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Chapitre 7: Effect of pH and Electrodialysis with Ultrafiltration Membrane (EDUF) Configuration on the Electromigration rate of Chitosan Oligomers

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Les résultats présentés au chapitre 6 ont démontré l’efficacité de l’électrodialyse avec des membranes d’ultrafiltration à séparer un mélange d’oligomères de chitosane. Basée sur les résultats obtenus au chapitre 6, une membrane d’ultrafiltration avec un seuil de coupure (MWCO) de 10000 Da a été sélectionnée pour la suite du projet. Dans cette partie de la thèse, l’effet du pH du milieu et du temps de traitement sur le taux d’électromigration des oligomères de chitosan et la possibilité de les séparer ont été étudiés.

La planification de ce travail et la rédaction de l’article ont été réalisées par l’étudiant Mohammed Aider qui est aussi auteur principal. Il a été supervisé avec beaucoup d’attention par le Dr. Laurent Bazinet qui a contribué au succès de ce travail par sa grande implication. Serge Brunet de ISM Biopolymer a contribué au succès de cet article par ses corrections et commentaires. Ils sont co-auteurs. Les résultats présentés dans ce chapitre sont en attente de soumission dans Journal of Membrane Science.

Dans cette partie, l'effet du pH, de la configuration de la cellule d'électrodialyse avec membrane ultrafiltration (EDUF), de la durée de traitement aussi bien que le degré de polymérisation des oligomères de chitosane sur le taux d'électromigration de chaque oligomère ont été étudiés. Une membrane d’ultrafiltration de 10000 Da a été utilisée. Les résultats ont montré que la configuration de la cellule avait un effet sur la possibilité de séparation des oligomères. Le taux d'électromigration de chaque oligomère de chitosane a diminué en augmentant le pH. La durée de traitement combinée avec le pH avait un effet sur la possibilité de séparation des oligomères en raison des différences de leurs mobilités électrophorétiques. Le dimère a montré le taux d'électromigration le plus élevé à toutes les valeurs de pH, suivis du trimère et du tétramère. Dans ce travail, en fonction du temps de traitement, il a été possible de produire un mélange composé du dimère et de trimère à pH 6, ou seulement une solution de dimère à pH 7. Il n'y avait aucune électromigration des oligomères de chitosane aux pH 8 et 9. Ce résultat était en conformité avec le résultat obtenu dans le deuxième sous objectif du volet fondamental.

Interest in chitosan oligomers of low molecular weights is growing fast because of their potential use as bioactive ingredients. Effect of pH, electrodialysis with ultrafiltration membrane (EDUF) cell configuration, processing time as well as chitosan oligomer chain length on electro-migration rate of chitosan oligomers through 10000 Da MWCO UF-membrane and their possible electro-separation was studied. It was shown that cell configuration influenced the possibility of separation of the oligomers, chitosan oligomer electro-migration rate decreased by increasing the pH of the medium, processing time influenced possibility of separation of the oligomers because of the differences of their electrophoretic mobilities. Hence, the dimer showed the highest electro-migration rate at all pH values, followed by trimer, and then by tetramer. It was shown that it was possible to separate dimer mixed with trimer at pH 6 or only the dimer at pH 7 according to the processing time. There was no electro-migration at pH 8 and 9.

Keywords: Chitosan oligomers; Dimer; Trimer; Tetramer; Electromigration; Electroseparation; Electrodialysis; Ultrafiltration.

7.1. Introduction

Chitosan D-glucosamine and its oligomers, especially dimers, trimers, tetramers, pentamers and hexamers, have recently attracted much attention because of their biological and physiological activities, such as antibacterial activity (Ming et al., 2006; Jeon et al., 2001; Jeon & Kim, 2000a-c), antitumor activity (Tokoro et al., 1988; Suzuki et al., 1986) and immunoenhancing effects (Hirano et al., 1991). They could be used also as substances for stimulation growth of Bifidobacterium biffidum and Lactobacillus sp as reported by Lee et al. (2002). Generally, chitosan D-glucosamine and oligosaccharides are produced by complete or partial hydrolysis of chitosan (Fukamizo et al., 1994; Izume & Ohtakara, 1987). Chitosan oligomers (from dimers to hexamers) are obtained as reaction intermediate products. Mainly, two methods are used for hydrolysis of chitosan: chemical and enzymatic hydrolyses (Akiyama et al., 1995). In the case of chemical hydrolysis, the reaction is performed under high temperatures and highly acidic medium. This hydrolysis reaction led to produce a large amount of chitosan D-glucosamine (chitosan monomer). Researchers reported difficulties in controlling the progress of the reaction (Akiyama et al., 1995) and the final product is a mixture of chitosan oligomers of different molecular weights (Li et al., 2005). In the other hand, enzymatic hydrolysis of chitosan although allowing a better control of the oligomers molecular weights, it remains that the end product is also a mixture of various fractions of different molecular weights (Kim & Rajapaksea, 2005; Kuroiwa et al., 2003; Kuroiwa et al., 2002).

