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Flocculation of Model Latex Particles by Chitosans of Varying Degrees of Acetylation Matthew Ashmore and John Hearn* Department of Chemistry and Physics, Nottingham Trent University, Clifton Lane, Nottingham NG11 8NS, England Received December 17, 1999. In Final Form: February 29, 2000 A series of chitosan samples spanning a wide range of molecular weights have been homogeneously acetylated to varying degrees and characterized. These have been used to flocculate a well-characterized “model colloid” polymer latex. The optimum flocculation concentrations have been identified using both residual turbidity and initial-rate methods. A charge neutralization mechanism with an enhancement of rate by a “charge patch” effect is shown to operate while both degree of acetylation and molecular weight can have an effect upon the optimum flocculation concentration.
Introduction Chitosan is the general name given to a continuum of copolymers of D-glucosamine and N-acetyl-D-glucosamine (Figure 1), which are soluble in dilute acid solution.1 They are the main derivatives of chitin (almost exclusively the N-acetyl-D-glucosamine polymer), which is the second most abundant polysaccharide in nature (following cellulose), being an important structural component of the exoskeletons of crustacea, mollusks, and insects. Chitin is insoluble in the usual aqueous and organic solvents but upon alkaline deacetylation yields the dilute acid-soluble cationic polyelectrolyte chitosan. It is becoming common practice to express the extent of acetylation of chitosan as the mole fraction acetylated, FA; e.g., FA ) 0.02 for 2% acetylated chitosan or 98% deacetylated chitin and would be written as chitosan[0.02]. Although chitosan does occur in nature, for example, in fungi, it is commercially produced from the chitin waste products of the seafood (crab, lobster, squid, etc.) processing industries.2 The fact that chitin is a renewable resource and chitosan is biodegradable,1 is nontoxic, and is an approved food additive3 means that they are environmentally friendly commodities, in contrast to synthetic polymers derived from petrochemical feedstocks.4 Research into applications for chitosan has grown phenomenally to cover diverse areas such as pharmaceuticals, biomaterials, cosmetics, food processing, and chelation of heavy metals.5 Chitosan is used as a flocculant on an industrial scale in activated sludge plants and in recovering proteins from food-processing factories.6 It has also been used in potable water applications,7 where it has the advantage over synthetic polymers whose use may be limited by levels of residual toxic monomers. Previously published studies of flocculation using chitosan have not always involved well-defined systems, e.g., where actual wastewater samples have been used8,9 or chitosan was characterized (1) Roberts, G. A. F. Chitin Chemistry; Macmillan Press: London, 1992; p 204. (2) Sandford, P. A. In Chitin & Chitosan; Skjak-Braek, G., Anthonsen, T., Sandford, P. A., Eds.; Elsevier Applied Science: London, 1989; p 51. (3) Hwang, D. C.; Damodaran, S. J. Agric. Food Chem. 1995, 43 (1), 33-37. (4) Kawamura, S. J. Am. Water Works Assoc. 1991, 83 (10), 88-91. (5) Roberts, G. A. F. Adv. Chitin Sci. 1997, 2, 22. (6) Thome, J. P.; Thys, I.; Hugla, J. L.; Patry, J.; Weltrowski, J. Chitin World 1994. (7) Huang, C., Chen, Y. J. Chem. Technol. Biotechnol. 1996, 66, 227232.
Figure 1. Repeat unit structures of chitin and chitosan including potential intramolecular hydrogen bonding.
by source rather than physicochemical characteristics.10 Studies have variously concluded that the predominant mechanism of flocculation occurred by bridging11 or that surface charge density effects were significant12 or unimportant.7,13 Some have found pH to be unimportant,7,14 while others have found molecular weight to be very significant.15 The aim of this paper is to report results from a model system in which both components are well characterized with a view to elucidating the mechanism of flocculation. Thus latex particles prepared by surfactantfree emulsion polymerization techniques having certified (8) Wu, A. C. M.; Brough, W. A. Proc. Int. Conf. Chitin/Chitosan, 1st 1977, 88. (9) Ganjidoust, H.; Tatsumi, K.; Yamagishi, T.; Gholian, R. N. Water Sci. Technol. 1997, 35 (2, 3), 291-296. (10) Castellanos-Perez, N.; Maldonado-Vega, A.; Fernadez-Villagomez, G.; Caffarel-Mendez, S. In ref 2, p 567. (11) Zhou, Y.; Huqun, L.; Shoucheng, D.; Weixia, Z.; Gonghui, Z. Wuli Huaxue Xuebao 1993, 9 (1), 77-84. (12) Agervist, I. Colloids Surf. 1992, 69 (2-3), 173-187. (13) Ordolff, D. Kiel. Milchwirtsch. Forshungsber. 1995, 47 (4), 339346. (14) Pinotti, A.; Bevilacqua, A.; Zaritzky, N. J. Food Eng. 1997, 32 (1), 69-81. (15) Domard, A.; Rinaudo, M.; Terrassin, C. J. Appl. Polym. Sci. 1989, 38, 1799-1806.
