CN103293236A - Method for measuring propylamine glutamine content - Google Patents
Method for measuring propylamine glutamine content Download PDFInfo
- Publication number
- CN103293236A CN103293236A CN2012105956525A CN201210595652A CN103293236A CN 103293236 A CN103293236 A CN 103293236A CN 2012105956525 A CN2012105956525 A CN 2012105956525A CN 201210595652 A CN201210595652 A CN 201210595652A CN 103293236 A CN103293236 A CN 103293236A
- Authority
- CN
- China
- Prior art keywords
- measuring
- glutamine
- triethylamine
- phase
- propylamine
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Images
Landscapes
- Investigating Or Analysing Biological Materials (AREA)
- Other Investigation Or Analysis Of Materials By Electrical Means (AREA)
Abstract
The invention discloses a method for measuring the propylamine glutamine content, belonging to the field of chromatographic analysis. In the method, octadecylsilane bonded silica is utilized as a chromatographic column stuffing, a mixed solvent of an organic phase and an aqueous phase of a phosphate buffer solution are utilized as mobile phases, and phenyl isothiocyanate is utilized as a derivatization reagent, wherein the organic phase is acetonitrile or methanol; the phosphate buffer solution is prepared in the following manner: adding water in 5ml of triethylamine to 1000ml, and adjusting the pH to 5.0-6.0 through phosphoric acid; in the mobile phase, the percent by volume of the aqueous phase is 30-50%. Compared with the prior art, the measuring method provided by the invention has the advantages of high analysis sensitivity, accurate and reliable result, and the like, thereby having good popularization and application values in the production field of propylamine glutamine injection.
Description
Technical field
The present invention relates to the stratographic analysis field, specifically utilize the method for post column derivatization high effective liquid chromatography for measuring alanyl glutamine in a kind of alanyl glutamine parenteral solution production run.
Background technology
(alanylglutamine dipeptide, Ala-Gln), (glutamine, donor Gln) remedied the Gln deficiency to N (2)-L-alanyl-L-glutamine, enlarged it clinically as the range of application of intravenous nutrition preparation as glutamine.Alanyl glutamine is an ingredient of parenteral nutrition, is applicable to the patient who needs to replenish glutamine, comprises the patient who is in kalabolism and hypermetabolism situation.
Molecular formula: C
8H
15N
3O
4, molecular weight: 188.18
In the prior art, the high efficiency chromatography method is one of important assay method of measuring the alanyl glutamine, as, the standard YBH06582005 of State Food and Drug Administration uses with amino bonded silica gel to be filling agent; Acetonitrile-0.05mol/L potassium phosphate buffer (pH4.0) (65: 35) is the phase that flows; The detection wavelength is 215nm.Document " pre-column derivatization HPLC method mensuration N (content of 2-L-alanyl-L-glutamine parenteral solution " (" Chinese biochemical drug magazine ", 2006,27 (6) 364-369) select for use the sodium acetate solution (adjusting pH5.50 with spirit of vinegar) of acetonitrile-0.1mol/L=44: 56 as the phase that flows, 30 ℃ of column temperatures, detecting wavelength is the detection analysis that 254nm carries out content; And document " triketohydrindene hydrate column front derivation method is measured the content of alanyl glutamine " (" contemporary Chinese application pharmacy ", 2011,07,669-670) adopt sulfonic group strong-acid Zeo-karb PEEK (4.6mm * 150mm), flowing is sodium potassium damping fluid mutually, flow velocity 0.45ml/min, ninhydrin solution flow velocity are 0.25ml/min.On the one hand and since the stability of alanyl glutamine a little less than, utilize above-mentioned three kinds to flow when measuring mutually, inferior separating effect or appearance time evening, be unfavorable for the content control of alanyl glutamine; On the other hand, the peak shape of alanyl glutamine is relatively poor in the national standard YBH06582005 of State Food and Drug Administration, when detecting certain error is arranged, and the time is longer.
Summary of the invention
Technical assignment of the present invention is at above-mentioned the deficiencies in the prior art, and the method for utilizing post column derivatization high effective liquid chromatography for measuring alanyl glutamine in a kind of alanyl glutamine parenteral solution production run is provided.
Technical assignment of the present invention is realized in the following manner: a kind of method of measuring the alanyl glutamine content, it is characterized in that: be chromatographic column filler with the octadecylsilane chemically bonded silica, be that the mixed solvent of water is as mobile phase with organic phase and phosphate buffered solution, mixed liquor with phenyl isothiocyanate, triethylamine, absolute ethyl alcohol, ultrapure water is derivative reagent
Described organic phase is acetonitrile or methyl alcohol;
Described phosphate buffered solution is made by following mode: 5ml adds water to 1000ml with triethylamine, and regulates pH between 5.0-6.0 with phosphoric acid;
In flowing mutually, the percent by volume of water is 30%～50%.
