974

Chemistry Letters Vol.35, No.8 (2006)

Direct Detection of Redox Reactions of Sulfur-containing Compounds
on Ferrite Nanoparticle (FP) Surface
Kosuke Nishio,1 Nobuyuki Gokon,2 Shingo Tsubouchi,1 Morihito Ikeda,1 Hiroki Narimatsu,1Satoshi Sakamoto,1 Yasuo Izumi,3 Masanori Abe,4 and Hiroshi Handa1
1Department of Biological Information, Graduate School of Bioscience and Biotechnology,
Tokyo Institute of Technology, Yokohama 226-8501
2
Department of Chemistry, Chemical Engineering Material Chemistry,
Graduate School of Science and Technology, Niigata University, Niigata 950-2181
3
Department of Environmental Chemistry and Engineering, Interdisciplinary Graduate Schoolof Science and Engineering, Tokyo Institute of Technology, Yokohama 226-8502
4
Department of Physical Electronics, Graduate School of Science and Engineering,Tokyo Institute of Technology, Tokyo 152-8552(Received May 1, 2006; CL-060516; E-mail: hhanda@bio.titech.ac.jp)
Direct observation of sulfur sites for sulfur-containing compounds (cysteine and its oxidative derivatives) on ferrite nanoparticles (FP) was performed by X-ray adsorption near edge
structure (XANES) measurements. XANES spectra of cysteine
derivatives conjugated with FP indicated that redox reactionsoccurred on the FP surface. Cysteine was completely oxidized
to cystine and cysteinesulﬁnic acid and cysteic acid were partially reduced. These studies revealed that the FP surface should
possess both an oxidation and reduction reactive center for sulfur-containing functional groups.

Recently, molecules possessing multiple anionic groups
such as citric acid, cysteine, and dimercaptosuccinic acid havebeen noticed because of their strong coordination to iron ion
on FP.1–3 These molecules could be adsorbed onto FP by simply
mixing an aqueous solution without any complicated process.
This easy adsorption process is quite important for various biotechnological applications such as bioseparation, magnetic resonance imaging (MRI), and hyperthermia.4 If there are functionalgroups that do not participate in immobilization to FP, they arethen exposed on the FP surface. As a result, the compound-conjugated FP possesses high dispersibility in liquid media. Thus,
they work as a dispersant and it is expected that they decorate
the FP surface. However, insuﬃcient understanding and lackof information about the binding conﬁguration of moleculesand surface adsorption reactions on FP have prevented researchand development toward various biological applications. Thereare two major reasons that make their analyses hard. First is
an extremely trace amount of these adsorbed molecules on FP
relative to the iron atoms in FP. Second is complicated reactivityof iron ions on FP relative to another molecule. Borghi et al.
reported a surface reductive dissolution of ferrite by mercaptocarboxylic acid.5 In adsorbing various molecules, an oxidation/reduction reaction occurred between a ferrite and the absorbing molecules making analysis of the reaction mechanism
especially complicated. To analyze a reaction mechanism on
ferrite, understanding of the binding conﬁguration of these molecules on ferrite is very important.
In our previous study, we found that cysteine, cystine, andcysteic acid were eﬃciently immobilized onto FP.6 Our result
indicated that there was a selective adsorption of sulfur atom
onto FP and/or a strong interaction between sulfur and iron on

the FP surface. Therefore, we focused on uncovering the surface
reaction between a sulfur functional group and an iron ion on FP.
We describe herein a chemical state of the sulfur atoms of cysteine and its analogs (cystine, cysteinesulﬁnic acid, and cysteicacid) adsorbed onto FP surface measured by XANES spectra.
We synthesized FP through conventional chemical coprecipitation. All procedures in preparation of samples were carriedout in glove box under N2 to prevent oxidation of FP. A 25mL portion of 0.1 M aqueous iron solution (FeCl2 /FeCl3 =1/2) was added into 50 mL of 0.2 M aqueous NaOH to give
FP suspension (Produced FP is a mixed solid solution between
Fe3 O4 and -Fe2 O3 and its size was around 8 nm.). Cysteine(75 mmol) or its derivatives (cystine, cysteinesulﬁnic acid, andcysteic acid) was dissolved in 0.2 M aqueous NaOH. Then,
1 mL of alkaline soution of cysteine or its derivatives was added
to the obtained FP suspension. After vigorous stirring for 1 h atroom temperature, cysteine-conjugated FP was collected magnetically and washed with MilliQ water. The conjugated FPwas dried under vacuum to give a black plate. After grindingof the plate-like FP under N2 , the FP powder was put on Kapton
tape to measure XANES spectra in ﬂuorescence detection modeat KEK-PF (Tsukuba, Japan) using beamline-9A with Si(111)
double crystal monochromator under ring conditions of 2.5
GeV and 348.5–347.3 mA. The entire path of the X-ray beam
was in He atmosphere. The monochromator was tuned to 65%
intensity, and rhodium-coated mirror was used both for suppressing higher harmonics. As standard sulfur K-edge XANES
spectrum, Na2 S2 O3 5H2 O was measured (2472.0 eV).
The XANES spectra of cysteine and its derivative-FP areshown in Figure 1, and the observed peak energies of sulfur
functional group are summarized in Table 1. The spectrum ofcysteine–FP gave two strong peaks (2472.7 and 2474.2 eV),which corresponded to each reference peak of cystine completely (2472.7 and 2474.2 eV, respectively).7 These spectra indicatethat a thiol group in cysteine was transformed into a disulﬁdeby oxidation to produce cystine on FP. This oxidation reaction(disulﬁde formation) was not observed in cysteine without FP.
XANES spectrum of cysteine–FP also provided signiﬁcant
information on the binding conﬁguration of cysteine oxidized
on FP. If there is any chemical bond between a sulfur atom
and an iron atom on the FP surface, there must be an energy shiftof sulfur K-edge in the XANES spectrum. Our XANES measurement of cysteine–FP exhibited no energy shift. Therefore, it is.

