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Journal of Stress Physiology & Biochemistry, Vol. 13, No. 4, 2017, pp. 81-87 ISSN 1997-0838 Original Text Copyright © 2017 by Basel Saleh
ORIGINAL ARTICLE
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Cadmium Biosorption Investigation from Aqueous Solutions with Ulva lactuca (Chlorophyta) and Padina pavonica (Phaeophyta) Seaweeds
Basel Saleh
Department of Molecular Biology and Biotechnology, Atomic Energy Commission of Syria, P.O. Box 6091, Damascus, Syria
*E-Mail: astientific@aec.org.sy
Received September 24, 2017
Fourier transform infrared spectroscopy (FTIR) analysis has been employed to identify various functional groups involved in cadmium biosorption into Ulva lactuca (Chlorophyta) and Padina pavonica (Phaeophyta) seaweeds exposed to different cadmium (Cd) (0, 2.5, 5 and 10 mg/L) concentrations for 4 days. Data revealed that Cd stress induced chemical structure alteration into studied seaweeds. Overall, for U. lactuca, carboxylic acids (C-O) & amides (C-N stretch) groups and aromatics groups (C-C stretch) showed a shift in wavelength with Cd stress referring that they may be play an important role in Cd biosorption process. Whereas, for P. pavonica, amides groups (C=O stretch), alcohols (O-H stretch) and phenols (H-bonded) groups showed a slight shift in wavelength indicating that these peaks could involve in Cd biosorption process. Thereby, FTIR analysis seems to be a potent tool for screening chemical structure alteration induced by Cd stress in marine ecosystems.
Key words: Seaweeds, Cadmium, Fourier transform infrared spectroscopy, Functional groups
Heavy metals impact on living organisms could be manifested by morphological, physiological, biochemical and molecular levels (Saleh 2017a). Biochemical analytical methods could help us to understanding binding metals mechanisms. This could be varied according to different factors e.g. species, metal ion nature, age and cell walls composition (Ovza and Ovez 2013). For seaweeds, this investigation seems to be simplest because seaweeds biomass was directly in contact with chemical solution and thereby, rapidly respond.
Cadmium (Cd) impact among various heavy metals, on living organisms in particularly on seaweeds, has extensively investigated in many researches (D'Souza et al., 2008; Lee et al., 2011; Ova and Ovez 2013; Saleh 2016a; b; c). Fourier transform infrared spectroscopy (FTIR) is considered as one of fewer potential valuable tools for screening and identification of various functional groups and organic compounds in plants and seaweeds (Lammers et al., 2009; Radhika and Mohaideen 2015). Its utility could be manifested by its simplicity and rapidity, nondestructive methods, direct analysis with no request of special sample preparation with low cost. Their advantages make it one of the most recent analytical techniques applied in characterization of different groups involved in plants and seaweeds and their interaction with wide ranges of heavy metal ions (Murphy et al., 2007; Ferreira et al., 2011; Fakhry 2013; Ghoneim et al., 2014; Radhika and Mohaideen 2015; Saleh 2017b).
To our knowledge, little informations have been published regarding chemical structure alteration in U. lactuca (Chlorophyta) and P. pavonica (Phaeophyta) seaweeds via Cd stress. Thereby, the current investigation aimed to characterize spectra of untreated compared to Cd-treated seaweeds using FTIR and consequently determine the main functional groups that could involve in Cd biosorption process.
MATERIALS AND METHODS
Seaweeds sampling
U. lactuca and P. pavonica samples were collected along the Syrian coast of the Mediterranean Sea. Samples collection was carried out from 34°37'734"N
latitude and 38°29'766''E longitude at 4 km North Lattakia - Syria. Samples were collected manually with disposable gloves; biomass was washed with seawater and then transported within a flask with 5 L seawater. Application of cadmium stress
Seaweeds cultivation, cadmium (0, 2.5, 5 and 10 mg/L Cd) treatment and experimental conditions were as recently reported by Saleh (2016a). Fourier transform infrared spectroscopy (FIIR) technique
A hundred mg of seaweed tissues for the both untreated and Cd-treated seaweeds were ground and 5 mL of 80% acetone were added. Tubes were kept at 4°C for 24 h, then they were centrifuged at 1400 g/ 2 min. Extracts were filtrated using Whatman filter papers as reported by Saleh (2015). These filtrated extracts were used as template for FTIR measurement using NXR FTIR (Thermo, USA) instruments.
