Helically Grooved Gold Nanoarrows: Controlled Fabrication, Superhelix, and Transcribed Chiroptical Switching

Abstract

Open AccessCCS ChemistryRESEARCH ARTICLE1 Sep 2021Helically Grooved Gold Nanoarrows: Controlled Fabrication, Superhelix, and Transcribed Chiroptical Switching Song Wang, Liheng Zheng, Wenjie Chen, Lukang Ji, Li Zhang, Wensheng Lu, Zheyu Fang, Fucheng Guo, Limin Qi and Minghua Liu Song Wang Beijing National Laboratory for Molecular Science (BNLMS), CAS Key Laboratory of Colloid, Interface and Chemical Thermodynamics, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 University of the Chinese Academy of Sciences, Beijing 100049 Google Scholar More articles by this author , Liheng Zheng State Key Laboratory for Mesoscopic Physics, Collaborative Innovation Center of Quantum Matter, Peking University, Beijing 100871 Google Scholar More articles by this author , Wenjie Chen Beijing National Laboratory for Molecular Science (BNLMS), CAS Key Laboratory of Colloid, Interface and Chemical Thermodynamics, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 University of the Chinese Academy of Sciences, Beijing 100049 Google Scholar More articles by this author , Lukang Ji Beijing National Laboratory for Molecular Science (BNLMS), CAS Key Laboratory of Colloid, Interface and Chemical Thermodynamics, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 Google Scholar More articles by this author , Li Zhang *Corresponding authors: E-mail Address: [email protected] E-mail Address: [email protected] Beijing National Laboratory for Molecular Science (BNLMS), CAS Key Laboratory of Colloid, Interface and Chemical Thermodynamics, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 Google Scholar More articles by this author , Wensheng Lu Beijing National Laboratory for Molecular Science (BNLMS), CAS Key Laboratory of Colloid, Interface and Chemical Thermodynamics, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 Google Scholar More articles by this author , Zheyu Fang State Key Laboratory for Mesoscopic Physics, Collaborative Innovation Center of Quantum Matter, Peking University, Beijing 100871 Google Scholar More articles by this author , Fucheng Guo Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, College of Chemistry, Peking University, Beijing 100871 Google Scholar More articles by this author , Limin Qi Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, College of Chemistry, Peking University, Beijing 100871 Google Scholar More articles by this author and Minghua Liu *Corresponding authors: E-mail Address: [email protected] E-mail Address: [email protected] Beijing National Laboratory for Molecular Science (BNLMS), CAS Key Laboratory of Colloid, Interface and Chemical Thermodynamics, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 University of the Chinese Academy of Sciences, Beijing 100049 Google Scholar More articles by this author https://doi.org/10.31635/ccschem.020.202000472 SectionsSupplemental MaterialAboutAbstractPDF ToolsAdd to favoritesDownload CitationsTrack Citations ShareFacebookTwitterLinked InEmail Plasmonic chiroptical materials, coupled by chirality and surface plasmons, have been attracting great interest due to their potential applications, such as photonics, chiral recognition, asymmetric catalysis and biosensing. Herein, we constructed a new chiral plasmonic nanostructure—helically grooved gold nanoarrows (HeliGNAs) by introducing l-/d-cysteine (l-/d-Cys) during the growth of the gold nanoarrows (GNAs). The variation of the concentration of l-/d-Cys leads to the HeliGNAs with a tunable plasmonic circular dichroism (PCD). Moreover, HeliGNAs can self-assemble into a superhelix at a higher concentration of l-/d-Cys, whose chirality can be transcribed to an achiral azobenzene derivative localized in the hotspots from the superhelix of HeliGNAs. A chiroptical switch is established based on the photoisomerization of azobenzene. This work provides not only a new way for fabricating helically featured gold nanostructure but also discloses interaction