Optimization of Organic Capping Material on Silver Nanoparticles by Chemical Reduction and Its Functionalization for Plasmonic Sensor Application

The application of metal nanoparticles in medicine and optoelectronic devices recently has shown remarkable results. In the application of biosensor, metal nanoparticles are functionalized by the used of organic materials such as thiol-derivatives and citrate because of their high affinity to metal such as gold and silver also those organic material is dispersed in water that can be applied directly to bio-materials. Herein, we present the optimization study of capping molecules Trisodium citrate and 3-mercaptopropionic acid (3-MPA) on silver nanoparticles (AgNP) by used of modified chemical reduction and exchange ligand methods to form stable AgNP. The optimization of AgNP was done by varying the ratio among the concentration of the precursor (AgNO3), reducing agent and capping materials. Our colloidal AgNP products show bright yellow to yellow-brownish colour with the plasmonic peak for citrate capped AgNP (Ag-Citrate) at 410-424 nm, 3-MPA capped AgNP (Ag-MPA) by direct method at 428-440 nm, and 3-MPA capped AgNP (Ag-MPA) by ligand exchange method at 436-462 nm. The chemical characteristics of our Ag-citrate and Ag-MPA show the differences coordination of COO- between free citrate or 3-MPA and after citrate or 3-MPA capped on Ag. In our experiment, the TEM images for both samples of AgNPs show the spherical shape with the range of diameter size 5-38 nm which depends on the given ratio of material concentration in the synthesis process. Our AgNP solution results with the open capping of citrate or 3-MPA then can be applied as probe sensor for plasmonic sensor application.


Introduction
Metal nanoparticles have interesting characteristics because of their unique optical properties called localized surface plasmon resonance (LSPR) which is different from their bulk materials.Recently, they have been used in numerous fields like medicine, biotechnology, materials research, and energy.LSPR is an effect that corresponds to the interaction between electromagnetic waves and the conductive matter of metal nanoparticles.In this case, the metal nanoparticles which have a size smaller than the wavelength of electromagnetic wave, resulted in collective oscillation of electron in conduction band.This interaction generates coherent localized plasmon oscillations with frequency that depends on the size and shape of the metal nanoparticle [1].Previous research has demonstrated that the properties of metal nanoparticles such as physical, optical, catalytic are strongly influenced by their size distribution, morphological shape, and surface properties.These properties can be modified by using various synthesis techniques, reducing agents, and stabilizers [2].
In general, there are three methods to prepare metal nanoparticles i.e physically, chemically, or biologically.The chemical reduction method needs three reactants such as metal precursor, a reducing agent, and capping agent.The reducing agent transforms metal ion from precursor into metal nanoparticles through reduction reaction.Meanwhile, the capping agent protects the core of nanoparticles by covering the outer layer and preventing aggregation of metal core.The morphology of the metal nanoparticles can be altered by adjusting the concentration and the dosage of the reagent [3].The stabilization of nanoparticles is frequently divided into two categories i.e electrostatic stabilization and steric stabilization.Anionic species, such as halides, carboxylates, or polyoxo-anions coordinate with metal particles to achieve electrostatic stability.Additionally, the development of an electrical double layer during electrostatic process results in coulombic repulsion between the nanoparticles.Steric stabilization is accomplished by preventing the diffusion of nanoparticles due to the presence of bulky steric stabilizers.Steric stabilizers usually are organic materials such as polymers or large cations like alkyl ammonium [2].
In the case of metal nanoparticles, it is essential to optimize the mass scale and synthesis proses of its metal precursor, capping agents and stabilizer [4].Tannic acid or citrate are typically used as reducing and stabilizing agents in the chemical production of metal nanoparticles.The primary benefit of using citrate in chemical reduction method is the possibility of further nanoparticle functionalization.This is because citrate molecules have a weak interaction with metal surfaces, making citrate ions easily exchangeable with other chemicals [5].Besides that, 3-mercaptopropionic acid (3-MPA) is one of the molecules used for surface functionalization because it has a thiol group and a carboxylate group which can be modified with other biomolecules [6].Yonezawa et.al have succeeded in injecting citrate and 3-MPA simultaneously into hydrogen tetrachloroaurate in the nanoparticle synthesis process [7].Dewi et.al.succeeded in carrying out the ligand exchange method on AuNPs using 3-MPA.In the results, it resulted in complete exchange of CTAB/C with 3-MPA and showed excellent colloidal stability [8].Specifically, silver nanoparticles (AgNPs) have many potential applications in medicine and biomedicine device, due to its antimicrobial properties, beside it has unique sharper plasmon characteristic in comparison to the gold nanoparticles, while the synthesis process of AgNPs is known to be more complex regarding to the less stability of colloidal formation.
In this study, we attempt to optimize the concentration parameters of Ag precursor and capping organic materials involved during the synthesis of AgNPs.In our case, we used capping materials of citrate to form citrate capped on silver nanoparticles (Ag-Citrate) and 3-MPA to form 3-MPA capped on silver nanoparticles (Ag-MPA).We investigated the effect of LSPR of AgNP solution by used of UV-Vis and FTIR spectroscopies and TEM for capturing the images.

