Sorption of paralytic shellfish toxins (PSTs) in algal polysaccharide gels

Algal Research 45 (2020) 101655

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Sorption of paralytic shellfish toxins (PSTs) in algal polysaccharide gels Dave Eldon B. Olano*, Lilibeth A. Salvador-Reyes*, Marco Nemesio E. Montaño, Rhodora V. Azanza

T

Marine Science Institute, University of the Philippines Diliman, Quezon City, 1101, Philippines

ARTICLE INFO

ABSTRACT

Keywords: Algal polysaccharides Paralytic shellfish toxins Sorption Carrageenan Alginate Agar Saxitoxin

Sorption mechanics of the paralytic shellfish toxins (PSTs), saxitoxin (STX) and neo-saxitoxin (neo-STX), on algal polysaccharide gels was characterized using surface chemistry models. Refined (RC), semi-refined (SRC) carrageenan and alginate showed sorption of STX and neo-STX. The sorption of PSTs on RC, SRC and alginate was affected by contact time and in part, temperature. From surface chemistry models, alginate followed a spontaneous endothermic physical monolayer sorption of STX and neo-STX. SRC and RC favoured the concurrence of physical and chemical monolayer sorption, being endothermic for SRC and exothermic for RC.

1. Introduction Paralytic shellfish toxins (PSTs) produced by dinoflagellates (Alexandrium spp., Pyrodinium spp., and Gymnodinium spp.) [9,11] and freshwater cyanobacteria (Microcystis spp., Anabaena spp., Cylindrospermopsis raciborskii and Aphanizomenon spp.) [7] are among the most potent neurotoxins. These molecules can cause a variety of effects such as nausea, vomiting; in severe cases, paralysis and death. PSTs such as saxitoxin (STX), neo-saxitoxin (neo-STX) and gonyautoxin (GTX) enter the food chain through bioaccumulation of PST causative organisms in shellfish, finfish and other marine organisms [18]. PSTs can persist in the water column, sediments and associated biota after the occurrence of generative blooms [8,19]. During blooms of PST causing organisms, saxitoxin concentration can reach up to 15 μg/L in natural waters [39], or in extreme cases, up to 600 μg/L [16]. A limit of 3 μg/L saxitoxin is indicated for drinking water [39]. The economic cost and health impacts associated with PSTs have prompted the design of control, mitigation and prevention strategies. HAB control strategies include the use of biological agents [7,34], chemicals [7], flocculants [30,38] and sorbent materials [8,9,21,39]. Large-scale chemical treatment using copper sulfate to control dinoflagellate blooms have been met with hurdles such as non-specificity of action, affecting co-occurring algae and other marine organisms [7,36]. PSTs are also resistant to ozonation and proved to be an ineffective control strategy [31,35]. The use of flocculants and sorbents have emerged as promising control strategies due to efficiency and practicality. Sorption studies have mainly focused on environmental

⁎

applications and wastewater treatment strategies. Oyster and chitin shell powder, clay, powdered activated charcoal, and κ-carrageenan are among the sorbents utilized for saxitoxin sorption. Incubation of oyster and chitin shell powder (200 mg) with 2–16 μg/L saxitoxin resulted in > 50% sorption after 18 h [26]. Adsorption of saxitoxin was shown to occur spontaneously with a pseudo-second order kinetics [26]. Further, sorption of STX in oyster and chitin shell powder was influenced by pH, with acidic conditions being least favourable, and suggested an ion exchange mechanism [26]. Sorption of saxitoxin (25 μg/L) in powdered activated charcoal on the other hand was influenced by the amount of sorbent and contact time [39]. Shorter incubation time of 2 h resulted in 48%, 51% and 77% removal of saxitoxin at pH 7.05, 8.07 and 10.7 [39]. Increasing the incubation time with activated charcoal to 24 h resulted to ∼100% sorption of saxitoxin at pH 8.07 and 10.7 [39]. A > 50% removal of saxitoxin (5 μM) was observed within 2 h of incubation with clay [8]. Clay-saxitoxin interaction fitted with a Freundlich isotherm and was suggested to occur via cation exchange mechanism [8]. Cañete and Montaño [9] reported the potential of κcarrageenan gel as sequestering agent of PSTs from Pyrodinium bahamense through a mouse bioassay for toxicity. After 7 min of incubation of P. bahamense extract with κ-carrageenan, a 55% reduction in toxicity was observed in the mouse bioassay [9]. Further, the toxin reduction from κ-carrageenan was affected by incubation time, polysaccharide concentration and gel surface area [9]. Sorption of κ-carrageenan was proposed to be through a cation exchange mechanism however, characterization of the sorption mechanics of PST in κ-carrageenan was not carried out.

Corresponding authors. E-mail addresses: [email protected] (D.E.B. Olano), [email protected] (L.A. Salvador-Reyes).

https://doi.org/10.1016/j.algal.2019.101655 Received 26 March 2019; Received in revised form 29 August 2019; Accepted 4 September 2019 2211-9264/ © 2019 Elsevier B.V. All rights reserved.