between chiral molecules/achiral plasmon, chiral plasmon/achiral molecules, thus deepening the understanding of plasmon-chirality-coupled circular dichroism. Download figure Download PowerPoint Introduction Chiral plasmonic nanomaterials, coupled by the plasmons and chirality, have been attracting great interest owing to their unprecedented optical properties in photonics, chiroptical switching, enantioselective recognition and separation, asymmetric catalysis, biological sensing, and diagnosis.1–7 The chiral plasmonic nanostructures have been generally created through a top-down lithography8,9 and physical vapor deposition,10 or a bottom-up surface modification11–13 and self-assembly.14,15 Besides the discrete chiral nanostructures, self-assembly provides an alternative way for fabricating chiroptical nanomaterials that can not only transfer the molecular chirality to a supra- and nanoscale level but also hierarchically amplify the chirality.16–18 Plasmonic nanoparticles attached to chiral biomolecules like DNA,19,20 protein21 and cellulose,22 chiral supramolecular assemblies,23,24 liquid crystal,25 polymers,26 and silica helices27 have been shown to exhibit well-defined plasmonic circular dichroism (PCD). Different from many organic or biological nanostructures, where helical features like double-helical DNA and α-helical protein are often observed, most of the chiral plasmons do not show helical features although they can show PCD. It was recently shown by Lee et al.28 that ornately chiral and chiroptically responsive stellated octahedra could be grown from gold nanocubes with the addition of enantiomeric cysteine. Gonzalez-Rubio et al.29 showed the micelle-directed chiral seeded growth of gold nanocrystals with sharp chiral wrinkles. Herein, we report the fabrication, self-assembly, and the chiroptical properties of a new nanostructure, helically grooved gold nanoarrows (HeliGNAs), as illustrated in Figure 1a. Starting from a gold nanorod, we found that when l-/d-cysteine (l-/d-Cys) was introduced as a chiral inducer to grow gold nanoarrows (GNAs),30 such HeliGNAs could be generated. Interestingly, the direction of the helical grooves on HeliGNAs followed the molecular chirality of l-/d-Cys. Moreover, the HeliGNAs could self-assemble into left- or right-handed superhelix, respectively, when the concentration of l-/d-Cys exceeded 10 μM. Remarkably, the strong electromagnetic fields or hotspots generated between the adjacent HeliGNAs could influence the optical properties of the deposited molecules at their surface. Subsequently, an achiral sodium (4-phenylazo-phenoxy)-acetate (AZONa) loading on the superhelix showed an induced circular dichroism (ICD), and a phototriggered chiroptical switch was fabricated based on the photoisomerization of AZONa (Figure 1b). This work provided not only a new strategy for fabricating helically featured plasmonic nanostructure but also disclosed interplay between chiral molecules/achiral plasmon, chiral plasmon/achiral molecules, thus deepening our understanding of chiral molecular coupling in plasmon. Figure 1 | Schematic illustration for the synthesis of chiral gold nanostructures. (a) GNAs and HeliGNAs synthesized by overgrowth of GNRs in various concentrations of l-Cys. At a higher concentration of l-Cys, the HeliGNAs can self-assemble to left-handed superhelix with the opposite PCD signals compared with the discrete GNAs and HeliGNAs. (b) The superhelix assembled by HeliGNAs induces an achiral AZONa to exhibit CD signals, and a chiroptical switch is fabricated based on photoisomerization of AZO moiety. GNAs, gold nanoarrows; HeliGNAs, helically grooved gold nanoarrows; PCD, plasmonic circular dichroism; AZONa, achiral sodium (4-phenylazo-phenoxy)-acetate; CD, circular dichroism; AZO, azobenzene. Download figure Download PowerPoint Experimental Methods Materials Tetrachloroauric(III) trihydrate (HAuCl4·3H2O, 99.99%), hexadecyltrimethylammonium bromide (CTAB, 98%), hexadecyltrimethylammonium chloride (CTAC, 96%), l-Cys (98%), and d-Cys (98%) were