.It will be discussed in detail in the next section about filtration ofbidispersed particles. It can be also seen in Fig. 2 that LnP is keptslightly higher in experiment E2 than in experiments E3 and E4.It is probably favoured by the same mechanism. Therefore, it canbe assumed that it could be helpful in some applications toperform a preliminary filtration of big particles to maintain ahigher flux during the further filtration of smaller particles.

3.2. Cake build-up during the filtration of monodispersed particles(experiment E1 and first part of experiment E2)

In the previous paper about monodisperse spherical particlessuspensions, the effect of the relative ratio “pore diameter/particlediameter” has been carefully studied and the notion of protectiveeffect was discussed [13]. A summary of the main results is brieflygiven in the following. At the beginning of cake formation,particles preferentially deposit over pores. The geometrical ratiobetween particle size (1 mm or 4.8 mm) and microsieves porediameter (0.8 or 2 mm pore size) and pitch (1.6 or 3 mm, respec-tively) modified the fraction of protected pores surrounding theplugged one. A 1 mm microsphere can block only one pore of themembrane. A 4.8 mm particle protects a certain number of sur-rounding pores depending on microsieve type (0.8 mm microsieveor 2 mm microsieve). This probably explains why the combinationof larger particles and smaller pores microsieve exhibits the lowestpermeability reduction. These aforementioned mechanisms areexemplified for experiment E1 and the first sequence of experi-ment E2 as it can be observed in the corresponding images andschemes of Figs. 3 and 4. It was also noticed that pore bridgingmechanisms, often discussed in the literature [6], still existed. Thatprobably explained why the 0.8 mmmicrosieve was not completelyfouled during the filtration of 1 mm microspheres.

During the full cake formation, cake morphology was investi-gated in terms of microsieve coverage and porosity. According tothe previous paper, cake porosity was evaluated from the surfaceoccupied by the particle along the z-axis using image processing[13]. The results reported in Table 3 reveal that identical suspen-sions lead to cakes of roughly the same morphology revealingquite compact deposits except in the first cake layer as alreadyillustrated in Figs. 3 and 4.

The analysis of the surface covered by particles along the cakethickness exhibits the existence of three regions: a germinationregion in contact with the membrane with moderate particleconcentration, a central high particle concentration region and asuperficial region, named capture region. These results were in

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Cake mass (μg)

4,8μm particles injection

1μm particles injection

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Fig. 2. Fouling caused during the dead end microfiltration of the differentsuspensions of microspheres on 0.8 mm pores diameter microsieve: relativepermeability vs. cake mass; (▴): Experiment E1, (●): Experiment E2, (◆): ExperimentE3, (■): Experiment E4.

I. Ben Hassan et al. / Journal of Membrane Science 466 (2014) 103–113106

agreement with the pioneer theoretical works of Schmitz et al. in1993 [16] and those reported by Meng et al. [5]. Whatever theparticle size, the cake organization appears to be quite similarexcept in the germination region.

3.3. Cake build-up during the filtration of bidispersed particles

During the filtration of real suspensions, the feed compositionmay vary from very fine particles to molecular aggregates togranular solids [6]. It is well known that particle size has a major

effect on fouling separation performance. The effect of particle sizeon both pore blockage and first cake layer formation should becarefully examined for process optimization. Moreover, in the caseof particles with a size distribution, their location and arrange-ment inside the cake could be correlated to their size [17]. To thisend, the procedure of successive filtration step by step was used tomake a CLSM analysis at microscopic scale, i.e. at particle scale(magnification 63� , zoom 6). Recall that conducting such refinedexperiments is a long and very complex task. Each filtration stepwith the associated CLSM analysis lasts about 1 h, thereby limitingthe number of filtration sequences.

3.3.1. Simultaneous filtration of bidispersed particles3.3.1.1. Initial filtration steps. In experiment E3, the very first stepsof filtration can be studied as few particles are injected at eachfiltration step. The final total mass deposited is low (less than

Fig. 3. Composite image of a 0.8 mm pore size microsieve fouled with 3 mg particle (A) and 7 mg of 1 mm particles (C) (first coming in yellow and the following in green)(objective 63� , zoom 6) and its schematic representations (B, D), respectively. (For interpretation of the references to color in this figure legend, the reader is referred to theweb version of this article.)

Fig. 4. Composite image of a 0.8 mm pore size microsieve fouled with (A) 4.8 mm particle (8 mg), its schematic representations (B, C, D), respectively, and composite image offirst arriving (green) and following (red). (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Table 3Effect of particle size on monodisperse cake porosity.

