Properties of complexes of galactomannan of Leucaena leucocephala and Al3+, Cu2+ and Pb2+

Bioresource Technology 89 (2003) 63–73

Properties of complexes of galactomannan of Leucaena leucocephala and Al3þ, Cu2þ and Pb2þ Simone Cristina Lombardi a, Ana Lucia Ramalho Merc^e b

b,*

a PIPE, UFPR, LACTEC––Pr, CEP:81531-990, Brazil Laborat orio de Equilıbrio Quımico, DQ, Centro Polit ecnico, Universidade Federal do Paran a, CEP:81531-990, Curitiba-Pr-Brasil, Curitiba CP 19081, Brazil

Received 18 March 2002; received in revised form 16 December 2002; accepted 29 December 2002

Abstract The use of biopolymers in many industrial processes is on the increase. The different interactions of biopolymers and electrolytes either in aqueous solutions or in solid state provide different physico-chemical properties and a simple correlation cannot be established. In this study, in order to determine the properties of the complexes of galactomannan of Leucaena leucocephala (gal) with the metal ions Al3þ and Pb2þ , toxic elements and Cu2þ , essential, the logs of the binding constants of the complexes formed in the aqueous solutions were calculated. Their rheological properties, their thermal behavior, the infrared characteristics and shape and form of the films formed by those complexes in solid state were also determined. The aqueous solutions properties have shown a better complexation between gal and Al3þ . The species distribution diagrams have shown an existence of complex species going from acidic to basic pH values. Infrared spectra have proved the complexations as well as the viscosity studies. Thermal stabilities in general were smaller in the complexed species than in the native biopolymers and the films obtained from aqueous solutions showed for Cu2þ the most different morphology compared to the biopolymer itself. A use can be suggested of this biopolymer in environmental remediations besides its already stablished industrial uses. Ó 2003 Elsevier Science Ltd. All rights reserved. Keywords: Metal complexes; Potentiometric titrations; TG-DSC; Infrared; Scanning electron microscopy; Biopolymer; Speciation

1. Introduction Industrial processes generate large quantities of wastewater. The food industry is an example of excessive water use producing a proportional wastewater that could be recycled either to the process or be released to the environment. The adsorption and/or the filtration processes using biopolymer are reported in the literature using cellulosic fibrous materials for the treatment of wastewaters (Chen et al., 2000). Many physical properties in food preparations such as their clarity, gel-forming ability and stability, among others, depend mainly on the effect of the interactions between the biopolymers in solution. The balance of hydrophobic and Coulomb interactions between the species plays a major role. Depending on their nature,

*

Corresponding author. E-mail address: [email protected] (A.L.R. Merc^e). URL: http://www.quimica.ufpr.br/leq.

pH, ionic strength, temperature, concentration and other minor variables, the proteins can attract or repel the polysaccharides in solution. In the former case, supramolecular soluble complexes are often formed. In the latter case, thermodynamic incompatibility and hence phase separation can arise (Delben and Stefancich, 1998; Delben and Stefancich, 1997). Also the use of biopolymers in unit operations to reduce the concentration of heavy metal ions emitted to the environment is related in the literature (Mundkur and Watters, 1993). Potentiometric titrations can be used to assess the interactions and macromolecular changes for linear polyelectrolytes in solution with their own counterions and polymeric solutions containing counterions of different valencies (Mouginot et al., 2000; Benegas et al., 1998). Highly industrial activities, such as aluminum smelting activity, wood products that are treated with copperbased preservatives, mine tailings where aluminum is a byproduct, old nonferrous factories in Europe which led to areas highly contaminated with metals like Zn, Cd,

0960-8524/03/$ - see front matter Ó 2003 Elsevier Science Ltd. All rights reserved. doi:10.1016/S0960-8524(03)00012-9

64

S.C. Lombardi, A.L.R. Merc^e / Bioresource Technology 89 (2003) 63–73

Cu and Pb, among many others are also responsible for contaminated wastewaters and soil. There are many studies concerning bioremediation for the purpose of biocleaning these types of wastewaters (Leung et al., 2000; Pascucci and Kowalak, 1999; Hard et al., 1999) as well as contaminated soils (Naidja et al., 2000; Diels et al., 1999) with a focus in the role of mineral colloids present in soil. The use of other materials to adsorb metal ions has also been examined (Singh et al., 2000; Kim et al., 1999; Bailey et al., 1999; Handra and Rustgi, 1998). The study of the binding constants and the speciation in aqueous solutions could help to solve this problem. The potentiometric titration was chosen to generate the data between galactomannan from Leucaena leucocephala and the metal ions Cu2þ , Al3þ and Pb2þ . This is a technique previously reported in the literature to provide via a proper choice of mathematical models, the formation constants for the complexes found in the equilibria as well as their speciation according to variations in pH values for biopolymers (Merc^e et al., 1998, 2000, 2001a,b, 2002; Noleto et al., 2002). Viscosity is a measure of the frictional resistance that a flowing liquid offers to an applied shearing force. The coefficient of viscosity (g) is the proportionality constant between this shearing force and the velocity gradient in the liquid. This technique provides an insight into the size and shape of flexible macromolecules, although some difficulties are found in interpreting results. However the influence of electrolytes on the viscosimetric properties of macromolecule is well recognized since it was demonstrated that viscosity was decreased in the presence of salt. This reduction was undoubtedly caused by a decrease in degree of dissociation of ionizing groups and associated contraction of the molecules. Also the expansion of polyelectrolytes alone in solutions is due to the ionization of functional groups as a result of increases in pH and/or dilution effects (Clapp et al., 1989). It is known that physico-chemical effects are only observed either in natural or synthetic compounds in solution. The associated contraction of the macromolecule can be taken for granted that the metal ion introduced into an aqueous system, by complexing to the binding sites of the macromolecule, decreases the viscosity (Clapp et al., 1989). Simple relationships between structures of compounds in solid-state and in solutions are not expected since dissolution of the substances in water may change their structure and composition dramatically. For kinetically labile metal complexes, the presence of several species in equilibrium with one another in the same solution makes the picture even more complicated. This is the reason why complexes must be further studied to complement the physico-chemical properties exhibited by them in the solid state (Burger et al., 1995).