purchased from Alfa Aesar, Shanghai, China. Silver nitrate (AgNO3, 99.9%), sodium oleate (NaOL, 98%), sodium borohydride (NaBH4, 98%), and l-ascorbic acid (AA, 99%) were purchased from Innochem, Beijing, China. Hydrochloric acid (HCl, 36–38 wt %) was purchased from Beijing Chemicals, Beijing, China. All the materials were used without further purification. All aqueous solutions in this investigation were prepared using Milli-Q water (18.2 MΩ·cm). Synthesis of GNAs with chiroptical properties GNAs were synthesized by a seed-mediated method as reported previously.30 Gold nanorods (GNRs) were synthesized as the seeds for the growth of GNAs. GNAs with chiroptical properties could be obtained by adding l-/d-Cys to the growth solution of GNAs. The detailed operation steps were presented as follows. First, a seed solution was prepared in an aqueous solution of CTAB (10 mL, 100 mM) by reduction of HAuCl4 (0.25 mL, 10 mM) with a freshly prepared ice-cold NaBH4 (600 μL, 10 mM) as a reducing agent. After vigorous stirring, the resultant seed solution was aged at 27 °C for 30 min. Second, for the preparation of the GNRs growth solution, NaOL (100 mL, 20 mM), CTAB (74 mL, 100 mM), and 26 mL water were mixed in a beaker flask under mild stirring and stored at 30 °C in a water bath, followed by the addition of 3.84 mL AgNO3 (10 mM). The mixed solution was kept undisturbed for 15 min at 30 °C, and then 10 mL HAuCl4 (10 mM) was added. After undisturbed standing for 90 min, the solution changed from yellow to colorless. Then 1.2 mL HCl (36–38 wt %) was injected. After another 15 min, 1.2 mL AA (20 mM) was added and vigorously stirred for 30 s. Subsequently, 80 μL seed solution was introduced. The growth solution was kept undisturbed for 12 h at 27 °C, and then GNRs can be obtained. GNRs were centrifuged twice (8000 rpm, 5 min), concentrated 10 times in volume and redispersed into aqueous CTAB solution (1 mM) as the seeds for the growth of GNAs. Finally, a growth solution of GNAs was prepared by adding 300 μL of 10 mM HAuCl4 into 10 mL of 100 mM CTAC, followed by the addition of 150 μL AgNO3 (10 mM) and 200 μL HCl (1 M). Au3+ was then reduced to Au+ by the rapid injection of 150 μL AA solution (100 mM). The solution was vigorously stirred for 30 s, and then the growth of GNAs was initiated by adding different concentrations of l-/d-Cys and 500 μL of GNRs seeds solution into the growth solution. The growth solution was placed in a 27 °C bath for 2 h. GNAs could be obtained when the brownish red solution gradually became blue. The solution was centrifuged twice (5000 rpm, 4 min) to remove unreacted reagents and was redispersed in a 1 mM CTAB solution for further characterization. Chiral induction HeliGNAs were prepared with 15 μM l-/d-Cys in overgrowth solution, and then concentrated 10 times in volume. The AZONa synthesized according to the method reported previously by our group was selected for our study.31 The AZONa was dissolved in water to generate a stock solution of 15 mM. The sample solutions were obtained by mixing 1 mL HeliGNAs and 50 μL AZONa solutions. The solutions were vigorously stirred for 30 s, and then incubated for 12 h to induce the chirality of AZONa. The AZONa was transformed from trans- to cis-isomer by 365 nm UV irradiation for 30 min. Numerical simulations Full-field electromagnetic wave simulations of the helical assembly of HeliGNAs (l-Cys, 15 μM) were performed by using the commercial finite element method (FEM) solver (COMSOL Multiphysics 5.3, COMSOL Co., Ltd., China), with the gap between adjacent HeliGNAs set at 2 nm. The simulation domain included structures with perfectly matched layers in x-, y-, and z-directions. The scattering field was chosen, and incident illumination was a circularly polarized wave with x-axis propagation direction. The permittivity data of gold were taken from sample data, and a minimum mesh size of 1 nm was used to refine the tip and spiral area of the HeliGNAs. Finite difference time domain (FDTD) method was used in the numerical simulation about