Bifunctional TiO₂/Ag₃PO₄/graphene composites with superior visible light photocatalytic performance and synergistic inactivation of bacteria

Abstract

In this work, bifunctional TiO₂/Ag₃PO₄/graphene (GR) composites have been prepared via the combination of ion-exchange method and hydrothermal approach, and the fabrication of “pizza-like” three-phase TiO₂/Ag₃PO₄/GR composites has been achieved through the electrostatic-driven assembly of positively-charged Ag⁺ on negatively-charged graphene oxide (GO) sheets, followed by the nucleation & controlled growth of Ag₃PO₄ and the deposition of Degussa P25 on the GO surface. Consequently, the hydrothermal treatment leads to the generation of TiO₂/Ag₃PO₄/GR composites with well-defined structures. The as-prepared composites exhibited highly efficient visible light photocatalytic activity toward organic dye molecule degradation and showed excellent bactericidal performance. This is the first report on the production of bifunctional three-phase metal oxide–Ag₃PO₄–GR composite materials with improved photocatalytic and antibacterial properties. The improved photocatalytic activity is attributed to the effective separation of photoexcited electron–hole pairs and fast charge transfer between components in the composite, while its excellent bactericidal performance is believed to come from intrinsic bacterial inactivation of Ag₃PO₄ and photo-induced antibacterial activity of active oxygen-containing radicals generated in the irradiated system. The proper molar ratio of Ag₃PO₄/TiO₂ and the added amount of GO in the precursor have been considered to play crucial roles in the formation of bifunctional composites with promising properties. The TiO₂/Ag₃PO₄/GR composite significantly decreases the percentage of expensive Ag-containing material while it reveals better photocatalytic and antibacterial performance than Ag₃PO₄, providing new insights into the low-cost, large-scale production of Ag₃PO₄-based function materials for practical applications.

---

1. Introduction

The past few decades have witnessed increasing environmental pollution, and exposures to environmental pollution remain a major source of health risk throughout the world. However, environmental pollution cannot be simply generated, and pollutants take many forms. They include not only inorganic and organic chemicals, but also bacteria and organisms. Despite the major efforts that have been made over the past few years to clean the environment, the search for new materials and technology to meet present demands in the removal and sterilization of a wide range of pollutants in water becomes even more urgent. Photocatalysis is an effective, economical and environment-friendly photooxidation process where the produced active oxygen-containing radicals are widely used to remove the contaminants by converting them to CO₂, H₂O,^1–5 etc. Recent advances in new photocatalytic materials and nanotechnology have demonstrated that highly efficient solar photocatalytic degradation of organic pollutants and bacteria in the presence of photocatalysts can be achieved. As a result, the design of highly active visible-light-responsive photocatalytic materials has attracted great attention and a range of visible light photocatalysts have been developed for the environmental decontamination.^6–13

Most recently, the pioneering work reported by Ye and co-workers has shown that silver orthophosphate (Ag₃PO₄) can achieve a quantum efficiency of nearly 90% for O₂ evolution from water under visible light irradiation.¹⁴ Moreover, it has been reported that Ag₃PO₄ demonstrates significant visible-light-driven photocatalytic activity in the degradation of organic pollutants in aqueous solution.¹⁵ Compared with several currently known visible light photocatalysts including doped TiO₂, BiVO₄, and AgX (X = Cl, Br), Ag₃PO₄ has a significantly higher photocatalytic efficiency and has been considered to be a promising candidate for practical applications due to its superior photooxidative capabilities by utilizing abundant solar light. However, several limitations of the Ag₃PO₄ photocatalytic system may restrict its practical use in energy and environmental sciences. Firstly, the Ag₃PO₄ photocatalyst is unstable upon photo-illumination and it is prone to be photoreduced and decomposed to weakly active Ag. The presence of generated black metallic Ag particles in the photocatalytic system would inevitably prevent visible light absorption and decrease its photocatalytic activity. Secondly, the use of a large amount of expensive silver-containing raw material in the present photocatalytic system strongly limits its large-scale production and practical application. Furthermore, Ag₃PO₄ normally possesses irregular microstructures and is insoluble in most solvents. The morphology, particle size, and specific highly reactive facet have been proved to play major roles in its highly efficient photocatalytic activity. Most recently, considerable efforts have been made to synthesize Ag₃PO₄ photocatalysts with well-defined morphologies including cubic Ag₃PO₄ microcrystals,¹⁶ hierarchical Ag₃PO₄ porous microcubes,¹⁷ dendritic Ag₃PO₄,¹⁸ concave trisoctahedral Ag₃PO₄ microcrystals,¹⁹ and Ag₃PO₄ tetrapods²⁰ and to design Ag₃PO₄-based composite photocatalysts by the combination of Ag₃PO₄ with different materials including metal oxides (TiO₂,^21–23 Fe₃O₄,²⁴ SnO₂ (ref. 25)), Ag,^26–30 AgX (X = Cl, Br and I),^31–33 carbon materials such as graphene oxide,^34–36 carbon quantum dots³⁷ and hydroxyapatite.^38,39 Despite tremendous efforts, it is still urgent and highly desirable to develop a facile and low-cost process for the large-scale production of Ag₃PO₄-based photocatalysts with enhanced stability and highly efficient photocatalytic activity.

Degussa P25, a commercially available TiO₂ nanomaterial, consists of two crystal forms of approximately 20% rutile and 80% anatase and has been widely used in photocatalytic studies due to its low cost, photocatalytic activity, stability and innocuousness. However, its visible-light-driven photocatalytic efficiency is generally low due to its wide band gap and the fact that it only absorbs lights in the ultraviolet region. Yao and co-workers reported the synthesis of heterostructured Ag₃PO₄/TiO₂ photocatalysts by the deposition of Ag₃PO₄ nanoparticles onto the P25 surface via an in situ precipitation method,²¹ and the obtained composite photocatalyst showed significantly improved photocatalytic degradation of organic dyes compared to that of pure Ag₃PO₄ and TiO₂. Better structural stability and recyclability of the photocatalyst under visible light irradiation were also observed.

