Fluorine-containing nuclear magnetic resonance displacement reagent for chiral recognition and detection of cyclic secondary amine

Abstract

The invention belongs to the technical field of nuclear magnetic resonance organic analysis, and discloses a fluorine-containing nuclear magnetic resonance displacement reagent for chiral recognition and detection of cyclic secondary amine. The structural general formula of the fluorine-containing displacement reagent is shown in the specification. The fluorine-containing displacement reagent and an analyte generate distinguishable characteristic fluorine spectrum signals through derivatization, and the analyte to be detected does not need to have ultraviolet absorption or crystallinity. The fluorine-containing probe is used for chemical sensing detection based on the nuclear magnetic resonance fluorine spectrum technology, the fluorine-containing probe has the capacity of being rapidly combined with cyclic secondary amine to generate a thiourea adduct of a diastereomer of a rigid structure, and generated characteristic fluorine spectrum resonance signals accurately correspond to all components in a complex system of organic secondary amine; high-precision directional identification can be realized, and the method has the capability of simultaneously detecting various different organic secondary amine molecules in a complex system. R1 is methyl, a monofluorine atom or trifluoromethyl, R2 is hydrogen, methyl, a monofluorine atom or trifluoromethyl, and at least one of R1 and R2 is a monofluorine atom or trifluoromethyl.

Description

Technical Field

[0001] This invention belongs to the field of nuclear magnetic resonance organic analysis technology, specifically relating to a class of fluorine-containing nuclear magnetic resonance shift reagents for chiral recognition and detection of cyclic secondary amines, used for nuclear magnetic resonance fluorine spectroscopy analysis of cyclic secondary amines. Background Technology

[0002] Chiral organic amines, especially secondary amines, possess excellent physiological activity and serve as important pharmacophores in many bioactive substances, making them a promising area for drug development. However, current methods for distinguishing and detecting chiral amines still face challenges. On one hand, rapid chiral detection capabilities need improvement; on the other hand, novel mechanisms for simultaneous in-situ detection of multiple components are still rare.

[0003] Nuclear magnetic resonance (NMR) typically cannot directly distinguish enantiomers. Only when enantiomers are in a chiral chemical environment can their NMR shift signals show identifiable differences. Two common methods for NMR identification are: (1) under certain conditions, a chiral molecule is chemically reacted with the enantiomer to form a diastereomer through derivatization; (2) a chiral solvation reagent is added to a sample containing the enantiomer to generate non-covalent interactions, forming a transient diastereomeric complex that makes the sample exhibit asymmetry in the external environment. However, chiral identification and differentiation of amine compounds often require prior derivatization, and the methods used for derivatization vary. For example, fluorinated chiral derivatization reagents for aldehydes or reactive esters that react with amino groups can achieve baseline separation using NMR fluorine spectrometry (Anal. Chem. 2022, 94, 1867-1873; Anal. Chem. 2022, 94, 7853-7860). Another approach involves pre-derivatizing the analyte with a fluorinated acyl chloride to convert it into a fluorinated amide or ester, followed by the addition of a chiral metal complex shift reagent, combining derivatization and solvation to distinguish chiral amine compounds (Anal. Chem. 2022, 94, 1441-1446). More recently, chiral cyclic palladium acetonitrile complexes have been used to dynamically bind fluorinated palladium probes to amine compounds, generating fluorine resonance signals for each enantiomer (JACS Au 2023, 3, 1348-1357; Anal. Chem. 2024, 96, 730-736; Anal. Chem. 2024, 96, 11448-11454). These methods have the following drawbacks: the need for purification and separation of the synthesized derivatives, or the challenging preparation of the shift reagents. Currently, there are no literature reports on the specific chiral differentiation and identification of cyclic secondary amines using highly active isocyanate-based fluorinated NMR shift reagents, making research on this topic of practical application value. Summary of the Invention

[0004] In view of the current state of technology, the purpose of this invention is to provide a class of isocyanate-based fluorinated nuclear magnetic resonance shift reagents for the highly active chiral recognition and specific detection of cyclic secondary amines, enabling the use of fluorinated nuclear magnetic resonance shift reagents for drug preparation screening, quality control, and screening of prohibited chemicals in biological samples.

