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Supplementary Information

In-Situ Formation and Immobilization of Gold Nanoparticles on Polydimethylsiloxane (PDMS) with

Catalase-Mimetic Activity

Hui Zhang and Kun-Lin Yang*

Department of Chemical and Biomolecular Engineering, National University of Singapore, 4

Engineering Drive 4, 117576, Singapore

*For correspondence

E-mail: cheyk@nus.edu.sg. Phone: (+65) 65166614. ORCID: 0000-0002-7958-9334.

Electronic Supplementary Material (ESI) for ChemComm.This journal is © The Royal Society of Chemistry 2020

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Contents

1. EXPERIMENTAL 3

2. FIGURES 4

3. TABLE 14

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1. Experimental

1.1. Materials

Benzoyl peroxide, potassium phosphate monobasic, potassium phosphate dibasic were purchased from

Sigma-Aldrich (Singapore). Hydrogen tetrachloroaurate(III) hydrate was obtained from Alfa Aesar

(U.S.A.). Sylgard 184 silicone elastomer base and curing agent for making polydimethylsiloxane

(PDMS) were procured from Dow Corning (U.S.A.). Stainless steel needles (0.90 × 38 mm) were

purchased from Terumo (Japan). All chemicals were used as received without further purification.

Deionized water was obtained from a Milli-Q system (U.S.A.).

1.2. Preparation of PDMS

PDMS was prepared by mixing 20.0 g of PDMS monomer with 2.0 g of curing agent. Next, the viscous

mixture was poured on a Petri dish and degassed for 1 h to remove bubbles. Then, the prepolymer

mixture was put in an oven at 65 °C for 5 h to solidify it. Finally, PDMS was removed from the Petri

dish and cut into small cubes with a dimension of ~ 1.2×0.8×0.5 cm prior to use.

1.3. Synthesis of AuNPs and immobilization

An aqueous solution containing 1 wt% of HAuCl4 was prepared and the pH was adjusted to 2~2.5. A

stainless steel needle was inserted into the solution to purge the solution with nitrogen gas continuously,

at a flow rate of 5~10 cm3/s. After the solution color changed from pale yellow to red, the needle and

nitrogen gas was removed, and a piece of PDMS cube was immersed into the solution. After the surface

of PDMS became red, the PDMS cube was removed from the solution and washed with DI water.

1.4. Measurement of Catalase-Mimetic Activity

Fifty microliter of 20 wt% hydrogen peroxide solution was added to 1 mL of 0.025 M phosphate-

buffered saline (PBS) at pH 7.4. Then, one piece of PDMS cube with the immobilized AuNPs was

placed into the solution and incubated in an oven at 37 °C for 30 mins.

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2. FIGURES

Fig. S1 Size distribution of AuNPs in the solution.

For the free AuNPs before the immobilization, we analyzed the particle size distribution by using

dynamic light scattering (DLS) as shown in Fig. S1, the average diameter is 40.6 nm, which is slightly

larger than that in the SEM image. This is probably because hydration diameter, not the physical

diameter, was measured in DLS. The AuNPs is positively charged with a zeta potential of +28.0 mV.

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Fig. S2. Detection of Fe2+ in the used HAuCl4 solution. (A) Original 1 mM KMnO4 solution. After it

mixed with (B) a new HAuCl4 solution, and (C) a used HAuCl4 solution. The yellow color indicates the

presence of Fe2+ in the used HAuCl4 solution.

We used KMnO4 reagent to check if Fe2+ was present. Fig. S2C shows that after the addition of the used

HAuCl4 solution to KMnO4, the red color of KMnO4 faded after 1 h. This result suggests that the

solution contained Fe2+ (which reduced KMnO4).

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Fig. S3. Detection of Fe3+ in the used HAuCl4 solution. (A) Original 1 mM phenol solution. After it

mixed with (B) a new HAuCl4 solution, and (C) a used HAuCl4 solution. No color change suggests that

no Fe3+ was present.

We used phenol reagent to check if Fe3+ was present. After the addition of the used HAuCl4 solution to

phenol, the solution did not change color (Fig. S3C). It shows that there was no Fe3+ in the solution

(Fe3+ is able to form red-color complex with phenol).

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ElementWeight

(%)Atomic (%)

C 29.30 44.56

O 24.35 27.81

Si 41.83 27.21

Au 4.53 0.42

Totals 100 100

Fig. S4 EDX of PDMS with immobilized AuNPs.

There is a possibility that iron species, rather than gold nanoparticles, were responsible for the

decomposition of H2O2. However, from the EDX data, we can see that there was a strong peak for Au

(4.53%), but there was no Fe. Thus, we ruled out the possibility that iron species were responsible for

the catalase-mimetic property.

