Pharmacology & Pharmacy, 2010, 1, 69-74
doi:10.4236/pp.2010.12010 Published Online October 2010 (
Copyright © 2010 SciRes. PP
The Study of Influence of Silica and Polyethilene
Glycols Organic-Inorganic Compounds on
Free-Radical Processes in Vitro
Olga G. Sitnikova1, Sergey B. Nazarov1, Irina V. Shikhanova2, Alexander V. Agafonov2,
Jean A. Dyuzhev1, Irina G. Popova1
1Ivanovo State Research Institute of Maternity and Childhood (RIMC), Ivanovo, Russia; 2Institute of Solution Chemistry, Russian
Academy of Sciences, Ivanovo, Russia.
Received July 20th, 2010; revised August 10th, 2010; accepted September 30th, 2010.
In this study investigation of influence of hybrid nanosilica-polyethylene glycols materials (molecular weight 1500,
6000 and 15000), prepared by sol-gel synthesis, on lipid peroxidation and antioxidant activity of human serum in vitro
was performed. Methods included chemiluminescence analysis and quantitative malonic dialdehyde estimation. It was
revealed that nanosilica-PEG materials with different molecular weight had certain biological activity. Powders of
SiO2-PEG 1500 and SiO2-PEG 6000 manifest prooxidant effects, whereas mesoporous (calcine) powders produced
antioxidant effects in blood serum in vitro.
Keywords: Free-Radical Oxidation, Nanosilica, Chemiluminescence, Malonic Dialdehyde
1. Introduction
The study of free-radical processes of lipid oxidation and
antioxidant system is an important problem of modern
biomedicine. Oxygen is a powerful oxidant and oxy-
gen-mediated reactions are the main sources of energy
for variety of biological species. Metabolic processes
produce reactive oxygen species (ROSs), free radicals,
peroxides, malonic dialdehyde, Schiff's bases, which
damage membrane structures and lead to oxidative stress,
being the causative factor for a lot of widely spread dis-
eases, including reproductive disorders and prenatal inju-
ries [1-3].
Concerning this the role of antioxidants is to neutralize
toxic products of free-radical lipid peroxidation. In con-
ditions of excessive peroxidation the capacity of antioxi-
dant defense may decrease due to insufficient endogenic
antioxidant production or unfavorable environment fac-
tors [3]. Various substances are known to have antioxi-
dant activity; their investigation is of certain importance
for both biology and medicine [4].
Nowadays silica compounds attract considerable at-
tention. Silica stimulates fibroblastic activity of mesen-
chyma, promoting granulation and scarring. The lack of
silica may lead to depression of leukocyte activity in in-
flammation, poor wound scarring, anorexia, pruritus,
tissue flexibility decrease, skin turgor decrease, vascular
permeability increase and haemorrhagia as a result. Apart
from this silica antioxidant activity stands [5].
Investigators of peroxidation showed that silica pow-
ders stimulated active oxygen species and free radicals
production in culture of epithelial cells, resulting in cas-
pase activation and apoptosis [6]. Some authors experi-
mentally found out that inhalation of crystalloid silica in
animals lead to oxidative stress, inflammation and alveo-
lar fibrosis [7,8].
As a catalysts and functional composite materials spe-
cial attention is attracted by matrix hybrid silica derivates.
In this study we presented physical-chemical properties
of silica-based materials and their influence on peroxida-
tion processes in human serum. Silica nanoparticles are
biologically inert, have high adsorption rate, thermal and
mechanic stability, and therefore are expected to mani-
fest antioxidant activity.
The aim of the work was to study the influence of or-
ganic-innorganic silica and polyethylene glycols com-
pounds on free-radical processes in vitro.