RESULTS AND DISCUSSION
Interaction of living organisms with heavy metals could be investigated based on morphological, physiological, molecular and biochemical methods. In the current investigation we focusing on functional groups changes in comparative study between untreated and Cd-treated U. lactuca and P. pavonica seaweeds under different Cd concentrations (0, 2.5, 5 and 10 mg/L Cd) using FTIR analysis.
Our data showed that Cd stress induced some changes in functional groups into U. lactuca (Fig. 1) and P. pavonica (Fig. 2) seaweeds.
From data presented in Fig. 1 and Fig. 2, estimated peaks and functional groups of untreated and Cd-treated seaweeds were summarized in Table 1 for U. lactuca and in Table 2 for P. pavonica.
In the case of U. lactuca, C-Cl stretch in alkyl halides groups and C-C stretch in aromatics groups (1420.5 cm-1) did not show shift in peak referring that these groups have not involved in Cd biosorption. Whereas, C=O stretch in amides groups, C-H stretch in alkanes groups and O-H stretch & H-bonded in alcohols and phenols groups slightly shifted indicating that all the previous groups could not involve in Cd
biosorption. Inversely to other groups such as C-Br stretch in alkyl halides groups, C-N stretch in aliphatic amines and carboxylic acids groups, C-C stretch in aromatics groups (1448 cm-1) seem to play a benefit role via Cd biosorption due to the noticeable shifting in wavelength.
As for P. pavonica, five [(=C-H oop bend (712.8 and 874.6 cm-1) in aromatics groups, C-O & C-N stretch in carboxylic acids and amides groups, C-C stretch in aromatics groups and C-H stretch in alkanes groups)] out seven peaks showed no differences appeared following Cd treatment indicating that these peaks have not involved in Cd biosorption. Whereas, C=O stretch in amides groups and O-H stretch & H-bonded in alcohols and phenols groups showed a slight shift in wavelength indicating that these peaks could involve in Cd biosorption process.
Previously, Murphy et al. (2007) reported that carboxyl was the main group involved in brown seaweeds metal sorption. Whereas, D'Souza et al. (2008) reported various functional groups involved in Cd sorption in Padina tetrastromatica brown seaweed exposed to Cd stress using FTIR analysis. The previous study revealed that N-H stretching vibrations (3350 cm-1), bending vibrations (1571 cm-1) and carbonyl of COOH (1710 cm-1) were the major functional groups appeared in treated P. tetrastromatica. The same study revealed also that C = O stretching of ester carbonyl (1764 cm-1) in control was shifted to 1760 cm-1 under Cd treatment. Indeed, a peak of 1577 cm-1 in control showed a down shift to 1571 cm-1 in treated seaweed due to N-H bending vibrations. As well as, Cd stress induced a peak of P = O asymmetric stretching vibrations of PO2-phosphodiesters (1220 cm-1).
Lee et al. (2011) reported different peaks appeared in Kappaphycus Alvarezii seaweed exposed to metal ions (Cu, Cd, Cr, Ni and Zn). In this respect, these peaks were 3420 cm-1 (-OH or -NH groups), 2930 and 1385 cm-1 (-CH3 groups), 2350 cm-1 (-C=N groups), 1646 and 1240 cm-1 (-COOH groups), 1050 cm-1 (-C-O groups), and 1640 and 1240 cm-1 (-COO groups) shifted to 1645 and 1235 cm-1 combined with appearance of new peak at 1730 cm-1 (-C=O groups). Ferreira et al. (2011) reported that hydroxyl, amino and amide groups
were mainly associated with heavy metals uptake (Ni, Zn and Pb) into dried Arthrospira (Spirulina) platensis and Chlorella vulgaris biomass.
