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Abstract
Light amplification in DNA systems is a promising technique for efficient and compact photonics devices in the future. Since highly fluorescent properties for hemicyanine dyes are caused by the interaction with DNA or a DNA-complex, their effects on basic optical properties were elaborated in phases of solution and solid films prepared with two methods. Results of absorption and fluorescence characteristics showed that the strongest light emission was achieved by the interaction with DNA-CTMA (cetyltrimethylammonium) rather than direct coupling to bare DNA. Laser performance was compared for films prepared with two means, indicating that this newly developed immersion method provided highly doped samples with better performance in intensity, threshold value and durability.
© 2017 Optical Society of America
1. Introduction
Since N. Ogata proposed electronic and photonic application of dye-functionalized DNA-surfactant complexes, vast studies were made to show their high performance in electroluminescent diodes, nonlinear optical devices, optical amplifiers, lasers and so on [1–4]. Amplified spontaneous emission (ASE) in the complex was demonstrated in 2000, before many types of dyes were incorporated to show their superior performance as laser devices [5–11]. The origin of the high capability was apparently based on fluorescence enhancement of the dyes interacting with DNA complexes. However, the root cause of the enhancement was not clarified very well. At an early stage, intercalation of the dyes into the space between base pairs in helical strand was considered to be the principal reason as evidenced for the case of ethidium bromide [12]. On the other hand, binding of the dyes to minor grooves of DNA was suggested to be an important factor as inferred from spectroscopic studies on cyanine dyes interacting with DNA [13,14]. Recently, we found that immersion of DNA-complex films into organic dye solutions provided highly doped complex films with high optical quality [15,16]. Films prepared with the method might incorporate dyes in outer space of DNA by electrostatic force, because DNA has already made strong coupling with surfactant. These films also showed strong fluorescence, good ASE performance and high durability under optical pumping.
Laser and ASE characteristics as well as optical properties such as absorption or fluorescence spectra depended on their fabrication process. In other words, it is possible to get insight into the interaction mechanism responsible for high performance and to control or optimize the device efficiency through preparation method. Therefore, it is worthwhile to investigate the relationship between preparation method and optical properties in detail. We have mainly studied laser characteristics of hemicyanine dyes embedded in DNA-cetyltrimethylammonium (CTMA) [6,15,16]. In this study, several types of hemicyanine dyes were used as active materials, and their spectroscopic properties were systematically studied as well as laser performance for films made by different methods. In the following section, sample preparation and experimental methods are described. Spectroscopic results on solutions and films are given in section 3, and lasing properties are summarized in section 4. Concluding remarks will be addressed in the last part.
2. Sample preparation and optical experimental setup
In this study, three types of hemicyanines were employed of which molecular structures are given in Fig. 1. Among them, 4-[4-(dimethylamino)styryl]-1-dococypyridinium (p-hemi22) has been used in our former studies, showing good laser performance [6,15]. Additionally two types of water-soluble short alkyl-chained hemicyanines were employed in this study, because their solubility to water and organic solvents made it possible to compare the interaction with DNA and with DNA-CTMA. Dyes and CTMA were purchased from Sigma-Aldrich and DNA was provided by the late N. Ogata. All of them were used without further purification. DNA-CTMA was precipitated by adding aqueous CTMA solution into DNA water solution as described in preceding works [1].
Fig. 1 Molecular structures of hemicyanines employed. (a) 4-[4-(dimethylamino)styryl]-1-methylpyridinium (p-hemi1), (b) 2-[4-(dimethylamino)styryl]-1-methylpyridinium (o-hemi1), and (c) 4-[4-(dimethylamino)styryl]-1-dococypyridinium (p-hemi22).
In order to study the effects from DNA and DNA-complex in solution, concentration of the dyes was designed to be 10−5 mol/L. DNA base pair concentration was adjusted from 0 to 5 x 10−4 mol/L, corresponding to 0 to 50 times greater than that of the dyes. DNA and the complex were added to the water and ethanol solutions, respectively. Absorption and fluorescence spectra were measured with a spectrometer V-670 (JASCO), and a fluorometer RF-5300PC (Shimadzu), respectively.