optical response of GNAs and HeliGNAs. The refractive index of the water was set at n = 1.33. For the dielectric function of gold, Johnson’s data were adopted. The geometric parameters of GNAs and HeliGNAs were the same as the practical parameters in experiments. Characterization UV–vis spectra were obtained using a Hitachi U-3900 spectrophotometer, Japan in quartz cuvettes (light path 1 mm). Circular dichroism (CD) spectra were detected on a JASCO J-815 spectrophotometer, Japan in quartz cuvettes (light path 1 mm). CD spectra of GNAs at different temperatures or different electric field intensities were also recorded on the JASCO J-815, Japan with corresponding accessories. Kuhn’s dis-symmetry factor (g-factor) was calculated from the measured extinction and CD values using: g-factor = 2 (AL − AR)/(AL + AR). Scanning electron microscopy (SEM) images of GNAs were recorded on a Hitachi S-4800 field emission SEM (FE-SEM) instrument with an accelerating voltage of 10 kV. The energy-dispersive X-ray spectroscopy (EDX) was measured on a Horiba EMAX x-act energy-dispersive spectroscope, which was attached to the Hitachi S-4800 system, Japan. Transmission electron microscopy (TEM) images and high-resolution TEM (HRTEM) images were captured using a JEM-2100F system, JEOL, Japan. Fourier transform infrared (FT-IR) spectra (KBr) were recorded on a JASCO FT/IR-660 Plus spectrophotometer, Japan with a wavenumber resolution of 4 cm−1. X-ray photoelectron spectroscopy (XPS) spectra were measured on an ESCALAB250XI electron spectrometer, Thermo Fisher Scientific, US from VG Scientific, US using 300 W Al Kα radiation. Dynamic light scattering (DLS) measurements were recorded on a Zetasizer Nano ZEN3600 (Malvern, UK) to investigate the particle size distribution of GNAs in solution. X-ray diffraction (XRD) analysis was performed on a Rigaku D/Max-2500, Japan X-ray diffractometer (Japan) with Cu Kα radiation (λ = 1.5406 Å), operated at a voltage of 40 kV and a current of 200 mA. The step size of XRD measurements was 0.026°. The X-ray scan speed was 4° min−1, and the scan range was 30–90°. Results and Discussion Chiral molecules directed HeliGNAs: fabrication and superhelix To obtain the HeliGNAs, an aqueous-based, two-step growth method was designed. In the first step, GNRs ( Supporting Information Figure S1a) with about 100 nm in length and 25 nm in diameter were synthesized using a seed-mediated method. Second, during the overgrowth with CTAC to facilitate the generation of GNAs,30l-/d-Cys was added to control the chirality of the GNAs. It was found that by simply changing the concentration of l-/d-Cys in the growth solution, the growth of the GNRs could be affected, leading to the emergence of HeliGNAs and a superhelix. SEM and TEM images (Figure 2 and Supporting Information Figure S2) show the morphology variation of GNAs as a function of l-/d-Cys concentration from 0.5 to 20 μM, while the concentration of HAuCl4 in the overgrowth solutions was kept at 0.3 mM. For simplification, these chiral GNAs are denoted as GNAs (l-/d-Cys, n μM). As shown in Figures 2a, 2e and 2i and Supporting Information Figures S2a and S2b, when the concentration of l-/d-Cys was <1 μM, the morphology of GNAs (l-/d-Cys, ∼1 μM) was almost the same as that of GNAs without Cys, where the GNAs showed smooth surfaces. As the concentration of l-/d-Cys increased to around 5 μM (Figures 2b, 2f and 2j), HeliGNAs (l-/d-Cys, 5 μM) with a basic arrow shape were obtained, while the four-wing shafts in the middle of the GNAs disappeared. Eventually, some helical grooves appeared. When carefully investigating the direction of these grooves, we found that the orientation of the grooves relied on the absolute configuration of Cys. The right-handed helical grooves were observed in the presence of l-Cys, while left-handed helical grooves were observed for d-Cys. As shown in Figures 2c and 2g, it was obvious that the opposite orientation of these grooves could be associated with utilization of l- and d-Cys, respectively. More geometric parameters