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Here, we expanded and explored the potential of the algal polysaccharide gels: agar, alginate, semi-refined (SRC) and refined (RC) carrageenans as sorbent material for PSTs. We quantified the sorption potential through batch sorption studies and characterized the PSTalgal polysaccharide gel interaction using surface chemistry models.

extracts and standards were analyzed using HPLC-FD as described by Lawrence and Menard [23] and as adapted by Cañete and Montaño [9]. The % recovery for CRM-STX-f and CRM-NEO-d are 100.00% and 99.99%, respectively. Toxin concentrations of STX and neo-STX were determined by integrating the peak areas and calculated using Eqs. (1) and (2).

2. Materials and methods

Rf =

2.1. Algal culturing

average peak area of standard standard conc.× V

(1)

PST peak area/PST Rf V × df

(2)

[PST] =

Mono-species algal cultures of Pyrodinium bahamense (PBCMZRVA042595) were obtained from the HAB laboratory of the Marine Science Institute, University of the Philippines (UP-MSI). Batch cultures of P. bahamense Mz were grown in modified F/2 medium for six weeks [17]. All media were prepared using sterilized (120 °C) and filtered seawater with 30 psu salinity. Aseptic techniques were observed during culture to avoid contamination. The cultures were maintained following the procedures of Azanza-Corrales et al. [4] until PST extraction.

where, Rf ≡ retention factor. V ≡ injection volume. df ≡ dilution factor. 2.5. Data analysis

2.2. PST extraction

Sorption efficiency of the algal polysaccharide gels was calculated Eq. (3) as adapted from Refs. [1,14]. Rate of sorption was calculated using Eq. (4).

PSTs were extracted from P. bahamense cultures according to the method of Oshima [32]. In brief, P. bahamense cells were lysed with 0.1 N CH3COOH and the resulting cell lysates were filtered. The pH of the filtrate was adjusted to pH 3.0 using 0.1 N CH3COOH to yield the PST crude extract.

Sorption Efficiency (%) =

[PST] i

[PSTf ]

[PSTi ]

× 100%

(3)

where,

2.3. Algal polysaccharide gel preparation and rheological characterization

[PSTi] ≡ initial PST concentration, [PSTf] ≡ final PST concentration.

Food-grade agar, κ-carrageenan (refined and semi-refined) and sodium alginate were obtained from Shemberg Philippines through the Seaweed Chemistry Laboratory and Pilot Plant, UP-MSI. Algal polysaccharides (agar and κ-carrageenan) were dissolved in warm (∼70 °C) distilled water with constant stirring to produce a 1.5% (w/v) polysaccharide solution. For alginate, 1% (w/v) CaCl2 solution was added dropwise to 1.5% (w/v) alginate solution [33] to form a gel. Gel strength was analyzed according to the procedure of Montaño and Pagba [27]. Viscosity was measured using a Brookfield Viscometer LDV-II+ Pro (equipped with LV 61 spindle at 30 rpm for 30 s) at 75 °C. Following characterization, a 20 mL-aliquot of the algal polysaccharide solutions were allowed to form a gel for 20 h. Polysaccharide gel discs (0.47 × 1.91 × 0.5 cm) were molded as in the procedure of Cañete and Montaño [9].

Rate of Sorption =

Sorption Efficiency time

(4)

Statistical analyses were done in Prism 7.00 and R 3.4.3 Statistical software. Sets were tested for normality using Shapiro-Wilk's test and homoscedasticity using Bartlett's test. Analysis of Variance (ANOVA) was done to compare more than two normally distributed independent sets. Both Tukey's HSD and Dunnett's test were used as post-hoc evaluation to assess significant differences among treatments, especially against the control. Since data sets are parametric, neither data transformations nor nonparametric tests were done. A 95% confidence interval (α = 0.05) was assumed in all statistical analyses. 2.6. Surface chemistry equations and models

2.4. Batch sorption test

2.6.1. Kinetic model Two basic kinetic (pseudo-first-order and pseudo-second-order) models, as adapted from [26,29,37], were used to model the batch sorption experimental data. The pseudo-first-order model was calculated according to Eq. (5):

A 1:10 10 mM phosphate buffer (pH 7.0): PST crude extract was prepared for the batch sorption experiment. Separate incubations for negative control (no algal polysaccharide discs), agar, alginate, RC and SRC were performed at 298 and 310 K, 50 rpm, with five algal polysaccharide discs in each setup. A 100 μL-aliquot of the PST solution was retrieved at t = 0, 5, 10, 30, 60, 120, and 180 min. Aliquots were subjected to toxin analysis using high performance liquid chromatography coupled to fluorescence detection (HPLC-FD). A series of five PST solutions (STX: 110.0 ± 60, 200 ± 50, 260 ± 50, 340 ± 40, 450 ± 30 μg/L; neo-STX: 100 ± 40, 170 ± 50, 260 ± 60, 360 ± 30, 500 ± 40 μg/L) were prepared by diluting appropriate volumes of PST stock solution with phosphate buffer, ex. for 20% PST solution: 20 mL PST stock solution +80 mL phosphate buffer at 298 and 310 K. These were subjected to batch sorption experiments using alginate, SRC and RC as indicated above.

log(qe

qt ) = log(qe )

k1 t 2.303

(5)

where qt is the sorption capacity at time t and k1 is the rate constant. The qe (at saturation) and qt (at specific time points) were calculated by using the formula:

q=

[PST] i

[PSTf ] × V w

(6)

where PSTi is the initial concentration of PST, PSTf is the final concentration PST, V is the volume of the sorbate (10 mL) and w is the mass of the sorbent (5 g). The calculated log(qe-qt) was plotted against contact time, t (mins). The best fit line and linearity (R2) were extracted from the plot. The rate constant, k1, was calculated according to Eq. (7).

