DNA-hexadecyltrimethyl ammonium chloride complex with enhanced thermostability as promising electronic and optoelectronic material
Recently DNA with hexadecyltrimethyl ammonium chloride (CTMA) complex has been included in some organic electronic devices. Thermal stability is one of key parameters required for successful applications in different electronic and optoelectronic devices. This work shows a possibility of enhancing thermal stability in this complexes and analyzes origin of this enhancement. Different techniques were applied to explore this issue of solid DNA-CTMA. Results of TGA analysis, DSC calorimetry, FTIR spectroscopy, and analysis of evolved gaseous products convince that chemical composition of DNA-CTMA complex remains fixed at temperatures <200–220 °C. In contrast, broadband dielectric spectroscopy applied to freshly prepared thin films of DNA-CTMA revealed at 150–160 °C a permanent and irreversible change of dielectric properties. This phenomenon may be attributed to a transformation affecting the microstructure. Some experiments were conducted also for the native DNA as a reference. We demonstrate that DNA-CTMA complex chemical composition is more stable at temperatures about 200 °C with respect to DNA which is very important for laser operated optoelectronic applications.
KeywordsDifferential Scanning Calorimetry Circular Dichroism Spectrum Differential Scanning Calorimetry Experiment CTMA Cetyltrimethylammonium Chloride
During the past decade biomaterials and their derivatives applied in solid filmform, began to be considered as promising electronic and optoelectronic material . Particular interest presents DNA as promising materials for biodegradable electronics. It was just demonstrated a significant number of working devices possessing DNA or its derivatives and other organic chemicals. For example owing to a rare combination of HOMO and LUMO energy positions, this biopolymer can be used as an effective electron blocker. A thin layer of DNA derivative introduced in the structure of an of an organic light emitting diode (OLED) has enhanced significantly its performance [1, 2, 3]. High dielectric constant combined with low electronic conductivity make DNA an interesting material for organic field effect transistors [4, 5]. Different dyes can be particularly efficiently intercalated and spatially separated in DNA matrix, what is of enormous interest for nonlinear optics [6, 7, 8], lasers technology [9, 10, 11] and electrochromic devices .
The majority of devices classified as “organic electronics” consist of a series of “thin” films, with thickness varying within 10…100 nm. The majority of standard techniques that transform polymeric materials in thin films are based on solvents, like spin-coating, dip coating, Dr. Blade or ink-jet printing. Biological functions of the native DNA limit the number of available solvents to water only. In order to use more volatile solvents, DNA needs to be converted in complex with an amphiphilic lipid, which plays the role of surfactant. Such a complex dissolves in a range of alcohols and some chlorinated solvents. In revenge it is insoluble in water, but even “well” dried specimens of DNA or its complexes contains some residual structural water, impossible to be removed [13, 14, 15, 16].
Properties of an electronic or a photonic device should be environmentally stable. This stability is determined by the stability of each of constituents. Generally polymeric materials are affected by environmental factors more intensively than inorganic compounds. Different aspects of DNA properties have been extensively studied during the past 60 years. These studies were carried out in aqueous environment due to interest of biological functions of DNA. However, there exist objective premises suggesting differences between properties of the native DNA in the solid and in aqueous solution, particularly when DNA is in complex with amphiphilic lipids.
Temperature is a key parameter for the stability of DNA properties. For example in water, upon heating, DNA double helix splits into single strands, what is referred to as melting . An increased temperature may be the source of DNA decomposition, as it was found for example in the case of DNA in bone material . The thermal decomposition depends on different factors and is not instantaneous. Heating makes the part of different protocols used for construction of many DNA consisting devices. Though, the question of DNA thermal stability is usually neglected. Usually discussion it is limited to a declaration that DNA thermal stability is superior to commercially available polymers like for example poly(methyl methacrylate) (PMMA). The source of such opinion arises from results obtained by standard polymer testing methods, like thermogravimetric analysis (TGA) or differential scanning calorimetry (DSC) [8, 19]. Commonly, the thermal stability is referred to the temperature, at which occurs loss of the mass other than entrapped solvent release. This temperature, situated between 200 and 220 °C, was found practically independent on surfactant type and its content in the material.
Excessive heating may occur also in other experiments. For example, high power laser light is absorbed and transferred to the bulk as the heat in experiments with DNA/dye composites studied for potentially occurring non-linear optical phenomena. Therefore, processes related to thermal annealing of DNA need to be well understand. It should be emphasized that relatively low photo-thermal stability is main restraining factor to apply them in optoelectronic devices with high power laser pulses.
In this work a range of complementary techniques were applied to describe quantitatively thermal annealing of the complex of DNA and hexadecyltrimethyl ammonium chloride (CTMA). So far, this complex may be considered as a standard in the field of DNA containing organic optoelectronics. Selected experiments were conducted also for the native DNA as reference.
