CN117003782A - TB-thiophene methylene malononitrile derivative and preparation and application thereof - Google Patents

TB-thiophene methylene malononitrile derivative and preparation and application thereof Download PDF

Info

Publication number
CN117003782A
CN117003782A CN202310984532.2A CN202310984532A CN117003782A CN 117003782 A CN117003782 A CN 117003782A CN 202310984532 A CN202310984532 A CN 202310984532A CN 117003782 A CN117003782 A CN 117003782A
Authority
CN
China
Prior art keywords
derivative
cells
thiophene
malononitrile
mol
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
CN202310984532.2A
Other languages
Chinese (zh)
Inventor
张鹏
梁燕妮
庄敏艳
张宇
苑睿
吴翚
宛瑜
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Jiangsu Normal University
Original Assignee
Jiangsu Normal University
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Jiangsu Normal University filed Critical Jiangsu Normal University
Priority to CN202310984532.2A priority Critical patent/CN117003782A/en
Publication of CN117003782A publication Critical patent/CN117003782A/en
Pending legal-status Critical Current

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
    • C07F5/00—Compounds containing elements of Groups 3 or 13 of the Periodic Table
    • C07F5/02—Boron compounds
    • C07F5/025—Boronic and borinic acid compounds
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K41/00—Medicinal preparations obtained by treating materials with wave energy or particle radiation ; Therapies using these preparations
    • A61K41/0057—Photodynamic therapy with a photosensitizer, i.e. agent able to produce reactive oxygen species upon exposure to light or radiation, e.g. UV or visible light; photocleavage of nucleic acids with an agent
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K49/00—Preparations for testing in vivo
    • A61K49/001—Preparation for luminescence or biological staining
    • A61K49/0013—Luminescence
    • A61K49/0017—Fluorescence in vivo
    • A61K49/0019—Fluorescence in vivo characterised by the fluorescent group, e.g. oligomeric, polymeric or dendritic molecules
    • A61K49/0021—Fluorescence in vivo characterised by the fluorescent group, e.g. oligomeric, polymeric or dendritic molecules the fluorescent group being a small organic molecule
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
    • C09K—MATERIALS FOR MISCELLANEOUS APPLICATIONS, NOT PROVIDED FOR ELSEWHERE
    • C09K11/00—Luminescent, e.g. electroluminescent, chemiluminescent materials
    • C09K11/06—Luminescent, e.g. electroluminescent, chemiluminescent materials containing organic luminescent materials
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N11/00—Investigating flow properties of materials, e.g. viscosity, plasticity; Analysing materials by determining flow properties
    • C—CHEMISTRY; METALLURGY
    • C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
    • C09K—MATERIALS FOR MISCELLANEOUS APPLICATIONS, NOT PROVIDED FOR ELSEWHERE
    • C09K2211/00—Chemical nature of organic luminescent or tenebrescent compounds
    • C09K2211/10—Non-macromolecular compounds
    • C09K2211/1018—Heterocyclic compounds
    • C09K2211/1025—Heterocyclic compounds characterised by ligands
    • C09K2211/1044—Heterocyclic compounds characterised by ligands containing two nitrogen atoms as heteroatoms
    • C—CHEMISTRY; METALLURGY
    • C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
    • C09K—MATERIALS FOR MISCELLANEOUS APPLICATIONS, NOT PROVIDED FOR ELSEWHERE
    • C09K2211/00—Chemical nature of organic luminescent or tenebrescent compounds
    • C09K2211/10—Non-macromolecular compounds
    • C09K2211/1018—Heterocyclic compounds
    • C09K2211/1025—Heterocyclic compounds characterised by ligands
    • C09K2211/1092—Heterocyclic compounds characterised by ligands containing sulfur as the only heteroatom
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N11/00—Investigating flow properties of materials, e.g. viscosity, plasticity; Analysing materials by determining flow properties
    • G01N2011/006—Determining flow properties indirectly by measuring other parameters of the system
    • G01N2011/008—Determining flow properties indirectly by measuring other parameters of the system optical properties

Landscapes

  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • General Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Veterinary Medicine (AREA)
  • Public Health (AREA)
  • Animal Behavior & Ethology (AREA)
  • Medicinal Chemistry (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Engineering & Computer Science (AREA)
  • Biochemistry (AREA)
  • Epidemiology (AREA)
  • Pathology (AREA)
  • Biomedical Technology (AREA)
  • Immunology (AREA)
  • General Physics & Mathematics (AREA)
  • Analytical Chemistry (AREA)
  • Molecular Biology (AREA)
  • Physics & Mathematics (AREA)
  • Materials Engineering (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Abstract