[0005] To achieve the objectives of this invention, a fluorinated NMR shift reagent based on isocyanate is first prepared and applied to the chemical sensing detection of fluorine in NMR spectroscopy. The fluorinated NMR shift reagent reacts directly with the analyte to produce a diastereomeric thiourea adduct with a specific structure, which is used for chiral analysis using NMR technology.

[0006] The specific technical solution of this invention is as follows:

[0007] The fluorine-containing nuclear magnetic resonance shift reagent for chiral recognition and detection of cyclic secondary amines described in this invention has the following general structural formula:

[0008]

[0009] R1 is methyl, monofluorine atom or trifluoromethyl, R2 is hydrogen, methyl, monofluorine atom or trifluoromethyl, and at least one of R1 and R2 is monofluorine atom or trifluoromethyl.

[0010] The following compounds are preferred:

[0011] R1=CH3,R2=2-F:probe 1;or ent-1

[0012] R1 = CH3, R2 = 3-F:probe 2

[0013] R1 = CH3, R2 = 4-F:probe 3

[0014] R1 = CH3, R2 = CF3:probe 4

[0015] R1 = CF3, R2 = H:probe 5

[0016] The fluorine-containing nuclear magnetic resonance shift reagent is achieved in the following manner:

[0017]

[0018] The preparation method is as follows:

[0019] Using fluorinated chiral primary amines as raw materials, sulfur phosgene reagent was added to toluene solvent and reacted at room temperature. After the reaction was completed, the samples were extracted and purified by column chromatography to obtain a class of fluorinated nuclear magnetic resonance probes based on isothiocyanate groups as functional groups.

[0020] This fluorine-containing nuclear magnetic resonance shift reagent enables highly sensitive detection of 28 cyclic secondary amines with important physiological activities. It can also be used for multi-component analysis, determination of chiral ligand purity, and quality control of natural products and other drugs.

[0021] The detection principle of this invention is as follows: The fluorine-containing NMR reagent, without any additives, derivatizes with a cyclic secondary amine to produce a diastereomeric thiourea adduct with a special structure, which is then used for chiral analysis using NMR technology. The product has a novel structure, and its general structural formula is:

[0022] R' = alkyl, aryl, ester, etc.

[0023] The advantages of this invention are as follows: 1. The fluorinated NMR shift reagent described in this invention is simple to synthesize and has a novel structure. 2. The fluorinated NMR shift reagent has good specificity and high sensitivity, enabling chiral analysis of 28 different structural types of cyclic secondary amines, and can be applied simultaneously in the multi-component chiral identification of 3 pairs of 6 chiral amines and the detection of chiral ligand purity. 3. This fluorinated NMR probe reacts rapidly with cyclic secondary amines to produce diastereomeric thiourea adducts. The bound products have high stability, and the resulting characteristic fluorine spectrum signals accurately correspond to each component in the complex system of organic secondary amines, enabling highly accurate directional identification. This method can simultaneously detect multiple different organic secondary amine molecules in complex systems, with high repeatability and accuracy, and can be well applied to the chiral differentiation and identification of cyclic secondary amines. Attached Figure Description

[0024] Figure 1 This is a diagram showing the distinguishing effect of the fluorine-containing NMR shift reagent of the present invention on different cyclic secondary amine analytes.

[0025] Figure 2 This is an application example of the fluorine-containing nuclear magnetic resonance shift reagent of the present invention for detecting the purity of chiral ligands and natural products.

[0026] Figure 3 This is a graph showing the accuracy analysis of the detection of chiral cyclic secondary amines using the fluorine-containing NMR shift reagent of this invention.

[0027] Figure 4 The diagram shows the effect of the fluorine-containing nuclear magnetic resonance shift reagent of the present invention being unable to recognize and distinguish chain primary amines, amides, pyridines, azoles, and thiol amino acids.

[0028] Figure 5 This is a single-crystal schematic diagram of the diastereomeric thiourea adduct with a special structure produced by the fluorine-containing NMR shift reagent of the present invention and the cyclic secondary amine.

