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Article

Synthesis of Silane-Based Poly(thioether) via Successive Click Reaction and Their Applications in Ion Detection and Cell Imaging

by
Zhiming Gou
,
Xiaomei Zhang
,
Yujing Zuo
and
Weiying Lin
*
Institute of Fluorescent Probes for Biological Imaging, School of Chemistry and Chemical Engineering, School of Materials Science and Engineering, University of Jinan, Shandong 250022, China
*
Author to whom correspondence should be addressed.
Polymers 2019, 11(8), 1235; https://doi.org/10.3390/polym11081235
Submission received: 19 June 2019 / Revised: 2 July 2019 / Accepted: 3 July 2019 / Published: 25 July 2019
(This article belongs to the Special Issue Fluorescent polymers for sensing and imaging)
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Abstract

:
A series of poly(thioether)s containing silicon atom with unconventional fluorescence were synthesized via successive thiol click reaction at room temperature. Although rigid π-conjugated structure did not exist in the polymer chain, the poly(thioether)s exhibited excellent fluorescent properties in solutions and showed visible blue fluorescence in living cells. The strong blue fluorescence can be attributed to the aggregation of lone pair electron of heteroatom and coordination between heteroatom and Si atom. In addition, the responsiveness of poly(thioether) to metal ions suggested that the selectivity of poly(thioether) to Fe3+ ion could be enhanced by end-modifying with different sulfhydryl compounds. This study further explored their application in cell imaging and studied their responsiveness to Fe3+ in living cells. It is expected that the described synthetic route could be extended to synthesize novel poly(thioether)s with superior optical properties. Their application in cell imaging and ion detection will broaden the range of application of poly(thioether)s.

1. Introduction

Poly(thioether) are important base polymers that exhibit excellent physical and chemical properties, such as good chemical and weather resistance, high impermeability, and high heat insulation properties [1]. These excellent properties made it to become a promising functional polymer for the application of optical materials, medical impression materials ion detection probes, energy storage and conversion materials [2,3,4]. Thus far, many methods have been developed for the ion detection, including microelectrode, absorption spectroscopy [5]. However, these methods are difficult to dynamically monitor cellular ion changes. By contrast, fluorescence spectroscopy has become a powerful tool for fluorescent labeling, ion sensing and cell imaging due to its high sensitivity, excellent selectivity and dynamic monitoring [6,7]. However, as one kind of functional polymer, poly(thioether)s are rarely applied in cell imaging, especially for Fe ion detection in living cells.
At present, two synthetic routes were adopted to prepare poly(thioether). One is the ring-opening polymerization of episulfide [8,9], and the other is thiol click polymerization between dithiols and dienes or dithiols and alkyne [10,11,12]. As one of most widely used and important reactions, the thiol click reaction has served as useful tool to fabricated dendrimer and block polymer [13,14], and also has been used for end-/side-group functionalization and surface modification [15,16].
Trimethylsilylacetylene (TMSA) is a versatile precursor to prepare important intermediates or unsaturated compounds with unique properties, and also to be used as a protecting group for functional modification. NaKa group and Miura group reported the synthesis of silyl-substituted thiophene derivatives and silyl-substituted fulvene derivatives based on TMSA, respectively [17,18]. Yokozawa et al. described the synthesis of ethynyl-functionalized poly(3-hexylthiophene) with one terminal ethynyl group, which has been very suitable for subsequent preparation of block copolymers via a click reaction [19]. Grirrane et al. synthesized a novel family of dipropargylamines through a double catalytic A3-coupling of primary amine, formaldehyde and TMSA, and subsequent deprotection of two terminal trimethylsilyl (TMS) groups endow them with high values of post-modification [20]. Furthermore, Alejandro and Pilar reported metal-monolayer-metal molecular electronic devices based on oligoynes (Me3Si–(C=C)4–SiMe3) through fluoride-induced deprotection of terminal trimethylsilyl (TMS) groups [21,22], and this nascent surface modification technique provided new perspectives for the fabrication of molecular electrochemical sensors.
Among the above reports, studies on poly(thioether)s using TMSA are rare. Here, this study reported the synthesis of functionalized poly(thioether)s containing silane via a two-step successive addition reaction (Scheme 1). Firstly, the addition of trimethylsilylacetylene (TMSA) to a dithiol with different molar ratio yielded a series of poly(thioether)s. Secondly, vinyl poly(thioether) (P1 and P2) was selected as intermediates, and reserved vinyl groups further reacted with thiol and finally gained the functionalized poly(thioether)s. The authors further explored their fluorescence properties and used them in cell imaging and ion detection. It was found that the post-functionalization of poly(thioether)s obviously enhanced their responsiveness to Fe3+ and avoided the disturbance of Fe2+. The obtained poly(thioether)s and post-functionalized products provided a simple route to synthesize sulfur-containing organosilicone polymer with superior optical properties and extended their application in the field of cell imaging and ion detection.