The invention provides a TB-thiophene methylene malononitrile derivative and preparation and application thereof, which are synthesized by taking p-bromoaniline, paraformaldehyde, 5-bromothiophene-2-formaldehyde, malononitrile and the like as raw materials through multi-step reaction. The product has larger Stokes shift (147 nm and 213nm respectively) in solution and solid state, has excellent solid state luminescence, and is used in THF/H 2 O=1/99 (v/v) solution forms uniform petal-shaped nano-aggregates with remarkable AIE properties; tool withThe pH value application range is wide, and the pH value can be applied to human physiological environments; fluorescence is enhanced by 6.1 times at methanol/glycerol=1/9 (v/v) compared with pure methanol, and the fluorescent dye has better response capability to viscosity; IC for HpeG2 cells and A549 cells under dark conditions 50 71.7 and 100. Mu. Mol.L, respectively ‑1 While under the illumination condition (428 nm), the inhibition rates of HpeG2 cells and A549 cells are respectively 9.2 and 19.9 mu mol.L ‑1 Shows very excellent photodynamic anticancer activity.

Description

TB-thiophene methylene malononitrile derivative and preparation and application thereof
Technical Field
The invention belongs to the field of chemical synthesis, and in particular relates tobase-thiophene methylene malononitrile derivatives, synthesis method and photodynamic anticancer activity thereof.
Background
Photodynamic therapy (Photodynamic Therapy, PDT) is a novel method for treating diseases such as tumors with photosensitizing drugs and laser activation. Compared with the traditional method, the method has the advantages of low side effect, minimally invasive, high safety, no drug resistance, high tumor destruction selectivity, easy combination with other therapies and the like, has better space-time precision and effectiveness in clinical research and treatment, and becomes an important emerging means for clinical tumor precision treatment.
The efficacy of photosensitizers is a central factor in determining the therapeutic effect of PDT, and along with the rapid development of PDT, the requirements of various photosensitizers, the efficiency of photosensitizers and the like are increasingly high. Aiming at the problems of poor visible absorption and penetrability and the like of the photosensitizer of the traditional PDT, more and more novel high-efficiency photosensitizers are developed. But is based onPhotosensitizers of The Base (TB) backbone are still in the primary stage and only one relevant report exists.
Thiophene is an electron-rich aromatic heterocycle and has good modifiable property. The thiophene derivatives have good photoelectric properties due to high electron mobility, low electrode potential and good conductivity, and are widely used in the design of biological diagnostic reagents, medicines, electronic and photoelectric equipment and conductive and electroluminescent materials.
The malononitrile molecule has two cyano groups with strong electron withdrawing, so that methylene of the malononitrile molecule is easy to form carbanion, thus the malononitrile molecule has high reaction activity and various reaction types. The arylmethylene malononitrile block prepared by Knoevenagal reaction of malononitrile and aldehyde has a longer conjugated system and strong electron-withdrawing capability, and is beneficial to improving the light absorption and emission capability of molecules after molecules are introduced.
Therefore, the invention designs and synthesizes the structure of D-pi-A by introducing electron-withdrawing thiophene methylene malononitrile fragment on TB skeletonbase-thiophene methylene malononitrile compounds.
Disclosure of Invention
Technical problems: the invention aims to provide a TB-thiophene methylene malononitrile derivative, and a preparation method and application thereof, wherein p-bromoaniline, paraformaldehyde, 5-bromothiophene-2-formaldehyde and malononitrile are used as raw materials, and the TB-thiophene methylene malononitrile derivative is synthesized through multi-step reactionbase-thiophene methylene malononitrile compounds and their use in viscosity identification and PDT treatment applications.
The technical scheme is as follows: the invention discloses a TB-thiophene methylene malononitrile derivative, which has the structural formula:
the invention relates to a preparation method of a TB-thiophene methylene malononitrile derivative, which comprises the following steps:
step 1, 4-bromoaniline reacts with paraformaldehyde to obtain a first intermediate, wherein the reaction formula is as follows:
step 2, reacting the first intermediate with n-butyllithium to obtain a second intermediate, wherein the reaction formula is as follows:
step 3, the second intermediate and 5-bromothiophene-2-formaldehyde are subjected to a coupling reaction to obtain a third intermediate, wherein the reaction formula is as follows:
step 4, the third intermediate and malononitrile are subjected to a coupling reaction to obtain a derivative, wherein the reaction formula is as follows:
the invention discloses an application of a TB-thiophene methylene malononitrile derivative in preparing a viscosity probe.