2.4.1. Toxin profiling and quantitation Certified Reference Materials (CRMs) of STX (CRM-STX-f, 66.3 ± 1.4 μmol/L) and neo-STX (CRM-NEO-d, 65.1 ± 2.1 μmol/L) were purchased from the National Research Council, Canada. PST

k1 = 2

2.303m

(7)

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where m ≡ slope of the line. The pseudo-second order kinetic model was calculated using the formula:

t 1 1 = + t qt k2 qe 2 qe

(8)

where qt is the sorption capacity at time t, t is the time (mins), and the k2 is the pseudo-second-order rate constant for sorption. The best fit line and linearity (R2) were obtained by plotting t/qt and time t in mins. The value of k2 was calculated and recorded using the formula:

k2 =

1 bqe 2

(9)

where b ≡ y-intercept 2.6.2. Isotherm models Experimental data were fitted using two of the most common isotherm models: Langmuir and Freundlich [5]. The equation describing the Langmuir model is shown in Eq. (10)

Ce C 1 = e + qe qmax qmax KL

(10)

where, Ce is the equilibrium toxin concentration of the PST solutions at varying concentrations after 180 mins of incubation, KL is the Langmuir equilibrium constant and qmax is the maximum sorption capacity. A plot of Ce/qe against Ce was used to determine the linear relationship, and KL and qmax were calculated accordingly using Eqs. (11) and (12), where m is the slope of the line and b, the y-intercept:

1 m

(11)

1 qmax b

(12)

qmax = KL =

Fig. 1. Time-course monitoring of STX reduction using algal polysaccharide gels at (A) 298 K and (B) 310 K. Data presented as mean ± SD, n = 3.

efficiency was assessed as the fraction of the total PST remaining in solution at the end of the incubation period. Alginate, SRC and RC showed time-dependent sorption efficiency at 298 K, with an increase in STX sorption with contact time (Fig. 1). In contrast, agar did not show any significant sorption of STX at all time points. Sorption in agar remained low and comparable to the negative control. The highest sorption rates for alginate and carrageenans were observed during the first 10 mins of incubation and gradually decreased. After 60 min, the rate of sorption remained almost constant at 0.1–0.2% STX/min for the three polysaccharide gels. There was a significant increase in sorption of STX in the algal polysaccharide gels (Fig. 1) with increased incubation temperature. Agar, which did not bind STX at 298 K, demonstrated activity after 30 min of incubation at 310 K. Alginate showed faster STX sorption at 310 K, resulting in a 31.8 ± 8.9% sorption efficiency after 30 min (Fig. 1). In contrast, the sorption efficiency of refined carrageenan was independent of temperature, neither affecting the rate nor sorption efficiency. The rate and efficiency of sorption in the negative control was negligible and remained constant over time. This implied the stability of STX during the observation period. Similar to STX, the sorption of neo-STX to alginate, SRC and RC showed time dependence (Fig. 2). Alginate, RC and SRC showed comparable sorption efficiencies after 30 min of incubation at 298 K. A gradual decrease in the rate of sorption of alginate, RC and SRC was observed until 60 min. After this time, the rate of sorption remained almost constant at 0.1–0.2% neo-STX/min for the three polysaccharide gels. An inverse relationship with temperature and neo-STX sorption in SRC and RC was observed. Longer incubation time was required for incubation at 310 K to obtain comparable sorption efficiency to the 298 K counterparts. Alginate showed a slight increase in sorption at 310 K. Agar also demonstrated low sorption of neo-STX. Negative control setups indicated stability of neo-STX.

Freundlich isotherm model was fitted using the equation:

log qe = log KF +

1 log Ce n

(13)

where, KF is the relative sorption capacity and n is related to the affinity of the sorbent to the sorbate. The parameters KF and n were obtained using the y-intercept and slope values on the trend line of the plot: log qe vs log Ce. 2.6.3. Thermodynamic models Gibb's free energy (ΔG°), enthalpy (ΔH°) and entropy (ΔS°) were used to describe the thermodynamics of PST sorption on algal polysaccharide gels. Enthalpy and entropy were obtained using the Van't Hoff equation shown in Eq. (14) from the slope and intercept of the ln (qe/Ce) versus 1/T plot [13].

ln(qe / Ce ) = ( S o/ R)

( H o/RT)

(14)

where, R is the universal gas constant (8.312 J/mol∙K) and T is the temperature at 298, 303 and 310 K. Gibb's free energy was derived using the Gibbs-Helmholtz equation as shown in Eq. (15).