Chemically derived graphene oxide (GO) with oxygen-containing functional groups has been proven to be a promising candidate for the construction of composite photocatalysts due to its solubility in solvents and its negatively-charged active sites on its high-surface-area sheets. Our previous report and other groups' work^34–36,40 confirm that the hybridization of Ag₃PO₄ with GO sheets not only results in the enhancement in the visible light absorption, but also leads to an improved visible light photocatalytic performance since GO sheets could facilitate charge transfer and suppress the recombination of photo-generated electrons and holes in the photocatalytic system. Our study further demonstrates that the generation of the Ag₃PO₄/GR composite photocatalyst by the hydrothermal treatment of the Ag₃PO₄/GO composite causes an obvious increase in its visible light photocatalytic activity.⁴¹

In consideration of the facts that improved stability, enhanced photocatalytic performance and low-cost of Ag₃PO₄ materials are all important factors responsible for stable and highly efficient Ag₃PO₄-based visible light photocatalysts. Herein, for the first time, we develop an efficient strategy for the fabrication of TiO₂/Ag₃PO₄/GR composites where the presence of TiO₂ and GR in the composite may effectively reduce the cost for the preparation of the composite while the hybridization of photocatalytic P25 and Ag₃PO₄ on highly conductive GR sheets favors the separation of photo-induced electrons and holes as well as charge transfer in the three-phase composite photocatalyst. The morphology, size, and visible-light photocatalytic behavior of the composites are investigated together with their structural and physicochemical properties. Furthermore, the TiO₂/Ag₃PO₄/GR composites are believed to have intrinsic antibacterial activity and enhanced photo-induced inactivation of bacterial cells under visible light irradiation. To the best of our knowledge, this is the first report concerning the fabrication of bifunctional Ag₃PO₄-based composites with improved visible light photocatalytic performance and enhanced antibacterial activity. The investigation provides a low-cost and effective approach for the large-scale production of Ag₃PO₄-based functional composite materials for the efficient removal of organic contaminants and bacterial inactivation under visible light irradiation.

2. Experimental section

2.1 Synthesis of samples

All reagents were purchased from Sinopharm Chemical Reagent Co. Ltd. (Shanghai, China) and used as received without further purification. Graphite oxide was synthesized from natural graphite by a modified Hummers' method with additional KMnO₄.^42,43 In a typical synthesis, the as-synthesized graphite oxide was first dispersed in distilled water, followed by ultrasonication for several hours to give GO dispersions. Then AgNO₃ solution was added into the above GO solution under magnetic stirring. After gentle stirring overnight, the ultrasonicated Degussa P25 aqueous dispersion was added dropwise into the Ag⁺–GO mixture, and the mixed solution was stirred for a further 30 min, followed by the addition of Na₂HPO₄ aqueous solution into the P25–Ag⁺–GO mixture. Upon the addition of Na₂HPO₄, yellowish-brown product precipitates were formed instantaneously. The reaction solution was then transferred into a 100 mL Teflon-lined stainless steel autoclave and kept in an oven at 180 °C for 24 h. The autoclave was left to cool naturally to room temperature; the obtained precipitate was collected by centrifugation, washed several times with deionized water and absolute ethanol, and dried at 60 °C under vacuum overnight. The reaction conditions for the preparation of hydrothermal composites are shown in Table 1, and the overall synthetic procedure for the generation of TiO₂/Ag₃PO₄/GR composites is illustrated in Scheme 1. Samples S1, S0.8, S0.6, S0.4, and S0.2 represent the samples prepared by using different molar ratios of Ag₃PO₄/TiO₂, while samples S-0, S-5, S-10, S-20, S-50, and S-100 stand for composites obtained in the presence of different GO amounts.

Table 1 Reaction conditions for the preparation of hydrothermal products

Sample	GO	AgNO3	P25	Na2HPO4·7H2O	M (Ag3PO4/TiO2)
S0	20 mg	9 mmol, 1.53 g		3 mmol, 0.804 g
S1	20 mg	9 mmol, 1.53 g	3 mmol, 0.24 g	3 mmol, 0.804 g	1
S0.8	20 mg	9 mmol, 1.53 g	3.75 mmol, 0.3 g	3 mmol, 0.804 g	0.8
S0.6	20 mg	9 mmol, 1.53 g	5 mmol, 0.4 g	3 mmol, 0.804 g	0.6
S0.4	20 mg	9 mmol, 1.53 g	7.5 mmol, 0.6 g	3 mmol, 0.804 g	0.4
S0.2	20 mg	9 mmol, 1.53 g	15 mmol, 1.2 g	3 mmol, 0.804 g	0.2
S-0		9 mmol, 1.53 g	3.75 mmol, 0.3 g	3 mmol, 0.804 g	0.8
S-5	5 mg	9 mmol, 1.53 g	3.75 mmol, 0.3 g	3 mmol, 0.804 g	0.8
S-10	10 mg	9 mmol, 1.53 g	3.75 mmol, 0.3 g	3 mmol, 0.804 g	0.8
S-20	20 mg	9 mmol, 1.53 g	3.75 mmol, 0.3 g	3 mmol, 0.804 g	0.8
S-50	50 mg	9 mmol, 1.53 g	3.75 mmol, 0.3 g	3 mmol, 0.804 g	0.8
S-100	100 mg	9 mmol, 1.53 g	3.75 mmol, 0.3 g	3 mmol, 0.804 g	0.8

---

Scheme 1 Schematic illustration of the formation of TiO₂/Ag₃PO₄/GR composites.