The Study of Influence of Silica and Polyethilene Glycols Organic-Inorganic Compounds on
Free-Radical Processes in Vitro
Copyright © 2010 SciRes. PP
2. Materials and Methods
2.1. Reagents for Silica Nanoparticles Synthesis
Tetraethoxysilane (TEOS) (C2H5O)4Si (high purity grade,
Ekos-1, Russia), diethylamine (С2Н5)2NH (moderate pu-
rity grade, Aldrich), polyethylene glycols [-OCH2CH2-]n
with molecular weight of 1500, 6000, 15000 (Aldrich)
and rectified ethanol (96 wt%) were used without further
2.2. Synthesis of Hybrid Organic-Inorganic
SiO2-Polyethylene Glycol Materials
The synthesis was carried out in ethanol media at tem-
perature of 20. Polyethylene glycol was dissolved in
required amount of water and was added to ethanol. Than
a catalyst of tatraethoxysilane hydrolysis – diethyl amine
– was added. The resulting mixture was homogenized for
15 min. After that small doses of tatraethoxysilane
(0.5-1.0 ml) were injected into the system using syringe.
Total synthesis time was 17 hrs. Particles were filtered
from the original solution and dried out on open air at
temperature of 70-100 until mass stability.
We investigated two types of hybrid organic-inorganic
SiO2-PEG materials: noncalcine and calcine in muffle
furnace (800, 2 hrs).
2.3. IR Spectroscopy
IR spectra of produced powders were recorded using
Avatar 360 FT-IR ESP spectrometer (wave range: 400-
4000 cm-1). Preliminary samples of powders were grind-
ed with KBr in agate mortar and pressed in discs.
2.4. Thermogravimetry
Thermogravimetric measuring were performed on ther-
moanalythical installation. The mass of a sample was
80-90 mg. The heating was carried out from room tem-
perature up to 1000 with rate of 5 per min.
2.5. Brunauer-Emmett-Teller (BET) Surface
Area Measurement
First step of BET analysis was heating of samples to
eliminate adsorbed impurities. Then argon at temperature
of 77K was adsorbed on materials to form a monomo-
lecular film on accessible area. The amount of adsorbed
argon served to determine the BET surface area.
In experiments we used native serum of 10 patients,
administered in obstetrical or gynecological clinics of
Ivanovo Research Institute of Maternity and Childhood.
We decided not to use whole blood to prevent coagula-
tion. Suspension of studied powders was added to 1 ml of
native serum, which was then incubated at 4 for 1 hr.
The ratio between powders and serum was 5.0 g/L for
mesostructured (noncalcine) powder and 50.0 g/L for
mesoporous (calcine) powder. At this stage of experi-
ments we didn’t try to evaluate pharmacological doses of
studied powders as the only aim of experiments was to
determine any possible pro- or antioxidant properties of
synthesized materials in vitro. The incubation tempera-
ture was found to limit bacterial activity. Native serum
without nanomaterials stood for control. After incubation
samples were centrifuged at 3000 rpm for 10 min. Su-
pernatants were transferred into clean dry tubes and un-
derwent analysis.
2.6. Estimation of Peroxidation Intensity by
Induced Chemiluminescence (CL)
The method was based on catalytic reduction of perox-
ides by bivalent ferric ions. Generated free radicals acted
as oxidation initiators in biological substrate. Free radi-
cals recombination followed by photon release, which
could be registered in 40 s. This was a period of maximal
intensity. We used hydrogen peroxide and ferric sulfate
as biochemiluminescence inductors. 0.1 ml of serum was
mixed with 0.4 ml phosphate buffer (pH 7.5), 0.4 ml fer-
ric sulfate (0.01 mM), 0.2 ml 2% hydrogen peroxide and
underwent CL. Intensity of luminescence was evaluated
by BHL-06M biochemiluminometer (Russia). Registered
parameters were as follows (Figure 1): peak intensity of
luminescense (Imax) – the highest intensity of lumines-
cense registered within first 40 seconds of Fenton’s reac-
tion; light sum (S) – the area under the curve of lumines-
cent signal; slope of the curve (tg α) – tangent of de-
crease signal angle.
Imax and S reflected potential for lipid peroxidation.
Antioxidant activity was characterized by tg α and coeffi-
cient K (Imax/S ratio).