2. Materials and Experiments

2.1. Materials

Trimethylsilylacetylene (TMSA), 2,2-dimethoxy-2-phenylacetophenone (DMPA), 1,2-ethanedithiol (EDT), and 3,6-dioxa-1,8-octanedithiol (DODT) were purchased from Aladdin Co. (Shanghai, China) and used directly. Furfuryl mercaptan, trimethoxysilylpropanethiol, and methyl 3-mercaptopropionate were supplied by Macklin biochemical (Shanghai, China) Co., Ltd. and used as received. All solvents were bought from Chemical Technology (Shanghai, China) Co., Ltd. and directly used without further purification.

2.2. Characterization and Measurements

NMR (1H and 13C) spectra were recorded by Bruker AVANCE 400 MHz using CDCl3 as solvents, and without tetramethylsilane as the internal standard. Thermogravimetric analysis (TGA) was measured using SDTQ 600 at 10 °C·min−1 under N2. The luminescence emission spectra of the poly(thioether)s were recorded by using a Hitachi F–4500 fluorescence spectrophotometer with Xe lamp as an excitation source. The data of molecular weights was measured by gel permeation chromatography (GPC) using a Waters 515 liquid chromatograph with a refractive index detector 2414 and using tetrahydrofuran (THF) as the elution solvent.