The invention relates to an application of a TB-thiophene methylene malononitrile derivative in preparing a medicine for photodynamic therapy of cancer.
The application of the photodynamic therapy medicine for cancer is aimed at inhibiting the HpeG2 cells of human liver cancer and A549 cells of human lung cancer.
The beneficial effects are that:
1. the synthesis method is simple and the post-treatment is convenient.
2. Has large Stokes displacement, excellent solid state luminescence and remarkable AIE properties; the pH value application range is wide, and the pH value can be applied to human physiological environments;
3. the product has good response to the viscosity, and has the potential of becoming a fluorescent probe for viscosity response;
5. for HpeG2 cells and A549 cells under dark conditionsCell IC 50 71.7 and 100. Mu. Mol.L, respectively -1 While under the illumination condition (428 nm), the inhibition rates of HpeG2 cells and A549 cells are respectively 9.2 and 19.9 mu mol.L -1 Shows excellent photodynamic anticancer activity and has potential of being developed into novel photodynamic anti-tumor photosensitive medicaments.
Drawings
FIG. 1 is a schematic illustration of the product derivatives of the examples 1 HNMR spectrogram;
FIG. 2 is a schematic illustration of the product derivative 7 of the example 13 C NMR spectrum;
FIG. 3a is an ultraviolet absorption spectrum of a third intermediate 6 in a different solvent;
FIG. 3b is a fluorescence emission spectrum of the third intermediate 6 in a different solvent;
FIG. 3c is the ultraviolet absorbance spectrum of derivative 7 in different solvents;
FIG. 3d is the fluorescence emission spectra of derivative 7 in different solvents;
FIG. 4 is a plot of (a) fluorescence emission spectra and (b) line graph of third intermediate 6 at different pH and (c) fluorescence emission spectra and (d) line graph of compound 7 at different pH;
FIG. 5 is a third intermediate 6 in different ratios of THF/H 2 (a) fluorescence emission spectra and (b) line graphs and derivatives 7 in O (v/v) in different ratios of THF/H 2 O (v/v) a (c) fluorescence emission spectrum and (d) a line graph;
FIG. 6 shows the THF/H ratio of derivative 7 2 SEM image at O (v/v) (a) THF/H 2 O=1/9 (v/v) and (b) THF/H 2 O＝1/99(v/v)；
FIG. 7 is a plot of (a) fluorescence emission spectra and (b) line for the third intermediate 6 at different viscosities and (c) fluorescence emission spectra and (d) line for the compound 7 at different viscosities;
FIG. 8 shows the fluorescence intensity of derivative 7 with different ions and molecules;
FIG. 9 is a plot of (a) fluorescence emission spectra versus (b) for derivative 7 at different temperatures; a line graph of (c) fluorescence emission spectrum and (d) after combination with egg white;
FIG. 10 is the phototoxicity and dark toxicity of derivative 7 on (a) A549 cells and (b) HpEG2 cells.
Detailed Description
The invention is further illustrated below with reference to examples.
Embodiments of the present invention are described in detail below. The examples described below are illustrative only and are not to be construed as limiting the invention. Those skilled in the art will appreciate that various changes and modifications can be made to the invention without departing from the spirit and scope thereof.
The following steps are as follows: 4-bromoaniline 1, paraformaldehyde 2, a first intermediate 3, a second intermediate 4, 5-bromothiophene-2-carbaldehyde 5, a third intermediate 6, a derivative 7.
base-thiophene methylene malononitrile compounds having the structural formula shown in the following table:
TABLE 1 structural formula of derivative 7
The invention also provides the novelA process for producing a base-thiophene methylene malononitrile compound, comprising:
in the embodiment, the catalyst is prepared from p-bromoaniline, paraformaldehyde, n-butyllithium, 5-bromo-thiophene-2-formaldehyde, malononitrile and the like serving as raw materials through a coupling reaction. The method comprises the following steps:
the 4-bromoaniline 1 reacts with paraformaldehyde 2 to obtain a first intermediate 3, the first intermediate 3 reacts with n-butyllithium to obtain a second intermediate 4, the second intermediate 4 reacts with 5-bromo-thiophene-2-formaldehyde to obtain a third intermediate 6, and the third intermediate 6 reacts with malononitrile to obtain a derivative 7 through coupling reaction.
The first intermediate 3 in the examples was prepared by the above synthetic method:
4-Bromoaniline (50.0 mmol) and paraformaldehyde (100.0 mmol) were successively added to a 200.0mL round-bottomed flask, the flask was warmed to-15℃in a low-temperature tank, and trifluoroacetic acid (100.0 mL, after completion of dropwise addition for about 30 min) was slowly added dropwise to the flask under stirring, followed by reaction at room temperature for 7 days. After completion of the reaction (TLC trace), the mixture was poured into ice water, ph=9-10 was adjusted with aqueous ammonia, cooled to room temperature, extracted with dichloromethane (50.0 ml×3) and dried by spinning to give the crude product. Acetone is added, the mixture is heated until the crude product is completely dissolved, recrystallized at room temperature, filtered by suction, and washed by acetone, thus obtaining a first intermediate 3.
