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Protocol for the synthesis of silver nanoparticles using sodium citrate and
sodium borohydride as reducing agents

Technical Report · March 2018


DOI: 10.13140/RG.2.2.12852.76168

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White papers in nanomaterials

PROTOCOL FOR THE SYNTHESIS OF SILVER


NANOPARTICLES USING SODIUM CITRATE AND
SODIUM BOROHYDRIDE AS REDUCING AGENTS

A. Pinedo-Flores, B. Alcázar and J. C. F. Rodríguez-Reyes

PROTOCOL FOR THE SYNTHESIS OF SILVER NANOPARTIC LES USING SODIUM


CITRATE AND SODIUM BOROHYDRIDE AS REDUCING AGENTS

Angela Pinedo-Flores*, Bryan Alcázar° and Juan Carlos F. Rodriguez-Reyes¥


Departments of Chemical Engineering* and Industrial Engineering°
Universidad de Ingeniería y Tecnología – UTEC, Jr. Medrano Silva 165, Lima 04, Peru
¥Corresponding author: jcrodriguez@utec.edu.pe

Table of Contents
Page
Disclaimer 1
1. Introduction: Principles of synthesis and application of nanoparticles 1
2. Materials, Reagents and Equipment needed 3
3. Protocol for synthesis and results of characterization 3
4. Characterization of nanoparticles. 4
5. Acknowledgements 5
6. References 6

DISCLAIMER

The present protocol is intended for use in a safe environment, such as a laboratory, and under
the supervision of professionals. Be aware that, in spite of the simplicity of the procedures,
there is an inherent risk in the handling of chemicals and in the disposal of residues.

1. INTRODUCTION: SYNTHESIS AND CHARACTERIZATION OF NANOPARTICLES

The matter in the nanoscale present properties which are completely different to those
observed in the macroscale (above 1 m). For example, while gold has been known to be an
inert, noble metal, in nanoparticle form it is an excellent catalyst, facilitating a broad range of
oxidation reactions.[1] Another example is that of silicon, which is a semiconductor in the
macroscale but in the nanoscale it behaves as a conductor (quantum dot).The known
antibacterial properties of silver (whose ions are able to interfere with key biological processes
in a bacteria) are enhanced when silver is used as nanoparticles, not only due to the slow
release of silver ions, but also due to the nanoparticle-mediated formation of reactive oxygen
species (ROS), which contribute to the destruction of bacteria. The more controlled, targeted
manner in which silver nanoparticles operate reduces the impact of silver in health and
aquatic life.[2] (Mavani, K .; Shah, M., 2013).
White papers in nanomaterials

PROTOCOL FOR THE SYNTHESIS OF SILVER


NANOPARTICLES USING SODIUM CITRATE AND
SODIUM BOROHYDRIDE AS REDUCING AGENTS

A. Pinedo-Flores, B. Alcázar and J. C. F. Rodríguez-Reyes

The synthesis of nanoparticles by chemical methods is popular as they require little


