On the impurities content in nanodiamonds evolve for

       On increasing the development of
nanomaterials for applications, their toxicology becomes of ever-increasing important. In addition, the
traditional factors govern the toxicological profile
of bulk materials, moving from micro to the nano-scale adds more dimensions: Apart
from elemental composition, potential toxicity of nanomaterials may depend on
their size, shape, and dispersion state. Traditional purification
techniques depict to produce highly pure large crystals or particles of bulk materials often fail for nanomaterials. In
addition, when the surface to volume ratio becomes larger, the contribution of
any contaminants that usually reside on the surface of particles becomes more noticable.
Therefore, control of purity and dispersion are no less important than
synthetic efforts pointed to develop the nanomedicine platforms and bringing
them to applications133. The
majority of impurities in detonation nanodiamond (DND) arise
from the material of the blast chamber, charge suspension device, and initiator
(usually Pb, Cu or Ag azide).
Carried out in a steel blast chamber, the explosion formed the ND-containing
detonation soot and originate metallic contaminations along with non-diamond
carbon. Hence, for most of its applications particullarly, in biology and medicine, DNDs
should be purified post-synthesis. A lots of
techniques has been proposed to purify NDs. Some of them could be used for NDs
of different origins, whether it is detonation DND
or high-temperature high-pressure ND (HPHTND)6,65. A liquid
phase oxidative treatment (with mineral acids, e.g. sulfuric,
nitric, and perchloric acids or mixtures there of)
seperate almost all graphitic and metal impurities. Nevertheless, some
non-carbon functional groups, e.g., sulfonic, are found as a result of liquid
oxidation. In contrast, gas phase oxidation techniques produce only oxygen-rich
carbon functional groups on the nanodiamonds  surface, although these methods needed
additional treatment in dilute acids to
remove metal impurities . Microwave-assisted liquid oxidation requires a lower temperature as compared to traditional liquid
oxidation, although the complete removal of metals is attain when EDTA
complexes are used. The nanodiamonds show no innate toxicity but may show toxicity that depends on the tailorable
surface properties of the material, intensify the need for testing all surface
modified nanodiamonds for their toxicity/biocompatibility.
Thus, it is critical to examine the impurities content in nanodiamonds evolve
for applications, although many researchers tacitly rely upon almost complete
removal of metal and non-diamond carbon contaminations after air and liquid
oxidation or liquid oxidation alone24,134. The
techniques for identifying the nanodiamonds purity and content of contaminants
include inductively-coupled plasma mass-spectrometry and elemental analysis,
SEM/TEM, XRD, XPS, and Raman spectroscopy1,2,4,135,136.

       In most of cases, one method is not enough
(for example, many manufacturers give the C:N:O content but
no phase composition of carbon, which is necessary to assess the content of
diamond and non-diamond phases in the material),
and complete characterization needs a combination of several techniques. The
notoriously strong aggregation of DNDs strictly limits their potential in many applications, involving
polymer-and metal-matrix compounds, as well as different applications. Since
the developing studies of nanodiamonds toxicity, various screenings have been execute to produce comprehensive toxicological profiles
of nanodiamonds (table 2).
These studies pointed to recognize the type of toxicity and the unexpressed
mechanisms. A general conclusion, which can be drawn from multiple toxicity
studies is that nanodiamonds derivatives from various origin and sizes do not ruin
the fundamental functions of cells, organs, and organisms in a acceptable range
of concentrations. At the same time,
there are various reports of nanodiamonds toxicity, which may be associated to
the use of poorly purified nanodiamonds. Table  5 outline the purification
protocols of nanodiamonds and the detailed contents of impurities (when
provided) along with toxicological outcomes. In most cases where
significant level of toxicity was describe, nanodiamonds have been
used as-received. In those studies where nanodiamonds has been purified (for
example, air or ozone oxidized and acid treatment to dissolve metals or liquid oxidized)
no remarkable toxicity was noticed. We suppose that if metallic and other impurities
were fully removed, the nanodiamond would be shown low or no toxicity in all
studies134,137,138. For
example, only slight apoptosis of HaCaT cells effect the membrane permeability changes, caspase activation, and
release of intracellular lactatedehydrogenase,
was noticed as a result of exposure to non-purified
as-received ND at 100 ?g ml?1
concentration139. On the
other hand, purified nanodiamond in same concentration
did not affect basal cellular toxicity of A549 cells140. Another
reason for induced cell death may be associated to unreasonably high
nanodiamonds concentrations used in the tests. For example, an increased level
of apoptosis has been noticed in both normal and cancer cells at 200–1000
?g ml?1
nanodiamonds. At the same time, the nanodiamonds concentrations below than 50 ?g
ml?1
no apparent toxicity was observed141. Nanodiamonds
did not persuade any cytotoxicity and inflammation in concentrations up to 50 ?g
ml?1,
as shown through examination of gene expression mechanisms, cell morphology,
immunotoxicity, and apoptosis142 .
Analysis of other effects (size, shape, and origin of nanodiamonds)
have shown that the concentration played the most important role in nanodiamonds
induced toxicity and inflammation143.

