JP2021031598A - Pyrene fluorochrome - Google Patents

Abstract

To provide a fluorochrome in which the maximum wavelengths of excitation light and fluorescence are relatively long wavelengths, and which is excellent in light stability.SOLUTION: A pyrene fluorochrome is represented by the formula (I) [where R1 is a C1-18 alkyl group or the like, X1 is >CR2R3 or the like (where R2 and R3 independently represent a C1-18 alkyl group), Y is a group having a specific polymethine structure].SELECTED DRAWING: None

Description

The present invention relates to a fluorescent dye having a relatively long maximum absorption wavelength and maximum fluorescence wavelength and excellent photostability.

A fluorescent dye is an organic compound having a highly π-conjugated structure, and when it is excited by absorbing light of a specific wavelength, its energy is not spent on vibration or rotational movement in the molecule, and light of a specific wavelength is used. Is released as. Fluorescence emission has different absorption wavelengths and emission wavelengths in this way, and because of its sensitivity to lasers and its ability to convert between light energy and electrical energy, it can be used as a dye for optical recording media or a photoelectric conversion dye. It is expected to be applied in various fields such as displays and solar cells. In addition, due to its high visibility, it is also used as a fluorescent labeling dye in the biotechnology field.

Ultraviolet light having high energy may be used as the absorbed light of the fluorescent dye, but in recent years, infrared light has been attracting attention because of its safety and high transparency. For example, Patent Document 1 discloses a silaxanthenium core-based fluorescent compound that is sufficiently redshifted by using a substituent selected to correspond to the far-infrared and NIR spectra. .. Patent Document 2 discloses a near-infrared fluorescent compound containing a pyridinium structure in a benzo [a] phenoxatin 5-substituted imino group. Patent Documents 3 and 4 disclose cyanine compounds useful as contrast components of near-infrared fluorescence contrast agents. However, in recent years, fluorescent dyes tend not to be newly developed.

The cyanine pigment structure, which is a typical fluorescent dye, has a heterocycle containing nitrogen at both ends of the polymethine skeleton. It is known that the absorption wavelength and the fluorescence wavelength can be shifted to the longer wavelength side by lengthening the polymethine skeleton. That is, every time the number of methine groups (−CH =) is increased by two, the absorption wavelength of the cyanine dye shifts to the long wavelength side of about 100 nm.

Special Table 2016-521254A Japanese Unexamined Patent Publication No. 2014-166975 Japanese Unexamined Patent Publication No. 2011-46663 Japanese Unexamined Patent Publication No. 2011-46662

As described above, fluorescent dyes in which the absorption wavelength and the fluorescence wavelength are shifted to the longer wavelength side have been developed. For example, it is known that the absorption wavelength and the fluorescence wavelength can be shifted to the longer wavelength side by lengthening the polymethine skeleton of the cyanine pigment. However, on the other hand, it is also known that when the polymethine skeleton is lengthened, the photostability decreases and the fluorescence intensity with respect to the excitation light decreases with time.
Therefore, an object of the present invention is to provide a fluorescent dye having a relatively long maximum wavelength of excitation light and fluorescence and excellent photostability.

The present inventors have conducted intensive studies to solve the above problems. As a result, it was found that by introducing a pyrene ring into the structure of the fluorescent dye, not only the absorption wavelength and the fluorescence wavelength are shifted to the long wavelength side, but also the photostability is surprisingly improved. The invention was completed.
Hereinafter, the present invention will be shown.

［式中、
Ｒ¹は、カルボキシ基、スルホ基、アジド基、またはエチニル基で置換されていてもよいＣ_1-18アルキル基を示し、
Ｘ¹は、＞ＣＲ²Ｒ³（式中、Ｒ²とＲ³は独立してＣ_1-18アルキル基を示す。）、−Ｏ−、または−Ｓ−を示し、
Ｙは、下記式（ｉ）〜（ｉｖ）で表される基から選択されるいずれかの基を示す。

（式中、
Ｘ²とＸ³は、独立して、＞ＣＲ¹⁰Ｒ¹¹（式中、Ｒ¹⁰とＲ¹¹は独立してＣ_1-18アルキル基を示す。）、−Ｏ−、または−Ｓ−を示し、
Ｚは＝Ｏまたは＝Ｓを示し、
Ｒ⁴〜Ｒ⁹は、独立して、カルボキシ基、スルホ基、アジド基、またはエチニル基で置換されていてもよいＣ_1-18アルキル基を示し、
ｌは、１以上、５以下の整数を示し、
ｍは、０以上、５以下の整数を示し、
ｎは、１以上、５以下の整数を示す。）］ [1] A pyrene fluorescent dye represented by the following formula (I).

