阅读说明：本技术 一种小儿复方氨基酸注射液含量检测方法 (Method for detecting content of compound amino acid injection for children ) 是由 龚若 孙文敬 于 2020-12-31 设计创作，主要内容包括：本发明公开了一种小儿复方氨基酸注射液含量检测方法。该方法采用自动在线衍生化与HPLC相结合的方法进行测定。本发明具有如下优点：流动相的配制操作简单,分析方法稳定,灵敏度高,可简便,高效、全面的检测小儿复方氨基酸注射液中的门冬氨酸、谷氨酸、丝氨酸、组氨酸、甘氨酸、苏氨酸、精氨酸、丙氨酸、酪氨酸、缬氨酸、色氨酸、苯丙氨酸、异亮氨酸、亮氨酸含量,有效检测控制小儿复方氨基酸注射液在制备、研究、检测及存放过程中的含量变化,可用于小儿复方氨基酸注射液的质量控制。(The invention discloses a method for detecting the content of a compound amino acid injection for children. The method adopts a method combining automatic on-line derivatization and HPLC for determination. The invention has the following advantages: the preparation operation of the mobile phase is simple, the analysis method is stable, the sensitivity is high, the content of aspartic acid, glutamic acid, serine, histidine, glycine, threonine, arginine, alanine, tyrosine, valine, tryptophan, phenylalanine, isoleucine and leucine in the compound amino acid injection for children can be simply, conveniently, efficiently and comprehensively detected, the content change of the compound amino acid injection for children in the preparation, research, detection and storage processes can be effectively detected and controlled, and the method can be used for quality control of the compound amino acid injection for children.)

1. The method for measuring the content of the compound amino acid injection for children is characterized by measuring by adopting a method combining automatic on-line derivatization and HPLC, wherein the chromatographic conditions are as follows: the chromatographic column is Agilent Zorbax Eclipse AAA (4.6X 150mm, 3.5 μm); the detection wavelength is 338 nm; the flow rate is 1.0 ml/min; the column temperature was 40 ℃; the mobile phase A is NaH₂PO₄And mixed solution of triethylamine, NaH₂PO₄The concentration of the mixed solution is 40mmol/L, the concentration of triethylamine is 0.03 wt%, and the pH value is adjusted to 6.8 by using a sodium hydroxide solution; the volume ratio of the mobile phase B is 40:40:20 of acetonitrile-ethanol-water mixture; elution was carried out with a gradient as follows:

time (minutes) Mobile phase A (%) Mobile phase B (%) 0 95 5 17 84 16 34 48 52 34.1 95 5 39 95 5

Technical Field

The invention belongs to the technical field of pharmaceutical analysis, and particularly relates to a method for detecting the content of a compound amino acid injection for children.

Background

The compound amino acid injection (18AA-I) for children is mainly used for supplying heat for parenteral nutrition when nutrition cannot be taken through the intestinal tract or the intake is insufficient in various post-traumatic high-metabolic states of premature infants, low-weight infants and newborns. "infants belong to a special period of human growth and development, and need special nutritional formula for nutritional support in pathological state. The formula of the compound amino acid injection 18AA-I for children particularly reduces the contents of phenylalanine, methionine and glycine, increases the dosage of cysteine, tyrosine, histidine and arginine, and can better meet the nutritional requirements of infants in special periods. "

The pharmacokinetics is that the tissue protein of human body is decomposed into amino acid on one hand, and the tissue protein is synthesized from the amino acid on the other hand, the continuous decomposition and synthesis are realized, the dynamic balance is kept, and the amino acid conversion is very rapid and frequent. The twenty amino acids that make up a protein all have common groups-amino and hydroxyl groups, with inherently similar metabolic processes. They all need to be deaminated to form ammonia and alpha-keto acid, and ammonia and carbon dioxide to form urea which is excreted by the kidney; the alpha-keto acid can provide energy, produce water and carbon dioxide to be discharged, and can be converted into sugar or fat.

The compound amino acid injection (18AA-I) for children is mainly suitable for children who can not take food through intestines and stomach due to digestive system diseases; children with hypoproteinemia caused by various diseases; children suffered from serious trauma, burn, septicemia and other imbalance of nitrogen balance in vivo; refractory diarrhea, malabsorption syndrome or parenteral nutrition of premature infants and low-weight infants, and has wide clinical application. The research on a method for detecting the content of the pediatric compound amino acid injection (18AA-I) with strong specificity and high sensitivity is very important.

Disclosure of Invention

The invention aims to provide a method for detecting the content of the compound amino acid injection for children, which can be used for quality control in the production of the compound amino acid injection for children.