In order to reach best mensuration effect, the column temperature during mensuration is preferably 30～40 ℃.
Described derivative reagent is preferably the mixed liquor of 400ml phenyl isothiocyanate, 400ml triethylamine, 2800ml absolute ethyl alcohol and 400ml ultrapure water.
1) it is an amount of to get alanyl glutamine, adds organic phased soln and the sample solution that contains 0.5mg among every 1ml is approximately made in dilution, as need testing solution;
2) flow rate of mobile phase being set is 0.8～1.2ml/min, is preferably 1ml/min; Detect wavelength 245nm; Select the C18 post for use, with phenyl isothiocyanate 400ml, triethylamine 400ml, absolute ethyl alcohol 2800ml, ultrapure water 400ml, mixing is post-column derivation reagent, the adjustment flow velocity is 0.25ml/min.；
3) get 1) need testing solution 10～50 μ l, preferred 20 μ l inject high performance liquid chromatograph, finish the mensuration of alanyl glutamine.
Compared with prior art, the wash-out separating effect of the inventive method is better, can make preferably this product separate with the impurity chromatographic peak and degree of separation better, appearance time is suitable, can effectively measure the content of alanyl glutamine, need not the advanced processing sample, the method simple and fast, the sensitivity for analysis height, the result accurately and reliably.Can be used for the quality control of alanyl glutamine.
Description of drawings
Accompanying drawing 1 is the chromatogram that utilizes the embodiment of the invention one measurement result;
Accompanying drawing 2 is the chromatograms that utilize the embodiment of the invention two measurement results.
Embodiment
Explain below the method for mensuration alanyl glutamine content of the present invention being done with specific embodiment with reference to Figure of description.
Embodiment 1
Instrument and condition
High performance liquid chromatograph: Waters2695 pump; The Waters2487 UV-detector
Chromatographic column: octadecylsilane chemically bonded silica is the chromatographic column (250mmx4.6mm, 5 μ m) of filling agent
Phase flows: acetonitrile-0.05mol/L potassium phosphate buffer (pH4.0) (65: 35) isocratic elution
Flow velocity: 1.0ml/min
Detect wavelength: 215nm
Column temperature: 40 ℃
Sampling volume: 20 μ l
Implementation step
Precision is measured this product 5ml, puts in the 100ml measuring bottle, adds the phase dilution that flows to scale, shakes up, and precision is measured 5ml again, puts in the 100ml measuring bottle, adds the phase dilution that flows to scale, shakes up, as need testing solution (making the solution that contains 0.5mg among every 1ml).Get 20 μ l and inject liquid chromatograph, the record chromatogram; Precision takes by weighing through 3 hours the about 25mg of L-alanyl-L-glutamine reference substance of 60 ℃ of drying under reduced pressure in addition, puts in the 50ml measuring bottle, adds to flow phased soln and be diluted to scale, shakes up, and solution is measured with method in contrast.Press external standard method with calculated by peak area, namely.
Get need testing solution and reference substance solution, carry out efficient liquid phase chromatographic analysis under above-mentioned chromatographic condition, the record chromatogram the results are shown in following table and accompanying drawing 1.
Retention time is that 15.968 minutes chromatographic peak is the chromatographic peak of alanyl glutamine among Fig. 1, and chromatogram shows that alanyl glutamine can be issued to good separating at same chromatographic condition with its impurity.
Instrument and condition
High performance liquid chromatograph: Waters2695 pump; The Waters2487 UV-detector
Chromatographic column: octadecylsilane chemically bonded silica is the chromatographic column (250mmx4.6mm, 5 μ m) of filling agent
Phase flows: acetonitrile-triethylamine 5ml adds water to 1000ml (transferring pH to 5.5 with phosphoric acid)=40: 60 etc.
