Nano Res.

Electronic Supplementary Material

Improved peroxidase-mimic property: Sustainable, high-efficiency interfacial catalysis with H2O2 on the surface of vesicles of hexavanadate-organic hybrid surfactants

Kun Chen1,2, Aruuhan Bayaguud1, Hui Li2, Yang Chu2, Haochen Zhang1, Hongli Jia1, Baofang Zhang2,

Zicheng Xiao3, Pingfan Wu3 (), Tianbo Liu2 (), and Yongge Wei1,4 ()

1 Department of Chemistry, Tsinghua University, Beijing 100084, China 2 Department of Polymer Science, University of Akron, Akron, Ohio 44325, USA 3 Institute of POM-based Materials, Hubei University of Technology, Wuhan 430065, China 4 State Key Laboratory of Natural and Biomimetic Drugs, Peking University, Beijing 100191, China

Supporting information to DOI 10.1007/s12274-017-1746-5

Materials and Methods

Reagents and Measurement. Propionic anhydride and myristic andydride were purchased from TCI. Bu4NBr

was purchased from Beijing Chemical Industry Group Co., Ltd. (Beijing, China). 4-Dimethylaminopyridine

(DMAP), and 3,3’,5,5’-tetramethylbenzidine (TMB) were purchased from Sigma-Aldrich. Triethylamine,

NaVO3·2H2O, pentaerythritol and other chemicals were purchased from Sinopharm Chemical Reagent Co., Ltd.

(Beijing, China). All the reagents were used without further purification. All aqueous solutions were prepared

by using ultrapure water (≥ 18 MΩ, Milli-Q, Millipore). All chemicals were of analytical grade and used without

purification except acetonitrile which was dried and distilled by refluxing in the presence of CaH2 prior to use.

The IR spectra of the products were recorded on a FTIR spectrometer (Perkin–Elmer, USA) on KBr pellets in

the range of 4000-400 cm-1 with resolution of 4 cm-1. 1H NMR spectra were obtained on a JNM-ECA300

spectrometer (JOEL, Japan) at 298 K. UV/Vis absorption spectra were recorded on a UN-2100s spectrometer

(Shimadzu, Japan) at 298K. Elemental analyses were carried out using a ThermoQuest FLASH-1112 instrument

(Thermo, USA). The electrospray mass spectra (ESI-MS) were measured on a Finngan LCQ Deca XP Plus

ion-trap mass spectrometer (San Jose, CA), and experiment was carried out in the negative-ion mode using

CH3CN as the solvent. Bright-field TEM was carried out with a JEOL-1230 electron microscope with an

accelerating voltage of 120 kV. A commercial Laser light scattering (LLS) spectrometer (Brookhaven) equipped

with a solid-state laser operating at 532 nm was applied for both Static Light Scatting (SLS) and Dynamic Light

Scattering (DLS) measurements. SLS experiments were performed over a broad range of scattering angles from

30° to 120° with 3° intervals. The radius of gyration (Rg) of the solute particles was directly calculated from a

partial Zimm plot with the software provided by Brookhaven Instruments Inc. The intensity-time correlation

Address correspondence to Pingfan Wu, pingfanwu-111@163.com; Tianbo Liu, tliu@uakron.edu; Yongge Wei, yonggewei@mail.tsinghua.edu.cn

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function was measured by DLS with a BI-9000AT multichannel digital correlator. The field correlation function

|g(1)(τ)| was analyzed by the CONTIN method to yield the distribution of the characteristic line width. The

normalized distribution function of the characteristic line width G(Γ) can then be obtained and applied to

determine the apparent translational diffusion coefficient. The hydrodynamic radius Rh and the size distribution

were measured by DLS. Rh is related to D by means of the Stokes-Einstein equation, Rh = kT/6πηD, in which k is

the Boltzmann constant and η is the viscosity of the solvent at temperature T. The particle-size distribution in

solution from a plot of ΓG(Γ) versus Rh was obtained from DLS measurement. A Rh,0 of the particles was

obtained by extrapolating Rh, app to zero-scattering angle.