Synthesis of AgNPs
The synthesis of Ag-Citrate as shown in the Figure 1 was done by following the modified reduction methods from Yonezawa et.al.and Pambudi et.al [7,9].Approximately 30 mL precursor solution made of dissolved AgNO3 is placed in three necked rounded flask and heated under reflux and stirred with magnetic bar.Once the precursor solution reached 85°C, solution of Trisodium citrate dihydrate was added into the solution.The concentration of the Trisodium citrate dihydrate solution varied from 0.5 wt% to 1.5 wt%.The mixed solution was continuously heated and stirred until the change in colour to yellow occurred.The mixture then cooled down to reached room temperature.The synthesis of Ag-MPA was done by used 2 kind of methods that are direct reduction and exchange ligand.The direct reduction method was done with similar step as used in the synthesis of Ag-Citrate, which was starting with heating precursor solution under reflux and stirred to reach 85°C and continue to add the mixture of Trisodium citrate dihydrate and 3-MPA into the solution.The concentration of Trisodium citrate dihydrate is fixed at the 0.75 wt% while the concentration of 3-MPA was varied from 12, 16, 20, 25, 30, 60 and 100 µM.The mixture then is cooled down to room.On the other case, the synthesis of Ag-MPA with exchange ligand method was done by using Ag-Citrate as precursor.First, Ag-Citrate solution is placed in vial under stirring and 3-MPA solution is then added to the vial with concentration ranging from 20 to 90 µM.The mixture is then stirred for 15 minutes.The result is then stored in a closed box at room temperature.
All the AgNPs solution were characterized by use of with UV-Vis spectrometer to get its optical characteristic and Fourier Transform Infrared (FTIR) spectrometer for its chemical characteristic, while the morphology of AgNP was captured by Transmission Electron Microscope (TEM).

Optical Properties of AgNP
As show in Figure 2, the absorbance spectra from UV-Vis spectrometer shows the plasmonic peak of Ag-citrate is on the range from 410 to 424 nm which indicate the formation of Ag-Citrate in the solution.The optimization of the synthesis of Ag-citrate was done with variation of trisodium citrate concentration from 0.25% to 1.5 wt%.In our case, the increasing of citrate concentration does not consistent to the blue-shifted phenomena of plasmonic peaks, but the broadening at high wavelength are clearly shown indicated the aggregation of nanoparticles.From the absorbance spectra of Ag-Citrate, it can be concluded that optimization of Ag-Citrate colloidal formation is found by used of citrate's concentration at 0.75wt% which resulted in the sharped band with the peak position at 424 nm.
On the other case, the synthesis results of Ag-MPA by direct reduction method with 7 variations of 3-MPA concentration (12, 15, 18, 25, 30, 60, and 100 μM) as capping solution is shown in Figure 3(a).We varied the concentration of 3-MPA in μM unit, since we get the material stock in exact concentration value.The UV-Vis spectra show plasmonic peaks in the range from 420 nm to 436 nm and the spectra also show that significant shifted of plasmonic peak only appear in certain concentrations.At the low concentration of 3-MPA (12,15,18,25 μM), the Ag-MPA solutions reveal similar plasmonic peaks to that of Ag-Citrate, while the biggest shifted was shown by the 30μM of 3-MPA concentration involved in the synthesis process which revealed the peak at 436 nm and the addition of high concentration of 3-MPA shows the unclear shifted phenomena.The optimization of Ag-MPA by used of exchange ligand method was done with 7 variations of 3-mercaptopropionic concentration (20, 30, 40, 50, 60, 70, and 90 μM).The UV-Vis spectra show that the increasing concentration of capping materials resulted in large red-shifted of the plasmonic peaks.The plasmonic peak of the Ag-Citrate was used in this exchange ligand method located around 424 nm, and after the addition of 3-MPA the peaks shifted to high wavelength and saturated at around 90 μM addition of MPA with plasmonic peak position at 462 nm as shown in Figure 3  exchange ligand method.