The degree wash-out
Column temperature: 40 ℃
Flow velocity: 1.0ml/min
Detect wavelength: 245nm
Derivative reagent: phenyl isothiocyanate 400ml, triethylamine 400ml, absolute ethyl alcohol 2800ml, ultrapure water 400ml, mixing faces with newly joining.Flow velocity is 0.25ml/min
Sampling volume: 20 μ l
Precision is measured this product 5ml, puts in the 100ml measuring bottle, adds the phase dilution that flows to scale, shakes up, and precision is measured 5ml again, puts in the 100ml measuring bottle, adds the phase dilution that flows to scale, shakes up, as need testing solution (making the solution that contains 0.5mg among every 1ml).Get 20 μ l and inject liquid chromatograph, the record chromatogram; Precision takes by weighing through 3 hours the about 25mg of L-alanyl-L-glutamine reference substance of 60 ℃ of drying under reduced pressure in addition, puts in the 50ml measuring bottle, adds to flow phased soln and be diluted to scale, shakes up, and solution is measured with method in contrast.Press external standard method with calculated by peak area, namely.Get need testing solution, carry out efficient liquid phase chromatographic analysis under above-mentioned chromatographic condition, the record chromatogram the results are shown in following table and accompanying drawing 2.
Retention time is that 8.147 minutes chromatographic peak is the chromatographic peak of alanyl glutamine among Fig. 2, good with impurity degree of separation on every side, by phenyl isothiocyanate as post-column derivation reagent, phenyl ring is introduced alanyl to be hung on the molecule of acid amides, form stable chemical constitution, be convenient to the assay of alanyl glutamine.Go out the retention time at peak than embodiment 1 early, also be better than embodiment 1 at aspects such as peak shape and theoretical cam curves, the accuracy of assay is higher.
Claims (3)
1. method of measuring the alanyl glutamine content, it is characterized in that: be chromatographic column filler with the octadecylsilane chemically bonded silica, be that the mixed solvent of water is as mobile phase with organic phase and phosphate buffered solution, mixed liquor with phenyl isothiocyanate, triethylamine, absolute ethyl alcohol, ultrapure water is derivative reagent
Described organic phase is acetonitrile or methyl alcohol;
Described phosphate buffered solution is made by following mode: 5ml adds water to 1000ml with triethylamine, and regulates pH between 5.0-6.0 with phosphoric acid;
In flowing mutually, the percent by volume of water is 30%～50%.
2. the method for mensuration alanyl glutamine content according to claim 1, it is characterized in that: the column temperature during mensuration is 30～40 ℃.
3. the method for mensuration alanyl glutamine content according to claim 1, it is characterized in that: described derivative reagent is the mixed liquor of 400ml phenyl isothiocyanate, 400ml triethylamine, 2800ml absolute ethyl alcohol and 400ml ultrapure water.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2012105956525A CN103293236A (en) | 2012-12-26 | 2012-12-26 | Method for measuring propylamine glutamine content |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2012105956525A CN103293236A (en) | 2012-12-26 | 2012-12-26 | Method for measuring propylamine glutamine content |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN103293236A true CN103293236A (en) | 2013-09-11 |
Family
ID=49094466
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2012105956525A Pending CN103293236A (en) | 2012-12-26 | 2012-12-26 | Method for measuring propylamine glutamine content |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103293236A (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111812259A (en) * | 2020-07-24 | 2020-10-23 | 费森尤斯卡比华瑞制药有限公司 | Method for detecting alanyl glutamine impurity in compound amino acid injection |
| CN113933403A (en) * | 2020-07-13 | 2022-01-14 | 苏州科伦药物研究有限公司 | Determination method of alanyl-glutamine related substances |
| CN115060827A (en) * | 2022-06-27 | 2022-09-16 | 辰欣药业股份有限公司 | Analysis method of related substances of alanyl glutamine injection |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1435691A (en) * | 2002-01-26 | 2003-08-13 | 中国科学院昆明动物研究所 | Method for determining amino acid in microdialysis sample by phenyl isothiocyanate derivatization method |
| CN1602850A (en) * | 2004-08-02 | 2005-04-06 | 陶灵刚 | N(2)-L-alanyl-L-glutamine injection and its preparation method |
| CN102406749A (en) * | 2010-11-22 | 2012-04-11 | 九芝堂股份有限公司 | HPLC (high-performance liquid chromatography) detection method capable of simultaneously detecting contents of six amino acids in donkey-hide glue blood-supplementing preparation |