CH2OH HO OH O O HO CH2

OH

O

O

OH O

O

O

OH OH CH2OH

n Fig. 1. Structure for a galactomannan mannose:galactose ratio of 2:1.

The objective of this present work was to determine, to measure and to characterize the interaction between the galactomannan of L. leucocephala (gal––refer to Fig. 1) and the metal ions Al3þ , Cu2þ and Pb2þ both in aqueous and solid state using potentiometric titrations, thermogravimetry and differential scanning calorimetry (TG-DSC), scanning electron microscopy (SEM) and infrared (IR) analysis. The metal ions were chosen in order to evaluate the complexing ability of the biopolymer galactomannan in remediating wastewaters contaminated with Al3þ and Pb2þ , two toxic metal ions and Cu2þ which has also deleterious effects when in great concentrations (Fra usto da Silva and Williams, 1997). The studies were made in aqueous solutions and in the solid complexes extracted from those aqueous solutions at different pH values. The solid complexes of gal and Cu2þ complementary results of the previously reported ones (Merc^e et al., 1998) in aqueous solution, are presented.

2. Methods The galactomannan studied in this work was extracted from seeds of L. leucocephala and thoroughly purified according to the literature and was kindly provided by Professor Maria Rita Sierakowski ([email protected]), UFPR, Brazil (Merc^e et al., 1998, 2000). The alditol acetates assay (Blakeney et al., 1983; Wolfrom and Thompson, 1963) provided the average mannose:galactose ratio of 1.6:1 for this galactomannan. The potentiometric titrations were carried out under an inert atmosphere of water–KOH saturated nitrogen (White-Martins––Brazil) in a water jacketed vessel maintained at 25:0  0:1 °C by a thermostated bath (Microquımica––Brazil, MQBTC––99-20) and ionic strength 0.100 mol/l (KNO3 ––Merck––Brazil). Data

S.C. Lombardi, A.L.R. Merc^e / Bioresource Technology 89 (2003) 63–73

were obtained from aliquots of fresh prepared 5 g/l of galactomannan solutions. A metrohm manual burette, 0:04  0:2 ml, was used to deliver the titrant, KOH (Merck––Brazil) solution 0.1 mol/l, previously standardized with potassium hydrogen phthalate (Carlo Erba––Italy), and the pH values were measured by an Analyzer (Brazil) glass electrode 1A01 and an Analyzer calomel reference electrode model 3A41 connected to a Micronal (Brazil) pH meter B-375. Both electrodes were previously calibrated with a standardized HCl with controlled ionic strength and KOH solutions following the literature (Martell and Motekaitis, 1992). The protonation and the stability constants were calculated using the microcomputer program Best7 (Martell and Motekaitis, 1992) and the distribution species diagram were drawn with the aid of SPE program (Martell and Motekaitis, 1992) which uses as input the output of Best7 program. All titrations were performed in triplicate for each metal to ligand ratio and for each metal ion. Metal salts p.a. grade (AlCl3  6H2 O (Reagen, Brazil), Cu(NO3 )2  3H2 O (Baker––USA) and Pb(NO3 )2 ––Merck––Germany) were used and standardized by conventional EDTA titrations (Schwarzenbach and Flaschka, 1969 for Al3þ ––Merc^e et al., 1998 for Cu2þ ––except Pb2þ solution which was standardized by atomic absorption spectrometry, Perkin Elmer spectrometer, model 4100 (Norwalk, CT, USA)). The statistical deviation values of the binding constants were obtained by calculating the mean of the deviations of all experimental points of duplicate or triplicate potentiometric titrations. It was calculated by the Best7 microcomputer program (Martell and Motekaitis, 1992). The viscosity of aqueous solutions of 5 g/l of galactomannan of L. leucocephala and metal ions were measured in a Brookfield Rheometer (Middleboro, MA, USA), model LVDV-3, at 150 rpm (speed) with varying pH values using CP 51 spindle (cone-plate). The starting solution of galactomannan was acidified with HNO3 aqueous solutions until pH ¼ 2:0 and afterwards a proper mass of solid metal ion was added to this solution in order to obtain the ligand to metal ratios of 1:1, 2:1 and 3:1. The pH was increased with appropriate additions of KOH (0.1 mol/l). The values of viscosity were taken at various speed rates, ranging from 30 to 180 rpm. The solid complexes obtained by precipitation of aqueous solutions by increasing the pH values until formation of solid products, were centrifuged after addition of ethanol from the original solution containing one mole of metal ion to one, two and three moles of ligand, separated and air dried accordingly (Merc^e et al., 2001a). The starting aqueous galactomannan solution was 5 g/l, the metal ions were added so as to give the required metal to ligand ratios, and the pH values were raised until precipitation occurred. Those pH values

