Beta-cyclodextrin derivative chiral stationary phase based on BINAM as well as preparation method and application of beta-cyclodextrin derivative chiral stationary phase

Abstract

The invention relates to the technical field of chiral stationary phase fillers, in particular to a BINAM-based beta-cyclodextrin derivative chiral stationary phase as well as a preparation method and application thereof. The structure of the BINAM-based beta-cyclodextrin derivative chiral stationary phase is as follows: a BINAM derivative is connected to the 6th site of beta-cyclodextrin and is bonded to the surface of silica gel through carbamate, and the structure is as shown in formula I in the specification. The novel beta-cyclodextrin CSP filler provided by the invention can be used for separating chiral compounds with various structure types, is suitable for different mobile phase systems of high performance liquid chromatography (HPLC), and also has good stability in mobile phases with strong polarity and high water content. The CSP filler provided by the invention can be used for splitting various chiral drugs such as imidazole antibacterial drugs, beta receptor blockers and the like and various triazole pesticides. And the requirements of daily analysis, production process and use process quality control and the like of chiral drugs and chiral pesticides can be met.

Description

Technical Field

[0001] The present invention relates to the technical field of chiral stationary phase fillers, and in particular to a BINAM-based β -Cyclodextrin derivative chiral stationary phase and its preparation method and application. Background Art

[0002] Chirality is one of the basic properties of life systems. Biomacromolecules such as enzymes, receptors, plasma proteins and ion channels all have chirality. Life itself is a chiral environment ( J. Am. Chem. Soc. 2024, 146, 26,17765–17772). When chiral drugs enter the human body, there may be stereoselective differences in chiral recognition between enantiomers and macromolecules in the body, resulting in different pharmacodynamics, pharmacokinetics and toxicology between enantiomers, and thus different pharmacological effects and adverse reactions. Nearly half of the drugs in clinical practice are chiral, but most of them are used clinically as racemates. Usually only one of the enantiomers exerts its efficacy, such as β -receptor blockers are an important class of drugs widely used in clinical practice. β -Beta-blockers contain at least one chiral center, and different enantiomers have significant differences in pharmacodynamics. S The activity of the -enantiomer is R -50-500 times the enantiomer. Except for timolol, chiral β - Most of the receptor blockers are racemic for clinical use, and the side effects such as dizziness and hallucinations caused by medication are considered to be related to R -Isomer related ( Molecules , 2021, 26: 468).

[0003] The research on chiral pesticides has also begun to attract widespread attention. Among racemic pesticides, half of them may be inactive. Spreading them on farmland not only pollutes the environment, but also causes waste of resources. According to statistics, among the more than 650 pesticides on the market, 173 are chiral, and there are only dozens of single enantiomer pesticides. With the continuous improvement of environmental safety requirements, the development of single enantiomer chiral pesticides has been put on the agenda ( Environ. Monitor. Assess . 2024, 196, 153.).

[0004] Therefore, establishing separation and analysis methods for chiral drug and chiral pesticide enantiomers is a key prerequisite for the pharmacodynamics, pharmacokinetics, interaction studies and quality control during the use of chiral drugs and chiral pesticide enantiomers, and has important practical significance for the research, development, evaluation and use of high-efficiency, low-toxicity and specific drugs / pesticides.

[0005] The HPLC chiral stationary phase method can be used not only for analysis but also for the preparation and semi-preparation of enantiomers due to its high efficiency, high sensitivity, wide application range, and the ability to be combined with other instruments. It has become the preferred method for the separation of chiral drug enantiomers and the in vivo and in vitro detection of chiral drugs.

[0006] β -Cyclodextrin ( β - cyclodextrins, β - CD), composed of 7 pyranose glucose units forming a conical structure, forming an inner hydrophobic cavity and an outer hydrophilic surface. This unique cavity structure can form an inclusion complex with chiral molecules and plays a decisive role in chiral separation. Chemical modification of the hydroxyl groups of cyclodextrin can not only change the shape and size of its cavity to meet various spatial requirements, but also the introduced substituents can provide different action sites. This is conducive to expanding the chiral separation flux of the stationary phase and improving its chiral selectivity. At present, most of the cyclodextrin chiral columns on the market are fully derivatized with carbamates on the hydroxyl groups of cyclodextrin, which has limited separation effects and is only suitable for the separation of some chiral drugs. 1,1'-Binaphthyl 2,2'-diamine (BINAM) is a typical axial chiral compound. This type of compound has a C2 symmetry axis and contains two identical naphthalene units. The two naphthalene rings hinder the free rotation of the 1,1'-bond, so the chiral binaphthyl molecule has a stable chiral configuration. In recent years, it has been widely used as a hot molecule in asymmetric catalysis and chiral synthesis research ( Tetrahedron 2024, 167, 134274.). Summary of the invention

[0007] The object of the present invention is to provide a method based on BINAM. β -Cyclodextrin derivative chiral stationary phase (CSP) and its preparation method and application. The present invention combines BINAM derivatives and β -Cyclodextrin, two traditional chiral selectors, are combined to prepare a new chiral stationary phase to enhance its chiral recognition ability and application range.

[0008] In order to achieve the above-mentioned object of the invention, the technical solution adopted by the present invention is: In the first aspect, a method is provided based on 1,1'-binaphthyl 2,2'-diamine (BINAM). β -Cyclodextrin derivative chiral stationary phase, β -A BINAM derivative is connected to the 6-position of cyclodextrin and bonded to the silica surface via carbamate. The structure is shown in Formula I:

[0009] Formula I R is selected from:

[0010] The configuration of BINAM is R -or S -Configuration.

