Supporting Information

Revitalizing Frens Method to Synthesize Uniform, Quasi-Spherical Gold

Nanoparticles with Deliberately Regulated Sizes from 2 to 330 nm

Haibing Xia,*,† Yujiao Xiahou, † Peina Zhang,† Wenchao Ding,† and Dayang Wang*,‡
†
State Key Laboratory of Crystal Materials, Shandong University, Jinan, 250100, P. R China;
‡
Department of Civil, Environmental and Chemical Engineering, School of Engineering, RMIT University,

Melbourne, VIC 3001, Australia.

Email: hbxia@sdu.edu.cn (H. X.) and dayang.wang@rmit.edu.au (D. W.)

Experimental Details

Materials. Hydrogen tetrachloroaurate(III) hydrate (HAuCl4, 99.9%, metals basis, Au >

49%, min), ascorbic acid (AA), sodium acetone dicarboxylate (SAD), reduced glutathione
(GSH), Tris (hydroxymethyl) aminomethane (Tris) and trisodium citrate dihydrate
(Na3C6H5O7, 99%) were purchased from Sinopharmacy Company (Shanghai, China). Silver
nitrate (AgNO3) was purchased from Alfa Aesar (99.95%, Tianjin, China). The chemicals
were used as received. The water in all experiments was prepared in a three stage Millipore
(A10) purification system and had a resistivity of > 18 MΩ cm−1. Prior to Au NP synthesis,
the aqueous solution of HAuCl4 (1 wt%) was prepared and stored in the fridge (4 °C). The
glassware and stir bars were thoroughly cleaned with aqua regia solutions consisted of HCl
(37%) and HNO3 (65%) with a volume ratio of 3:1, followed by thorough rinsing with H2O.
Caution: Aqua Regia solutions are dangerous and must be used with extreme care and never
stored the solutions in closed containers.
Synthesis of Au NPs via Frens Method. A given volume of the aqueous solutions of
citrate (1 wt%) was added into the boiling aqueous solutions of HAuCl4, followed by further
reflux of the reaction solution for 30 min to yield Au NPs. The total volume of the reaction
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solution was set as 50 mL and the concentration of HAuCl4 in the reaction solution was set as
0.01 wt%. The volume of the aqueous citrate solutions added was varied to adjust the sizes of
the Au NPs obtained. The experimental details were summarized in Table S1 and Figure S1.

Table S1. Summary of the experimental details for synthesis of Au NPs by Frens method.a
Volume of citrate (1 wt%) (mL)b 0.19c 0.21 0.3 0.5 0.75 1.5
d
Final citrate concentration (wt %) 0.0038 0.0042 0.006 0.01 0.015 0.03
Rc/a 0.52 0.58 0.82 1.4 2.1 4.1
NP size (nm) 75-174 51-142 45-103 35-78 19-35 14-18
a
The concentration of HAuCl4 in the reaction solution was 0.01 wt%.
b
The volume of the aqueous solution of sodium citrate (1 wt%) added into the boiling aqueous solution of
HAuCl4 (0.01 wt%).
c
Nanowire-like Au NPs would be obtained when the volume of citrate was less than 0.19 ml.
d
The concentration of citrate in the boiling reaction solution.

Synthesis of Ultra-small Au NPs via GSH-assisted Pre-mixing Method. To

synthesize 2 nm uniform Au NPs, 0.5 mL of the aqueous solution of HAuCl4 (1 wt%) and