(3) First intermediate 3 (5.0 mmol) was added to a 100mL round bottom flask, after three times of air extraction, the flask was placed in a low temperature tank, the temperature was adjusted to-78 ℃, 20.0mL of anhydrous tetrahydrofuran was added to the flask under stirring, 2.5mL of n-butyllithium was added dropwise, after 1 hour of reaction under argon protection, 0.6mL of trimethyl borate was added dropwise, and then the flask was left to react at room temperature for 4 hours. After completion of the reaction by TLC, extraction with dichloromethane (30.0 mL. Times.3) was performed and the crude product was obtained by spin-drying. Purification of the crude product by column chromatography (V PE :V EA =5:1) to afford second intermediate 4 (65%).
(4) A second intermediate 4 (1.0 mmol), 5-bromothiophene-2-carbaldehyde (1.2 mmol), tetrakis (triphenylphosphine) palladium (20% mmol,0.03 g) and K was taken 2 CO 3 (0.2 mmol) was added sequentially to a 100mL round bottom flask, 20mL anhydrous toluene was added under argon protection, and the reaction was carried out at 108℃for 24h. After completion of the reaction (TLC trace), quench with water, extract with dichloromethane (10.0 mL. Times.3), the organic phase was taken over Na 2 SO 4 The crude product obtained by spin drying after drying was purified by column chromatography (V Petroleum ether :V Acetic acid ethyl ester =6:1) to afford third intermediate 6 (65%).
(5) A third intermediate 6 (1.0 mmol), malononitrile (1.2 mmol) and absolute ethanol 10.0mL were added sequentially to a 100mL round bottom flask and reacted at 70℃for 6h under the protection of argon. After completion of the reaction (TLC trace), quench with water, extract with dichloromethane (10.0 mL. Times.3), the organic phase was taken over Na 2 SO 4 The dried crude product was purified by column chromatography (V Petroleum ether :V Acetic acid ethyl ester =3:1) to give derivative 7 (77%).
Derivative 7 has the formula: c (C) 23 H 17 BN 4 O 2 S
The Chinese name is: (8- (5- (2, 2-dicyanovinyl) thiophen-2-yl) -6H,12H-5, 11-xylylene [ b, f ] [1,5] diazo-2-yl) boronic acid
English is named:
(8-(5-(2,2-dicyanovinyl)thiophen-2-yl)-6H,12H-5,11-methanodibenzo[b,f][1,5]diazocin-2-yl)boronic acid
appearance: red solid
Melting point: 184.2-184.8 DEG C
Nuclear magnetic resonance hydrogen spectrum: 1 H NMR(400MHz,CDCl 3 )δ7.73(s,1H),7.64(d,J＝4.0Hz,1H),7.49(d,J＝8.1Hz,1H,Ar-H),7.39-7.38(m,1H,-OH),7.30(d,J＝4.0Hz,1H),7.24(s,1H,-OH),7.21-7.15(m,3H,Ar-H),7.00(d,J＝7.2Hz,1H,Ar-H),6.93(d,J＝7.5Hz,1H,Ar-H),4.76-4.72(m,2H,-CH 2 -bridge),4.38-4.20(m,4H,TB-CH 2 *2).
nuclear magnetic resonance carbon spectrum: 13 C NMR(100MHz,DMSO-d 6 )δ155.06,144.55,131.35,120.50,109.10,107.78,66.06,56.83,56.63,40.49,40.28,40.07,39.96,39.65,39.44,39.23.
mass spectrometry: HRMS (ESI) m/z calcd for C 23 H 17 BN 4 O 2 S[M+H] + :425.1242；found,425.1268.
Optical Properties
The solvation effect of the compounds of the invention was tested, and the specific test protocol is as follows:
the compound 6 and the derivative 7 were purified with Methanol (Methanol), dimethyl sulfoxide (DMSO), N-Dimethylformamide (DMF), tetrahydrofuran (THF), ethyl acetate (ethyl acetate), chloroform (CHCl) 3 ) Toluene (tolene), n-Hexane (n-Hexane) to give a concentration of 1×10 -5 mol·L -1 And testing the ultraviolet absorption spectrum and the fluorescence emission spectrum of the working fluid. As shown in fig. 3a-3 d.
As can be seen from fig. 3a-3d, the uv absorption spectra of the third intermediate 6 and derivative 7 are not significantly different in each solvent; and the third intermediate 6 and the derivative 7 are relatively strong in fluorescence intensity due to higher solubility in a medium-small polar solvent.
The ultraviolet absorption and fluorescence emission spectra of the third intermediate 6 and the derivative 7 in THF solution and their solid state fluorescence emission spectra were tested, and the specific test protocol is as follows:
weighing 10 -5 mol of the second intermediate 4, the third intermediate 6 and the derivative 7 are taken up in THF to a concentration of 1X 10 -5 And (3) testing ultraviolet absorption, fluorescence emission and solid state fluorescence emission spectra of the fluorescent dye. The spectral data of the second intermediate 4, the third intermediate 6 and the derivative 7 are shown in table 2.
Table 2 Spectrum data (THF) for second intermediate 4, third intermediate 6 and derivative 7
a Ultraviolet absorption wavelength (slit is 2.5/5 nm) in the solution; b molar extinction coefficient epsilon=a/bC in 1×10 5 L·mol -1 ·cm -1 ； c Fluorescence emission wavelength in the solution; d stokes shift in solution; e relative fluorescence quantum yield (reference: quinine sulfate); f fluorescent brightness in L.mol -1 ·cm -1 ； g Solid state excitation wavelength (slit 5/5 nm); h solid state fluorescenceA wavelength of the radiation; i solid Stokes displacement.
As can be seen from table 2, the fluorescence properties of derivative 7 were changed as follows compared to the starting second intermediate 4, third intermediate 6:
(1) Solution and solid lambda of derivative 7 em Obvious red shift occurs, and the displacement of the solution and the solid Stokes (147 nm and 213nm respectively) is obviously increased;