[0011] In the second aspect, a method based on 1,1'-binaphthyl 2,2'-diamine (BINAM) is provided. β - A method for preparing a cyclodextrin derivative chiral stationary phase, comprising the following steps: S1. Dry β -cyclodextrin is dissolved in an aqueous sodium hydroxide solution, and a solution of p-toluenesulfonyl chloride in acetonitrile is added to react at room temperature or under ice bath conditions. After the reaction is completed, impurities are filtered out, and the filtrate is adjusted to pH≈7 with hydrochloric acid (HCl, 30-38%), and refrigerated at 4°C overnight to precipitate. The solid is filtered and recrystallized twice with a small amount of hot water, filtered and dried to obtain a mono-6-p-toluenesulfonyl- β -cyclodextrin (TsO- β CD).

[0012]

[0013] Formula II S2. Take the TsO- β CD was added to ammonia water, stirred and reacted at 40-70°C for 10-24h, and then most of the solvent was removed under reduced pressure, and then dropped into acetone solution to form a precipitate, which was filtered and dried to obtain 6-amino-6-deoxy- β -Cyclodextrin (NH2- β CD).

[0014]

[0015] Formula III S3. BINAM was dissolved in anhydrous dichloromethane, 4-nitrobenzene isocyanate was added to react, and compound AM-1 was obtained by purification by silica gel column chromatography, as shown in Formula IV.

[0016]

[0017] Formula IV S4. Dissolve compound AM-1 in anhydrous dichloromethane, add phenyl isocyanate containing different substituents to react, and purify by silica gel column chromatography to obtain compound AM-2, as shown in Formula V.

[0018]

[0019] Formula V R is selected from:

[0020] S5. Place compound AM-2 in a round-bottom flask, add iron, ethanol and hydrochloric acid, and heat to react for 5 to 12 hours. After the reaction is completed, adjust the pH to 7 with saturated sodium bicarbonate (NaHCO3), extract with ethyl acetate (EtOAc) three times, combine the EtOAc layers, remove the solvent by vacuum distillation, and purify by silica gel column chromatography to obtain compound AM-3, as shown in Formula VI.

[0021]

[0022] Formula VI R is selected from:

[0023] S6. AM-3 was dissolved in ethyl acetate or dichloromethane, thiophosgene and saturated NaHCO3 aqueous solution were added, and after the reaction was completed, the ethyl acetate layer was separated with a separatory funnel, anhydrous sodium sulfate was added and dried, the solution was distilled under reduced pressure, and the compound AM-4 was purified by silica gel column chromatography, as shown in Formula VII:

[0024] Formula VII R is selected from:

[0025] S7. Compound AM-4 (Formula VII), NH2-CD (Formula III) and a catalytic amount of 4-dimethylaminopyridine (DMAP) were added to N,N-dimethylformamide (DMF) for reaction for 15-30 hours, then dropped into acetone to form a precipitate and filtered, the solid was washed with acetone for several times, and vacuum dried at 60°C to obtain a product as shown in Formula VIII. β -Cyclodextrin-BINAM derivative (AM-CD).

[0026]

[0027] Formula VIII R is selected from:

[0028] S8. AM-CD and isocyanatepropyltriethoxysilane were added to DMF to react for a period of time, and activated silica gel was added to continue the reaction. After cooling to room temperature, the solid was filtered, and the obtained solid was washed repeatedly with DMF and methanol for several times, and then vacuum dried to obtain a BINAM-based compound as shown in Formula I. β - Cyclodextrin derivative CSP filler.

[0029] Furthermore, in S1 βThe molar mass ratio of -cyclodextrin and p-toluenesulfonyl chloride is 1:0.8~1.5; the concentration of the sodium hydroxide aqueous solution is 5~10 mol / L, and the amount of acetonitrile solution is 2~5 mL of acetonitrile per gram of p-toluenesulfonyl chloride; the reaction is carried out at room temperature and the reaction time is 5~12 h.

[0030] Further, the amount of ammonia in S2 is per gram of TsO- β Add 20~40mL of ammonia water to CD.

[0031] Furthermore, the configuration of BINAM in S3 is R -or S -configuration, and the molar mass ratio of it to p-nitrobenzene isocyanate is 1:1~2; the reaction temperature is room temperature, and the reaction time is 2~10 h.

[0032] Further, the phenyl isocyanates with different substituents in S4 are: 4-NitrophenylIsocyanate, 4-Chlorophenyl Isocyanate, p-Tolyl Isocyanate, Phenyl Isocyanate, 4-Methoxyphenylisocyanate, 4-(Trifluoromethyl)PhenylIsocyanate, 4-(Dimethylamino)phenylisocyanate, 1-(Tert-Butyl)-4-Isocyanatobenzene, 3,5-Dimethylphenylisocyanate, 4-(Trifluoromethyl)-4-isocyanatobenzene ... Isocyanate), 3,5-dichlorophenyl Isocyanate, the molar mass ratio of AM-1 to phenyl isocyanate with different substituents is 1:1~2; the reaction temperature is room temperature, and the reaction time is 2~10 h.

[0033] Furthermore, the molar mass ratio of AM-2 to iron in S5 is 1:5~20; the ratio of ethanol to hydrochloric acid is 4:3 (v / v); the reaction temperature is 60~80 ℃; and the reaction time is 10~24 h.

[0034] Furthermore, the molar mass ratio of AM-3 to thiophosgene in S6 is 1:1~2; the amount of saturated NaHCO3 aqueous solution used is 10 mL per gram of AM-3; the reaction is carried out at room temperature or in an ice bath, and the reaction time is 0.5~2 h.

[0035] Furthermore, NH2- β The molar mass ratio of CD, AM-4 and DMAP is 1:1~2:0.1; the reaction is carried out under nitrogen protection; the amount of solvent DMF is based on 1:1 per gram of NH2- β Add 10-20 mL of CD and the reaction time is 20-24 h.

[0036] Furthermore, all reactions in S8 were carried out under nitrogen protection; the molar mass ratio of AM-CD to triethoxypropyl isocyanate was 1:2~5; the reaction time was 4~10 h; the reaction temperature after adding activated silica gel was 110~120°C, and the reaction time was 20~36 h.