2.3. Synthesis of Poly(thioether)

The synthetic route and structure of poly(thioether)s have been shown in Scheme 1. The detailed procedure for P1 was as follows: TMSA (0.98 g, 10.0 mmol), EDT (0.75 g, 8.0 mmol) and DMPA (1 wt%) were placed in a glass vessel. The above mixture was exposed to a UV light source (365 nm, 100 w) with stirring for 20 min at room temperature. The crude product was washed three times with cold methanol and then reduced pressure distillation. The obtained light-yellow solid was P1. Yield: 96%. A series of poly(thioether)s (P2, P3, P4, P5, and P6) were synthesized according to the same above reaction condition and the treating process.
The data of P1: 1H-NMR (400 MHz, CDCl3, ppm): δ = 0.05–0.20 (-SiCH3), 2.05 (-SiCHCH2S-), 2.69–3.14 (-SiCHCH2S-, -SCH2CH2S-), 5.72 and 5.77 (s, -SiCH2=CH2S-), 6.47 and 6.52 (s, -SiCH2=CH2S-). 13C-NMR (400 MHz, CDCl3, ppm): δ = −3.0 (SiCH3), 31.6 (-SiCHCH2S-), 32.8 and 36.5 (-SCH2CH2S-, -SiCHCH2S-), 110.7 and 133.0 (-SiCH2=CH2S-).
The data of P2: 1H-NMR (400 MHz, CDCl3, ppm): δ = 0.07–0.13 (-SiCH3), 2.07 (-SiCHCH2S-), 2.75–2.95 (-SiCHCH2S-, -SCH2CH2O-), 3.03–3.08 (-OCH2CH2O-), 3.61–3.70 (-SCH2CH2O-), 6.49 and 6.53 (-SiCH2=CH2S-). 13C-NMR (400 MHz, CDCl3, ppm): 2.06–2.07 (SiCH3), 29.7 and 32.89 (-OCH2CH2S-), 31.7 (-SiCHCH2S-), 36.7 (-SiCHCH2S-), 70.7 (-OCH2CH2O-), 70.9 (-SCH2CH2O-), 110.9 and 133.1 (-SiCH2=CH2S-). Yield: 95%.
The data of P3: 1H-NMR (400 MHz, CDCl3, ppm): δ = 0.06–0.19 (m, -SiCH3), 2.07 (-SiCHCH2S-), 2.79–3.09 (-SiCHCH2S-, -SCH2CH2S-). 13C-NMR (400 MHz, CDCl3, ppm): −3.0 (SiCH3), 24.6 (-SCH2CH2SH), 31.6 (-SiCHCH2S-), 32.9 (-SCH2CH2S-), 33.7 (-SCH2CH2SH), 36.5 (-SiCHCH2S-). Yield: 97%.
The data of P4: 1H-NMR (400 MHz, CDCl3, ppm): δ = 0.07–0.13 (-SiCH3), 2.07 (-SiCHCH2S-), 2.73–2.91 (-SiCHCH2S-, -SCH2CH2O-), 3.02–3.06 (-OCH2CH2O-), 3.62–3.68 (-SCH2CH2O-). 13C-NMR (400 MHz, CDCl3, ppm): 2.06–2.07 (SiCH3), 29.8 and 32.9 (-OCH2CH2S-), 31.8 (-SiCHCH2S-), 36.9 (-SiCHCH2S-), 70.8 (-OCH2CH2O-), 70.9 (-SCH2CH2O-). Yield: 93%.
The data of P5: 1H-NMR (400 MHz, CDCl3, ppm): δ = 0.05–0.16 (m, -SiCH3), 1.35 (-SCH2CH2SH), 2.05 (-SiCHCH2S-), 2.69–3.05 (-SiCHCH2S-, -SCH2CH2S-). 13C-NMR (400 MHz, CDCl3, ppm): −1.76 (SiCH3), 24.6 (-SCH2CH2SH), 31.6 (-SiCHCH2S-), 32.9 (-SCH2CH2S-), 33.7 (-SCH2CH2SH), 36.5 (-SiCHCH2S-). Yield: 92%.
The data of P6: 1H-NMR (400 MHz, CDCl3, ppm): δ = 0.07–0.16 (-SiCH3), 1.34 (-OCH2CH2SH), 2.09 (-SiCHCH2S-), 2.74–3.07 (-SiCHCH2S-, -SCH2CH2O-), 3.62–3.64 (-OCH2CH2O-), 3.70–3.75 (-SCH2CH2O-, -OCH2CH2SH). 13C-NMR (400 MHz, CDCl3, ppm): 2.07 (SiCH3), 21.0 (-OCH2CH2SH), 29.6 and 32.9 (-OCH2CH2S-), 67.1–71.3 (-OCH2CH2O-, -SCH2CH2O-, -OCH2CH2SH). Yield: 94%.