10.1021/la991648w CCC: $19.00 © 2000 American Chemical Society Published on Web 04/27/2000
Flocculation of Latex Particles by Chitosans
surface charges of strong acid sulfate groups were chosen as the disperse phase. Chitosans of varying degrees of acetylation were prepared by homogeneous reacetylation of a highly deacetylated sample so as to produce random copolymers (heterogeneous deacetylation tends to produce block sequences), and these were characterized with regard to the degree of acetylation and intrinsic viscosity. Experimental Section Materials. Chitosan samples designated as being “oligomeric”, “medium molecular weight”, and “high molecular weight”, subsequently coded O, M, and H, respectively, were kindly supplied by Professor G.A.F. Roberts, Department of Fashions and Textiles, Nottingham Trent University. A 400 nm surfactantfree polystyrene latex with a certified surface charge density (Product No. 1-400; Batch No. 2-63-46.186,1) was obtained from the Interfacial Dynamics Corp., Portland, OR. All water used was doubly distilled from an all-Pyrex apparatus. Acetic acid was of GPR grade (99.5%) supplied by BDH, Poole, Dorset, U.K. SLR grade sodium chloride (99.6% after drying), AR grade methanol (>99%), SLR grade ammonia (0.88, 35% NH3), SLR grade sodium acetate (>99%), and AR grade cetyltrimethylammonium bromide (CTAB) (>99%) were supplied by Fisher Scientific, Loughborough, Leicestershire, U.K. Acetic anhydride (>98%) was supplied by Sigma Aldrich, Poole, Dorset, U.K. When solutions of cationic materials were prepared and used, all vessels were conditioned to allow for losses by adsorption onto the walls;16,17 i.e., the vessels were first equilibrated with a solution of the desired concentration, which was then discarded and replaced with a new solution at the desired concentration. Preparation of Chitosans. The medium molecular weight and high molecular weight chitosan samples were prepared as follows: The starting material was initially dissolved in 0.1 M acetic acid, and the mixture was filtered through polyester monofilament to remove insoluble impurities. The material was then precipitated by addition of ammonia to pH 8. The precipitated chitosan was then washed with doubly distilled water until neutral to litmus, steeped in methanol overnight, and, after further filtration, dried under vacuum at 40 °C. The oligomeric chitosan, supplied as the hydrochloride salt, was neutralized in methanolic ammonia, washed with water and then with methanol, and dried under vacuum at 40 °C. Homogeneous N-Acetylation of Chitosans. A 98% deacetylated sample of starting material (4 g) was placed in a 1 L stoppered conical flask and then dissolved in 400 mL of 0.1 M acetic acid. A methanolic solution of acetic anhydride was prepared such that 10 mL contained a 1:0.1 molar ratio of chitosan amine groups to acetic anhydride. The required volume of acetic anhydride solution was pipetted into a 500 mL measuring cylinder and made up to 400 mL with methanol. This solution was added to the chitosan solution with continuous stirring, and the resultant solution was stirred for a further 20 min before being transferred to a water bath at 25 °C, where it remained for 24 h. After removal from the water bath, the chitosan was reprecipitated by addition of ammonia to pH 8. The material was then filtered off through polyester monofilament, washed with water until neutral to litmus, and steeped in methanol before final filtration and drying under vacuum at 40 °C. Chitosans with degrees of acetylation between 2% and 57% were prepared by this method. Characterization of the Chitosans. Determination of N-Acetyl Content. The method for determining N-acetyl content by chitosan adsorption of dye was based on the procedure developed by Maghami and Roberts:18 A 5 × 10-3 M (1.75 g L-1) solution of C.I. Acid Orange 7 [Orange II; 4-(2-hydroxy-1naphthylazo)benzenesulfonic acid, sodium salt] in 0.1 M acetic acid was prepared. Quantities of 0.05 g of chitosan were accurately weighed and placed in each of three clean, dry 250 mL Quickfit (16) Connor, P.; Ottewill, R. H. J. Colloid Interface Sci. 1971, 37 (3), 642-651. (17) Gregory, J.; Sheiham, I. Br. Polym. J. 1974, 6, 47-59. (18) Maghami, G. G.; Roberts, G. A. F. Makromol. Chem. 1988, 189, 2239-2243.