instrumental and are relatively inexpensive. The most common way to synthesize silver
nanoparticles is via chemical reduction (Khan, Z. et al., 2011), where the reduction of silver
ions take place in controlled conditions,[3,4] which can limit the nucleation of reduced
(neutral) silver atoms:
+ 0
𝐴𝑔(𝑎𝑐) + 𝑒 − → 𝐴𝑔(𝑠) (1)
Controlled conditions include i) the use of low concentrations of silver ions (usually in the
milimolar, mM, range) so that growing nuclei have a large mean free path and, therefore, less
probability to continue nucleating, ii) a stirring sufficiently fast so that the dilution of an
incoming silver solution is instantaneous, but sufficiently slow that impedes the clashing and
growing of silver nuclei, and iii) the use of a capping agent able to confer a surface charge to
the growing nanoparticle, favoring repulsion between nanoparticles. For example, as citrate
or borohydride moieties are adsorbed on the surface of the nanoparticles, they become
“negatively charged” and can repel other nanoparticles. Borohydride and citrate can also work
as reducing agents, completing the redox reaction shown in (1) as:
+ 0 + +
4𝐴𝑔(𝑎𝑐) + 𝐶6 𝐻5 𝑂7 𝑁𝑎3 (𝑎𝑐) + 2𝐻2 𝑂(𝑙) → 4 𝐴𝑔(𝑠) + 𝐶6 𝐻5 𝑂7 𝐻3 (𝑎𝑐) + 3𝑁𝑎(𝑎𝑐) + 𝐻(𝑎𝑐) +
𝑂2 (𝑔) (2)
+ − 0
𝐴𝑔(𝑎𝑐) + 𝐵𝐻4(𝑎𝑐) → 𝐴𝑔 (𝑠) + 0.5 𝐻2(𝑔) + 0.5 𝐵2 𝐻6(𝑔) (3)
Other variables include temperature, the presence of additional species, the volumes involved
in the synthesis, etc. Nanoparticles are relatively simple to synthesize. In fact, the challenges
associated with nanoparticle production are not related to the synthetic aspect, but instead
in the reproducibility of results and in the development of methods to stabilize them.
Since the properties of nanoparticles are size- and shape- dependent, characterization of
nanoparticles is a crucial aspect to ensure that the appropriate size has been reached, which
depends on the application. Tools for characterization include electron microscopies, such as
scanning electron microscopy (SEM) and transmission electron microscopy (TEM);
spectroscopies, such as Raman and ultraviolet-visible absorption spectroscopy (UV-vis) and
also diffraction light scattering (DLS). Often, the most affordable instrument is the UV-vis
spectrometer. This technique takes advantage of the fact that the electrons on the surface of
a metallic nanoparticle can be in resonance with visible light at specific wavelenghts. This
effect, called surface plasmon resonance (SPR), originates the absorption of light, which is
seen in a UV-vis spectrum.[5] The wavelenght of maximum absorption ( max) is dependent
on the shape and size of the nanoparticle. For spherical silver nanoparticles, for example,
values of max close to 400 nm indicate a size of 5-10 nm, and higher values of max are
observed as nanoparticle grow larger. On the other hand, the intensity of the absorption signal
provides a good indication of the concentration of nanoparticles, although a careful
calibration is often required and only a semi-quantitative value can be reached.
This protocol describes the synthesis of silver nanoparticles (Ag NPs) using borohydride and
citrate as reducers (Equations 2 and 3 above). The results will be characterized by UV-vis
spectrometry and scanning electron microscopy (SEM).
White papers in nanomaterials

PROTOCOL FOR THE SYNTHESIS OF SILVER


NANOPARTICLES USING SODIUM CITRATE AND
SODIUM BOROHYDRIDE AS REDUCING AGENTS

A. Pinedo-Flores, B. Alcázar and J. C. F. Rodríguez-Reyes

2. MATERIALS, REAGENTS AND EQUIPMENT NEEDED

2.1. Reagents
 Deionized water
 Silver nitrate, AgNO3(99.8% - J.A. Elmer)
 Sodium citrate (99.9% -Movilab)
 Sodium borohydride, NaBH4 (>98%, Sigma-Aldrich)
 Sodium hydroxide (NaOH, 0.1M, Merck)
 Nitric acid (HNO3, 0.1%, J.T. Baker)

2.2. Materials and Equipment


 Volumetric flasks 50, 100, 250 and 500 mL.
 Burets, 50 mL (preferably made of dark glass to impede the light-induced
decomposition of AgNO3; alternatively, it can be covered with aluminum foil)
 Beakers, 50, 100,500 and 1000 mL
 Thermometer
 Erlenmeyer flasks, 250 mL
 Micropipet, 100-1000 μL
 Pipet, 10 mL
 Stirring/heating plate
 Digital scale
 Test tubes

3. PROTOCOL FOR THE SYNTHESIS OF SILVER NANOPARTICLES

3.1. Synthesis of nanoparticles using sodium borohydride as reducer

i. Place 20 mL of a solution AgNO3 1 mM in a buret.


