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Separation and preparation method of a tetrapeptide impurity in alanyl glutamine injection

A technology of alanyl glutamine and injection, applied in separation methods, preparation of test samples, separation of solid adsorbents and liquids, etc.

Active Publication Date: 2019-01-01
HANGZHOU MINSHENG PHARM CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the particularity of the molecular structure of alanylglutamine, it is easy to introduce by-products with similar structures during high-temperature sterilization. Due to the similarity of their chemical structures, traditional separation methods such as extraction, recrystallization, distillation and TLC are difficult separate it
Analytical HPLC can only obtain microgram-level products. In order to obtain high-quality alanyl glutamine injection, it is necessary to extract its main impurities and confirm its structure

Method used

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  • Separation and preparation method of a tetrapeptide impurity in alanyl glutamine injection
  • Separation and preparation method of a tetrapeptide impurity in alanyl glutamine injection
  • Separation and preparation method of a tetrapeptide impurity in alanyl glutamine injection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Example 1. Pre-separation of a tetrapeptide trace impurity in alanyl glutamine injection:

[0041] Dissolve 400.0 g of the lyophilized powder of alanyl glutamine injection in 500 ml of distilled water, filter (through a filter membrane with a pore size of 0.45 μm), and collect the filtrate for later use.

[0042] The preparative chromatographic separation conditions are:

[0043] High performance liquid chromatograph model: Hanbang NP7010

[0044] Chromatographic column: 50×250mm, filled with aminopropyl bonded silica gel as the stationary phase filler, the particle size of the filler is 20μm, and the pore size is

[0045] Flow rate: 50ml / min.

[0046] Detection wavelength: 215nm.

[0047] Mobile phase: acetonitrile-ammonium formate buffer (40:60 by volume, wherein the ammonium formate concentration in the ammonium formate buffer is 20 mmol / l, and adjusted to pH 4.0 with formic acid).

[0048] Elution method: isocratic elution.

[0049] Loading amount: 40.0g (50m...

Embodiment 2

[0053] Example 2, Purification and separation of tetrapeptide impurities in alanyl glutamine injection

[0054] Dissolve 4.0 g of the crude tetrapeptide impurity product in Example 1 with 50 ml of distilled water, filter (through a filter membrane with a pore size of 0.45 μm), and collect the filtrate for future use.

[0055] Preparative chromatographic refining separation conditions:

[0056] High performance liquid chromatograph model: Waters 600

[0057] Chromatographic column: 20×250mm, filled with aminopropyl bonded silica gel as the stationary phase filler, the particle size of the filler is 5 μm, and the pore size is

[0058] Flow rate: 10ml / min.

[0059] Detection wavelength: 215nm.

[0060] Mobile phase: acetonitrile-formic acid buffer (50:50 by volume, wherein the formic acid concentration of the formic acid buffer is 0.1%).

[0061] Elution method: isocratic elution.

[0062] Loading amount: 400mg (5ml).

[0063] Purification process: equilibrate the chromat...

Embodiment 3

[0069] Example 3, Structural Confirmation of Tetrapeptide Impurities in Alanyl Glutamine Injection

[0070] High Resolution Mass Spectrometry (HR-MS):

[0071] Mass spectrometer model: Agilent 6210TOF-MS.

[0072] Determination mode: electrospray positive ion mode.

[0073] The high-resolution electrospray mass spectrometry results of the target impurity (that is, the white powdery solid tetrapeptide impurity obtained in Example 2) in positive ion mode are shown in image 3 , which produces protonated ions [M+H] + m / z 418.1944, the corresponding element composition is C 16 h 28 N 5 o 8 (418.1932), error is -2.78ppm, resulting in potassium adduct ion [M+K] + m / z 456.1498, the corresponding element composition is C 16 h 27 KN 5 o 8 (456.1491), the error is -1.63ppm, according to the above data, it can be confirmed that the molecular formula of the target impurity is C 16 h 27 N 5 o 8 , and the data results are shown in Table 2.

[0074] Table 2. High-resolution m...

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Abstract

The invention discloses a separation preparation method of tetrapeptide impurity in an alanyl-glutamine injection. The separation preparation method includes following steps: 1), adopting a preparation-type high performance liquid chromatography method to pre-separate the tetrapeptide impurity in the alanyl-glutamine injection, wherein the high performance liquid chromatography method includes using aminopropyl bonded silica gel as a stationary phase and acetonitrile-ammonium formate buffering liquid as a moving phase to perform isocratic elution, and a molecular formula of the tetrapeptide trace impurity is C16H27N5O8; 2), adopting the preparation-type high performance liquid chromatography method to refine and purify a tetrapeptide impurity crude product, wherein the high performance liquid chromatography method includes using aminopropyl bonded silica gel as a stationary phase and acetonitrile-ammonium formate buffering liquid as a moving phase to perform isocratic elution; collecting and combining target distillate, and lyophilizing to obtain a purified tetrapeptide impurity.

Description

technical field [0001] The invention belongs to the technical field of drug impurity research, and in particular relates to a method for separating and preparing a tetrapeptide trace impurity in alanyl glutamine injection. Background technique [0002] Chemical drugs will produce some adverse reactions in clinical use, which are not only related to the pharmacological activity of the drug itself, but also have a lot to do with the impurities in the drug. Therefore, for chemical drugs, each national pharmacopoeia has a special general rule for impurity testing in the appendix. The second part of the Chinese Pharmacopoeia has set up guidelines for drug impurity research in the appendix since (2005). The State Food and Drug Administration has also issued the "Chemical Technical guidelines for drug impurity studies. For the impurities in the drug, the quality control limit must be established according to its physiological activity, including: the impurities in the drug are eff...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N1/28B01D15/10
CPCB01D15/10G01N1/28G01N2001/2893
Inventor 柯云翠朱义周娜
Owner HANGZHOU MINSHENG PHARM CO LTD