2.2 Characterization

The morphologies of the as-synthesized products were examined by field-emission scanning electron microscopy (FESEM, JEOL, JSM-7001F), transmission electron microscopy (TEM, JEOL, JEM-2100) and atomic force microscopy (AFM, MFP-3D SA). The phases of the obtained products were collected on a Bruker D8 Advance X-ray diffractometer (Cu Kα radiation, λ = 0.15406 Å) in a 2θ range from 10° to 80° at room temperature. Raman experiments were performed using a DXR spectrometer using the 532 nm laser line, and measurements were made in backscattering geometry. UV-visible diffuse reflectance spectra were recorded within a 200–800 nm wavelength range using a Shimadzu UV2450 spectrometer.

2.3 Photocatalytic experiments

The photocatalytic activities of products were valued by the decomposition of organic dyes under visible-light irradiation. The optical system for the photocatalytic reaction was composed of a 350 W Xe lamp and a cut-off filter (λ > 420 nm). Organic dye solutions (100 mL, 10⁻⁵ mol L⁻¹) containing 50 mg of samples were put in a sealed glass beaker, first ultrasonicated for 10 min and then stirred in the dark for 30 min to ensure absorption–desorption equilibrium. After visible light illumination, 4 mL of samples were taken out at regular time intervals (2 min) and separated through centrifugation (10 000 rpm, 10 min). The supernatants were analyzed by recording variations of the absorption band maximum in the UV-vis spectra of the dye molecule by using a Lambda 25 UV/Vis spectrophotometer.

2.4 Evaluation of antibacterial activities of samples

Bacteria were cultivated in nutrient broth at 37 °C for 18 h in a rotary shaker until reaching a stationary growth phase. The as-prepared cells were then resuspended and diluted to the required cell density of around 10⁷ colony-forming units per milliliter (CFU mL⁻¹) with sterilized saline solution (0.9% NaCl). The antibacterial activity of the composites was tested on six bacterial strains: Escherichia coli, Staphylococcus aureus, Salmonella typhi, Pseudomonas aeruginosa, Bacillus subtilis, and Bacillus pumilus from ATCC. All of the bactericidal experiments were performed at room temperature and repeated three times in order to give an average value; the measured data for each set of experiments were expressed with the mean and standard deviation.

The minimum inhibitory concentrations (MIC) of each of the composites were determined against all test strains. Various concentrations (6.25 ppm, 12.5 ppm, 25 ppm, 50 ppm, 100 ppm, and 200 ppm) of the composites were mixed with freshly sterilized (121 °C, 15 min) nutrient broth (tempered at 37 °C) in glass tubes (15 mm × 15 mm), and consequently 100 μL of pre-cultured strain (initial concentration 10⁶ CFU mL⁻¹) was added into each tube by micropipette. All the tubes were then placed in a precision constant temperature incubator at 35 °C for 48 h where a high-pressure mercury lamp (Philips HPR 125 W) was used as light source. MIC was determined as the lowest composite concentration that resulted in complete inhibition in nutrient broth. For the minimum bactericidal concentration (MBC), the mixtures of a series of concentrations (MIC, 2MIC, 4MIC, …) of composite dispersions with test strains were drawn with one loop full streak-inoculated to the nutrient agar plates. All the plates were then placed in the same incubator equipped with the same light source at 35 °C for 48 h. MBC was determined as the lowest composite concentration that appeared without any colonies observed on the plates.

In order to further investigate the effect of the antibacterial composite on the bacteria cells, the best sample S0.8 was chosen as the best antibacterial composite from the MIC and MBC results and the colonies were counted to determine the viable bacterial numbers after being incubated. In a typical process, PBS buffer was first prepared from a mixture of 0.2 M NaH₂PO₄ and 0.2 M Na₂HPO₄ aqueous solutions. S0.8 dispersion with the concentration of 200 ppm was then mixed with sterilized PBS solution, followed by the addition of pre-cultured strain to reach the cell concentration of 10⁶ CFU mL⁻¹. All the tubes were then incubated in a temperature-controlled rotary shaker at 20 °C for different times (0, 0.5 h, 1 h, 2 h, 4 h, and 8 h). For TEM characterizations of untreated and treated samples, 10 μL of each specimen dispersion was first loaded on TEM copper grids and then stained with tungstophosphoric acid aqueous solution. The air-dried copper grids were examined using the TEM (JEOL JEM-2100) as described earlier.

3. Results and discussion

The morphological features of TiO₂/Ag₃PO₄/GR composites were evaluated by SEM, TEM and HRTEM characterization and the results are shown in Fig. 1. SEM observations shown in Fig. 1a and b reveal the formation of pizza-like composite structures. The agglomeration of micro-sized Ag₃PO₄ particles and TiO₂ nanoparticles was observed on the surface of graphene sheets. Two distinct particles were found in the high-magnification SEM image of the composite shown in Fig. 1b. Larger particles ranging from 0.3 μm to 0.8 μm are assigned to as-prepared Ag₃PO₄ crystals and smaller nanoparticles are attributed to hydrothermally treated Degussa P25. Moreover, the EDX pattern of TiO₂/Ag₃PO₄/GR composites was also recorded and shown in Fig. 1b (inset), showing the presence of several signals from Ti, Ag, P, C and O. The corresponding TEM image of the TiO₂/Ag₃PO₄/GR composites was shown in Fig. 1c and S1† where the particle size characteristics of each component are clearly identified. Irregular micro-sized Ag₃PO₄ particles and smaller TiO₂ nanoparticles were found to be covered by a thin graphene sheet. In addition, a HRTEM image was recorded on the selected area of Fig. 1c, and the high-magnification observation shown in Fig. 1d and S1† clearly confirms the presence of thin and wrinkled graphene layers. The interplanar spacing of 0.35 nm was clearly determined, which corresponds to the (101) crystallographic plane of TiO₂, in good agreement with anatase TiO₂ (JCPDS no. 21-1272).