Recently, it was shown that it is possible to separate chitosan oligomers using electrodialysis with ultrafiltration (EDUF) process. EDUF is a hybrid electroseparation process, which combines conventional electrodialysis with ultrafiltration membranes (Aider et al., 2007a; Labbé & Bazinet, 2006; Poulin et al., 2006a; Bazinet et al., 2005a, b; Labbé et al., 2005). The ultrafiltration membrane is stacked as molecular barrier in the electrodialysis cell and the driving force of the process is an applied external electric field. Various ultrafiltration membranes with different molecular weight cut-offs (MWCO) were tested with an aim of studying the effect of the UF-membranes MWCO on separation of chitosan oligomers (Aider et al., 2007a). It was shown that the MWCO had a significant effect on the chitosan oligomers electromigration rates but also on the nature of the migrated molecules. It was shown that electro-separation duration time has significant effect on electro-migration kinetic of each migrated chitosan oligomer (Aider et al., 2007a).

Chitosan oligomers contain one or more amine groups in their chain which can carry an electric charge and some pH conditions. Electrophoretic mobility is one of the most important behaviors of chitosan oligomers. This characteristic depends on the oligomers degree of polymerization, ionic strength and pH of the medium. Effect of pH is directly related to the protonation/deprotonation phenomenon of the amine groups. In acidic media, the amine group is highly protonated and the degree of protonation decreases by increasing pH. Recently, electromigration behavior of chitosan D-glucosamine and oligomers with degree of polymerization from 2 to 6 in dilute aqueous systems containing either NaCl and KCl salts at 0.01, 0.05 and 0.1 M at pH values from 2 to 9 was evaluated. The results showed that the electromigration of the chitosan D-glucosamine and oligomers did not change by changing the type of salt in the running medium and that the pH had a significant effect on the direction of migration under external electric field. In addition, the increase in the ionic strength of the medium caused a significant decrease on the absolute value of the electrophoretic mobility and the highest values of the electromobility were observed in water (Aider et al., 2006a). In other study, the effect of the concentration of a chitosan oligomer mixture on its electrophoretic behavior was studied as a function of pH and ionic strength. It was shown that the concentration and pH value have significant effect on the average electrophoretic mobility of the chitosan oligomers mixture. At a concentration of 3%, the ionic strength did not show any effect on the electromigration behavior of the chitosan oligomers mixture. By decreasing the concentration of the chitosan oligomers mixture, ionic strength showed a significant effect on the average electrophoretic mobility (Aider et al., 2006a,b).

Considering that potential applications of chitosan oligomers require pure or highly enriched fractions of defined molecular weight, it appears important to develop more effective means for production on a large scale of these bioactive molecules. In this context, the aim of this study was to investigate the effect of pH and EDUF cell configuration on chitosan oligomers electroseparation performances.

A MicroFlow type electrodialysis cell (ElectroCell AB, Täby, Sweden) with an effective area of 10 cm2 was used. The EDUF cell configurations [Anode-AEM1-MUF-AEM2-Cathode] and [Cathode-AEM1-MUF-AEM2-Anode] defines three closed compartments (Figures 7. 1 and 7.2).

The first compartment was for NaCl aqueous solution (20 g/L) as electrode rinse solution. The second was for the chitosan oligomer aqueous solution and the third one for KCl aqueous solution (2 g/L). Each compartment was connected to a separate external reservoir to allow recirculation of the solution. The solutions were circulated using three centrifugal pumps and the flow rates in each compartment were controlled by flow meters. The anode was a dimensionally stable electrode (DSA) to oxygen and the cathode was a 316 stainless steel electrode. For all experiments, the anode/cathode voltage difference was supplied by a variable 0–30V power source (HPD 30-10SX, Xantrex, Burnaby, BC, Canada). Once the EDUF cell was assembled, electrodes spacing was 2.0 ± 0.1 cm.