1 mm particles (E1) 4.8 mm particles (E2, sequence 1)

0.42 0.45

I. Ben Hassan et al. / Journal of Membrane Science 466 (2014) 103–113 107

17 mg) although the number of filtration steps is 5. Thereby, poreblocking mechanisms and progressive coverage of the microsievecan be accurately examined. As can be expected, the behavior offirst 4.8 mm and 1 mm particles deposited is in agreement with theobservations already made and described in Section 1. Indeed, itcan be seen in Fig. 5 that single particles are perfectly centeredover some pores and these pores are thus presumably blocked andclosed, i.e. the fluid cannot flow anymore.

Significant accumulations of small particles can be observed indifferent places over the microsieve. Small agglomerates werealready observed in the case of the filtration of only 1 mm particles(experiment E1) where bridging of some pores was also found. Thelargest accumulations of small particles mainly take place aroundthe large particles deposited, where fluid can still easily flowaround particles. Indeed, as schematically illustrated in Fig. 6,6 pores around each 4.8 mm particle centered over one pore,remain completely open as they are inaccessible to 1 mm particles.Since small particles aggregate around the larger ones, the flowtowards these aggregates should remain high and it can beassumed that the permeability reduction mainly results from poreblockage by isolated small particles instead of small particleaccumulation.

Now let us look at the next filtration steps which correspond tothe beginning of cake growth. The apparent fraction of themicrosieve filtration area covered by particles, αc, vs. total cakemass is plotted in Fig. 7 and compared to permeability reductionLnp. It should be noticed that a low value of αc (about 25%) issufficient to obtain a dramatic value of Lnp (about 15%). This stillconfirms that permeability reduction is driven by pore blockagemechanism at the beginning of filtration. When other particles

settle, αc significantly increases to reach 70% at the end ofexperiment E3. Accordingly the calculation of the mean cakethickness shows that only one to two layers of the 4.8 mm particleswere finally deposited. This is in agreement with the low ratio(1/25) between the number of 4.8 mm and 1 mm particles in thefeed solution of experiment E3. Finally, it can also be deduced fromFig. 7 that the aggregates of small particles accumulated aroundlarge particles tend to grow in both directions since the increase ofαc becomes smooth as far as mass deposited increases. This isconfirmed by the variations of the fraction of the area occupied byparticles, αS, along the z-axis showing that the cake thickens andslightly grows during the initial filtration steps (Figs. 8 and 9).Now, let us examine more carefully the area occupied by the4.8 mm and 1 mm particles (Fig. 9). If the smaller particles con-tribute equally to cake thickening and cake growth (Fig. 7), thelarger ones seem more involved in cake thickening as reported byFig. 9 where the respective fraction of area occupied by eachparticle type is plotted along the z-axis. However, the largeparticles are few because the overall number of particles injectedduring experiment E3 has remained quite low and the ratiobetween 4.8 mm and 1 mm particles in the feed solution is low(1/25). Thus, the subsequent thickness of the cake at the end ofexperiment is in the order of magnitude, of about three largeparticle diameters. Moreover, the shape of the αS vs. z-curve(Fig. 8) indicates that the cake contains only two regions (seeSection 1, [5]): the germination region and the superficial regionwhose thicknesses are of about one and two large particlediameters, respectively. The central region of high particle

Fig. 5. Composite images of microfiltration cake formed by bidisperse suspension:1.68 mg of 1 mm particles and 4.8 mm particles.

Fig. 6. Schematic representations of the microsieve fouled with bidisperse suspensions.

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Fig. 7. Evolution of the apparent fraction of the microsieve filtration area coveredby particles, αc, and the permeability reduction (Lnp) during experiment E3.

I. Ben Hassan et al. / Journal of Membrane Science 466 (2014) 103–113108

concentration does not yet exist. This is confirmed by the max-imum value of αS which only equals 0.4, giving a high void fractionof 0.6. It would not make sense to give a porosity value of the cakeat this stage.

3.3.1.2. Multilayer cake formation. Contrary to experiment E3 thatwas stopped after the injection of a quite low mass of particles(17 mg), experiment E4 was performed at higher mass injected(24 mg) in order to follow the formation of a multilayered cake. As

expected, the initial filtration steps of experiment E4 are found tobe quite similar to the ones of experiment E3. As illustrated inFig. 10, 1 mm particles mainly accumulate around the 4.8 mmparticles (yellow square) or block some pores of the microsieve.It should be noted that the larger particles also progressivelyaccumulate in these aggregates initially formed by the smallerparticles around one large particle (blue triangle). Then, bothparticle types contribute to the growth and thickening of theaggregates. These observations still confirm that the highest flowrate is maintained around the first 4.8 mm particles deposited and,thus, strengthen the assumption of the existence of the remainingopen pores around them.