Go = H o

T So

(15)

3. Results 3.1. Contact time and temperature affect the rate and efficiency of sorption of STX and neo-STX on algal polysaccharide gels Sorption of STX and neo-STX in agar, alginate, SRC and RC was quantified at 298 and 310 K for 180 min. P. bahamense crude extract was incubated with the algal polysaccharide gel discs. Sorption 3

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3.3. Interaction of STX and neo-STX on algal polysaccharide gels follow the Langmuir isotherm model Batch sorption experiments using five different PST concentrations at equilibrium contact time of 180 min and temperatures of 298 and 310 K were used for the isotherm modeling. By plotting Ce/Qe versus Ce, the Langmuir constant KL and maximum sorption capacity (qmax) of the algal polysaccharides were estimated. On the other hand, the Freundlich constant KF and intensity of sorption (n) were estimated from the intercept and the slope of the logQe versus logCe plot, respectively. Suitability of the isotherm model for each algal polysaccharide gel was determined from the R2 values. Sorption of STX and neo-STX on all algal polysaccharide gels followed the Langmuir isotherm model at both temperatures (Fig. 5, Table 3). Linearity of the Langmuir model for STX sorption ranged from 0.8464 to 0.9795, compared to Freundlich's, which ranged from 0.0110 to 0.6584 (Table 3, Supporting Information). Likewise, the Langmuir model plots for neo-STX sorption in algal polysaccharide gels yielded R2 of 0.8864 to 0.9516 (Table 3, Supporting Information). From the KL and qmax at 298 K, SRC gave the highest affinity and sorption capacity for STX, while for neo-STX, alginate showed preferential sorption (Table 3). The increase in temperature influenced the affinity of the algal polysaccharide gels for STX and neo-STX, being favourable to the increased affinity and qmax of alginate (Table 3). However, an opposite trend was observed for the sorption of STX and neo-STX in carrageenans. 3.4. STX and neo-STX sorption on algal polysaccharide gels are spontaneous and thermodynamically stable From the slope and intercept of Fig. 6, ΔH° and ΔS° were calculated, respectively. Consequently, using Eq. (15), ΔG° was determined. STX and neo-STX sorption in all algal polysaccharide gels indicated spontaneity of the sorbent-sorbate interaction, as indicated by ΔG° < 0 kJ/ mol (Table 4). The ΔG° for STX and neo-STX sorption in alginate and RC ranged from −50 to 0 kJ/mol, suggesting a spontaneous physical sorption. ΔG° values for SRC were outside the specified range for both physical and chemical sorption. Moreover, as the temperature increased, the reactions continued to remain spontaneous (Table 4). Based on the ΔH° values for STX and neo-STX sorption in alginate and SRC, the process would likely involve an endothermic physical sorption (Table 4). Negative ΔH° values for STX and neo-STX sorption in RC is indicative of an exothermic physical sorption process. All algal polysaccharide gels had positive ΔS° values, which corresponds to increased randomness at the sorbent-sorbate interface (Table 4).

Fig. 2. Time-course monitoring of neo-STX reduction using algal polysaccharide gels at A. 298 K and B. 310 K. Data presented as mean ± SD, n = 3.

3.2. Sorption kinetics of STX and neo-STX in alginate and carrageenans is influenced by temperature Sorption of STX and neo-STX in alginate, refined and semirefined carrageenan was characterized using kinetic, isothermic and thermodynamic modeling. Batch sorption experimental data was modelled using Lagergren pseudo-first-order and pseudo-second-order kinetic models. By plotting log(qe-qt) versus t, the first-order rate constant k1 and theoretical equilibrium capacity (qe) were estimated from the slope and intercept, respectively. On the other hand, the second-order rate constant k2 and the theoretical equilibrium capacity (qe) were estimated from the intercept and the slope of the t/qt versus t plot. The linearity of the plot, R2, served as the primary criteria for the fitness of the kinetic models. Further, agreement between theoretical and experimental qe values provided validation on the suitability of the kinetic model. Sorption of STX in alginate, SRC and RC (Fig. 3, Table 1) at 298 and 310 K showed good linearity using the pseudo-first-order model (R2 = 0.9224–0.9947). Calculated and experimental qe values in the pseudo-first-order model also showed good agreement (Table 1). At 298 K, neo-STX sorption on alginate preferred the pseudo-first-order kinetic model (R2 = 0.8245) (Table 2, Fig. 3). At 310 K, alginate displayed preference for both pseudo-first-order and pseudo-second-order models, having comparable R2 for both models. The same trend was observed for neo-STX sorption on SRC and RC, acceptable linearity for both pseudo-first-order and pseudo-second-order kinetic models were observed with increased incubation temperature (Figs. 3 and 4 , Table 2). Since linearity of both pseudo-first-order and pseudo-secondorder models were high, concurrence of physical and chemical sorption may be possible [28].

4. Discussion Several studies have explored the use of sorbents as a strategy to mitigate PSTs [8,9,26,31,39]. In our study, we used surface chemistry through kinetic, isothermic and thermodynamic models to characterize the sorption mechanics, dynamics and retention of PSTs on algal polysaccharide gels. Kinetic studies were used to provide hints on the sorption mechanism [1] as pseudo-first-order kinetics implies physisorption [21,41], while pseudo-second-order kinetics implies chemisorption [21,45]. Sorption isotherms were used to describe a substance's mobility [24] and retention on a solid substance at various concentrations [21]. The Langmuir and Freundlich isotherm described monolayer and multilayer binding, respectively [21,26,41). Thermodynamic studies were performed to investigate temperature effects on the sorption process. Gibbs free energy (ΔG°) describes spontaneity of sorbent-sorbate interactions, enthalpy (ΔH°) quantifies the heat evolved or gained during the sorption process [42], while entropy (ΔS°) measures the randomness of the interaction [2,41].