Electroseparation of chitosan oligomers treatments with EDUF system were carried-out on 200 ml of chitosan oligomer solution in a batch process, in both cell configurations at 6 pH values (4, 5, 6, 7, 8 and 9). These pH values were selected following our previous work on the effect of the pH value on chitosan oligomer electrophoretic mobility (Aider et al., 2006a,b). A constant voltage of 5 V was used as previously used by Aider et al. (2007a). The chitosan oligomer solution was obtained by dissolving chitosan oligomer mixture in HPLC grade water to obtain a final concentration of 3%. Its initial pH was adjusted to the desired values by adding 1 N HCl or NaOH. The pH of the 200 ml KCl solution in the adjacent compartment was also adjusted and was the same one as in the chitosan oligomer solution compartment. Samples of chitosan oligomer and KCl solutions were collected at the beginning of the process before applying the external electric field and every 60 min during the 4 h-treatment. During treatment, pH and conductivity values were recorded in the KCl and chitosan oligomer solutions. Measurements of the current intensity were used for the calculation of the total system resistance. Thickness and electrical conductivity of the membranes were measured before and after each treatment to check for their integrity and verify their potential fouling. All samples were analyzed by HPLC to determine the electromigration rate of the three chitosan oligomers. Current efficiency is an important parameter for assessing the suitability of any electroseparation process for practical application. The overall current efficiency for the studied chitosan oligomers (dimer, trimer and tetramer), was defined as the current carried by each oligomer relative to the total electricity used during the EDUF treatment.

The pH of the medium, processing time, chitosan oligomer chain length as well as EDUF cell configuration had significant effects on chitosan oligomer electromigration rates (P<0.001).

According to configuration-1 in the mode [Anode-MEA-MUF-MEA-Cathode], Figures ( 7. 3- 7. 5) show the evolution of the electromigration rates of chitosan oligomers through 10000 Da MWCO UF-membrane EDUF system as function of time under different pH conditions of the medium. For all pH values, the dimer showed the highest electromigration rates compared to the others chitosan oligomers (Figure 7. 1). Moreover, by increasing pH of the medium, electromigration rate of the oligomers decreased significantly.

At pH 4, the dimer showed a linear increase of its electromigration rate in time with average values of 2.47 ± 0.99% at τ =1h and 11.50 ± 4.34% at 4h of treatment. These data are in good agreement with those obtained in our previous work under the same conditions (Aider et al., 2007). At pH 5, the dimer showed similar behaviors as those recorded at pH 4. By increasing the pH up to 6, at τ = 1 and 2h, dimer showed electromigration rates similar to those recorded at pH 4 and 5. But, at τ = 3 and 4 h of electroseparation treatment, dimer electromigration rates were significantly lower than those recorded at pH 4 and 5. By increasing pH of the medium up to 7, dimer continued to migrate through the UF-membrane. At τ = 1 and 2h, it was the only one oligomer which crossed the UF-membrane with average electro-migration rates of 1.63 ± 0.81 and 3.78 ± 1.46%, respectively.

Electromigration behavior of the trimer as function of time and pH is shown in Figure 7. 4. At pH 4, trimer showed electromigration rates which varied between 1.47 ± 0.66 and 8.53 ± 1.45% at τ = 1 and 4h, respectively. At pH 5, the trimer showed similar electromigration rates as those recorded at pH 4. By increasing pH of the medium up to 6, the trimer showed electromigration rates similar to those recorded at pH 4 and 5 at only between τ =1 and 3h. At the end of the electroseparation process (τ = 4h), trimer showed an electromigration rate different from those recorded at pH 4 and 5 corresponded to the same time. By increasing pH up to 7, trimer had migrated through the 10000 Da MWCO UF-membrane used only after 3 and 4h of treatment with low electromigration rates at with average values of 0.67 ± 0.14 and 0.84 ± 0.43%, respectively.

Tetramer electromigration rates are shown in Figure 7. 5 as function of time and solution pH values. At pH 4, it migrated through the 10000 Da MWCO UF-membrane only after 3h of treatment with average electromigration rates of 2.62 ± 0.34 and 4.26 ± 0.31%, respectively, for τ = 3 and 4h. At pH 5, tetramer migrated through the ultrafiltration membrane only after 4h of treatment and at very low electromigration rate with an average value of 0.32 ± 0.55%. By increasing the pH up to 6 and 7, the tetramer did not migrate through the 10000 Da MWCO ultrafiltration membrane used.