Now have a look at the specific role of the 1 mm particles thatthey are much more numerous than the 4.8 mm particles, 25/1 and125/1 for experiments E3 and E4, respectively. The apparentfraction of the microsieve filtration area covered by particles, αc,has been specifically plotted vs. 1 mm particle mass deposited inFigs. 7 (experiment E3) and 11 (experiment E4). Recall that only the14 filtrating stripes (6.3 mm2) are the filtration area. For a givenvalue of 1 mm particle mass deposited, say 3 mg (identified by thestraight lines drawn in the figures), the corresponding value of αcremains the same in both experiments (0.7). This is an importantfinding showing that smaller particles predominately contribute tocake thickening and subsequently facilitate the progressive cakehomogenization.

It can also be seeing in Fig. 13 that the cake progressivelythickens to get apparently homogeneous in terms of spatialdistribution over the microsieve. Surprisingly the spatial distribu-tion of the two types of particles seems to become also homo-geneous during the growth of the cake (Fig. 13C) and only drivenby the very low ratio (1/125) between the number of 4.8 mm and1 mm particles in the feed suspension. At this filtration step, newarriving particles are no longer in contact with the heterogeneousgermination region. The growth of the multilayer cake becomesuniform all over the microsieve. The associated value of theapparent fraction of the microsieve filtration area covered byparticles, αc, reaches then 1 (Fig. 11).

Taking benefit of the CLSMmethod, the morphology of the cakecan then be carefully examined using the variations of the fractionof the area occupied by particles, αS along the z-axis. We havealready discussed in another paper [13] that the global approachusing cake mass and cake thickness to calculate cake porosity [8]was not relevant especially for cakes of low thickness. Indeed, twodrawbacks should be mentioned: (i) the germination region andthe superficial region of the cake are not taken into account and(ii) the calculation depends on the definition of the cake thickness.This is particularly true in the present case of the filtration ofbidispersed suspensions. As mentioned by other authors [18],three different regions in the cake have been demonstrated (see

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z (μ

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αs

3.31 μg 6.92 μg 10.33 μg 14.12 μg 17.37 μg

Fig. 8. Evolution of the fraction of the area occupied by the particles in each imagealong the z axis vs. total cake mass during experiment E3.

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αs

2.74μg 0.57μg

8.55μg 1.78μg

14.38μg 2.99μg

Fig. 9. Evolution of the fraction of the area occupied by each type of particles ineach image along the z axis vs. cake mass: 1 mm particles: (empty symbols), 4.8 mmparticles: (full symbols); total cake mass (◆, ◇) 3.31 mg, (▲, △) 10.33 mg, (●, ○)17.37 mg.

Fig. 10. Composite images of microfiltration cake formed by bidisperse suspension; total cake mass: (A): 3.36 mg, (B): 6.01 mg, (C): 10.83 mg (Experiment E4). (Forinterpretation of the references to color in this figure, the reader is referred to the web version of this article.)

I. Ben Hassan et al. / Journal of Membrane Science 466 (2014) 103–113 109

Section 1 and [13]). Here, these three regions are still wellillustrated in Fig. 12. It can be noticed that the formation of thegermination region for experiment E4 is in good agreement withthe one previously discussed for experiment E3. Indeed, thez-variations of αS are similar during the first steps of filtration(mass deposited lower than 15 mg). In this first region, αS progres-sively increases as mass deposited increases to get 0.6, which isthe constant value associated with the central region of the cakeand that corresponds approximately to the void ratio of randompacking of spheres. From this filtration step, the cake consists ofthe three aforementioned regions: (i) the germination region ofabout 10 mm (twice the larger particle diameter), (ii) the highconcentration region where the porosity is about 0.4 (say 1–αS)and (iii) the superficial region of about 10 mm (twice the largerparticle diameter). This morphological analysis of the cake formedby bidispersed particles exhibits an analog result as the one foundin the case of the filtration of monodispersed particles [13].However, it should be mentioned that the value of 0.4 is anapproximate value of porosity because it is a hard task to performaccurate measurements of the area occupied by 1 mm particles.