| CN102749397A (en) * | 2012-07-19 | 2012-10-24 | 天津金耀集团有限公司 | Analysis method of compound amino acid and dipeptide injection |
-
2012
- 2012-12-26 CN CN2012105956525A patent/CN103293236A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1435691A (en) * | 2002-01-26 | 2003-08-13 | 中国科学院昆明动物研究所 | Method for determining amino acid in microdialysis sample by phenyl isothiocyanate derivatization method |
| CN1602850A (en) * | 2004-08-02 | 2005-04-06 | 陶灵刚 | N(2)-L-alanyl-L-glutamine injection and its preparation method |
| CN102406749A (en) * | 2010-11-22 | 2012-04-11 | 九芝堂股份有限公司 | HPLC (high-performance liquid chromatography) detection method capable of simultaneously detecting contents of six amino acids in donkey-hide glue blood-supplementing preparation |
| CN102749397A (en) * | 2012-07-19 | 2012-10-24 | 天津金耀集团有限公司 | Analysis method of compound amino acid and dipeptide injection |
Non-Patent Citations (6)
| Title |
|---|
| MARIA GOMEZ-MINGOT ET AL.: "Development of a novel analytical approach combining the quantification of amino acids, organic acids and glucose using HPLC-UV-Vis and HPLC-MS with screening via NMR", 《ANAL. METHODS》 * |
| 张云楚 等: "柱前衍生化HPLC法测定N(2)-L-丙氨酰-L-谷氨酰胺注射液的含量", 《中国生化药物杂志》 * |
| 朱彭龄: "磷酸三乙胺-乙腈流动相体系反相分离2,4-二硝基氟苯-氨基酸衍生物", 《分析测试技术与仪器》 * |
| 瞿其曙 等: "柱前衍生法在氨基酸分析测定中的应用", 《化学进展》 * |
| 诸敏 等: "茚三酮柱后衍生法测定丙氨酰-谷氨酰胺的含量", 《中国现代应用药学》 * |
| 鲍玉洲 等: "用高效液相法行人眼组织氨基酸含量测定", 《眼科研究》 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113933403A (en) * | 2020-07-13 | 2022-01-14 | 苏州科伦药物研究有限公司 | Determination method of alanyl-glutamine related substances |
| CN111812259A (en) * | 2020-07-24 | 2020-10-23 | 费森尤斯卡比华瑞制药有限公司 | Method for detecting alanyl glutamine impurity in compound amino acid injection |
| CN115060827A (en) * | 2022-06-27 | 2022-09-16 | 辰欣药业股份有限公司 | Analysis method of related substances of alanyl glutamine injection |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN104483406B (en) | The high-efficiency liquid chromatography method for detecting of L-arginine-α-ketoglutaric acid | |
| CN102749397A (en) | Analysis method of compound amino acid and dipeptide injection | |
| CN103487518B (en) | Impurity detection method and content determination method for clindamycin hydrochloride for injection | |
| CN104280475A (en) | Method for detecting content of amino acid and glycyl glutamine in compound amino acid (15) dipeptide (2) injection | |
| CN104965041A (en) | High performance liquid chromatography detection method for parecoxib sodium isomer | |
| CN102288687B (en) | Method for analyzing and detecting impurities in ornithine aspartate | |
| CN102590393B (en) | Method for simultaneously determining contents of multiple ingredients in multivitamin amino acid chewing pill | |
| CN106841408B (en) | The rapid detection method of methionine sulfoxide in a kind of amino acid injection | |
| CN103293236A (en) | Method for measuring propylamine glutamine content | |
| CN111323527B (en) | Method for simultaneously measuring various psychotropic drugs by using composite two-dimensional liquid chromatography | |
| CN105388225A (en) | Analysis and detection method for UDPC in medicinal preparation containing citicoline sodium | |
| CN103134892A (en) | Analytical method of compound amino acid (15) double peptide (2) injection | |
| CN105223280A (en) | A kind of black-bone chicken increases the assay method of amino acid content in emulsion formulation | |
| CN106383185B (en) | The high-efficiency liquid chromatography method for detecting of carbobenzyloxy-L-alanine | |
| CN101458235B (en) | Matrine liquid chromatography measuring method | |
| CN111323503A (en) | Method for measuring content of compound amino acid injection | |
| CN102636582B (en) | Method for determining content of diminazene and antipyrine in diminazene particle | |
| CN100480698C (en) | Method for inspecting hypotensive medicine | |
| CN105181859A (en) | Cysteine hydrochloride in acetaminophen injection and test method of degradation product cystine | |
| CN103175930B (en) | A kind of HPLC analytical method measuring sodium sulphite content | |
| CN103063794A (en) | Content detecting and control method of epalrestat tablets | |
| CN103995062B (en) | Method for testing exenatide and impurities thereof by using high performance liquid chromatography | |
| CN102901783A (en) | Content determination method for pranoprofen eye drops | |
| CN102928525A (en) | Method for detecting related substance II of cefpirome in cefpirome sulfate/sodium chloride injection | |
| CN106018573A (en) | Method for detecting purity of tipiracil hydrochloride in TAS-102 |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20130911 |
|
| WD01 | Invention patent application deemed withdrawn after publication |