Figure 1. Typical kinetics of serum chemiluminescense. The
registered parameters are as follows: peak intensity (Imax),
light sum (S(t)), the slope of the curve (tg α).
The Study of Influence of Silica and Polyethilene Glycols Organic-Inorganic Compounds on
Free-Radical Processes in Vitro
Copyright © 2010 SciRes. PP
2.7. Malonic Dialdehyde Serum Concentration
The method was based on reaction between malonic
dialdehyde (MDA) and thiobarbituric acid [9]; the prod-
uct of reaction had pink color and was identified by
SF-46 spectrophotometer (Russia) on wave length 523
3. Results and Discussion
Qualitative and quantitative structure of materials was
investigated by IR spectroscopy and thermogravimetric
Thermograms of hybrid organic-inorganic SiO2-PEG
material (molecular weight 15000) are presented on Fig-
ure 2.
Burning of polymer component from matrix SiO2-PEG
15000 hybrid is described by the peak with maximum at
291 on differential thermogravimetric diagram. The
amount of organic phase was 39 wt% in sample. Water
content was 13.5 wt%. The curve of differential thermal
analysis did not reveal any thermal effects.
Figure 3 shows IR spectra of hybrid organic-inorganic
(a) SiO2-PEG 15000 and (b) material calcined at 800.
There were characteristic oscillations of Si-OH and
Si-O-Si found in spectra. Also, in areas of wavenumbers
3600-3500, 1640 and 400-200 cm-1 oscillations of ad-
sorbed H2O, isolated and linked OH groups were found.
In spectrum of hybrid organic-inorganic SiO2-PEG
15000 oscillations of alkyl CH3, CH2 and CH groups
[10-19] proved the presence of organic phase in compos-
Intensive flat halo on X-ray diagram of hybrid SiO2-
PEG 15000 material reflected its amorphism (Figure 4).
Results of sedimentation gave data on morphology of
hybrid material powders. Sizes of particles of hybrid
SiO2-PEG composite determined by sedimentation be-
fore and after calcining were (Figure 5) as follows. Hy-
brid material: 2-300 µm, with the prevalence of 24 and
150 µm fractions; tempered material: 2-110 µm, with
modal size 28 µm. Specific areas of noncalcine and cal-
cine hybrid organic-inorganic SiO2-PEG 15000 material
powders were discovered by thermal desorption of argon
(Table 1).
Thus, there were synthesized two groups of composite
materials with different structure: organic-inorganic
compounds with amorphous structure and mesostructural
materials with system of ordered mesochannels, filled by
molecular organic templates. All materials were calcined
at 800 for creating of high-porous silica specimens
with the same structural features as their noncalcine an-
cestors. Investigation of pro- and antioxidant properties
Figure 2. Thermograms of hybrid organic-inorganic SiO2-
PEG 15000 material.
Table 1. Specific surface areas of hybrid SiO2-PEG 15000
and calcined at 800 materials.
Material Specific surface area by
BET, m2/g
SiO2-PEG 15000 (noncalcine) 91
SiO2-PEG 15000 (calcine) > 800
of these powders was carried out in vitro.
Biochemical study revealed different influence of in-
vestigated powders on intensity of peroxidation in human
serum (Tables 2 and 3). Parameters of chemilumines-
cence revealed in native serum were estimated as 100%.
Adding of SiO2-PEG powders with molecular weight
1500 and 6000 to experimental system lead to increased
chemiluminescence, which proved high ROS and free
radical (R-, OH-, RO-, RO2-, O2-) production in Fenton's
reaction [20,21]. Recombination of radicals formed un-
stable tetraoxide, which disintegrated with photon emis-
sion and raised lightsum S (plus 23%, p = 0.0081 and
18%, p = 0.0131, comparing with controls, respectively
for 1500 and 6000 powders), fast flash Imax (plus 26%, p
= 0.0459 for 1500 powder). Investigation of SiO2-PEG
with molecular weight 15000 didn’t reveal any signifi-
cant CL changes. Study of mesoporous (calcine) SiO2-
PEG powder revealed its antioxidant activity, which was
developed by decreasing of peroxidation processes inten-
sity in Samples 3 and 5 (Table 3). We found increasing
of tg α (plus 15%, p = 0.0510) and decreasing of MDA.