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Fig. 3. Pseudo-first-order kinetic modeling of A. STX and B. neo-STX on algal polysaccharide gels at 298 K and 310 K. Data presented as mean log(qe-qt) from triplicate runs.

Carrageenans followed a physical monolayer sorption of STX and neo-STX at 298 K. At higher temperature, carrageenans showed best fit to both pseudo-first-order and pseudo-second-order kinetic models. This implied the propensity of carrageenans for both physical and chemical sorption processes, following monolayer STX and neo-STX binding at 310 K. The first 10 min of incubation in RC showed the highest rate of PST sorption through potential physical sorption. Pseudo-first-order kinetics is best suited for the first 20–30 min of interaction time [25]. Saturation was observed, starting at 60 min postincubation. Chemisorption, a stronger chemical interaction than physisorption, may have played a vital role in the saturation of PSTs in RC with longer incubation time. The sorption of PSTs in carrageenans may be due to the interaction of negatively charged sulfate groups that electrostatically interact with the positively-charged amine groups of

PSTs [9], and the hydrogen bonding and complexation between the amine and sulfate moieties of carrageenan [10,43]. The thermodynamic parameters revealed spontaneous, exothermic physical sorption processes for RC, while spontaneous, endothermic physical and chemical processes occurred for SRC. Carrageenans showed higher rates and affinities for STX over neo-STX, indicating the propensity of this algal polysaccharide gel for the sorption of STX. PSTs have several amine groups that can potentially be protonated, thereby becoming cationic depending on the pH of the solution. Shi et al. [39] reported that STX shifted to a mixture of mono-cationic and di-cationic species at pH 7.1. On the other hand, neo-STX contains an additional hydroxy group making it more anionic than STX. The anionic nature of neo-STX than STX could potentially contribute to electrostatic repulsion and consequently, the better preference of carrageenans for STX sorption.

Table 1 Parameters for Pseudo-first and Pseudo-second-order Sorption Kinetics of Saxitoxin in Algal Polysaccharide Gels at 298 K and 310 K. Temp (K)

Alginate Refined Carrageenan Semi-refined Carrageenan

298 310 298 310 298 310

qe (exp)

452.05 304.80 648.89 286.55 471.46 327.36

Pseudo-first-order

Pseudo-second-order

k1

qe (theor)

R2

k2

qe (theor)

R2

0.0122 0.0124 0.0154 0.0147 0.0196 0.0234

474.79 257.69 672.05 271.77 476.10 358.34

0.9624 0.9224 0.9947 0.9783 0.9672 0.9597

26.44 17.40 102.57 16.76 61.10 25.12

769.23 294.12 1000.0 344.83 555.56 357.14

0.2333 0.7918 0.5379 0.8382 0.7932 0.8840

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Table 2 Parameters for Pseudo-first and Pseudo-second-order Sorption Kinetics of Neo-saxitoxin in Algal Polysaccharide Gels at 298 and 310 K. Temp (K)

Alginate Refined Carrageenan Semi-refined Carrageenan

298 310 298 310 298 310

qe (exp) (ng/g)

Pseudo-first-order

562.69 480.46 584.78 503.62 373.28 434.91

0.008 0.021 0.010 0.018 0.014 0.010

k1

Pseudo-second-order

qe (theor) (ng/g) 478.63 420.63 436.52 486.97 250.67 312.03

Isothermic analysis also indicated that the increase in temperature negatively affects the interaction of PSTs with carrageenans and may potentially pose as a limitation. Semi-refined carrageenan followed the same trend as refined carrageenan, with preferential sorption of STX and reduced performance with increased incubation temperature. Refined and semi-refined carrageenans possess similar structure but different purity levels. Semi-refined carrageenan did not undergo purification hence, salt residues and other carbohydrates such as cellulose may still be present. These impurities may antagonistically affect the interaction of semi-refined carrageenan with PSTs and hence, affect the sorption efficiency, sorption capacity and affinity. These mechanistic studies support the observed reduction in toxicity levels of P. bahamense extract with κ-carrageenan treatment. We further evaluated the potential of the algal polysaccharides alginate and agar, for PST sorption. Agar showed low sorption of STX and neo-STX, likely due to the weak interaction between agar and PSTs.

2

R

k2

0.8245 0.9088 0.8435 0.9850 0.7685 0.9268

94.65 132.95 88.91 258.34 191.99 40.62

qe (theor) (ng/g) 588.24 555.56 555.56 588.24 416.67 370.37

R2 0.6257 0.9026 0.8138 0.9668 0.9844 0.8643

Agar has predominantly hydroxy groups that can only allow weak hydrogen bonding with PSTs. Moreover, the high gel strength of agar can cause loss of chain flexibility, reduction of mobility of chelating groups and their accessibility, and decreased swelling capacity [10,40]. In contrast, alginate showed comparable sorption potential with carrageenan, albeit with a different mechanism and selectivity. This is hitherto the first report of the sorption potential of alginate for PSTs. Alginate, similar to carrageenan, has cation-exchange properties. The carboxylic acid moieties of alginate can form electrostatic interactions with the cationic moieties of STX and neo-STX. Alginate showed best fit for a pseudo-first-order kinetics and Langmuir isotherm model. This implied preference for a physical, monolayer adsorption of STX and neo-STX in alginate at 298 K and 310 K. Thermodynamics further corroborated the results of the kinetic model as the ΔG° and ΔH° values reflected a spontaneous, endothermic physical sorption process. Adsorption is normally an exothermic process [21]; however, alginate

Fig. 4. Pseudo-second-order kinetics modeling of A. STX and B. neo-STX on algal polysaccharide gels at 298 and 310 K. Data presented as mean t/qt from triplicate runs.

