Degradation of Microcystin-LR in water: Hydrolysis of peptide bonds catalyzed by maghemite under...

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Applied Catalysis B: Environmental 160–161 (2014) 597–605

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Applied Catalysis B: Environmental

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egradation of Microcystin-LR in water: Hydrolysis of peptide bondsatalyzed by maghemite under visible light

anfen Fanga,b, Yu Zhangb, Wanhong Maa,∗, David M. Johnsona,b, Ying-ping Huanga,b,∗

Innovation Center for Geo-Hazards and Eco-Environment in Three Gorges Area, Yichang 443002, Hubei Province, ChinaEngineering Research Center of Eco-environment in Three Gorges Reservoir Region, Ministry of Education, China Three Gorges University,ichang 443002, China

r t i c l e i n f o

rticle history:eceived 2 February 2014eceived in revised form 27 April 2014ccepted 11 May 2014vailable online 4 June 2014

eywords:aghemiteicrocystin-LR

a b s t r a c t

Maghemite, an abundant iron-oxide mineral, was investigated as a visible light catalyst for decomposingand mineralizing Microcystin-LR (MC-LR) in water using hydrogen peroxide (H2O2) as the initial oxidant.Interestingly, of the twelve intermediates identified by LC–MS, half were products of MC-LR peptide bondhydrolysis rather than the oxidation products typical of photocatalytic oxidation (PCO). The catalytichydrolysis of MC-LR by maghemite and visible light has not been previously reported. Detailed examina-tion of intermediates provided insight into the catalytic pathway. Electron spin resonance (ESR) spectrarevealed that visible light irradiation does not increase hydroxyl radical (•OH) production from H2O2

decomposition. Visible light appears to play a new role in this unique PCO system, generating Brønsted•

ydrolysiseptide bonds

sites on the maghemite surface via a metal ligand charge transfer (MLCT), rather than producing OHfrom H2O2. Because opening the cyclic peptide structure detoxifies MC-LR, this catalytic system could beuniquely efficient in detoxifying MC-LR in the presence of relatively high levels of natural organic matter(NOM). This work introduces a new means of selectively and effectively detoxifying MCs in water usingan inexpensive, environmentally friendly and bio-compatible mineral.

. Introduction

Microcystins (MCs) are cyclic peptides released by cyanobacte-ia during algal blooms in eutrophic waters and among the mostommon of the cyanotoxins [1,2]. The cyclic peptide structure,hared across MCs, was first characterized in 1986 [3]. MCs inhibithosphatase PP1 and PP2A, produce lesions in the liver and pro-ote cancer [4–7]. Microcystin-LR (MC-LR, Fig. 1) is one of the most

oxic and widely distributed MCs in natural waters and has beensed as a model contaminant for MCs in numerous investigations8]. Health-related cases in humans and animals caused by MC-LRere little reported until a fatal human incident occurred in 1996

9]. The World Health Organization (WHO) has set the provisional

The stability of the cyclic peptide structure prevents effectiveemoval of MC-LR by conventional water or wastewater treatment.

∗ Corresponding author at: Innovation Center for Geo-Hazards and Eco-nvironment in Three Gorges Area, Yichang 443002, Hubei Province, China. Tel.:86 717 6397488; fax: +86 717 6395966.

E-mail addresses: huangyp@ctgu.edu.cn, chem ctgu@126.com,ingpinghuang@126.com (Y.-p. Huang).

Among the advanced oxidation technologies (AOTs), photocat-alytic oxidation (PCO) is the most promising treatment technology[11,12]. For example, both TiO2 photocatalysis [11,13,14] andthe UV-Fenton system [15] effectively degrade MCs in waterby generating highly reactive hydroxyl radicals (•OH). Recently,a range of heterogeneous catalysts, including various iron andmanganese oxides, have been used to degrade refractory organiccompounds by activating H2O2 to produce •OH [16,17]. However,most PCO processes require UV irradiation, severely restrictingPCO application for environmental remediation because sunlightcontains only 4–5% UV. Some iron-bearing mineral oxides areactivated by visible light due to metal ligand charge transfer(MLCT) on the surface [18], but the •OH yield depends on surfaceFe(III)/Fe(II) catalytic recycling and is typically low [19]. To date,iron-bearing minerals have rarely used visible light to enhancedegradation of organic pollutants such as MC-LR [20]. Further-more, trace water pollutants such as MCs are particularly difficultto remove by •OH oxidation alone because •OH is not selectiveand other natural organic matter (NOM) is typically present at