Whereas, Fakhry (2013) reported that 3250, 2550, 3400, 1200, 3000, 1670 and 1128 cm-1 referring to OH, COOH, NH2, S=O, C-H, C=O and C-O groups, respectively were involved in dye sorption into P. pavonica.
Otherwise, Ova and Ovez (2013) reported that carboxylate groups were included in heavy metals absorption (Cd, Ni, Pb, Zn, Fe and Cr) into Nitzschia closterium biomass. As well as, Naspu et al. (2014) reported functional groups involved in Cd biosorption into Padina sp. modified with PAA/HCl (mP-PAA/HCl) using FTIR analysis. The previous study revealed a peak of 3322.78 cm-1 yielded by interaction between PAA/HCl and seaweed. As well as, peaks at 1607.52 cm-1 (N-H group), 2943.68 cm-1 (C-H stretch) and 1040.11 cm-1 (C-N group) were involved in biosorption process.
Whereas, Ghoneim et al. (2014) characterized Cd adsorption in dried U. lactuca by identified the various functional groups involved in this process. The previous investigation indictaed the presence of C-H bond of methyl, methylene and methoxy, C-H stretching, C-C stretching, C-O and C-O stretching groups. Moreover, the previous study mentioned some changes in the hydroxyl and amino groups position's due to Cd stress.
Moreover, Radhika and Mohaideen (2015) applied FTIR technique to screnning different functional groups in two U. lactuca and Gracilaria corticata seaweeds. Indeed, Ponnanikajamideen et al. (2014) identified the different groups presented in different organic solvents characterized Padina tetrastromatica biomass. The previous study indicated the presence of OH, N-H and C-N stretching, -C-H bending vibrations, -C=C-stretching vibrations and - O-H stretching vibrations groups.
Other study (Sulaymon et al., 2013) however, investigated various functional groups involved in Pb, Cd, Cu, and As ions biosorption with dried different green (Chlorophyta) and blue-green (Cyanophyta) seaweeds species, using FTIR technique. The previous study revealed that interaction of seaweeds with metal ions from metal solution induced different functional
groups such as: amine groups (3414 cm-1), alkyl (2966 and 2943 cm-1) , carboxylic groups of C=O or ester groups (1797 cm-1), amide groups of C=O (1647 cm-1), carboxylate groups of COO- ( 1427 cm-1), sulfonate groups of S=O and fatty acids COO- groups (1300 cm-1), and organic phosphate groups of P-O-C (1033 cm-1).
Moreover, Dekhil et al. (2011) applied the same technique to investigate different functional groups appeared into dried Caulerpa racemosa green seaweed exposed to Pb and Cd ions. The previous study revealed
the appearance of O-H bending group (1420 cm-1) for the two ions, amide group slightly shifted to N-H stretching (3410 to 3396 cm-1) with Cd and appearance of new peak (3340 cm-1) corresponding to N-H stretching group with Pb. Indeed, the same study revealed that some groups such as C-O stretching group in the amide group did not involve in metals sorption. While, other one like C-O in amino group could exhibited an essential role in metals sorption process.
Wave number (cm"1)
Figure 1. FT-IR spectra of untreated U. lactuca (a), 2.5 mg/L (b), 5 mg/L (c) and 10 mg/L (d) Cd after 4 days exposure.
Table 1. FT-IR transmission frequencies (cm-1), estimated peaks and functional groups of untreated and Cd-treated U. lactuca seaweed.
IR frequency (cm-1) Bond Functional groups C T1 T2 T3
515-690 C-Br stretch Alkyl halides 613.2 585.9 580.4 582.1
550-850 C-Cl stretch Alkyl halides 848.3 848.3 848.4 848.4
1020-1250 C-N stretch Aliphatic amines 1063 1063.8 1055.6 1054.8
1000-1350 C-O; C-N stretch Carboxylic Acids, Amides 1125.1 1258.4 1265 1259.7
1400-1500 C-C stretch Aromatics 1420.5 1420.5 1420.5 1420.5
1400-1500 C-C stretch Aromatics 1448 1423.5 1423.4 1540.9
1600-1670 C=O stretch Amides 1635.4 1636.1 1633.1 1637
2850-3000 C-H stretch Alkanes 2927.6 2926.5 2927.1 2927.1
3200-3500 O-H stretch, H-bonded Alcohols, phenols 3431.1 3440.5 3440.5 3431.1
Notes. C: Control, T1: 2.5 mg/L Cd, T2: 5 mg/L Cd and T3: 10 mg/L Cd.