(2) The solution of derivative 7 has significantly increased relative fluorescence quantum yield.
This is probably due to the fact that electron withdrawing group thiophene methylene malononitrile promotes the electron flow in the molecule, reduces the whole energy of the molecule, makes the fluorescence emission easier, the maximum fluorescence emission wavelength longer and Stokes displacement increased;
(3) The combined action of c=c and two-c≡n limits intramolecular rotation, so that the whole molecule has a highly distorted structure, thereby enhancing solid-state light emission intensity.
The above results demonstrate that combining the TB backbone with thiophene and methylene malononitrile fragments can amplify the advantages of both in light-emitting properties, a new way to obtain products with excellent light-emitting properties.
pH response
The third intermediate 6 is prepared to have a concentration of 1X 10 by using THF as a solvent -4 mol·L -1 Respectively weighing 1.0mL of a third intermediate 6 working solution into 9 volumetric flasks of 10.0mL, then respectively adding 1.0mL of a buffer solution with pH value of 2.2-10.0 (citric acid/disodium hydrogen phosphate system is selected when the pH value is 2.2-8.0, sodium bicarbonate/sodium carbonate system is selected when the pH value is 9.0-10.0), and THF (total volume) to ensure that the concentration is 1X 10) -5 mol L -1 Its fluorescence emission spectrum (. Lambda.) was measured ex =380 nm, slit: 5/5nm, FIGS. 4a-4 b). The method of preparing the reagent of derivative 7 is consistent with that of third intermediate 6 (lambda ex =430 nm, slit: 5/5nm, FIGS. 4c-4 d).
As can be seen from fig. 4a-4d, the fluorescence intensity of the third intermediate 6 and the derivative 7 remained almost constant in the pH range of 2.2-10.0, indicating that the third intermediate 6 and the derivative 7 have a wide pH application range.
AIE Property
Since the third intermediate 6 is easily soluble in THF and poorly soluble in water, the third intermediate 6 is formulated to have a concentration of 1×10 - 4 mol·L -1 1.0mL of the working solution is respectively measured in 10 volumetric flasks of 10.0mL, then 0.0-9.0mL of double distilled water is respectively added into the 10 volumetric flasks of 10.0mL, and THF is added to constant volume so that the concentration is 1 multiplied by 10 -5 mol·L -1 (THF/H 2 O (v/v) is 1/9-9/1 in turn), 6 is prepared to have the concentration of 1 multiplied by 10 -3 mol·L -1 100.0 mu L of the working solution is measured in a 10.0mL volumetric flask, 9.9mL of double distilled water is added into the 10.0mL volumetric flask, and THF is added to the volumetric flask to fix the volume so that the concentration is 1 multiplied by 10 - 5 mol L -1 So that THF/H 2 O=1/99 (v/v). The fluorescence emission spectra were measured as shown in FIGS. 5a-5b (lambda ex =365 nm, slit: 5/10 nm).
The method of preparing the reagent of derivative 7 is consistent with that of third intermediate 6 (lambda ex =430 nm, slit: 5/5nm, FIGS. 5c-5 d).
As shown in fig. 5a-5d, the fluorescence of the third intermediate 6 decreases with the increase of the polarity of the solution, and ACQ phenomenon appears; and when the water content of the derivative 7 is 10% -90%, fluorescence is continuously reduced due to the TICT effect, and as the water content is continuously increased, the derivative 7 is aggregated at the water content of 99% due to limited intramolecular vibration and limited intramolecular rotation in an aggregation state, so that the fluorescent light intensity reaches a peak value, and the fluorescent light has good AIE performance and is suitable for stable imaging of human body environment.
Compound 7 was observed in THF/H using Scanning Electron Microscopy (SEM) 2 O=1/9 (v/v) and THF/H 2 Topographical features at o=1/99 (v/v), as shown in fig. 6a-6 b.
As can be seen from fig. 6a-6 b: compound 7 in THF/H 2 The molecules exhibit an amorphous state when O=1/9 (v/v) and are in THF/H 2 O=1/99 (v/v) the molecules present uniform petal-shaped nano-aggregates due to rapid aggregation, the average diameter of the molecules being 10 μm.
Viscosity response
The viscosity responsiveness of the third intermediate 6 and derivative 7 was tested: methanol as solvent6 is prepared to have the concentration of 1 multiplied by 10 -4 mol·L -1 Is a working fluid of (a). Taking 10 volumetric flasks of 10.0mL, adding 0.0-9.0mL glycerol respectively, taking 1.0mL working solution into the volumetric flasks, and metering methanol to volume to make its concentration 1×10 -5 mol·L -1 Fluorescence emission spectra (volume ratio of methanol/glycerol: 1/9 to 10/0 in order) were measured as shown in 7a to 7b (lambda ex =365 nm, slit: 5/10 nm).
The method of preparing the reagent of compound 7 corresponds to the third intermediate 6 (λ ex =430 nm, slit: 5/10nm, FIGS. 7c-7 d).
As can be seen from fig. 7, the fluorescence intensities of the third intermediate 6 and the derivative 7 increase with an increase in viscosity. This may be due to the increased viscosity resulting in a hindered intramolecular movement of the third intermediate 6. However, the segment of the aromatic methyl malononitrile in the derivative 7 further limits the intramolecular movement, so that the capability of the segment of the aromatic methyl malononitrile to respond to the viscosity is stronger, which indicates that the product derivative 7 has the potential of becoming a fluorescent probe with the viscosity response.