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Fig. 5. Langmuir isotherm modeling of A. STX and B. neo-STX on algal polysaccharide gels at 298 and 310 K. Data presented as one point per replicate. Table 3 Parameters for the Langmuir isotherm models of STX and neo-STX onto algal polysaccharide gels at 298 and 310 K. Temp (K)

Alginate Refined Carrageenan Semi-refined Carrageenan

298 310 298 310 298 310

STX

Neo-STX

KL

qmax (ng/g)

R2

KL

qmax (ng/g)

R2

51.45 265.28 176.00 96.00 227.21 85.01

715.82 1033.06 1266.30 886.52 1045.15 980.28

0.8877 0.9316 0.9795 0.9394 0.8876 0.9253

96.36 308.49 96.00 47.48 73.90 38.27

1179.11 1404.49 886.52 753.58 1053.41 837.38

0.9181 0.9093 0.9394 0.8864 0.8975 0.9516

sorption was shown to prefer an endothermic process [3,15]. This preference can be attributed to chemical reactions or bonds involved in the sorption process [3]. Since kinetic and thermodynamic data established physical sorption or weak forces, i.e. van der Waal's force [42], as the primary driving force of the sorption process, the observed endothermic reaction may not be due to chemical reactions or bond formation, but may only be due to the protonation and deprotonation of the functional groups of alginate [6,12]. In contrast to carrageenan, alginate has comparable affinity for sorption of STX and neo-STX. Also, the sorption in alginate was weakly dependent on the temperature. These suggest that alginate may be a more promiscuous and versatile sorbent compared to carrageenan. The integration of kinetic, thermodynamic and isothermic characterization highlighted key differences in

sorption preference and binding characteristics of carrageenan and alginate. While algal polysaccharide gels showed weaker monolayer physical sorption mechanism compared to other sorbents such as oyster and chitin materials, they offer flexibility and versatility that can be utilized to increase the sorption capacity. Modification and fortification of algal polysaccharide gels can further be done to enhance its sorption capacity and subsequently, its affinity and rate of PST sorption [20,22,44]. Carrageenan and alginate are also relatively inexpensive materials that are classified as GRAS. Their ability to bind STX and neo-STX opens the possibility of use as sorbent for PST removal in drinking water and gastrointestinal decontamination during poisoning.

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Table 4 Thermodynamic parameters for the sorption of STX and neo-STX on algal polysaccharide gels. ΔH° (kJ/mol)

ΔS° (J/mol∙K)

Temperature (K)

ΔG° (kJ/mol)

0.137

97.11

−0.140

38.31

Semi-refined Carrageenan

0.600

201.28

298 303 310 298 303 310 298 303 310

−28.80 −29.29 −29.96 −11.56 −11.75 −12.01 −59.38 −60.39 −61.80

Neo-STX Alginate

0.132

97.19

−0.290

5.719

0.979

288.41

298 303 310 298 303 310 298 303 310

−28.83 −29.32 −30.00 −19.94 −20.22 −20.63 −84.97 −86.41 −88.43

STX Alginate Refined Carrageenan

Refined Carrageenan Semi-refined Carrageenan

Authors' contributions to the manuscript DEO, LSR, RVA, and MNM designed the experiment. DEO conducted the experiments. DEO and LSR analyzed and interpreted the data, and wrote the manuscript. DEO, LSR, MNM and RVA reviewed the data and discussion, and edited the manuscript. LSR overall supervision. Declaration of Competing Interest No conflicts applicable. Acknowledgments The authors acknowledge funding from the Department of Science and Technology – Philippine Council for Agriculture, Aquatic and Natural Resources Research and Development (DOST-PCAARRD) through the research program Hazard Detection and Mitigation Tools for Opportunistic Algal Blooms in a Changing Environment. We thank A. Yñiguez, J. K. Andres and J. Mendoza for technical assistance and M. Murphy for critically reviewing the manuscript. This is MSI Contribution No. 466. Appendix A. Supplementary data

Fig. 6. Thermodynamic plots of A. Alginate B. RC and C. SRC. Data presented as mean ln(qe/Ce) from triplicate runs.

Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.algal.2019.101655.

5. Conclusion

References

This study assessed the mechanics, dynamics and retention of PSTs on algal polysaccharide gels. Alginate and carrageenans showed varying STX and neo-STX sorption mechanism and characteristics. Alginate followed a spontaneous endothermic physical monolayer sorption with STX and neo-STX. Carrageenans showed favourable sorption for STX that is negatively influenced by temperature. Semirefined carrageenan had the propensity to favor the concurrence of physical and chemical monolayer sorption. On the other hand, refined carrageenan showed the ideal sorption conditions. Surface chemistry studies indicated that the binding of PSTs on refined carrageenan is favourable and exothermic in the occurrence of a physical and chemical monolayer sorption.

