Algal Flocculation with Synthetic Organic Polyelectrolytes
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1 APPuF.D MIcRomoLoDY, Dec. 1969, p Vol. 18, No American Society for Microbiology Printed in U.S.A. Algal Flocculation with Synthetic Organic Polyelectrolytes MARK W. TENNEY, WAYNE F. ECHELBERGER, JR., RONALD G. SCHUESSLER, AND JOSEPH L. PAVONI Department of Civil Engineering, University of Notre Dame, Notre Dame, Indiana Received for publication 23 April 1969 The feasibility of removing algae from water and wastewater by chemical flocculation techniques was investigated. Mixed cultures of algae were obtained from both continuous- and batch-fed laboratory reactors. Representative cationic, anionic, and nonionic synthetic organic polyelectrolytes were used as flocculants. Under the experimental conditions, chemically induced algal flocculation occurred with the addition of cationic polyelectrolyte, but not with anionic or nonionic polymers, although attachment of all polyelectrolyte species to the algal surface is shown. The mechanism of chemically induced algal flocculation is interpreted in terms of bridging phenomena between the discrete algal cells and the linearly extended polymer chains, forming a three-dimensional matrix that is capable of subsiding under quiescent conditions. The degree of flocculation is shown to be a direct function of the extent of polymer coverage of the active sites on the algal surface, although to induce flocculation by this method requires that the algal surface charge must concurrently be reduced to a level at which the extended polymers can bridge the minimal distance of separation imposed by electrostatic repulsion. The influence of ph, algal concentration, and algal growth phase on the requisite cationic flocculant dose is also reported. The removal of algae from various types of water, such as public water supplies and effluents from algal wastewater treatment systems, is of extreme importance if continual maintenance of a high level quality of water is to be insured. The presence of algae in water not only creates nuisance conditions (unsightly blooms, taste and odors, turbidity, clogging of filters, etc), but, in the strictest sense, algae must also be considered as potential organic and inorganic pollutants. Among the various methods that have been suggested for algal separation are autoflocculation, centrifugation, chemical treatment, filtration, flotation, ion-exchange, sedimentation, and straining. The purpose of this research was to investigate the removal of algae from their suspending liquid by chemical flocculation techniques. The available literature pertaining to chemical flocculation of algae, although limited, indicates the apparent feasibility of utiliing this technique for algal removal. In studies involving the agglomeration of Chlorella cells with synthetic organic polyelectrolytes, Cohen, Rourke, and Woodward 'Present address: Technical Service and Development, The Dow Chemical Co., Midland, Mich (2) observed that good flocculation was achieved with a cationic polyelectrolyte, whereas no flocculation was obtained with the anionic polymer. Excellent aggregation of Chlorella and Scenedesmus was also reported with cationic polyelectrolytes by Golueke, Oswald, and Gee (4), indicating that the requisite polyelectrolyte dose was insensitive to ph in the range of 5. to 1.4, but that increasing concentrations of dispersed algal cells increased the requisite polyelectrolyte dose proportionately. Field-scale pilot plant tests by the Dow Chemical Co. have served to demonstrate further the feasibility of using cationic polyelectrolytes as algal flocculants. Aside from the work of Golueke, Oswald, and Gee (4), which begins to delineate the parameters involved in the chemical flocculation of algae, little other quantitative data are available on this subject. Thus, in the present work, primary consideration was given not only to determining the effectiveness of selected polyelectrolytes as algal flocculants, but also to evaluating the basic mechanism of the algal-flocculant interaction and the associated parameters affecting this interaction. Most algal cells (with the exception of certain Downloaded from on May 15, 218 by guest
2 966 TENNEY ET AL. APPL. MICROBIOL. filamentous forms) fall in the sie range of 5 to 5 jam. Although larger than true colloids (1 to 1 nm), algae nevertheless possess many surface properties similar to true colloids. Discrete algal cells are known to form stable microbial suspensions, possess a chemically reactive cellular surface, and possess a net negative surface charge (5) due to the ioniation of functional ionogenic groups. Since the stability of algal suspensions apparently depends on the forces interacting between the particles themselves as well as on the forces interacting between the particles and the water, algae can be considered as hydrophilic bio-colloids. A theoretical model describing the mechanism of algal flocculation would parallel that proposed