The conclusion was that nanosilica-PEG compounds with
different molecular weight possessed specific oxidant
and antioxidant activity. SiO2-PEG 1500 and SiO2-PEG
6000 powders lead to activation of free-radical oxidation,
The Study of Influence of Silica and Polyethilene Glycols Organic-Inorganic Compounds on
Free-Radical Processes in Vitro
Copyright © 2010 SciRes. PP
Figure 3. IR spectra of noncalcine (a) and calcine (b) hybrid organic-inorganic SiO2-PEG 15000 materials.
Table 2. Parameters of peroxidation (MDA) and induced chemiluminescence in blood serum after adding of
mesostructured (noncalcine) SiO2-PEG powders (5 mg/mL) with different molecular weight (M).
Parameters Statistics n = 5 M=1500 M=6000 M=15000
MDA, % M ± m 102.67 ± 21.40 86.67 ± 8.69 98.33 ± 10.81
S, % M ± m 123.00 ± 2.08** 118.00 ± 2.08** 102.0 ± 8.39
Imax, % M ± m 126.00 ± 5.77* 115.33 ± 8.69 105.33 ± 10.91
tg α, % M ± m 125.67 ± 6.77 110.33 ± 4.06 103.00 ± 13.20
K, % M ± m 95.67 ± 4.48 92.33 ± 4.33 97.33 ± 3.38
Note: Here and further asterix reflects significant differences among estimated groups (*р < 0,05; **р < 0,01).
Table 3. Parameters of peroxidation (MDA) and induced chemiluminescence of blood serum after adding of
mesoporous (calcine) SiO2-PEG powders (50 mg/mL) with different molecular weight (M).
M = 1500 M = 6000 M = 15000
Parameters Statistics n = 5
1 2 3 4 5 6
MDA, % М ± m 94.60 ± 9.49 86.60 ± 6.9889.00 ± 7.7793.20 ± 10.5489.60 ± 3.79* 100.20 ± 4.10
S, % М ± m 97.80 ± 3.69 93,00 ± 6.89103.80 ± 3.6298.40 ± 6.2796.10 ± 1.91 109.80 ± 5.19
Imax, % М ± m 104.80 ± 5.80 103.00 ± 8.96111.20 ± 8.73103,40 ± 12.8698.20 ± 7.68 150.50 ± 29.70
tg α, % М ± m 116,80 ± 9.73 128.60 ± 15.41115.20 ± 5.17*114.20 ± 17.11112.80 ± 4.64* 173.25 ± 43.16
К, % М ± m 100,40 ± 2.79 109.00 ± 2.65*99.20 ± 1.8399.60 ± 6.0294.20 ± 2.48 120.00 ± 22.76
Note: Numbers 1, 2, 3, 4, 5, 6 stands for different preparations of mesoporous (calcine) SiO2-PEG with different molecular weight (1500, 6000, 15000).
The Study of Influence of Silica and Polyethilene Glycols Organic-Inorganic Compounds on
Free-Radical Processes in Vitro
Copyright © 2010 SciRes. PP
Figure 4. X-ray diagram of hybrid organic-inorganic SiO2-
PEG 15000 material.
050100 150 200 250 300
noncalcine material
calcine material
fraction content,%
particle size, m
Figure 5. Particle size distribution in noncalcine and calcine
hybrid organic-inorganic SiO2-PEG 15000 materials.
whereas mesoporous (calcine) materials with molecular
mass 6000 and 15000 manifested antioxidant properties
in human serum due to their large specific surface area (>
800 m2/g). These findings can be useful for creating of
pharmacological preparations.
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The Study of Influence of Silica and Polyethilene Glycols Organic-Inorganic Compounds on
Free-Radical Processes in Vitro
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