[During the ceremony,
R ¹ _{represents a C 1-18} alkyl group optionally substituted with a carboxy group, a sulfo group, an azide group, or an ethynyl group.
X ¹ indicates> CR ² R ³ (in the formula, R ² and R ³ independently indicate a C _1-18 alkyl group), −O−, or −S−.
Y represents any group selected from the groups represented by the following formulas (i) to (iv).

(During the ceremony,
X ² and X ³ independently indicate> CR ¹⁰ R ¹¹ (in the formula, R ¹⁰ and R ¹¹ independently represent a C _1-18 alkyl group), -O-, or -S-. ,
Z indicates = O or = S,
R ^{4 to} R ⁹ independently represent a _{C 1-18} alkyl group optionally substituted with a carboxy group, a sulfo group, an azide group, or an ethynyl group.
l indicates an integer of 1 or more and 5 or less,
m indicates an integer of 0 or more and 5 or less,
n represents an integer of 1 or more and 5 or less. )]

[2] The pyrene fluorescent dye according to the above [1], wherein Y is a group represented by the formula (i).
[3] The pyrene fluorescent dye according to the above [2], wherein ^{X 2} is> CR ¹⁰ R ^11.
[4] The pyrene fluorescent dye according to the above [2] or [3], wherein ^{R 4} is an unsubstituted C _{1-18 alkyl group.}
[5] The pyrene fluorescent dye according to any one of the above [1] to [4], wherein ^{X 1} is> CR ² R ^3.
[6] The pyrene fluorescent dye according to any one of the above [1] to [5], wherein ^{R 1} is an unsubstituted C _{1-18 alkyl group.}

In the present invention, the "C _1-18 alkyl group" refers to a linear or branched monovalent saturated aliphatic hydrocarbon group having 1 or more carbon atoms and 18 or less carbon atoms. For example, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, s-butyl group, t-butyl, n-pentyl, n-hexyl, n-octyl, n-decane, n-dodecane, n-tetradecane. , N-hexadecane, n-octadecane and the like. When steric hindrance is taken into consideration, a C _1-10 alkyl group is preferable, a C _1-6 alkyl group or a C _1-4 alkyl group is more preferable, and a C _1-2 alkyl group is even more preferable. When it has a substituent such as a carboxy group, a C _1-6 alkyl group is preferable, and a C _2-4 alkyl group is more preferable. When it is desired to increase the lipophilicity of the pyrene fluorescent dye, a C _6-18 alkyl group is preferable, a C _8-18 alkyl group is more preferable, and a C _10-16 alkyl group is even more preferable.

The pyrene fluorescent dye according to the present invention has a relatively long maximum absorption wavelength and a maximum fluorescence wavelength, and is excellent in light stability. For example, according to the examples described later, the maximum absorption wavelength and the maximum fluorescence wavelength are shifted to the long wavelength side of about 100 nm with respect to the conventional squalane dye having the same structure except that it does not have a pyrene ring. Moreover, the light stability is improved. Therefore, the pyrene fluorescent dye according to the present invention may replace the conventional fluorescent dye used in various fields.

^{FIG. 1 is a 1} H-NMR spectrum of an example of a pyrene fluorescent dye according to the present invention. FIG. 2 shows an example of a pyrene fluorescent dye according to the present invention, and an absorption wavelength spectrum and a fluorescence wavelength spectrum of a conventional fluorescent dye. FIG. 3 is a graph showing an example of a pyrene fluorescent dye according to the present invention and the photostability of a conventional fluorescent dye.

