3 Results and discussion
The novel RCNs were synthesized
by soft-template hydrothermal method. As indicated in the TEM image
(Figure 1 ), it was apparently demonstrated the products were
constituted principally by rod-shaped carbon particles with relatively
uniform dispersion and regular particle size. And the average size of
the carbon rods was measured to be approximately 85-120nm in length, and
35-50nm in width. This novel morphology might be originated from the
rodlike glucose-F127 monomicelles form, in which the polymerization,
dehydration and nucleation of glucose took place, and finally form the
rod-shaped nano structures.
The hydrodynamic diameter of RCNs was then measured by DLS. As shown inFigure S1, the size of
the particles was calculated to be 110nm, which was consistent with the
TEM results. Furthermore, the stability test of the RCNs was
characterized by DLS over 2 weeks, as shown inFigure S2 . It could be
seen that the size exhibited almost negligible changes, which proved
that the RCNs possess good stability in aqueous solution system.
The surface functional groups of carbon nanomaterials were further
analyzed by FT-IR. Figure 2 showed that the absorption peak at
3600-3200cm-1 was the stretching vibration of hydroxyl
O-H, the absorption peak at 3000-2800cm-1 was the
stretching vibration of saturated C-H, the absorption peak at
1770-1680cm-1 was the stretching vibration of ester
carbonyl C=O, and the absorption peak at 1250-1000cm-1was the stretching vibration of ester C-O.
The UV-vis-NIR spectra of the obtained RCNs in aqueous solution was
shown in Figure 3 , which
revealed that the samples possessed high absorption in the NIR region
around 650-900nm. The excellent NIR absorption characteristic of the
samples encouraged us to explore their photothermal properties. The RCNs
aqueous solution with different concentrations were irradiated under an
808nm laser at 2.5W/cm2 power density. As shown in theFigure 4 , under 808nm laser, the water temperature only
increased by 3°C within 5 min. And it could be seen that the temperature
of the RCNs solution increased more and more rapidly as the
concentration increased. When the concentration was 50μg/mL, the
temperature could raise to 45℃ in 5 min, while the concentration was
100μg/mL, it could raise to 54℃. The photothermal conversion efficiency
of RCNs was calculated to be about 23.7 % using the reported
literature. In the course of 6 cycles of photothermal heating and
natural cooling (Figure S3 ), the photothermal heating effect
did not change significantly, indicating that the RCNs had good
photothermal stability.
In order to improve the therapeutic effect of the nanomaterials toward
tumors, the nanocomposite with integrated thermo-chemotherapy
performance was fabricated, while the classic DOX was used as the model
anti-tumor drug. By the analysis of the DOX@RCNs, the drug loading rate
of DOX for the nanocomposites was calculated to be 17.4%. The
photothermal heating test was further arranged for the DOX@RCNs at the
concentration of 50μg/mL. It could be seen in the Figure 5 that
the temperature could raise to 39℃ in 5 min, and 52℃ in 10 min, which
showed excellent photothermal heating effect.
The drug release experiment under 808 nm laser radiation was explored,
it could be seen from Figure 6 that the release rate of DOX in
the composites could achieve at 34.6% in 60 min, after 2h, the rates
was calculated to be 64.7%. In the comparative experiments without
irradiation, the corresponding releasing rates were calculated to be
5.7% and 7.2%, respectively. It could be concluded that infrared
radiation can significantly increase the release of DOX, further realize
integrated treatment of hyperthermia and chemotherapy.
Considering that low toxicity is a necessary condition for nano-drugs,
the cell viability of MCF-7 cells incubated with different
concentrations of RCNs and DOX@RCNs were conducted. The cell viability
of MCF-7 cells incubated with two novel nano-drugs was shown inFigure 7 . It could be observed that the carbon nanomaterials
exhibit excellent biocompatibility, and have little cytotoxicity to
MCF-7 cells up to a concentration of 300μg/mL. For comparison, the
cytotoxicity for DOX@RCNs was slightly stronger. The cell survival rate
decreased gradually with the increasing of the concentration of the
sample, when the concentration was 300μg/mL, the cell survival rate was
78.6%.
Both the RCNs and DOX@RCNs were further explored their potential
antitumor activity under NIR irradiation against MCF-7 cells. It could
be seen that there was almost no cytotoxicity at the concentration of
50μg/mL for both RCNs and DOX@RCNs without NIR irradiation
(Figure 7 ). In contrast, under NIR irradiation, two novel
nano-drugs exhibited enhanced cytotoxicity against MCF-7 cells
(Figure S4 ). It should be noted that no obvious cell apoptosis
occurred after the MCF-7 cells were irradiated in the culture medium
without the nanomaterials. The relative cell viabilities were calculated
to be 50.2% at the RCNs concentration of 50μg/mL after 5 min of
continuous irradiation (2.5W/cm2), suggesting the
excellent photothermal toxicity effect of the RCNs. For the DOX@RCNs
samples, benefiting from the effective NIR-sensitive release of DOX,
DOX@RCNs exhibited stronger cytotoxicity than that of RCNs, while the
relative cell viabilities decreased to 23.6% at the same test
conditions.
In addition, confocal fluorescence microscopy was used to explore the
toxicity effects of the nanomaterials (Figure 8) . DAPI, which
could strongly bind to DNA, is a blue fluorescent dye used for
fluorescence microscopy observation of living and fixed cells. In
addition, fluorescein-dUTP is a green fluorescent dye which is used to
label apoptotic cells. In this research, in order to display the cell
apoptosis promoted by nanomaterials, DAPI/fluorescein-dUTP assays were
performed to intuitively observe live and apoptotic cells. After 4 min
irradiation, it could be seen that obvious apoptosis appeared in the
MCF-7 cells for the RCNs group compared with the control ones, however,
much more extensively apoptosis occurs for the DOX@RCNs group. This
phenomenon was consistent with the MTT results.