Langmuir, Vol. 16, No. 11, 2000 4907 conical flasks. A 100 mL portion of dye solution was pipetted into each flask plus a fourth, control flask containing no polymer. All flasks were then stoppered and placed in a water bath at 60 °C for 20 h. The contents of each flask were filtered through glass wool to remove the polymer residue, and the filtrates were allowed to cool. The amount of dye adsorbed by each chitosan sample was then determined, following appropriate dilution, by comparing the absorbance at 484 nm of the test solution with that of the control solution. Then, via the Beer-Lambert law, the N-acetyl content was assessed on the basis of the 1:1 stoichiometry between protonated amine groups and the negatively charged groups on the dye. Molecular Weight. The molecular weight of each chitosan sample was determined by viscometry to check that no degradation of the polymer occurred during reacetylation. The method followed was that described by Wang, Bo, Li, and Qin:19 A range of standard chitosan solutions were prepared in 0.2 M acetic acid/0.1 M sodium acetate solution. Viscosity measurements of the polymer solutions and pure solvent were carried out using an Ubbelohde viscometer at 30 ( 0.05 °C in a thermostated water bath. The flow times of the solutions permitted the intrinsic viscosity [η] to be determined for each solution. The intrinsic viscosity was then related to the molecular weight by use of the Mark-Houwink equation
[η] ) KMvR
(1)
where K and R are constants dependent upon the polymer, solvent, and temperature but independent of the molecular weight. K and R were calculated using the equations determined in the above study:
K ) 1.64 × 10-30(DD)14.0 -2
R ) -1.02 × 10 (DD) + 1.82
(2) (3)
where DD is the percentage degree of deacetylation of the chitosan sample. Flocculation Studies. Initial Rates. The kinetics of flocculation by polyelectrolytes shows three characteristic time scales: ta for polymer adsorption, tr for rearrangement of the adsorbed polymer, and tc for particle collision. While tr is of the order of a few seconds,20 ta and tc have been estimated using classical second-order reaction kinetics.21,22 To work in the equilibrium flocculation regime (tc . tr),20 so that physical adsorption and rearrangement precede flocculation, the particle number density should be kept below 1010/cm3 . Although absolute aggregation rates can only be derived for very small particles, monitoring the initial increase in the turbidity of an aggregating dispersion can provide useful data to compare the initial rates of coagulation or flocculation.23 The initial rates of increase in turbidity of 400 nm polystyrene latex upon aggregation were estimated by following the increase in absorbance at 550 nm using a UNICAM 8720 UV/vis spectrophotometer in the kinetics mode. A cuvette containing 2 mL of 500-fold diluted stock latex plus 0.5 mL of a modifier solution (to control pH, ionic strength, etc.) was placed in the spectrophotometer. A 0.5 mL quantity of coagulant or flocculant solution was added smoothly using a 500 µL syringe with a needle that had been ground flat at the end. Absorbance increment versus time curves were recorded over several minutes and subsequently plotted. The initial gradients of the curves, ∆A/dt were determined graphically. The ratio of the initial flocculation rate to the maximum initial coagulation rate was plotted against the polymer concentration, expressed in terms of the moles of chitosan monomer, and hence positive charge added. (19) Wang, W.; Bo, S.; Li, S.; Qin, W. Int. J. Biol. Macromol. 1991, 13, 281-285. (20) Pelssers, E. G. M.; Cohen Stuart, M. A.; Fleer, G. J. Colloids Surf. 1989, 38, 15. (21) Gregory, J. In The Effects of Polymers on Dispersion Properties; Tadros, Th. F., Ed.; Academic Press: London, 1982; p 301. (22) Gregory, J. Colloids Surf. 1988, 31, 231. (23) Gregory, J. In NATO Advanced Research Workshop on Nanoparticles in Solids and Solutions; NATO: Szeged, Hungary, 1996; p 37.