by LaMer and Healy (7) for the flocculation of colloidal particles by polymers. This model would postulate that polymer molecules will attach themselves (either by electrostatic or chemical forces, or both) to the surface of the algae at one or more sites, and that part of the chain will extend out into the bulk of the solution. When these extended chain segments make contact with vacant sites on other algal cells (or other similarly extended chain segments), bridges are created which will serve to form a threedimensional algal-polymer matrix. Under quiescent conditions, this matrix will subside from suspension, thus allowing for the discharge of a clear supernatant liquid. MATERIALS AND METHODS Both laboratory-scale batch- and continuous-flow algal culturing reactors were used. The initial algal inoculum was obtained from the inside wall of an aeration tank at the South Bend Wastewater Treatment Plant and was continually maintained in the laboratory throughout this study. All of the algal cultures were sustained on a standard liquid algal medium (6) with the predominant species being of the Chlorophyta. Culture illumination was provided through a bank of fluorescent lamps (Westinghouse Type F2T12/GRO) developed specifically to stimulate photosynthesis. This light source imparted an intensity of 4 ft-c to the culture surface with an energy dominance in the blue and red regions of the visible light spectrum. Commercially available synthetic organic polyelectrolytes were examined with respect to their suitability as algal flocculants. Organic flocculants used were a cationic polyamine with an approximate molecular weight of 5 X 16 (Dow Chemical Co., C-31), an anionic polystyrene with a molecular weight in the range 8 X 17 to 1 X 18 (Dow Chemical Co., A-21), and a nonionic polyacrylamide with an approximate molecular weight of 3 X 16 (Nalco Chemical Co., N-67). For all of the flocculation tests, standard jar testing procedures were used. After the flocculant addition to 25 ml of algal suspension, E >- s 8_ / W 6 o 4 U. Z 2 4 COEF. OF CORRELATION (r).994 w o Z ') KJELDAHL NITROGEN CONCENTRATION - mg/i FIG. 1. Calibration curve; relationship between cationic polyelectrolyte (Dow, C-31) and Kjeldahl nitrogen concentration. there was a 2-min rapid mix at 8 rev/min (blade speed) followed by a 15-min slow mix at 3 rev/min (blade speed) and a 1-hr period of quiescent settling. A constant ph, as suggested by Stumm and Morgan (1), was maintained during the test period by automatic titration. The extent of algal flocculation was evaluated through measurement of the residual turbidity (spectrophotometric absorbance at 69 nm) in the supernatant liquid after the settling period. Flocculation curves for the various test conditions were developed by correlating the residual turbidity information with the increasing flocculant dosage. Concurrently the algal surface charge was approximated by microelectrophoretic mobility measurements (1) and the extent of polymer coverage on the algal surface was determined by measuring the concentration of residual polymer in solution. Residual cationic polyelectrolyte concentrations were determined by Kjeldahl nitrogen (Fig. 1), and residual anionic and nonionic polyelectrolyte concentrations were determined by organic carbon analyses (Chemical Oxygen Demand). Algal cell concentrations were determined by analysis of the dry solids concentration expressed as milligrams per liter by using a.45-;tm membrane filter with drying at 13 C for 1 hr. RESULTS AND DISCUSSION Polymer interactions with algae: cationic polyelectrolyte. The extended segment theory of polymer flocculation would predict that to achieve optimal flocculation of a given algal suspension, an optimal concentration of polyelectrolyte must be added. Addition of polyelectrolyte concentrations either greater or less than the optimum will not achieve the degree of flocculation associated Downloaded from on May 15, 218 by guest
3 VOL. 18, 1969 ALGAL FLOCCULATION 967 with the optimal polyelectrolyte dose. Such a relationship for a cationic polyelectrolyte is shown in Fig. 2A; the optimal dose can be observed to occur at approximately 2.5 mg/liter. In accordance with this theory, the degree of flocculation will be determined by the extent to which the algal surface has been covered with polyelectrolyte. Polyelectrolyte concentrations less than optimum will result in insufficient bridging to withstand the shearing forces imposed by the conditions of agitation. Conversely, if too many surface sites on the suspended algal particles are covered with polymer segments, bridging will be hindered and in the limit, when all the adsorptive sites are occupied, bridging will be totally prevented because of electrostatic or static hinderance, or both. Confirmation of this interpretation is given in Fig. 2C and D. Figure 2C shows the relationship between the concentration of polymer added and the concentration of polymer measured in solution; the difference between the two representing