Gold nanoparticles (AuNPs) have been used in various applications in high-technology fields such as organic photovoltaics, sensory probes, therapeutic agents, drugdelivery in biological and medical applications, electronic conductors and catalysis due totheir unique optical properties. These microscopic particles, which are smaller than 100 nmin size, give off vibrant colors when interacting with light, and such interactions arecontrolled by the size and the environment around the surface of the gold nanoparticles. Inaddition, these gold nanoparticles have also shown to bind to many other organic moleculesin the self-assembled monolayer (SAM), which is defined as the molecular assembliesformed by adsorption and are organized into more ordered domains (figure 1 below). Inthis self-assembled monolayer system, the organic molecule which consists of a head groupand a functional group linked by a “tail” binds to the substrate (gold surfaces) via the headgroup. Thiols and disulfides as the most common head groups have shown to bind to goldsurfaces, although the mechanism is not entirely understood. Based on the ab initiocalculations of disulfides, the Gronbeck group (2000) proposed that thiol molecules have astrong interaction with the gold surface due to the formation of the strongly bound gold-thiolate (Au-S) bond.The unique properties of these gold nanoparticles in their interaction with lightalong with their biocompatibility allow them to be helpful in biomedical applications. Theyhave recently emerged as nanocarriers in target-specific delivery of therapeutic agents orsmall drug molecules. With the aim of utilizing these gold nanoparticles as potential drugcarriers, the Wang group focused their research on the interaction of the gold nanoparticlesand the photochemistry of 6-thioguanine (6-TG), the small thiopurine drug molecules thatbelong to the class of thiobase drugs. These drugs are known for their anticancer, anti-inflammatory and immunosuppressant activity. 6-TG molecules are usually incorporatedinto DNA or RNA via metabolism but long-term use of these drugs has a high potential ofhaving skin cancer. Specifically, 6-TG molecules absorb UVA (ultraviolet A) rays verystrongly at the wavelength of 342 nm and photosensitizes as the absorbed energy istransferred to the molecular oxygen to generate singlet oxygen (1O2) from the triplet state(3O2). The generated singlet oxygen molecules are then likely to react with 6-TG to produceguanine-6-sulfonate (GSO3) molecules, which becomes an irreversible block in DNAreplication and transcription since they are incapable of binding to DNA (Scheme 1). As aresult, 6-TG can cause oxidative damage to DNA molecules. Previous research by Podsiadloet al (2008) has shown that AuNPs promote intracellular uptake of the 6-TG drug moleculesand enhance the antiproliferation activity of 6-TG in cancer cells due to the formation of thestrong Au-S covalent bond. The Wang group examined the photophysical andphotochemical modifications of 6-TG due to their loading onto gold nanoparticles as anattempt to prevent 6-TG photocytotoxicity. Based on the ultraviolet irradiation experimentsand ab intio calculations, they have shown that the AuNPs can prevent the photoreaction of6-TG to produce singlet oxygen and block DNA damage by regulating its photochemical andphotophysical properties.In order to test the interaction between the gold nanoparticles and 6-TG, the AuNPsmolecules were first synthesized from the reaction between tetracloroaurate (III)trihydrate and sodium citrate tribasic dehydrate, resulting in a gold solution of dispersedspherical particles with an average diameter of 15 ± 2 nm according to transmissionelectron microscopy (TEM) characterization. The loading of 6-TG onto AuNPs was preparedby incubating 6-TG molecules with AuNPs for 5 minutes as the color of the gold solutionchanges from wine red to purple. The excess unbound 6-TG molecules were filtered bycentrifugation and the supernatant was removed. The precipitate was redispersed in thesame volume of the solution and the process was repeated three times. UV-Vis spectroscopywas used to determine the final concentration of the 6-TG-AuNP-solution through theabsorbance of 6-TG. Samples of 6-TG integrated into single-strand DNA (TG-ss-DNA) anddouble-strand DNA (TG-ds-DNA) binding to AuNPs were also prepared in the similarmanner, however, the incubation time required two hours instead of 5 minutes. The finalconcentrations of these samples were also confirmed by UV-Vis absorbance to be 1.55 and1.37 ?M for TG-ss-DNA and TG-ds-DNA respectively. Regarding the laser irradiation assay,the third harmonic (355 nm – UVA ray) and fourth harmonic (266 nm – UVC ray) of aContinuum Surelite II Nd:YAG laser beams were used to irradiate through the sampleprepared in a 1 cm quartz cuvette for a certain amount of time and afterwards the sampleswere analyzed for UV-Vis absorption and fluorescence. In addition, computational analysisnamed SA-CASSCF was also incorporated in order to explain the molecular mechanism ofAuNPs and 6-TG via the geometry optimization of the ground state (S0), the lowest threeexcited singlet states (S1min, S2min, and S3min), and the lowest two excited triplet states (T1minand T2min).The photochemical reaction of only 6-TG (no AuNPs were used) in the presence ofUVA and UVC excitation as the control was manifested by the UV-Vis absorbance of 6-TG(Figure 1). As the 266 nm and 