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Table 1. Extent of Acetylation of Chitosan Samples Determined by Dye Adsorption sample
mol wt
mole fraction
designation
oligomer 1 2 3 4 5 6 1H 2H
oligomer medium medium medium medium medium medium high high
0.021 ((1.07%) 0.019 ((1.32%) 0.081 ((1.52%) 0.141 ((1.47%) 0.240 ((2.26%) 0.329 ((1.56%) 0.570 ((3.57%) 0.172 ((0.52%) 0.382 ((1.64%)
FA[0.02]O FA[0.02]M FA[0.08]M FA[0.14]M FA[0.24]M FA[0.33]M FA[0.57]M FA[0.17]H FA[0.38]H
Table 2. Intrinsic Viscosities of Chitosans and Calculated Quantities sample
[η]/mL s-1
mol wt
DP
FA[0.02]O FA[0.02]M FA[0.08]M FA[0.14]M FA[0.24]M FA[0.33]M FA[0.17]H
6.05 611 570 573 474 836 932
1.90 × 103 5.27 × 105 5.32 × 105 6.03 × 105 8.07 × 105 1.92 × 106 1.12 × 106
12 3255 3237 3614 4615 10980 6661
Residual Turbidity. A 1 mL portion of the latex, previously diluted 500-fold as above, was pipetted into a 1.5 mL microcentrifuge tube together with 0.25 mL of the nappropriate modifier solution. A 0.25 mL quantity of flocculant solution was added smoothly using a 500 µL syringe, and the tube was capped, inverted twice slowly, and left overnight. Each tube was then centrifuged at 1000 rpm for 5 min. The supernate from each tube was transferred to a UV cuvette, and the absorbance of the solution was determined at 550 nm. Electrophoresis. A 0.5 mL quantity of latex, diluted 500-fold as before in distilled water, and 0.5 mL of 0.25 M acetic acid were pipetted into a 15 mL Pyrex phial. A 0.5 mL portion of chitosan solution was introduced by syringe, and 10 mL of 0.5 M acetic acid was added to stop any flocculation. Electrophoretic mobilities were determined using a Coulter Delsa 440SX ζ potential analyzer.
Results and Discussion Characterization. Chitosans. The mole fractions of acetylation of chitosan samples as determined by the dye adsorption technique are shown in Table 1 together with estimates of experimental error. Except for those of the highest degree of acetylation, errors fell below 2.5%. Table 2 shows the intrinsic viscosities of the chitosan samples. These were used in conjunction with K and R values, derived from the Wang equations (2) and (3) for varying degrees of acetylation, to calculate the molecular weights or degrees of polymerization. In Table 2, it can be seen that, as the degree of acetylation increases for a given chitosan specimen, the degree of polymerization (DP) also increases when it would have been expected to remain constant. It was for this reason, following an ultracentrifugation study in which DP did remain constant, that Wang recently24 concluded that none of the literature values of K and R for chitosans are reliable. As will be shown for our study, however, accurate quantitative values of the molecular weights are not in fact essential; rather, it is more important that a wide range of molecular weight samples were employed. Latex. The latex was supplied at a number density of 2.4 × 1012 particles/mL and a charge content of 5.0 µequiv/ g, with an error, estimated from conductometric titration data kindly supplied by Dr. J. Goodwin, Interfacial Dynamics Corp., to be (4%. It had been prepared by a surfactant-free emulsion polymerization technique25 and (24) Wang, W. Personal communication.