598 Y. Fang et al. / Applied Catalysis B: Environmental 160–161 (2014) 597–605

CHN NH2

D-Glu Mdha

D-MeAsp

Fig. 1. The structure of Microcystin-LR (MC-LR). Adda is (2s,3s,8s,9s)-3-amino-9-methoxy-2,6,8-trimethyl-10-phenyldeca-4(E),6(E)-dienoic acid; d-Glu isdl

MtbpsMbicahisi[tsttb[hphH

ahauc[hscteafp

20 30 40 50 60 70

2 Theta ( )

Fig. 2. XRD pattern of maghemite. Note: the positions and intensities of the diffrac-

-glutamic acid; Mdha is N-methyl-dehydroalanine; d-Ala is d-alanine; l-Leu is-Leucine; d-Me-Asp is d-erythro-ˇ-methlaspartic acid; l-Arg is l-arginine.

In the search for a selective and effective means of degradingC-LR, hydrolysis of the cyclic peptide bonds could be a break-

hrough in visible light assisted PCO. Selectivity is inherently highecause multiple peptide bonds are rare in most NOM and organicollutants. Detoxification is effective because catalytic hydroly-is of a peptide bond opens the stable macrocyclic structure ofC-LR, altering the binding site and significantly decreasing inhi-

ition of the phosphatase, PPA1 [4]. Dionysiou and co-workers [11]ndicated that complete MC-LR transformation by TiO2/UV photo-atalysis still leaves significant toxicity (PPA1 inhibition remainst nearly 100%). Catalytic conversion of MC-LR by TiO2 leads toydroxylation of the Adda double bonds and does not prevent bind-

ng of the Mdha moiety of MC-LR to PPA1. In contrast, when amall amount of H2O2 was added to this TiO2/UV system, PPA1nhibition by MC-LR degradation products rapidly approached zero11]. This could be attribute to oxidative hydrolysis of MC-LR pep-ide bonds mediated by Brønsted sites generated by H2O2 on theurface of the TiO2 catalyst. It has been demonstrated previouslyhat ring opening of cyclic peptide bonds of Microcystin-LR by bac-erial microcystinase effectively renders the compound nontoxicy reducing the interaction with the target protein phosphatase21]. Recently, N-protected and free Adda amino residues from theydrolysis of Microcystin-LR was proved nontoxicity [22]. Com-ared with mineralization, detoxification of MCs by photocatalyticydrolysis of peptide bonds obviously requires small quantities of2O2, making MC treatment in water more economic.

Hydrolysis of peptide bonds can be catalyzed by Brønsted acidsnd bases and metal complexes. Glycylglycine, for example, isydrolyzed with a half-life of approximately 2 days in 1 M NaOHnd 150 days in 1 M HCl at 25 ◦C, but with a half-life of several yearsnder neutral pH conditions [23,24]. Peptide bonds are rapidlyleaved by complexes of copper, molybdenum and zirconium25–27], but metal complexes are expensive and some producearmful environmental effects. Recently, peptide bond hydroly-is at the water–pyrite interface under extreme thermodynamiconditions has been reported [28,29] and the results are relevanto biogeochemistry and environmental chemistry. Maghemite, forxample, has great potential for decontaminating soils, sediments,nd water [30]. However, production of Brønsted sites on the sur-ace of maghemite in a PCO system that catalyze hydrolysis of MCeptides has not been reported.