Interference experiment
Checking common cation Fe 3+ 、Al 3+ 、Na + 、Ca 2+ 、Cu 2+ 、Cr 3+ And K + The method comprises the steps of carrying out a first treatment on the surface of the Anionic CO 3 2- 、HCO 3 - 、CH 3 COO - 、PO 4 2- 、SO 4 2- 、SCN - And HS (high speed) - The method comprises the steps of carrying out a first treatment on the surface of the Effects of biological thiols Cys, hcy and GSH and 90% glycerol (left to right 2-18,1 as blank) on the fluorescence emission spectrum of derivative 7 as shown in FIG. 8 (lambda) ex =430 nm, slit: 10/10 nm).
The result shows that after various anions and cations and biological thiols are added, the fluorescence intensity of the derivative 7 is basically unchanged, but the fluorescence intensity of the derivative 7 in 90% glycerol is obviously enhanced, which proves that the derivative 7 can realize specific detection of viscosity in a complex biological environment.
Protein aggregation assay
The formation of dense polypeptide chains during misfolding and aggregation of proteins can lead to viscosity changes. The physicochemical properties of the protein are changed after the protein is denatured, and hydrophobic groups in the molecule are exposed, so that the condensation speed of the protein is increased, the protein is separated out from the aqueous solution, and the viscosity of the protein is increased. Therefore, the viscosity change in the protein aggregation process is simulated by adopting the egg white thermal denaturation process, and the viscosity change in the denaturation process is monitored by using the derivative 7 to observe the change of the fluorescence property.
The fluorescence change of the derivative 7 at 0-100deg.C was first tested (FIGS. 9a-9 b), and the result shows that the fluorescence intensity of the derivative 7 at 0-100deg.C has no significant change, indicating that it has good thermal stability (lambda) ex =430 nm, slit: 5/10 nm).
The derivative 7 is mixed with a proper amount of egg white, the protein gradually gathers and the viscosity increases along with the temperature rise, and the fluorescence of the derivative 7 gradually increases, so that the derivative 7 can be used for monitoring the viscosity change when the protein gathers and has the potential of becoming a viscosity response probe for the protein gathers (figures 9c-9 d).
In vitro photodynamic therapy
Taking human non-small cell lung cancer (A549) cells and human liver cancer (HepG 2) cells as models, and detecting cytotoxicity of the derivative 7 on the A549 cells and the HepG2 cells by adopting an MTT method. A549 cells and HepG2 cells were seeded in 96-well plates (1×10) -5 mu.L of medium was added to each well, CO at 37 ℃ 2 After 24h incubation in the incubator, different concentrations of derivative 7 were added to the inoculated cells for 24h incubation. The microwell plates were then rinsed 3 times with PBS buffer and 10. Mu.L of MTT solution was added to each well for an additional 4h of incubation. Removing culture medium in the wells, adding 150 μl of DMSO into each well to dissolve blue-violet formazan (formazan) crystals in the cells, placing on a shaking table, and shaking at low speed for 5-7min to dissolve the crystalline substance sufficiently. And finally, measuring absorbance values of each hole at 560nm and 670nm by adopting an enzyme-linked immunosorbent assay. Cytotoxicity was calculated by the following formula:
Viability％＝[∑(A i /A 0 ×100)/n]
in which A i Absorbance values for different concentrations of the compound, respectively; a is that 0 Average absorbance values for control wells without added compound; n (=3)) Three parallel experiments are shown.
The MTT method was used to detect the dark toxicity and phototoxicity of derivative 7 on HpeG2 cells and A549 cells. The light source was 430nm, the control group was not subjected to light treatment, and absorbance values at 560nm and 670nm of each well were measured using an enzyme-linked immunosorbent assay. Dark toxicity and phototoxicity of derivative 7 on A549 and HpeG2 cells are shown in FIGS. 10a-10 b.
TABLE 5 half-maximal inhibitory amount (IC) of derivative 7 against two cells 50 )
Half inhibition rates of the derivative 7 on HepG2 and A549 cells are shown in Table 5, and the result shows that the derivative 7 has low dark toxicity and high phototoxicity on both HpeG2 and A549 cells, and has excellent PDT effect. Of particular interest, derivative 7 has very low dark toxicity to a549 cells (IC 50 >100.0μmol·L -1 ) While the phototoxicity is higher (IC 50 ＝19.9μmol·L -1 ) Indicating that it has the potential of developing into anti-liver cancer drugs.
The foregoing is only a preferred embodiment of the invention, it being noted that: it will be apparent to those skilled in the art that various modifications and adaptations can be made without departing from the principles of the present invention, and such modifications and adaptations are intended to be comprehended within the scope of the invention.