Methyl-mercaptopropionate, trimethoxysilylpropanethiol and furfuryl mercaptan were selected and used to the functionalization of P1 and P2. Taking methyl-mercaptopropionate as an example, the above obtained P1, methyl-mercaptopropionate (4.0 mmol) and DMPA (1 wt%) were dissolved in THF (2 mL) and reacted following the above-mentioned procedure. The crude product was washed three times with cold methanol and then reduced pressure distillation. The obtained light-yellow solid was P1-1. Yield: 93%. A series of functionalized poly(thioether)s (P1-1, P1-2, P1-3, P2-1, P2-2, and P2-3) were synthesized according to the same procedure.
The data of P1-1: 1H NMR (400 MHz, CDCl3): 0.03–0.09 (SiCH3), 2.21 (SiCHCH2S), 2.47–2.51(SCH2CH2COOCH3), 2.60–2.99 (SiCHCH2S, SCH2CH2S), 3.61 (COOCH3). 13C-NMR (400 MHz, CDCl3, ppm): −3.01 (SiCH3), 29.6 (SCH2CH2COCH3), 31.61 (SCH2CH2O), 32.7 (SiCHCH2S), 33.4 (SCH2CH2COCH3), 36.5 and 38.1 (SiCHCH2S), 69.8 (OCH2CH2S), 172.1 (COOCH3).
The data of P1-2: 1H NMR (400 MHz, CDCl3): 0.01–0.10 (SiCH3), 2.23 (SiCHCH2S), 2.51 and 3.18 (SiCHCH2S), 2.58–3.01 (SCH2CH2S), 3.33–3.43 (SCH2CO), 6.27 and 6.40 (C=CH2CH2CH2O), 7.58 (C=CH2CH2CH2O). 13C NMR (400 MHz, CDCl3): −1.75 (SiCH3), 30.0 (SCH2CO), 31.0 and 31.6 (SCH2CO), 32.6 and 34.6 (SiCHCH2S), 36.4(SiCHCH2S), 106.9–111.1 (-C=CHCH=CHO-), 142.3 (-C=CHCH=CHO-), 151.4 (-C=CHCH=CHO-). Yield: 94%.
The data of P1-3: 1H NMR (400 MHz, CDCl3): 0.01–0.15 (SiCH3), 0.56–0.74 (OSiCH2CH2CH2S), 1.53–1.66 (OSiCH2CH2CH2S), 2.17–2.26 (SiCHCH2S), 2.43-2.52 and 3.18 (SiCHCH2S), 2.63–3.04 (SCH2CH2S), 3.41–3.50 (OCH3). 13C NMR (400 MHz, CDCl3): −2.0 (SiCH3), 8.35 (SiCH2CH2CH2S), 9.65 (SiCH2CH2CH2S), 31.4 and 31.7 (SCH2CH2S, SiCHCH2S), 33.1 (SiCH2CH2CH2S), 36.7 (SiCHCH2S), 48.9 and 50.2 (OCH3). Yield: 93%.
The data of P2-1: 1H NMR (400 MHz, CDCl3): 0.07–0.10 (SiCH3), 2.23 (SiCHCH2S), 2.59–2.65 (SCH2CH2O), 2.66–2.76 (SiCHCH2S), 2.78–2.85 (SCH2CH2CO,), 3.52 (OCH2CH2O), 3.54–3.60 (SCH2CH2O), 3.61 (OCH3). 13C-NMR (400 MHz, CDCl3, ppm): −1.89 and −0.53 (SiCH3), 29.8 (SCH2CH2COCH3), 31.61 (SCH2CH2O), 32.9 (SiCHCH2S), 33.6 (SCH2CH2COCH3), 36.7 and 38.3 (SiCHCH2S), 51.9 (COOCH3), 69.8 (OCH2CH2O), 70.9 (OCH2CH2S), 172.3 (COOCH3). Yield: 92%.
The data of P2-2: 1H NMR (400 MHz, CDCl3): 0.01–0.09 (SiCH3), 2.24 (SiCHCH2S), 2.67–2.76 (SCH2CH2O), 2.51 and 2.76–2.92 (SiCHCH2S), 3.52–3.54 (SCH2CO), 3.55–3.59 (OCH2CH2O), 3.75–3.95 (SCH2CH2O), 6.26 and 6.38 (-C=CHCH=CHO-), 7.57 (-C=CHCH=CHO-). 13C NMR (400 MHz, CDCl3): −1.74 (SiCH3), 29.7 (SCH2CO), 31.0 and 31.8 (SCH2CH2O), 32.8 and 34.7 (SiCHCH2S), 36.6(SiCHCH2S), 70.0 (OCH2CH2O), 70.8 (SCH2CH2O), 106.9–111.0 (-C=CHCH=CHO-), 142.7 (-C=CHCH=CHO-), 151.8 (-C=CHCH=CHO-). Yield: 94%.
The data of P2-3: 1H NMR (400 MHz, CDCl3): 0.07–0.10 (SiCH3), 0.58–0.72 (OSiCH2CH2CH2S), 1.55–1.62 (SiCH2CH2CH2S), 2.24 (SiCHCH2S), 2.43–2.52 and 2.99 (SiCHCH2S), 3.42 (SCH2CO), 3.48–3.59 (OCH2CH2O), 3.50–3.54 (OCH3), 3.42–3.59 (SCH2CH2O). 13C NMR (400 MHz, CDCl3): −1.9 (SiCH3), 8.33 (SiCH2CH2CH2S), 9.67 (SiCH2CH2CH2S), 31.3 and 31.7 (SCH2CH2O, SiCHCH2S), 32.9 (SiCH2CH2CH2S), 36.7 (SiCHCH2S), 48.9 and 50.1 (OCH3), 70.0 (OCH2CH2O), 70.9 (SCH2CH2O). Yield: 91%.