In the work reported here, maghemite was used as a vis-ble light catalyst to degrade MC-LR in water at neutral pHsing H2O2 as the initial oxidant. Hydrolytic cleavage of thetable ring structure facilitates oxidative decomposition. The

tion lines of �-Fe2O3 (JCPDS file 39-1346) are indicated by tic marks just above thehorizontal axis.

unique degradation products of MC-LR were identified using liquidchromatography-mass spectroscopy (LC–MS) and •OH formationwas monitored using electron spin resonance (ESR). Our resultsare quite different from previously investigated PCO systems:(1) catalytic hydrolysis reactions are important to the degrada-tion of MC-LR by the maghemite photocatalyst and (2) visiblelight irradiation does not increase production of •OH from H2O2decomposition but increases H2O2 utilization efficiency to 100%for MC-LR degradation. The maghemite PCO system reported hereintroduces a promising new approach for selectively and effec-tively detoxifying MCs in treatment of water with high levelsof NOM.

2. Materials and methods

2.1. Reagents

Maghemite samples were provided by the Central-SouthResearch Institute of Metallurgical Geology (Hubei, China). Themaghemite was washed (1.6 m3 water/t mineral) to remove nat-ural contaminants, ground and sieved (100-mesh) to obtain crystalparticles ≤150 �m. The maghemite was characterized using X-raydiffraction (XRD) (Brucker D8 Advance II, Germany) with a CuK� radiation source and a scan rate of 2.4◦/min from 20◦ to 80◦.Results indicate the typical hexagonal structure of maghemite withunit-cell parameters a = b = c = 8.351 A. The characteristic peaks ofthe primary components (Fig. 2) are designated with asterisks(*) and they compare well with the reference material (�-Fe2O3,JCPDS 39-1346). The specific surface area was 1.68 m2/g as deter-mined by BET surface area analysis (Micromeritics ASAP 2020,USA).

MC-LR (95%; FW 995.2 g/mol) was purchased from ExpressTechnology Co., Ltd (China). A 50 mg/L MC-LR standard solution wasobtained by dissolving 0.25 mg MC-LR in 5 mL of methanol (HPLCgrade). The standard was stored at −20 ◦C and 5.0 mL test solutionscontaining 2.0 mg/L MC-LR were prepared by mixing 0.2 mL MC-LRstandard solution with 4.80 mL of water (autoclaved by Milli-Q®

Synthesis A10, Millipore Corp., Billerica, MA, USA). Standard Ful-vic acid sample (adjusted by Nordic lake fulvic acid obtained from,

Y. Fang et al. / Applied Catalysis B: Environmental 160–161 (2014) 597–605 599

–48.

.1–4

–47.

.1–4

–22.

.3–2

–25.

.0–3

–46.

.0–5

–49.

.5–2

–27.

8.7–

–10.

–17.

–27.

c 1, c

e 1e 2

h 1h 2

i 1i 2

).0 2 4 6 8

Fig. 3. MC-LR (2.0 mg/L) degradation curves at neutral pH in maghemite (240 mg/L)−4

(methanol/H2O, v/v 0.2/4.8). (a) dark/maghemite/MC-LR; (b) vis/MC-LR/H2O2; (c)dark/maghemite/MC-LR/H2O2; (d) vis/maghemite/MC-LR/H2O2.

2.2. Photocatalytic process

Photocatalytic reactions were carried out in a PCO chamber(Xujiang Mechanical and Electrical Factory, Nanjing). The visiblelight source was a 250-W xenon lamp (Institute of Electric Lightsource, Beijing, China) positioned inside a cylindrical Pyrex reac-tor and surrounded by a circulating water jacket for cooling. Toensure illumination by visible light only, a cutoff filter was placedoutside the Pyrex jacket to eliminate radiation with � < 420 nm.10 mL of 2.0 mg/L MC-LR and 2.4 mg of maghemite powder wasplaced in the Pyrex reaction vessel. Prior to irradiation, the sus-pensions were magnetically stirred in the dark for 2 h to ensureadsorption/desorption equilibrium established. Hydrogen perox-ide (H2O2) was then added to the reaction vessel to give an initialconcentration of 1.0 × 10−4 mol/L and the pH was adjusted to

0 5 10 15 20 250.0

TOC/TOC0

Fig. 4. Change in TOC during MC-LR (2.0 mg/L) degradation at neutral pH inmaghemite (240 mg/L) and H2O2 (1 × 10−4 mol/L) under the following conditions:5 mL MC-LR solution (methanol/H2O, v/v 0.2/4.8). (a) dark/maghemite/MC-LR;(b) vis/MC-LR/H2O2; (c) dark/maghemite/MC-LR/H2O2; (d) vis/maghemite/MC-LR/H2O2.