A person skilled in the art can produce the pyrene fluorescent dye according to the present invention according to a conventionally known method for producing a cyanine dye. For example, the pyrene fluorescent dye represented by the formula (I), in which Y is the group represented by the formula (i), can be produced by the following synthetic scheme. Hereinafter, Q represented by the formula (P) may be abbreviated as “Q (P)”.

^{In the above reaction, the pyrene-squaric acid dye (I 1} ) is produced by reacting the pyrene compound (II) with the squaric acid derivative (III) in the presence of a base in a solvent.

Solvents, pyrene compounds (II) and showed moderate solubility in squaric acid derivative (III), is not particularly as long as they do not and to inhibit a reaction limited, for example, methanol, ethanol, and 1-butanol C _{1 -4} Alcohols; aromatic hydrocarbon solvents such as benzene, toluene, chlorobenzene; and mixed solvents thereof.

Examples of the base include organic bases such as quinoline and pyridine, and inorganic bases such as sodium acetate.

The reaction conditions may be adjusted as appropriate. For example, the amount of the pyrene compound (II) with respect to the aquaric acid derivative (III) may be 1.9 times mol or more and 2.1 times mol or less. The amounts of the pyrene compound (II) and the squaric acid derivative (III) in the reaction solution may be appropriately adjusted according to the solubility and the like. The amount of the base used is preferably an excess amount. The reaction temperature can be 150 ° C. or lower from room temperature, and the reaction may be carried out under heating and reflux. The reaction time may be until the raw material compound is consumed by thin layer chromatography or the like, or may be determined by a preliminary experiment or the like, and may be, for example, 1 hour or more and 60 hours or less.

The pyrene fluorescent dye (I) having Y as a group (ii) to (iv) can be produced by the following synthetic scheme. In the following, the synthetic scheme ^{of the pyrene fluorescent dye (I 2} ) in which Y is the group (ii) is typically shown.

The pyrene-cyanine fluorescent dye (I ² ) can be produced under the same conditions as the ^{pyrene-squarine fluorescent dye (I 1} ) except that the polymethine compound (IV) is used instead of the squaric acid derivative (III). Further, in the pyrene-merocyanine fluorescent dye (I ³ ) and the pyrene-LDS fluorescent dye (I ⁴ ), one end of the polymethine compound (IV) is replaced with an oxopyrimidine ring or an aniline ring, respectively, instead of the polymethine compound (IV). It can be produced in the same manner as the ^{pyrene-cyanine fluorescent dye (I 2} ) except that these compounds and the pyrene compound (II) are used in a molar ratio of about 1: 1.

The pyrene compound (II) can be produced by applying a conventionally known indole synthesis method, benzoxazole synthesis method, or benzothoxazole synthesis method. However, when X ¹ is> CR ² R ³ , the following method using a metal catalyst (Matyas Turkey et al., Organic & Biomolecular Chemistry, 2010, 8, 5576-5582) is very efficient.

When X ¹ is −O—, hydroxypyrene is acylated, acylhydroxypyrene is obtained from the obtained acylated product by a Fries rearrangement reaction, and the acylhydroxypyrene is oxime-ized and then subjected to a transfer cyclization reaction to obtain an oxazole ring. To form. If X ¹ is -S-, use pyrene thiol instead of hydroxypyrene.

The pyrene fluorescent dye (I) and the intermediate compound may be purified by a known method, or the intermediate compound may be used in the next reaction only by crude purification or not by purification depending on the purity. In addition, the reactive functional group may be appropriately protected and deprotected.

Squaric acid pigments, also called squarylium pigments, are classified as cyanine pigments, but they have a squaric acid (square acid) moiety in the central part of the polymethine conjugated system, and a cationic group and an anionic group coexist in one molecule. It has a unique structure called a zwitterionic structure. Squalane dyes are used as charge generators for electrophotographic photosensitive members, sensitizing dyes for organic solar cells, and the like.

The indocyanine pigment has a structure having a heterocycle containing nitrogen at both ends of the polymethine skeleton. One nitrogen is ammonium with a cationic structure and has a role as an electron acceptor, and the other nitrogen is a tertiary amine and has a role as an electron donor. It usually exists as a salt with anions. Examples of the anion include halide ion, sulfonic acid ion, perchlorate ion, tetrafluoroborate, hexafluoroantimonate and the like. Indocyanine pigments are used as pigments for optical recording media and the like.