For the investigation of lasing properties, sample films were formed with two methods based on spin coating. In ‘conventional method’, DNA-CTMA was dissolved in ethanol along with hemicyanines and stirred for one day at room temperature. Concentration of the complex was 60g/L and amount of the dyes was typically adjusted to be 5wt% in solid polymer. Films of 1μm thickness were obtained with spin-coating on glass substrates. For ‘immersion method’, non-doped DNA-CTMA films prepared with the spin-coating were immersed in dye solutions typically for one day. The method was inspired by the study of You et al. who applied it to non-filmed complex samples [17]. In this study, acetone was used as a solvent, although it slightly softened the films. Water could be another option, but dye adsorption from water solution was not very efficient in this case, contrary to rhodamine dyes [18,19].
For the measurements of emission spectra and output intensity to determine threshold value, we made the optical setup to form population grating in the sample films, because the method did not need extra fabrication processes, making it possible to do the measurements with the identical condition. Second harmonics of Nd3+:YAG laser with 10Hz repetition was used for pumping. The laser beam was split into two branches with a beam splitter to make interference on the samples, and incident angles were adjusted manually with two mirrors. In our experiment, half angle was varied around 40 degree to obtain second order Bragg grating [16,20]. Intensity and the spectra of emission from substrate edge were recorded by a spectrometer coupled with an optical fiber head.
3. Optical characteristics of hemicyanines interacting with DNA and DNA-CTMA
Absorption spectra of p-hemi1 and o-hemi1 aqueous solutions were measured with a constant dye concentration while varying additive DNA amounts. Result given in Fig. 2(a) indicated that the addition of DNA reduced the absorption magnitude of p-hemi1 and also shifted its peak position to the longer wavelength side. On the other hand, no significant change was observed when DNA-CTMA was added into ethanol solutions, although slight absorption decrement was found as shown in Fig. 2(b). Similar behavior was given for o-hemi1 case. For the case of p-hemi22 in ethanol, no shift was caused by DNA-CTMA.
Fig. 2 Dependence of absorption spectra for p-hemi1 (1 x 10−5 M) on the concentration of (a) DNA and (b) DNA-CTMA. Solvents were (a) water and (b) ethanol.
Fluorescence spectra for the same solutions of p-hemi1 are given in Fig. 3. Fluorescence intensity increased significantly with the addition of DNA in water solution without spectral shift. Slight blue shift was caused by DNA-CTMA when the dyes were dissolved in ethanol, although the change of spectrum was not very large. Similar wavelength shift was observed for o-hemi1, but was less significant for p-hemi22.
Fig. 3 Dependence of fluorescence spectra for p-hemi1 on the concentration of (a) DNA and (b) DNA-CTMA obtained with the same solutions used for Fig. 2. Excitation wavelength was 440nm for both.
Large shift of absorption peaks and the reduction of their magnitudes in hemicyanines with pure DNA suggested the strong dye-DNA interaction in the ground state. Existence of isosbestic point (in particular clearly observed for o-hemi1) indicated that two forms were in equilibrium, that is, free non-emissive state and bound emissive state. Dependence of emission intensity on DNA concentration is shown in Fig. 4, which shows that the spectral shift and emission enhancement occurred with identical DNA ratio, which means that the emission enhancement, spectral shift and peak magnitude reduction could be assigned to the interaction with DNA. These three effects had been observed in ethidium bromide intercalated in DNA strands [12]. Although the possibility of intercalation could not be denied, we temporarily attribute the enhancement to the groove binding, mainly because Kumar et al. concluded that main interaction mode for p-hemi1 (DSMI) was groove binding [21]. To get further insight into the interaction mode, circular dichroism measurement is in progress.
Fig. 4 Relationship between fluorescence intensity and DNA molar ratio for hemicyanines interacting with DNA or DNA-complex in solutions. Intensities are normalized by absorbed excitation light flux.
When the dyes were mixed with DNA-CTMA, no remarkable shift was observed but peak intensity slightly reduced, maybe due to the change of local field made by surrounding surfactant molecules. On the other hand, fluorescence spectral shift was caused by the interaction with the complex, suggesting that the dye experienced strong influence in excited states. Magnitudes of the shift were 12, 18, and 6nm for p-hemi1, o-hemi1, and p-hemi22, respectively. If the enhancement and shift were originated from the proximity of conjugated part of the dye to DNA, long alkyl chain presumably hampered approaching of chromophore to DNA strand for p-hemi22. However, since no difference was found for the three cases in the behavior shown in Fig. 4, the enhancement could not be attributed to direct binding to DNA strand. DNA-CTMA could be a good template for suppressing non-radiative relaxation, although the detail of the process is still unknown.