65

were for Al3þ , ligand to metal ratios of 1:1, 2:1 and 3:1, 4.52, 6.75 and 8.50, respectively; for Cu2þ , ligand to metal ratios of 1:1 and 2:1, 7.12 and 8.88, respectively, and for aqueous solutions of Pb2þ and galactomannans, in the ligand to metal ratios of 1:1 and 1:2, the final pH values were 5.50 and 8.10, respectively. These solid complexes were used in the infrared and thermal analysis. The IR spectra were recorded with a FTIR doublebeam spectrometer (Bomem––Hartmann & Braun, Quebec, Que., Canada) in a KBr (Merck––USA, spectroscopic grade) matrix which always contained 1% m/m of sample in an attempt to perform quantitative analysis. Samples were scanned 16 times between 450 and 4050 cm1 . The thermogravimetry-differential scanning calorimetry (TG-DSC) analysis were recorded in a simultaneous thermal analysis STA 409 EP (Netzsch, Selb, Bayern, Germany) under air, from 21 to 520 °C, 2 °C/min, using an open cylindrical aluminum sample pans (4 mm diameter, 2 mm high). For the scanning electronic microscopy, solutions of galactomannan and the metal ion in the same ratios as used in the potentiometric titrations were dried on a thin glass plate on which a film was deposited. Aqueous solutions of 5 g/l of galactomannan of L. leucocephala and each metal ion at ionic strength of 0.10 mol/l (KNO3 ) were dried at room temperature on glass plates. For the metal ion Al3þ , the ligand to metal ratios were 1:1, 2:1 and 3:1 with the pH values adjusted to 3.15, 3.14 and 3.31, respectively. For Cu2þ , ligand to metal ratios of the solutions were 1:1, 2:1 at pH values of 3.68 and 4.06, respectively, and for the metal ion Pb2þ , ligand to metal ratios of 1:1 and 2:1, pH values of 3.30 and 3.07, respectively. The samples were coated for 1 min with gold to obtain a uniform thickness and scanning electron micrographs were taken using a Philips SEM model 505 microscope (Eindhoven, MD, Netherland). 2.1. Data treatment For the galactomannan studied, the mannose:galactose average ratio of 1.6:1, was treated as a set of monomers in the calculations performed to obtain the binding constants using the protonation constants values in the literature for either the monomer mannose or galactose (Martell and Smith, 2001). The protonation equilibrium of the monomeric sugar unit, referred from now on as L, of the biopolymer with one –OH group represented is shown in Eq. (1). 

O–L þ Hþ ¢ HO–L

log Ka1 ¼ 12:3  0:1

ð1Þ

or L þ H ¢ HL

log Ka1 ¼ 12:3  0:1

0

ð1 Þ

Eqs. (2) and (3) show a monomeric sugar unit having two –OH deprotonated groups, the second protonation

66

S.C. Lombardi, A.L.R. Merc^e / Bioresource Technology 89 (2003) 63–73

constant obtained by mathematical calculations with program Best7. 

O–L–O þ Hþ ¢ HO–L–O

log Ka1 ¼ 12:3  0:1 ð2Þ

or L þ H ¢ HL

log Ka1 ¼ 12:3  0:1

HO–L–O þ Hþ ¢ HO–L–OH

0

ð2 Þ

log Ka2 ¼ 10:6  0:1 ð3Þ

or HL þ H ¢ H2 L

log Ka2 ¼ 10:6  0:1

0

ð3 Þ

Taking into account that all experimental errors are minimized and a special procedure for the pH electrodes standardization is done, the microcomputer program Best7 (Martell and Motekaitis, 1992) can be employed to treat the data of potentiometric titrations. Best7 carries out calculations with an algorithm which calculates pH directly and minimizes the sum of the weighted squares of  log½Hþ  residuals. Program Best basically solves for the set of equilibrium constants corresponding to the model selected, making it possible to explore all aspects and variations of the model. In the calculation steps, the sigma fit, represented as the statistical variation in the equilibrium constant, is calculated along with the main algorithm and is the mean result of the weighted sum of the squares of the deviations in pH values of all points obtained compared to the calculated ones, in how many performed potentiometric titrations replicates. The millimoles employed in the calculations were calculated using the molecular weight of a monomer as the alditol acetate assay did not exhibit any of the other main constituents of the biopolymer. The hydrolysis constants of the metal ions were obtained from published values (Baes and Mesmer, 1976). The BEST7