Claims (5)

1. A TB-thiophene methylene malononitrile derivative characterized by the structural formula:
2. a process for the preparation of a TB-thiophene methylenemalononitrile derivative according to claim 1, comprising the steps of:
step 1, 4-bromoaniline (1) reacts with paraformaldehyde (2) to obtain a first intermediate (3), and the reaction formula is as follows:
step 2, the first intermediate (3) reacts with n-butyllithium to obtain a second intermediate (4), and the reaction formula is as follows:
step 3, the second intermediate (4) and 5-bromothiophene-2-formaldehyde (5) are subjected to a coupling reaction to obtain a third intermediate (6), wherein the reaction formula is as follows:
step 4, the third intermediate (6) and malononitrile are subjected to a coupling reaction to obtain a derivative (7), wherein the reaction formula is as follows:
3. use of a TB-thiophene methylenemalononitrile derivative according to claim 1 for the preparation of a viscosity probe.
4. Use of a TB-thiophene methylene malononitrile derivative according to claim 1 for the preparation of a medicament for photodynamic treatment of cancer.
5. The use according to claim 4, wherein the use of the medicament for photodynamic therapy of cancer is directed to the inhibition of human liver cancer HpeG2 cells and human lung cancer a549 cells.
CN202310984532.2A 2023-08-07 2023-08-07 TB-thiophene methylene malononitrile derivative and preparation and application thereof Pending CN117003782A (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN202310984532.2A CN117003782A (en) 2023-08-07 2023-08-07 TB-thiophene methylene malononitrile derivative and preparation and application thereof