ii. Place 20 mL of NaBH4 2 mM in a 250 mL Erlenmayer flask with a magnetic pill.
iii. Place the Erlenmayer flask on a heating/stirring plate, heating to 70° C while stirring at a
moderate rate (~100 rpm).
iv. Add dropwise the AgNO3 solution until a light yellow color is observed and is stable. It
corresponds to ~10-12 mL of the solution.
v. Storing – option a. Cool down the solution to room temperature. Store in sealed
containers, preferably opaque and with as little air as possible. Theoretically, the mass
of silver in suspension is approximately 1mg.
vi. Storing – option b. Add 1g of activated carbon or any other support to the nanoparticle
suspension (step v.) under stirring. Samples can be taken to microscopy
characterization.
White papers in nanomaterials

PROTOCOL FOR THE SYNTHESIS OF SILVER


NANOPARTICLES USING SODIUM CITRATE AND
SODIUM BOROHYDRIDE AS REDUCING AGENTS

A. Pinedo-Flores, B. Alcázar and J. C. F. Rodríguez-Reyes

3.2. Synthesis of Ag nanoparticles using sodium citrate as reducer

i. Place 20 mL of a solution AgNO3 2 mM in a buret.


ii. Place 100 mL of sodium citrate 7 mM in a 250 mL Erlenmayer flask with a magnetic pill.
This solution should be at pH 9; pH is adjusted by adding dropwise the solutions of
NaOH and/or HNO3.
iii. Place the Erlenmayer on a heating/stirring plate, heating to 100° C (boiling), stirring
vigorously (~400 rpm).
iv. Add dropwise the AgNO3 solution until a light yellow color is observed and is stable. It
corresponds to ~8-12 mL of the solution.
v. Storing – option a. Cool down the solution to room temperature. Store in sealed
containers, preferably opaque and with as little air as possible. Theoretically, the mass
of silver in suspension is approximately 1mg.
vi. Storing – option b. Add 1g
of activated carbon or any
other support to the
nanoparticle suspension
(step v.) under stirring.
Samples can be taken to
microscopy
characterization.

4. CHARACTERIZATION

The protocol described above


has been determined after
changing several parameters
such as the concentration of the
reducer and the precursor, the
pH and the temperature. The
first, simplest way to identify
the success of the synthesis is
by visual inspection. For
example, Fig. 1 shows the
changes in color during the
synthesis of Ag NPs using citrate
as reducer. As increasing
Fig. 1. Progress of the Ag NPs synthesis as AgNO3 is
volumes of silver nitrate are
added to 100 mL of 7 mM citrate solution at pH 9.
added the solution turns
White papers in nanomaterials

PROTOCOL FOR THE SYNTHESIS OF SILVER


NANOPARTICLES USING SODIUM CITRATE AND
SODIUM BOROHYDRIDE AS REDUCING AGENTS

A. Pinedo-Flores, B. Alcázar and J. C. F. Rodríguez-Reyes

yellow, indicative of the


formation of nanoparticles of
sizes below 10 nm. Volumes
added above 10 mL result in a
change in color to brown and
finally to grey. These changes
are, however, only qualitative
and more precise strategies
need to be considered. Our
group has used UV-VIS
spectroscopy as a quick gauge
to choose the right values for a
determined parameter.
Spectra for a solution
containing nanoparticles were Fig. 2. UV-vis absorption spectrometry of Ag NPs
collected after synthesis and suspensions using AgNO3 and citrate at different pH. All
we checked for the intensity of other parameters were kept constant throughout
the absorption peak (related to experiments.
the concentration of
nanoparticles), the full width at
half maximum (related to the
dispersion of nanoparticle
sizes, where the narrower the
peak the closer to a unique
nanoparticle size) and the peak
position (related to the
nanoparticle size). We present
some of the results we
obtained.
4.1. Impact of the pH in the
synthesis of Ag NPs using
citrate. Figure 2 shows the UV-
vis spectra of the suspension of Fig. 3. UV-vis absorption spectrometry of Ag NPs
Ag NPs obtained after suspensions using citrate at pH 9 and borohydride as
performing the synthesis at pH reducers. The same volumen of AgNO3 was employed in
7, 9 and 11. At pH 7 there is a both experiments.
small amount of Ag NPs
obtained, as determined from the small intensity of the spectrum. At pH 9 there is a higher
production of Ag NPs, characterized by a broad peak at around 414 cm-1. However, increasing
the pH to 11 increases the size distribution of nanoparticles, as evidenced by the broadening
of the signal. Since there is a shift of the signal to 424 cm -1, it can be assumed that at pH 11
White papers in nanomaterials