Fig. 1 SEM images (a, b, EDX inset) and TEM images (c, d, HRTEM inset) of as-prepared TiO₂/Ag₃PO₄/GR composites.

Fig. 2a shows the XRD pattern of as-synthesized TiO₂/Ag₃PO₄/GR composites. Two diffraction peaks marked by “•” can be readily indexed as the (101) and (211) planes of anatase (JCPDS no. 21-1272), and the rest of the diffraction peaks can be identified to the body-centered cubic phase of Ag₃PO₄ (JCPDS no. 06-0505). No obvious characteristic diffraction peaks of GR or GO are observed in the XRD pattern of TiO₂/Ag₃PO₄/GR composites. The phenomenon might be ascribed to low diffraction intensities of GR and GO compared to those of crystalline Ag₃PO₄ and TiO₂, as well as the tiny amount graphene oxide employed in the reactants. It is well known that Raman spectroscopy plays an important role in determining the detailed structure of graphitic materials. Thus, Raman spectra of GO, Ag₃PO₄, and as-synthesized TiO₂/Ag₃PO₄/GR composites were recorded. Two distinct bands at 1355 cm⁻¹ and 1595 cm⁻¹ are observed in the Raman spectrum of GO (Fig. S2a†), corresponding to the D and G bands of graphite, respectively. Several characteristic bands corresponding to the vibrations of Ag₃PO₄ (Fig. S2b†) appear in the region less than 1200 cm⁻¹, which can be assigned to different modes of Ag₃PO₄ sample including the external modes, the bending vibration of the tetrahedral PO₄ ionic group, and the symmetric stretch of P–O–P and O–P–O bonds. It can be seen from Fig. 2b that the spectrum of the composite also exhibits two peaks at around 1350 cm⁻¹ (D band) and 1590 cm⁻¹ (G band). Moreover, other peaks ranging from 100–1200 cm⁻¹ are believed to come from Ag₃PO₄ and TiO₂. Generally, the intensity ratio of the D and G bands (I_D/I_G) is applied to evaluate the disorder degree in the graphitic layers and average size of the sp² domains of the graphitic materials. The I_D/I_G value of GO is estimated to be about 0.92, while an increased I_D/I_G value ratio of 1.05 was observed in the TiO₂/Ag₃PO₄/GR composite, indicating that less defects formed in the graphitic layers, which suggests the reduction of GO to GR upon hydrothermal treatment in the composite.

Fig. 2 XRD pattern (a) and Raman spectra (b) of TiO₂/Ag₃PO₄/GR hybrids; UV-vis diffuse reflectance spectra of different photocatalysts (c) and different amounts of GO as precursors (d).

It is believed that visible light absorption properties of catalysts are crucial in determining their photocatalytic performance, especially for the photocatalytic pollutant or bacterial degradation under visible light irradiation. Thus diffuse reflectance spectra of different samples were recorded and are shown in Fig. 2c. It is notable that pure Ag₃PO₄ showed good absorbance in the whole region. The hybridization of Ag₃PO₄ with P25 (S-0) resulted in the obvious decrease in the absorbance, whereas the introduction of 20 mg GO in precursors (S0) caused a slight increase in the absorbance. The TiO₂/Ag₃PO₄/GR composite S0.8 where 20 mg GO is employed and the molar ratio of Ag₃PO₄/TiO₂ is 0.8 exhibited better absorbance than pure Ag₃PO₄ in the visible range (500–800 nm) and the composite S-0 in the whole UV-visible range. However, its absorbance is obviously lower than that of S0 sample where 20 mg GO is incorporated with Ag₃PO₄ in the absence of P25, implying that the introduction of GO into the composite definitely favors the enhanced visible light absorbance while the employment of TiO₂ has a negative effect on the visible light absorbance of the composite. It is also shown from Fig. 2d that the presence of different amounts of GO in the precursor affects the optical property of light absorption for the TiO₂/Ag₃PO₄/GR obviously. The added GO has been reduced to GR upon the hydrothermal treatment by losing the majority of functional groups, and the generated GR induces the increased light absorption intensity particularly in the visible region, as observed in all of six composites (S-0, S-5, S-10, S-20, S-50, S-100, and S-200) with different addition amounts of GO. The presence of GR in the composites leads to a continuous absorption band in the visible light region, which is in good agreement with the color of the samples.

The photocatalytic activities of TiO₂/Ag₃PO₄/GR composites were first evaluated by decomposing RhB under visible light irradiation. It is clearly shown from Fig. 3a that the sample S0 (Ag₃PO₄/GR composite) exhibits the lowest photocatalytic activity of less than 80% in 12 min. When 0.24 g TiO₂ was introduced into the Ag₃PO₄/GR composite, the corresponding three-phase composite S1 showed enhanced photocatalytic activities of more than 95% in 12 min. Further addition of TiO₂ leads to the decrease in the molar ratio of Ag₃PO₄/TiO₂ from 1 to 0.8, resulting in the generation of the TiO₂/Ag₃PO₄/GR composite (S0.8) with the highest photocatalytic activity of around 95% in 6 min and almost 100% in 10 min. However, TiO₂/Ag₃PO₄/GR composites with lower molar ratios of Ag₃PO₄/TiO₂ than 0.8 (S0.6, S0.4, S0.2) showed decreased photocatalytic efficiencies, and all the three-phase TiO₂/Ag₃PO₄/GR composites demonstrate higher photocatalytic performance than the two-phase Ag₃PO₄/GR composite (S0). The above results imply that the proper molar ratio of Ag₃PO₄/TiO₂ plays a crucial role in determining the photocatalytic activities of TiO₂/Ag₃PO₄/GR composites. Furthermore, when the optimal molar ratio of 0.8Ag₃PO₄/TiO₂ is fixed, the effect of added GO on photocatalytic activities of TiO₂/Ag₃PO₄/GR composites was investigated, and the results are shown in Fig. 3b. The Ag₃PO₄/TiO₂ composite (S-0) reveals the photocatalytic efficiency of around 75% in 12 min in the absence of GO. When 5 mg GO was introduced in the Ag₃PO₄/TiO₂ composite, the photocatalytic activity of the composite S-5 was increased to nearly 85% in 12 min, and a further increase in GO to 10 mg leads to an enhanced photocatalytic activity of more than 95% in 12 min. The highest photocatalytic efficiency of around almost 100% in 10 min was achieved when 20 mg GO was used. However, the addition of a higher amount than 20 mg GO (50 mg, 100 mg) causes a negative effect on the photocatalytic activity of the TiO₂/Ag₃PO₄/GR composite, and the photocatalytic efficiencies of two samples (S-50 and S-100) are even lower than that of the two-phase Ag₃PO₄/TiO₂ composite (S-0), implying the appropriate added GO amount also affects the photocatalytic performance of TiO₂/Ag₃PO₄/GR composites significantly and the best added GO amount in this study is 20 mg.

