阅读说明：本技术 一种雷美替胺对映异构体检测方法及其质量控制标准 (Ramelteon enantiomer detection method and quality control standard thereof ) 是由 陆华龙 于 2018-06-28 设计创作，主要内容包括：本发明公开了一种雷美替胺对映异构体检测方法及其质量控制标准。本发明采用高效液相色谱法,通过使用正相色谱柱,有机溶剂试剂作为流动相,根据雷美替胺消旋体对雷美替胺中的对映异构体进行定位,能够对存在于各类剂型中的雷美替胺进行其对映异构体检查,提高现有的雷美替胺的质量标准,为今后制定雷美替胺的对映异构体测定标准提供依据。该检测方法具有较高的准确度和精密度,耐用性强,适合工业上用于雷美替胺对映异构体的检查及质量控制。(The invention discloses a ramelteon enantiomer detection method and a quality control standard thereof. The invention adopts the high performance liquid chromatography, uses the normal phase chromatographic column and the organic solvent reagent as the mobile phase, positions the enantiomer in the ramelteon according to the ramelteon racemate, can check the enantiomer of the ramelteon existing in various dosage forms, improves the quality standard of the existing ramelteon, and provides a basis for the determination standard of the enantiomer of the ramelteon in the future. The detection method has high accuracy and precision and strong durability, and is suitable for the detection and quality control of ramelteon enantiomer in industry.)

1. A ramelteon enantiomer detection method is characterized by comprising the following steps:

1) Preparation of system suitability test solution: dissolving and diluting the ramelteon racemate reference substance with a mobile phase to prepare a solution of 0.1-0.5 mg/ml serving as a system applicability test solution;

2) Preparation of a test solution: dissolving and diluting ramelteon with a mobile phase to prepare a ramelteon solution of 0.2-1.0 mg/ml as a test solution;

3) Preparation of control solution: precisely measuring a proper amount of a test solution, adding a mobile phase to dilute and prepare a ramelteon solution of 0.01-0.1 mg/ml as a reference solution;

4) Chromatographic conditions and system applicability test: adopting a chiral column CHIRALPAK AD-H with a diameter of 4.6 multiplied by 250mm, and using n-hexane-ethanol as a mobile phase, wherein the volume ratio of the n-hexane to the ethanol in the mobile phase is 80-95/20-5, the flow rate is 0.6-1.4 ml/min, the detection wavelength is 200-220 nm, and the column temperature is 20-35 ℃;

5) Ramelteon enantiomer determination: respectively and precisely measuring 2-50 mul of the system applicability test solution, the test sample solution and the control solution, injecting into a liquid chromatograph, recording a chromatogram, and calculating the content of the enantiomer by peak area according to an external standard method.

2. The method for detecting ramelteon enantiomer of claim 1, which comprises the following steps:

1) Preparation of system suitability test solution: taking 50mg of ramelteon racemate reference substance, precisely weighing, placing into a 100ml volumetric flask, adding a mobile phase for dissolving and diluting to a scale, and taking the reference substance as a system applicability test solution;

2) Preparation of a test solution: taking 50mg of ramelteon, precisely weighing, placing in a 100ml volumetric flask, adding a mobile phase for dissolving and diluting to a scale, and taking the solution as a test solution;

3) preparation of control solution: precisely measuring 1ml of a test solution, placing the test solution in a 100ml volumetric flask, adding a mobile phase for dissolving and diluting to a scale, and taking the scale as a reference solution;

4) the chromatographic condition and system applicability test adopts a chiral column CHIRALPAK AD-H of 4.6 multiplied by 250mm, n-hexane-ethanol is used as a mobile phase, wherein the volume ratio of the n-hexane to the ethanol in the mobile phase is 90/10, the flow rate is 1.0ml/min, the detection wavelength is 210nm, and the column temperature is 25 ℃; the theoretical plate number is not lower than 2500 calculated according to the ramelteon peak, and the separation degree of the ramelteon peak and the enantiomer peak thereof in the system adaptability test conforms to the specification;

5) ramelteon enantiomer determination: injecting 20 mul of system applicability solution into a liquid chromatograph, wherein the separation degree of the ramelteon and the ramelteon enantiomer is in accordance with the requirement; respectively injecting 20 μ l of the enantiomer control solution and the sample solution into a liquid chromatograph, and recording chromatogram; if the enantiomer peak is detected in the test solution spectrum, the content of the enantiomer is calculated by the peak area according to an external standard method.