of HeliGNAs are provided in Supporting Information Figure S3. With increases in the concentration of l-/d-Cys, the length of the two pyramidal heads of the GNAs gradually decreased, and more pitch number appeared at the middle position of HeliGNAs with smaller pitch distance. For the HeliGNAs (l-Cys, 5–20 μM), the depth of helical grooves was around 4 nm, and the aspect ratio was around 2.3, with only small fluctuations. Figure 2 | Morphology of GNAs (l-/d-Cys, n μM) obtained with different concentrations of l-/d-Cys and 0.3 mM HAuCl4. SEM images of (a) GNAs (l-Cys, 1 μM), (b and c) HeliGNAs (l-Cys, 5 μM), (d) HeliGNAs (l-Cys, 15 μM), (e) GNAs (d-Cys, 1 μM), (f and g) HeliGNAs (d-Cys, 5 μM), and (h) HeliGNAs (l-Cys, 15 μM). Insert: the description of helical grooves orientation for HeliGNAs (l-Cys, 5 μM) and HeliGNAs (d-Cys, 5 μM), respectively, marked by blue frame in (c) and (g). TEM images of (i) GNAs (l-Cys, 1 μM), (j) GNAs (l-Cys, 5 μM), (k) HeliGNAs (l-Cys, 15 μM), and (l) HeliGNAs (d-Cys, 15 μM). GNAs, gold nanoarrows; SEM, scanning electron microscopy; HeliGNAs, helically grooved gold nanoarrows; TEM, transmission electron microscopy. Download figure Download PowerPoint HRTEM was used to analyze the atomic lattice structure of GNAs. As shown in Supporting Information Figure S4, the exposed lattice fringes with distance of 0.24 nm of pyramidal heads and the middle part of GNAs (l-Cys, 1 mM) were observed, which could be assigned to the {111} planes. For the HeliGNAs (l-Cys, 5 μM), although the helical grooves formed, the crystal facets were consistent with GNAs (l-Cys, 1 mM), which could be assigned to the {111} planes. From the XRD patterns in Supporting Information Figure S5, the intensity of the (111) plane was much stronger than other crystal planes. Therefore, the gold nanocrystal grew along the (111) plane, which was fully consistent with the results observed by TEM. As we previously reported, the GNAs (l-/d-Cys, 0.5–10 μM) could be assembled as superstructures with an orderly arrangement on solid interfaces by solvent evaporation, which was driven by the entropic forces favoring close packing and shape complementarity (Figures 2a, 2b, 2e, and 2f).30 It was interesting to find that, as the concentration of l-/d-Cys (15–20 μM) further increased, a large number of one-dimensional (1D) nanochains were observed instead of the close packing. Through careful investigation, these HeliGNAs were found to further assemble into a superhelix, as shown in Figures 2d and 2h and Supporting Information Figures S6 and S8. Moreover, the helicity of the superhelix was strongly related to the helicity of HeliGNAs or the configuration of the surface modified Cys. l- and d-Cys finally resulted in the formation of left- and right-handed superhelices, respectively. TEM images further confirmed the helical assembly of HeliGNAs under high concentrations of l- and d-Cys, and the helical orientation of the superhelix depended on the configuration of Cys (Figures 2k and 2l and Supporting Information Figures S7 and S9). We inferred that Cys was the main factor for the formation of the superhelix. FT-IR and XPS spectra ( Supporting Information Figures S10 and S11) confirmed that the Cys molecules were tethered to the surface of HeliGNAs by the Au–S bond. At a high Cys concentration, these Cys molecules on the surface of HeliGNAs might be linked to the HeliGNAs via hydrogen bond or electrostatic interactions. Owing to the chiral nature of Cys molecules, adjacent HeliGNAs approaching each other would follow the chiral geometry to decrease the steric hindrance to form the superhelix. It should be noted that for other chiral inducers, glutathione (GSH) was found to lead to the helical grooves formation on GNAs surface as well ( Supporting Information Figure S12a). In sharp contrast, after replacing Cys with mercaptoacetic acid during overgrowth, the helical grooves could not form on the surface of GNAs ( Supporting Information Figure S12b), which indicated that the chiral inducer (Cys or GSH) played an