[0037] Furthermore, the preparation method of the activated silica gel in S8 is as follows: the chromatographic stationary phase silica gel is heated to reflux with a 10% hydrochloric acid solution, cooled and washed with distilled water until pH ≈ 7, and vacuum dried at 120°C for 12 hours to obtain the obtained product, wherein the amount of the hydrochloric acid solution is determined by 10-20 mL per gram of silica gel. The reaction temperature is 100-120°C, and the reaction time is 10-20 hours.

[0038] In a third aspect, a BINAM-based β -Application of cyclodextrin derivative chiral stationary phase in the separation of chiral compounds. The chiral compounds that can be separated include: imidazole antibiotics, β Beta-blockers, nimodipine, aminoglutethimide, equol and triazole chiral pesticides.

[0039] The beneficial effects of the present invention are: 1. The raw materials of the product of the present invention are widely available, the reaction conditions are mild and the cost is low.

[0040] 2. The present invention combines BINAM derivatives with β -CSP filler prepared by combining cyclodextrin, plays a role in β - While the cyclodextrin inclusion complex is acting, BINAM derivatives can provide more action sites, which is different from the underivatized β -Compared with cyclodextrin CSP, the chiral separation effect and separation range of cyclodextrin stationary phase are greatly enhanced.

[0041] 3. Compared with the traditional polymer CSP, the CSP filler prepared by the present invention has a clear structure. BINAM derivatives are characterized by nuclear magnetic resonance 1 H NMR, 13 C NMR combined with mass spectrometry (MS) was used for structure determination. β -The structures of cyclodextrin derivatives and CSP fillers were characterized by elemental analysis (EA), thermogravimetric analysis (TG), etc.

[0042] 4. In the product of the present invention, the cyclodextrin derivative is bonded to the silica gel through a urea bond, has good stability, and can be applied to normal phase, reverse phase and polar mobile phase HPLC. It is resistant to acid and alkali (pH = 4~10) and high pressure (40 MPa), and can still maintain good chiral separation performance in highly polar and high water content mobile phases.

[0043] 5. The CSP filler prepared by the present invention can be applied to the separation of chiral compounds of various structural types, and can separate imidazole antibiotics, β Receptor blockers, nimodipine, aminoglutethimide, equol and a variety of triazole chiral pesticides, etc., can meet the daily analysis and production of these chiral drugs and chiral pesticides, and the quality control during clinical use. BRIEF DESCRIPTION OF THE DRAWINGS

[0044] Figure 1 The synthetic route diagram of the embodiment of the present invention; Figure 2 This is the chromatogram of enantiomeric separation of 9 chiral drugs in Example 1; Figure 3 This is the chromatogram of the enantiomer separation of 6 chiral pesticides in Example 1. DETAILED DESCRIPTION

[0045] The specific implementation modes of the present invention are described below so that those skilled in the art can understand the present invention. However, it should be clear that the present invention is not limited to the scope of the specific implementation modes. For those of ordinary skill in the art, as long as various changes are within the spirit and scope of the present invention as defined and determined by the attached claims, these changes are obvious, and all inventions and creations utilizing the concept of the present invention are protected.

[0046] Example 1 Reference Figure 1 , a method based on ( S )-BINAM β - A method for preparing a cyclodextrin derivative chiral stationary phase, comprising the following steps: (1) Preparation of activated silica gel Add 20.0 g of chromatographic stationary phase silica gel and 200 ml of 10% hydrochloric acid solution into a 500 ml round-bottom flask, heat and reflux for 24 h, cool to room temperature, wash with distilled water until pH ≈ 7, and vacuum dry at 120°C for 12 h to obtain activated silica gel.

[0047] The chromatographic stationary phase silica gel used was produced by Qingdao Meigao Chemical Co., Ltd., China, with a particle size of 5 m m, specific surface area 214m 2 / g.

[0048] (2) 6-amino-6-deoxy-β - Preparation of cyclodextrin 50g β -cyclodextrin was dispersed in 100 ml pure water, and 16 ml 8.25 mol / L NaOH aqueous solution was added to dissolve it. Under ice bath condition, 8 g p-toluenesulfonyl chloride acetonitrile solution was added, and the reaction was carried out at room temperature for 12 h. Impurities were filtered, and the filtrate was adjusted to pH ≈ 7 with HCl. The precipitate was precipitated at 4 ° C overnight, filtered, and the precipitate was recrystallized twice with a small amount of hot water, and vacuum dried to obtain mono-6-p-toluenesulfonyl- β -cyclodextrin (TsO- β CD), yield 10%. The structure is shown in formula II.

[0049]

[0050] Formula II Take 5 g of dry mono-6-toluenesulfonyl- β -cyclodextrin, add 165 mL of concentrated ammonia water, stir and react at 50 ° C for 20 h, then remove most of the solvent under reduced pressure, drop into acetone to form a white flocculent precipitate, filter and dry to obtain 6-amino-6-deoxy- β -Cyclodextrin (NH2- β CD), yield 95%. The structure is shown in formula III.

[0051]

[0052] Formula III (3) Preparation of BINAM derivatives 2g ( S )-BINAM was dissolved in 20 mL of dichloromethane, 1.15 g of p-nitrobenzene isocyanate was added, and the mixture was stirred at room temperature for 4 h. The crude product was purified by normal phase silica gel column chromatography and eluted with petroleum ether-ethyl acetate (3:1) to obtain a yellow solid product AM-1 with a yield of 70%. The structure is shown in Formula IV.

[0053]

[0054] Formula IV 2 g of compound AM-1 was dissolved in 20 mL of dichloromethane, 1.1 g of p-nitrobenzene isocyanate was added, and the mixture was stirred at room temperature for 4 h. The crude product was purified by normal phase silica gel column chromatography and eluted with petroleum ether-ethyl acetate (3:1) to obtain a yellow solid product AM-2a with a yield of 90%. The structure is shown in Formula V-a.