42.5 µL of the aqueous solution of AgNO3 (0.1 wt%) was mixed under stirring. Into the
resulting HAuCl4/AgNO3 mixture solution, 1.5 mL of the aqueous solution of citrate (1 wt%)
was added; the citrate-to-HAuCl4 molar ratio was 4:1. The total volume of the
HAuCl4/AgNO3/citrate mixture solution was brought to 2.2 mL by adding water. After
incubation at ambient condition for 6 min, 0.3 mL of the aqueous solution of GHS (0.05 M)
was added into the HAuCl4/AgNO3/citrate mixture solution. Note that the GSH addition may
cause a rapid color change of the mixture solution from light yellow to brown and then back
to faint yellow within 10 s. After further incubation of 5 min, the
HAuCl4/AgNO3/citrate/GSH mixture solution was quickly injected into the boiling water
under vigorous stirring, followed by a color change of the reaction solution from colorless to
faint yellow in 10 min. The reaction solution was further refluxed for 45 min under stirring to
warrant the complete formation of Au NPs.
The similar protocol was applied to produce 4 nm uniform Au NPs except that the
citrate-to-HAuCl4 molar ratio decreased to 3:1 and the incubation of HAuCl4/AgNO3/citrate
mixture solution was 8 min prior to the GSH addition. Note that in order to guarantee the
reproducible synthesis of 2 or 4 nm Au NPs, the water needed to be heated to the vigorous

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boiling state,S1 before the addition of the HAuCl4/AgNO3/citrate/GSH mixture solution. The
experimental details were summarized in Table S2.

Table S2. The experimental details of the synthesis of Au NPs with the sizes in the range of
2- 4 nm by GSH-assisted premixing method.a
NP size Volume of citrate Final citrate concentration Tp-m Volume of GSH (0.05 Rc/a
(nm) (1wt%) (mL)b (wt%)c (min) M) (mL)d
2 1.5 0.03 6 0.3 4:1
4 1.1 0.022 8 0.3 3:1
a
0.5 mL of the aqueous solution of HAuCl4 (1 wt%) and 42.5 µL of the aqueous solution of AgNO3 (0.1 wt%)
were used for preparation of the pre-mixture solution and the concentration of HAuCl4 in the boiling reaction
solution was 0.01 wt%.
b
The volume of the aqueous solution of sodium citrate (1 wt%) was used to mixed with the pre-formed aqueous
solution of HAuCl4/AgNO3 mixtures.
c
The concentration of citrate in the boiling reaction solution.
d
The volume of the aqueous solution of GSH (0.05 M) was used to mixed with the pre-formed aqueous solution
of HAuCl4/citrate/AgNO3 mixtures.

Synthesis of Monodisperse, Quasi-spherical Au NPs in the Size Range of 6 - 39 nm

at Size Interval of 3 nm via Pre-mixing Method. 0.5 mL of the aqueous solution of HAuCl4
(1 wt%) and 42.5 µL of the aqueous solution of AgNO3 (0.1 wt%) were mixed under stirring.
Into the resulting HAuCl4/AgNO3 solution, a given volume of the aqueous solution of citrate
(1 wt%) was added under stirring; the volume of the citrate solution was varied in order to
adjust the citrate-to-HAuCl4 molar ratio in the reaction media. Water was added into the
resulting HAuCl4/AgNO3/citrate mixture solution to bring the total solution volume to 2.5
mL. After incubation at the room temperature for a given time (pre-mixing time), the
resulting HAuCl4/AgNO3/citrate mixture solution was quickly injected into 47.5 mL of
boiling water under vigorous stirring. The color of the reaction solution turned from colorless
to grayish within 10 s and then gradually to dark grey, purplish red and pellucid wine red
within 3 min. The reaction solution was further refluxed for 30 min under stirring to warrant
formation of uniform quasi-spherical Au NPs. After the removal of the heat source, the
reaction solution was cooled to room temperature under gentle stirring. The experimental
details were summarized in Table S3.