Figure 2. Rate of coagulation of 400 nm latex by CTAB relative to the rate of rapid coagulation (4) and residual absorbance ([) for varying charge ratio additions of CTAB.
was described26 as ultraclean following dialysis and to contain sulfate and hydroxyl surface groups with an insignificant number of carboxyl groups. It is known27 that sulfate-only latices can be prepared by using sodium bicarbonate buffers in conjuction with persulfate initiators. If such a reaction proceeds to high conversion of monomer to polymer, so as to avoid complications in the cleaning process,28 then a relatively short period of dialysis may be all that is needed to produce a clean model colloid latex. Without the use of an acid-washing step or the use of an ion-exchange resin, some of the counterions of the sulfate groups may be, e.g., sodium ions rather than protons. It has been suggested that functional groups on the end of polymer chains being repelled away from the latex surface form a “hairy” layer, causing, e.g., anomalous maxima in electrophoretic mobilities as a function of ionic strength, so that the particles are less than ideal model colloids.29 Other explanations have been proposed, including surface conductance effects or preferential ion adsorption,30 while Zhao et al.31 have described, on the basis of photon correlation spectroscopy data, the surface layer on particles of the type and surface charge density used here as being, in any case, relatively flat. In our work, the decision was made to use the particles as received rather than to, for example, heat them under pressure to above the glass transition temperature of polystyrene so as to desorb or adsorb the “hairy” chains.32 To do so would have risked sulfate group hydrolysis and latex instability together with a change in the certified surface charge density. Stability Studies. Typical results from both initialrate and residual turbidity studies are shown in Figures 2-6. The initial rates of flocculation are shown relative to the rate of rapid coagulation at a sodium chloride concentration of 1 M, i.e., well above the critical coagulation concentration of the latex of 0.5 M. Significant features of all the curves obtained are summarized in Table 3 with respect to the stoichiometric ratio of protonated amine groups, added as chitosan, relative to the anionic sulfate charge on the latex. The table lists the stoichiometric ratios (25) Goodwin, J. W.; Hearn, J.; Ho, C. C.; Ottewill, R. H. Br. Polym. J. 1973, 5, 347. (26) Product Guide 96/97; Interfacial Dynamics Corp.: Portland, OR, 1997. (27) Chainey, M.; Hearn, J.; Wilkinson, M. C. J. Colloid Interface Sci. 1987, 117 (2), 477-484. (28) Wilkinson, M. C.; Hearn, J.; Steward, P. A. Adv. Colloid Interface. Sci. 1999, 81, 77-165. (29) Seeburgh, J. E.; Berg, J. C. Colloids Surf., A 1995, 100, 139153. (30) Marlow, B. J.; Rowell, R. L. Langmuir 1991, 7, 2970-2980. (31) Zhao, J.; Brown, W. Langmuir 1996, 12, 1141-1148. (32) Bastos-Gonzalez, D.; Hildago-Alvarez, R.; De Las Nieves, F. J. J. Colloid Interface Sci. 1996, 177, 372-379.
Flocculation of Latex Particles by Chitosans
Figure 3. Rate of flocculation relative to the rate of rapid coagulation (4) and residual absorbance ([) for varying charge ratio additions of 2% acetylated oligomeric chitosan.
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Figure 6. Rate of flocculation relative to the rate of rapid coagulation (4) and residual absorbance ([) for varying charge ratio additions of 38% acetylated high molecular weight chitosan. Table 3. Critical Flocculation (and in the Case of CTAB Coagulation) Concentrations, in Terms of the Ratios of Amine to Sulfate Groups, for Each Sample
Figure 4. Rate of flocculation relative to the rate of rapid coagulation (4) and residual absorbance ([) for varying charge ratio additions of 2% acetylated medium molecular weight chitosan.
Figure 5. Flocculation ranges of 14% ([), 33% (4), and 57% (×) acetylated medium molecular weight chitosan determined by residual absorbance.
