JOURNAL OF BIOSCIENCE AND BIOENGINEERING Vol. 103, No. 6, 509–513. 2007 DOI: 10.1263/jbb.103.509
© 2007, The Society for Biotechnology, Japan
Batch and Continuous Packed Column Studies of Cadmium Biosorption by Hydrilla verticillata Biomass Sushera Bunluesin,1 Maleeya Kruatrachue,1* Prayad Pokethitiyook,1 Suchart Upatham,2 and Guy R. Lanza3 Department of Biology, Faculty of Science, Mahidol University, Rama 6 Road, Bangkok 10400, Thailand,1 Faculty of Science, Burapha University, Chonburi 20130, Thailand,2 and Environmental Science Program, 312 Stockbridge Hall, University of Massachusetts, Amherst, MA 01003, USA3 Received 16 August 2006/Accepted 19 February 2007
The removal of heavy metal ions by the nonliving biomass of aquatic macrophytes was studied. We investigated Cd biosorption by dry Hydrilla verticillata biomass. Data obtained in batch experiments indicate that H. verticillata is an excellent biosorbent for Cd. Cd was rapidly adsorbed and such adsorption reached equilibrium within 20 min. The initial pH of the solution affected Cd sorption efficiency. Results obtained from the other batch experiments conformed well to those obtained using the Langmuir model. The maximum adsorption capacity qmax for H. verticillata was 15.0 mg/g for Cd. The breakthrough curve from the continuous flow studies shows that H. verticillata in the fixed-bed column is capable of decreasing Cd concentration from 10 to a value below the detection limit of 0.02 mg/l. The presence of Zn ions affected Cd biosorption. It can be concluded that H. verticillata is a good biosorbent for treating wastewater with a low concentration of Cd contaminants. [Key words: Hydrilla verticillata, biosorption, packed column study]
Heavy metals once they contaminate the environment have permanent adverse ecological effects. Therefore, heavy-metal contamination in the environment has become an area of increasing concern. With increasing environmental awareness and the toughening of governmental policies, it has become necessary to develop new environmentally friendly ways to clean up contaminants using low-cost methods and materials (1). Biosorption is a process that utilizes dead or living biomass to sequester toxic heavy metals and is particularly useful for the removal of contaminants from industrial effluents. Biosorption processes are particularly suitable for the treatment of wastewater containing low heavy metal ion concentrations (2). Adsorbent materials (biosorbents) derived from suitable biomass can be used for the effective removal and recovery of heavy metal ions from wastewater streams. Aquatic plants, both living and dead, are heavy-metal accumulators. Therefore, the application of aquatic plants to the removal of heavy metals from wastewater has gained increasing interest (3). Some freshwater macrophytes including Potamogeton lucens, Salvinia hergozi, Eichhornia crassipes, Myriophyllum spicatum, Cabomba sp., Ceratophyllum demersum have been investigated for their potential in heavy-metal removal (4–7). Their mechanisms of metal removal by biosorption can be classified as extracellular accumulation/precipitation, cell surface sorption/precipitation, and intracellular accumulation (8). These mechanisms
can result from complexation, metal chelation, ion exchange, adsorption and microprecipitation (7). The biomasses of aquatic plants, algae and plant materials are biological resources available in large quantities and can be used for the development of biosorbent materials. The biosorption of Cd ions by different living and nonliving biomasses has been extensively studied (9–14). Hydrilla verticillata is a submerged aquatic weed that can grow up to the surface and form dense mats in all bodies of water. However, only few studies of Cd removal by H. verticillata have been reported. H. verticillata has shown promising sorbent characteristics for the removal of Cd ions in our preliminary studies. Therefore, in this study, the biosorption of Cd ions by H. verticillata was studied, and the effects of different experimental parameters on Cd biosorption, such as contact time, initial pH and Cd concentration, were also investigated. Moreover, the adsorption of metal ions on H. verticillata dry biomass packed in a column was also examined. These studies are considered fundamental for further studies involving the scaling-up of the process under actual conditions. MATERIALS AND METHODS Biosorbents The aquatic macrophyte H. verticillata was obtained from natural ponds and grown in 30% Hoagland’s nutrient solution (15) under controlled conditions (25 ±2°C and 12 h-12 h light-dark cycle). Three-week-old H. verticillata specimens from a starting culture were collected and washed with distilled water, dried in an oven for 12 h at 100°C, and ground into powder. The