Fig. 3 Visible light photocatalytic activities of TiO₂/Ag₃PO₄/GR composites with different molar ratios (a) and different added amounts of GO (b) toward RhB; reactive species trapping experiments (c).

Furthermore, reactive species trapping experiments were performed to investigate active oxidizing species in the photocatalytic process where three different chemicals, p-benzoquinone (BZQ, a O₂⁻˙ radical scavenger), disodium ethylenediaminetetraacetate (Na₂EDTA, a hole scavenger) and tert-butanol (a ˙OH radical scavenger), were employed. The experimental results (Fig. 3c) indicated that when 5 mM Na₂–EDTA as well as 5 mM BZQ was introduced to the above photocatalytic system, the photocatalytic activity of the hybrids was intensively suppressed with the degradation efficiency, decreasing from 100% to 20% in 10 minutes. Moreover, the presence of tert-butanol in the TiO₂/Ag₃PO₄/GR photocatalytic system showed negligible effect on its excellent photocatalytic activity. The reactive species trapping results indicate that the photo-induced active holes and superoxide ions make major contributions to the highly efficient photocatalytic performance. In addition to the photocatalytic performance toward RhB, two organic dyes MB and MO were also chosen as model pollutants to evaluate the photocatalytic efficiency of the as-prepared TiO₂/Ag₃PO₄/GR photocatalyst, and the result is shown in Fig. S3.† It can be clearly seen that, when MB was employed, a relatively higher photocatalytic efficiency was observed, while an obvious lower photocatalytic activity was achieved once MO was introduced. It is suggested that the removal of the majority of organic pollutants RhB or MB under visible light irradiation was obtained in 6 min, and a longer irradiation time contributes less to its photocatalytic efficiency. However, the elimination of organic pollutant MO was found to occur gradually at regular intervals of 2 min, achieving a photocatalytic efficiency of around 60% in 6 min and almost 100% in 12 min.

It is well known that Ag-based materials are effective biocides against numerous kinds of bacteria and fungi, and besides, photo-generated oxidative radicals from irradiated TiO₂-based materials are capable of inhibiting the growth of bacteria in the photocatalytic process. Thus, it is reasonable to assume that the obtained TiO₂/Ag₃PO₄/GR composite could be a promising candidate for the disinfection of water by synergistic effects from photocatalytic inactivation of microorganisms and direct bacterial inhibition/killing. For the first time, the intrinsic antibacterial and photocatalytic disinfection of different composite materials were investigated, and the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of different samples under light irradiation are shown in Table 2. It is clearly shown that six samples with different molar ratios (Ag₃PO₄/TiO₂) all exhibited excellent bacterial inhibition activities against four different bacteria. MIC values against S. aureus, S. typhi and P. aeruginosa are observed to be lower than 100 ppm, while a relatively higher value of 100 ppm is found against E. coli. The bactericidal activities of the above samples are further confirmed by MBC test results; notably, the majority of bacteria can be completely killed in a concentration equivalent to the MIC value, implying that the bacteria can be killed simultaneously soon after the growth has been inhibited. When the amount of GO was changed in the precursor, where the molar ratio of Ag₃PO₄/TiO₂ was kept at 0.8, six samples still demonstrate excellent antibacterial activities against four bacteria. Similar lower MIC and MBC values against three bacteria, S. aureus, S. typhi and P. aeruginosa, were obtained while E. coli is still an exception. With the increase in the added amount of GO in the precursor from 0 to 5 mg, 10 mg, and 20 mg, the MIC or MBC of the corresponding sample against three bacteria was observed to be further decreased to a lower value, indicating that the introduction of an increased amount of GO causes enhanced bacterial inhibition or bactericidal activity of the sample. However, when more than 20 mg GO was employed in the precursor, as-prepared TiO₂/Ag₃PO₄/GR composites exhibit slightly poorer bacterial inhibition or bactericidal activity toward three bacteria, implying that the presence of a higher percentage of GR reduced from GO upon hydrothermal treatment has a negative effect on the antibacterial activity of the TiO₂/Ag₃PO₄/GR composite. It is generally accepted that the direct bacterial activity of Ag₃PO₄ should result from dissolved Ag⁺, and the photocatalytic inactivation of bacteria also makes a major contribution to excellent antibacterial performance of TiO₂/Ag₃PO₄/GR composites. Under visible light irradiation, the synergistic effects from Ag⁺ and photo-induced oxidative species lead to total and efficient bacterial removal. For the TiO₂/Ag₃PO₄/GR samples prepared from precursors with different amounts of GO, it is proposed that the proper addition of GO in the precursor may improve the solubility/dispersibility of the sample, which favors membrane penetration of the antibacterial composite into the host cell and leads to inactivation of bacteria more efficiently. However, a further increase in the added amount of GO in the precursor results in a higher percentage of reduced GR in the TiO₂/Ag₃PO₄/GR composite, due to the fact that GR only presents limited antibacterial activity. Consequently, the TiO₂/Ag₃PO₄/GR composite with a higher percentage of GR demonstrates decreased bacterial inhibition or bactericidal effects.