for the following features: (a) the maximum in the rate curve; (b) the minimum ratio at which significant flocculation occurs chosen consistently throughout at 0.5 absorbance unit; (c) the maximum ratio at which significant flocculation ceases chosen consistently throughout at 0.5 absorbance unit; (d) the center of the flocculation region, i.e., the mean of values from features b and c; (e) the breadth of the flocculation region at 0.5 absorbance unit; (f) the range of experimental errors at true 1:1 stoichiometry, allowing for error in the charge on the latex and in dispensing a given volume of latex together with the error in the measured degree of acetylation of the chitosan and its volumetric delivery. As an aid to discussing the results for chitosan, data obtained with the cationic surfactant CTAB are first considered. As has been demonstrated previously,16,33 it
sample
max rate
min conc
max conc
center
breadth
error range
CTAB FA[0.02]O FA[0.02]M FA[0.14]M FA[0.33]M FA[0.57]M FA[0.17]H FA[0.38]H
1.05 1.01 0.97 1.11 1.12 1.28 1.06 0.96
1.00 0.93 0.76 0.97 1.01 1.12 0.85 0.89
1.15 1.09 1.09 1.23 1.24 1.34 1.14 1.14
1.08 1.01 0.93 1.10 1.13 1.23 1.00 1.02
0.15 0.16 0.33 0.26 0.25 0.16 0.29 0.25
0.94-1.06 0.93-1.07 0.93-1.07 0.93-1.07 0.93-1.07 0.91-1.09 0.94-1.06 0.93-1.07
can be seen (Figure 2) that the latex is first destabilized by charge neutralization when cationic headgroups adsorb onto the anionic surface groups followed by restabilization with a positive charge as a consequence of adsorption via the hydrophobic tails. The maximum in the initial rate of flocculation falls within the error range for 1:1 stoichiometry, while the center point of the residual turbidity trough falls just outside that range. Earlier studies using CTAB16 have reported surface charge neutralization to occur at 1:1 stoichiometry as shown by zero electrophoretic mobility. The shapes of adsorption isotherms involving equilibrium aqueous-phase concentrations suggest that there would be some partition of the CTAB between the particle surface and the aqueous phase, and indeed a homologous series of alkyltrimethylammonium bromide surfactants showed a Traube’s rule effect. It might be expected then that the concentration of CTAB required for an optimum effect would be slightly above stoichiometric although exact neutrality may not be required since the latex with a low residual negative charge may only provide a small energy barrier to aggregation. A 5% enhancement of rate above that for rapid coagulation is observed, an effect which has been reported previously34 and ascribed to an increased hydrophobic effect as functional surface groups are neutralized and hydrophobic hydrocarbon tails are added to the surface. We now discuss results involving chitosan as the flocculant shown in Figures 3-6 and Table 3. As a polyion, its energy of adsorption onto an oppositely charged surface is high, and it would be expected to adsorb in a relatively flat configuration.35,36 Again, charge reversal is seen to occur, leading to restabilization. Such overcompensation of surface charges by polycations has been predicted (33) Zhao, J.; Brown, W. Langmuir 1995, 11, 2944-2950. (34) Gregory, J. Water Sci. Technol. 1993, 27 (10), 1-17. (35) Claesson, P. M.; Ninham, B. W. Langmuir 1992, 8, 1406-1412. (36) Kasper, D. R. Ph.D. Thesis, California Institute of Technology, 1971.
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theoretically,37,38 confirmed experimentally,39,40 and attributed41,42 to a lack of flexibility of the polyelectrolyte chains having more closely spaced charges than those on the particle. We can note that a knowledge of chitosan molecular weight is not essential in establishing the number of cationic charges added, since it is the molar ratio of D-glucosamine to N-acetyl-D-glucosamine in the sample that is needed and was established in the characterization section. For the 98% deacetylated oligomeric chitosan (Figure 3), there is concordance between optimum flocculation concentrations as judged by the maximum in the rate and the center point of the turbidity trough which, at 1.01, is comfortably within the error range for 1:1 stoichiometry. There is a 12% enhancement of rate upon charge neutralization compared with that for rapid coagulation and a significantly larger increase than was found with CTAB. This type of rate enhancement has been attributed by Kasper36 and by Gregory43 to opposite charge attractions arising from the “charge patch” nature of polyelectrolyte adsorption. The positive charges on the polyelectrolyte chain are usually more closely spaced than are the center to center distances between negative charges on the particle surface. Thus, when the polymer molecule is adsorbed in a relatively flat configuration on the particle surface to give a “mosaic pattern”, a local excess of positive charges are introduced while the rest of the surface is free of flocculant and carries the original negative charge. These regions of opposite charge may then lead to Coulombic attractions between the particles, leading to a flocculation rate above that of simple coagulation. For synthetic polyelectrolytes in water, rate enhancements of the order of 2-fold were reported by Gregory.43 In the case of the chitosans, the use acid solutions produces a screening effect, reducing the electrostatic