output results are used as input parameters to program SPE (Martell and Motekaitis, 1992) to draw the distributions diagrams. For the metal ion Al3þ the mathematical model that best suited the equilibrium data was the taking of two protons by this metal ion of a monomeric sugar unit of the biopolymer when forming the complexed species (refer to Eqs. (2) and (3)). Cu2þ also displaces two protons from the ligand (Merc^e et al., 1998) while the metal ion Pb2þ could displace only one proton of gal (refer to Eq. (1)). These two mathematical models employed have been described previously (Merc^e et al., 1998, 2000, 2001a; Martell and Hancock, 1996; Ciardelli et al., 1995).

3. Results and discussion The potentiometric pH profiles of galactomannan and the metal ions Al3þ and Pb2þ are depicted in Figs. 2 and 3. For Cu2þ refer to previous reports (Merc^e et al., 1998, 2001a). The binding constants for the complexes species are in Table 1. It can be seen from these values that the strongest Lewis acid is the one which complexes the best, followed by an intermediate and the least strong, respectively being, Al3þ , Cu2þ and Pb2þ for the species ML. The logarithm value of the binding constants for the ML2 species was almost the same value as for the ML species, probably because there is no significant steric hindrance to the formation of ML2 . In the case of Al3þ this assumption is reinforced by the detection of ML3 . The species distribution diagrams in Figs. 4 and 5, gal and Al3þ and gal and Pb2þ , respectively, revealed the formation of complexed species in the acid region for Al3þ , starting at pH around 4.0, with ML and existing in at least one complexed species in all pH range until 11.0. For gal and Pb2þ (Fig. 5), the formation of complexed

Fig. 2. Potentiometric pH profile of galactomannan 0.09 mmol of L. leucocephala (gal) and in the presence of the metal ion Al3þ , ligand to metal ratios of 1:1, 2:1 and 3:1. T ¼ 25:0 °C; I ¼ 0:100 mol/l (KNO3 ).

S.C. Lombardi, A.L.R. Merc^e / Bioresource Technology 89 (2003) 63–73

67

12

10

pH

8

6

4

2 0.00

0.05

0.10

0.15

0.20

0.25

0.30

a (n mole KOH/n mole ligand) gal 0.3 mmol

gal 1:1 Pb2+

gal 2:1 Pb2+

Fig. 3. Potentiometric pH profile of galactomannan 0.3 mmol of L. leucocephala (gal) and in the presence of the metal ion Pb2þ , in the ligand to metal ratios of 1:1 and 2:1. T ¼ 25:0 °C; I ¼ 0:100 mol/l (KNO3 ).

Table 1 Logarithms for the binding constants of the complexes between galactomannan and Al3þ , Cu2þ (Merc^e et al., 1998) and Pb2þ log K––galactomannan of L. leucocephala

Al3þ

Pb2þ

Cu2þ

½ML=½M ½L ðb1 Þ ½ML2 =½ML ½L ðb2 Þ ½ML3 =½ML2  ½L ðb3 Þ

16.53  0.09 (16.53) 16.37  0.09 (32.90) 11.20  0.09 (44.11)

7.90  0.09 (7.90) 7.88  0.09 (15.78) n.d.

15.6  0.1 (15.6) n.d. n.d.

n.d.: not detected.

Fig. 4. Species distribution diagram of 0.09 mmol solution of galactomannan (L) with Al3þ (M, 0.03 mmol) of pH values from 2.0 to 10.0. Percentage of a species present, with the metal ions set at 100%. Hx represents (OH ).

species initiated only after pH 5.0, and ML2 existed until at least pH 10.0. The species distribution of Cu2þ and gal revealed a similar behavior (Merc^e et al., 2001a).

In Figs. 6–8 the thermal profiles of the solid complexes of galactomannan and the metal ions Al3þ , Pb2þ and Cu2þ are shown, respectively. The final value of mass percentage for the biopolymer alone is a negative

68

S.C. Lombardi, A.L.R. Merc^e / Bioresource Technology 89 (2003) 63–73

Fig. 5. Species distribution diagram of 0.3 mmol solution of galactomannan (L) with Pb2þ (M, 0.15 mmol) of pH values from 2.0 to 10.0. Percentage of a species present, with the metal ions set at 100%. Hx represents (OH ).