Table 2 MIC and MBC results for as-prepared composites

Sample	MIC (MBC)
	E. coli	S. aureus	S. typhi	P. aeruginosa
S0	100 (100)	50 (50)	6.25 (6.25)	6.25 (12.5)
S1	100 (100)	12.5 (25)	6.25 (12.5)	12.5 (12.5)
S0.8	100 (100)	6.25 (12.5)	6.25 (12.5)	12.5 (12.5)
S0.6	100 (100)	12.5 (25)	12.5 (12.5)	12.5 (25)
S0.4	100 (100)	25 (50)	25 (25)	12.5 (50)
S0.2	100 (100)	50 (50)	50 (50)	25 (50)
S-0	100 (100)	50 (100)	25 (50)	25 (25)
S-5	100 (100)	50 (50)	25 (25)	12.5 (25)
S-10	100 (100)	25 (12.5)	12.5 (25)	12.5 (12.5)
S-20	100 (100)	6.25 (12.5)	6.25 (12.5)	12.5 (12.5)
S-50	100 (100)	25 (25)	25 (50)	25 (25)
S-100	100 (100)	25 (50)	50 (50)	25 (50)

---

On the basis of the above photocatalytic and antibacterial results, the TiO₂/Ag₃PO₄/GR sample S0.8 where the added GO was 20 mg and the molar ratio of Ag₃PO₄/TiO₂ was 0.8 has been evaluated as the best three-phase composite with highly efficient photocatalytic performance and excellent antibacterial activities. In order to further understand the synergistic effects of the TiO₂/Ag₃PO₄/GR composite S0.8, control experiments including MIC (MBC) of the composite S0.8 with E. coli in dark, E. coli under the light irradiation, and mixtures of S0.8 with E. coli under the light irradiation were conducted, and the results are shown in Table 3.

Table 3 MIC and MBC results of control experiments against bacteria

Sample	MIC (MBC)
	S0.8 with E. coli in dark	E. coli under the light	S0.8 with E. coli under the light
E. coli	>800 (>800)	>800 (>800)	100 (100)
S. aureus	>800 (>800)	>800 (>800)	6.25 (12.5)
S. typhi	>800 (>800)	>800 (>800)	6.25 (12.5)
P. aeruginosa	>800 (>800)	>800 (>800)	12.5 (12.5)

---

Furthermore, the determination of rapidity and bactericidal duration of the sample S0.8 has been assessed by time-kill analysis. It is shown from Fig. 4 that treatment with 200 ppm aqueous dispersion of S0.8 demonstrated a strong bactericidal effect on different kinds of bacteria. Considerably, within the first 2 h, the bacterial population was observed to decrease dramatically from above 6–6.5 log CFU mL⁻¹ of the control to 2.1–2.4 log CFU mL⁻¹ for S. aureus and B. subtilis, and around 1.0 log CFU mL⁻¹ for P. aeruginosa and B. pumilus. When the time was prolonged to 4 h, bacterial counts continued to decrease and the number of cells of all bacteria fluctuated around 1 log CFU mL⁻¹. After 8 h, the bacterial population was completely inactivated, and the data are not listed in Fig. 4.

Fig. 4 Time-kill analysis of the composite S0.8 against different bacteria.

Furthermore, TEM characterization was used to investigate the morphological changes of typical Gram-negative bacteria E. coli and Gram-positive bacteria S. aureus cells before and after the disinfection reaction. As shown in Fig. 5a and c, the E. coli and S. aureus cells remained in a good state in the absence of the antibacterial sample S0.8. However, when the composite was introduced into the system and the composite–bacteria mixture was incubated, it is evident from Fig. 5b and d that obvious cell damages or membrane deformation could be observed for both bacteria, in which regular rod-like or spherical cellular shapes, as well as bigger cell sizes are found to disappear. TEM observations imply that the TiO₂/Ag₃PO₄/GR composite may react with membrane proteins to damage the cell membrane, resulting in the inactivation of its relative function. It is suggested that the addition of GO in the precursor improved the solubility of the composite in antibacterial experiments, and the high-surface-area GR sheet reduced from GO could adsorb and gather the bacteria onto its surface, resulting in enhanced interactions between bacteria and active bactericidal components on GR sheets.

Fig. 5 TEM images of the untreated (a and c) and treated E. coli and S. aureus cells (b and d).