Synthesis of Na2[V6O13{(OCH2)3CCH2OH}2] (Na21). Compound of 1 was synthesized according to previous

reports with minor modifications [1, 2]. An amount of 10.4 g NaVO3·2H2O was dissolved in 100 mL deionized

water. 1 M HCl was dropwise added until pH = 3, and then 3.2 g pentaerythritol was added to the solution. The

mixture was stirred at 80 °C for 48 h and then filtrated. 80 mL of the dark red filtrate was carefully added to a

solution of tetrabutylammonium bromide (0.2 g/mL Bu4NBr aqueous solution) and orange solid was collected

by centrifugation. The product (Bu4N)21 was washed by 100 mL deionized water for three times and dried for

storage. The residue filtrate was evaporated the solvent and collected the crystals for usage. The obtained

crystals are Na21 and characterized. FT-TR [(KBr) ν/cm-1]: 2960(m), 2873(m), 1636(w), 1481(m), 1384(w), 1127(m),

1071(s), 1035(s), 945(vs), 811(s), 796(m), 721(s), 582(m). UV/Vis (in water): λmax = 352 nm, 247 nm.

Synthesis of Na2[V6O13{(OCH2)3CCH2OOCCH2CH3}2] (Na22). A mixture of 1.26 g (1 mmol) (Bu4N)21, 0.20 g (2

mmol) propionic anhydride, 0.01 g DMAP, 0.20 g (2 mmol) triethylamine and 20 mL CH3CN were stirred at 40 °C

for 48 h. After cooled to room temperature, the solution was poured into 50 mL deionized water and the red

precipitates were collected by filtration. Na22 was obtained by cation exchange resin. Elemental analysis for:

V6O23Na2C16H26; calc: C: 20.49%, H: 2.79%; found: C: 20.64%, H: 2.85%. 1H NMR (DMSO-d6, 300Hz) δ = 4.94 (s,

12H), 3.93 (s, 4H), 2.30 (m, 4H, J = 7.5 Hz), 0.99 (t, 6H, J = 7.5 Hz). ESI-MS ((Bu4N)21 in MeCN, negative):

[V6O13{(OCH2)3CCH2OOCCH2CH3}2]2-, m/zcal = 446.00, m/zobs = 446.09; {(Bu4N)[V6O13{(OCH2)3CCH2OOCCH2CH3}2]}-,

m/zcal = 1134.47, m/zobs = 1133.83. FT-TR [(KBr) ν/cm-1]: 2959(m), 2873(m), 1744(s), 1633(w), 1482(m), 1382(w),

1239(w), 1125(m), 1068(s), 1040(s), 952(vs), 807(s), 719(s), 582(m). UV/Vis (in MeCN): λmax = 352 nm, 246 nm.

Synthesis of Na2[V6O13{(OCH2)3CCH2OOC(CH2)12CH3}2] (Na23). The synthesis of 3 is similar to that of 2, except

the use of myristic andydride instead of propionic anhydride. The product of Na23 was obtained by cation

exchange resin. Elemental analysis for: V6O23Na2C38H70; calc: C: 36.61%, H: 5.66%; found: C: 36.52%, H: 5.61%. 1H NMR (CDCl3, 300Hz) δ = 5.21 (s, 12H), 3.99 (s, 4H), 2.26 (m, 4H, J = 7.5 Hz), 1.25 (m, 44 H), 0.86 (t, 6H, J = 7.1 Hz).

ESI-MS ((Bu4N)21 in MeCN, negative): [V6O13{(OCH2)3CCH2OOC(CH2)12CH3}2]2-, m/zcal = 600.22, m/zobs = 600.45;

{(Bu4N)[V6O13{(OCH2)3CCH2OOC(CH2)12CH3}2]}-, m/zcal = 1442.91, m/zobs = 1442.53. FT-TR [(KBr) ν/cm-1]: 2937(m),

2853(m), 1716(s), 1633(w), 1449(m), 1376 (w), 1241(w), 1122(m), 1068(s), 1036(s), 952(vs), 808(s), 720(s), 581(m).

UV/Vis (in MeCN): λmax = 351nm, 245nm.

Figure S1 ORTEP representation of the cluster anion of 3 (50% probability ellipsoid, H atoms represented in ball mode): V, yellow; C, gray; O, red; H, light gray.

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Table S1 KDApp of the 1-TMB complex at different pH.

pH KDApp (μM)

4.5 3.4

6.5 5.7

8.5 10.4

Figure S2 Schematic illustration of the structure of TMB and the catalytic reaction products and the peroxidase-like activity of the hexavanadate clusters is pH (A), temperature (B), and H2O2 concentration (C) dependent, as well as influenced by their solution behaviors (D). Experiments were carried out at 35 °C using 5 μg hexavanadate clusters in a reaction volume of 1.0 mL PBS buffer, pH 6.5, with 416 μM TMB as the substrate, 98 mM H2O2 as the oxidation agent, unless otherwise stated. The maximum point in each curve (a–c) was set as 100%. A) Hexavanadate clusters show a wide pH adaptability of 3.5~7.0. B) Hexavanadate clusters show a wide temperature adaptability around 30~80 °C. C) 0.05~0.3 M H2O2 concentration is required to reach maximal peroxidase activity.