* Corresponding author. e-mail:
[email protected] phone: +66-2-201-5483 fax: +66-2-354-7166 509
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biomass was passed through a 1-mm sieve to remove any particles greater than 1 mm in size before use and stored in a desiccator. This was carried out to obtain small particles since the smaller the particle size the higher the surface area. Metal solution The test solutions containing single Cd or Zn ions were prepared by diluting 1000 mg/l of Cd(NO3)2 ⋅4H2O standard solution (Merck, Darmstadt, Germany) or 1000 mg/l of Zn(NO3)2 standard solution (BDH, London, UK) with deionized water to the desired concentrations (1, 10, and 100 mg/l Cd and 10 mg/l Zn). The pH of the working solution was maintained by adding 1 M HNO3 or 1 M NaOH. All the chemicals used were of analytical reagent (AR) grade. Batch biosorption study The biosorption capacity of H. verticillata was determined by shaking 100 ml of Cd solutions of various concentrations (1, 10, 100 mg/l Cd) in a 250-ml flask, with homogeneously dry biomass of different weights (0.3, 0.5, 0.7, and 0.9 g). The mixture was shaken on a rotary shaker (IKA KS501 digital; Janke & Kunkel GmbH & Co. KG, Staufen, Germany) at 150 rpm for 1 h at 25 ±2°C. Biomass was separated from the metal solution by passing through a Whatman 0.45-µm membrane filter and the filtrate was subjected to residual Cd concentration determination. For the determination of metal biosorption rate, the filtrate was analyzed for residual Cd after contact periods of 3, 15, 30, 60, 90 and 120 min. The effect of pH on Cd sorption by H. verticillata was determined by adjusting the metal solution at different pHs 1, 3, 5, 7 and 9. All the experiments were performed in triplicate. A metal- and biomass-free blank was used as a control. The data used to derive the Langmuir constants were obtained by varying the initial Cd concentration while keeping the biomass in each sample constant. The preliminary results showed that equilibrium was obtained after 1 h of the run. Thus, the equilibrium period of 1 h during the sorption experiment was defined as an equilibrium condition. The reusability of the biosorbent was also studied. Dry H. verticillata biomass of different weights (1, 2, and 3 g) was packed in a glass column (1 cm ID, 50 cm column length). The packed bed volumes obtained were 5, 10 and 17 ml, respectively. For each experiment, 50 ml of 10 mg/l Cd solution (pH 5) was passed through the packed column; the effluent was then collected and analyzed for Cd concentration. For desorbtion process, 50 ml of 0.1 HCl was used to rinse the Cd-laden biomass, followed by distilled water. The desorbed and regenerated H. verticillata column was used again for two more cycles. Packed bed column continuous flow studies Packed bed experiment was conducted at room temperature (25 ± 2°C) in a 1-cm ID glass column, packed with 0.5 and 1 g of H. verticillata dry biomass to obtain bed volumes of 3 and 4.3 ml, respectively. Solutions of 10 mg/l Cd (pH 5) were pumped through the column at a flow rate of 9.8 ml/min using a peristaltic pump (model 77200-62; Cole Palmer Instrument Company, Barrington, IL, USA). The samples were periodically collected and analyzed for metal concentrations. The biosorption saturation capacity of the packed column was reached at the stage when no Cd sorption occurred or the outlet concentration reached 10 mg/l. For the determination of the synergistic/antagonistic effect of cations present in the influent metal solution, Zn and Cd of the same concentration (10 mg/l) were passed through 1 g of the biomass on the packed column. Effluents were collected periodically and analyzed for Cd and Zn concentrations. Metal analysis After biosorption, the pH of the filtrate was adjusted to a value below 2 and the residual concentrations of the metal ions were analyzed by flame atomic absorption spectrophotometry (SpectraAA 55B; Variance, Melbourne, Australia). All the pieces of glassware used were soaked in 10% nitric acid overnight and rinsed with distilled water. Preliminary experiments had shown that Cd losses due to adsorption onto the flask walls and filter paper were negligible. The Cd uptake q (mg metal ion/g dry biomass)
was calculated by the simple concentration difference method using q = (CI − CF)V/M
(1)
where CI and CF are the initial and final metal concentrations (mg/l), respectively; V is the volume of sample solution (l); and M is the dry weight of the added biomass (g). The linearized equation form of the Langmuir model used to evaluate maximum metal uptake is expressed as CF 1 CF = + q qmaxb qmax
(2)
where b is a constant related to the energy of adsorption/desorption and qmax is the maximum uptake.