355 nm laser, which corresponds with the UVA and UVC light,irradiate the aqueous solution of 6-TG with excess O2, the photoactivation occurs andthrough time the amount of 6-TG decreases, as can be seen by the decrease in absorbance at342 nm (Figure 1b,d). On the other hand, as the reaction continues, the amount of GSO3produced increases, as can be seen by the increasing fluorescence intensity at 408 nm (Fig 1a,c). GSO3 is produced due to the oxidation of 6-TG by the singlet oxygen 1O2, which ispreviously the result of UV excitation of 6-TG (Scheme 1). Both UVA and UVC rays caninduce the formation of singlet oxygen and also the oxidation of 6-TG by 1O2 to produce GSO3,which leads to DNA oxidative damage.As the photoactivation of 6-TG by UV excitation was seen in the control study,AuNPs were examined for its potential drug carrier ability. When 6-TG is added to the goldsolution, the loose citrate shell of the AuNPs could be replaced through the exchangereaction and thus the Au-S covalent bond is formed (Figure 3g). The negatively chargedcitrate can be used to stabilize the AuNPs, but as it dissociates, the AuNPs aggregate. TheAuNPs activity can be analyzed in their two distinct states: monodispersion or aggregation.The monodispersion of gold nanoparticles can be confirmed by the UV-Vis absorbance at520 nm while their aggregation can be seen at 700 nm. When the 266 nm laser (UVC ray)was used to irradiate the AuNP-6-TG solution, the increasing fluorescence intensity of the408 nm band for the GSO3 formation was still observed (Figure 3a), indicating that thephotoactivation and oxidation of 6-TG still took place to produce GSO3 similar to the control,however the intensity of the band is much less compared to the control one (about 860 a.uin Figure 1a and 140 a.u in Figure 3a respectively). The UV-Vis spectrum of the AuNPsshows an increasing absorbance peak 520 nm and a decreasing peak at 700 nm (Figure 3b).This means that the gold nanoparticles no longer aggregate upon UVC excitation and theAuNPs returns to the monodispersion state. The Au-S bond formation during the pre-incubation of 6-TG with gold nanoparticles may have been destroyed due to the O-S bondthroughout the irradiation time. AuNPs cannot prohibit 6-TG photooxidation reaction in thepresence of UVC light. However, in the case of UVA excitation at 355 nm, the fluorescenceband at 408 nm of GSO3 was not seen over irradiation time, implying that GSO3 is notproduced and the photooxidation of 6-TG did not occur (Figure 3c). In terms of the state ofthe gold nanoparticles, the negligible change in the absorbance at both 520 nm and 700 nmmeans that the gold nanoparticles remain aggregated throughout the irradiation time(Figure 3d). These results have shown that the loading of 6-TG onto gold nanoparticlesinhibits 6-TG from being oxidized by 1O2 to produce GSO3 and thus DNA oxidative damagewill be prevented (Figure 3g). In addition, since the AuNPs absorb light broadly from UV tovisible region, they can still be excited under the laser irradiation of 266 and 355 nm, acontrol experiment of AuNP excitation was conducted to test whether AuNP excitationalone can affect 6-TG photooxidation. The authors found that the excitation of AuNP has noeffect on the photooxidation of 6-TG because no fluorescence intensity of GSO3 was detected(Figure 3e). Moreover, no disaggregation of AuNPs was found, since the peaks at 520 nmand 700 nm saw no change in absorbance (Figure 1f).The mechanisms of the gold nanoparticles affecting the photochemical properties ofab initio calculations for the lowest three excited singlet states (S1min/S1, S2min/S1 andS3min/S3) and the lowest three triplet states (T1min/T1, T2min/T1 and T3min/T3) wereperformed. The S2 excitation state under UVA excitation (355 nm) corresponds to the C=S(purine) bond deformation as can be seen with the elongation of C=S bond from ? to?*orbital of 6-TG (Figure 4). When 6-TG is pre-incubated with gold nanoparticles, and theC=S bond of 6-TG is removed and the C-S-Au bond is formed. As a result, the 355 nm lightcannot excite 6-TG to the S2 state anymore once 6-TG is bound to AuNPs. On the other hand,the UVC rays (266 nm) corresponds to the S3 excitation, which localizes to the C=N bondand results in the dihedral angel change in the N3?C2?N11?H12 bond (Figure 4). Since theUVC excitation does not correspond to the S2 state localized to the C=S bond but instead theS3 state, pre-incubation of 6-TG and AuNPs cannot prevent 6-TG from being photoactivatedto S3 and thus 6-TG molecules can still be photooxidized to produce GSO3. The Au-S bondformation only modifies the S2 excitation and thus blocks UVA photochemistry.In conclusion, the loading of 6-TG onto gold nanoparticles can be a potentialpathway in the inhibition of GSO3 formation under UVA phototoxicity of 6-TG. Suchelimination of the UVA photooxidation of 6-TG has crucial biological and medicinalapplications. UVA takes up to 95% of the UV radiation in sunlight, and since 6-TG is a UVAchromophore, patients who have to use 6-TG drug molecules for long-term treatment havehigh chance of skin sensitivity and cancer risks. The application of AuNPs as a potentialdrug carrier for 6-TG can efficiently prohibit 6-TG phototoxicity under UVA excitation andthus eradicate the side effects of 6-TG drug treatment. Furthermore, AuNPs mightpotentially become a useful application in the removal of side effects for the class ofthiobase drugs.