Technical Field

The invention relates to a ramelteon enantiomer detection method and a quality control standard thereof, belonging to the technical field of pharmaceutical analysis.

Technical Field

Ramelteon is a melatonin receptor agonist, has high affinity for melatonin MT1 and MT2 receptors, and is more selective than the MT3 receptor. Ramelteon shows full agonist activity in cells expressing human MT1 or MT2 receptor in vitro. The activity of ramelteon at the MT1 and MT2 receptors makes it sleep-promoting, since these receptors, through the action of endogenous melatonin, are thought to be involved in the maintenance of the underlying normal wake-sleep cycle of the circadian rhythm. Ramelteon has no appreciable affinity for GABA receptor complexes or for binding neuropeptides, cytokines, 5-hydroxytryptamines, dopamine, norepinephrine, acetylcholine and opioid receptors. Ramelteon also did not interfere with many of the selected enzyme activities in one standard plate screen.

The main metabolite M-II of ramelteon has activity, the affinity to human MT1 and MT2 receptors is one fifth and one tenth of the parent molecule respectively, and the activity is 17-25 times lower than that of ramelteon in an in vitro functional test. Although M-II is less active than the parent drug at the MT1 and MT2 receptors, M-II circulates at higher concentrations than ramelteon, 20-100 times higher than the average systemic exposure produced by the parent drug. M-II has weak affinity for the 5-hydroxytryptamine 5-HT2B receptor, but no appreciable affinity for other receptors or enzymes. Similar to ramelteon, M-II does not interfere with the activity of many endogenous enzymes. All other known metabolites of ramelteon are inactive. Because the lorcaserin is a single enantiomer, the control of the content of impurities in the raw material medicine, particularly the content of the enantiomer, has important significance for reducing the adverse reaction. Based on the detection result, the invention provides a ramelteon enantiomer detection method and a quality control standard thereof.

Wherein the structural formula of the ramelteon is as follows:

disclosure of Invention

In order to accurately reflect the content of the enantiomer in the ramelteon raw material medicine and provide reasonable basis for the formulation of quality standard so as to better control and master the product quality and improve the safety of clinical medication, the invention provides a ramelteon enantiomer detection method and a quality control standard thereof. The detection method has high accuracy and precision and strong durability, and is suitable for the detection and quality control of ramelteon enantiomer in industry.

The invention realizes the purpose through the following technical scheme:

ramelteon is a melatonin receptor agonist with the chemical name (S) -N- [2- (1,6,7, 8-tetrahydro-2H-indeno [5,4-b ]]Furan-8-yl) ethyl]Propionamide. The ramelteon enantiomer of the salt of the invention is in mirror symmetry with ramelteon, and the chemical formula of the ramelteon enantiomer is C₁₆H₂₁NO₂ The chemical name of the compound is (R) -N- [2- (1,6,7, 8-tetrahydro-2H-indeno [5, 4-b)]Furan-8-yl) ethyl]Propionamide. The invention provides a ramelteon enantiomer detection method, which comprises the following steps:

1) Preparation of system suitability test solution: dissolving and diluting the ramelteon racemate reference substance with a mobile phase to prepare a solution of 0.1-0.5 mg/ml serving as a system applicability test solution;

2) Preparation of a test solution: dissolving and diluting ramelteon with a mobile phase to prepare a ramelteon solution of 0.2-1.0 mg/ml as a test solution;

3) Preparation of control solution: precisely measuring a proper amount of a test solution, adding a mobile phase to dilute and prepare a ramelteon solution of 0.01-0.1 mg/ml as a reference solution;

4) Chromatographic conditions and system applicability test: adopting a chiral column CHIRALPAK AD-H with a diameter of 4.6 multiplied by 250mm, and using n-hexane-ethanol as a mobile phase, wherein the volume ratio of the n-hexane to the ethanol in the mobile phase is 80-95/20-5, the flow rate is 0.6-1.4 ml/min, the detection wavelength is 200-220 nm, and the column temperature is 20-35 ℃;

5) Ramelteon enantiomer determination: respectively and precisely measuring 2-50 mul of the system applicability test solution, the test sample solution and the control solution, injecting into a liquid chromatograph, recording a chromatogram, and calculating the content of the enantiomer by peak area according to an external standard method.