600 Y. Fang et al. / Applied Catalysis B: Enviro

0:1 10: 1 20:1 50:10

CFA/CMC-LR

46.8% 47.9%

Fig. 5. Effect of FA on the removal efficiency of MC-LR.240 mg/L maghemite, cH2O2 =1 × 10−4 mol/L, cMC-LR = 2.0 mg/L, cFA/cMC-LR = 0:1, 10:1; 20:1 and 50:1, neutral pH,irradiated by visible light for 4 h. Standard Fulvic acid sample (adjusted by NordiclS

2uwrt4twG

5sloar

ake fulvic acid obtained from 1R105F) was purchased from International Humicubstance Society (HISS).

atalyst particles. The filtrate was analyzed using the methodsescribed below.

.3. Analytical methods

MC-LR was determined using HPLC (Waters 600 HPLC, Waters998 photodiode-array detector, USA) with a Kromasil C18 col-mn (4.6 mm × 150 mm, 10 �m particle size). The mobile phaseas water and methanol (65:35, v/v) containing 0.05% trifluo-

oacetic acid (TFA). The flow rate, injection volume, columnemperature and detection wavelength were 0.8 mL/min, 20 �L,0 ◦C and 238 nm, respectively. Under these conditions, the elu-ion time for MC-LR was ∼8.9 min. Total organic carbon (TOC)as determined with a TOC analyzer (Jena N/C2100 TOC analyzer,ermany).

Hydroxyl radical levels were monitored by ESR (Brucker ESP00E, Germany) with DMPO (5,5-dimethyl-1-pyrroline-N-oxide)pin trapping. The light source was a Quanta-Ray ND:YAG pulsed

0 10 20 0.0

Intens.

ig. 6. Total ion chromatogram (TIC) obtained during MC-LR (2.0 mg/L) degradation at night irradiation.

nmental 160–161 (2014) 597–605

Reaction intermediates were monitored using LC/MS (AgilentG1315D DAD/amazon SL, USA) with full scanning (m/z 300–1200)in the positive ion mode. Gradient elution was used with formicacid (pH = 2.6) and acetonitrile as the mobile phase. Gradientelution was programmed as 0–20% acetonitrile (10 min) fol-lowed by an increase to 35% (10 min), 60% (15 min), and 80%(10 min).

Toxicity testing of MC-LR before and after the treatment: 19male and 19 female Kunming mice (18–22 g, specific pathogen-free) were used for the study. All mice were housed in animalrooms. These mice were maintained on a 12 h light/dark cycleand had no restriction of eating or drinking. The research wasperformed in accordance to the “Guide for the Care and Use of Labo-ratory Animals” (China Three Gorges University). The untreated andtreated (24 h degradation) MC-LR solutions (2 mg/L) were admin-istered by gavage injection (i.g.) or intraperitoneal injection (i.p.)at a dose of 80 �g/kg body weight after the animals had fastedfor 18 h. The test animals were randomly divided into 4 groups,each group including an equal number of male and female mice;group A (control group, i.p., 8 mice), group B (untreated solution,i.g., 10 mice), group C (untreated solution, i.p., 10 mice) and groupD (treated solution (24 h), i.p., 10 mice). Saline was injected intoanimals in the control group and all animals were released fromfasting after being injected. The toxic response following i.g. injec-tion is significantly less than the response produced by i.p. [31].injection. All animals were observed daily throughout the 7 day testperiod.

3. Results and discussion

3.1. Photocatalytic degradation of MC-LR

The ability of maghemite to photocatalytically degrade MC-LR under different conditions is displayed in Fig. 3. Degradationof MC-LR without H2O2 was negligible in the dark after 8 h(a), indicating that maghemite alone does not catalyze oxida-tive degradation or hydrolysis of MC-LR. Nearly 20% of MC-LRwas decomposed under irradiation when H2O2 was added with-out maghemite (b). In the dark, a small quantity of MC-LR wasdecomposed by the maghemite/H2O2 system (c) with reaction rateconstant k = 0.04 min−1, according to pseudo-first order kinetics.The maghemite catalyst activates H2O2 and partially degrades MC-LR without light irradiation. When the system containing bothmaghemite and H2O2 was irradiated with visible light, a very rapid