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Table S3. Summary of the experimental details for synthesis of monodisperse,
quasi-spherical Au NPs with sizes from 6 nm to 39 nm by premixing method.a
NP size Volume of citrate Final citrate concentration Tp-m Rc/a
(nm) (1wt%) (mL)b (×103 wt%)c (min)
6±0.5 2.5 50 16 6.9
9±0.5 1.5 30 8 4.1
12±1 1.5 30 5 4.1
15±1 1.1 22 4.5 3.0
18±1 0.7 14 4 2.0
21±2 0.85 17 5 1.6
24±2 0.75 15 5.5 1.4
27±3 0.7 14 5 1.3
30±3 0.65 13 4.5 1.2
33±3 0.35 7 2.5 1.0
36±3 0.3 6 2.5 0.8
39±3 0.25 5 2 0.7
a
0.5 mL of the aqueous solution of HAuCl4 (1 wt%) and 42.5 µL of the aqueous solution of AgNO3 (0.1 wt%)
were used for preparation of the pre-mixture solution. While that was 0.015 wt% for synthesis of Au NPs with
sizes in the range of 21 – 30 nm, the concentration of HAuCl4 in the boiling reaction solution was 0.01 wt% for
synthesis of Au NPs with other sizes.
b
The volume of the aqueous solution of sodium citrate (1 wt%) was used to mixed with the pre-formed aqueous
solution of HAuCl4/AgNO3 mixtures.
c
The concentration of citrate in the boiling reaction solution.

In order to study of the effect of TB on the color change during citrate reduction of
HAuCl4 to Au NPs, 2.0 mL of TB buffer (0.1 M) was added into boiling water before
addition of the HAuCl4/citrate/AgNO3 mixture solution.

Synthesis of Monodisperse, Quasi-spherical, Au NPs in the Size Range of 40 - 95 nm

by Tris-assisted Pre-mixing Method. 0.5 mL of the aqueous solution of HAuCl4 (1 wt%)
and 42.5 µL of the aqueous solution of AgNO3 (0.1 wt%) were mixed under stirring. Into the
resulting HAuCl4/AgNO3 solution, a given volume of the aqueous solution of citrate (1 wt%)
was added under stirring; the volume of the citrate solution was varied in order to adjust the
citrate-to-HAuCl4 molar ratio in the reaction media. Water was added into the resulting
HAuCl4/AgNO3/citrate mixture solution to bring the total solution volume to 1.5 mL. After
incubation for a given time (pre-mixing time), 2.0 mL of Tris buffer (0.10 M) was added into
the resulting HAuCl4/AgNO3/citrate mixture solution. After 1.0 min incubation under
stirring, the resulting HAuCl4/AgNO3/citrate/Tris mixture solution was quickly injected into

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boiling water under vigorous stirring, leading to rapid color change of the reaction solution
turned from colorless to pink, powdery violet, and ruby red within 1 min. Afterwards, the
reaction solution further refluxed for 30 min under stirring to warrant the formation of Au
NPs. The experimental details were summarized in Table S4.

Table S4. Summary of the experimental details for synthesis of Au NPs with size range from
40 nm to 95 nm by TB-assisted premixing method.a,b The final concentration of TB and auric
acid is 0.048 wt% and 0.01 wt%, respectively.
NP size (nm) 40 ± 3 60 ± 3 82±5 95± 6
Volume of citrate (1wt%) (mL)c 0.5 0.4 0.35 0.3
Final citrate concentration (wt%)d 0.01 0.008 0.07 0.006
Rc/a 1.4 1.1 1.0 0.8
Tp-m / min 5 4 4 3
a
0.5 mL of the aqueous solution of HAuCl4 (1 wt%) and 42.5 µL of the aqueous solution of AgNO3 (0.1 wt%)
were used for preparation of the pre-mixture solution. The concentration of HAuCl4 in the boiling reaction
solution was 0.01 wt%.
b
2 mL of the Tris buffer solution (0.10 M) was used to mixed with the pre-formed aqueous solution of
HAuCl4/citrate/AgNO3mixtures. The concentration of Tris in the boiling reaction solution was 0.048 wt%.
c
The volume of the aqueous solution of sodium citrate (1 wt%) was used to mixed with the pre-formed aqueous
solution of HAuCl4/AgNO3 mixtures.
d
The concentration of citrate in the boiling reaction solution.