2 Materials and methods
2.1 Nucleic acids and chemicals
DNA extracted from salmon roe and milt sacs, was purchased from Chitose Institute of Science and Technology (CIST) in Hokkaido, Japan. The exact details of purification technique applied at mass scale isolation are proprietary. Purity claimed by the supplier was c.a. 96 % with a protein content of c.a. 2 %. In an earlier study we found also traces of 3d-metals . The CIST-DNA has been the subject of many works on DNA based electronics  and so far can be considered as a standard material in this domain. According to many authors the average molecular weight of the CIST-DNA is centered at 8 MDa . However, our tests by agarose gel electrophoresis revealed broad bands, structureless and without a detectable maximum. Consequently, “a few MDa” seems to be the most reliable estimate for polydispersed molecular weight of the CIST-DNA. All the other chemicals [n-Butanol, analytical grade and cetyltrimethylammonium chloride (≥98 %)] were supplied by Sigma-Aldrich and used as received. Throughout this work only Millipore water was used.
2.2 Synthesis of the DNA complex
DNA was dissolved in water (1.5 g to 250 ml H2O) on magnetic stirrer. Visually clear solution was sonicated in an ultrasonic bath at room temperature for 10 min. This procedure decreased the molecular mass of DNA and the viscosity of the solution. It is believed that the cationic surfactant can easier access the surface of shorter DNA helices owing to limited tertiary structure adapted by the polymer. In result of sonication, DNA molecular weight was reduced to few hundred base pairs (as determined by agarose gel electrophoresis). It was verified that a longer sonication didn’t further decrease molecular weight. Next, the solution was precipitated with aliquot of the surfactant CTMA solution, maintaining DNA to CTMA 1:1 wt. ratio. The precipitate was abundantly rinsed with water, dialyzed on 0.45 µm PTFE filter and dried in a vacuum dryer.
2.3 Preparation of samples
DNA and DNA-CTMA powders were finely ground in agate mortar. Portions of both powders were re-dissolved at 10 wt% in Millipore water (DNA) and butanol (DNA-CTMA). After being filtered, the solutions were poured in PTFE moulds and let evaporate in the ambient air. The obtained films were c.a. 2 μm thick and were separated from the substrate. All the samples, prior to measurements, were additionally dried in primary vacuum for 24 h at ambient temperature. If necessary, they were stored in air tight vials filled with argon and in presence of silica gel.
2.4 Experimental techniques
Thermally triggered mass loss was explored by thermogravimetric analysis (TGA) using Netzsch TG 209 apparatus. The samples were heated at constant rates in flux of argon or oxygen. The effect of the heating rate (varied between 5 and 20 K/min) was negligible. The first specimens of DNA and DNA-CTMA powders were heated from the ambient temperature up to 600 °C. According to the obtained results, four temperatures (190, 230, 270 and 400 °C) were arbitrary chosen as “critical”. Thereafter, the analytical runs was repeated with fresh specimens, but the heating was stopped at a one of these “critical” temperatures. The residues were carefully collected from the analytic pan, ground with KBr and pressed in pellets. Afterwards, infrared spectra were collected using Spectrum 65 Perkin Elmer FTIR spectrometer with spectral resolution 1 cm−1. A supplementary study was carried out through FTIR analysis of the gases released by the heated samples versus temperature. This type of measurements were performed using thermal analyzer STA F3 449 Jupiter Netzsch coupled to TENSOR 27 Bruker FTIR spectrometer.
Differential scanning calorimetry (DSC) scans were recorded employing DSC Mettler Toledo 822e calorimeter, the instrument that use two-furnace method. One of the furnaces was loaded with a sealed pan with the titled specimen, while the other with an empty sealed have been served as reference. In some cases, the lid of the DSC pan was punctured to let out vapors released during the heating. The sample and reference compartments were continuously purged with high-purity argon to prevent the sample from rehydration. Samples were heated and cooled cyclically few times. Starting from 25 °C the sample was heated at a constant ramp either (either 20, 10 K/min or K/min) up to 190 °C and finally cooled down back to the initial temperature. Such cycles, consisting of two runs (heating and cooling), were repeated. In the further text, the runs will be referred by the cycle number.
Circular dichroism is considered as an ultimate evidence that DNA exists in the form of the double helix and not as a bunch of separated single strands. Circular dichroism spectra of thermally annealed DNA-CTMA samples were measured at room temperature on a JASCO J-815 (Japan) spectropolarimeter.
3 Results and discussion
An analysis of graphs in Fig. 4, leads to conclusion that chemical decomposition occurs as a multi-step process. A one thing different between spectra of samples heated at 25 °C and at 190 °C is a slight increase of the band intensity at wavelength 1700 cm−1. Such a trend is reverse to that observed as result of dehydration . It is surprising because an excessive dehydration is expected due to thermally forced removal of the structural water, inherently built in DNA structure.
More visible changes occur between spectra recorded at 190 °C and at 230 °C. In DNA spectrum even bands characteristic of sugar and bases disappear. DNA-CTMA spectrum preserves some initial characteristics, but the intensity of c.a. 1060 cm−1 backbone vibrations is much decreased. Probably a fraction of the initial DNA-CTMA still preserves its structure. Spectra of the samples heated at 400 °C are featureless without any characteristics of DNA.
