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NANOSYSTEMS: PHYSICS, CHEMISTRY, MATHEMATICS, 2016, 7 (1), P. 153-157
Study of antioxidant activity of fullerenols by inhibition of adrenaline autoxidation
I. A. Dubinina1,2, E. M. Kuzmina1, A.I. Dudnik2, N. G. Vnukova1,2, G.N. Churilov1,2, N. A. Samoylova3
1 Siberian Federal University, Krasnoyarsk, Russia 2Kirensky Institute of Physics SB RAS, Krasnoyarsk, Russia 3Leibniz Institute for Solid State and Materials Research, Dresden, Germany
churilov@iph.krasn.ru
PACS 81.05.ub DOI 10.17586/2220-8054-2016-7-1-153-157
In this paper, we describe application of the adrenaline autoxidation reaction to determine the antioxidant activity of fullerenols C6o, C70 their mixture with higher fullerenol and endohedral fullerenol Y@C82. It was shown that the adrenaline autoxidation reaction can be applied to determine the antioxidant activity of fullerenols. The antioxidant activity of C70 fullerenol was higher than that of C60 fullerenol Additionally, the antioxidant activity of Y@C82 fullerenol was higher than that of C70 fullerenol.
Keywords: fullerenol, antioxidant activity, endohedral fullerenol, adrenaline autoxidation. Received: 20 November 2015
1. Introduction
Polyhydroxylated fullerenes have been developed as the main derivative of fullerenes with excellent biocompatibility and biofunctionality [1,2]. Indeed, C6o fullerenols have been found to have outstanding antioxidant activity, which scavenge almost all reactive oxygen species (ROS) and reactive nitrogen species (RNS) [3].
Endohedral metallofullerenes, i.e. molecules in which a fullerene encapsulates a metal atom(s), have shown great promise for use in biomedical science. Although C60 has been the most commonly studied fullerene in biological systems, a few endohedral materials have been synthesized using C60 as a cage molecule despite the limited interior volume of C60. Therefore, most endohedral metallofullerenes are synthesized using C82 or higher molecular weight ffullerenes (presumably with larger interior volumes). Our group has prepared many C82 ffullerene derivatives. Y@C82 is one of the most important molecules in the metallofullerene family [4]. Yttrium endohedral metallofullerenol (e.g., Y@C82(OH)22) is a ffunctionalized fullerene with yttrium trapped inside the C82 fullerene cage. It was reported that the chemical and physical properties of endohedral metallofullerenols depend on the number and position of the hydroxyl groups on the ffullerene cage [4]. These results demonstrated that modifying the outer cage of Y@C82 with a number of hydroxyl groups tunes the electronic properties of the inner metal atom as well as the electron density and polarizability of the electrons localized on the fullerene's cage.
A number of fullerenes, fullerenols, and endohedral metallofullerenols have been shown to be capable of scavenging free radicals [3,5]. However, much less is known about the antiox-idant activity of Y@C82(OH)22. It has not been determined if the ROS-scavenging capability of Y@C82(OH)22 is higher than that of other functionalized fullerenes. In this work, we applied
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the adrenaline autoxidation reaction to determine the antioxidant activity of fullerenols Ceo and their mixture with higher and endohedral fullerenols.
2. Methods
2.1. Preparation of fullerenols
Fullerene mixture with endohedral Y-fullerene was synthesized at the set up with high frequency (HF) arc plasma for sputter graphitic electrodes (graphitic rods) [6,7]. Graphitic rods, with an axial hole in which Y2O3 was inserted, were annealed at 1800 °C for 20 min. Arc current was 190 A, frequency - 66 kHz, and chamber pressure was 98 kPa. Extraction was carried out using CS2.
Synthesized fullerenes were separated by liquid chromatography with turbostratic graphite (graphite with an interplanar distance 3.42 A) as stationary phase and toluene/hexane (4:6) mixture as mobile phase (or pure toluene in case of C70 elution). Endohedral Y-containing fullerene was separated using HPLC chromatography (Agilent 1200-series with Buckyprep column).
The experimental preparation of fullerenols was achieved under acidic conditions by treatment with nitric acid.
2.2. Antioxidant activity study
Antioxidant activity (AA) of fullerenols was measured according to Hasanova et. al [8] by the inhibition of adrenaline autoxidation. The reaction mixture contained 0.1 cm3 of 0.1 % adrenaline hydrochloride, 4 cm3 carbonate buffer, pH 10.55, and 0.1 cm3 of fullerenol solution. The increase in absorbance due to adrenaline autoxidation was monitored at 347 nm and the percentage inhibition of the maximal rate of increase in absorbance was determined. According to [8] more than 10 % inhibition rate shows the AA of investigated solution. This method has been developed for measuring AA of vegetative gathering and for the first time applied for fullerenols. The AA was calculated as follows:
AA = (D - ^M00, % (1)
where D1 is optical density of pure adrenaline and D2 is optical density of adrenaline with fullerenol.
3. Results
Synthesized Y-containing fullerene mixture was investigated by HPLC (Fig. 1). We collected fraction with Y-containing endofullerene (30 - 40 min) and investigated it by mass-spectrometry (Bruker BIFLEXTM III Time-of-Flight mass spectrometer with laser desorption) (Fig. 2). The fullerene mixture was shown to consist of only one type of endohedral metallo-fullerene - Y@C82. The fullerene mixture was separated and 3 fractions were received - pure C60 (99 %), C70 with higher fullerenes (92 and 8 %, respectively), and higher fullerenes with 6 % of Y@C82.
Fullerene fractions were hydroxylated and solutions with concentrations 2 mg/ml of fullerenols were produced.
Figure 3 shows the dynamics for the adrenaline autoxidation reaction. The decrease in the optical density showed the inhibitory effect of the fullerenols. To rate the influence of fullerenols on adrenaline autoxidation process, different exposure times were chosen - 30 s, 3 min, 5 min. Calculation of AA showed that all fullerenols displayed great AA. The AA significantly increased with increasing concentration of higher fullerenols (Fig. 4).
Study of antioxidant activity of fullerenols...
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Fig. 1. Chromatogram of fullerene mixture with Y-endofullerene
Fig. 2. Mass-spectrum of fullerene fraction with Y-endofullerene (30 - 40 min)
4. Conclusion
Fullerenols were investigated to see if they inhibit the adrenaline autoxidation reaction and thus show great antioxidant activity. Increasing the higher fullerenol content, as well as Y@Cg2 content, resulted in an increase in the antioxidant activity of fullerenol solution.
Acknowledgements
The work was supported by the Russian Foundation for Basic Research 15-03-06786.
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Fig. 3. Optical density dependence on reaction time. 1 - pure adrenaline, 2 -C6o fUllerenol, 3 - C70 and higher fullerenols and 4 - endohedral and higher fullerenols
Fig. 4. Comparative antioxidant activity of fullerenols (where 100 % of AA means that adrenaline autoxidation doesn't occur, and 0 % means that there is no inhibitory effect). Statistically significant changes in AA level (ANOVA, p < 0.05, n = 3)
Study of antioxidant activity of fullerenols.
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