RESULTS AND DISCUSSION Effects of biomass quantity and Cd concentration on biosorption The sorption rate of Cd by H. verticillata was very high. Most of the metal biosorption occurred during the first 3–5 min (Fig. 1). The sorption equilibrium was also achieved within a short period of 30 min. The equilibrium was reached within 20 min of contact similarly to that in the case of Zn, Pb and Cu adsorptions by C. demersum (16). There are many parameters that determine adsorption rate, such as the structural properties of the sorbent (e.g., porosity, surface, morphology and swelling degree), and the existence of other metal ions that compete for the same active complexation sites. The Cd and Cr adsorptions onto Wolffia globosa were very rapid and more efficient when low metal concentrations were tested (17). This rapid kinetics has practical significance because it facilitates the use of smaller reactor volumes ensuring efficiency and economy. A similar rapid metal biosorption has been reported by several researchers (18, 19) At a low Cd concentration (1 mg/l) the biosorption efficiency was very high (Fig. 2). As Cd concentration increased, the efficiency progressively decreased. At the same Cd concentration, the increase in the amount of biomass did not increase biosorption efficiency. Except at 100 mg/l, the biosorption efficiency of H. verticillata increased when biomass was increased. The rate of increase in the degree of Cd removal was not proportional to the increase in H. verticillata biomass. This may be attributed to the interference
FIG. 1. Time course of Cd removal by H. verticillata dry biomass. Conditions: 0.5 g; pH 5; 150 rpm; 25 ± 2°C; initial Cd concentration, 10 mg/l.
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FIG. 3. Effects of pH on biosorption of 10 mg/l of Cd solution using H. verticillata dry biomass. Conditions: 0.5 g; contact time, 1 h; 150 rpm; 25 ± 2°C. FIG. 2. Effects of weight of H. verticillata dry biomass on biosorption of Cd at different concentrations. Conditions: 150 rpm; contact time 30 min; pH 5; 25 ± 2°C. Symbols: empty bars, 1 mg/l; solid bars, 10 mg/l; gray bars, 100 mg/l.
between binding sites at higher metal concentrations (20). This interference is due to the net Cd ion availability around the microenvironment of the biomass and the electrostatic interaction between ions at nonlimiting sorption sites on the biomass, which make the ions compete for available sites. Hence, the adsorption capacity decreases with time (21). In contrast, the high Cd adsorption efficiency at a lower biomass was due to higher metal ion-to-biosorbent ratio. This ratio decreased as biosorbent quantity increased, which subsequently resulted in a decrease in Cd biosorption rate (20). Meanwhile, the increase in Cd concentration decreased Cd biosorption rate at all weights of Cicer arientinum dry biomass used. This decrease may be due to the saturation of the sorption sites (22). The efficiencies of Pb, Zn and Cr removals by Hemidesmus indicus were also reduced when the initial metal concentration was increased up to 250 mg/l. These observations can be explained by the fact that at very low concentrations of metal ions, the ratio of the sorptive surface area to the total metal available area is high; thus, there is a great chance for metal removal. When metal ion concentration is increased, binding sites become more rapidly saturated as the amount of biomass remains constant (23). Effect of pH The pH of a solution is one of the most important parameters on the biosorption of metal ions (24). Metal removal was significantly affected by the initial pH of the solution. The biosorption of Cd in a very highly acidic medium (pH 1) was observed to be negligible (Fig. 3). At pHs lower than 3, Cd removal was inhibited possibly as result of the competition between hydrogen and Cd ions for sorption sites, with an apparent preponderance of hydrogen ions (11). Cd removal reached equilibrium at pH 3 and maintained this state up to pH 9. However, pH 5 was found to be optimum for biosorption. Ion exchange is the main mechanism responsible for metal ion biosorption. The main functional groups in ion exchange reactions at neutral pH are the carboxyl group present in plant tissues. Metal binding probably occurs with weakly acidic carboxyl groups (pKa range of 3.5–5.5) of plant cell wall constituents (5). In the W. globosa-Cd system, the degree of metal biosorption also increased when the initial pH increased. The maximum ad-
FIG. 4. Linearized Langmuir adsorption isotherm of Cd for H. verticillata.
