br temperature of solutions with different concentrations in
temperature of solutions with different concentrations increased to 26.7, 34.6, 41.8, and 49.7 °C, respectively. In order to evaluate the photothermal stability of BPNs-PDA-PEG-PEITC, the 100 μg/mL solu-tion was irradiated with 808 nm NIR laser for 5 min. After turning off the NIR laser, the solution was naturally cooling to room temperature. The heating-cooling test was repeated for five times, the variation of temperature was shown in Fig. 2d. In addition, the BPNs-PDA-PEG-PEITC which stored at 4 °C for 7 days presents the similar temperature rise when put in the same exposure conditions (Fig. 2e) suggesting BPNs-PDA-PEG-PEITC with good photothermal stability. Besides, a vital parameter to evaluate photothermal capability was photothermal con-version efficiency (η). Based on the calculation formula reported by previous literature (See Calculation Formula in Supporting informa-tion), τ can be got through linear relationship between cooling time and −ln(θ) (Fig. 2f) and the photothermal conversion efficiency was calculated as 34.9%. Compared with other PTT materials, such as the η
2.3. ROS characterization
The photodynamic activity of BPNs-PDA-PEG-PEITC was measured using electron spin resonance (ESR) spectroscopy which could detect the generation of ROS. Singlet oxygen (1O2) is a type of ROS. 2,2,6,6-tetramethylpiperidine (TEMP), a 1O2 trapping agent, was chosen to verify the generation of 1O2 by BPNs-PDA-PEG-PEITC through ESR spectroscopy under light irradiation (λ ≥ 600 nm) . The 1:1:1 tri-plet signal characteristic with a hyperfine splitting constant (aN = 16.2 G) in the ESR spectroscopy (Fig. 2g) for the BPNs-PDA-PEG-PEITC were in accordance with the characteristics of 2,2,6,6-
tetramethylpiperidine-N-oxyl (TEMPO). The signal characteristics dis-appeared in the absence of BPNs-PDA-PEG-PEITC (blank) . The generated 1O 2 could be consumed by sodium azide (NaN3). After adding NaN3, the characteristics of TEMPO didn’t appear which was consistent with blank confirming the generation of 1O2 was from BPNs-PDA-PEG-PEITC. 1,3-diphenylisobenzofuran (DPBF), a probe molecule, could react with 1O2 presenting a reduced Lycopene intensity at 410 nm which was used to further evaluate the generation of 1O2. It could be found that an obvious absorption reduction when the BPNs-PDA-PEG-PEITC was irradiated at 660 nm laser (Fig. 2h and i) sug-gesting that the photodynamic activity of BPNs-PDA-PEG-PEITC. In the absence of BPNs-PDA-PEG-PEITC, there was no obvious change of ab-sorption intensity at 410 nm (Fig. 2i) under the irradiation at 660 nm laser. The fluorescent dye 2,7-dichlorodihydrofluorescein diacetate (H2DCFHDA), a ROS probe, could generate green fluorescence after being oxidized by ROS which was used to further prove intracellular ROS production under irradiation . As shown in Fig. S8, a very weak fluorescence signal was presented of the MCF-7/ADR incubated with BPNs-PDA-PEG-PEITC without irradiation. In contrast, the cells after irradiated with 660 nm light exhibited a strong green fluores-cence, demonstrating the intracellular production of ROS.
2.4. The hemocompatibility of samples
The bioapplication of nanocarrier will be finally performed through intravenous injection, so it is essential to evaluate the hemocompat-ibility of the nanocarrier. After red blood cells (RBCs) interacted with nanomaterials, its morphological change is a vital factor to evaluate the hemocompatibility of nanomaterials for bioapplication. RBCs presents fragmentation or aggregation indicating that incompatible nanoma-terials interacted with RBCs. As shown in Fig. 3a, the RBCs interacted
Fig. 1. (a–d) TEM images of the BPNs, BPNs-PDA, BPNs-PDA-PEG, and BPNs-PDA-PEG-PEITC. (e) Size distribution of BPNs-PDA-PEG-PEITC. (f) FI-IR spectra of BPNs, BPNs-PDA, BPNs-PDA-PEG, and BPNs-PDA-PEG-PEITC. (g) Scanning TEM and EDS elemental mappings (P, N, and S, respectively) pictures of BPNs-PDA-PEG-PEITC. (h) XPS spectra of BPNs and BPNs-PDA-PEG-PEITC.
with pure PBS (negative control) which presented a normal circular shape. In contrast, the RBCs contacted with water (positive control) showed an obvious morphological destruction (Fig. S9). The micro-graphs of RBCs incubated with BPNs-PDA-PEG-PEITC with different concentrations (Fig. 3b–f) retained the same circular morphology, in-dicating the samples have no influence on RBCs shape. Hemolysis rate, a vital parameter, could be also used to evaluate the hemocompatibility of biomaterials. Hemolysis will lead to hemoglobin-release which causes the thrombosis. Hemolysis rate < 5% is permissible for bioma-terials [60,61]. BPNs-PDA-PEG-PEITC at different concentrations pre-sented hemolysis rate from 0.45% to 0.81% (Fig. 3g), even though at a high concentration the sample still had a mild influence on hemolysis. Activated partial thromboplastin time (APTT), thrombin time (TT), and prothrombin time (PT) are commonly used to measure the clotting time of materials . The APTT, TT, and PT data of BPNs-PDA-PEG-PEITC (Fig. 3h–j) at the concentration range of 100, 200, 400, 800 and 1600 µg/mL was close to the time of control groups indicating the ex-cellent hemocompatibility of BPNs-PDA-PEG-PEITC. These results de-monstrated the BPNs-PDA-PEG-PEITC could be injected via vein. In addition, the hemocompatibility of BPNs-PDA-PEG-PEITC/DOX was also estimated. Because free DOX can cause hemolysis , BPNs-PDA-PEG-PEITC/DOX showed a higher hemolysis than BPNs-PDA-PEG-