The detection method of ramelteon enantiomer specifically comprises the following steps:

1) Preparation of system suitability test solution: taking 50mg of ramelteon racemate reference substance, precisely weighing, placing into a 100ml volumetric flask, adding a mobile phase for dissolving and diluting to a scale, and taking the reference substance as a system applicability test solution;

2) Preparation of a test solution: taking 50mg of ramelteon, precisely weighing, placing in a 100ml volumetric flask, adding a mobile phase for dissolving and diluting to a scale, and taking the solution as a test solution;

3) Preparation of control solution: precisely measuring 1ml of a test solution, placing the test solution in a 100ml volumetric flask, adding a mobile phase for dissolving and diluting to a scale, and taking the scale as a reference solution;

4) The chromatographic condition and system applicability test adopts a chiral column CHIRALPAK AD-H of 4.6 multiplied by 250mm, n-hexane-ethanol is used as a mobile phase, wherein the volume ratio of the n-hexane to the ethanol in the mobile phase is 90/10, the flow rate is 1.0ml/min, the detection wavelength is 210nm, and the column temperature is 25 ℃; the theoretical plate number is not lower than 2500 calculated according to the ramelteon peak, and the separation degree of the ramelteon peak and the enantiomer peak thereof in the system adaptability test conforms to the specification;

5) Ramelteon enantiomer determination: injecting 20 mul of system applicability solution into a liquid chromatograph, wherein the separation degree of the ramelteon and the ramelteon enantiomer is in accordance with the requirement; respectively injecting 20 μ l of the enantiomer control solution and the sample solution into a liquid chromatograph, and recording chromatogram; if the enantiomer peak is detected in the test solution spectrum, the content of the enantiomer is calculated by the peak area according to an external standard method.

The invention also provides a quality control standard of ramelteon enantiomer, wherein the content of ramelteon enantiomer is measured by the detection method. When the detection method is adopted for determination, the content of the ramelteon enantiomer cannot exceed 0.5 percent. In order to control the product quality more strictly and ensure the medication safety, the standard of enantiomer inspection is improved to be not more than 0.2 percent.

Experimental example 1: mobile phase screening in assay methods

The present inventors screened the mobile phase for ramelteon enantiomer examination. The test results are shown in Table 1.

Table 1 ramelteon enantiomer check chromatographic condition selection

Experimental results show that when the mobile phase is n-hexane-ethanol (90: 10, V/V), the obtained HPLC chromatogram has good main peak pattern and proper retention time, and the separation of the main peak and the impurity peak achieves better effect. N-hexane-ethanol (90: 10, V/V) was therefore determined as the final mobile phase condition for the ramelteon isomer impurity check.

Experimental example 2: detection wavelength selection

Taking a proper amount of ramelteon and a proper amount of ramelteon racemate reference substances, respectively dissolving and diluting the reference substances by using a mobile phase to obtain solutions with the concentration of about 5 mu g/ml, and respectively scanning within the wavelength range of 200-400 nm by using an ultraviolet-visible spectrophotometry (0401 in the four-part general rule of Chinese pharmacopoeia 2015). Scanning results show that ramelteon has maximum absorption at the wavelength of 210nm, so that 210nm is selected as the detection wavelength for isomer detection.

Drawings

Figure 1 is a chromatogram of the separation of ramelteon from its enantiomers wherein an enantiomer of ramelteon is present with a tR of 15.345 and ramelteon is present with a tR of 10.925. The detection wavelength of channel 1 of detector A is 210 nm.

Detailed Description

The present invention is further described by the following specific examples, but those skilled in the art should understand that the examples are not intended to limit the present invention in any way, and the technical scope of the present invention is subject to the claims.