Results in Fig. 4 showed that required amount of DNA-CTMA for the highest emission was smaller than pure DNA. For the interaction with DNA fluorescence intensity gradually increased until DNA ratio exceeded 30 times of the dyes, while steep increase from 3 fold and saturation at about 10 fold of molar amount characterized the interaction with DNA-CTMA in ethanol solutions. Since 30 times or more DNA was required to make emission intensity saturate, it means that such large amounts of DNA were necessary to store molecules in sites far from each other enough to prevent aggregation or strand distortion. On the other hand, higher concentration of the dyes could be incorporated into DNA-CTMA, and the fact is preferable for applications usually utilizing the complex films. Such a high storing capacity of DNA-CTMA was also demonstrated for an azobenzene derivative and explained through so-called semi-intercalation model [22].
As given in Fig. 5, absorption peak of p-hemi1 and o-hemi1 in films strongly depended on fabrication method. Dye concentration of these films was 5wt% for all. In both cases, immersion made films gave longer wavelength about 20~30nm than those for conventionally made ones. Emission peaks also gave similar shifts. It is not very clear what the difference between the two methods induced such a big change, because in both cases dye molecules were incorporated into the matrix with post process where strong coupling of DNA and CTMA has already made.
Fig. 5 Absorption and emission spectra under 440nm excitation for p- and o-hemi1 prepared with conventional and immersion methods. Fluorescence curves were normalized with peak values.
Absorption and fluorescence spectra for the films with various dye concentration were measured. Absorption intensity was proportional to dye concentration for p- and o-hemi1 prepared with the two ways, meaning that designated amounts of the dye were incorporated in the films even with the immersion. While the spectral shape did not depend on the concentration so much for the conventionally made samples, the peak positions for immersion samples moved to shorter side with the concentration. The shift direction suggested the molecular interaction in H-type alignment. Fluorescence quenching with concentration also supported the existence of such intermolecular interactions, because the quenching were more significant for the immersion made films. Therefore, molecules seemed to be well dispersed for the conventional case, but the presumed aggregation did not have negative effects for laser performance as shown in the next section.
4. Laser oscillation from hemicyanine doped complex films
Laser oscillation was demonstrated with all the samples prepared by the two methods and their threshold intensities are summarized in Table 1. The values about 0.07~0.2mJ/cm2 were the lowest among hemicyanines we studied and very close to the record achieved by Yu et al. with a persistent grating [23]. Emission spectra indicated lasing from two p-hemi1 samples and the linewidth was as narrow as 2nm.
Table 1. Threshold values and lasing lifetime for dye doped DNA complex films prepared with two methods.
Durability was estimated from the evolution of emission spectrum obtained by continual measurement under the pumping with the intensity of 0.5mJ/cm2. Emission spectra for o-hemi1 devices made by the two ways were compared in Fig. 6, showing that wavelength purity was superior for immersion films as no ASE was included. The other panel of Fig. 6 shows the temporal evolution of emission intensity. Initial intensity was several hundred times stronger for the immersion made devices. The same tendency was observed for all three types of the dyes.
Fig. 6 (a) Emission spectra for two o-hemi1 devices observed under the pumping above the threshold. (b) Spectral peak intensity as a function of time.
Reduction of laser intensity occurred in minutes and the intensity became less than one tenth before the cease of lasing. Succeeding ASE also stopped in 1~8min., followed by usual fluorescence emission of which intensity decayed with much slower rate. Transition from the ASE to fluorescence was estimated from width change of the emission spectra. In our former work, ASE continued more than one hour for similar devices and under similar pumping condition to this case [18]. In order to maintain lasing in population grating scheme, enough gain modulation for DFB was required as well as basal gain necessary for amplification. Shorter ASE lifetime might be caused by continual stimulated emission during lasing, because the reduction was relatively more remarkable for the immersion made samples giving strong emission.
5. Conclusion
Absorption spectrum change observed in hemicyanine solutions with the addition of DNA indicated that strong coupling occurred in the ground state. On the other hand, change in fluorescence spectrum with the addition of DNA-complex suggested the interaction in excited states. The most significant emission enhancement was observed in the dye interacting with the DNA complex in ethanol solutions. Optical properties of the dyes in solid state DNA-complex films depended on the manufacturing mean. Films made by immersion method showed superior laser properties as low threshold, high intensity, and good durability for all three types of hemicyanines we examined, compared to those made with usual way.
Acknowledgments
A part of this work was conducted with equipment supported by Nanotechnology Platform Program (Synthesis of Molecules and Materials) of MEXT, Japan. We thank to Daiki Ochi for providing a program for emission data analysis.
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