It is reasonably expected that the as-synthesized three-phase TiO₂/Ag₃PO₄/GR composite S0.8 displays highly efficient visible-light-driven photocatalytic activity and demonstrates excellent bactericidal performance against different kinds of bacteria based on the above results and discussion. Due to the presence of graphene and heterogeneous structures, it is necessary to understand the possible mechanism for the photocatalytic decoloration of organic dyes and inactivation of bacteria. As a result, a schematic diagram of the charge separation and transfer in the TiO₂/Ag₃PO₄/GR composite under visible light irradiation is proposed and is shown in Scheme 2. Generally, several factors are responsible for the enhanced photocatalytic decoloration and bactericidal performance. Firstly, the introduction of GO into the precursor plays an important structure-directing role in the generation of well-defined “pizza-like” TiO₂/Ag₃PO₄/GR composites. The presence of negatively-charged GO sheets may absorb positively-charged Ag⁺ on the large-surface-area GO sheets by the electrostatically-driven assembly. Due to the nature of TiO₂ in a dispersion pH greater than 6, when Degussa P25 is added into the Ag⁺–GO mixture, ultrasonicated TiO₂ nanoparticles can be adsorbed onto the Ag⁺ surface. The subsequent addition of PO₄³⁻ results in the formation and the controlled growth of Ag₃PO₄ on the GO surface, suggesting the construction of well-defined three-phase TiO₂/Ag₃PO₄/GO structures. The following hydrothermal treatment at 180 °C for 24 h caused little effect on the structure of the composite where the reduction of GO to GR occurs. Compared with pure Ag₃PO₄, TiO₂ and two-phase TiO₂/Ag₃PO₄ composite, as-prepared TiO₂/Ag₃PO₄/GR composites demonstrate better solubility/dispersibility, which makes a primary contribution to the irradiated photocatalytic process and antibacterial experiments under visible light. Secondly, either GO or GR has a large surface area, and the TiO₂/Ag₃PO₄/GR composite exhibits higher adsorption capacity of organic dyes than Ag₃PO₄, TiO₂ and two-phase TiO₂/Ag₃PO₄ composite. Moreover, the presence of black GR in the TiO₂/Ag₃PO₄/GR composite enhances the absorbance in the visible light region as shown in Fig. 2d. Both the improved adsorption of pollutant/bacteria and effective visible light utilization are helpful for the enhanced photocatalytic performance. Furthermore, most importantly, the formation of the heterostructures is believed to play key roles in the highly efficient photocatalytic performance and antibacterial activities. It is well-known that an effective charge separation/transfer is crucial for the enhancement in photocatalytic activities. For the TiO₂/Ag₃PO₄/GR composite, potentials of both the conduction band and valence band of TiO₂ are more negative than those of Ag₃PO₄ (conduction band: 0.45 eV, valence band: 2.45 eV). Under visible light irradiation, TiO₂ nanoparticles possess a large band gap of 3.0 eV that cannot absorb visible light under the present conditions with filtered λ > 420 nm, while the valence band (VB) and the conduction band (CB) of Ag₃PO₄ can be separated easily. Due to the presence of conductive GR sheets, it can serve as an effective acceptor of the photoexcited electrons. Hence, the photogenerated CB electrons of Ag₃PO₄ can be transferred to GR sheets in the TiO₂/Ag₃PO₄/GR composite. The transportation and mobility of electrons on GR sheets is very rapid in the specific π-conjugated structure, and thus, the efficient electron transfer from Ag₃PO₄ to GR sheets keeps electrons away from the Ag₃PO₄. More photo-generated electrons and holes are produced by continuously working in this way, effectively suppressing the charge recombination and improving the photocatalytic activity. Meanwhile, well-separated electrons in GR sheets can be trapped by the absorbed oxygen in GR surface to generate reactive oxygen species (ROSs), such as superoxide anions (O₂⁻), and the produced active radical species can decompose dye molecules into CO₂, H₂O, etc. and attack specific bacteria. In addition, the photo-induced holes on the surface Ag₃PO₄ particles may significantly accelerate the photocatalytic degradation of organic dyes or bacteria. It is also confirmed from XRD patterns of different samples before and after recycled photocatalytic experiments in Fig. S3† that the presence of GR in the TiO₂/Ag₃PO₄/GR composite effectively protects Ag₃PO₄ from being decomposed into metallic Ag, suggesting better stability and recyclability of TiO₂/Ag₃PO₄/GR composite in the photocatalytic process.

Scheme 2 Diagram of the mechanism of photocatalytic degradation of organic dye molecules and bacteria under visible light irradiation.

4. Conclusion

In summary, we have demonstrated an effective hydrothermal approach for the fabrication of TiO₂/Ag₃PO₄/GR composites. The TiO₂/Ag₃PO₄/GR composite shows highly efficient photocatalytic degradation activity toward organic dye molecules and also exhibits excellent antibacterial activity against common bacteria. Bifunctional TiO₂/Ag₃PO₄/GR composites illustrate improved visible light photocatalytic performance and enhanced antibacterial activity compared with bare Ag₃PO₄, TiO₂ and two-phase composites, due to the generation of composite materials. By adjusting the molar ratio of Ag₃PO₄/TiO₂ and the added amount of GO, the photocatalytic and antibacterial activities of the composites can be regulated. As a result, this novel bifunctional TiO₂/Ag₃PO₄/GR composite may find promising applications in environmental protection and water disinfection.

Acknowledgements

This work was financially supported by the National Natural Science Foundation of China (51102116, 11102075, 51302112), Natural Science Foundation of Jiangsu (BK2011480, BK2011534) and Open Fund of Key Laboratory for Intelligent Nano Materials and Devices of the Ministry of Education (INMD-2014M02).