30 40 50 Time [min]

a, b, c

eutral pH in maghemite (240 mg/L) and H2O2 (1 × 10−4 mol/L) after 5 h of visible

CHN NH2

C49H74 N10O12

m/z 995

D-Glu Mdha

D-MeAsp

C44 H69N9O10

m/z 884

product d

=D-Me Asp*

=D-Ala

CHN NH2

C35H53N7O8

m/z 700

produ ct b

NH2C39H62 N6O9

m/z 759product g

C39H62N6O8

m/z 743

produ ct a

Guanidy l oxi dation

Guanidy l an d

Carboxyl oxidation

=D-Glu

=D-Glu*=Mdha*

Carboxy l oxidati on

CHN NH2

a. Cataly tic hydrolys is of MC-LR p eptide bo nds wit hout Adda oxi dat ion

Fig. 7. Proposed reaction sequences for photocatalytic hydrolysis and partial oxidative hydrolysis of MC-LR. (a) Catalytic hydrolysis of MC-LR peptide bonds without Addao e Add

auMrwtip(

xidation; (b) catalytic hydrolysis of MC-LR peptide bonds after hydroxylation of th

isible light and maghemite to selectively degrade MC-LR by pep-ide bond hydrolysis.

The primary decomposition reaction could be either the cat-lytic hydrolysis of peptide bonds or the catalytic oxidation of thensaturated bonds of MC-LR. The primary decomposition route ofC-LR was determined by comparing TOC removal under various

eaction conditions since hydrolysis of MC-LR peptide bonds aloneould not lead to a change in TOC. Results are shown in Fig. 4 and

he general trend is consistent with the data from HPLC analysis ofntermediates (see Table 1). There was little change in TOC in theresence of maghemite alone (a) for 25 h. The other two controls,b) and (c), produced similar (∼15%) reductions in TOC resulting

a benzene ring.

from the degradation (∼25%) of MC-LR. This indicates that H2O2decomposes to produce •OH with and without irradiation. The •OHinduces degradation similar to that of the Fenton reaction and oxi-dizes most organic compounds to CO2 and H2O. However, this TOCreduction under visible light irradiation (Fig. 4d) was significantlyenhanced (up to 58% TOC removal after 25 h), approaching 100%H2O2 utilization efficiency, based on Eq. (1).

C49H74N10O12 + 148H2O2 → 49CO2 + 180H2O + 10HNO3 (1)

The maghemite catalyst significantly improves the efficiencyof H2O2 utilization, even in the presence of methanol (∼1.6 M,about 106 times the concentration of MC-LR). To gain insight on

602 Y. Fang et al. / Applied Catalysis B: Environmental 160–161 (2014) 597–605

Fig. 7. (Continued ).

CHN NH2

C49H74N10O12

m/z 995

C49H76N10O14

m/z 1029

produ ct e1-3

C37H58N10O12

m/z 83 5

product j

C34H54N10O12

m/z 795

produc t k

C33H54N10O12

m/z 783

product l

CHN NH2

C37H58N10O13

m/z 85 1

prod uct f

Fig. 8. Intermediates that belong to traditional oxidative pathways for MC-LR degradation by the maghemite system.

he decomposition of MC-LR, the position of the hydrolyzed MC-LReptide bonds were determined by examining the decomposition

ntermediates with LC–MS.

egradation efficiency was investigated. Fulvic acid (FA) was usedo represent the NOM present in practical treatments [32] anddded to the maghemite photocatalytic system used to degrade

MCLR. As shown in Fig. 5, a low concentration of FA (cFA/cMC-LR < 10)has a negligible effect on the degradation of MC-LR. Even whenthe FA concentration was enhanced to 50 times of that of MC-LR,

6 nvironmental 160–161 (2014) 597–605

uiiIpimhhd

dPpacouapoPmd

y1sMab(aip•

t(Maaildi(tp

3280 33 00 332 0 3340 3360 3380 3400

Magnetic Field(G)