[0029] Figure 6The spectrum of a stable diastereomeric thiourea adduct was generated by the combination of a fluorine-containing NMR shift reagent and the analyte 1-phenyltetrahydroisoquinoline. Detailed Implementation

[0030] The present invention will be further described below with reference to the accompanying drawings and specific embodiments, so that those skilled in the art can better understand and implement the present invention. The following embodiments are used to illustrate the present invention, but do not limit the scope of the present invention.

[0031] Example 1: Synthesis of a fluorinated NMR shift reagent for chiral recognition and detection of cyclic secondary amines:

[0032]

[0033] 1 mmol of chiral primary amine was dissolved in 10 mL of toluene solvent, 2 mmol of potassium carbonate was added, and the mixture was stirred for 0.5 hours. Then 1.5 mmol of phosgene was added, and the reaction was allowed to proceed for 12 hours. Thin-layer chromatography was used to detect the completeness of the reaction. The mixture was extracted sequentially with ethyl acetate, saturated sodium bicarbonate, and saturated brine. After drying, the solvent was removed by rotary evaporation to obtain a fluorinated nuclear magnetic resonance shift reagent.

[0034] Its characterization data are as follows:

[0035] Probe 1:(R)-configuration:yellow liquid,157.5mg,87%yield. (c 0.1, in CHCl3). 1 H NMR (400MHz, CDCl3) δ7.49–7.41(m,1H),7.37–7.26(m,1H),7.23–7.17(m,1H),7.11–7.03(m,1H),5.23(q,J=6.7Hz,1H),1.68(d,J=6.7Hz,2H). 13 C NMR (101MHz, CDCl3) δ160.35,157.89,129.94,129.85,127.47,127.34,126.88,126.84,124.72,124.68,115.85,115.64,51.23,23.81. 19 F NMR(376MHz,CDCl3)δ-119.40.HRMS(ESI)m / z:[MH] - calcd for C9H7FNS180.0283,found180.0251.

[0036] Probe ent-1:(S)-configuration:yellowliquid,159.3mg,88%yield. (c 0.1,in CHCl3). 1 H NMR(400MHz,CDCl3)δ7.49–7.41(m,1H),7.37–7.26(m,1H),7.23–7.17(m,1H),7.11–7.02(m,1H),5.23(q,J=6.8Hz,1H),1.69(d,J=6.7Hz,3H). 13 C NMR(101MHz,CDCl3)δ160.35,157.89,129.94,129.86,127.47,127.34,126.89,126.85,124.73,124.69,115.86,115.65,51.25,23.81. 19 F NMR(376MHz,CDCl3)δ-119.40.HRMS(ESI)m / z:[M-H] - calcd for C9H7FNS180.0283,found180.0278.

[0037] Probe 2:(R)-configuration:yellow liquid,83%yield. (c0.1,in CHCl3). 1 H NMR(400MHz,CDCl3)δ7.35(td,J=7.8,5.7Hz,1H),7.10(d,J=7.7Hz,1H),7.05(q,J=3.4Hz,1H),7.01(d,J=8.5Hz,1H),4.92(q,J=6.8Hz,1H),1.68(d,J=6.8Hz,3H). 13 C NMR(101MHz,CDCl3)δ164.21,161.75,142.73,142.66,130.61,130.52,121.12,121.09,115.28,115.07,112.79,112.56,56.54,56.52,24.84. 19 F NMR(376MHz,CDCl3)δ-111.71.HRMS(ESI)m / z:[M-H] - calcd for C9H7FNS 180.0283,found 180.0279.

[0038] Probe 3:(R)-configuration:yellow liquid,86%yield. (c0.1,in CHCl3). 1 H NMR(400MHz,CDCl3)δ7.30(dd,J=8.6,5.3Hz,2H),7.07(t,J=8.6Hz,2H),4.90(d,J=6.8Hz,1H),1.66(d,J=6.8Hz,3H). 13 C NMR(101MHz,CDCl3)δ163.67,161.22,136.09,136.06,127.28,127.20,115.93,115.72,56.45,24.96. 19 F NMR(376MHz,CDCl3)δ-113.76.HRMS(ESI)m / z:[M-H] - calcd for C9H7FNS180.0283,found 180.0271.