[0055]

[0056] Formula V-a 2 g of compound AM-2a (Formula Ⅴ-a) and 1.8 g of iron were placed in a round-bottom flask, 20 ml of EtOH and 15 ml of HCl were added, and the mixture was reacted in an oil bath at 75 °C for 12 h. The pH was then adjusted to 7 with NaHCO3, and the mixture was extracted three times with EtOAc. The solvent was removed by distillation under reduced pressure, and the crude product was purified by normal phase silica gel column chromatography. The yellow solid product AM-3a was eluted with petroleum ether-ethyl acetate (1:1) with a yield of 20%. The structure is shown in Formula Ⅵ-a.

[0057]

[0058] Formula VI-a 0.5 g of compound AM-3a was dissolved in 5 mL of ethyl acetate, and 0.12 mL of thiophosgene and 5 mL of saturated NaHCO3 aqueous solution were added. The mixture was reacted in an ice bath for 0.5 h. The ethyl acetate layer was separated with a separatory funnel and dried with anhydrous sodium sulfate. The solution was distilled under reduced pressure. The crude product was purified by normal phase silica gel column chromatography and eluted with petroleum ether-ethyl acetate (8:1) to obtain a brown solid product AM-4a with a yield of 95%. The structure is shown in Formula VII-a.

[0059]

[0060] Formula VII-a AM-4a NMR and mass spectrometry data: 1 H NMR (600 MHz, Chloroform- d ) d 8.14 (d, J = 9.0Hz, 1H), 7.92 (d, J = 11.1 Hz, 2H), 7.88 (m, 1H), 7.83 (m, 2H), 7.78 (m, 3H),7.36 (q, J = 7.5 Hz, 2H), 7.19 (m, 2H), 7.14 (m, 1H), 7.09 (m, 3H), 7.01 (d, J =8.5 Hz, 1H), 6.95 (d, J = 8.5 Hz, 1H), 6.82 (s, 1H), 6.79 (s, 4H). 13 C NMR (150MHz, CDCl3) d154.10, 152.75, 144.93, 142.07, 136.48, 135.54, 134.97, 134.85,132.94, 132.84, 131.38, 131.11, 129.84, 129.81, 128.47, 128.40, 127.50,127.46, 126.86, 126.40, 125.93, 125.69, 125.40, 125.14, 125.12, 123.09,122.87, 122.09, 121.51, 118.10. MS (ESI) calculated for [M+H] + 625.

[0061] (4) BINAM-based β - Preparation of Cyclodextrin Derivatives Take 0.4g of compound AM-4a and 0.5g of NH2- β CD was dissolved in 20 mL DMF, 10 mg DMAP was added, and the mixture was reacted at 85 °C for 24 h under nitrogen protection. After the reaction was completed, it was dropped into 200 mL acetone to precipitate. m m) was filtered and washed with acetone for several times, and dried under vacuum at 60°C for 24 h to obtain a white solid product AM-CDa with a yield of 70% and a structure as shown in Formula VIII-a.

[0062]

[0063] Formula VIII-a (5) Based on BINAM β - Preparation of Cyclodextrin Derivative CSP Filler Take 0.6g AM-CDa and dissolve it in DMF, add 0.22mL 3-isocyanatepropyltriethoxysilane, and stir and react at 65℃ for 6 hours under nitrogen protection. Then add 2.2g activated silica gel, heat to 115℃ and react at constant temperature for 24 hours. The filtered solid is repeatedly washed with DMF, methanol and acetone in turn, and vacuum dried at 100℃ to obtain CSP filler, the structure of which is shown in formula Ⅰ-a. Elemental analysis shows that its carbon content is 3.06%, hydrogen content is 1.27%, and nitrogen content is 0.19%; the weight loss rate of thermogravimetric analysis is 8.76%.

[0064]

[0065] Formula Ⅰ-a (6) Packing of chiral chromatographic columns The chiral stationary phase was loaded into the empty column by homogenization method. First, the empty column (150×4.6 mm) was connected to the lower end of the homogenization tank, 2.1 g of CSP filler was weighed into a 50 mL beaker, 30 mL of homogenate (methanol) was added, and it was ultrasonicated for 1 min to make it a suspension, and then transferred to the homogenization tank. Methanol was used as the displacement liquid, and it was quickly filled into the empty column at a pressure of 40 MPa and maintained for 30 min to prepare a chiral chromatographic column.

[0066] Example 2 A method based on ( S )-BINAM β - A method for preparing a chiral stationary phase of a cyclodextrin derivative, the steps of which are substantially the same as those of Example 1, except that the reagent reacting with AM-1 is p-chlorophenyl isocyanate.

[0067] The NMR and mass spectrometry data of AM-4b are: 1 H NMR (600 MHz, Chloroform- d ) d 7.89 (d, J =9.1 Hz, 1H), 7.85 (m, 3H), 7.81 (t, J = 7.9 Hz, 2H), 7.37 (m, 2H), 7.20 (s,1H), 7.13 (q, J = 6.9 Hz, 2H), 7.06 (s, 1H), 6.95 (d, J = 8.5 Hz, 1H), 6.92 (d, J =8.5 Hz, 1H), 6.87 (d, J = 8.5 Hz, 2H), 6.84 (s, 1H), 6.79 (s, 1H), 6.75 (d, J =8.5 Hz, 2H), 6.62 (d, J = 8.3 Hz, 2H), 6.56 (d, J = 8.3 Hz, 2H). 13 C NMR (150 MHz,CDCl3) d154.12, 153.61, 136.71, 135.82, 135.20, 135.11, 132.78, 132.74,131.19, 131.08, 129.57, 129.10, 128.42, 128.40, 127.32, 126.41, 126.32,125.71, 125.60, 125.23, 125.10, 122.99, 122.62, 122.46, 122.21, 121.69,121.24. MS (ESI) calculated for [M+H] + 614. The BINAM- β -The structure of cyclodextrin CSP is shown in formula Ⅰ-b. Elemental analysis shows that its carbon content is 3.52%, hydrogen content is 1.63%, and nitrogen content is 0.43%; the weight loss rate of thermogravimetric analysis is 6.26%.