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Figure S3 Aggregation of 1 and 2 induced by TMB in the catalytic process. Left photograph: from left to right, 416 μM TMB solution with 9.5 mM H2O2, TMB and H2O2 incubated with 5 μg 1 and 2 for 3 min, and with 3 for 4 h, respectively. Right photograph: precipitates of 1 at the bottom of tube after 3000 rpm centrifugation (left), and clear solution of 3 (right).

Figure S4 IR spectra of TMB (gray line), Na22 (red line), and precipitates collected from the mixture solution of compound 2 and TMB.

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Figure S5 A) Time-resolved scattering intensity curve showing the precipitation process of 6 μM 1 (black squares) in the presence of 416 μM TMB in the first 200 s. B) Time-resolved scattered intensity curve showing the precipitation process of 6 μM 1 (black squares) and 4 μM 3 (red circles) in the presence of 416 μM TMB.

Figure S6 A) CONTIN analysis of the DLS data showing that the assemblies having a radius Rh of 94 nm. B) An average Rg value of 96 nm is obtained from the SLS study with a Zimm plot. A ratio of Rg/Rh = 1 indicates the formation of hollow, spherical vesicles in solution.

Figure S7 1H NMR spectra of TMB (blue line) in DMSO-d6, TMB & assemblies of 3 (green line) in 2 v/v% DMSO-d6/D2O, and assemblies of 3 (red line) in 2 v/v% DMSO-d6/D2O.

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Figure S8 Steady-state kinetic assay and catalytic mechanism of hexavanadate clusters. A–F) The velocity (v) of the reaction was measured by using 5 μg hexavanadate clusters in 1.0 mL PBS buffer (pH 6.5) at 35 °C. Error bars shown represent the standard error derived from three repeated measurements. The concentration of H2O2 was 98 mM and the TMB concentration was varied (A–C). The concentration of TMB was 416 μM and the H2O2 concentration was varied (D–F). G–H) Double-reciprocal plots of activity of hexavanadate clusters at a fixed concentration of one substrate versus varying concentration of the second substrate for H2O2 and TMB. The y-axis values are the observed absorbance values.

Table S2 Comparison of the three hexavanadates as HRP-mimic peroxidase. Km is the Michaelis constant. Vmax is the maximal reaction rate. kcat is the maximum number of substrate molecules converted to the product per enzyme molecule per second. The constant kcat/Km is a measure of how efficiently an enzyme converts a substrate into the product.

Catalyst Substrate Km (mM) Vmax (M/s) kcat (s-1) kcat/Km (s-1 mM-1)

1 TMB 0.032 ± 0.007 2.15 ± 0.28 × 10-7 0.018 0.56

1 H2O2 63.35 ± 10.43 8.58 ± 0.89 × 10-7 0.071 0.0011

2 TMB 0.026 ± 0.005 1.91 ± 0.22 × 10-7 0.018 0.69

2 H2O2 40.38 ± 6.64 6.79 ± 0.62 × 10-7 0.063 0.0016

3 TMB 0.021 ± 0.003 2.09 ± 0.16× 10-7 0.026 1.23

3 H2O2 43.61 ± 6.15 7.22 ± 0.59 × 10-7 0.090 0.0021

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Table S3 Comparison of the kinetic parameters of several nanostructures and HRP (TMB as a substrate). Km is the Michaelis constant, Vmax is the maximal reaction rate.

Catalyst Km (mM) Vmax (M/s) Reference

Fe3O4 MNPs 0.098 3.44 × 10-8 3

GO-COOH 0.0237 3.45 × 10-8 4

Pt48Pd52-Fe3O4 dumbbells 0.079 9.36 × 10-8 5

PtNTs 0.0186 1.179 × 10-7 6

PW12 0.11 4.31× 10-7 7

HRP 0.062 3.61× 10-8 5

1 0.032 2.15 × 10-7 This work

2 0.026 1.91 × 10-7 This work

3 0.021 2.09 × 10-7 This work

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