Recently, numerous of non-invasive characterization techni-ques were developed in order to better understand the basicmechanisms governing particle deposition and following cakeformation. For instance, the laser based method allowed Mendretet al. to measure the local thickness of a clay cake on a planemembrane [9]. Furthermore, ultrasonic time domain reflectometry

(UDTR) was used by Li et al. to measure kaolin cake's thickness[19]. However, the internal cake characteristics could not bedetermined by classical characterization techniques. Compared toother methods, one of the main features of the CLSM is that itallows the 3D reconstruction of the cake fouling, thanks to Image J3D viewer. This tridimensional reconstruction helps analyze theshape of the deposit formed over the filtration stripes. In Fig. 13,three composite images made from the microsieve observation inbright field mode and the cake 3D reconstruction in fluorescencemode can be seen. It shows the topography of the cake formed ona 0.8 mm microsieve at different times of filtration associated withdifferent masses of particles deposited. These representations arethe most realistic views of the cake. It confirms the randomdistribution of the particles on the filtering stripes at the begin-ning of the run. At the end of filtration, all the filtration area iscovered.

3.4. Filtration of small particles after a first filtration of largeparticles (experiment E2)

The mechanisms that take place during the beginning ofmonodispersed particles cake formation were already discussedin Section 1, especially in the case of the filtration of 4.8 mmparticles. In experiment E2, a mass of 43.60 mg of 4.8 mm particleswas injected before the injection of 1 mm particles. Images of thecake obtained after this first filtration are given in Fig. 14. Accord-ingly, the z-variations of αS (fraction of area occupied by theparticles) show that the cake consists of approximately two layersof particles (Fig. 15 (full triangles)). The maximum value of αS

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0.8

0.9

1

1.1

0

0.2

0.4

0.6

0.8

1

0 1 2 3 4 5 6 7 8 9 10 11 12

Total cake mass (μg)

αcL p*

1μm particles mass (μg)

Fig. 11. Evolution of the apparent fraction of the microsieve filtration area coveredby particles, αc, (∎) and the permeability reduction (◆) of the 0.8 mm microsievefouled by a bidisperse suspension during experiment E4 particles vs. cake mass.

0

5

10

15

20

25

30

35

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70

z(μm

)

αs

3.36μg 8.50μg 10.84μg

15.16μg 20.64μg 23.78μg

Fig. 12. Evolution of the fraction of the area occupied by the particles in each imagealong the z-axis vs. total cake mass during experiment E4.

Fig. 13. Composite image of 3 D construction microfiltration bidispersed cake atcake mass (A) 3.36 mg and (B) 8.50 mg and (C) 16.96 mg (objective 63� , zoom 1)(Experiment E4).

I. Ben Hassan et al. / Journal of Membrane Science 466 (2014) 103–113110

associated with 43.60 mg of large particles deposited is a little bithigher than 0.5. This indicates that the germination region isalmost achieved. Only a few voids at the cake surface are stillpresent. Further large particle injections would lead to the growthand the thickening of the cake as already found [13].

Now let us examine what happens during the filtration of smallparticles on the cake formed by large particles. As can be observedin Fig. 14B and C, small particles first migrate towards the voids ofthe cake that they may cross because of their small size. Then, theytend to thicken the cake and some of them eventually reach themembrane surface. This is confirmed by the z-variations of αSplotted in Figs. 15 and 16. In particular the curves that correspondto the first steps of filtration of 1 mm particles (empty squares andtriangles) in Fig. 16 exhibit an approximate value of αS equal to0.03 at z¼0, which reveals that some small particles reach the

membrane surface during the first steps of the filtration of thesmall particle suspension. It can also be observed that, due tosmall particles, αS increases in the germination region. Indeed, thecake is thickened by small particle accumulation in the remainingvoid space. As the void spaces get filled, small particles accumulateat the surface of the cake formed by large particles during thefurther steps of filtration. They progressively smooth the cakesurface by filling the space between the large particles (Fig. 16C).It is well exemplified by the z-variations of αS (Fig. 15) associatedwith these steps of filtration. This confirmed that the surface of thecake becomes smoother. Finally, the cake is expected to grow dueto further accumulation of small particles and at this stage thecake obtained with the addition of the small particles should behomogenous. Unfortunately it was impossible to keep on continu-ing the experiment for validation because the transmembrane

Fig. 14. Series of composite images of microfiltration cake formed by the bidisperse suspension at cake mass (A) 43.6 mg, (B) 45.6 mg, (c) 57.8 mg during experiment E2;objective 63� , zoom 1 (1), zoom 6 xy and yz views (2,3), respectively.