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Fig. S5 Post treatments of PDMS and their effects on the immobilization of AuNPs. (A) control, (B)

HCl, (C) NaOH, (D) NaClO, (E) NaBH4, (F) curing agent, and (G) ethanol containing benzoyl peroxide.

Various post treatments of PDMS were done, including acid (HCl), base (NaOH), oxidizing (NaClO,

benzoyl peroxide) and reducing (NaBH4) agents. Fig. S5G shows that AuNPs could not be immobilized

on the surface of PDMS, indicating that the treatment of benzoyl peroxide inhibited the immobilization

of AuNPs. Similarly, after PDMS was immersed in a NaClO solution, the immobilization was also

supressed (Fig. S5D). On the other hand, when reducing agents such as curing agent (Fig. S5F), more

AuNPs can be immobilized. However, the effect of reducing agents such as NaBH4 (Fig. S5E) was not

as good as expected. This is probably because PDMS is hydrophobic in nature, it is difficult to treat the

surface with an aqueous solution of NaBH4. The surface of PDMS de-wetted quickly when it was taken

out from the solution.

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Fig. S6 Immobilization of AuNPs on PDMS prepared by using different monomer/curing agent ratios.

(A) monomer/curing agent =10:1 and the PDMS was exposed to air for one month to oxidize the

remaining curing agent, (B) monomer/curing agent =10:1, (C) monomer/curing agent =10:1, and both

top and bottom surface of PDMS were rough, (D) monomer/curing agent =10:1.4, (E) monomer/curing

agent =10:0.7. The controlled experiments suggest that curing agent plays a critical role in the

immobilization of AuNPs.

To further confirm the role of curing agent, we prepared PDMS with different monomer/curing agent

ratios. Compared with the normal PDMS (monomer/curing agent = 10:1), PDMS with more curing

agent (Fig. S6D), the bulk of PDMS became red. For the PDMS with less curing agent (Fig. S6E), the

red color was very light. This result indicates the excessive curing agent is responsible for the

immobilization of AuNPs.

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Fig. S7 The SEM of the AuNPs immobilized on PDMS for (A) 1 h (diameter = 36.3 ± 9.4 nm) and (B)

3 h (diameter = 30.3 ± 3.1 nm).

More AuNPs appeared because of the curing agent. At the same time, the AuNPs became smaller and

more uniform, probably going through a ripening process.

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Fig. S8 SEM of pristine PDMS at the (A) bottom surface and (B) top surface.

The roughness is different for the top and bottom surfaces. There is a possibility that the AuNPs can

only be immobilized on the bottom surface because of the roughness. To analyse the possibility, we

prepared a piece of PDMS with both the top and bottom were rough (Fig. S6C). However, the AuNPs

still only immobilized on the bottom surface. Thus, we ruled out the possibility.

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Fig. S9. (A) UV spectra of H2O2 solutions with different concentration. (B) A calibration curve for H2O2

concentration based on the absorbance values at 240 nm. The concentration of H2O2 were recorded by

monitoring the absorbance at 240 nm. (C) Decomposition rate of H2O2 at different H2O2 concentrations.

(D) Lineweaver–Burk plot.

The Michaelis-Menten equation (1) is known as the general equation for steady-state enzyme kinetics,

(1)𝑣=

𝑉𝑚𝑎𝑥[𝑆]

𝐾𝑚+ [𝑆]

(2)

1𝑣=

𝐾𝑚𝑉𝑚𝑎𝑥

×1[𝑆]

+1

𝑉𝑚𝑎𝑥

where Vmax is the maximum initial velocity, Km is the Michaelis-Menten constant, and [S] is the

substrate concentration. We also fitted our experimental data with Michaelis-Menten reaction kinetics.

Vmax and Km were estimated to be 4.9×10-5 M/s and 1.2 M from the Lineweaver-Burk plot (R2=0.98)

in Fig. S9D.

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3. TABLE

Table S1. Comparison of Vmax and Km for mimetic and natural catalase.

No. Catalyst Km (mM) Vmax (M/s) Ref.

1 Azobenzene-modified silica encapsulated

AuNPs

UV:117

Vis:252

11.77×10-8

4.98×10-8

1

2 AuNPs on doxorubicin encapsulated

poly(lactic-co-glycolic acid) vehicle

-- -- 2

3 AgPt NPs 62.98 6.1×10-6 3

4 Non-Heme Oxoiron(IV) Complex 1390 -- 4

5 RuO2 NPs 400 -- 5

6 Catalase from bovine liver 54.30 1.62×10-5 6

7 Catalase from P. mirabilis 537 -- 7

8 AuNPs 1200 4.9×10-5 This work

We compared Vmax and Km for different catalase enzymes and catalase-mimetic materials including

gold nanoparticles in Table S1. The Km value in this work is larger than the stablised AuNPs in solution

or natural catalase, indicating a lower affinity to the substrate. However, the Vmax value is comparable

to others.

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