The merocyanine dye exhibits a large absorption coefficient and is used as a light absorber, sensitizer, dye, etc. for optical filters, photosensitive photographic materials, dyes, paints, inks, electrophotographic photosensitive members, toners, heat sensitive materials, etc. It is widely used in recording papers, transfer ribbons, optical recording dyes, solar cells, photoelectric conversion elements, semiconductor materials, clinical test reagents, dyes for laser treatment, dyeing, and the like.

The LDS dye laser is promising as a light source for the DIAL system because the LDS dye has a relatively long life and can be easily replaced with Ti: Sapphire.

Further, the pyrene-squarine dye (I ¹ ), the pyrene-cyanine fluorescent dye (I ² ), the pyrene-merocyanine fluorescent dye (I ³ ), and the pyrene-LDS fluorescent dye (I ⁴ ) according to the present invention have maximum absorption wavelengths. The maximum fluorescence wavelength is shifted to the long wavelength side, and excellent photostability is exhibited. In addition, the pyrene fluorescent dye of the present invention, which is highly water-soluble by substituting a carboxy group or a sulfo group with _{a C 1-18 alkyl group, can be used in a living body for bioimaging and photodynamic treatment.} obtain.

Hereinafter, the present invention will be described in more detail with reference to examples, but the present invention is not limited by the following examples as well as the present invention, and appropriate modifications are made to the extent that it can be adapted to the gist of the above and the following. Of course, it is possible to carry out, and all of them are included in the technical scope of the present invention.

Example 1: Synthesis of Pyrene-Squareine Dye

(1) Synthesis of Compound 1 1-aminopyrene (3 g, 13.8 mmol), RuCl ₃ · H ₂ O (28.6 mg, 0.138 mmol), and xantphos (239.6 mg, 0.414 mmol) were placed in an eggplant flask. After replacing the gas phase in the flask with argon, 2,3-butanediol (1.27 mL, 13.8 mmol) was added, and the mixture was heated at 110 ° C. for 1 hour, further heated to 180 ° C., and heated for 2 days. After returning the temperature of the reaction product to room temperature, it was dissolved in chloroform and the insoluble matter was filtered off. After concentrating the filtrate under reduced pressure, the target product as a yellow solid was isolated by silica gel column chromatography (eluent: chloroform) (yield: 2.62 g, yield: 70%). The formation of the target product was ^{confirmed by 1 1} H-NMR.

(2) Synthesis of Compound 2 Pd ₂ (dba) ₃ (170.3 mg, 0.186 mmol) and P (2-furyl) ₃ (85.9 mg, 0.37 mmol) were placed in a two-necked eggplant flask and inside the flask. The gas phase of the flask was substituted with argon. Further, allyl methyl carbonate (1.69 mL, 14.9 mmol) and dehydrated dichloromethane (18.6 mL) were added, and the mixture was stirred at room temperature for 10 minutes. This solution was added to a separately prepared solution of compound 1 (2 g, 7.43 mmol) in dehydrated dichloromethane (18.6 mL), and the mixture was stirred at room temperature for 3 hours. After removing the solvent with an evaporator, the target product, which is an orange to brown viscous liquid, was isolated by silica gel column chromatography (eluent: ethyl acetate / hexane = 1/3) (yield: 1.25 g). , Yield: 55%). The formation of the target product was ^{confirmed by 1 1} H-NMR.

(3) Synthesis of Compound 3 Compound 2 (1.2 g, 3.88 mmol) and Pd / C (120 mg, 10 w%) were placed in a two-necked eggplant flask, the gas phase in the flask was replaced with hydrogen, and then methanol (1.2 g, 3.88 mmol) was placed. 19.4 mL) was added. Further, hydrogen gas was bubbled into the same solution, and the mixture was stirred at room temperature for 4 hours. After removing the insoluble matter with a fold-folded filter paper, the filtrate was concentrated under reduced pressure, and the target product as an orange solid was isolated by silica gel column chromatography (eluent: hexane / ethyl acetate = 5/1) (yield: 954 mg, yield: 79%). The formation of the target product was ^{confirmed by 1 1} H-NMR.