The method for measuring the content of the pediatric compound amino acid injection adopts a method combining automatic on-line derivatization and HPLC, and the chromatographic conditions are as follows: the chromatographic column is Agilent Zorbax Eclipse AAA (4.6X 150mm, 3.5 μm); the detection wavelength is 338 nm; the flow rate is 1.0 ml/min; the column temperature was 40 ℃; the mobile phase A is NaH₂PO₄And mixed solution of triethylamine, NaH₂PO₄The concentration of the mixed solution is 40mmol/L, the concentration of triethylamine is 0.03 wt%, and the pH value is adjusted to 6.8 by using a sodium hydroxide solution; the volume ratio of the mobile phase B is 40:40:20 of acetonitrile-ethanol-water mixture; elution was carried out with a gradient as follows:

time (minutes)	Mobile phase A (%)	Mobile phase B (%)
			0	95	5
17	84	16
			34	48	52
34.1	95	5
			39	95	5

The invention has the following advantages: the preparation operation of the mobile phase is simple, the analysis method is stable, the sensitivity is high, the content of aspartic acid, glutamic acid, serine, histidine, glycine, threonine, arginine, alanine, tyrosine, valine, tryptophan, phenylalanine, isoleucine and leucine in the compound amino acid injection for children can be simply, conveniently, efficiently and comprehensively detected, the content change of the compound amino acid injection for children in the preparation, research, detection and storage processes can be effectively detected and controlled, and the method can be used for quality control of the compound amino acid injection for children.

Detailed Description

The present invention is described in further detail below with reference to specific embodiments, which are given for the purpose of illustration only and are not intended to limit the scope of the invention.

Example 1

Solution preparation:

1) sodium hydroxide solution: 4.3g of sodium hydroxide was taken, dissolved in water to 100ml, and shaken to obtain a sodium hydroxide solution.

2) 9.6g of anhydrous sodium dihydrogen phosphate is precisely weighed, placed in a 2L beaker, dissolved by adding 2000ml of water, 0.6ml of triethylamine is dropwise added, the pH value is adjusted to 6.8 by using sodium hydroxide solution, and the mixture is uniformly stirred, filtered, and subjected to ultrasonic treatment to be used as a mobile phase A.

3) Weighing 400ml of acetonitrile, 400ml of ethanol and 200ml of water, uniformly mixing, carrying out suction filtration and ultrasonic treatment to obtain a mobile phase B.

Amino acid controls were weighed as follows:

serial number	Name (R)	Sample weighing (mg)
			1	Histidine hydrochloride (C)6H9N3O2·HCl·H2O)	12.3
2	Glycine (C)2H5NO2)	16.2
			3	Arginine hydrochloride (C)6H14N4O2·HCl)	14.45
4	Phenylalanine (C)9H11NO2)	14.15
			5	Leucine (C)6H13NO2)	18.95
6	Aspartic acid (C)4H7NO4)	23
			7	Glutamic acid (C)5H9NO4)	39.4
8	Serine (C)3H7NO3)	13.4
			9	Threonine (C)4H9NO3)	39.4
10	Alanine (C)3H7NO2)	37.6
			11	Valine (C)5H11NO2)	27
12	Tryptophan (C)11H12N2O2)	21.2
			13	Isoleucine (C)6H13NO2)	34
14	Tyrosine (C)9H11NO3)	11

Putting the weighed No. 1-6 reference substance into a 50ml measuring flask, adding water to dissolve and dilute the reference substance to a scale, and shaking up to obtain a reference solution I; placing the weighed No. 7-13 reference substance into a 100ml measuring flask, adding water to dissolve and dilute the reference substance to a scale, and shaking up to obtain a reference solution II; the weighed No. 14 reference substance is placed in a 100ml measuring flask, dissolved and diluted to the scale by adding water, and shaken up to be used as a reference solution III. Precisely sucking 10ml of the control solution I, 5ml of the control solution II and 1ml of the control solution III respectively, placing the reference solutions in a same 100ml measuring flask, and adding water to a constant volume to obtain reference solution.

Precisely sucking 1ml of the compound amino acid injection (18AA-I) for children into a 100ml measuring flask, adding water to dilute to scale, and shaking up to obtain the compound amino acid injection.

Chromatographic conditions are as follows:

the type of the chromatographic column: agilent Zorbax Eclipse AAA (4.6 x 150mm, 3.5 um);

mobile phase: a: 40mmol/L NaH2PO4+ 0.03% triethylamine (adjusted to pH 6.8 with sodium hydroxide solution);

mobile phase B: acetonitrile-ethanol-water (40:40: 20);

detection wavelength: 338 nm;

flow rate: 1.0 ml/min;

column temperature: 40 ℃;

the gradient elution was as follows:

time (minutes)	Mobile phase A (%)	Mobile phase B (%)
			0	95	5
17	84	16
			34	48	52
34.1	95	5
			39	95	5

The detection instrument adopts an Agilent 1260 type high performance liquid chromatograph.

The peak appearance sequence of amino acid on the chromatographic column is as follows: aspartic acid, glutamic acid, serine, histidine, glycine, threonine, arginine, alanine, tyrosine, valine, tryptophan, phenylalanine, isoleucine, leucine.

The above is only a preferred embodiment of the present invention, and it should be noted that it is possible for those skilled in the art to make various changes and modifications without departing from the inventive concept of the present invention, and these changes and modifications are all within the scope of the present invention.