04 Y. Fang et al. / Applied Catalysis B: E

.2. Identification of intermediates by LC–MS

After a reaction time of 5 h at least twelve intermediate prod-cts could be identified in the total ion chromatogram (TIC), shown

n Fig. 6. That every of them changes with different reaction timess further observed by TIC in details and summarized in Table 1.n Table 1, the elution times, mass to charge ratios (m/z), andeak intensities at various irradiation times are shown for the 12

ntermediates and substrate MC-LR. Among all 12 detected inter-ediates, six resulted from hydrolysis of peptide bonds (a–d, g and

), while the other six are typical PCO products (e, f and i–l). Theydrolysis products were rarely reported in the literatures on PCOecomposition of MC-LR in water.

The hydrolysis of MC-LR peptide bonds is rapid and loss of a-MeAsp fragment produces intermediate product d (m/z 884).roduct b (m/z 700) is then formed by hydrolytic cleavage of theeptide bond connecting Mhda and d-Ala. Products a (m/z 743)nd g (m/z 759), the primary hydrolysis products, are remnantsontaining d-Glu and Mdha (see Fig. 7a). With products d and b, noxidation of MC-LR occurs. The conjugated diene in Adda readilyndergoes oxidative hydroxylation, but remains intact in products, b, d and g; the photocatalytic system preferentially hydrolyseseptide bonds. In the minor products d and g, the guanidyl groupsn l-Arg is converted to a terminal amine group, consistent withCO decomposition of the Adda segment [12]. However, they areinor components in the TIC of LC–MS and are formed even in the

ark.Product i1–2 (m/z 1013), with two isomers, contains the

ydroxylated phenyl ring of Adda (Fig. 6b). Product c1–2 (m/z18; only one of the two possible isomers was detected) and1–2 (m/z 829) result from successive catalytic hydrolysis of theeptide bonds connecting d-MeAsp to l-leu and Mdha to d-AlaFig. 7b).

The peak for product b (m/z 700), rising from the base line to aaximum and then returning to the baseline (Table 1), indicates

he cyclic structure of MC-LR was cleaved by catalytic hydrolysis.The common oxidation products in a PCO process, such as prod-

cts e, j–l, f (m/z 1029, 835, 795, 783, 851), were also detectedithout concomitant hydrolysis of peptide bonds (Fig. 8).

It is well known that species e (m/z 1029) is formed by dihydrox-lation of the unsaturated bonds of Adda by •OH attack, forming the, 2 and 1, 4 isomers (see right side of Fig. 8) [33]. However, in theystem under investigation, dihydroxylation is not important forC-LR degradation (see Fig. 6 and Table 1). Product f (m/z 851) is

n oxidation product formed, not by hydrolysis of peptide bonds,ut by carbonylation of the methyl group on Meha of product jm/z 835) as shown in Fig. 8. Most of the oxidation products, suchs product a–d, g and h, were not similar to those produced by typ-cal PCO, so the present system must involve alternate degradationathways. Intermediate product fragments that do not occur in aOH dominated process of MC-LR degradation can help explain theey role of catalytic hydrolysis in efficient use of H2O2 and visibleight irradiation.

The last four products a, c, d and g (m/z 743, 918, 884, 759, respec-ively), produced by hydrolysis of peptide bonds in the MC-LR ringsee the last line of Table 1) are more difficult to eliminate than

C-LR itself. This indicates that the catalyst preferentially inter-cts with the intact cycle structure of MC-LR, composed of sevenmino acids linked by peptide bonds (Fig. 1). After the macrocycles opened even by a single hydrolysis, the maghemite catalyst hasittle ability to promote further decomposition. Opening the cycleetoxifies MC-LR, the toxicity of MC-LR was significantly decreased

n the photocatalytic degradation process in the toxicity testingsee Fig. S1). Therefore, the selectivity of the maghemite catalystoward the cyclic structure makes this catalytic system particularlyromising.

Fig. 9. ESR spectral changes of DMPO-•OH adducts with time of irradiation.Maghemite powder: 240 mg/L; [DMPO] = 1.3 mol/L; [H2O2] = 1 × 10−4 mol/L; [MC-LR] = 2.0 mg/L.