In all cases, by increasing pH up to 8 and 9, no oligomer had migrated through the 10000 Da MWCO UF-membrane.

EDUF cell configuration-2 in the mode [Cathode-AEM-UFM-AEM-Anode] (Figure 7. 2) was used in order to verify if electromigration of oligomers through 10000 Da MWCO UF-membrane is possible towards the anode in particular at pH between 7 and 9, where it was previously demonstrated that some chitosan oligomers have a small negative charge and thus could migrate towards the cathode (Aider et al., 2006a,b). The same pH conditions as those tested for configuration-1 [Anode-AEM-UFM-AEM-Cathode] were carried out with configuration-2 (Figure 7. 2) to allow chitosan oligomers electro-migration towards the anode. HPLC analyses of samples from KCl solution did not show any presence of chitosan oligomers. So, using a concentration of 3% of initial chitosan oligomers solution and with a configuration-2 [Cathode-AEM-UFM-AEM-Anode], chitosan oligomers electromigration was not possible in 4 h of EDUF treatment.

The effect of the pH on chitosan oligomers electromigration through 10000 Da MWCO UF-membrane is directly related to the protonation and deprotonation phenomenon of the chitosan oligomers amine group, which is pH dependent. Chitosan oligomer electrophoretic mobility depends on the degree of protonation of the amine group. At pH values 4 and 5, the oligomers showed the highest electromigration rates because of the high protonation level of the amine functional group. These results are in good agreement with data reported by Aider et al. (2006) where it was shown that in the pH range between 4 and 6, there is no significant difference between average electrophoretic mobilities of chitosan oligomer mixture which is the same one as that used in the present study (Aider et al., 2006a). By increasing pH of the medium up to 7, electromigration rate of all chitosan oligomers decreased significantly compared with data recorded at pH between 4 and 6. This was caused by high deprotonation level (Juang & Hao, 2002)of the amine function, which had as consequence to decrease the electrophoretic mobility of the oligomers. Electrophoretic mobility is an intrinsic characteristic of each chitosan oligomer and the electromigration rate depends on this property. At pH 8 and 9 no oligomer had migrated through the 10000 Da MWCO UF-membrane because of the complete deprotonation of the amine function. This result is also in good agreement with data reported by Aider et al. (2006) where it was found that at pH 8 and 9, electrophoretic mobility of chitosan oligomer mixture composed of dimer, trimer and tetramer is motionless. All these results recorded at different pH values show that the migration of chitosan oligomers through 10000 Da MWCO UF-membrane was caused by the effect of the electric field on charged amine groups and not by the diffusion. If diffusion acted as driving force, chitosan oligomer migration at pH 8 and 9 would have been possible.

The pH of the medium (P<0.001) and processing time (P<0.001) have significant effect on electrical conductivity in both chitosan oligomer mixture and KCl solutions during electromigration of chitosan oligomers in cell configuration-1 (Figures 7.6 and 7.7). Whatever the pH of the medium, the tendencies were the same ones. Taking processing time as independent variable of the electroseparation process, electric conductivity of chitosan oligomer solution decreased (Figure 7.6) with time, whereas in the KCl solution it increased (Figure 7.7). Differences between initial values of electrical conductivity of chitosan oligomer solutions were caused by the added quantity of HCl or NaOH (according to the case) necessary to readjust solution pH to the desired initial value (between 4 and 9). This phenomenon was not observed with the KCl solution. This could be explained by the buffering capacity of chitosan oligomer solution which required more HCl or NaOH to reach the desired initial pH value compared to KCl solution which has low buffering capacity because of its composition (only water and KCl). Data analyses showed that in the pH interval from 4 to 6 there was no significant difference between conductivity decrease in chitosan oligomer solution and its increase in KCl compartment in the interval of time between 0 and 4h of the process. Same results were found for pH range from 7 to 9. The differences did not exceed experimental error. All these observations could be attributed to the initial conditions of the electroseparation process. In both chitosan oligomer and KCl compartments pH had a same initial value. In a previous study where pH was readjusted only in the chitosan oligomer compartment, Aider et al. (2006) showed that the decrease of electric conductivity of chitosan oligomer solution was lower than its increase in the KCl compartment. It was explained that this phenomenon was caused by the migration of the H+ ions from chitosan oligomer solution towards KCl compartment and that they had contributed to increase the overall KCl solution electric conductivity. Mainly, this is due to the high electrophoretic mobility of H+ ions considered as the most mobile ion (Koneshan et al., 1998).