Morphology of AgNP
The physical appearance of the AgCA solution as shown in Figure 4(a) change slightly from the original yellow colour into yellow-brownih colour which indicated the aggregation of solution even though the solution still looks transparent and also UV-Vis spectra in the Figure 2 shows the plasmonic characteristic with some shoulder peak at high wavelength.The aggregation may be formed during the stirring treatment in the synthesis for various concentrations of citrate and in our experiment, the  The TEM images of our synthesized Ag-Citrate and Ag-MPA is shown in Figure 5(a) and 5(b) by used of 1 wt% of citrate's concentration.The chemical reduction method that we used in the synthesis of AgNP show mostly spherical shape with average diameter ~18 nm for Ag-Citrate and Ag-MPA reveal diameter size ~10 nm.In the TEM images, it can be seen also some small AgNP which is less than 5 nm in diameter size.In the experiment, we purified the AgNPs solutions with high speed centrifugation at 8000 rpm for 15 min to rinse out extra citrate and others Ag ion.However, this causes quite poor particle aggregation, marked by a change in the colour of the solution to clear and the presence of precipitate.Nevertheless, our synthesized AgNPs (Ag-Citrate and Ag-MPA) are quite stable for 1 month if we keep in the refrigerator.

Chemical characterization of AgNPs.
Since the FTIR spectrometer measure the vibration modes of capping organic materials i.e citrate and 3-MPA related to the conformation on Ag metal, we tried to compare the result to that of free citrate and 3-MPA as shown in Figure 6.The specific band assignments of the FTIR spectra results are identified by use the reference from our previous study [9] and other research [10] and we summaries in Table 1.In the measurement of FTIR spectrometer, we used sample pellet by dried-up AgNPs solution about 200 µL into KBr powder to form transparent pellet and we used transmittance mode for measurement in air environment.The S-H stretching band that appeared in bulk 3-MPA is disappeared in all Ag-MPA synthesis products.This is caused by the dissociation of the S-H bond and the adsorption of the S atom from 3-MPA onto the metal surface which indicated the binding of the thiol group to the AgNP surface.The strong peak of carboxylic acid (-COOH) stretching mode at 1712 cm -1 from free 3-MPA is found to decrease after 3-MPA is anchoring on Ag, but the carboxylate asymmetric stretching (vasym COO-) at 1589 cm -1 and symmetric stretching (vsym COO-) at 1389 cm -1 are clearly observed in the Ag-CA and Ag-MPA that slightly shifted to low frequencies in comparison to that of free Citrate and 3-MPA.In the case of Ag-MPA, the carboxylate (COO-) modes are overlapped between COO-from citrate and 3-MPA.

Summary
In this works, we successfully synthesize both Ag-Citrate and Ag-MPA by using chemical reduction method and exchange ligand method.The formation of Ag-Citrate and Ag-MPA can be identified from the appearance of yellow colour solution for Ag-CA and yellow-brownish colour for Ag-MPA, together with spherical shape about 18 nm and 10 nm in diameter size, respectively as measured by TEM images.
Absorbance spectra for Ag-CA and Ag-MPA show unique plasmonic band with maximum peak position from 410-440 nm with various concentration of 3-MPA.In our case, the optimized formation of Ag-CA occurs with involvement of 0.75 wt% of Citrate during synthesis process, while the optimized condition for Ag-MPA solution occur when we involve the concentration 3-MPA at 30 µM.The chemical characterization shows the disappearance of S-H bond from bulk 3-MPA that indicate new bond of S ion with Ag.Further, Ag-citrate and Ag-MPA show the differences coordination of COO-between free citrate or 3-MPA and after citrate or 3-MPA capped on Ag.Finally, based on Ag-citrate and Ag-MPA solution result with open capping of citrate or 3-MPA, it can be applied as probe sensor for plasmonic sensor application by capturing the other organic molecule or biomolecule on surface of our synthesized AgNPs.

Figure 3 .
Figure 3. Absorbance spectra of Ag-MPA by used of (a) direct synthesis method and (b) exchange ligand method.

10th
Asian Physics Symposium (APS 2023) Journal of Physics: Conference Series 2734 (2024) 012043 optimum condition is found at citrate 0.75 wt% where the colour of AgCA solution is slightly yellowbrown colour.In the case of Ag-MPA solution as shown in Figure 4(b) dan 4(c), the colour change by the addition of various concentration of 3-MPA does not clearly show at low concentration of 3-MPA but at the concentration of 90-100mM, it can be observed that the colour changes to watery brownish colour.

Figure 6 .
Figure 6.FTIR spectra of Ag-Citrate and Ag-MPA in comparison to the free Citrate and 3-MPA.

Table 1 .
Band assignment of Ag-Citrate and Ag-MPA in comparison to the free Citrate and 3-MPA.