the concentation of polymer attached to the algal surface. It can be noted (in particular from the insert to Fig. 2C) that up to the polymer concentration of approximately 3 mg/liter, all of the added polyelectrolyte is attached to the algal surface and no residual concentration is in solution. Above this concentration, however, increasing amounts of polymer are I- D EJ WJZ o c r Z.5 D. SURFACE COVERAGE 1 CE w.3-24, , IL. H < ALGAL CONCG 2 mg/1 U. u-.1 2OD CONCENTRATION OF CATIONIC POLYELECTROLYTE - mg/i FIG. 2. Algal flocculation with cationic polyelectrolyte (Dow, C-31). m or en) co E 76 1; m F CONCENTRATION OF ANIONIC POLYELECTROLYTE - mg/i FIG. 3. Algal flocculation with anionic polyelectrolyte (Dow, A-21). observed in solution and eventually the two curves are parallel, indicating that the algal surface is now completely covered with polymer. Figure 2D (the ordinate differences of Fig. 2C) represents the extent of algal surface coverage as a function of increasing polymer dosage; optimal flocculation occurs at approximately 5% surface coverage. EA I I A. FLOCCULATION CURVE B. MICRO- ELECTROPHORETIC MOBILITY ALGAL CONC. 15mg/ ph = 7. Additional evidence in support of this model can be obtained from measurements of the change in the net algal surface charge resulting from increasing polyelectrolyte concentrations; attachment of the cationic polyelectrolyte to the negative algal cell would predict a proportional charge reduction of the algal-polymer species. As shown in Fig. 2B, charge reduction (and reversal) is obtained from the attachment of the cationic species; however, no further change in charge results after complete polymer coverage of the algal surface. In view of the low optimal ph for algal flocculation with cationic polyelectrolytes, the question may arise whether such ph adjustment will conceivably cause leakage of nitrogenous constituents from the algal suspension, thereby giving false values for concentrations of polymer in solution and, consequently, erroneously calculated values for surface coverage. To determine whether such a leakage occurred, Kjeldahl nitrogen analyses were performed on the filtrate from six algal suspensions which were adjusted to ph values ranging from 2 to 12 without cationic polymer addition. No nitrogenous leakage was evidenced in this study. Anionic and nonionic polyelectrolytes. Under the conditions of our experimentation, no algal flocculation was achieved with the anionic and nonionic polyelectrolytes selected. In both instances, Downloaded from on May 15, 218 by guest
4 " TENNEY ET AL. APPL. MICROBIOL. however, polymer-algal surface coverage relationships similar to those with a cationic polyelectrolyte were observed. Figures 3A and B represent the flocculation curve and micro-electrophoretic mobility, respectively, for algal cells dosed with anionic polyelectrolyte. Although no flocculation is obtained with this anionic polyelectrolyte, the fact that the anionic polymer is attached to the w algal surface is apparent from the corresponding <.4 increase in negative micro-electrophoretic mobility. The data obtained with a nonionic polyelectrolyte are shown in Fig. 4. The tendency for Cl) M.3 the algae to remain dispersed within the dosages tested is indicated in Fig. 4A. Measurement of.2 residual polymer concentration (Fig. 4C) reveals that the nonionic polymer is being adsorbed on.1 ALGALCN.27m/ the algal surface in a similar fashion to the cationic polymer (Fig. 4D). As would be anticipated, Fig. 4B indicates that no change in net surface charge of the algae occurs from the attachment of the nonionic polyelectrolyte. FIG. 5. Effect of ph on algal flocculation with DH It is apparent from these data that the mechanism of anionic and nonionic polymer bonding to liter; Dow, C-31). constant dosage of cationic polyelectrolyte (1 mg/ the algal surface is controlled by chemical forces rather than electrostatic forces; hydrogen bonding, anion interchange with adsorbed anions (such suspensions in the presence of established polymer The tendency for the algae to remain in stable as OH-), or interaction with cations on the surface bonding would clearly indicate the inability of the attached polymer segments to form colloid surface have been proposed as possible attachment mechanisms in related systems (9). bridges. In all probability, the length of the extended polymer segments are insufficient to bridge the limiting (or minimal) distance separating the W.3 particles (established by the electrostatic repulsion c<.2 A. FLOCCULATION CURVE of the like particle charges). Thus, to avoid electrostatic interference, the net surface charge on (n. the algae must be reduced to a value which will.. allow the individual particles to approach one > -1. B. MICRO- ELECTROPHORETIC MOBILITY another at a distance less than the length of the extended polymer segment. [It can be noted in m ' -2. Fig. 2A and B, for example, that the net algal surface charge in the one of flocculation has been C. RESIDUAL POLYMER '1-8 / appreciably reduced from that shown in Fig. 3B E Xr CONC. OF / 8-6-)S POLYMER and 4B.] In this context, algal flocculation with C; anionic and nonionic polyelectrolytes could be 642oo44-'S~ feasible under different environmental conditions, such as agitation and varying electrolyte concentrations (especially polyvalent cations). O* * t ~ ~ ~~~~ IN SOLUTION Parameters affecting algal flocculation with - 5 D. SURFACE COVERAGE cationic polyelectrolytes. The requisite polymer E dosage in any flocculation process is influenced by 4 many variables. In our studies, the effect of three o 3 parameters on the requisite cationic polyelectrolyte dosage was evaluated (ph, algal 2 ALGAL CONC. 15mg/I concen- L. ph U_ CONCENTRATION OF NON-IONIC POLYELECTROLYTE-mg/l FIG. 4. Algal flocculation with nonionic polyelectrolyte (Nalco, N-67). tration, and algal growth phase). As will be discussed subsequently, the effect of these parameters is pronounced; it should be noted, however, that other parameters (e.g., degree and extent of agitation, algal sie, temperature, electrolyte concentration, etc.) would also be expected to influence Downloaded from on May 15, 218 by guest
5 VOL. 18, 1969 ALGAL FLOCCULATION _ o POLYMER CONCENTR. - mg/i.3.2 X /5i CONCENTRATION OF CATIONIC POLYELECTROLYTE - mg/i FIG. 6. Effect of algal concentration on requisite cationic poyelectrolyte concentration. the flocculation of algae with synthetic organic polyelectrolytes. ph. In any flocculation process, consideration must be given to hydrogen ion concentration as it will influence both the surface charge density of the colloid and the action of the polyelectrolyte (e.g., extent of coiling, degree of ioniation, charge density, etc.). Figure 5, for example, indicates the effect of ph on algal flocculation while maintaining a constant concentration of cationic polyelectrolyte. It can be observed from Fig. 5 that most effective flocculation is obtained in the lower ph ranges (i.e., 2 through 4). Microelectrophoretic mobility measurements (in the absence of any polyelectrolyte addition) indicate that in this ph range algal cells exhibit their lowest net surface charge. Our studies indicate that in neutral and basic ph ranges algae possess a net negative surface charge, whereas increasing hydrogen ion concentration decreases (and reverses) the net negative surface charge; an isoelectric point is observed in the ph range 2 through 3. Considering also that in this low ph range cationic polyelectrolytes will be highly charged and well extended, it may be observed that the optimal ph for algal flocculation with cationic polyelectrolyte will be established at the level at which (i) electrostatic repulsion forces are the lowest and (ii) polymeric bridging will be facilitated due to greater polymer expansion. Algal concentration. Interpretation of algal flocculation with synthetic organic polyelectrolytes in terms of algal-polymer surface coverage relationships would predict that variation in algal concentration (algal surface area) would appreciably influence the concentration of polyelectrolyte required for a given degree of flocculation. When a specific fraction of surface coverage dictates a certain degree of flocculation, increasing the surface area of the colloids by increasing their concentration correspondingly should increase the concentration of polyelectrolyte required to achieve the same degree of flocculation. Figure 6 shows the flocculation curves for three different algal concentrations, and the insert to Fig. 6 reveals the definite stoichiometry between algal concentration and polymer dosage for optimal flocculation (5% surface coverage). Algal growth phase. Another parameter which affects the flocculation of algae with polymers and is unique to bio-colloidal systems is the effect the growth phase of microorganisms. The previously reported data (Fig. 2-6) all relate to the flocculation of dispersed algae obtained from a chemostatically operated or continuous-flow type of culturing reactor, thus insuring a constant algal growth stage. Because the surface properties of algae are known to vary with their growth phase (5), a batch system, initially seeded with a population from the continuous flow reactor, was established. This system allowed for the withdrawal of samples at various stages in the algal growth curve for subsequent flocculation analysis. Figure 7A shows a typical growth curve for this system, and Fig. 7B indicates the concentration of polymer required for 5% flocculation (normalied to a cell concentration of 3 mg/liter) as a function of time (or growth stage). Comparison of Fig. 7A and B indicates that to achieve the same degree of flocculation (e.g., 5%) the least amount of flocculant is required in the late log and early declining growth phases. Consistent with the previous observation that the chemical reactivity of the algal surface is such to allow positive bonding of nonionic and anionic polymers, it would appear plausible to postulate that other molecular constituents could similarly be sorbed on the algal surface. The production of high molecular weight extracellular metabolites by algae is documented in the literature (3, 8). Experimental extraction of these extracellular polymers throughout the algal growth curve was undertaken using ethyl alcohol extraction techniques similar to those of Ueda et al. (11). As Fig. 8 shows, the rapid accumulation of this extracellular material occurs exclusively during the endogenous phase of algal growth. Other Downloaded from on May 15, 218 by guest