[0068]

[0069] Formula I-b Example 3 A method based on ( S )-BINAM β - A method for preparing a chiral stationary phase of a cyclodextrin derivative, the steps of which are substantially the same as those of Example 1, except that the reagent reacting with AM-1 is p-toluene isocyanate.

[0070] The NMR and mass spectrometry data of AM-4c are: 1 H NMR (600 MHz, Chloroform- d ) δ 8.44 (d, J = 9.1 Hz, 1H), 8.13 (d, J = 8.9 Hz, 1H), 7.98 (d, J = 8.8 Hz, 1H), 7.95 (d, J =9.1 Hz, 1H), 7.89 (d, J = 8.2 Hz, 1H), 7.87 (d, J = 8.1 Hz, 1H), 7.44 – 7.41 (m,1H), 7.36 (d, J = 8.0 Hz, 2H), 7.25 – 7.22 (m, 1H), 7.18 – 7.15 (m, 1H), 7.03(t, J= 9.7 Hz, 3H), 6.92 (d, J = 8.3 Hz, 3H), 6.78 (d, J = 7.0 Hz, 3H), 6.68 (d, J =7.9 Hz, 2H), 6.53 (s, 1H), 6.46 (s, 1H), 2.21 (s, 3H). 13 C NMR (150 MHz, CDCl3) d 154.98, 153.05, 137.37, 135.56, 134.93, 132.79, 132.63, 131.19, 130.55,129.75, 129.60, 129.50, 128.22, 127.34, 127.18, 126.41, 126.20, 125.70,125.62, 125.30, 124.98, 124.64, 122.92, 122.40, 121.26, 120.55, 20.84. MS (ESI) calculated for [M+H] + 594. The BINAM- β -The structure of cyclodextrin CSP is shown in formula Ⅰ-c. Elemental analysis shows that its carbon content is 2.61%, hydrogen content is 1.87%, and nitrogen content is 0.32%. The weight loss rate of thermogravimetric analysis is 6.58%.

[0071]

[0072] Formula I-c Example 4 A method based on ( S )-BINAM β - A method for preparing a chiral stationary phase of a cyclodextrin derivative, the steps of which are substantially the same as those of Example 1, except that the reagent reacting with AM-1 is phenyl isocyanate.

[0073] The NMR and mass spectrometry data of AM-4d are: 1 H NMR (600 MHz, Chloroform- d ) d 8.28 (d, J =9.0 Hz, 1H), 8.05 (d, J = 9.0 Hz, 1H), 7.93 (m, 2H), 7.86 (m, 2H), 7.39 (m,3H), 7.22 (d,J = 7.8 Hz, 1H), 7.16 (t, J = 7.7 Hz, 1H), 7.00 (t, J = 8.3 Hz, 4H), 6.93 (m, 6H), 6.86 (m, 2H), 6.80 (d, J = 7.7 Hz, 2H). 13 C NMR (150 MHz, CDCl3) d 154.50, 153.20, 137.10, 136.80, 135.36, 134.91, 134.52, 132.77, 132.70,131.19, 130.66, 129.59, 129.09, 128.26, 128.21, 127.27, 127.18, 126.41,126.19, 125.79, 125.66, 125.27, 125.14, 124.81, 124.62, 123.02, 121.54,121.41, 120.73. MS(ESI)calculated for [M+H] + 580. The BINAM- β The structure of -cyclodextrin CSP is shown in formula Ⅰ-d. Elemental analysis shows that it contains 2.54% carbon, 2.62% hydrogen and 0.50% nitrogen. The weight loss rate of thermogravimetric analysis is 6.09%.

[0074]

[0075] Formula I-d Example 5 A method based on ( S )-BINAM β - A method for preparing a chiral stationary phase of a cyclodextrin derivative, the steps of which are substantially the same as those of Example 1, except that the reagent reacting with AM-1 is 4-methoxyphenyl isocyanate.

[0076] The NMR and mass spectrometry data of AM-4e are: 1 H NMR (600 MHz, Chloroform- d ) d 8.13 (m,1H), 7.94 (m, 1H), 7.84 (m, 2H), 7.79 (t, J = 8.3 Hz, 2H), 7.36 (m, 3H), 7.17(t,J = 7.7 Hz, 1H), 7.09 (t, J = 8.2 Hz, 1H), 6.98 (d, J = 8.5 Hz, 1H), 6.85 (d, J =8.4 Hz, 2H), 6.76 (m, 4H), 6.69 (m, 1H), 6.57 (m, 1H), 6.49 (d, J = 8.4 Hz,2H), 6.40 (d, J = 8.7 Hz, 2H), 3.70 (s, 3H). 13 C NMR (150 MHz, CDCl3) d 153.99,152.02, 136.05, 134.35, 134.22, 133.65, 131.64, 131.48, 130.04, 129.61,128.44, 128.38, 127.19, 126.12, 126.02, 125.11, 124.76, 124.44, 124.10,124.01, 123.67, 121.81, 120.89, 120.52, 119.85, 113.24, 54.30. MS (ESI) calculated for [M+H] + 610. The BINAM- β The structure of -cyclodextrin CSP is shown in formula Ⅰ-e. Elemental analysis shows that it contains 4.08% carbon, 1.18% hydrogen and 0.78% nitrogen. The weight loss rate of thermogravimetric analysis is 7.95%.

[0077]

[0078] Formula I-e Example 6 A method based on ( S )-BINAM β - A method for preparing a chiral stationary phase of a cyclodextrin derivative, the steps of which are substantially the same as those of Example 1, except that the reagent reacting with AM-1 is 4-trifluoromethylphenylisocyanate.