There was significant effect of pH (P<0.001), processing time (P<0.001), interaction pH*treatment time (P*τ) (P<0.001) on the global EDUF system electric resistance.

Figure 7.8 shows the evolution of the global system electric resistance at different pH values as a function of processing time, according to configuration-1 [Anode-AEM-UFM-AEM-Cathode] (Figure 7.1). In general, the tendency was the same one in the range of the studied pH values. Global system electric resistance decreased with time. At all pH values, during first 30 min system electrical resistance decreased drastically, thereafter, the

decrease was less important during the following 30 min. Thereafter, throughout the electroseparation process, the global system electric resistance continued to decrease but with a weak slope. EDUF system exploited at pH 5 showed the highest electric resistance in comparison with the other pH values. Systems exploited at pH 4, 6 and up showed identical behavior. The EDUF system with pH 9 showed the lowest global electric resistance compared to all other EDUF systems.

All these results could be explained as follows: in all cases, HCl or NaOH was used to adjust pH to the desired value. Contribution of H+ and OH- ions to increase solution electrical conductivity (as consequence to decrease its electric resistance) is higher than any other ion. However, H+ is the most mobile ion followed by OH- ion (H+ twice more mobile) (Koneshan et al., 1998). So, EDUF system with pH 5 showed the highest electric resistance because the initial chitosan oligomer solution had pH 5.4 ± 0.1, and to adjust pH solution up to a value of 5, less quantity of HCl (source of ion H+) was necessary comparatively to that used to decrease pH down to 4. Quantity of hydrogen ions added in the case of pH 4 was more important than that added to obtain pH 5 and as consequence, contribution of hydrogen ions to increase electric conductivity of the solution at pH 4 was more important compared to the case with pH 5. As result, electric resistance at pH 5 was the highest one. For the other EDUF systems at the other pH values, initial electric resistance was also lower then that at pH 5 because different quantities of NaOH (source of ion OH-) were added to adjust the pH up to desired values (6, 7, 8 and 9). Less quantity of NaOH was needed to reach pH 6 compared with the quantities added to obtain pH 7, 8 and 9. This was the reason why the initial system electric resistance was higher at pH 6. The contribution of OH- ions to the overall system conductivity was more important in the case when pH values between 7 and 9 were used.

For the configuration-2, the increase of overall system electric resistance was caused mainly by the demineralization of the KCl compartment (decrease of electrical conductivity in this compartment).

For the configuration-2 (Figure 7. 2), in all cases, global system electrical resistance increased throughout the process. With this configuration, there was no electro-migration of chitosan oligomers through 10000 Da MWCO UF-membrane; therefore, data were not shown.

All these results could be explained as follows: in all cases, HCl or NaOH was used to adjust pH to the desired value. Contribution of H+ and OH- ions to increase solution electrical conductivity (as consequence to decrease its electric resistance) is higher than any other ion. However, H+ is the most mobile ion followed by OH- ion (twice more mobile) (Koneshan et al., 1998). So, EDUF system with pH 5 showed the highest electric resistance because the initial chitosan oligomer solution had pH 5.4 ± 0.1, and to adjust pH solution up to a value of 5, less quantity of HCl (source of ion H+) was necessary comparatively to that used to decrease pH down to 4. Quantity of hydrogen ions added in the case of pH 4 was more important than that added to obtain pH 5 and as consequence, contribution of hydrogen ions to increase electric conductivity of the solution at pH 4 was more important compared to the case with pH 5. As result, electric resistance at pH 5 was the highest one. For the other EDUF systems at the other pH values, initial electric resistance was also lower then that at pH 5 because different quantities of NaOH (source of ion OH-) were added to adjust the pH up to desired values (6, 7, 8 and 9). Less quantity of NaOH was needed to reach pH 6 compared with the quantities added to obtain pH 7, 8 and 9. This was the reason why the initial system electric resistance was higher at pH 6. The contribution of OH- ions to the overall system conductivity was more important in the case when pH values between 7 and 9 were used.

For the configuration-2, the increase of overall system electric resistance was caused mainly by the demineralization of the KCl compartment (decrease of electrical conductivity in this compartment).