I. Ben Hassan et al. / Journal of Membrane Science 466 (2014) 103–113 111

pressure was too high. It is noticeable that the overall maximumvalue of αS is of the order of 0.6, giving a porosity of 0.4 (Fig. 15).This value is the same as the one obtained in experiment E4 in thehigh concentration region of the cake. Moreover, αS becomesfinally equal to 0.6 on a range of z, presuming the beginning of ahomogeneous growth of the cake during the further steps offiltration.

4. Conclusion

In this work, the initiation and building of cakes formed duringthe filtration of bidisperse fluorescent particles suspensions werestudied. in situ analyses at the particle level were performedaccording to a previous described method based on CLSM.

During the filtration of bidisperse suspensions, the particledistribution over the membrane for the first cake layer and thecake build-up were investigated in terms of microsieve coverage,permeability reduction and porosity.

As the particles arrive at the membrane surface, the behavior offirst 4.8 mm and 1 mm particles is similar to the one witnessedduring the filtration of monodisperse suspensions. Moreover smallparticles seem to preferentially accumulate around the largeparticles deposited close to the place where pores surrendering

the blocked pores are still open and, then, the fluid can easily flow.Then, the scenario of multilayer cake formation was studied stepby step. The evolution of the apparent fraction of the microsievefiltration area covered by particles showed that 1 mm particlespredominately contribute to cake thickening and facilitate theprogressive cake homogenization.

Moreover, the shape of the area occupied by particles along thez-axis still demonstrated three regions with different concentra-tions: germination, central and superficial zone. The cake porosityin the central region was about 0.4.

Finally, the small particles were filtrated after the deposition ofa cake formed with the large ones. Small particles migration in thevoids of the cake formed by the larger particles is observed.

In future works, various issues will be addressed, especially theanalysis of the fouling cakes obtained during the filtration ofbiological suspensions using a mix of fluorescent microorganisms.On the other hand, the effect of the membrane properties on cakebuild-up, as suggested in the present paper, when two size cellsare mixed will also need to be better explored.

Acknowledgments

This work would not have been possible without the financialsupport of Action Intégrée Franco-Tunisienne du Ministère desaffaires étrangères et Européennes français et du ministère del'Enseignement Supérieur, de la recherche Scientifique tunisien.We thank Christophe ELLERO (LISBP) for his technical support andCecile POUZET and Aurélie LE RU from IFR40 confocal platform(Auzeville France) for their technical assistance.

Nomenclature

αc apparent fraction of filtration area covered byparticles

αs fraction of the area occupied by the particles ineach image along the z-axis

Td mean cake thickness (mm)Lcp water permeability at a given time of the filtration

experiment (L h�1 m�2 bar�1)L0p water permeability of the clean microsieve

(L h�1 m�2 bar�1)LnP permeability reductionJ permeate flux (m s�1)R hydraulic resistance to filtrate flow (m�1),ΔP transmembrane pressure (Pa)μ liquid viscosity (Pa s)ε cake porositySc apparent area of the surface occupied by the cake

on the image (mm2).Sm filtration area in the observed image (mm2).S(z) area of the surface occupied by the particles at each

image of the stackSf6 area of the filtrating area at zoom 6Mc cake mass (kg)MT total cake mass (kg)N1 number of 1 mm particles deposited in experiment

E1N3 number of 1 mm particles deposited in experiment

E3N4 number of 1 mm particles deposited in experiment

E4ρc particles density (kg m�3)

0

5

10

15

20

25

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70

z (μ

m)

α s

7.89μg43.60μg45.59μg ( 43.60+1.99)47.12μg (43.60+3.52)50.89μg (43.60+7.29)52.39μg (43.60+8.79)54.07μg (43.60+10.47)57.33μg (43.60+13.73)

Fig. 15. Evolution of the fraction of the area occupied by the particles along the z-axis vs. cake mass during the filtration of monodisperse suspension of 4.8 mmparticles (full symbols) and bidisperse suspensions (empty symbols).

0

5

10

15

20

25

0.00 0.10 0.20 0.30 0.40 0.50 0.60

z (μ

m)

α s

43.60μg 1.99μg 3.52μg 7.29μg 8.79μg 10.47μg 13.73μg

Fig. 16. Evolution of the fraction of the area occupied by each type of particlesalong the z-axis vs. cake mass of monodisperse suspensions: 4.8 mm particles: (fullsymbols), 1 mm particles: (empty symbols).

I. Ben Hassan et al. / Journal of Membrane Science 466 (2014) 103–113112

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