[1] Z. Aly, A. Graulet, N. Scales, T. Hanley, Removal of aluminum from aqueous solutions using PAN-based adsorbents: characterization, kinetics, equilibrium and thermodynamic studies, Environ. Sci. Poll. Res. 21 (5) (2014) 3972–3986. [2] Z. Aly, V. Luca, Uranium extraction from aqueous solution using dried and pyrolyzed tea and coffee wastes, J. Radioanal. Nucl. Chem. 295 (2) (2013) 889–900. [3] R. Aravindhan, N.N. Fathima, J.R. Rao, B.U. Nair, Equilibrium and thermodynamic studies on the removal of basic black dye using calcium alginate beads, Coll. Surf. 299 (1–3) (2007) 232–238. [4] R.A. Azanza-Corrales, S. Hall, Isolation and culture of Pyrodinium bahamense var. compressum from the Philippines, in: T.J. Smayda, Y. Shimizu (Eds.), Toxic Phytoplankton Blooms in the Sea, Elsevier, Amsterdam, 1993, pp. 725–730. [5] N. Barka, M.E. Makhfouk, Removal of methylene blue and Eriochrome black T from aqueous solutions by biosorption on Scolymus hispanicus L.: kinetics, equilibrium

8

Algal Research 45 (2020) 101655

D.E.B. Olano, et al.

solutions, Chem. Central J. 6 (2012) 86. [27] M.N.E. Montano, C.V. Pagba, Agar Processing and Characterization, DOSTPhilippine Council for Aquatic and Marine Research and Development and UPMarine Science Institute, 1996. [28] V.S. Munagapati, Equilibrium isotherms, kinetics, and thermodynamics studies for congo redadsorption using calcium alginate beads impregnated with nano-goethite, Ecotoxicol. Environ. Saf. 141 (2017) 226–234. [29] A.E. Ofomaja, Sorption dynamics and isotherm studies of methylene blue uptake on palm kernel fiber, Chem. Eng. J. 126 (2007) 35–43. [30] I.S. Orizar, P.P. Rivera, M.L. San Diego-McGlone, R.V. Azanza, Harmful algal bloom (HAB) mitigation using ball clay: effect on non-target organisms, J. Envi. Sci. Man. 1 (2013) 36–43. [31] P.T. Orr, G.J. Jones, G.R. Hamilton, Removal of saxitoxins from drinking water by granular activated carbon, ozone and hydrogen peroxide-implications for compliance with the Australian drinking water guidelines, Water Res. 38 (2004) 4455–4461. [32] Y. Oshima, Post-column derivation HPLC methods for paralytic shellfish poisons, in: G.M. Hallegraff, D.M. Anderson, A.D. Cembella (Eds.), Manual on Harmful Marine Microalgae, UNESCO, 1995. [33] L.H. Pignolet, A.S. Waldman, L. Schechinger, G. Govindargin, J.S. Nowich, The alginate demonstration: polymers, food science, and ion-exchange, J. Chem. Educ. 75 (11) (1998) 1430 1998. [34] L.V. Padilla, M.L. San Diego-McGlone, R.V. Azanza, Preliminary results on the use of clay to control Pyrodinium bloom - a mitigation strategy, Sci. Diliman 18 (1) (2006) 35–42. [35] J. Rositano, G. Newcombe, B. Nicholson, P. Sztajnbok, Ozonation of NOM and cyanobacterial toxins in four treated waters, Water Res. 35 (2001) 23–32. [36] G.A. Rounsefell, J.E. Evans, Large-scale experimental test of copper sulfate as control for the Florida red tide, US Fish and Wildlife Service Special Scientific Report 270 (1958). [37] M.M.A. Salleh, D.K. Mahmoud, A.W. Wan Abdul Karim, A. Idris, Cationic and anionic dye adsorption by agricultural solid wastes: a comprehensive review, Desalination 280 (2011) 1–13. [38] M.R. Sengco, J.A. Hagstrom, E. Graneli, D.M. Anderson, Removal of Prymnesium parvum (Haptophyceae) and its toxins using clay minerals, Harmful Algae 4 (2005) 261–274. [39] H. Shi, J. Ding, T. Timmons, C. Adams, pH effects on the adsorption of saxitoxin by powdered activated carbon, Harmful Algae 19 (2012) 61–67. [40] D. Shiftan, F. Ravenell, M.A. Mattescu, R.H. Marchessault, Change in the V/B polymorph ration and T1 relaxation of epichlorohydrin crosslinked high amylose starch exipient, Starch 52 (2010) 186–195. [41] N.P. Sotirelis, C.V. Chrysikopoulos, Interaction between graphene oxide nanonparticles and quartz sand, Environ. Sci. Technol. (2015) 13413–13421. [42] H.N. Tran, S. You, H. Chao, Thermodynamic parameters of cadmium adsorption onto orange peel calculated from various methods: a comparison study, J. Environ. Chem. Eng. 4 (2016) 2671–2682. [43] R.L. Veroy, M.N.E. Montano, M.L.B. de Guzman, E.C. Laserna, G.J.B. Cajipe, Studies on the dye binding of heavy metals to algal polysaccharides from Philippine seaweeds, I: carrageenan and the binding of lead and cadmium, Bot. Mar. 23 (1980) 59–62. [44] Y. Wu, H. Qi, C. Shi, R. Ma, S. Liu, Z. Huang, Preparation and adsorption behaviors of sodium alginate-based adsorbent-immobilized β-cyclodextrin and graphene oxide, RSC Adv. 7 (2017) 31549–31557. [45] S.A. Idris, K.M. Alotaibi, T.A. Peshkur, P. Anderson, M. Morris, L.T. Gibson, Adsorption kinetic study: effect of adsorbent pore size distribution on the rate of Cr (VI) uptake, Microporous Mesoporous Mater. 165 (2013) 99–105.

