HPLC (high-performance liquid chromatography) detection method capable of simultaneously detecting contents of six amino acids in donkey-hide glue blood-supplementing preparation

A technology for amino acids and preparations, applied in the field of HPLC determination of six amino acids in donkey glue blood-enriching preparations, which can solve problems such as complex processing methods, and achieve simple sample processing methods, low requirements for operating proficiency, and good reproducibility sexual effect

Active Publication Date: 2013-08-28
JIUZHITANG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Although the 2010 edition of the Pharmacopoeia included the determination of the amino acid content of donkey-hide gelatin raw materials, it was limited to a single medicinal material, and the sample processing method was complicated. Only L-hydroxyproline, glycine, alanine, and L-proline were determined. The content of 4 kinds of amino acids

Method used

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  • HPLC (high-performance liquid chromatography) detection method capable of simultaneously detecting contents of six amino acids in donkey-hide glue blood-supplementing preparation
  • HPLC (high-performance liquid chromatography) detection method capable of simultaneously detecting contents of six amino acids in donkey-hide glue blood-supplementing preparation
  • HPLC (high-performance liquid chromatography) detection method capable of simultaneously detecting contents of six amino acids in donkey-hide glue blood-supplementing preparation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Example 1 Preparation of 6 Amino Acid Mixed Standards

[0036]Take appropriate amount of L-glutamic acid reference substance, L-hydroxyproline reference substance, glycine reference substance, L-alanine reference substance, L-arginine reference substance, L-proline reference substance, and accurately weigh , add 0.1mol / L hydrochloric acid solution to make each 1ml contain L-glutamic acid 73.1μg, L-hydroxyproline 86.4μg, glycine 0.1609mg, L-alanine 64.8μg, L-arginine 59.3μg , a mixed solution of 93.9 μg of L-proline to obtain mixed standard substances of 6 kinds of amino acids.

Embodiment 2

[0037] Example 2 Preparation of test solution

[0038] Method 1. Grind donkey glue blood-enriching granules to be tested into powder, take 0.25g, put it in an ampoule bottle, weigh it accurately, add 5ml of 5mol / l hydrochloric acid, put it on an alcohol blowtorch for drawing and sealing, and ultrasonically dissolve it for 30 minutes, 130 ℃ hydrolysis for 1 hour. Let cool, transfer to an evaporating dish, wash with 10ml of water several times, put the washing liquid into an evaporating dish, and evaporate to dryness in a water bath. Add 0.1mol / L hydrochloric acid solution to dissolve the residue, transfer it to a 25ml measuring bottle, add 0.05mol / L hydrochloric acid solution to dilute to the mark, shake well, centrifuge at 3000r / min for 5 minutes and take the supernatant to obtain the solution to be tested.

[0039] Method 2. After grinding the donkey glue blood-enriching granules to be tested into powder, take 0.25g and put it in an ampoule bottle, weigh it accurately, add...

Embodiment 3

[0041] Example 3 derivatization

[0042] Take 2ml each of the above-mentioned standard solution and the solution to be tested, put them in 10ml centrifuge tubes respectively, add 0.1mol / L phenylisothiocyanate (PITC)-acetonitrile solution (take 120μl phenylisothiocyanate in a 10ml volumetric flask 1.0ml, 1mol / L triethylamine-acetonitrile solution (take 1.4ml of triethylamine in a 10ml volumetric flask, dilute with acetonitrile and add to the mark, shake 1.0ml, vortexed for 1min, left at room temperature for 1 hour, added 4ml of n-hexane, shaken, vortexed for 1min, left for 10 minutes, removed the lower layer solution, filtered through a microporous membrane to obtain the derivatized standard solution and derivatized test solution.

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Abstract

The invention relates to a HPLC (high-performance liquid chromatography) detection method capable of simultaneously detecting contents of six amino acids in a donkey-hide glue blood-supplementing preparation. The method comprises the following steps: providing a donkey-hide glue blood-supplementing preparation sample; providing an amino acid standard sample; respectively adding the donkey-hide glue blood-supplementing preparation sample and the amino acid standard sample into phenylisothiocyanate-acetonitrile solution; uniformly mixing and standing, and respectively adding n-hexane; uniformlymixing and standing once again, respectively extracting lower-layer solutions, and filtering to obtain a derivatized donkey-hide glue blood-supplementing preparation sample solution and a derivatizedamino acid standard sample solution; sucking the derivatized donkey-hide glue blood-supplementing preparation sample solution and the derivatized amino acid standard sample solution, and injecting into a liquid chromatograph to perform amino acid detection. According to the invention, the contents of six major amino acids in the donkey-hide glue blood-supplementing preparation are detected by adopting reversed-phase high-performance liquid-phase, the sample treatment method is simple, the detection method has good specificity, repeatability and accuracy, and the internal quality of donkey-hide glue blood-supplementing granules can be well controlled.

Description

technical field [0001] The invention relates to an application method of quality control in the field of pharmacy, in particular to an HPLC determination method for simultaneously determining the contents of six amino acids in donkey glue blood-tonifying preparations. Background technique [0002] In the prescription of donkey-hide gelatin blood-tonifying preparations, donkey-hide gelatin is the king drug, accounting for the largest proportion. It is a traditional Chinese blood-tonifying medicine, and the protein and amino acids in donkey-hide gelatin are the nutritional basis for its tonic effect. [0003] In the standards issued by the Ministry of Traditional Chinese Medicine, the inspection items of Lvjiao Buxue Granules include the inspection of the total nitrogen content of donkey-hide gelatin. The determination method of total nitrogen can only roughly reflect the amount of nitrogen-containing compounds in the sample, and cannot well reflect the situation of protein an...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N30/02G01N30/06G01N30/36A61K36/804A61K9/16A61P7/06A61K35/36
Inventor 欧金秀谷陟欣熊灿娟朱丽王实强张妮瑜毛威
Owner JIUZHITANG
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