[0079] The NMR and mass spectrometry data of AM-4f are: 1 H NMR (600 MHz, Chloroform- d ) d 8.55 (d, J=9.0 Hz, 1H), 8.49 (d, J = 8.9 Hz, 1H), 8.37 (t, J = 10.6 Hz, 2H), 8.29 (m, 2H),7.92 (m, 1H), 7.83 (q, J = 7.0 Hz, 2H), 7.70 (d, J = 7.9 Hz, 3H), 7.63 (q, J = 7.0Hz, 2H), 7.43 (m, 5H), 7.35 (m, 1H), 7.26 (m, 4H). 13 C NMR (150 MHz, CDCl3) d .98, 124.56, 124.54, 124.22, 124.12, 124.06, 124.00,123.91, 122.11, 121.47, 121.43, 120.73, 120.55, 120.07, 118.23. MS (ESI) calculated for [M+H] + 648. The BINAM- β The structure of -cyclodextrin CSP is shown in formula Ⅰ-f. Elemental analysis shows that it contains 2.61% carbon, 1.87% hydrogen and 0.71% nitrogen. The weight loss rate of thermogravimetric analysis is 5.58%.

[0080]

[0081] Formula I-f Example 7 A method based on ( S )-BINAM β - A method for preparing a chiral stationary phase of a cyclodextrin derivative, the steps of which are substantially the same as those of Example 1, except that the reagent reacting with AM-1 is 4-(dimethylamino)phenyl isocyanate.

[0082] The NMR and mass spectrometry data of AM-4g are: 1H NMR (600 MHz, Chloroform- d ) δ 8.34 (d, J = 9.1 Hz, 1H), 8.20 (d, J = 4.6 Hz, 1H), 7.95 (d, J = 8.9 Hz, 1H), 7.90 (t, J =9.0 Hz, 2H), 7.84 (d, J = 8.2 Hz, 1H), 7.66 (m, 1H), 7.53 (m, 1H), 7.40 (m,2H), 7.26 (m, 1H), 7.25 (m, 1H), 7.15 (d, J = 8.3 Hz, 1H), 7.09 (m, 3H), 7.03(d, J = 8.5 Hz, 1H), 6.97 (d, J = 8.6 Hz, 2H), 6.86 (d, J = 8.8 Hz, 2H), 6.81 (m,1H), 6.39 (m, 2H), 2.93 (s, 6H). 13 C NMR (150 MHz, CDCl3) d .84, 137.63, 135.95, 135.07, 134.26, 132.78, 132.55, 131.72, 130.34, 129.79,129.77, 128.60, 128.40, 128.32, 127.29, 127.22, 126.40, 126.25, 126.22,125.88, 125.58, 125.34, 124.86, 124.81, 123.38, 123.25, 120.36, 120.22,119.43, 118.30, 110.97, 39.98. MS(ESI)calculated for [M+H] + 623. The BINAM- β The structure of -cyclodextrin CSP is shown in formula Ⅰ-g. Elemental analysis shows that it contains 3.88% carbon, 1.94% hydrogen and 0.85% nitrogen. The weight loss rate of thermogravimetric analysis is 7.76%.

[0083]

[0084] Formula I-g Example 8 A method based on ( S )-BINAM β - A method for preparing a chiral stationary phase of a cyclodextrin derivative, the steps of which are substantially the same as those of Example 1, except that the reagent reacting with AM-1 is 1-(tert-butyl)-4-isocyanate benzene.

[0085] The NMR and mass spectrometry data of AM-4h are: 1 H NMR (600 MHz, Chloroform- d ) δ 8.15 (d, J = 9.1 Hz, 1H), 8.11 (d, J = 8.9 Hz, 1H), 7.80 (m, 4H), 7.64 (m, 2H), 7.58 (s,1H), 7.33 (t, J = 5.6 Hz, 1H), 7.28 (t, J = 7.4 Hz, 1H), 7.15 (t, J = 7.4 Hz, 2H),7.09 (m, 3H), 7.02 (s, 1H), 6.95 (m, 3H), 6.74 (d, J = 8.8 Hz, 2H), 6.71 (d, J =8.9 Hz, 2H), 1.14 (s, 9H). 13 C NMR (150 MHz, CDCl3) d .75, 126.09,125.94, 125.65, 125.43, 125.40, 125.33, 125.01, 124.53, 123.81, 122.75,121.06, 120.36, 119.08, 34.87, 30.96. MS (ESI) calculated for [M+H] + 636. The BINAM- βThe structure of -cyclodextrin CSP is shown in formula Ⅰ-h. Elemental analysis shows that it contains 3.43% carbon, 1.11% hydrogen and 0.62% nitrogen. The weight loss rate of thermogravimetric analysis is 6.36%.

[0086]

[0087] Formula I-h Example 9 A method based on ( S )-BINAM β - A method for preparing a chiral stationary phase of a cyclodextrin derivative, the steps of which are substantially the same as those of Example 1, except that the reagent reacting with AM-1 is 3,5-dimethylphenylisocyanate.

[0088] The NMR and mass spectrometry data of AM-4i are: 1 H NMR (600 MHz, Chloroform- d ) d 8.27 (d, J =9.0 Hz, 1H), 8.01 (d, J = 8.9 Hz, 1H), 7.92 (d, J = 8.9 Hz, 1H), 7.88 (d, J = 9.0Hz, 1H), 7.83 (s, 1H), 7.80 (d, J = 8.2 Hz, 1H), 7.55 (s, 1H), 7.38 (t, J = 7.9Hz, 1H), 7.32 (t, J = 8.0 Hz, 1H), 7.17 (t, J = 8.3 Hz, 1H), 7.11 (t, J = 7.7 Hz,1H), 6.98 (d, J = 8.4 Hz, 2H), 6.92 (d, J = 8.6 Hz, 2H), 6.89 (d, J = 8.6 Hz, 1H),6.83 (d, J = 8.8 Hz, 3H), 6.69 (m, 1H), 6.58 (m, 1H), 6.42 (m, 2H), 2.02 (s, 6H). 13 C NMR (150 MHz, CDCl3) d7, 137.29, 136.59,135.37, 135.01, 134.37, 132.81, 132.70, 131.22, 130.53, 130.50, 129.52,128.32, 128.19, 128.16, 127.14, 127.04, 126.99, 126.54, 126.34, 126.07,125.73, 125.59, 125.49, 125.35, 125.02, 124.97, 124.88, 123.36, 122.84,121.37, 120.44, 120.04, 119.43, 119.34, 21.08. MS (ESI) calculated for [M+H] + 608. The BINAM- β -The structure of cyclodextrin CSP is shown in formula Ⅰ-i. Elemental analysis shows that its carbon content is 2.48%, hydrogen content is 2.93%, and nitrogen content is 0.78%. The weight loss rate of thermogravimetric analysis is 6.81%.