[0039] Probe 4:(R)-configuration:yellow liquid,88%yield. (c0.1,in CHCl3). 1 H NMR(400MHz,CDCl3)δ7.72(d,J=8.0Hz,1H),7.65(dd,J=7.9,6.1Hz,2H),7.44(t,J=7.6Hz,1H),5.32(q,J=6.6Hz,1H),1.69(d,J=6.6Hz,3H). 13 C NMR(101MHz,CDCl3)δ139.19,132.84,128.35,127.50,126.69,126.39,126.03,125.97,125.92,125.86,125.35,122.62,53.11,53.08,25.53. 19 F NMR(376MHz,CDCl3)δ-58.54.HRMS(ESI)m / z:[M-H] + calcd for C 10 H7F3NS230.0251,found 230.0247.Probe 5:(R)-configuration:yellowliquid.88%yield. (c 0.11,CHCl3). 1H NMR (400MHz, CDCl3) δ7.28 (s, 5H), 5.00 (q, J = 6.4Hz, 1H). 13 C NMR (101MHz, CDCl3) δ141.08,130.37,130.22,129.07,127.83,126.80,123.99,121.18,118.37,62.78,62.45,62.11,61.78. 19 F NMR(376MHz,CDCl3)δ-75.19.HRMS(ESI)m / z:[MH] - calcd forC9H5F3NS216.0095, found 216.0087.

[0040] Example 2: Fluorine-containing NMR shift reagents are used for chemical sensing detection based on NMR fluorine spectroscopy. When the fluorine-containing probe reagent Probe 1 is mixed with optically pure, racemic, or optically active non-optically pure analytes in an organic solvent, strong fluorine spectral signals can be observed during derivatization, and the characteristic fluorine spectral signals of different analytes can be clearly distinguished.

[0041] A racemic cyclic secondary amine analyte at a concentration (10-60 mM) was dissolved in deuterated chloroform, and a fluorinated NMR probe solution (10 mM) was prepared simultaneously. Then, 100 μL of the fluorinated NMR probe solution (10 mM) and 500 μL of the analyte solution (10 mM) were mixed, preferably at a probe-to-analyte volume ratio of 1:5. The mixture was then transferred to an NMR tube and immediately subjected to NMR analysis. 19 F NMR nuclear magnetic resonance analysis was performed under the same acquisition parameters. 19 The emission frequency of the F atom nucleus was 376 MHz (after 32 cumulative scans), yielding highly sensitive NMR fluorine spectra of 28 cyclic secondary amines with important physiological activities, such as... Figure 1 As shown. Examples of applications of the fluorinated nuclear magnetic resonance shift reagent Probe 1 in detecting the purity of chiral ligands and natural products include... Figure 2 As shown in Table 1, the chiral ee value analysis data for 1-phenyltetrahydroisoquinoline are presented below, and their accuracy is as follows: Figure 3 As shown.

[0042]

[0043]

[0044] Conversely, the fluorinated nuclear magnetic resonance shift reagent Probe 1 does not exhibit any recognition or differentiation effect with a series of chain primary amines, amides, pyridines, azoles, or thiol amino acids, such as... Figure 4 As shown.

[0045] Example 3: Analysis of stable diastereomeric thiourea adducts generated by the binding of fluorinated NMR shift reagents with cyclic secondary amines.

[0046] This invention designs and synthesizes a series of chiral fluorine-containing NMR shift reagents. When these NMR probe molecules (taking Probe 1 as an example) bind to chiral analytes with different configurations, they rapidly generate rigid and stable diastereomeric thiourea adducts. Combined with the high sensitivity of fluorine to its surrounding environment, the stable thiourea adducts produce distinguishable fluorine NMR signals, thereby achieving configuration determination and enantiomeric composition detection of the chiral analyte, achieving a second-level response for detection upon mixing. This method does not require the sample to have UV absorption or fluorescence, and does not require sample purification for chiral analysis. Simultaneously, this method can also achieve precise chiral identification of six cyclic secondary amines in a mixture. Leveraging the rapid and highly sensitive characteristics of fluorine NMR detection, these fluorine-containing chiral NMR shift reagents can achieve specific chiral analysis of cyclic secondary amines. This chemical sensing method is simple, rapid, and accurate. Based on these advantages, this method has significant research and application value in drug preparation screening, quality control, and screening of prohibited chemicals in biological samples. The reaction process is shown below:

[0047]

[0048] R' = alkyl, aryl, ester, etc.