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN202310984532.2A CN117003782A (en) 2023-08-07 2023-08-07 TB-thiophene methylene malononitrile derivative and preparation and application thereof

Publications (1)

Publication Number Publication Date
CN117003782A true CN117003782A (en) 2023-11-07

Family

ID=88570706

Family Applications (1)

Application Number Title Priority Date Filing Date
CN202310984532.2A Pending CN117003782A (en) 2023-08-07 2023-08-07 TB-thiophene methylene malononitrile derivative and preparation and application thereof

Country Status (1)

Country Link
CN (1) CN117003782A (en)

Similar Documents

Publication Publication Date Title
Chen et al. A highly selective naked-eye and fluorescent probe for fluoride ion based on 1, 8-naphalimide and benzothizazole
Wang et al. Highly sensitive and selective fluorometric off–on K+ probe constructed via host–guest molecular recognition and aggregation-induced emission
Wang et al. A novel p-aminophenylthio-and cyano-substituted BODIPY as a fluorescence turn-on probe for distinguishing cysteine and homocysteine from glutathione
JP5526124B2 (en) Near-infrared fluorescent compound
Bai et al. Construction of an NIR and lysosome-targeted quinoline-BODIPY photosensitizer and its application in photodynamic therapy for human gastric carcinoma cells
Wang et al. Tetrahydro [5] helicene-based fluorescent probe for rapid and sensitive detection of bisulfite in living cells
CN114853656B (en) Carbazole derivative with AEE characteristic, preparation method and application
CN114591632B (en) Azaindole-hemicyanine dye, and synthetic method and application thereof
WO2023092814A1 (en) Amino-substituted chromenoquinoline type fluorescent marker, and preparation and use thereof
CN110804009A (en) Chemiluminescent substrate with high chemiluminescent intensity, long wavelength and good stability, and preparation method and application thereof
CN111793371B (en) 3, 5-asymmetrically modified BODIPY near-infrared fluorescent dye and preparation method thereof
Zhou et al. Rational engineering of novel rhodamines with stable far red to NIR emission and high brightness for in vivo imaging
Yang et al. Novel flavonoid derivatives and their corresponding rare earth complexes: Synthesis and luminescent properties
CN111303111B (en) Huang Jing near-infrared two-region dye, preparation method and fluorescence imaging application
Qin et al. DAD type based NIR fluorescence probe for monitoring the cysteine levels in pancreatic cancer cell during ferroptosis
Li et al. Two pH-responsive fluorescence probes based on indole derivatives
CN113416196B (en) benzothiadiazole-TB compound and synthesis method and application thereof
CN117003782A (en) TB-thiophene methylene malononitrile derivative and preparation and application thereof
CN114773875B (en) Azaindole-squaraine dye, and synthetic method and application thereof
CN113501836A (en) Star BODIPY near-infrared fluorescent dye and preparation method thereof
CN117003759A (en) TB-pyridine cyano vinyl derivative with viscosity response and synthesis and application thereof
CN117024462A (en) sulfonamide-naphthalimide-TB derivative and synthetic method and application thereof
CN113735864B (en) D-benzothiadiazole-TB (-D) derivative and synthetic method and application thereof
CN113583033B (en) benzothiadiazole-TB-fluoroboron complex and synthesis method and application thereof
CN117402165A (en) pyridine-TB derivatives, and synthetic method and application thereof

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination