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High-efficiency liquid-phase chromatography detection method of L-arginine-alpha-ketoglutarate

A high-performance liquid chromatography and ketoglutaric acid technology, applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of poor precision, low accuracy, and decreased column efficiency of spectrophotometry, and achieve good precision and Accuracy, labor saving, quality controllable effect

Active Publication Date: 2015-04-01
JING JING PHARMA
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  • Abstract
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  • Claims
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Problems solved by technology

[0003] In the prior art, Zhongjing Hongwu et al. have disclosed the use of spectrophotometry to measure α-ketoglutarate (simultaneous determination of α-ketoglutarate and pyruvic acid by spectrophotometry, Journal of Tongji University, 1986, 14 (4): 531- 536), but the accuracy of spectrophotometry is poor, the accuracy is not high, and only the content of α-ketoglutaric acid can be determined
[0004] Mao Ying et al disclosed the determination of ketoglutaric acid content by high performance liquid chromatography (HPLC method for the determination of ketoglutaric acid content, China Pharmaceutical Affairs, 2008, 22 (10): 598-599), which used Agilent ODS C 18 column, with 0.5% (NH 4 ) 2 HPO 4 -H 3 PO 4 The buffer solution is the mobile phase, the flow rate is 1.0ml / min, and the ultraviolet detection wavelength is 233nm, but this method can only be used to determine the content of ketoglutarate, and cannot be used for L-arginine-α-ketoglutarate. Content detection
[0005] In the determination of arginine content under the category of ceftazidime for injection in the 2010 edition of the Chinese Pharmacopoeia, dihydroxypropylsilane-bonded silica gel was used as a filler, and acetonitrile-pH2.0 phosphate buffer (weighed ammonium dihydrogen phosphate 1.15 g, add 800ml of water to dissolve, adjust the pH value to 2.0 with phosphoric acid, then dilute to 1000ml with water, mix well) (750:250) as the mobile phase, the detection wavelength is 206nm, but this method can only measure L-arginine Content
When an amino column is used for the detection of acidic substances, the presence of acidic substances means the presence of protons, which will protonate the slightly negatively charged amino functional groups, resulting in some retention properties for certain types of analytes after a period of use. Changed or manifested in the decline of column efficiency, affecting the accuracy of content determination

Method used

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  • High-efficiency liquid-phase chromatography detection method of L-arginine-alpha-ketoglutarate
  • High-efficiency liquid-phase chromatography detection method of L-arginine-alpha-ketoglutarate
  • High-efficiency liquid-phase chromatography detection method of L-arginine-alpha-ketoglutarate

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Chromatographic conditions

[0039] Mobile phase: 0.05mol / L sodium dihydrogen phosphate buffer (use 0.05mol / L phosphoric acid to adjust the pH value to 3.4)

[0040] Mobile phase flow rate: 1.0ml / min

[0041] Detection wavelength: 205nm

[0042] Column temperature: 25°C

[0043] Experimental steps:

[0044] (1) Prepare mobile phase, adjust pH value, filter and degas;

[0045] (2) Adjust the conditions of the high-performance liquid chromatograph, and start to balance the chromatographic column;

[0046] (3) Configuration of reference substance and test solution:

[0047] Reference substance solution: Weigh 34.0 mg of L-arginine reference substance and 29.0 mg of α-ketoglutarate reference substance into a 100ml measuring bottle, add mobile phase to dilute to the mark, and shake well to obtain reference substance solution.

[0048]Test solution: Weigh 64 mg of L-arginine-α-ketoglutarate (the molar ratio of L-arginine to α-ketoglutarate is 1:1) and place it in a 100m...

Embodiment 2

[0054] Chromatographic conditions

[0055] Mobile phase: 0.05mol / L sodium dihydrogen phosphate buffer (use 0.05mol / L phosphoric acid to adjust the pH value to 3.4)

[0056] Mobile phase flow rate: 1.0ml / min

[0057] Detection wavelength: 205nm

[0058] Column temperature: 25°C

[0059] Experimental steps:

[0060] (1) Prepare mobile phase, adjust pH value, filter and degas;

[0061] (2) Adjust the conditions of the high-performance liquid chromatograph, and start to balance the chromatographic column;

[0062] (3) Configuration of reference substance and test solution:

[0063] Reference substance solution: Weigh 68.0 mg of L-arginine reference substance and 29.0 mg of α-ketoglutarate reference substance into a 100ml measuring bottle, add mobile phase to dilute to the mark, and shake well to obtain reference substance solution.

[0064] The test solution: Weigh 96 mg of L-arginine-α-ketoglutarate (the molar ratio of L-arginine to α-ketoglutarate is 2:1) and place it in a...

Embodiment 3

[0088] The difference with Example 1 is that the chromatographic conditions:

[0089] Mobile phase: 0.05mol / L potassium dihydrogen phosphate buffer (use 0.05mol / L phosphoric acid to adjust the pH value to 3.4);

[0090] Mobile phase flow rate: 0.5ml / min;

[0091] Detection wavelength: 200nm;

[0092] Column temperature: 15°C.

[0093] Configuration of reference substance and test solution:

[0094] Reference substance solution: Weigh 16 mg of L-arginine reference substance and 14 mg of α-ketoglutarate reference substance in a 100ml measuring bottle, add mobile phase to dilute to the mark, and shake well to obtain reference substance solution.

[0095] For the test solution, weigh 30 mg of L-arginine-α-ketoglutarate (the molar ratio of L-arginine to α-ketoglutarate is 1:1) and place it in a 100ml measuring bottle, add flow Phase dissolved and diluted to the mark, shake well, that is.

[0096] Perform HPLC analysis to determine the content of L-arginine and α-ketoglutarate ...

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Abstract

The invention discloses a high-efficiency liquid-phase chromatography detection method of L-arginine-alpha-ketoglutarate, and relates to the technical field of column chromatography. According to chromatographic condition, a chromatographic column adopts a C18 chromatographic column; a mobile phase adopts a 0.02mol / L to 0.08mol / L phosphate buffer solution with pH value of 3.0 to 4.0; the flow rate of the mobile phase is 0.5 to 1.3 ml / min; the detection wavelength is 200nm to 210nm; the temperature of the chromatographic column is 15 to 30 DEG C, and the content of L-arginine and the content of alpha-ketoglutarate in L-arginine-alpha-ketoglutarate or preparations of the L-arginine-alpha-ketoglutarate are determined by carrying out the high-efficiency liquid-phase chromatographic analysis. The content of L-arginine and the content of alpha-ketoglutarate in L-arginine-alpha-ketoglutarate or preparations of the L-arginine-alpha-ketoglutarate can be accurately determined by virtue of one-step high-efficiency liquid-phase chromatographic detection, and the two components can be well separated; moreover, the balance time of the C18 chromatographic column is short and is about 30 to 40 minutes, and the working efficiency can be improved.

Description

technical field [0001] The invention relates to the technical field of column chromatography. Background technique [0002] L-arginine-α-ketoglutaric acid is an amino acid salt health product with increasing sales volume in the international market. It is mainly used to improve human protein and enhance human physique. There are two specifications of 1:1 and 2:1 (the molar ratio of arginine and ketoglutaric acid). The liver-protecting drugs developed by pharmaceutical companies can directly and quickly provide nutrition for liver dysfunction, provide energy for liver cells, and quickly maintain the normal function and health of the liver. [0003] In the prior art, Zhongjing Hongwu et al. have disclosed the use of spectrophotometry to measure α-ketoglutarate (simultaneous determination of α-ketoglutarate and pyruvic acid by spectrophotometry, Journal of Tongji University, 1986, 14 (4): 531- 536), but the spectrophotometric method has poor precision and low accuracy, and ca...

Claims

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Application Information

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IPC IPC(8): G01N30/02G01N30/06
Inventor 李斌水何瑞红郭志建张柳
Owner JING JING PHARMA
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