Fig. 6. Thermal profile––TG-DSC of the solid products extracted from an aqueous solution of pH ¼ 4:5–5.5 of 10 moles of galactomannan and Al3þ in the molar ratios ligand to metal 1:1 and 2:1.

value due to a bias in the equipment in correcting the base line. The main observed thermal effects (Merc^e et al., 2001a,b) numbered from 1 to 4 can be described as follows with the respective temperatures stated in Table 2. In all cases, after the buoyancy effects on the TG balance, at the very beginning of the run, there was an endothermic loss of adsorbed water in the biopolymer and its complexes (numbered 1). The second and third exothermic transitions could be due to changes in conformation and breakage of some branches of the biopolymer between 180 and 310 °C for all cases, as the TG associated curves show a substantial mass loss. A final exothermic transition (numbered 4) occurred due to oxidative degradation of the sample. Final destruction occurred in the temperature range of 390–440 °C. The

boundaries of the four steps depended on the nature of the complexes. For Al3þ (Fig. 6), the thermal events shifted to smaller temperatures than in gal alone. The thermal events 2 and 3 partially disappeared in the complexes. The same events are presented by gal and gal complexed to Cu2þ (Fig. 7) in the ligand to metal ratio of 1:1, but in smaller temperatures. In the ligand to metal 2:1 ratio, there was only the oxidative degradation step (390 °C–– number 4). For Pb2þ (Fig. 8), the thermal event number 2 disappeared in both metal to ligand ratios studied. The degradation temperature and thermal event number 3 shifted to smaller values than for the biopolymer alone. In all complexed system studied after the oxidation degradation step, the TG curves presented a varied

S.C. Lombardi, A.L.R. Merc^e / Bioresource Technology 89 (2003) 63–73

69

Fig. 7. Thermal profile––TG-DSC of the solid products extracted from an aqueous solution of pH ¼ 6:5–7.0 of 10 moles of galactomannan and Cu2þ in the molar ratios ligand to metal 1:1 and 2:1. In the abscissa it is represented the time.

Fig. 8. Thermal profile––TG-DSC of the solid products extracted from an aqueous solution of pH ¼ 6:5–7.0 of 10 moles of galactomannan and Pb2þ in the molar ratios ligand to metal 1:1 and 2:1.

Table 2 Temperatures of thermal events of gal and the complexes of gal and Al3þ , Cu2þ and Pb2þ Event number

Gal alone (°C)

Gal–Al3þ (°C) a

1 2 3 4

55 280 300 430

n.d.: not detected. a Ligand to metal ratio.

Gal–Pb2þ (°C)

Gal–Cu2þ (°C)

1:1

2:1

1:1

2:1

1:1

2:1

n.d. n.d. 415

240 n.d. 395

n.d. 245 405

n.d. 245 390

190 225 390

n.d. n.d. 390

70

S.C. Lombardi, A.L.R. Merc^e / Bioresource Technology 89 (2003) 63–73

Table 3 Values of viscosity (g  0:5) (mPa) at 50 rpm (speed) of aqueous solutions of 5 g/l of galactomannan of L. leucocephala (gal), with varying pH values and metal ions pH range

Al3þ + Leu

Cu2þ + Leu

Pb2þ + Leu

1:1

2:1

3:1

1:1

2:1

1:1

2:1

3.0–3.5 3.5–4.0 4.0–4.5 4.5–5.0 5.0–5.5 5.5–6.0

30.2 27.4 n.d. n.d. n.d. n.d.

33.8 30.0 n.d. n.d. n.d. n.d.

28.7 25.9 n.d. n.d. n.d. n.d.

26.4 26.1 25.4 24.0 24.0 n.d

24. 0 23.3 23.8 22.6 21.8 21.0

29.7 25.2 20.0 14.8 14.5 n.d.

29.4 23.7 22.4 19.7 17.5 n.d.

n.d.: not determined.

percentage of metal oxides left behind depending on the metal ion employed. This was expected since there is still organic matter and some oxides left. The values for the viscosity (mPa) of the aqueous solutions of galactomannans and the metal ions in different ligand to metal ratios obtained at a speed of 50 rpm are presented in Table 3. The values for the viscosity of the galactomannan alone and metal ions solutions are 16.0 and 1 mPa, respectively. The various viscosity values obtained with varying speeds exhibited nearly identical patterns, but the best set of values were at 50 rpm where the formation of insoluble products were more delayed. The complexation between the metal ions studied and gal can be seen in the decreasing viscosity values as the pH values were raised to the values where complexes were formed and insolubility products of the system did not prevent measurements. It is known that solution parameters such as pH, ionic strength, the presence of metal ions to complex with the polymer, concentration of the biopolymer and the nature of the solvent can determine the size and shape of the particles. The micrographs at a magnification of 2300 of gal alone and gal and the metal complexes with Al3þ and Cu2þ , ligand to metal ratios of 1:1,

Fig. 9. Micrograph of gal, magnification of 2300 .

are shown in Figs. 9–11, respectively. The shape and particle arrangements in the SEM micrographs within the magnification used to obtain the images of the obtained films were allowed to dry from the aqueous so-

Fig. 10. Micrograph of gal and Al3þ , ligand to metal ratio of 1:1, magnification of 2300 .

Fig. 11. Micrograph of gal and Cu2þ , ligand to metal ratio of 1:1, magnification of 2300 . The bubble in the right corner was accidentally formed when the electron beam reached the film.