2.4. Fluorescence Property and Ion Detection

Poly(thioether)s were dissolved in ethanol (0.3 mg/mL) and their fluorescent emission spectra were measured under excitation wavelengths of 330 nm.
The responsiveness of poly(thioether)s to ions were detected by adding corresponding chlorine salt (100 μL, 1 mmol/L) to poly(thioether)s solution (900 μL, 0.2 mg/mL).
The HeLa cells were incubated with 4.0 μg of poly(thioether)s for 20 min at 37 °C and washed with phosphate buffer solution (PBS). The cell images of poly(thioether)s in HeLa cells were observed under a confocal microscope after adding new culture medium. In addition, the responsiveness of poly(thioether)s to Fe3+ in HeLa cells were measured by successively adding 10 μL, 30 μL, 50 μL and 70 μL Fe3+ ion (1 mmol/L), respectively. The response process was observed in situ under confocal microscopy.

3. Results and Discussion

3.1. Synthesis of Poly(thioether)s and Post-Functionalization

A series of poly(thioether)s were synthesized using different molar ratio (1:0.8, 1:1.0, and 1:1.2) between TMSA and dithiol (Scheme 1). In view of the same reaction condition and purification step, 1,2-ethanedithiol based poly(thioether)s (P1, P3, and P5) was selected to be analyzed and their 1H NMR have been shown in Figure 1. The characteristic peak of alkenyl groups (-CH=CH-) appeared at approximately 5.75 ppm and 6.5 ppm only existed in the polymer when the molar ratio was 1:0.8. By the same token, the characteristic peak of sulfhydryl groups (-SH) appeared at approximately 1.7 ppm and was presented when the molar ratio was 1:1.2. The appearance of the peak at 1.7 ppm when the molar ratio was 1:1 could be attribute to the low boiling point of TMSA, which resulted in the volatilization of part of TMSA in the reaction process. In addition, the thiol-alkyne reaction proceeded under a radical step-growth mechanism involving a two step addition. Therefore, the dithiol radicals can react with alkynyl groups from different active sites and generate two types of product structures (α-addition and β-addition) which exist simultaneously in the polymer backbone. The appearance of peak at 1.35 ppm belonged to α-addition products. However, a small amount of α-addition could not have had a significant influence on the fluorescent property of poly(thioether)s. This is because the integral structure of linear polymer chain is likely to remain unchanged. 1H-NMR data indicates that the thiol-alkyne click reaction is a simple and an efficient approach to prepare poly(thioether)s with a special functional group by simply changing the molar ratio. Further, the remained end functional group could be used for further modification.
Theoretically, the reaction with a molar ratio of 1:1 is likely to react completely and maximize the molecular weight of polymers. However, the obtained polymers in this situation did not have reactive sites. When one monomer is excessive, the non-stoichiometric alkyne-to-dithiol molar ratio may result in a handful of functional groups in chain end of polymers. The polymers can possess short polymer chains and low molecular weight. In addition, these maintained functional groups could be used for post-functionalization and naturally various polymers with interesting properties. Carbon-carbon double bonds were maintained in chain end when the molar ratio was 1:0.8, and unreacted sulfhydryl groups were maintained when the molar ratio was 1:1.2. Moreover, the excessive dithiol can lead to the polymers possessing shorter chain lengths than excessive alkyne. Therefore, among the three different molar ratios, the sequence of their molecular weight may be as follows: Mw (1:1) > Mw (1:0.8) > Mw (1:1.2). This speculation has been confirmed by the test outcome. The molecular weight of polymers was measured by GPC and the detailed data have been depicted in Table 1.
The thiol-alkyne reaction proceeded via radical step-growth involving two successive addition processes. The first addition generates an intermediate that contains alkenyl group, and subsequent thiol-alkene reaction yields poly(thioether)s. When one monomer is excessive, the non-stoichiometric alkyne-to-dithiol molar ratio can result in a handful of functional groups in the chain end of polymers. Poly(thioether)s containing alkenyl (P1 and P2) were selected to further modification. A series of functionalized poly(thioether)s (P1-1, P1-2, P1-3, P2-1, P2-2, and P2-3) were synthesized with different sulfhydryl compounds via thiol-alkene click reaction (Scheme 1), and a detailed synthetic procedure was described in the experimental section. The modified sulfhydryl compound can bring new and interesting properties to poly(thioether)s, although the number of modified functional groups is very small.