attraction, as a result of the increased ionic strength involved. Structural studies44 suggest that the cationic charges in fully deacetylated chitosan are spaced at 1 nm while the negative charges on the latex, assuming hexagonal close packed circular “parking areas”, are 2.5 nm apart. Thus, charge patches with a net positive charge are to be expected. For the oligomeric chitosan, more patches will be formed than would be the case for a higher molecular weight material. It was observed during residual turbidity studies that flocs formed by the oligomer were weaker and more redispersible than was the case for higher molecular weight materials, and this was interpreted as reflecting the more disperse nature of the charge interaction. Next, considering the four samples of medium molecular weight chitosan of increasing degrees of acetylation, it can be seen (Figure 4 and Table 3) that, for the chitosan[0.02]M sample, results fall just within the error band for 1:1 stoichiometry while significant flocculation commences at a much lower ratio than for the oligomer. The rate enhancement is greater at some 30%. Thereafter (Table 3 and Figure 5), with increasing degrees of acetylation of the medium molecular weight chitosan, increasing amounts of flocculant are required with chitosan[0.14]M (37) van der Steeg, H. G. M.; Cohen Stuart, M. A.; de Keizer, A.; Bijsterbosch, B. Langmuir 1992, 8, 2538. (38) Linse, P. Macromolecules 1996, 29, 326. (39) Shubin, V.; Samoshina, Yu.; Menshikova, A.; Evseeva, T. Colloid Polym. Sci. 1997, 275, 655. (40) Tanaka, H.; Odberg, L.; Wagberg, L.; Lindstrom, T. J. Colloid Interface Sci. 1990, 134, 217. (41) Shubin, V.; Linse, P. Macromolecules 1997, 30, 5944. (42) Denoyel, R.; Durand, G.; Lafuma, F.; Audebert, R. J. Colloid Interface Sci. 1990, 139, 281. (43) Gregory, J. J. Colloid Interface Sci. 1976, 1, 35-44. (44) Minke, R.; Blackwell, J. J. Mol. Biol. 1978, 120, 167.
Ashmore and Hearn
Figure 7. Reduction of the maximum relative rate of flocculation as a function of added sodium chloride for 2% acetylated medium molecular weight chitosan.
at 1.10, chitosan[0.33]M at 1.13, and chitosan[0.57]M at a 1.23 stoichiometric ratio, reflecting an increasing inefficiency of neutralization with increased acetylation. As the linear charge density on the molecules decreases, more molecules are involved in the neutralization. Also, with increasing acetylation, the backbone of the molecule becomes stiffer19 as a consequence of intramolecular hydrogen bonding between acetamido (-NHCOCH3) and hydroxymethyl (HOCH2-) moieties (see Figure 1). The stiffer molecules are less able to configure themselves to match the template set by the charges on the particle surface. More molecules are involved, forming more patches but of lower excess cationic charge density. The breadth of the flocculation region decreases as these molecules become less effective flocculants and the number of marginally stable particles, which may contribute to the flocculation zone, in the more protracted time span of the residual turbidity experiment, decreases. It would also be anticipated that the charge density in a patch would be greater for molecules with the greater linear charge density and this is indeed reflected in the enhancements in the aggregation rates. The high molecular weight chitosan samples at degrees of acetylation of 17% (Table 3) and 38% (Figure 6) showed, within the limits of experimental uncertainty, a return to 1:1 stoichiometry, where these degrees of acetylation for the medium molecular weight material would have been expected to show lower efficiency. Kasper,36 modeling the charge patch mechanism, suggested that the polymer adsorption density, i.e., the ratio of the weight of polymer per patch to the area of the patch, is proportional to the molecular weight raised to the power of one-third. Thus, these patches would be expected to be of greater efficiency than for the medium molecular weight materials of the same degree of deacetylation. Again, the rate enhancement was greater at the higher linear charge density while the breadths of the flocculation regions were rather similar. That the enhancement of rates observed in this work is a consequence of charge patch effects is supported by the data shown in Figure 7, where it can be seen that, as the ionic strength is increased, by additions of sodium chloride, so the rate enhancement decreases. The increased ionic strength shields the charges so that attractions are reduced and the rate falls to that of rapid coagulation. Electrophoretic mobility data in Figure 8 for chitosan[0.02]M are in agreement, within the bounds of experimental error, with zero mobility occurring at 1:1 stoichiometric addition. For chitosan[0.57]M, zero mobility occurs at a little above the 1:1 stoichiometric ratio but again is in agreement with the results from the flocculation experiments. In the electrophoresis experiments, the particles remain in an unflocculated condition
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Langmuir, Vol. 16, No. 11, 2000 4911
little qualitative difference to the following argument, then using Katchalsky’s equation47
pKa ) pH + log[(1 - x)/x]
(5)
x ) degree of dissociation
Figure 8. Electrophoretic mobility data as a function of charge ratio for 2% (4) and 57% ([) acetylated medium molecular weight chitosan.