S.C. Lombardi, A.L.R. Merc^e / Bioresource Technology 89 (2003) 63–73

lutions at a certain pH value and varying metal ion concentrations. These resulted in a sheet-like structure of varying thickness exhibiting homogeneous grain-like particles in the obtained films for gal, gal and Pb2þ and more homogeneous in gal and Al3þ . For gal and Cu2þ it was seen straight marks on the obtained film (Fig. 11). The water solubility of gal and the metal ions played an important role in the final obtained film. The pH values of the starting solutions were chosen according to the distribution species diagrams obtained using potentiometric titrations. These showed the presence of at least one complexed species. These chosen pH values were sometimes on the borderline where insoluble and/or hydrolysis products were formed, so when the metal ion concentration increased the formation of those insoluble products entrapped in the centre of the glass plate. This also was favored by the supersaturation reached by the solution during air drying. It was seen that the homogeneity of the film formed was not affected in other areas of the glass plate but the centre in all ligand to metal ratios obtained, according to the stoichiometry of the complexed species found. The IR technique, albeit not powerful when analysing carbohydrates due to accidental degeneracy of the spectra, can provide some structural information when comparing the spectra of the native biopolymer and when in the presence of metal ions. It was recently demonstrated (Merc^e et al., 2001a; Rao et al., 2000) how elucidative an IR spectra can potentially be in the complexation of both saccharides and/or ascorbic acid with iron based on FTIR and UV–Vis studies even when only one difference in vibration of one substituent in the complexed biopolymer structure compared to the biopolymer alone.

71

In monosaccharides the primary hydroxyl substituents are the most chemically reactive. The IR obtained spectra were mainly an attempt to quantitatively monitor the decrease in the –OH stretching band, around 3400 cm1 . Although water absorption by either the solid complexes or by the KBr pellets could hinder the IR effect of diminishing the –OH band due to a sharing of this binding site with a Lewis acid, in all obtained IR spectra with gal and the metal ions studied the intensity of the –OH band decreased. The region of 1300–1500 cm1 covers the scissoring and wagging type of bending modes of –CH2 and –CH3 groups, as well as to the plane bending, hydroxyl modes. The –CH2 and –CH3 groups do not participate in hydrogen bond formation but –CH2 is a part of the –CH2 OH group, which produces hydrogen bonds. Also the region of 1640–1650 cm1 refers to water in aqueous derived solutions of carbohydrates, and 1384–1390 cm1 refers to inorganic compounds. The 1384 cm1 inorganic assigned band, seen in the IR spectra of Cu2þ and Pb2þ , was not present in the gal––Al3þ solid studied complexes. This is due to a great concentration of the counter ion NO 3 employed in the making of the complexes where nitrate p.a. salts were used, except for the metal ion aluminum, for which chloride derived salt were used. Also it can be inferred that gal –OH groups have a greater affinity to this latter harder metal ion and that the obtained solid complexes were isolated under such experimental conditions not favoring the formation of insoluble hydrolysis products. The IR spectra of gal––Al3þ have shown a decrease in the –OH band, although due to absorption of moisture the solid 1:1 ligand to metal ratio appeared greater in the

100

80

%T

60

40

20

0 500

1000

1500

2000

2500

3000

3500

wavelength (1/cm) gal

gal 1:1 Cu2+

gal 2:1 Cu2+

Fig. 12. IR spectra of gal and Cu2þ in ligand to metal ratios of 1:1 and 2:1.

4000

72

S.C. Lombardi, A.L.R. Merc^e / Bioresource Technology 89 (2003) 63–73

100 90 80 70

%T

60 50 40 30 20 10 0 500

1000

1500

2000

2500

3000

3500

4000

wavelength (1/cm) gal

gal 1:1 Pb2+

gal 2:1 Pb2+

Fig. 13. IR spectra of gal and gal and Pb2þ in ligand to metal ratio of 1:1 and 2:1.

3400 cm1 region than the 2:1 ligand to metal ratio, however still being smaller than the gal alone. The infrared spectra of gal alone and gal and Cu2þ are shown in Fig. 12. For some unexpected reason the solid extracted from the aqueous solution in which metal to ligand ratio was 1:1 and the pH adjusted to 7.12, became very dark. This indicated that almost all metal ion present hydrolysed, became insoluble or the resulting solid could not be dried properly, yielding a biased quantitative IR spectra. The 2:1 ligand to metal ratio however, exhibited a decrease in the –OH stretching band and a variation in the shape of the 2930 and 2975 cm1 band. The infrared spectra of the complexes of gal and Pb2þ as well as native gal are shown in Fig. 13. There is an expected decrease in the intensity of the –OH band when one goes from the ligand to metal ratio of 2:1 to 1:1, as in the 1:1 ligand to metal ratio more –OH groups are involved in the complexation leaving less –OH groups prone to vibrate. Also variations in the shape and width of the 2930 and 2975 cm1 bands were observed. The region between 2850 and 2950 cm1 , also indicative of –CH and –CH2 stretching bands is changed in all three obtained spectra. This is probably due because the –CH2 groups, although not being direct participants in hydrogen bonds formation, are part of the primary alcohol of the monomeric sugar units of the biopolymer (–CH2 OH) which produces hydrogen bonds. As the complexation occurs this special wavenumber region is changed, indicating that the main complexation site is through C-6 (Noleto et al., 2002; Mukhopadhyay et al., 2000; Hanna et al., 1999). Based on data of potentiometric titration and on the IR results, the structure in Fig. 14 can be proposed for a complexed gal where the C-6 –OH basic site is always involved in the formation of any complexed species.