3.2. Fluorescent Properties

Poly(thioether)s (P1, P2, P3, P4, P5, and P6) and functionalized poly(thioether)s (P1-1, P1-2, P1-3, P2-1, P2-2, and P2-3) emit a blue color in the solution and their emission spectra excited by 330 nm as shown in Figure 2. P1, P3, and P5 were synthesized with different molar ratios between TMSA and EDT, their fluorescence intensity (Figure 2a) decreased sharply as the molar ratio of dithiol increased. While P2, P4, and P6 were synthesized with different molar ratios between TMSA and DODT, their fluorescence intensity (Figure 2b) slightly decreased as the molar ratio of dithiol increased. The unconventional fluorescence can be attributed to the aggregation of long pair electrons of heteroatom (O atom and S atom). Meanwhile, the coordination bonds of O → Si and S → Si were both conducive to cause strong blue photoluminescence in poly(thioether)s [23,24]. The aggregation and coordination in poly(thioether)s containing the O atom (P2, P4, and P6) were stronger than poly(thioether)s without the O atom (P1, P3, and P5). The interaction between heteroatom and silicon atom resulted in the significant difference of fluorescence intensity among poly(thioether)s (P1, P3, and P5) and negligible difference of fluorescence intensity among poly(thioether)s (P2, P4, and P6).
The introduction of sulfhydryl compounds can bring new properties to poly(thioether)s (P1 and P2). The end modified poly(thioether)s demonstrated different trends of fluorescent intensity. The functionalized poly(thioether)s (P1-1, P1-2, and P1-3) based on P1 showed obvious fluorescence quenching after the end-modification (Figure 2c). The introduction of three sulfhydryl compounds all resulted in the decreasing of fluorescence intensity, especially to methyl-mercaptopropionate. The functionalized poly(thioether)s (P2-1, P2-2, and P2-3) based on P2 showed slight fluorescence enhancement after the end-modification (Figure 2d). The introduction of three sulfhydryl compounds did not show a significant difference to the fluorescence intensity of P1. The difference between two types of functionalized poly(thioether)s indicated that the end modified functional groups had an obvious influence on the fluorescent property of poly(thioether)s, although the number of modified functional groups was very small. The different effect of the end modified groups to P1 and P2 could be attributed to the structure of the polymer chain. The oxygen atom in the main chain enhanced the structural stability of P2 and weakened the influence of the end modified groups on the fluorescent properties of poly(thioether)s. The above results indicated that the fluorescent properties of functionalized poly(thioether)s were influenced by the end modified groups and the structure of the polymer chain.

3.3. Ion Detection and Cell Imaging

As one types of most important bioactive substances in biological systems, metal cations are vital to the stability and effectivity of the immune system. To study the responsiveness of poly(thioether)s to various cations, a series of chloride salts (Al3+, Ba2+, Ca2+, Co2+, Cu2+, Fe2+, Fe3+, Mg2+, Ni2+, and Sn2+) were added to the solutions of poly(thioether)s (P1 and P2) and their end-modified derivatives. As shown in Figure 3a,e, the fluorescence intensity of P1 and P2 both decreased at different levels after adding metal ions, and especially for Fe2+ and Fe3+. This behavior of fluorescence quenching of poly(thioether)s after adding metal ions can be attributed to the coordination between heteroatom (O atom and S atom) and metal ion, especially for Fe3+. This coordination destroyed the interaction between heteroatom and Si atom, and subsequently decreased the aggregation extent of the long pair electrons of heteroatom. These two factors described above resulted in the fluorescence quenching of poly(thioether)s.
In addition, the end-modified with different sulfhydryl compounds resulted in significant differences between their responsiveness to Fe2+ and Fe3+. The introduction of methyl 3-mercaptopropionate did not bring obvious changes of P1-1 to iron ions (Fe2+ and Fe3+) compared to P1 (Figure 3b), while P2-1 had negligible selectivity to Fe2+ and Fe3+ compared to P2 (Figure 3f). However, the introduction of furfuryl mercaptan and trimethoxysilylpropanethiol significantly enhanced the selective responses of P1 and P2 derivates to Fe3+. It can be observed that P1 derivates (Figure 3c,d) and P2 derivates (Figure 3g,h) all exhibited good selective to Fe3+. The above results suggested that end-modified functional groups with a very small amount could obviously change the fluorescent properties of poly(thioether)s.
Regarding the good selectivity of functionalized poly(thioether)s to Fe3+, this study further explored their fluorescence behaviors in living cells. First, the HeLa cells were stained with poly(thioether)s (P1 to P6) for 20 min at 37 °C and washed with PBS solution. Their confocal fluorescence images were observed and depicted in Figure 4. The images show visible blue fluorescence in the dark filed and mainly distributed in the cytoplasm. The distribution was due to the way the poly(thioether)s entered into the cell. The polymers enter the cell by simple diffusion. The blue fluorescence of poly(thioether)s belonged to unconventional fluorescence because rigid π-conjugated structures and heterocycle structures did not exist in the obtained poly(thioether)s. Furthermore, silicon atoms in the polymer chain could enhance their fluorescent properties due to their empty 3d orbital [25]. The fluorescence intensity of poly(thioether)s containing the O atom (P2, P4 and P6) in cells were stronger than poly(thioether)s without containing the O atom (P1, P3 and P5). This may relate to the structure of the polymer chain. The O atom in the main chain enhanced the coordination in polymers, therefore poly(thioether)s containing O atom exhibited stronger fluorescence in living cells. In addition, it can be observed that some poly(thioether)s absorbed on the surface of the cells in the bright filed. The results suggest that poly(thioether)s could easily enter living cells and could be absorbed on the surface of the cells.
To further investigate the responsiveness of poly(thioether)s to Fe3+ in living cells, P1 and P2 derivates were selected as a sample and their fluorescent quenching behaviors were observed in situ under confocal microscopy. HeLa cells were stained only with poly(thioether)s and their cell images were observed under confocal microscopy. The responsiveness of poly(thioether)s to Fe3+ in HeLa cells were measured by successively adding Fe3+ ions. The fluorescence intensity of HeLa cells in the blue field was calculated by confocal microscopy. Confocal fluorescence images of P1 and their derivates were observed as shown in Figure 5. With the increasing Fe3+ concentrations, the fluorescence intensity of P1 remained roughly unchanged. Meanwhile, the end-modification with different sulfhydryl compounds brought obvious responsiveness to Fe3+. P1-1 showed slightly fluorescence quenching after adding successive concentrations of Fe3+. P1-2 and P1-3 both exhibited obvious fluorescence quenching with the increasing Fe3+ ions, especially for P1-3. Similarly, the fluorescence intensity of P2 remained roughly the same with the increasing Fe3+ concentrations (Figure 6). Three types of end-modified poly(thioether)s show a similar trend in the change of fluorescence quenching.