Indeed, volatile compounds other than water, were released not earlier than at temperatures exceeding 210 °C (DNA) and 217 °C (DNA-CTMA). Starting with the ambient temperature, a series of close, narrow lines of water vapor can be seen in the FTIR spectra between 1300 and 2000 cm−1. A particularly sharp release of water vapor was observed between 70 and 140 °C with a maximum centered at 110 °C.
Differential scanning calorimetry (DSC) offers supplementary information on DNA thermodynamics. DSC of solid DNA has not often been reported. Available data appeared mainly in bibliography discussing hydration of DNA [27, 28, 29, 30, 31]. DSC of DNA-CTMA was reported even less frequently [32, 33]. Interpretation and comparison between already published results (sometimes substantially different) is further complicated by the fact of different experimental procedures implemented by the authors. One of important issues, already pointed out , is gas-tightness of pans where the specimen resides during DSC experiment. Vapors is released from the sample condensate inside the pan when temperature returns to the ambient value. It is not evident that water can rehydrate DNA in such a manner that the initial state of the material would be recovered. In result, during the next heating cycle a bi-phasic system will be studied. The solution to this inconvenience is a not hermetic pan. In such a case all vapors are removed during the first heating run by the inert gas purging the heating compartment. In this study lids of DSC pans were punctured to make them non-hermetic.
The bound water may act as a natural plasticizer  in many biopolymers. The observed reproducibility of DNA-CTMA glass transition means that either some of the bound water remains confined despite heating up to 190 °C (what would be surprising) or CTMA plasticizes DNA (what is a more realistic explanation). The exact value of Tg will be depend on many factors. Other authors situate Tg of DNA-CTMA, found by DSC, at much higher temperatures, something like 150 °C . However, in the last work precisions on details of the experiment are not explained.
A logical description of the observed phenomenon must include rearrangement of the microstructure. The material, initially remains stiff and temperature practically does not affects its relaxation. After reaching 100 °C it becomes gradually viscoelastic and the maximum of relaxation moves towards higher frequencies. At the extreme end of the temperature range occurs a phase transition. In result, the material becomes more stiff, however not as stiff as it was initially at lower temperatures. This new structure persists despite consecutively repeated heating/cooling cycles.
Nevertheless, inspection of the graphs in Fig. 14 shows, that the CD spectra preserved their overall shape at temperatures as high as at least 180 °C.
We have established that DNA-CTMA complex chemical composition is more stable at temperatures about 200 °C with respect to traditional ones which is very important for laser operated optoelectronics applications. Such novel material preserves its dielectric properties regardless further heating what may additionally confirm their enhanced thermal stability. The DNA-CTMA complex which initially is insensitive to temperature, at 150–160 °C permanently changes its dielectric properties. The most probable explanation is a new microstructure adapted by the material. The observed reproducibility of DNA-CTMA glass transition means that either some of the bound water remains confined despite heating up to 190 °C (what would be surprising) or CTMA plasticizes the DNA (what is a more realistic explanation). The Tg is shifted towards higher temperatures as hydration of the sample decreased. However, at an extremely low hydration (one water molecule per nucleotide) the characteristic step is disappeared. This absence of glass transition is in agreement with DNA thermogram if one suppose that all water was removed from the sample during the first heating run. The previous research have shown the “first run” DSC thermograms of solid DNA, that contain visible singularities between 40 and 80 °C. These singularities were attributed to denaturation of DNA. In the current study such facts were not reproduced. A DNA specimen was subjected to two heating/cooling cycles situated between 25 and 90 °C. Between the end of heating run and the beginning of cooling run, the sample was conditioned at 90 °C for 15 min. Generally in this temperature range it was absent in this case. The general description of the observed phenomenon must include rearrangement of the microstructure. The material, initially remains stiff and temperature practically does not affects its relaxation. After reaching 100 °C it becomes gradually viscoelastic and the maximum of relaxation moves towards higher frequencies. At the extreme end of the temperature range occurs a phase transition. In result, the material becomes more stiff, however not as stiff as it was initially at lower temperatures. This new structure persists despite consecutively repeated heating/cooling cycles.
This project was partially financed by the Polish Ministry of Science and Higher Education. The research was carried out with the equipment purchased thanks to the financial support of the European Regional Development Fund in the framework of the Polish Innovation Economy Operational Program (Contract No. POIG.02.01.00-12-023/08).
- 1.B. Sun, D. Liang, X. Li, P. Chen, J. Mater. Sci.: Mater. Electron. 27, 3957–3962 (2016)Google Scholar
- 32.E.M. Heckman, C.M. Bartsch, P.P. Yaney, G. Subramanyam, F. Ouchen, J.G. Grote, in Materials Science of DNA, 1st edn., ed. by J.-I. Jin, J. Grote (CRC Press, Boca Raton, 2011), pp. 180–229Google Scholar
- 33.L.E. Johnson, L.N. Latimer, S.J. Benight, Z.H. Watanabe, D.L. Elder, B.H. Robinson, C.M. Bartsch, E.M. Heckman, G. Depotter, K. Clays, Proc. SPIE 8464, 1–10 (2012)Google Scholar
Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.