sorption of Cd occurred at an initial pH of 7 (17). Meanwhile, the effects of equilibrium pH on Cu sorptions onto P. lucens, S. hergozi, and E. crassipes were similar. Maximum removal was attained between pHs 5.5 and 6.6 when Cu was mainly in Cu2+ form. No sorption occurred at very low pHs when Cu was hydrolyzed to either neutral or negative species (5). Biosorption capacity The maximum Cd adsorption capacity, qmax, was fitted and calculated through the linearized form of the Langmuir isotherm. For H. verticillata dry biomass, qmax was 15.0 mg Cd/g dry weight, the binding constant (b) was 2.92 and the correlation coefficient (r2) was 0.997 (Fig. 4). The Langmuir equation assumes that (i) the solid surface presents a finite number of identical sites that are genetically uniform, (ii) there are no interactions between adsorbed species, indicating that the amount adsorbed has no influence on adsorption rate and, (iii) a monolayer is formed when the solid surface reaches saturation (11). Reusability of H. verticillata The efficient removal of adsorbed metals from H. verticillata biomass is necessary to ensure their long-term use for repeated extraction-elution cycles. It has been reported that HCl shows higher Cd desorption activity in black gram husk biomass than HNO3, H2SO4 and EDTA (22). Moreover, HCl is well documented as an efficient desorbing agent for Cd, U, Cu (25). Therefore, in this study, 0.1 M HCl was used as a desorbing agent in three cycles of Cd adsorption-desorption. After the first cycle, Cd biosorption by the H. verticillata biomass was re-
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TABLE 1. Adsorption-desorption of Cd by H. verticillata packed column using 0.1 M HCl as desorbing agent Dry biomass (g) 1 2 3
Cycle 1 2 3 1 2 3 1 2 3
Cd adsorption (%) 100 100 95.62 100 85.04 98.32 100 94.31 80.29
duced at all weights of plants (Table 1). The adsorption efficiency also gradually decreased at all weights with the lowest (80.3%) being in the third cycle of the 3-g biomass experiment. The reason for this is that HCl could destroy the structure of polysaccharides, which are binding sites for heavy metals, including the hydrogen bonding capacity of the biomass, simultaneously with the hydrolysis of polysaccharides (26). Fixed-bed column with continuous flow A fixed-bed column packed with H. verticillata biomass was designed to operate a continuous liquid flow system for Cd biosorption. Fixed-bed breakthrough curves at two different weights of dry biomass (0.5 and 1 g) were obtained to illustrate the capability of column operation (Fig. 5). The dry H. verticillata biomass showed good Cd sorption ability. The adsorption column containing 1 g (dry weight) of the biomass could purify 10 mg/l Cd solution even below the detection limit of 0.02 mg/l before the breakthrough occurred at both weights of dry biomass used. The Cd influent-effluent equilibria at zero sorption by H. verticillata at 0.5 and 1 g (dry weight) were reached at volumes of 4200 and 7800 ml, respectively. Other fixed-bed column studies have also demonstrated similar sorption efficiencies of algal, microbial and plant biomasses (22, 27, 28). For the determination of the interference caused by the presence of other divalent cations, Zn was added to the Cd solution, which was then passed through the column for sorption study. The presence of Zn had an antagonistic effect on Cd sorption (Fig. 6). The Cd breakthrough point of the mixed solution was at 3900 ml, whereas, that of the Cdalone solution with effluent Cd concentration was at 7800 ml. Many functional groups on the cell wall and cell membrane are not specific, and different cations compete for binding sites. It has been reported that metal removal is enhanced when the ionic radii of metal cations affect ion exchange and adsorption (29). The change in influent flow rate also had almost no effects on Cd removal due to the rapid sorption of Cd on the biosorbent (30). Conclusion This study showed that H. verticillata dry biomass could be used as an efficient biosorbent material for the treatment of Cd ion contaminants in aqueous solutions. Batch adsorption studies showed that H. verticillata can adsorb Cd based on the Langmuir model at a maximum adsorption capacity of 15.0 mg Cd/g dry weight. Adsorption capacity was pH-dependent and the kinetics of such adsorption was very rapid with an 80% biosorption efficiency
FIG. 5. Biosorption of Cd in fixed-bed column with continuous flow at different sorbent biomasses. Conditions: initial Cd concentration 10 mg/l; pH 5; actual flow rates 10.91 ml/min for 0.5 g and 9.88 ml/min for 1 g; 25 ±2°C. Symbols: diamonds, 0.5 g of biomass; circles, 1 g of biomass.
FIG. 6. Biosorption of Cd in mixed Cd and Zn solution in fixedbed column with continuous flow. Conditions: initial Cd and Zn concentration, 10 mg/l; pH 5; actual flow rate 12.96 ml/min; 1 g of dry biomass used; 25 ± 2°C. Symbols: triangles, Cd; squares, Cd + Zn.
occurring within 3 min. H. verticillata dry biomass can be used efficiently in fixed-bed operation. For the development of a continuous fixed-bed column bioreactor system for onsite operations, it is also necessary to determine its adsorption efficiency for other heavy metals. ACKNOWLEDGMENTS This work was supported by the Royal Golden Jubilee Ph.D. program of the Thailand Research Fund, and in part by the grants from Post-Graduate Education, Training and Research Program in Environmental Science, Technology and Management under Higher Education Development Project of the Commission on Higher Education, Ministry of Education, and Mahidol University, Bangkok, Thailand.
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