3.4. Thermogravimetric Analysis (TGA)

To evaluate the thermal stability of poly(thioether)s, P1 and P2 were selected and their thermodynamics properties were measured by TGA analysis at 10 °C·min−1 in N2. The TGA data indicated that the poly(thioether)s exhibited high thermal stability (Figure 7). The thermal decomposition temperature (Td) of P1 and P2 were at approximately 290 °C and 320 °C, respectively. P2 possessed higher Td than P1, which indicated that the presence of oxygen atoms in the polymer chain increased the thermal performance of poly(thioether)s. In addition, P1 exhibited two degradation steps. The first weight loss step of P1 before 250 °C could be ascribed to the sublimation of low molecular weight of poly(thioether)s. The second weight loss of P1 can be attributed to the decomposition and sublimation of the main chain. The high Td of P1 and P2 suggested that the good thermal stability of poly(thioether)s, which was prepared by simple thiol-alkyne click reaction at room temperature.

4. Conclusions

A series of silane-based poly(thioether)s and functionalized poly(thioether)s were synthesized via successive thiol click reaction. These poly(thioether)s exhibited excellent fluorescent properties in solutions and showed visible blue fluorescence in living cells. The responsiveness of poly(thioether)s to metal ions can be adjusted by end-modifying with different sulfhydryl compounds. The functionalized poly(thioether)s showed high selectivity to Fe3+ detection and still exhibited responsiveness to Fe3+ ions in living cells. The synthetic route of poly(thioether)s described herein could be extended to synthesize novel organosilicone fluorescent materials, and their application in ion detection and cell imaging can extend the application field of poly(thioether)s.

Author Contributions

W.L. directed the research, Z.G. and Y.Z. designed the experiments, X.Z. and Z.G. performed the experiments, Z.G., Y.Z. and W.L. wrote the paper.

Funding

This work was financially supported by NSFC (21877048, 21472067, 21672083), Taishan Scholar Foundation (TS201511041), the startup fund of University of Jinan (309-10004, 160100332, 140200321), and Natural Science Foundation of Shandong Province (ZR2018BB022).

Conflicts of Interest

There are no conflict to declare.