it can be predicted that, at pH 5, the association will remain high, at 97%, so that only 3% more polyelectrolyte might be needed at this pH, while, at pH 6.5, the 50% association suggests that twice the amount used at pH 3 might be required, and if the argument is continued to pH 7, then a 4-fold increase would be predicted. For pHs 3, 5, 6, and 7, the experimental results are at least in qualitative accord with these predictions although, beyond pH 6.6, some chitosan precipitation is to be expected.1 Thus, while doubts about the appropriate choice and the constancy of the pKa make quantitative comparisons difficult, it can be seen that the efficiency of chitosan as a flocculant decreases markedly as neutral pH is approached.
(4)
Conclusions Flocculation of a model colloid latex, having a very typical surface charge density, using well-characterized chitosans of varying degrees of acetylation, has been shown to proceed by a charge neutralization mechanism. Enhancement of the rate of flocculation compared with the rate of fast coagulation suggests that a “charge patch” mechanism is in operation. At high degrees of deacetylation, i.e., at high linear charge densities, optimum flocculation occurs at similar weight percent concentrations of the chitosan independent of its molecular weight. The trend, with an increasing degree of acetylation of the chitosan, is for a reduction of efficiency as the backbone of the chitosan chain becomes more rigid and the use of a high molecular weight material does then show some advantage. Chitosans require acid conditions to act as cationic polyelectrolytes and to retain their solubility, and thus their efficiency is considerably reduced as neutral pH is approached. For practical use, there would be advantages, in terms of the concentration by weight of flocculant required, in using chitosans of a high degree of deacetylation under acid conditions. As shown above, however, chitosans can produce charge neutralization over a wider range of degrees of acetylation and pH, albeit with reduced efficiency. In the assessment of the overall cost of using this renewable resource and nontoxic flocculant, the amount of preprocessing required for optimal performance may warrant comparison with the cost of the increased dose required when used under less ideal conditions.
lies predominantly (99.97%) to the left. The acid dissociation constant for polyelectrolytes is not actually constant since, e.g., the ease of dissociation of the conjugate acid group -NH3+ will be increased in the presence of similar adjacent groups, thereby increasing Ka and decreasing pKa. If, however, the pKa is approximated to be constant at 6.5,45,46 although other realistic choices would make
Acknowledgment. We wish to sincerely thank Prof. George Roberts for providing chitosan samples and for advice on their characterization, Dr. Jim Goodwin, President of the Interfacial Dynamics Corp., for providing titration data on the latex, and Mr. Rowland Crouch of Beckman Coulter for help in obtaining electrophoretic mobility data.
Figure 9. Effect of changing pH upon the charge ratio required for flocculation using 2% acetylated medium molecular weight chitosan: [ ) pH 3; 4 ) pH 5; × ) pH 6; ] ) pH 7.
as a consequence of the high dilution used. They retain the charge patch heterogeneity of positive and negative charges coexisting on individual particles, thereby causing some uncertainty as to the likely location of a Stern plane.36 The greater overcompensation shown by the stiffer chitosan is manifest in the attainment of a positive mobility greater than that for chitosan[0.02]M. Figure 9 shows the influence of pH upon the optimum flocculation concentration for chitosan[0.02]M, and a similar pattern of results was found for chitosan[0.33]M and chitosan[0.57]M. At pH 3, the equilibrium
-NH3+ + H2O h -NH2 + H3+O
(45) Domard, A. Int. J. Biol. Macromol. 1987, 9, 98. (46) Rinaudo, M.; Domard, A. In ref 2, p 71.
LA991648W (47) Katchalsky, A. J. Polym. Sci. 1954, 12, 159.