[0089]

[0090] Formula I-i Example 10 A method based on ( S )-BINAM β - A method for preparing a chiral stationary phase of a cyclodextrin derivative, the steps of which are substantially the same as those of Example 1, except that the reagent reacting with AM-1 is 3,5-dichlorophenyl isocyanate.

[0091] The NMR and mass spectrometry data of AM-4g are: 1 H NMR (600 MHz, Chloroform- d ) δ 8.09 (d, J = 9.0 Hz, 1H), 8.04 (d, J = 9.0 Hz, 1H), 7.95 (d, J = 9.0 Hz, 1H), 7.85 (t, J =8.2 Hz, 2H), 7.76 (d, J = 8.2 Hz, 1H), 7.46 (d, J = 11.8 Hz, 2H), 7.38 (t, J= 7.5Hz, 1H), 7.33 (t, J = 7.5 Hz, 1H), 7.14 (t, J = 7.7 Hz, 1H), 7.10 (t, J = 8.0 Hz,1H), 7.01 (m, 2H), 6.88 (m, 9H). 13 C NMR (150 MHz, CDCl3) d .73, 122.48, 122.17, 121.80,122.69, 117.30. MS (ESI) calculated for [M+H]+ 648. The BINAM- β The structure of -cyclodextrin CSP is shown in formula Ⅰ-j. Elemental analysis shows that it contains 3.37% carbon, 1.15% hydrogen and 0.33% nitrogen. The weight loss rate of thermogravimetric analysis is 6.23%.

[0092]

[0093] Formula I-j Application Example 1: Based on BINAM β -Application of Cyclodextrin CSP in High Performance Liquid Chromatography The chiral stationary phase described in Formula I of the present invention has good stability and wide chiral selectivity, and can be used to separate a series of enantiomeric compounds in high performance liquid chromatography. For the convenience of description, the high performance liquid chromatograph used in this embodiment is Agilent 1260, but this does not limit the present invention.

[0094] The chiral stationary phase described in Example 1 was used to prepare a chiral chromatographic column, which can separate most of the β Receptor blockers (Table 1), imidazole antibiotics, nimodipine, aminoglutethimide, equol, ketoprofen, thalidomide and other chiral drugs (Table 1) and most triazole pesticides (Table 2) can be separated in reverse phase flow.

[0095] Chromatographic conditions: flow rate 0.8 mL / min, room temperature.

[0096] in k 1 is the retention factor of the first eluting enantiomer, which is given by the formula k 1 = (t1-t0) / t0, where t1 and t2 are the elution times of the first and second peaks, respectively, and t0 is the dead time, which is 2.34 min here; α is the selection factor, and the calculation formula is α = k 2 / k 1 R s is the separation degree, and the calculation formula is R s = 2(t2-t1) / (W1+W2), where W1 and W2 are the peak widths of the first and second eluting enantiomers, respectively.

[0097] Table 1 BINAM- synthesized in Example 1 β - Results of separation of chiral drugs by cyclodextrin CSP

[0098] a. Mobile phase: ACN / 0.3%FA (15 / 85) b. Mobile phase: ACN / 0.1%FA (10 / 90) c. Mobile phase: ACN / MeOH / TEA / AcOH (95 / 5 / 1.2 / 0.8, v / v / v / v) d. Mobile phase: MeOH / 0.1% FA (40 / 60) e. Mobile phase: ACN / H2O (10 / 90) f. Mobile phase: MeOH / H2O (30 / 70) g. Mobile phase: MeOH / H2O (50 / 50) Table 2 BINAM- synthesized in Example 1 β - Results of separation of chiral pesticides by cyclodextrin CSP

[0099] a Mobile phase: MeOH / H2O (35 / 65) b Mobile phase: MeOH / H2O (45 / 55) c Mobile phase: MeOH / H2O (50 / 50) d Mobile phase: ACN / 0.1% FA (10 / 90) In summary, the present invention provides a novel β -Cyclodextrin CSP filler can separate chiral compounds of various structural types, is suitable for different mobile phase systems of high performance liquid chromatography (HPLC), and has good stability in mobile phases with strong polarity and high water content. The CSP filler provided by the present invention is capable of separating imidazole antibiotics, β Receptor blockers and other chiral drugs and triazole pesticides. It can meet the needs of daily analysis, production process and quality control during the use of these chiral drugs and chiral pesticides.

[0100] It will be apparent to those skilled in the art that the present invention is not limited to the details of the exemplary embodiments described above, and that the present invention can be implemented in other specific forms without departing from the spirit or essential features of the present invention. Therefore, the embodiments should be considered exemplary and non-restrictive in all respects, and the scope of the present invention is defined by the appended claims rather than the above description, and it is intended that all changes falling within the meaning and scope of the equivalent elements of the claims be included in the present invention.

[0101] In addition, it should be understood that although the present specification is described according to implementation modes, not every implementation mode contains only one independent technical solution. This description of the specification is only for the sake of clarity. Those skilled in the art should regard the specification as a whole. The technical solutions in each embodiment may also be appropriately combined to form other implementation modes that can be understood by those skilled in the art.

Claims

1. A BINAM-based β -Cyclodextrin derivative chiral stationary phase, characterized in that β -A BINAM derivative is connected to the 6-position of cyclodextrin and bonded to the silica surface via carbamate. The structure is shown in Formula I: Formula I R is selected from: The configuration of BINAM is R -or S -Configuration.