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Polymers 11 01235 sch001 550
Scheme 1. The synthesis route of functionalized polythioether containing silane.
Scheme 1. The synthesis route of functionalized polythioether containing silane.
Polymers 11 01235 sch001
Polymers 11 01235 g001 550
Figure 1. 1H NMR data of (a) P1, (b) P3, and (c) P5.
Figure 1. 1H NMR data of (a) P1, (b) P3, and (c) P5.
Polymers 11 01235 g001
Polymers 11 01235 g002 550
Figure 2. Fluorescence spectra of poly(thioether)s (a) P1, P3 and P5, (b) P2, P4 and P6, and functionalized poly(thioether)s (c) P1-1 to P1-3 and (d) P2-1 to P2-3 at 0.2 mg/mL final concentration in ethanol solution (slits = 10 nm).
Figure 2. Fluorescence spectra of poly(thioether)s (a) P1, P3 and P5, (b) P2, P4 and P6, and functionalized poly(thioether)s (c) P1-1 to P1-3 and (d) P2-1 to P2-3 at 0.2 mg/mL final concentration in ethanol solution (slits = 10 nm).
Polymers 11 01235 g002
Polymers 11 01235 g003 550
Figure 3. Fluorescence intensity response of functionalized poly(thioether)s (a) P1, (b) P1-1, (c) P1-2, (d) P1-3, (e) P2, (f) P2-1, (g) P2-2, and (h) P2-3 to different metal cations (0.1 mM) in ethanol solution. (λex = 330 nm, slits = 10 nm).
Figure 3. Fluorescence intensity response of functionalized poly(thioether)s (a) P1, (b) P1-1, (c) P1-2, (d) P1-3, (e) P2, (f) P2-1, (g) P2-2, and (h) P2-3 to different metal cations (0.1 mM) in ethanol solution. (λex = 330 nm, slits = 10 nm).
Polymers 11 01235 g003
Polymers 11 01235 g004 550
Figure 4. Confocal fluorescence images of (a) P1, (b) P2, (c) P3, (d) P4, (e) P5, and (f) P6 in HeLa cells. bar = 20 μm.
Figure 4. Confocal fluorescence images of (a) P1, (b) P2, (c) P3, (d) P4, (e) P5, and (f) P6 in HeLa cells. bar = 20 μm.
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Polymers 11 01235 g005 550
Figure 5. Confocal fluorescence images and quantified relative fluorescence intensity of P1 and their derivates after adding successive concentrations of Fe3+ ions in HeLa cells. bar = 20 μm. The statistical analyses were performed with student’s t-test (n = 4). *3.37 × 10−8 < p < 1.57 × 10−4 and the error bars represent standard deviations (± S. D.).
Figure 5. Confocal fluorescence images and quantified relative fluorescence intensity of P1 and their derivates after adding successive concentrations of Fe3+ ions in HeLa cells. bar = 20 μm. The statistical analyses were performed with student’s t-test (n = 4). *3.37 × 10−8 < p < 1.57 × 10−4 and the error bars represent standard deviations (± S. D.).
Polymers 11 01235 g005
Polymers 11 01235 g006 550
Figure 6. Confocal fluorescence images and quantified relative fluorescence intensity of P2 and their derivates after adding successive concentrations of Fe3+ ion in HeLa cellss. bar = 20 μm. The statistical analyses were performed with student’s t-test (n = 4). *5.2 × 10−9 < p < 2.39 × 10−6 and the error bars represent standard deviations (± S. D.).
Figure 6. Confocal fluorescence images and quantified relative fluorescence intensity of P2 and their derivates after adding successive concentrations of Fe3+ ion in HeLa cellss. bar = 20 μm. The statistical analyses were performed with student’s t-test (n = 4). *5.2 × 10−9 < p < 2.39 × 10−6 and the error bars represent standard deviations (± S. D.).
Polymers 11 01235 g006
Polymers 11 01235 g007 550
Figure 7. Thermogravimetric analysis (TGA) curves of P1 and P2.
Figure 7. Thermogravimetric analysis (TGA) curves of P1 and P2.
Polymers 11 01235 g007
Table
Table 1. Gel permeation chromatography (GPC) data for the poly(thioether)s (P1 to P6).
Table 1. Gel permeation chromatography (GPC) data for the poly(thioether)s (P1 to P6).
SampleMn
(g.mol−1)		Mw
(g.mol−1)		Mz
(g.mol−1)		PDIMz/Mw
P13210400049101.241.23
P23400432053801.271.24
P3942014950210201.581.40
P42280044460783601.951.76
P51860199021501.071.08
P62340280034401.191.23

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MDPI and ACS Style

Gou, Z.; Zhang, X.; Zuo, Y.; Lin, W. Synthesis of Silane-Based Poly(thioether) via Successive Click Reaction and Their Applications in Ion Detection and Cell Imaging. Polymers 2019, 11, 1235. https://doi.org/10.3390/polym11081235

AMA Style

Gou Z, Zhang X, Zuo Y, Lin W. Synthesis of Silane-Based Poly(thioether) via Successive Click Reaction and Their Applications in Ion Detection and Cell Imaging. Polymers. 2019; 11(8):1235. https://doi.org/10.3390/polym11081235

Chicago/Turabian Style

Gou, Zhiming, Xiaomei Zhang, Yujing Zuo, and Weiying Lin. 2019. "Synthesis of Silane-Based Poly(thioether) via Successive Click Reaction and Their Applications in Ion Detection and Cell Imaging" Polymers 11, no. 8: 1235. https://doi.org/10.3390/polym11081235

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