6 97 TENNEY ET AL. work in our laboratory has shown these polymeric molecules to be comprised of long chain polysaccharides, proteins, and nucleic acids, and are probably of sufficient length to form bridges between algal particles. Hence, microbially produced polymers initially could be enhancing the flocculation, whereas in later growth stages the accumulation of this material could be acting as a protective colloid. Within the limitations of this study, chemically induced algal flocculation occurred with the addition of a cationic synthetic organic polyelectrolyte but not with the anionic or nonionic polymers. The interpretation of the mechanism of chemically induced algal flocculation may be described in terms of a bridging phenomenon between the discrete algal cells and the linearly extended polymer chains, forming a three-dimensional matrix that is capable of subsiding under quiescent conditions. Results indicate that chemically induced algal flocculation is a direct function of the extent of polymer coverage of the active sites on the algal cell surface, provided that the algal surface charge is reduced to a level which permits the extended polymer chains to bridge the minimal distance of separation established by electro- 5 A. ALGAL GROWTH CURVE DECLINING 4 LOG. GROWTH GROWTH ENDOGENOUS RESPIRAT'N or_, 1 J ca6 I. 3 22_ 1 l l l l l l l B. CONC. OF POLYMER REQ'D. FOR 5% FLOCCUL2N TIME - DAYS FIG. 7. Effect of growth phase ont algalflocculation with cationic polyelectrolyte (Dow, C-31) TIME - DAYS APPL. MICROBIOL FIG. 8. Accumulation of extracellular metabolites during algal growth. static repulsion. Optimal algal flocculation occurs at approximately 5% coverage of algal cell surface, with stabiliation and restabiliation occurring at lower and higher surface coverage ratios, respectively. Algal flocculation with cationic polymers is most effective at low ph levels due to reduced electrostatic repulsion between the colloids and improved opportunity for polymeric bridging because of greater extension of the polymer chains. A definite stoichiometric relationship is indicated between algal cell concentration and requisite cationic polymer dosage for optimal flocculation; however, the flocculation of algae with cationic polymer is definitely influenced by the growth phase of the algae, with optimal conditions for flocculation in the late log and early declining growth phases. ACKNOWLEDGMENTS This investigation was supported by Department of Health, Education, and Welfare, Environmental Control Administration research grant UI-33, Public Health Service traineeship grant ES to R. G. S. from the Division of Environmental Health Sciences, National Institutes of Health, and Federal Water Pollution Control Administration research fellowship 5-Fl-WP-38, to J. L. P. The kindness of Dow Chemical Co. and Nalco Chemical Co. for supplying samples of synthetic organic polyelectrolytes and information is appreciated. LITERATURE CITED 1. Black, A. P., and A. L. Smith Determination of the mobility of colloidal particles by micro-electrophoresis. J. Amer. Water Works Ass. 54: Cohen, J. M., G. A. Rourke, and R. L. Woodward Natural and synthetic polyelectrolytes as coagulant aids. J. Amer. Water Works Ass. 5: Fogg, G. E. and R. A. Lewin (ed.) Physiology and biochemistry of algae. Academic Press Inc., New York. 4. Golueke, C. G., J. W. Oswald, and H. K. Gee Ser. Report no Sanitary Engineering Research Liboratory, University of California, Berkeley. Downloaded from on May 15, 218 by guest
7 VOL. 18, 1969 ALGAL FLOCCULATION Ives, K. J Electrokinetic phenomena of planktonic algae. Proc. Soc. Water Treat. Exam. 5: Kat, W. A., and J. Myers Nutrition and growth of several blue green algae. Amer. J. Bot. 42: LaMer, V. K., and T. W. Healy Adsorption-flocculation reactions of macromolecules at the solid-liquid interface. Rev. Pure Appl. Chem. 13: Lewin, R. A Extracellar polysaccharides of green algae. Can. J. Microbiol. 2: Sommerauer, A., D. L. Sussman, and W. Stumm The role of complex formation in the flocculation of negatively charged sols with anionic polyelectrolytes. Kolloid-Z. Z. Polymere. 225: Stumm, W., and J. J. Morgan Chemical aspects of coagulation. J. Amer. Water Works Ass. 54: Ueda, S., K. Fujita, K. Komatsu, and Z. Nakashima Polysaccharide produced by gens pulularia. L. Production of PoLysaccharide by growing cells. Appl. Microbiol. 11: Downloaded from on May 15, 218 by guest
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