[0049] When the cyclic secondary amine is analyte 1-phenyltetrahydroisoquinoline, the resulting adduct is restricted from free rotation due to the rigidity of the nitrogen atoms on the ring. A schematic diagram of the single crystal of the rigid diastereomeric adduct is shown below. Figure 5 As shown, its fluorine NMR signal exhibits diastereomeric peaks at -118.60 ppm and -118.88 ppm, respectively, and the resulting signal chemical shift is stable and does not change with concentration, indicating that the probe binds to the cyclic secondary amine and the measured product has strong stability. Figure 6 As shown.

[0050] The physical characterization data of the diastereomeric adduct thiourea are as follows:

[0051] R, RI: white solid, 92.8% yield, Mp=165.8–166.1℃, (c0.1,in CHCl3). 1H NMR(400MHz,CDCl3)δ7.38–7.27(m,4H),7.25–7.02(m,10H),6.19(d,J=7.9 Hz,1H),5.94(q,J=7.3 Hz,1H),3.96(t,J=6.2 Hz,2H),2.99–2.78(m,2H),1.49(d,J=7.0 Hz,3H). 13 C NMR(101 MHz,CDCl3)δ181.12,162.30,159.87,140.92,136.49,135.07,130.27,130.14,129.26,129.21,129.02,128.93,128.64,128.18,128.11,127.63,127.49,126.90,126.67,124.50,124.46,116.13,115.92,62.62,52.85,44.23,28.14,21.68. 19 FNMR(376 MHz,CDCl3)δ-118.60.HRMS(ESI)m / z:[M+H] + calcd for C 24 H 24 FN2S 391.1644,found 391.1649.

[0052] R,S-I:white solid,93.8%yield,M.p.=163.5–163.8℃, (c0.1,in CHCl3). 1 H NMR(400 MHz,CDCl3)δ7.34–7.26(m,3H),7.26–7.14(m,8H),7.09–6.93(m,2H),6.22(d,J=7.5 Hz,1H),5.92(q,J=6.8 Hz,1H),4.00–3.86(m,2H),2.96–2.78(m,2H),1.60(d,J=7.0 Hz,3H). 13C NMR(101 MHz,CDCl3)δ181.10,162.16,159.73,140.98,136.41,134.99,130.30,130.17,129.17,129.12,128.83,128.62,128.23,128.10,127.59,127.47,126.95,126.63,124.30,124.27,115.96,115.74,62.50,52.66,44.01,28.12,21.96. 19 F NMR(376 MHz,CDCl3)δ-118.88.HRMS(ESI)m / z:[M+H] + calcd for C 24 H 24 FN2S391.1644,found 390.1649.。

Claims

1. A fluorine-containing nuclear magnetic resonance shift reagent, characterized in that, Its general molecular formula is as follows: R1 is methyl, monofluorine atom or trifluoromethyl, R2 is hydrogen, methyl, monofluorine atom or trifluoromethyl, and at least one of R1 and R2 is monofluorine atom or trifluoromethyl.

2. The fluorine-containing nuclear magnetic resonance shift reagent as described in claim 1, characterized in that, Choose the following compounds: R1 = CH3, R2 = 2-F R1 = CH3, R2 = 3-F R1 = CH3, R2 = 4-F R1 = CH3, R2 = CF3 R1 = CF3, R2 = H.

3. The application of the fluorine-containing nuclear magnetic resonance shift reagent as described in claim 1, characterized in that, It was applied to chemical sensing detection in nuclear magnetic resonance fluorine spectroscopy for chiral identification and detection of cyclic organic secondary amines.

4. The application of the fluorine-containing nuclear magnetic resonance shift reagent as described in claim 3, characterized in that, The cyclic secondary amine has the following general structural formula: R' is an alkyl, aryl, or ester group.

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