CN106645536A - Method for determining content of low-molecular-weight PEG modifier - Google Patents

Method for determining content of low-molecular-weight PEG modifier Download PDF

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CN106645536A
CN106645536A CN201710013691.2A CN201710013691A CN106645536A CN 106645536 A CN106645536 A CN 106645536A CN 201710013691 A CN201710013691 A CN 201710013691A CN 106645536 A CN106645536 A CN 106645536A
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molecular
temperature
low
peg
column
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CN106645536B (en
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施立钦
邱从平
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Ningbo Polytechnic
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/02—Column chromatography
    • G01N30/88—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/02—Column chromatography
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/02—Column chromatography
    • G01N30/04—Preparation or injection of sample to be analysed
    • G01N30/06—Preparation
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/02—Column chromatography
    • G01N30/62—Detectors specially adapted therefor
    • G01N30/64—Electrical detectors
    • G01N30/68—Flame ionisation detectors
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/02—Column chromatography
    • G01N2030/022—Column chromatography characterised by the kind of separation mechanism
    • G01N2030/025—Gas chromatography
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/02—Column chromatography
    • G01N30/04—Preparation or injection of sample to be analysed
    • G01N30/06—Preparation
    • G01N2030/062—Preparation extracting sample from raw material
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/02—Column chromatography
    • G01N30/88—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86
    • G01N2030/8809—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86 analysis specially adapted for the sample
    • G01N2030/884—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86 analysis specially adapted for the sample organic compounds
    • G01N2030/885—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86 analysis specially adapted for the sample organic compounds involving polymers

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  • Physics & Mathematics (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Pathology (AREA)
  • Investigating Or Analysing Biological Materials (AREA)
  • Medicinal Preparation (AREA)

Abstract

The invention discloses a method for determining the content of a low-molecular-weight PEG modifier. The method comprises the steps of firstly building a PEG determination standard curve through an FID detector, adding a proper amount of pure water to a to-be-detected liquid and diluting the to-be-detected liquid into proper concentration, wherein by adopting a capillary column, carrier gas is high-purity nitrogen, the column flow is 1mL/min and the temperature of a vaporizing chamber is 250 DEG C; by adopting the FID detector, measuring the peak area under the conditions that the temperature is 250 DEG C and the column temperature is 150 DEG C; and obtaining the content of the low-molecular-weight PEG modifier in the to-be-detected liquid according to the standard curve. Quantitative detection and analysis are carried out by adopting a gas chromatography, and the built method is fast, simple and accurate, and can be widely used for content determination, analysis and detection of the low-molecular-weight PEG modifier or medicine.

Description

Low-molecular-weight PEG dressing agent content assaying method
Technical field
The invention belongs to Pharmaceutical Analysis technical field.
Background technology
Polyethylene glycol (PEG) is a kind of pH neutral, nontoxic, and the parent with unique physicochemical property and good biocompatibility Water high molecular polymer, the only a few for being also Jing FDA approvals can be with one of synthetic polymer of internal injection.When PEG is coupled to When on protein, polypeptide, small molecule organic drugs or nano particle shell, immune clearance and the kidney of pharmaceutical preparation can be reduced Dirty quick elimination, extends the circulation time in vivo of medicine, reduces the toxicity of medicine.As new pharmaceutic adjuvant, the matter of PEG Amount control and absorption in vivo, distribution and discharge process for PEG chemical drug things design with evaluation with highly important Meaning.
A kind of new feature PEG dressing agent of diethylene glycol dressing agent, be in hitherto known polymer albumen and Cell absorbs the minimum polymer of level, and due to the nontoxic and good biocompatibility of diethylene glycol, the drug modification is by work The diethylene glycol of change is coupled on medicine by chemical method, improves the physicochemical property and BA of medicine, improves medicine The property such as thing dynamics and drug effect, significantly increases the dissolubility and stability of medicine, reduces immunogenicity and antigenicity, reduces poison Side effect, the effects such as increased in vivo bioactivity and curative effect, this technology started to be applied to protein (peptides), enzyme, In antibody and small-molecule drug.
At present the analysis for PEG is frequently with radioactive label method, colorimetric method, Western Blot and high performance liquid chromatography Method (HPLC), but these methods exist it is clearly disadvantageous.For example, cost is too high when radioactive label method is used to mark PEG, and The methodology certification label efficiency of maturation is not yet set up at present, also illustrates the moving in vivo of the PEG after mark without contrast test Whether mechanical behavior there occurs change；Colorimetric method and Western Blot methods can not all obtain accurate quantitative result, and the latter It is by determining the amount of antibody combined with PEG PEG to be carried out quantitatively indirectly.In view of the poor selectivity of antibody, may and sample Present in endogenous interfering material combine, therefore be not suitable for the quantitative analysis of PEG in the complex biological sample such as blood plasma or tissue； HPLC methods adopt RI-detector, and analysis time is long, and lower limit of quantitation is only 50mg/mL, are not enough to the biological trace in vivo of analysis The PEG levels of amount.
The content of the invention
It is an object of the invention to provide a kind of low-molecular-weight PEG dressing agent content assaying method.
To reach above-mentioned purpose, the technical solution used in the present invention is：
A kind of low-molecular-weight PEG dressing agent content assaying method, is detected, detecting step is using gas chromatography：
1) PEG bioassay standard curves are initially set up by fid detector, using capillary column, carrier gas is High Purity Nitrogen, post stream Speed is 1mL/min, and temperature of vaporization chamber is 250 DEG C, and using fid detector, 250 DEG C of temperature, column temperature is 150 DEG C, and sample size is 1uL, determine concentration be 1,2,3,4, the standard liquid of 5ug/mL, obtain PEG bioassay standard curves, peak area is linearly closed with concentration System is good, R2=0.992；
2) appropriate pure water is added to be diluted to debita spissitudo liquid to be detected, using capillary column, carrier gas is High Purity Nitrogen, column flow For 1mL/min, temperature of vaporization chamber is 250 DEG C, and using fid detector, 250 DEG C of temperature, column temperature is measure peak face under the conditions of 150 DEG C Product；
3) according to calibration curve, the content of low-molecular-weight PEG dressing agent in liquid to be detected is obtained.
Further, it is the PEG dressing agents within 2000 that described low-molecular-weight PEG dressing agent refers to molecular weight.
Further, described low-molecular-weight PEG dressing agent be diethylene glycol methyl propionate, diethylene glycol ethyl propionate, Any one in diethylene glycol n-butyl propionate.
Beneficial effect：
The present invention carries out quantitative detecting analysis, institute according to the architectural characteristic of low-molecular-weight PEG dressing agent using gas-chromatography Construction method is quick, easy, accurately, is expected to using the PEG dressing agents of low-molecular-weight or detecting containing measure analysis for medicine.
Description of the drawings
Fig. 1 is the calibration curve of diethylene glycol methyl propionate in a kind of embodiment 1.
Specific embodiment
The present invention is described in further detail with reference to the accompanying drawings and detailed description.
Embodiment 1
The measure of diethylene glycol methyl propionate
(1) standard items storing solution:Diethylene glycol methyl propionate standard items 100mg is accurately weighed, is dissolved in water and constant volume is arrived 1000mL, obtains 100ug/mL respective standard storing solutions, in being stored in 4 DEG C of refrigerator.
(2) standard solution：Respectively draw 1,2,3,4,5mL certain volumes standard reserving solution in 100mL volumetric flasks, plus Water constant volume obtain respective concentration for 1,2,3,4,5ug/mL titers (now with the current).
Testing sample process：Accurate measuring 1ml prepare liquids add appropriate pure water to be diluted to concentration for 1-5ug/mL debita spissitudos.
Using capillary column, carrier gas is High Purity Nitrogen, and column flow is 1ml/min, and temperature of vaporization chamber is 250 DEG C, is examined using FID Device, 250 DEG C of temperature are surveyed, column temperature is 150 DEG C, peak area (R2 good with concentration linear relationship>0.99, average recovery 98.3- 100.5%.
Standard liquid is using 1,2,3,4,5ug/mL concentration, draw calibration curve linearly dependent coefficient R2=0.992, be loaded The rate of recovery 98.3-100.5% its lowest detection is limited to 0.59ug/mL (S/N=3), and detection precision is 0.65%.
Appropriate pure water is added to be diluted to debita spissitudo liquid to be detected, using capillary column, carrier gas is High Purity Nitrogen, and column flow is 1mL/min, temperature of vaporization chamber is 250 DEG C, and using fid detector, 250 DEG C of temperature, column temperature is measure peak face under the conditions of 150 DEG C Product, according to calibration curve, obtains the content of low-molecular-weight PEG dressing agent in liquid to be detected.
The determinand adopted in above-described embodiment is diethylene glycol methyl propionate, the results showed, molecular weight be 2000 with Interior PEG dressing agents can obtain preferable result using said method.
Although being illustrated to embodiments of the present invention in specification, these embodiments are intended only as prompting, Should not limit protection scope of the present invention.It is equal that various omissions, displacement and change are carried out without departing from the spirit and scope of the present invention Should be comprising within the scope of the present invention.