3.3. Proposed degradation pathway

To clarify the role of visible light irradiation in the presentmaghemite photocatalytic system, either accelerating H2O2 splitinto •OH or other catalytic reaction, ESR with spin-trapping (DMPO)was used to observe the production and change of •OH free-radicalduring both the dark reaction and visible light catalytic reaction(Fig. 9). The DMPO-•OH adduct signature is a signal with intensityratio 1:2:2:1 and its presence is a clear indication that •OH forms inthe dark. Visible light irradiation did little to increase signal inten-sity, indicating that excitation by visible light does not enhancethe formation of •OH. This is highly consistent with the findingsof Kwan and Voelker [34]; Given that the significantly enhancedMC-LR degradation (Figs.3 and 4c vs d) caused by visible light irra-diation, now it was very distinct that •OH free-radical formed onthe maghemite surface is not the chief contributions to degradationof MC-LR in the present photocatalytic system.

Based on ESR observations of •OH and the fact that catalyti-cally hydrolyzed products dominate in MC-LR decomposition byLC–MS, it is postulated that H2O2 interacts with the maghemitesurface, which is then activated by visible light to producetransient Brønsted base sites and high valence iron (Eq. (3)).Brønsted base sites and the Fe(IV) O species on the maghemitesurface then hydrolyze the peptide bonds and oxidize MC-LR(Eq. (4)). This mechanism would not produce more •OH undervisible light irradiation because the H2O2 decomposition, cat-alyzed by Fe(II)/ Fe(III) (Eq. (2b)), proceeds very rapidly, evenin the dark. Visible light plays a crucial role in the formation ofBrønsted base sites (Eq. (3)) and regeneration of active ferric sites(Eq. (4b)).

≡ Fe(III)H2O2 →≡ Fe(III) (OOH) + H+ (2a)

H2O2≡Fe(II)/≡Fe(III)−→ •OH + −OH(dark reaction, no increase in

•OH with illumination) (2b)

≡ Fe(III) (OOH) + Fe(III)adjacentvis−→ ≡ Fe(IV) O

+ ≡ Fe(III) −OH(Bronsted base) (3)

≡ Fe(IV) O + MC-LRvis−→ →≡ Fe(II) = HO-MC-LR (4a)

Mfpviottstao

tS(SacT

[[[[[[[

Y. Fang et al. / Applied Catalysis B: E

Note: Fe(III) represents active ferric sites on the maghemiteurface, to which H2O adsorbs or coordinates prior to catalyticction.

To completely elucidate this possible mechanism, more directvidence will be necessary; for example, in situ analysis ofe(III)/Fe(II) transformation by XPS and FTIR spectroscopy. We areorking fast to approach to this mechanism.

. Conclusions

Maghemite was employed as a photocatalyst to decomposeC-LR in water. Based on H2O2 consumption, both MC-LR trans-

ormation and TOC removal occurred at near 100% efficiency. Theroduction of •OH in the dark was the same as under visible light,ery atypical of PCO systems. Twelve intermediate products weredentified by LC–MS and half originated from catalytic hydrolysisf the peptide bonds of MC-LR. Based on these results, we postulatehat H2O2 is used to generate both high valence iron (Fe(IV) O) andhe Brønsted base, Fe(III) −OH, via a MLCT process. The catalyticystem selectively detoxifies MC-LR by predominantly hydrolyzinghe peptide bonds of the ring structure. Because the maghemite cat-lyst targets peptide bonds, detoxification of MC-LR in the presencef NOM appears feasible.

cknowledgments

This work was funded by the National Natural Science Founda-ion of China (Nos. 21377067, 21207079 and 21177072), Naturalcience Foundation for Innovation Group of Hubei Province, ChinaNo. 2009CDA020) and Hubei Provincial Department of Educationcience Research Project Youth Talent Project (Q20141208). Welso would like to thank Dr. Zhao Bo of Department of Pharma-ology, Medical Science College (CTGU, China) for his assistance inoxicity Testing.

ppendix A. Supplementary data

Supplementary data associated with this article can beound, in the online version, at http://dx.doi.org/10.1016/.apcatb.2014.05.016.

nmental 160–161 (2014) 597–605 605

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