References

1. J. L. Wang and L. J. Xu, Crit. Rev. Environ. Sci. Technol., 2012, 42, 251.
2. J. C. Sin, S. M. Lam, A. R. Mohamed and K. T. Lee, Int. J. Photoenergy, 2012, 2012, 185159,  DOI:10.1155/2012/185159.
3. C. P. Silva, M. Otero and V. Esteves, Environ. Pollut., 2012, 165, 38.
4. B. Limburg, E. Bouwman and S. Bonnet, Coord. Chem. Rev., 2012, 256, 1451.
5. S. M. Lam, J. C. Sin, A. Z. Abdullah and A. R. Mohamed, Desalin. Water Treat., 2012, 41, 131.
6. H. Tong, S. X. Ouyang, Y. P. Bi, N. Umezawa, M. Oshikiri and J. H. Ye, Adv. Mater., 2012, 24, 229.
7. A. Kubacka, M. Fernandez-Garcia and G. Colon, Chem. Rev., 2012, 112, 1555.
8. Q. J. Xiang, J. G. Yu and M. Jaroniec, Chem. Soc. Rev., 2012, 41, 782.
9. L. W. Zhang and Y. F. Zhu, Catal. Sci. Technol., 2012, 2, 694.
10. D. H. Pei and J. F. Luan, Int. J. Photoenergy, 2012, 2012, 262831,  DOI:10.1155/2012/185159.
11. A. Di Paola, E. Garcia-Lopez, G. Marci and L. Palmisano, J. Hazard. Mater., 2012, 211, 3.
12. E. Casbeer, V. K. Sharma and X. Z. Li, Sep. Purif. Technol., 2012, 87, 1.
13. X. Q. An and J. C. Yu, RSC Adv., 2011, 1, 1426.
14. Z. G. Yi, J. H. Ye, N. Kikugawa, T. Kako, S. X. Ouyang, H. Stuart-Williams, H. Yang, J. Y. Cao, W. J. Luo, Z. S. Li, Y. Liu and R. L. Withers, Nat. Mater., 2010, 9, 559.
15. Y. Bi, S. Ouyang, N. Umezawa, J. Cao and J. Ye, J. Am. Chem. Soc., 2011, 133, 6490.
16. Y. P. Bi, H. Y. Hu, S. X. Ouyang, G. X. Lu, J. Y. Cao and J. H. Ye, Chem. Commun., 2012, 48, 3748.
17. Q. H. Liang, W. J. Ma, Y. Shi, Z. Li and X. M. Yang, CrystEngComm, 2012, 14, 2966.
18. Y. P. Bi, H. Y. Hu, Z. B. Jiao, H. C. Yu, G. X. Lu and J. H. Ye, Phys. Chem. Chem. Phys., 2012, 14, 14486.
19. Z. B. Jiao, Y. Zhang, H. C. Yu, G. X. Lu, J. H. Ye and Y. P. Bi, Chem. Commun., 2013, 49, 636.
20. J. Wang, F. Teng, M. D. Chen, J. J. Xu, Y. Q. Song and X. L. Zhou, CrystEngComm, 2013, 15, 39.
21. W. F. Yao, B. Zhang, C. P. Huang, C. Ma, X. L. Song and Q. J. Xu, J. Mater. Chem., 2012, 22, 4050.
22. R. Y. Liu, P. G. Hu and S. W. Chen, Appl. Surf. Sci., 2012, 258, 9805.
23. S. B. Rawal, S. D. Sung and W. I. Lee, Catal. Commun., 2012, 17, 131.
24. G. P. Li and L. Q. Mao, RSC Adv., 2012, 2, 5108.
25. L. L. Zhang, H. C. Zhang, H. Huang, Y. Liu and Z. H. Kang, New J. Chem., 2012, 36, 1541.
26. Y. P. Liu, L. Fang, H. D. Lu, L. J. Liu, H. Wang and C. Z. Hu, Catal. Commun., 2012, 17, 200.
27. Y. P. Liu, L. Fang, H. D. Lu, Y. W. Li, C. Z. Hu and H. G. Yu, Appl. Catal., B, 2012, 115, 245.
28. Y. P. Bi, H. Y. Hu, S. X. Ouyang, Z. B. Jiao, G. X. Lu and J. H. Ye, J. Mater. Chem., 2012, 22, 14847.
29. Y. P. Bi, H. Y. Hu, S. X. Ouyang, Z. B. Jiao, G. X. Lu and J. H. Ye, Chem.–Eur. J., 2012, 18, 14272.
30. W. Teng, X. Y. Li, Q. D. Zhao, J. J. Zhao and D. K. Zhang, Appl. Catal., B, 2012, 125, 538.
31. Y. Hou, F. Zuo, Q. Ma, C. Wang, L. Bartels and P. Y. Feng, J. Phys. Chem. C, 2012, 116, 20132.
32. J. Cao, B. D. Luo, H. L. Lin, B. Y. Xu and S. F. Chen, J. Hazard. Mater., 2012, 217, 107.
33. Y. P. Bi, S. X. Ouyang, J. Y. Cao and J. H. Ye, Phys. Chem. Chem. Phys., 2011, 13, 10071.
34. Q. H. Liang, Y. Shi, W. J. Ma, Z. Li and X. M. Yang, Phys. Chem. Chem. Phys., 2012, 14, 15657.
35. L. Liu, J. C. Liu and D. D. Sun, Catal. Sci. Technol., 2012, 2, 2525.
36. G. D. Chen, M. Sun, Q. Wei, Y. F. Zhang, B. C. Zhu and B. Du, J. Hazard. Mater., 2013, 244–245, 86.
37. H. C. Zhang, H. Huang, H. Ming, H. T. Li, L. L. Zhang, Y. Liu and Z. H. Kang, J. Mater. Chem., 2012, 22, 10501.
38. X. T. Hong, X. H. Wu, Q. Y. Zhang, M. F. Xiao, G. L. Yang, M. R. Qiu and G. C. Han, Appl. Surf. Sci., 2012, 258, 4801.
39. J. J. Buckley, A. F. Lee, L. Olivi and K. Wilson, J. Mater. Chem., 2010, 20, 8056.
40. H. Y. Cui, X. F. Yang, Q. X. Gao, H. Liu, Y. Li, H. Tang, R. X. Zhang, J. L. Qin and X. H. Yan, Mater. Lett., 2013, 93, 28.
41. X. F. Yang, H. Y. Cui, Y. Li, J. L. Qin, R. X. Zhang and H. Tang, ACS Catal., 2013, 3, 363.
42. D. C. Marcano, D. V. Kosynkin, J. M. Berlin, A. Sinitskii, Z. Z. Sun, A. Slesarev, L. B. Alemany, W. Lu and J. M. Tour, ACS Nano, 2010, 4, 4806.
43. X. F. Yang, H. Y. Ding, D. Zhang, X. H. Yan, C. Y. Lu, J. L. Qin, R. X. Zhang, H. Tang and H. J. Song, Cryst. Res. Technol., 2011, 11, 1195.

---

Footnote

† Electronic supplementary information (ESI) available. See DOI: 10.1039/c4ra01559b

---