Copyright  2006 The Chemical Society of Japan

Chemistry Letters Vol.35, No.8 (2006)

975
Table 1. Observed peaks energies in cysteine and its derivatives conjugated to FP and observed sulfur K-edge energy peaks(reference peaks) in cysteine and its derivative

Normalized Intensity

SampleCysteine–FP
Cystine–FP
Cysteine sulﬁnic acid–FP
Cysteic acid–FP

Cysteine–FP

Cysteine
CystineCysteine sulﬁnic acid
Cysteic acid

Cystine–FP

Cysteinesulfinic acid–FP
Cysteine
CystineCysteinesulfinic acid
Cysteic acid

Cysteic acid–FP

2465

2470

State

2475
2480
Energy / eV

2485

2490

Figure 1. Sulfur K-edge XANES spectra of cysteine, cystine,cysteinesulﬁnic acid, and cysteic acid-conjugated FP, as well
as reference spectra of the unconjugated parent compounds.
The black triangles indicate pre-edge feature (peak) appearedin cysteinesulﬁnic acid– and cysteic acid–FP.
expected that oxidized cysteine (or cystine itself) molecule binds
to FP through carboxyl and/or amino groups. This result is consistent with the reported reductive dissolution of iron oxide by
thiol-containing molecules.8 Since no characteristic change
was observed in the XANES spectrum of cystine–FP, a redox
reaction between cystine and FP did not occur.
The main XANES peaks in cysteinesulﬁnic acid–FP (2477.4eV) and cysteic acid–FP (2481.3 eV) did not change comparedto those of cysteinesulﬁnic acid and cysteic acid, respectively.
Their sulfur functional groups did not react with FP. However,we observed an interesting weak pre-edge feature in cysteinesulﬁnic acid–FP and cysteic acid–FP. The pre-edge feature in
XANES spectrum of cysteinesulﬁnic acid–FP appeared at2472.7 and 2474.2 eV (Figure 1), which corresponds to theintense peaks of disulﬁde in cystine. Cysteic acid–FP showed
an apparent pre-edge feature at 2476.7 eV (Figure 1), which
was slightly lower than that of sulﬁnic acid. This peak did notcorrespond to any cysteine derivative (Table 1). This featureindicates the presence of reduced compounds derived from cysteic acid. Although we could not identify this species at present,
this might be a deoxidized product with a sulfur functionalgroup. From these results, we assumed that these peaks would
indicate the existence of a reactive center on FP surface wheresulﬁnic acid and/or sulfonic acid functional groups were deoxidized.
In summary, we directly observed an oxidative transforma-Peak energy/eV
2472.7, 2474.2
2472.7, 2474.2
2472.7, 2474.2, 2477.4
2476.7, 2481.3–SHa–S–S–a–SO2 Ha–SO3 Ha2473.42472.7, 2474.2
2477.42481.3a–SH, –S–S–, –SO2 H, and –SO3 H are corresponding to cysteine, cystine, cysteinesulﬁnic acid, and cysteic acid, respectively. The sulfur K-edge XANES spectrum of Na2 S2 O3 5H2 O
was measured as standard (2472.0 eV).

.

tion of cysteine into cystine and a reduction of cysteinesulﬁnicacid to a disulﬁde compound, as well as cysteic acid to a reduced
compound on FP surface in XANES. Disulﬁde did not undergo
oxidation and/or reduction. Cysteine was oxidized to cystinecompletely on FP and cysteinsulﬁnic acid and cysteic acid werereduced partially on FP. These results would mean that a surface
redox potential energy of FP is quite close to that of disulﬁdegroup. Currently, examination of the XANES spectra of othersulfur-containing compounds is underway in an eﬀort toelucidate binding information between FP and sulfur-containingcompounds.
This work was supported by a Grant for Research andDevelopment Projects in Cooperation with Academic Institutions from the New Energy and Industrial Technology Development Organization (NEDO). We also thank Ms. Yuka Masaike,Ms. Akiko Tsuchitani, Mr. Yuusuke Mochizuki, Mr. Shotaro
Nomura at Tokyo Institute of Technology for helpful advice
and discussion.
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Published on the web (Advance View) August 5, 2006; doi:10.1246/cl.2006.974