important role in the formation of helical grooves on GNAs. Chiroptical properties of the HeliGNAs UV–vis and CD spectra were used to investigate the optical properties of the chiral GNAs. As shown in Supporting Information Figure S13, the longitudinal surface plasmonic resonances (LSPR) absorption peak of GNRs was at 900 nm, and the transverse surface plasmonic resonances (TSPR) absorption peak was at 508 nm. Comparing with UV-vis spectrum of GNRs, the LSPR peak of GNAs (l-Cys, 0.5 μM) first showed blue shift to 801 nm, and the TSPR peak had a red shift to 584 nm with a shoulder peak at 535 nm (Figure 3a). With increasing concentration of l-Cys, the LSPR absorption peak of GNAs (l-Cys, ∼10 μM) exhibited in a further blue shift to 720 nm, and the TSPR absorption peak became one peak and exhibited a red shift to 542 nm. The variation of UV spectra could be attributed to the change of aspect ratio of GNAs.32 As shown in Supporting Information Figure S3c, since the aspect ratio became smaller with increasing concentrations of l-Cys, the LSPR absorption bands showed a blue shift. Obviously, the cross section of GNAs (l-Cys, <1μM) exhibited anisotropy (see Supporting Information Figure S3b), and the TSPR absorption bands had two peaks because of the difference between D1 and D2. However, the HeliGNAs (l-Cys, 5 μM) had a relatively consistent diameter and so only exhibited one TSPR absorption peak.33 The numerical simulation results, as shown in Supporting Information Figure S14, were consistent with the experimental results. This demonstrated that the two peaks in the TSPR band of GNAs originated from the anisotropic cross section. When the concentration of l-/d-Cys exceeded 10 μM, the LSPR absorption peak of HeliGNAs (l-Cys, 15 and 20 μM) significantly widened and dramatically decreased, which might be caused by the self-assembly of HeliGNAs (l-Cys, 15 and 20 μM) in solution. It has been reported that the aggregation or assembly of nanorods could cause the hypochromic and widening effect of the plasmonic peak.34,35 Apart from this, a weak absorption peak appeared around 280 nm which might correspond to the plasmon resonance of a small amount of silver atoms, which were reduced and deposited on the surface of GNAs, as shown in Supporting Information Figure S15.36 Figure 3 | Chiroptical properties of HeliGNAs synthesized with different concentrations of l-Cys from 0.5 to 20 μM and 0.3 mM HAuCl4. (a) UV–vis spectra. (b) CD spectra. Anisotropic factors g of CD peaks corresponding to (c) TSPR and (d) LSPR of GNAs with different concentrations of l- and d-Cys. HeliGNAs, helically grooved gold nanoarrows; GNAs, gold nanoarrows; CD, circular dichroism; TSPR, transverse surface plasmonic resonances; LSPR, longitudinal surface plasmonic resonances. Download figure Download PowerPoint The deposition of Ag atoms on the surface of GNAs was also a necessary condition for the anisotropic growth of GNAs. According to a previous paper, the deposition of Ag atoms could lead to selective passivation of the {111} facets, resulting in different deposition rates of Au atoms on different crystal facets.37 As shown in Supporting Information Figure S16, we could not obtain the GNAs without AgNO3 in overgrowth solution. When the d-Cys was used in the overgrowth solution, the UV–vis spectra of the synthesized GNAs (d-Cys, n μM) were identical with that of GNAs (l-Cys, n μM) ( Supporting Information Figure S17a). The chiroptical activity of GNAs was investigated by CD spectra. As shown in Supporting Information Figure S18, the GNAs without Cys modification did not exhibit any PCD. However, obvious PCD of GNAs was obtained even when the concentration of Cys was as low as 0.5 μM (Figure 3b and Supporting Information Figure S17b). As shown in the CD spectra of GNAs in Figure 3b, the CD intensity and profiles were strongly dependent on the concentration of l-Cys. When the concentration of l-Cys was low (0.5 and 1 μM), a positive peak from 500 to 700 nm corresponded to TSPR absorption, and a negative peak from 700 to 900 nm