and thermodynamics, J. Taiwan Inst. Chem. Eng. 42 (2) (2011) 320–326. [6] J. Bajpai, R. Shrivastava, A.K. Bajpai, Dynamic equilibrium studies on adsorption of Cr(VI) ions onto binary bio-polymeric beads of cross linked alginate and gelatin, Coll. Surf. 236 (2004) 81–90. [7] D. Boesch, D. Anderson, R. Horner, S. Shumway, P. Tester, T. Whiteledge, Harmful algal blooms in coastal waters: Options for prevention, control and mitigation, NOAA Coastal Ocean Program 10 (1996) 21–28. [8] J.M. Burns, S. Hall, J.L. Ferry, The adsorption of saxitoxin to clays and sediments in saline waters, Water Res. 43 (2009) 1899–1904. [9] S.J.P. Canete, M.N.E. Montano, k-Carrageenan gel as agent to sequester paralytic shellfish poison, Mar. Biotechnol. 4 (2002) 565–570. [10] G. Crini, Recent developments in polysaccharide-based materials used as adsorbents in wastewater treatment, Prog. Polym. Sci. 30 (2005) 38–70. [11] European Food Safety Authority (EFSA), Marine biotoxins in shellfish - Saxitoxin group, EFSA J. 1019 (2009) 1–76. [12] M.A. Fontecha-Camara, M.V. Lopez-Ramon, M.A. Alvarez-Merino, C. MorenoCastilla, About the endothermic nature of the adsorption of the herbicide diuron from aqueous solutions on activated carbon fiber, Carbon 44 (2006) 2330–2356. [13] Q. Fu, Y. Deng, H. Li, J. Liu, H. Hu, S. Chen, T. Sa, Equilibrium, kinetic and thermodynamic studies on the adsorption of the toxins of Bacillus thuringiensis subsp. kurstaki by clay minerals, Appl. Surf. Sci. 255 (8) (2009) 4551–4557. [14] Z.N. Garba, I. Bello, A. Galadima, A.Y. Lawal, Optimization of adsorption conditions using central composite design for the removal of copper (II) and lead (II) by defatted papaya seed, Karbala Int. J. Modern Sci. 2 (2016) 20–28. [15] C. Gok, Biosorption of uranium(VI) from aqueous solution using calcium alginate beads, J. Hazard. Mat. 168 (1) (2009) 369–375. [16] M. Grachev, I. Zubkov, I. Tikhonova, et al., Extensive contamination of water with saxitoxin near the dam of the Irkutsk Hydropower Station reservoir (East Siberia Russia), Toxins 10 (10) (2018) 402. [17] R.R.L. Guillard, Culture of phytoplankton for feeding marine invertebrates, in: W.L. Smith, M.H. Chanley (Eds.), Culture of Marine Invertebrate Animals, Plenum Press, New York USA, 1975, pp. 26–60. [18] S. Hall, G. Striachartz, E. Moxzydlowski, A. Ravindran, P.B. Reichardt, The saxitoxins: Sources, chemistry and pharmacology, in: S. Hall, G. Strichartz (Eds.), Marine Toxins: Origin, Structure and Molecular Pharmacology, American Chemical Society, Washington DC, 1990. [19] A.G. Haubois, V.M. Bricelj, J. Naar, Transfer of brevetoxins to a tellinid bivalve by suspension and deposit-feeding and its implications for clay mitigation of Karenia brevis blooms, Mar. Biol. 151 (5) (2007) 2003–2012. [20] R. Karthik, S. Meenakshi, Removal of Cr(VI) ions by adsorption onto sodium alginate-polyaniline nanofibers, Int. J. Biol. Macromol. 72 (2015) 711–717. [21] R. Kecili, C.M. Hussain, Mechanism of adsorption of nanomaterials, Nanomater. Chromatogr. (2018) 89–115. [22] T. Kubo, Y. Tominaga, F. Watanabe, K. Kaya, K. Hosoya, Selective adsorption on water-souluble ionic compounds by interval immobilization technique based on molecular imprinting, Anal. Sci. 24 (2008) 1633–1636. [23] J.F. Lawrence, C. Menard, Liquid chromatographic determination of paralytic shellfish poisons in shellfish after pre-chromatographic oxidation, J. Assoc. Off. Anal. Chem. 74 (6) (1991) 1006–1012. [24] G. Limousin, J.P. Gaudet, L. Charlet, S. Szenknect, V. Barthes, M. Krimissa, Sorption isotherms: a review on physical bases, modeling and measurement, Appl. Geochem. 22 (2007) 249–275. [25] G. McKay, Y.S. Ho, J.C.Y. Ng, Biosorption of copper from waste waters: a review, Sep. Purif. Methods 28 (1) (1999) 87–125. [26] S.P. Melegari, W.G. Matias, Preliminary assessment of the performance of oyster shells and chitin materials as adsorbents in the removal of saxitoxin in aqueous

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