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        The immunotoxicity, that results in an
increased secretion of chemokines and cytokines, has been estimate for nanodiamonds
derivatives with many cell types. After exposure to ND-COOH, themesenchymal
stem cells did not change the secretion of cytokines, chemokines, and growth
factors144. Cell
oxidative stress is another indicator of cellular toxicity. The oxidative
stress produce by nanodiamonds derivatives is cell specific.
For example, no signs of oxidative stress have been observed in neuroblastoma
cells, macrophages, keratinocytes, and PC-12 cells . On the other hand,
lymphocytes and endothelialcells have shown nanodiamonds induced oxidative
stress that can be partially associated to the use of as-received non-purified
nanodiamonds145. A concept
of adsorption, distribution, metabolism, and elimination (ADME)
is broadly used to analyse carbon nanomaterials bioavailability, tissue
distribution, metabolism, and excretion from the body . NDs labeled with 188Re,
125I, and 18F radionuclides have been used in biosafety
explorative studies on mice and rats. Alike to other nanomaterials, exposure
routes to nanodiamonds can impact toxicity146. Two
studies of pulmonary delivery of nanodiamonds report counter results that show
the both toxic and non-toxic properties of nanodiamonds in lungs and other
organs, together with cardiovascular system. This controversy can be detect
back to a dose-dependent nature of the noticed effects or to differences in
purity of nanodiamonds used. In another example, the controversial nanodiamonds
toxicity towards blood could be described by impurities and poor purification,
since the researchers reported rupture of the membranes of white and red blood
cells using non-purified as-received nanodiamonds101. On the
other hand, no signs of hematological toxicity were observed with acid purified
BASD detonation nanodiamonds. Substantial attention has been paid to the developmental toxicity of nanodiamonds. Classic
test systems using embryos of Xenopus laevis, and
Danio rerio (zebrafish) to determine teratogenic and
embryogenic potential of nanodiamonds have been reported147. Xenopus
embryos turned out to be sensitive to nanodiamonds surface functionalization,
which sometimes resulted in a low survival rate due to developmental
abnormalities. Dose-dependent toxicity of nanodiamonds was reported for the zebrafish model143. Particle
aggregation and inability of sub-vertebrate species to cleanse nanodiamonds
from their body at higher nanodiamonds concentrations can donate to the
reported nanodiamonds toxicity in these cases148. The fate
of nanodiamonds carbon in other organisms, and particularly, in animals raises
questions. Unlike metal nano-particles, nanodiamonds cannot be digested or
dissolved in the human body78. To
conclude, nanodiamonds is undoubtedly a perspective nanomaterial with less concerns regarding its toxicity as compared
to other carbon materials, as well as CTAB-coated gold nanoparticles, semiconductor quantum dots, etc. Some contradictions in the experimental results
noticed for nanodiamonds can be descibed by the impurities content, and thus a
proper purification of as-received nanodiamonds
is utterly important. The level of impurities as well as surface chemistry of nanodiamonds
must be taken into account during analysing the nanodiamonds toxicological profile.
In particular, heavy metals, graphitic and amorphous carbon, ceramic and other
potentially harmful impurities have to be removed14.

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