2. The BINAM-based method of claim 1 β - A method for preparing a cyclodextrin derivative chiral stationary phase, characterized in that: The steps include: S1. β -cyclodextrin reacts with p-toluenesulfonyl chloride to obtain mono-6-p-toluenesulfonyl- β -Cyclodextrin TsO- β CD; Formula II S2. Take the TsO- β CD is added to ammonia water, and the mixture is stirred at 40-70°C for 10-24 hours to obtain 6-amino-6-deoxy- β -Cyclodextrin NH2- β CD; Formula III S3. BINAM was dissolved in anhydrous dichloromethane, p-nitrobenzene isocyanate was added to react, and the compound AM-1 as shown in Formula Ⅳ was obtained by purification by silica gel column chromatography; Formula IV S4. Compound AM-1 is dissolved in anhydrous dichloromethane, phenyl isocyanate containing different substituents is added to react, and purified by silica gel column chromatography to obtain compound AM-2 as shown in formula V; Formula V R is selected from: S5. Add iron, ethanol and hydrochloric acid to compound AM-2, heat the reaction for 5 to 12 hours, and purify by silica gel column chromatography to obtain compound AM-3 as shown in formula VI; Formula VI R is selected from: S6. AM-3 was dissolved in ethyl acetate or dichloromethane, thiophosgene and saturated aqueous NaHCO3 were added, and the mixture was purified by silica gel column chromatography to obtain compound AM-4 as shown in formula VII; Formula VII R is selected from: S7. Compound AM-4 of formula VII and compound NH2- β CD and a catalytic amount of DMAP are added to DMF and reacted for 15 to 30 hours to obtain the product shown in Formula VIII. β - Cyclodextrin-BINAM derivative AM-CD; Formula VIII R is selected from: S8. AM-CD and isocyanatepropyltriethoxysilane were added to DMF to react for a period of time, and activated silica gel was added to continue the reaction. After cooling to room temperature, the solid was filtered, and the obtained solid was washed repeatedly with DMF and methanol for several times, and then vacuum dried to obtain a BINAM-based compound as shown in Formula I. β -Cyclodextrin derivative CSP filler.

3. The method according to claim 2 based on BINAM β - A method for preparing a cyclodextrin derivative chiral stationary phase, characterized in that: The method of S1 specifically comprises the following steps: Will β -cyclodextrin is dissolved in an aqueous sodium hydroxide solution, and a solution of p-toluenesulfonyl chloride in acetonitrile is added to react at room temperature or under ice bath conditions. After the reaction is completed, impurities are filtered out, the filtrate is adjusted to pH with hydrochloric acid, and refrigerated overnight to precipitate. The solid is filtered and recrystallized several times with a small amount of hot water, filtered and dried to obtain a mono-6-p-toluenesulfonyl- β -Cyclodextrin TsO- β CD; β The molar mass ratio of -cyclodextrin and p-toluenesulfonyl chloride is 1:0.8~1.5; the concentration of the sodium hydroxide aqueous solution is 5~10 mol / L, and the amount of acetonitrile solution is 1 gram of p-toluenesulfonyl chloride dissolved in 2~5 mL of acetonitrile.

4. The method according to claim 2 based on BINAM β - A method for preparing a cyclodextrin derivative chiral stationary phase, characterized in that: The configuration of BINAM in S3 is R -or S -configuration, the molar mass ratio of BINAM to p-nitrobenzene isocyanate is 1:1~2; the reaction temperature is room temperature, and the reaction time is 2~10 h.

5. The BINAM-based method according to claim 2 β - A method for preparing a cyclodextrin derivative chiral stationary phase, characterized in that: The phenyl isocyanates containing different substituents in S4 include: p-nitrobenzene isocyanate, p-chlorophenyl isocyanate, p-toluene isocyanate, phenyl isocyanate, 4-methoxyphenyl isocyanate, 4-trifluoromethylphenyl isocyanate, 4-(dimethylamino)phenyl isocyanate, 1-(tert-butyl)-4-isocyanatebenzene, 3,5-dimethylphenyl isocyanate, and 3,5-dichlorophenyl isocyanate. The molar mass ratio of AM-1 used to the phenyl isocyanate containing different substituents is 1:1~2; the reaction temperature is room temperature, and the reaction time is 2~10 h.

6. The BINAM-based method according to claim 2 β - A method for preparing a cyclodextrin derivative chiral stationary phase, characterized in that: The molar mass ratio of AM-2 to iron in S5 is 1:5~20; the ratio of ethanol to hydrochloric acid is 4:3 (v / v); the reaction temperature is 60~80 ℃; and the reaction time is 10~24 h.

7. The BINAM-based method according to claim 2 β - A method for preparing a cyclodextrin derivative chiral stationary phase, characterized in that: The molar mass ratio of AM-3 to thiophosgene in S6 is 1:1~2; the amount of saturated NaHCO3 aqueous solution used is 10 mL per gram of AM-3; the reaction is carried out at room temperature or in an ice bath, and the reaction time is 0.5~2 h.

8. The BINAM-based method according to claim 2 β - A method for preparing a cyclodextrin derivative chiral stationary phase, characterized in that: NH2- in S7 β The molar mass ratio of CD, AM-4 and DMAP is 1:1~2:0.1; the reaction is carried out under nitrogen protection; the amount of solvent DMF is based on 1:1 per gram of NH2- β Add 10~20 mL of CD; the reaction time is 20~24 h.

9. The BINAM-based method according to claim 2 β - A method for preparing a cyclodextrin derivative chiral stationary phase, characterized in that: All reactions in S8 were carried out under nitrogen protection; the molar mass ratio of AM-CD to triethoxypropyl isocyanate was 1:2~5; the reaction time was 4~10 h; the reaction temperature after adding activated silica gel was 110~120°C, and the reaction time was 20~36 h.

10. The BINAM-based method of claim 1 β -Application of cyclodextrin derivative chiral stationary phase in separation of chiral compounds, characterized in that: The chiral compounds that can be separated include: imidazole antibiotics, β Beta-blockers, nimodipine, aminoglutethimide, equol and triazole chiral pesticides.

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