(4) Synthesis of Compound 4 Compound 3 (694 mg, 2.23 mmol), iodomethane (2 mL), and acetonitrile (11.2 mL) were placed in an eggplant flask and heated at 80 ° C. for 24 hours. The precipitate was collected by filtration to obtain the desired product as a solid yellow powder (yield: 512 mg, yield: 51%). The formation of the target product was ^{confirmed by 1 1} H-NMR.

(5) Synthesis of Pyrene-Squareine Dye (PYSQ) Compound 4 (200 mg, 0.44 mmol), 3,4-diethoxy-3-cyclobutene-1,2-dione (32 μL, 0.22 mmol), and 1 in an eggplant flask. A mixture of −butanol / pyridine = 1/1 (1.1 mL / 1.1 mL) was added, and the mixture was heated under reflux at 120 ° C. for 21 hours. The reaction mixture was concentrated under reduced pressure and isolated by silica gel column chromatography (eluent: dichloromethane / methanol = 99/1). Further washing with hot methanol gave the desired product as a green powder solid (yield: 40.6 mg, yield: 13%). ^{The 1} H-NMR spectrum of the obtained PYSQ is shown in FIG.

Test Example 1: Fluorescence characteristics The fluorescence characteristics of the pyrene-squareine dye (PYSQ) synthesized in Example 1 were evaluated. For comparison, the fluorescence characteristics of the conventionally known squalane dye (SQ) were also evaluated in the same manner. The results are shown in Table 1 and FIG.

As shown in the results shown in Table 1 and FIG. 2, PYSQ showed a light absorption / fluorescence wavelength that was nearly 100 nm longer than that of SQ. The brightness (product of molar extinction coefficient and fluorescence quantum yield) was about the same as that of SQ, which is called a high-luminance dye.

Test Example 2: Photostability test The pyrene-squareine dye (PYSQ) synthesized in Example 1 was dissolved in toluene, irradiated with excitation light of 657 nm, and the time change of fluorescence intensity was monitored. Further, for comparison, the concentration of the solution of the conventionally known squalane dye (SQ) was adjusted so that the absorbance at 657 nm was the same, and then the change in fluorescence intensity over time was similarly monitored. The results are shown in FIG.
As shown in the results shown in FIG. 3, after 3 hours of light irradiation, the fluorescence intensity of SQ was attenuated to about 83% with respect to the initial state, while it was maintained at 90% or more in PYSQ. Therefore, it was found that the photostability of PYSQ is superior to that of SQ. SQ is a dye having far superior light stability as compared with commercially available dyes such as Nile Red.

Claims (6)

A pyrene fluorescent dye represented by the following formula (I).

[During the ceremony,
R ¹ _{represents a C 1-18} alkyl group optionally substituted with a carboxy group, a sulfo group, an azide group, or an ethynyl group.
X ¹ indicates> CR ² R ³ (in the formula, R ² and R ³ independently indicate a C _1-18 alkyl group), −O−, or −S−.
Y represents any group selected from the groups represented by the following formulas (i) to (iv).

(During the ceremony,
X ² and X ³ independently indicate> CR ¹⁰ R ¹¹ (in the formula, R ¹⁰ and R ¹¹ independently represent a C _1-18 alkyl group), -O-, or -S-. ,
Z indicates = O or = S,
R ^{4 to} R ⁹ independently represent a _{C 1-18} alkyl group optionally substituted with a carboxy group, a sulfo group, an azide group, or an ethynyl group.
l indicates an integer of 1 or more and 5 or less,
m indicates an integer of 0 or more and 5 or less,
n represents an integer of 1 or more and 5 or less. )] The pyrene fluorescent dye according to claim 1, wherein Y is a group represented by the formula (i). The pyrene fluorescent dye according to claim 2, wherein X ² is> CR ¹⁰ R ^11. The pyrene fluorescent dye according to claim 2 or 3, wherein R ⁴ is an unsubstituted C _{1-18 alkyl group.} The pyrene fluorescent dye according to any one of claims 1 to 4, wherein X ¹ is> CR ² R ^3. The pyrene fluorescent dye according to any one of claims 1 to 5, wherein R ¹ is an unsubstituted C _{1-18 alkyl group.}

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