Fig. 14. Proposed structure for a galactomannan man:gal ratio of 2:1 complexed to a metal ion in a 1:1 ligand to metal ratio.

The physico-chemical properties exhibited by gal and the metal ions Al3þ , Cu2þ and Pb2þ enhance the possibility of using gal as an effective chelating agent for either industrial purposes such as paint and ink industries or in remediation of the studied metal ions in wasterwaters (Ng et al., 2002; Jianlong et al., 2000; Petruzzelli et al., 2000; Berbenni et al., 2000). Also the residual Al3þ used in the municipal water treatments, could be removed by employing gal to complex this excess toxic metal ion that is eventually present in the so called cleaned water. Those aspects will be the subject of future studies.

Acknowledgements We thank UFPR, Lactec––Pr for the infrared analysis and the Microscopy Laboratory of UFPR, specially

S.C. Lombardi, A.L.R. Merc^e / Bioresource Technology 89 (2003) 63–73

Professor Daura Regina Eiras Stofella, and Professor Maria Rita Sierakowski for the viscosity analysis.

References Baes Jr., C.F., Mesmer, R.E., 1976. The Hydrolysis of Cations. JohnWiley & Sons, New York, USA. Bailey, S.E., Olin, T.J., Bricka, M., Adrian, D.D., 1999. A review of potentially low-cost sorbents for heavy metals. Wat. Res. 33, 2469– 2479. Benegas, J.C., Cleven, R.F.M.J., van den Hoop, M.A.G.T., 1998. Potentiometric titration of poly (acrylic acid) in mixed counterion systems. Chemical binding of Cd ions. Anal. Chim. Acta 369, 109– 114. Berbenni, P., Pollice, A., Canzizni, R., Stabile, L., Nobili, F., 2000. Removal of iron and manganese from hydrocarbon contaminated groundwaters. Bioresour. Technol. 74, 109–114. Blakeney, A.B., Harris, P.J., Henry, R.J., Stone, B.A., 1983. A simple and rapid preparation of alditol acetates for monosaccharide analysis. Carbohydr. Res. 113, 291–299. Burger, K., Nagy, L., Gyurcsik, B., 1995. The structure of metal complexes of small models of glycoproteins in aqueous solution. J. Mol. Liq. 65/66, 213–219. Chen, L.A., Carbonell, R.G., Serad, G.A., 2000. Recovery of proteins and other biological compounds from food processing wastewaters using fibrous materials and polyelectrolytes. Wat. Res. 34 (2), 510– 518. Ciardelli, F., Tsuchida, E., W€ ohrle, D. (Eds.), 1995. Macromolecule–– Metal Complexes. Springer, Berlin. Clapp, C.E., Emerson, W.W., Olness, A.E., 1989. Sizes and shapes of humic substances by viscosity measurements. In: Hayes, M.H.B., MacCarthy, P., Malcolm, R.L., Swift, R.S. (Eds.), Humic Substances. II. In Search of Structure. John Wiley and Sons, Great Britain, pp. 497–514 (Chapter 17). Delben, F., Stefancich, S., 1997. Interaction of food proteins with polysaccharides. I. Properties upon mixing. J. Food Eng. Sci. 31, 325–346. Delben, F., Stefancich, S., 1998. Interaction of food polysaccharides with ovalbumin. Food Hydrocoll. 12 (3), 291–299. Diels, L., De Smet, M., Hooyberghs, L., Corbisier, P., 1999. Heavy metals bioremediation of soil. Mol. Biotechnol. 12 (2), 149–158. Fra usto da Silva, J.J.R., Williams, R.J.P., 1997. The Biological Chemistry of the Elements. The Inorganic Chemistry of the Life. Clarendon Press, Oxford, UK, pp. 266, 397–399, 515–516, 536– 539, 542, 546–547, 549. Handra, R.C., Rustgi, R., 1998. Biodegradable polymers. Prog. Polym. Sci. 23, 1273–1335. Hanna, A.A., Basta, A.H., El-Saied, H., Abadir, I.F., 1999. Thermal properties of cellulose acetate and its complexes with some transition metals. Polym. Degrad. Stab. 63, 293–296. Hard, B.C., Walther, C., Babel, W., 1999. Sorption of aluminum by sulfate-reducing bacteria isolated from uranium mine tailings. Geomicrobiol. J. 16 (4), 267–275. Jianlong, W., Hanchang, S., Yi, Q., 2000. Wastewater treatment in a hybrid reactor (HBR): effect of organic loading rates. Process Biochem. 36, 297–303. Kim, Y.H., Park, J.Y., Yoo, Y.J., Kwak, J.W., 1999. Removal of Lead using xanthated marine brown alga, Undaria pinnatifida. Process Biochem. 34, 647–652. Leung, W.C., Wong, M.F., Chua, H., Lo, W., Yu, P.H.F., Leung, C.K., 2000. Removal and recovery of heavy metals by bacteria isolated from activated sludge treating industrial effluents and municipal wastewaters. Water Sci. Technol. 41 (12), 233–240. Martell, A.E., Hancock, R.D., 1996. In: Fackler Jr., J.P. (Ed.), Metal Complexes in Aqueous Solutions. Plenum Press, New York, p. 205.