Claims (3)

1. a kind of low-molecular-weight PEG dressing agent content assaying method, it is characterised in that detected using gas chromatography, is examined Surveying step is：
1) PEG bioassay standard curves are initially set up by fid detector, using capillary column, carrier gas is High Purity Nitrogen, and column flow rate is 1mL/min, temperature of vaporization chamber is 250 DEG C, and using fid detector, 250 DEG C of temperature, column temperature is 150 DEG C, and sample size is 1uL, is surveyed Determine concentration be 1,2,3,4, the standard liquid of 5ug/mL, obtain PEG bioassay standard curves, peak area is good with concentration linear relationship It is good, R2=0.992；
2) appropriate pure water is added to be diluted to debita spissitudo liquid to be detected, using capillary column, carrier gas is High Purity Nitrogen, and column flow is 1mL/min, temperature of vaporization chamber is 250 DEG C, and using fid detector, 250 DEG C of temperature, column temperature is measure peak face under the conditions of 150 DEG C Product；
3) according to calibration curve, the content of low-molecular-weight PEG dressing agent in liquid to be detected is obtained.
2. low-molecular-weight PEG dressing agent content assaying method according to claim 1, it is characterised in that：Described low point It is the PEG dressing agents within 2000 that son amount PEG dressing agents refer to molecular weight.
3. low-molecular-weight PEG dressing agent content assaying method according to claim 2, it is characterised in that：Described low point Son amount PEG dressing agents are any one in diethylene glycol methyl propionate, diethylene glycol ethyl propionate, diethylene glycol n-butyl propionate Kind.
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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN112858687A (en) * 2020-12-30 2021-05-28 宁波职业技术学院 Serum amyloid protein A detection reagent and preparation method thereof

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WO2010090596A1 (en) * 2009-02-04 2010-08-12 Agency For Science, Technology And Research Hollow silica particle with a polymer thereon
CN102226792A (en) * 2011-04-01 2011-10-26 广州市谱尼测试技术有限公司 Detection method for 2-methoxyethanol, 2-ethoxyethanol, 2-methoxyethanol acetate and 2-ethoxyethanol acetate
CN102998387A (en) * 2012-11-23 2013-03-27 江苏中烟工业有限责任公司 Method for determining ethylene glycol monomethyl ether, glycol ether, ethylene glycol ether acetate, glycol and diglycol in food wrap paper
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CN104614451A (en) * 2014-12-08 2015-05-13 宁波北仑大港科技创新服务中心 Methyl 3-(2-(2-hydroxyethoxy)ethoxy)propanoate modification agent detection method

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN112858687A (en) * 2020-12-30 2021-05-28 宁波职业技术学院 Serum amyloid protein A detection reagent and preparation method thereof
CN112858687B (en) * 2020-12-30 2023-09-15 宁波职业技术学院 Serum amyloid A detection reagent and preparation method thereof

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