Synthesis of Uniform Au NPs in the Size Range of 105 - 330 nm via TB-assisted
Turkevich Method. 2.0 mL of TB buffer solution (0.10 M) was added into boiling water (47
ml). After boiling for 5.0 min, 300 µL of the aqueous solution of citrate (1.0 wt%) was
injected into the boiling TB solution, followed by addition of 0.50 mL of HAuCl4 aqueous
solution (1.0 wt%) after 2.0 min. The whole reaction solution was further refluxed for 1.0 h
under stirring to warrant the formation of Au NPs. The experimental details were summarized
in Table S5.

Table S5. Summary of the experimental details for synthesis of uniform Au NPs with size
range from 105 nm to 330 nm by TB-assisted Turkevich method.a,b The TB concentration is
0.048 wt% and its concentration is 0.06 wt% for Au NPs with sizes of 330 nm. The HAuCl4
concentration is always 0.01 wt%.
NP size (nm) 105±12 125±15 166±17 240±28 330±32
Volume of citrate (1 wt%) (mL)c 300 150 100 50 50
Final citrate concentration ( ×103 wt%)d 6 3 2 1 1
Rc/a 0.8 0.4 0.27 0.14 0.14
a
The concentration of HAuCl4 in the boiling reaction solution was 0.01 wt%.

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b
The concentration of Tris in the boiling reaction solution was 0.06 wt% for synthesis of 330 nm Au NPs, while
that was kept as 0.048 wt% for synthesis of the Au NPs with other sizes.
c
The volume of the aqueous solution of sodium citrate (1 wt%) was used to mixed with the pre-formed aqueous
solution of HAuCl4/AgNO3 mixtures.
d
The concentration of citrate in the boiling reaction solution.

Characterization. UV-Vis absorption spectroscopy was recorded on a Varian Cary 50

spectrophotometer by using quartz cuvettes with 10 mm path length. The quantitative
analysis of HAuCl4 consumption was carried out according to the method reported in
literature.S2,S3 Briefly, the aliquots of the aqueous solutions of HAuCl4/AgNO3/citrate
mixtures were taken out and added into the aqueous solutions of NaCl (0.90 M)/HCl (0.10
M) mixtures in ice water bath to quench the reaction. AuCl4- ions show a sharp absorption
peak at 314 nm due to the charge transfer to solvent band at pH 1.5. According to Beer’s law,
the concentrations of AuCl4- ions were determined by the intensity of the absorption peak at
314 nm.
TEM and HRTEM images were obtained with a JEOL JEM-2100F transmission electron
microscope operating at an acceleration voltage of 200 kV. DLS analysis was performed at
room temperature on a Malvern Zetasizer Nano ZS using a He-Ne laser with a wavelength of
632 nm. The aliquots of the aqueous solution of HAuCl4/AgNO3/citrate mixtures at different
time during incubation at room temperature were taken out and added into the aqueous
solution of sodium mercaptobenzoate (50 mM) to stabilize the reaction intermediates for DLS
and Zeta-potential measurements.

References:
S1. Ding, W.; Zhang, P.; Li, Y.; Xia, H.; Wang, D.; Tao, X. ChemPhysChem, 2015, 16, 447 –
454.
S2. Peck, J. A.; Tait, C.D.; Swanson, B. I.; Brown Jr., G. E. Geochimica et Cosmochimica
Acta 1991, 55, 671-676.
S3. Xia, H.; Bai, S.; Hartmann, J.; Wang, D. Langmuir 2010, 19, 3585-3589.

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Figure S1. TEM images of the Au NPs prepared at Rc/a of 4.1 (a), 2.1 (b), 1.4 (c), 0.82 (d),
0.58 (e), and 0.52 (f), by Frens method. The experimental details are listed in Table S1.