73

Martell, A., Motekaitis, R., 1992. The Determination and Use of Stability Constants. VCH, New York. Martell, A.E., Smith, R.M., 2001. NIST Critical Stability Constants of Metal Complexes, version 6.0; NIST Database 46, Gaithersburg, MD, USA. Merc^e, A.L.R., Lombardi, S.C., Mangrich, A.S., Reicher, F., Szpoganicz, B., Sierakowski, M.R., 1998. Equilibrium studies of galactomannan of Cassia fastuosa and Leucaena leucocephala and Cu2þ using potentiometry and EPR spectroscopy. Carbohydr. Pol. 35 (1), 13–20. Merc^e, A.L.R., Fernandes, E., Mangrich, A.S., Sierakowski, M.R., 2000. Evaluation of the complexes of galactomannan of Leucaena leucocephala and Co2þ , Mn2þ , Ni2þ and Zn2þ . J. Braz. Chem. Soc. 11 (3), 224–231. Merc^e, A.L.R., Landaluze, J.S., Mangrich, A.S., Szpoganicz, B., Sierakowski, M.R., 2001a. Complexes of arabinogalactan of Pereskia aculeata and Co2þ , Cu2þ , Mn2þ and Ni2þ . Bioresour. Technol. 76, 29–37. Merc^e, A.L.R., Fernandes, E., Mangrich, A.S., Sierakowski, M.R., Szpoganicz, B., 2001b. Fe(III)––galactomannan solid and aqueous complexes. Potentiometric, EPR spectroscopy and thermal data. J. Braz. Chem. Soc. 12 (6), 791–798. Merc^e, A.L.R., Carrera, L.C.M., Romanholi, L.K.S., Recio, M.A.L., 2002. Aqueous and solid complexes of iron(III) with hyaluronic acid. Potentiometric titrations and infrared spectroscopy studies. J. Inorg. Biochem. 89, 212–218. Mouginot, Y., Morlay, C., Cromer, M., Vittori, O., 2000. Potentiometric study of copper(II) and nickel(II) complexation by a cross-linked poly (acrylic acid) gel. Anal. Chim. Acta 407, 337– 345. Mukhopadhyay, A., Kolehmainen, E., Rao, C.P., 2000. Lanthanide–– saccharide chemistry: synthesis and characterization of Ce(III)–– saccharide complexes. Carbohydr. Res. 324, 30–37. Mundkur, S.D., Watters, J.C., 1993. Polyelectrolyte-enhanced ultrafiltartion of copper from a waste stream. Sep. Sci. Technol. 28 (5), 1157–1168. Naidja, A., Huang, P.M., Bollag, J.M., 2000. Enzyme––clay interactions and their impact on transformations of natural and anthropogenic organic compounds in soil. J. Environ. Qual. 29 (3), 677–691. Ng, J.C.Y., Cheung, W.H., MacKay, G., 2002. Equilibrium studies of the sorption of Cu(II) ions onto chitosan. J. Colloid Interf. Sci. 255, 64–74. Noleto, G.R., Merc^e, A.L.R., Iacomini, M., Gorin, P.J.A., Soccol, V.T., Oliveira, M.B.M., 2002. Effects of a Lichen galactomannan and its vanadyl (IV) complexes on peritoneal macrophages and leishmanicidal activity. Mol. Cell. Biochem. 233, 73–83. Pascucci, P.R., Kowalak, A.D., 1999. Metal distribution in complexes with Chlorella vulgaris in seawater and wastewater. Water Environ. Res. 71 (6), 1165–1170. Petruzzelli, D., Volpe, A., Di Pinto, A.C., Passino, R., 2000. Conservative technologies for environmental protection based on the use of reactive polymers. React. Funct. Polym. 45, 95–107. Rao, C.P., Geetha, K., Raghauan, M.S.S., Sreedhara, A., Tokunaga, K., Yamaguchi, T., Jadhav, V., Ganesh, K.N., Krishnamoorthy, T., Ramaiah, K.V.A., Bhattacharyya, R.K., 2000. Transition metal saccharide chemistry and biology: syntheses, characterization, solution stability and putative bio-relevant studies of iron-saccharide complexes. Inorg. Chim. Acta 297, 373–382. Schwarzenbach, G., Flaschka, H., 1969. Complexometric Titrations. Methuen & Co Ltd, UK, p. 192. Singh, S., Pradhan, S., Rai, L.C., 2000. Metal removal from single and multimetallic systems by different biosorbent materials as evaluated by differential pulse anodic stripping voltammetry. Process Biochem. 36, 175–182. Wolfrom, M.L., Thompson, A., 1963. Acetylation. Methods Carbohydr. Chem., vol. 2, New York, pp. 211–215.