Table 2. FT-IR transmission frequencies (cm 1), estimated peaks and functional groups of untreated and Cd-treated P. pavonica seaweed.
IR frequency (cm-1) Bond Functional qroups C T1 T2 T3
690-900 =C-H oop bend Aromatics 712.8 712.8 712.7 712.8
690-900 =C-H oop bend Aromatics 874.6 874.5 874.8 874.7
1000-1350 C-O; C-N stretch Carboxylic Acids, Amides 1059.1 1059.3 1059.1 1058.9
1400-1500 C-C stretch Aromatics 1428.4 1427.5 1428.8 1428.5
1600-1670 C=O stretch Amides 1641.9 1644.4 1634.6 1638.5
2850-3000 C-H stretch Alkanes 2923.3 2923.1 2923.2 2924.8
3200-3500 O-H stretch, H-bonded Alcohols, phenols 3411.9 3414.7 3427.7 3419.5
Notes. C: Control, T1: 2.5 mg/L Cd, T2: 5 mg/L Cd and T3: 10 mg/L Cd.
Figure 2. FT-IR spectra of untreated P. pavonica (a), 2.5 mg/L (b), 5 mg/L (c) and 10 mg/L (d) Cd after 4 days exposure.
Other recent investigation (Saleh 2016a) focused on physiological impact of Cd stress into U. lactuca green and P. pavonica brown seaweeds for 4 days. The previous investigation revealed that Cd treatment reduces studied physiological parameters (specific growth rate, chlorophyll a & b, total chlorophyll and osmotic potential values) into the both seaweeds. In this respect, this decline was more noticeable in U. lactuca compared to P. pavonica. Indeed, carotenoids content decreased into U. lactuca and inversely increased into P. pavonica. These compounds could be considered as
an antioxidant due to their important role as a defenses mechanism against reactive oxygen species (ROS) induction by heavy metals exposure; suggesting that P. pavonica could declare more tolerant to Cd treatment compared to U. lactuca.
Moreover, Saleh (2016b) reported DNA changes induced by Cd in U. lactuca for 4 days based on Random Amplified Polymorphic DNA (RAPD) marker. The previous study revealed that genomic template stability (GTS%) value increased with Cd concentration increased. Similar recent findings were also reported by
Saleh (2016c) in P. pavonica treated with similar Cd concentrations using also the same marker.
More recently, Saleh (2017b) reported biosorption of four metal ions (Pb, Cu, Cd and Zn) after 5 days exposure into U. lactuca seaweed using FTIR analysis. The previous study revealed that O-H, N-H, -C-O, P-O-C, S-O, C-C, -C-C, -C-OH, C-H and C-I groups, were the main functional groups involved in metal ions biosorption process. Moreover, Saleh (2017a) reviewed physiological and molecular impacts of various heavy metal pollutants in plants and seaweeds. The previous review revealed the importance of some plants and seaweeds as a cheap potent source for heavy metals bioremediation.
CONCLUSION
In the current study, FTIR technique has been employed to monitoring and identifying the functional groups in two seaweeds (U. lactuca and P. pavonica) under different Cd concentrations. From data presented herein, for U. lactuca, C-O & C-N stretch in carboxylic acids and amides groups and C-C stretch in aromatics groups seem to play an important role in Cd biosorption. Whereas, C=O stretch in amides groups, O-H stretch in alcohols groups and H-bonded in phenols groups
pavonica.
ACKNOWLEDGEM ENT
I thank Professor Othman Ibrahim (Director General of AECS) and Professor MirAli Nizar (Head of Molecular Biology and Biotechnology Department) for their support and to the Plant Biotechnology Group for technical assistance.
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