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Figure S2. Temporal evolution of size variation of intermediate products in the aqueous
solutions of HAuCl4/citrate/AgNO3 mixtures with Rc/a of 6.9 (a) and 0.8 (b) during incubation
at room temperature. When Rc/a is 6.9, the concentrations of HAuCl4 and Ag+ are 0.167 wt%
and 1.42 × 10-3 wt%, respectively. When Rc/a is 0.8, the concentrations of HAuCl4 and Ag+ are
0.2 wt% and 1.7 × 10-3 wt%, respectively.

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Figure S3. Temporal evolution of the concentration of auric ion in the aqueous solutions of
HAuCl4/citrate/AgNO3 mixtures with Rc/a of 6.9 (●), 4.1 (▲) and 0.8 (▼) during incubation
at room temperature. When Rc/a is 4.1 (▲) and 0.8 (▼), the concentrations of HAuCl4 and
Ag+ are 0.2 wt% and 1.7 × 10-3 wt%, respectively. When Rc/a is 6.9 (●),the concentrations of
HAuCl4 and Ag+ are 0.167 wt% and 1.42 × 10-3 wt%, respectively.

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Figure S4. Digital photos of the temporal color evolution of the reaction solution after adding
the aqueous solution of HAuCl4 into the boiling aqueous solution of citrate for synthesis of
Au NPs by Turkevich method. R is 6.9. The concentrations of citrate and HAuCl4 are 0.05
wt% and 0.01 wt%, respectively in the boiling reaction solution.

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Figure S5. TEM image of the intermediate products extracted from the aqueous solutions of
HAuCl4/citrate/AgNO3 mixtures with Rc/a of 6.9 during incubation at room temperature. The
intermediate products are aggregates of tiny Au NPs with sizes of 1.5 – 2.5 nm.

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Figure S6. Plot of the hydrodynamic size (a) and zeta-potential (b) of the particles obtained
in the reaction solution after adding the aqueous solution of HAuCl4/citrate/AgNO3/GSH
mixtures into boiling water. Rc/a is 4.1 and Tp-m is 6 min. The experimental details are
summarized in Table 1 and Table S2.

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Figure S7. Temporal evolution of the zeta-potential of the particles obtained in the reaction
media during incubation of the aqueous solution of HAuCl4/citrate/AgNO3 mixtures with Rc/a
of 0.8 at room temperature The experimental details are summarized in Table 1 and Table S3.

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Figure S8. UV-Vis spectra of the Au NPs with sizes increasing in increments of 3 nm from 6
nm to 39 nm prepared at Rc/a decreasing from 6.9 to 0.69 by pre-mixing method. The
experimental details are listed in Table 1 and Table S3. The absorption spectra are
normalized at 400 nm for easy comparison. The TEM images of the Au NPs are shown in
Figure 4.

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Figure S9. UV-Vis spectra of the Au NPs with sizes of 40 ± 3 (a), 60 ± 3 (b), 82±5 (c), and
95 ± 6 nm (d), obtained at Rc/a of 1.4, 1.1, 1.0, and 0.8 by TB–assisted pre-mixing Turkevich
method. The absorption spectra are normalized at 400 nm for easy comparison. The
experimental details are listed in Table 1 and S4. The TEM images of the Au NPs are shown
in Figure 5.

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Figure S10. UV-Vis spectra of the Au NPs with sizes of 105 ± 12 nm (a), 125 ± 15 nm (b),
166 ± 17 nm (c), 240 ± 28 nm (d) and 330 nm ± 32 nm (e), respectively obtained at Rc/a of
0.8 (a), 0.4 (b), 0.27 (c), 0.14 (d) and 0.14 by TB–assisted Turkevich method. The
experimental details are listed in Table 1 and S5. The absorption spectra are normalized at
400 nm for easy comparison. The TEM images of the Au NPs are shown in Figure 6.

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