Figure 7. 9 shows the current efficiency for each chitosan oligomer as function of pH. Data showed that the current efficiency η was very low. At all pH values, the dimer had the highest current efficiency. At pH 4 and 5, no significant difference was found between the current efficiencies (P>0.126) with an average value that did not exceed 9%. By increasing the pH of the medium, current efficiency decreased and reached zero value at pH 8 and 9 because of the null electrophoretic mobility. The current efficiency related to the trimer had a general behavior similar to that of the dimer but with lower values at each corresponding pH. No significant difference was found between the η values at pH 4 and 5. A decrease of the current efficiency was observed by increasing pH of the medium. No efficiency was recorded at pH 8 and 9. The tetramer showed the lowest current efficiency and was equal to zero at pH 6.

These low values of the current efficiency η could be explained by the relatively low electrophoretic mobilities of the chitosan oligomers in comparison with the minerals present in the solution and by UF-membrane resistance to the oligomers migration. The major part of the electric current was carried by the minerals because of their high mobilities. Current efficiency values of 5.996, 0.336, 4.739, 0.037 and 42.140% for butyric, valeric, adipic, caproic and oxalic acids, respectively, were reported (Nagarale et al., 2006; Wang et al., 2006). The particularity of their work was the use of anion exchange membrane as a separation barrier whereas in our case, an UF-membrane was used. In the other hand, current efficiencies for chlorogenic acid, scopoletin and rutin of 0.019, 0.000 and 0.062%, respectively, were reported by Bazinet et al. (2005b). In their work, an ultrafiltration membrane of 500 Da MWCO was used.

Comparison of the 10000 Da MWCO ultrafiltration membrane thickness, electric conductivity before and after each electrodialysis with ultrafiltration membrane (EDUF) operation according to pH of the EDUF process and cell configuration were used for membrane fouling evaluation (Tables 7.1 and 7.2). There was no significant difference between membrane electric conductivity and thickness before and after EDUF treatments (P > 0.05) independently of pH or cell configuration. All differences did not exceed experimental error. These results indicate that there was no (or weak) membrane fouling which could be removed by rinsing membrane after each operation or by storing in distilled water between EDUF runs. Same results were found for the anion exchange membranes used during the EDUF treatments. Results of comparison of their characteristics (membrane thickness and electrical conductivity) before and after 4 h of electroseparation treatment (Table 7.2) showed that no significant difference was detected (P>0.129). Before using this membranes, (τ = 0 h), the average electrical conductivity was κ = 4.645 ± 0.121 ms.cm-1, and thickness l = 0.131 ± 0.001 mm. After 4h of treatment, the average electrical conductivity was κ= 4.501 ± 0.149 ms.cm-1, and membrane thickness l = 0.130 ± 0.001 mm. These results indicated that there was no fouling of the anion exchange membranes and that they would keep their full integrity. These results are in good agreement with those reported by Poulin et al. (2007) and Aider et al. (2007a).

This study showed that the electromigration rate of chitosan oligomers of low molecular weight (dimer, trimer and tetramer) migrate through a 10000 Da MWCO UF-membrane was different dependently on pH of the medium and processing time. The highest electromigration rate was obtained in the pH range of 4 and 6. Effect of pH was related to the protonation/deprotonation phenomenon of chitosan oligomer amine function. This protonation/deprotonation behavior directly influenced the electrophoretic mobility of each oligomer and consequently the separation of some fractions through retardation of the electromigration of other molecules. Hence, it was possible to obtain pure solution of dimer at pH 7 after 2h because it was the only chitosan oligomer which migrated through the 10000 Da MWCO UF-membrane, or to produce fraction composed only by the dimer and trimer without tetramer at pH 6 after 2 hours of electroseparation. The results showed also that it is possible to separate mixture of chitosan oligomers of low molecular weight by combination of pH and electroseparation duration.

Absence of chitosan oligomer electromigration at pH 8 and 9, and in the [Cathode-AEM1-MUF-AEM2-Anode] cell configuration showed that the passage of the oligomers through 10000 Da MWCO UF-membrane was caused by the effect of the applied external electric field on the charged molecules and not to the diffusion.

Further investigations are necessary to understand effect of hydrodynamic conditions and/or electric field on electromigration of chitosan oligomers and possibility of their separation.

© Mohammed Aider, 2007