PROTOCOL FOR THE SYNTHESIS OF SILVER


NANOPARTICLES USING SODIUM CITRATE AND
SODIUM BOROHYDRIDE AS REDUCING AGENTS

A. Pinedo-Flores, B. Alcázar and J. C. F. Rodríguez-Reyes

there are Ag NPs of larger sizes than at pH 9. Thus, it is clear that the optimal pH is 9 as it
provides a significant amount of Ag NPs without a broad distribution of sizes.
4.2. Citrate vs. borohydride as reducer. Figure 3 shows the difference in the UV-vis spectra
collected using borohydride and citrate. From the difference in intensities, it is clear that
borohydride produces more nanoparticles than citrate. Moreover, from the narrowness of the
borohydride spectra it is deduced that the nanoparticle size is also more uniform with
borohydride.

Fig. 4. Scanning Electron Microscopy (SEM) images: A) Activated carbon exposed to a citrate
solution (without Ag NPs); B) and C) Ag NPs supported on activated carbon with citrate as a
reducer; D) Ag NPs supported on activated carbon with borohydride as a reducer.
White papers in nanomaterials

PROTOCOL FOR THE SYNTHESIS OF SILVER


NANOPARTICLES USING SODIUM CITRATE AND
SODIUM BOROHYDRIDE AS REDUCING AGENTS

A. Pinedo-Flores, B. Alcázar and J. C. F. Rodríguez-Reyes

4.3. Scanning Electron Microscopy (SEM) images. Figure 4 shows a micrograph of activated
carbon exposed to citrate solution with and without Ag NPs. The size of nanoparticles
detected is around 80 nm. Micrographs of activated carbon impregnated with Ag NPs from
borohydride solutions are not shown due to the fact that borohydride modified the
morphology of the activated carbon (it did not show a porous structure such as in Fig. 4), likely
due to the reaction of borohydride and adsorption of by-products on the surface of activated
carbon.

5. ACKNOWLEDGEMENTS

This work has been supported by the Universidad de Ingeniería y Tecnología – UTEC and
Cleveland Clinic through a research grant for joint research projects. Ms. Karinna Visurraga
(UTEC) is acknowledged for administrative and technical support. Dr. Vijay Krishna (Cleveland
Clinic), Mr. Luis Palomino and Mr. Miguel Gakiya (UTEC) are thanked for useful discussions.

6. REFERENCES

[1] T. A. Baker, X. Liu, and C. M. Friend, “The mystery of gold’s chemical activity: local bonding,
morphology and reactivity of atomic oxygen.,” Phys. Chem. Chem. Phys., vol. 13, no. 1, pp.
34–46, 2011.
[2] K. Mavani, M. Shah, “Synthesis of silver nanoparticles by using sodium borohydride as a
reducing agent”, Int. J. Eng. Res. Tech., vol. 2, no. 3, pp. 1-5, 2013.
[3] S. Solomon, M. Bahadori, A. V Jeyarajasingam, S. A. Rutkowsky, C. Boritz, and L. Mulfinger,
“Synthesis and Study of Silver Nanoparticles,” J. Chem. Educ., vol. 322, no. 2, pp. 322–325,
2007.
[4] Z. Khan, S. A. Al-Thabaiti, A. Y. Obaid, A. O. Al-Youbi, “Preparation and characterization of
silver nanoparticles by chemical reduction method”, Colloids Surf. B., vol. 82, no.2, pp. 513-
517,2011.
[5] X. Lu, M. Rycenga, S. E. Skrabalak, B. Wiley, and Y. Xia, “Chemical synthesis of novel plasmonic
nanoparticles.,” Annu. Rev. Phys. Chem., vol. 60, pp. 167–92, 2009.

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