attributed to LSPR absorption appeared. When the concentration of l-Cys increased (5 and 10 μM), the CD profiles were similar to those in a low Cys concentration. However, the CD intensity was obviously enhanced because of the formation of helical grooves ( Supporting Information Figure S3e). It is worth noting that, with the l-Cys concentration (15 and 20 μM) further increasing, the CD spectra of HeliGNAs (l-Cys, 15 and 20 μM) exhibited opposite PCD signals compared with the GNAs (l-Cys, 0.5–10 μM) (Figure 3b). The CD spectra had a negative TSPR peak from 500 to 720 nm and a positive LSPR peak around 880 nm. We speculated that the inversion of CD signals was caused by the self-assembly of HeliGNAs in solution. This result was consistent with the aforementioned SEM and UV–vis spectra. Moreover, for HeliGNAs (l-Cys, 20 μM), a new PCD band appeared from 230 to 370 nm, which could be attributed to the plasmon resonance of Ag on the surface of HeliGNAs.36 Supporting Information Tables S1 and S2 show the content of Au and Ag atoms of GNAs, which was analyzed by EDX. With the increase of Cys, the content of Ag on the surface of HeliGNAs also increased, and the PCD signal of Ag was obtained. The CD spectra of GNAs with d-Cys as additive were opposite to the CD spectra of GNAs (l-Cys, 0.5–20 μM) ( Supporting Information Figure S17b). The anisotropic factor (g-factor), which was used for quantitative comparison of chiroptical properties among different GNAs, is shown in Figures 3c and 3d and Supporting Information Table S3. With the concentration of l- and d-Cys increasing, g-factor enhancement and chiral inversion could be clearly observed. We speculated that the superhelix of HeliGNAs formed in a high concentration of Cys had already assembled in solution. Thus, the superhelix could not be observed in the superstructure of GNAs prepared under low Cys concentration on the solid interface. To clarify whether the superhelix was formed in solution or not, the DLS of GNAs was measured, as shown in Supporting Information Figure S19. This showed only a small difference in the size distribution of the GNAs when the Cys concentration was in the range of 0.5–10 μM. One peak was around 30 nm, and the other was around 130 nm, which could be ascribed to the diameter and length of the GNAs (l-/d-Cys, 0.5–10 μM), respectively. However, the size distribution of HeliGNAs (l-/d-Cys, 15 and 20 μM) showed distinct changes, which only had a main peak around 150 nm and a very weak peak around 4800 nm. Therefore, we inferred that large aggregates existed in solution. These indicated that the high concentration (15 and 20 μM) of l- and d-Cys might further link the HeliGNAs into assemblies in the solution. To confirm the contribution of chiral grooves to the enhancement of PCD signals, we also monitored the variation of CD profiles with different concentrations of HAuCl4 when the concentration of l-/ d-Cys was kept at 5 μM. The amount of HAuCl4 in the overgrowth solution could also influence the final morphology and PCD response of the HeliGNAs. For comparison, five types of HeliGNAs were synthesized in the overgrowth solution with concentrations of HAuCl4 from 0 to 0.4 mM. From the SEM images in Figures 4a–4d and Supporting Information Figure S20 and geometric parameters in Supporting Information Figure S21, the morphology variation of HeliGNAs was observed. The GNRs maintained their original morphology in the overgrowth process without HAuCl4. When the concentration of HAuCl4 was 0.1 mM, the HeliGNAs only had two small pyramidal heads and shallow grooves in the middle part. When a higher HAuCl4 concentration was used, the two pyramidal heads grew larger, and the depth of helical grooves increased, with an obvious increase in diameter and a decrease in aspect ratio and pitch number. The changes in the morphology of HeliGNAs caused the transformations in the absorption spectra as shown in Figure 4e and Supporting Information Figure S22. The decrease in the aspect ratio ( Supporting Information Figure S21b) resulted in